StemCell Therapy

Abstract

In recent years, there has been an explosion of interest in stem cells, not just within the scientific and medical communities but also among politicians, religious groups and ethicists. Here, we summarize the different types of stem cells that have been described: their origins in embryonic and adult tissues and their differentiation potential in vivo and in culture. We review some current clinical applications of stem cells, highlighting the problems encountered when going from proof-of-principle in the laboratory to widespread clinical practice. While some of the key genetic and epigenetic factors that determine stem cell properties have been identified, there is still much to be learned about how these factors interact. There is a growing realization of the importance of environmental factors in regulating stem cell behaviour and this is being explored by imaging stem cells in vivo and recreating artificial niches in vitro. New therapies, based on stem cell transplantation or endogenous stem cells, are emerging areas, as is drug discovery based on patient-specific pluripotent cells and cancer stem cells. What makes stem cell research so exciting is its tremendous potential to benefit human health and the opportunities for interdisciplinary research that it presents.

Keywords: adult stem cells, ES cells, iPS cells, cell-based therapies, drug discovery

The human body comprises over 200 different cell types that are organized into tissues and organs to provide all the functions required for viability and reproduction. Historically, biologists have been interested primarily in the events that occur prior to birth. The second half of the twentieth century was a golden era for developmental biology, since the key regulatory pathways that control specification and morphogenesis of tissues were defined at the molecular level (Arias 2008). The origins of stem cell research lie in a desire to understand how tissues are maintained in adult life, rather than how different cell types arise in the embryo. An interest in adult tissues fell, historically, within the remit of pathologists and thus tended to be considered in the context of disease, particularly cancer.

It was appreciated long ago that within a given tissue there is cellular heterogeneity: in some tissues, such as the blood, skin and intestinal epithelium, the differentiated cells have a short lifespan and are unable to self-renew. This led to the concept that such tissues are maintained by stem cells, defined as cells with extensive renewal capacity and the ability to generate daughter cells that undergo further differentiation (Lajtha 1979). Such cells generate only the differentiated lineages appropriate for the tissue in which they reside and are thus referred to as multipotent or unipotent (figure1).

Origin of stem cells. Cells are described as pluripotent if they can form all the cell types of the adult organism. If, in addition, they can form the extraembryonic tissues of the embryo, they are described as totipotent. Multipotent stem cells have the ability to form all the differentiated cell types of a given tissue. In some cases, a tissue contains only one differentiated lineage and the stem cells that maintain the lineage are described as unipotent. Postnatal spermatogonial stem cells, which are unipotent in vivo but pluripotent in culture, are not shown (Jaenisch & Young 2008). CNS, central nervous system; ICM, inner cell mass.

In the early days of stem cell research, a distinction was generally made between three types of tissue: those, such as epidermis, with rapid turnover of differentiated cells; those, such as brain, in which there appeared to be no self-renewal; and those, such as liver, in which cells divided to give two daughter cells that were functionally equivalent (Leblond 1964; Hall & Watt 1989). While it remains true that different adult tissues differ in terms of the proportion of proliferative cells and the nature of the differentiation compartment, in recent years it has become apparent that some tissues that appeared to lack self-renewal ability do indeed contain stem cells (Zhao et al. 2008) and others contain a previously unrecognized cellular heterogeneity (Zaret & Grompe 2008). That is not to say that all tissues are maintained by stem cells; for example, in the pancreas, there is evidence against the existence of a distinct stem cell compartment (Dor et al. 2004).

One reason why it took so long for stem cells to become a well-established research field is that in the early years too much time and energy were expended in trying to define stem cells and in arguing about whether or not a particular cell was truly a stem cell (Watt 1999). Additional putative characteristics of stem cells, such as rarity, capacity for asymmetric division or tendency to divide infrequently, were incorporated into the definition, so that if a cell did not exhibit these additional properties it tended to be excluded from the stem cell list. Some researchers still remain anxious about the definitions and try to hedge their bets by describing a cell as a stem/progenitor cell. However, this is not useful. The use of the term progenitor, or transit amplifying, cell should be reserved for a cell that has left the stem cell compartment but still retains the ability to undergo cell division and further differentiation (Potten & Loeffler 2008).

Looking back at some of the early collections of reviews written as the proceedings of stem cell conferences, one regularly finds articles on the topic of cancer stem cells (McCulloch et al. 1988). However, these cells have only recently received widespread attention (Reya et al. 2001; Clarke et al. 2006; Dick 2008). The concept is very similar to the concept of normal tissue stem cells, namely that cells in tumours are heterogeneous, with only some, the cancer stem cells, or tumour initiating cells, being capable of tumour maintenance or regrowth following chemotherapy. The cancer stem cell concept is important because it suggests new approaches to anti-cancer therapies (figure2).

The cancer stem cell hypothesis. The upper tumour is shown as comprising a uniform population of cells, while the lower tumour contains both cancer stem cells and more differentiated cells. Successful or unsuccessful chemotherapy is interpreted according to the behaviour of cells within the tumour.

As in the case of tissue stem cells, it is important that cancer stem cell research is not sidetracked by arguments about definitions. It is quite likely that in some tumours all the cells are functionally equivalent, and there is no doubt that tumour cells, like normal stem cells, can behave differently under different assay conditions (Quintana et al. 2008). The oncogene dogma (Hahn & Weinberg 2002), that tumours arise through step-wise accumulation of oncogenic mutations, does not adequately account for cellular heterogeneity, and markers of stem cells in specific cancers have already been described (Singh et al. 2004; Barab et al. 2007; O'Brien et al. 2007). While the (rediscovered) cancer stem cell field is currently in its infancy, it is already evident that a cancer stem cell is not necessarily a normal stem cell that has acquired oncogenic mutations. Indeed, there is experimental evidence that cancer initiating cells can be genetically altered progenitor cells (Clarke et al. 2006).

In addition to adult tissue stem cells, stem cells can be isolated from pre-implantation mouse and human embryos and maintained in culture as undifferentiated cells (figure1). Such embryonic stem (ES) cells have the ability to generate all the differentiated cells of the adult and are thus described as being pluripotent (figure1). Mouse ES cells are derived from the inner cell mass of the blastocyst, and following their discovery in 1981 (Evans & Kaufman 1981; Martin 1981) have been used for gene targeting, revolutionizing the field of mouse genetics. In 1998, it was first reported that stem cells could be derived from human blastocysts (Thomson et al. 1998), opening up great opportunities for stem cell-based therapies, but also provoking controversy because the cells are derived from spare in vitro fertilization embryos that have the potential to produce a human being. It is interesting to note that, just as research on adult tissue stem cells is intimately linked to research on disease states, particularly cancer, the same is true for ES cells. Many years before the development of ES cells, the in vitro differentiation of cells derived from teratocarcinomas, known as embryonal carcinoma cells, provided an important model for studying lineage selection (Andrews et al. 2005).

Blastocysts are not the only source of pluripotent ES cells (figure1). Pluripotent epiblast stem cells, known as epiSC, can be derived from the post-implantation epiblast of mouse embryos (Brons et al. 2007; Tesar et al. 2007). Recent gene expression profiling studies suggest that human ES cells are more similar to epiSC than to mouse ES cells (Tesar et al. 2007). Pluripotent stem cells can also be derived from primordial germ cells (EG cells), progenitors of adult gametes, which diverge from the somatic lineage at late embryonic to early foetal development (Kerr et al. 2006).

Although in the past the tendency has been to describe ES cells as pluripotent and adult stem cells as having a more restricted range of differentiation options, adult cells can, in some circumstances, produce progeny that differentiate across the three primary germ layers (ectoderm, mesoderm and endoderm). Adult cells can be reprogrammed to a pluripotent state by transfer of the adult nucleus into the cytoplasm of an oocyte (Gurdon et al. 1958; Gurdon & Melton 2008) or by fusion with a pluripotent cell (Miller & Ruddle 1976). The most famous example of cloning by transfer of a somatic nucleus into an oocyte is the creation of Dolly the sheep (Wilmut et al. 1997). While the process remains inefficient, it has found some unexpected applications, such as cloning endangered species and domestic pets.

A flurry of reports almost 10 years ago suggested that adult cells from many tissues could differentiate into other cell types if placed in a new tissue environment. Such studies are now largely discredited, although there are still some bona fide examples of transdifferentiation of adult cells, such as occurs when blood cells fuse with hepatocytes during repair of damaged liver (Anderson et al. 2001; Jaenisch & Young 2008). In addition, it has been known for many years that adult urodele amphibians can regenerate limbs or the eye lens following injury; this involves dedifferentiation and subsequent transdifferentiation steps (Brockes & Kumar 2005).

The early studies involving somatic nuclear transfer indicated that adult cells can be reprogrammed to pluripotency. However, the mechanistic and practical applications of inducing pluripotency in adult cells have only become apparent in the last 2 or 3 years, with the emergence of a new research area: induced pluripotent stem cells (iPS cells). The original report demonstrated that retrovirus-mediated transduction of mouse fibroblasts with four transcription factors (Oct-3/4, Sox2, KLF4 and c-Myc; figure1) that are highly expressed in ES cells could induce the fibroblasts to become pluripotent (Takahashi & Yamanaka 2006). Since then, rapid progress has been made: iPS cells can be generated from adult human cells (Takahashi et al. 2007; Yu et al. 2007; Park et al. 2008a); cells from a range of tissues can be reprogrammed (Aasen et al. 2008; Aoi et al. 2008); and iPS cells can be generated from patients with specific diseases (Dimos et al. 2008; Park et al. 2008b). The number of transcription factors required to generate iPS cells has been reduced (Kim et al. 2008); the efficiency of iPS cell generation increased (Wernig et al. 2007); and techniques devised that obviate the need for retroviral vectors (Okita et al. 2008; Stadtfeld et al. 2008). These latter developments are very important for future clinical applications, since the early mice generated from iPS cells developed tumours at high frequency (Takahashi & Yamanaka 2006; Yamanaka 2007). Without a doubt, this is currently the most exciting and rapidly moving area of stem cell research.

In all the publicity that surrounds embryonic and iPS cells, people tend to forget that stem cell-based therapies are already in clinical use and have been for decades. It is instructive to think about these treatments, because they provide important caveats about the journey from proof-of-principle in the laboratory to real patient benefit in the clinic. These caveats include efficacy, patient safety, government legislation and the costs and potential profits involved in patient treatment.

Haemopoietic stem cell transplantation is the oldest stem cell therapy and is the treatment that is most widely available (Perry & Linch 1996; Austin et al. 2008). The stem cells come from bone marrow, peripheral blood or cord blood. For some applications, the patient's own cells are engrafted. However, allogeneic stem cell transplantation is now a common procedure for the treatment of bone marrow failure and haematological malignancies, such as leukaemia. Donor stem cells are used to reconstitute immune function in such patients following radiation and/or chemotherapy. In the UK, the regulatory framework put in place for bone marrow transplantation has now an extended remit, covering the use of other tissues and organs (Austin et al. 2008).

Advances in immunology research greatly increased the utility of bone marrow transplantation, allowing allograft donors to be screened for the best match in order to prevent rejection and graft-versus-host disease (Perry & Linch 1996). It is worth remembering that organ transplantation programmes have also depended on an understanding of immune rejection, and drugs are available to provide effective long-term immunosuppression for recipients of donor organs. Thus, while it is obviously desirable for new stem cell treatments to involve the patient's own cells, it is certainly not essential.

Two major advantages of haemopoietic stem cell therapy are that there is no need to expand the cells in culture or to reconstitute a multicellular tissue architecture prior to transplantation. These hurdles have been overcome to generate cultured epidermis to provide autologous grafts for patients with full-thickness wounds, such as third-degree burns. Proof-of-principle was established in the mid-1970s, with clinical and commercial applications following rapidly (Green 2008). Using a similar approach, limbal stem cells have been used successfully to restore vision in patients suffering from chemical destruction of the cornea (De Luca et al. 2006).

Ex vivo expansion of human epidermal and corneal stem cells frequently involves culture on a feeder layer of mouse fibroblastic cells in medium containing bovine serum. While it would obviously be preferable to avoid animal products, there has been no evidence over the past 30 years that exposure to them has had adverse effects on patients receiving the grafts. The ongoing challenges posed by epithelial stem cell treatments include improved functionality of the graft (e.g. through generation of epidermal hair follicles) and improved surfaces on which to culture the cells and apply them to the patients. The need to optimize stem cell delivery is leading to close interactions between the stem cell community and bioengineers. In a recent example, a patient's trachea was repaired by transplanting a new tissue constructed in culture from donor decellularized trachea seeded with the patient's own bone marrow cells that had been differentiated into cartilage cells (Macchiarini et al. 2008).

Whereas haemopoietic stem cell therapies are widely available, treatments involving cultured epidermis and cornea are not. In countries where cultured epithelial grafts are available, the number of potential patients is relatively small and the treatment costly. Commercial organizations that sell cultured epidermis for grafting have found that it is not particularly profitable, while in countries with publicly funded healthcare the need to set up a dedicated laboratory to generate the grafts tends to make the financial costbenefit ratio too high (Green 2008).

Clinical studies over the last 10 years suggest that stem cell transplantation also has potential as a therapy for neurodegenerative diseases. Clinical trials have involved grafting brain tissue from aborted foetuses into patients with Parkinson's disease and Huntington's disease (Dunnett et al. 2001; Wright & Barker 2007). While some successes have been noted, the outcomes have not been uniform and further clinical trials will involve more refined patient selection, in an attempt to predict who will benefit and who will not. Obviously, aside from the opposition in many quarters to using foetal material, there are practical challenges associated with availability and uniformity of the grafted cells and so therapies with pure populations of stem cells are an important, and achievable (Conti et al. 2005; Lowell et al. 2006), goal.

No consideration of currently available stem cell therapies is complete without reference to gene therapy. Here, there have been some major achievements, including the successful treatment of children with X-linked severe combined immunodeficiency. However, the entire gene therapy field stalled when several of the children developed leukaemia as a result of integration of the therapeutic retroviral vector close to the LMO2 oncogene locus (Gaspar & Thrasher 2005; Pike-Overzet et al. 2007). Clinical trials have since restarted, and in an interesting example of combined gene/stem cell therapy, a patient with an epidermal blistering disorder received an autologous graft of cultured epidermis in which the defective gene had been corrected ex vivo (Mavilio et al. 2006).

These are just some examples of treatments involving stem cells that are already in the clinic. They show how the field of stem cell transplantation is interlinked with the fields of gene therapy and bioengineering, and how it has benefited from progress in other fields, such as immunology. Stem cells undoubtedly offer tremendous potential to treat many human diseases and to repair tissue damage resulting from injury or ageing. The danger, of course, lies in the potentially lethal cocktail of desperate patients, enthusiastic scientists, ambitious clinicians and commercial pressures (Lau et al. 2008). Internationally agreed, and enforced, regulations are essential in order to protect patients from the dangers of stem cell tourism, whereby treatments that have not been approved in one country are freely available in another (Hyun et al. 2008).

Three questions in stem cell research are being hotly pursued at present. What are the core genetic and epigenetic regulators of stem cells? What are the extrinsic, environmental factors that influence stem cell renewal and differentiation? And how can the answers to the first two questions be harnessed for clinical benefit?

Considerable progress has already been made in defining the transcriptional circuitry and epigenetic modifications associated with pluripotency (Jaenisch & Young 2008). This research area is moving very rapidly as a result of tremendous advances in DNA sequencing technology, bioinformatics and computational biology. Chromatin immunoprecipitation combined with microarray hybridization or DNA sequencing (Mathur et al. 2008) is being used to identify transcription factor-binding sites, and bioinformatics techniques have been developed to allow integration of data obtained by the different approaches. It is clear that pluripotency is also subject to complex epigenetic regulation, and high throughput genome-scale DNA methylation profiling has been developed for epigenetic profiling of ES cells and other cell types (Meissner et al. 2008).

Oct4, Nanog and Sox2 are core transcription factors that maintain pluripotency of ES cells. These factors bind to their own promoters, forming an autoregulatory loop. They occupy overlapping sets of target genes, one set being actively expressed and the other, comprising genes that positively regulate lineage selection, being actively silenced (Jaenisch & Young 2008; Mathur et al. 2008; Silva & Smith 2008). Nanog stabilizes pluripotency by limiting the frequency with which cells commit to differentiation (Chambers et al. 2007; Torres & Watt 2008). The core pluripotency transcription factors also regulate, again positively and negatively, the microRNAs that are involved in controlling ES cell self-renewal and differentiation (Marson et al. 2008).

As the basic mechanisms that maintain the pluripotent state of ES cells are delineated, there is considerable interest in understanding how pluripotency is re-established in adult stem cells. It appears that some cell types are more readily reprogrammed to iPS cells than others (Aasen et al. 2008; Aoi et al. 2008), and it is interesting to speculate that this reflects differences in endogenous expression of the genes required for reprogramming or in responsiveness to overexpression of those genes (Hochedlinger et al. 2005; Markoulaki et al. 2009). Another emerging area of investigation is the relationship between the epigenome of pluripotent stem cells and cancer cells (Meissner et al. 2008).

Initial attempts at defining stemness by comparing the transcriptional profiles of ES cells, neural and haemopoietic stem cells (Ivanova et al. 2002; Ramalho-Santos et al. 2002) have paved the way for more refined comparisons. For example, by comparing the gene expression profiles of adult neural stem cells, ES-derived and iPS-derived neural stem cells and brain tumour stem cells, it should be possible both to validate the use of ES-derived stem cells for brain repair and to establish the cell of origin of brain tumour initiating cells. Furthermore, it is anticipated that new therapeutic targets will be identified from molecular profiling studies of different stem cell populations.

As gene expression profiling becomes more sophisticated, the question of what is a stem cell? can be addressed in new ways. Several studies have used single cell expression microarrays to identify new stem cell markers (Jensen & Watt 2006). Stem cells are well known to exhibit different proliferative and differentiation properties in culture, during tissue injury and in normal tissue homeostasis, raising the question of which elements of the stem cell phenotype are hard-wired versus a response to environmental conditions.

One of the growing trends in stem cell research is the contribution of mathematical modelling. This is illustrated in the concept of transcriptional noise: the hypothesis that intercellular variability is a manifestation of noise in gene expression levels, rather than stable phenotypic variation (Chang et al. 2008). Studies with clonal populations of haemopoietic progenitor cells have shown that slow fluctuations in protein levels can produce cellular heterogeneity that is sufficient to affect whether a given cell will differentiate along the myeloid or erythroid lineage (Chang et al. 2008). Mathematical approaches are also used increasingly to model observed differences in cell behaviour in vivo. In studies of adult mouse interfollicular epidermis, it is observed that cells can divide to produce two undifferentiated cells, two differentiated cells or one of each (figure3); it turns out that this can be explained in terms of the stochastic behaviour of a single population of cells rather than by invoking the existence of discrete types of stem and progenitor cell (Clayton et al. 2007).

The stem cell niche. Stem cells (S) are shown dividing symmetrically to produce two stem cells (1) or two differentiated cells (D) (2), or undergoing asymmetric division to produce one stem cell and one differentiated cell (3). Under some circumstances, a differentiated cell can re-enter the niche and become a stem cell (4). Different components of the stem cell niche are illustrated: extracellular matrix (ECM), cells in close proximity to stem cells (niche cells), secreted factors (such as growth factors) and physical factors (such as oxygen tension, stiffness and stretch).

There is strong evidence that the behaviour of stem cells is strongly affected by their local environment or niche (figure3). Some aspects of the stem cell environment that are known to influence self-renewal and stem cell fate are adhesion to extracellular matrix proteins, direct contact with neighbouring cells, exposure to secreted factors and physical factors, such as oxygen tension and sheer stress (Watt & Hogan 2000; Morrison & Spradling 2008). It is important to identify the environmental signals that control stem cell expansion and differentiation in order to harness those signals to optimize delivery of stem cell therapies.

Considerable progress has been made in directing ES cells to differentiate along specific lineages in vitro (Conti et al. 2005; Lowell et al. 2006; Izumi et al. 2007) and there are many in vitro and murine models of lineage selection by adult tissue stem cells (e.g. Watt & Collins 2008). It is clear that in many contexts the Erk and Akt pathways are key regulators of cell proliferation and survival, while pathways that were originally defined through their effects in embryonic development, such as Wnt, Notch and Shh, are reused in adult tissues to influence stem cell renewal and lineage selection. Furthermore, these core pathways are frequently deregulated in cancer (Reya et al. 2001; Watt & Collins 2008). In investigating how differentiation is controlled, it is not only the signalling pathways themselves that need to be considered, but also the timing, level and duration of a particular signal, as these variables profoundly influence cellular responses (Silva-Vargas et al. 2005). A further issue is the extent to which directed ES cell differentiation in vitro recapitulates the events that occur during normal embryogenesis and whether this affects the functionality of the differentiated cells (Izumi et al. 2007).

For a more complete definition of the stem cell niche, researchers are taking two opposite and complementary approaches: recreating the niche in vitro at the single cell level and observing stem cells in vivo. In vivo tracking of cells is possible because of advances in high-resolution confocal microscopy and two-photon imaging, which have greatly increased the sensitivity of detecting cells and the depth of the tissue at which they can be observed. Studies of green fluorescent protein-labelled haemopoietic stem cells have shown that their relationship with the bone marrow niche, comprising blood vessels, osteoblasts and the inner bone surface, differs in normal, irradiated and c-Kit-receptor-deficient mice (Lo Celso et al. 2009; Xie et al. 2009). In a different approach, in vivo bioluminescence imaging of luciferase-tagged muscle stem cells has been used to reveal their role in muscle repair in a way that is impossible when relying on retrospective analysis of fixed tissue (Sacco et al. 2008).

The advantage of recreating the stem cell niche in vitro is that it is possible to precisely control individual aspects of the niche and measure responses at the single cell level. Artificial niches are constructed by plating cells on micropatterned surfaces or capturing them in three-dimensional hydrogel matrices. In this way, parameters such as cell spreading and substrate mechanics can be precisely controlled (Watt et al. 1988; Thry et al. 2005; Chen 2008). Cells can be exposed to specific combinations of soluble factors or to tethered recombinant adhesive proteins. Cell behaviour can be monitored in real time by time-lapse microscopy, and activation of specific signalling pathways can be viewed using fluorescence resonance energy transfer probes and fluorescent reporters of transcriptional activity. It is also possible to recover cells from the in vitro environment, transplant them in vivo and monitor their subsequent behaviour. One of the exciting aspects of the reductionist approach to studying the niche is that it is highly interdisciplinary, bringing together stem cell researchers and bioengineers, and also offering opportunities for interactions with chemists, physicists and materials scientists.

Almost every day there are reports in the media of new stem cell therapies. There is no doubt that stem cells have the potential to treat many human afflictions, including ageing, cancer, diabetes, blindness and neurodegeneration. Nevertheless, it is essential to be realistic about the time and steps required to take new therapies into the clinic: it is exciting to be able to induce ES cells to differentiate into cardiomyocytes in a culture dish, but that is only one very small step towards effecting cardiac repair. The overriding concerns for any new treatment are the same: efficacy, safety and affordability.

In January 2009, the US Food and Drug Administration approved the first clinical trial involving human ES cells, just over 10 years after they were first isolated. In this trial, the safety of ES cell-derived oligodendrocytes in repair of spinal cord injury will be evaluated (http://www.geron.com). There are a large number of human ES cell lines now in existence and banking of clinical grade cells is underway, offering the opportunity for optimal immunological matching of donors and recipients. Nevertheless, one of the attractions of transplanting iPS cells is that the patient's own cells can be used, obviating the need for immunosuppression. Discovering how the pluripotent state can be efficiently and stably induced and maintained by treating cells with pharmacologically active compounds rather than by genetic manipulation is an important goal (Silva et al. 2008).

An alternative strategy to stem cell transplantation is to stimulate a patient's endogenous stem cells to divide or differentiate, as happens naturally during skin wound healing. It has recently been shown that pancreatic exocrine cells in adult mice can be reprogrammed to become functional, insulin-producing beta cells by expression of transcription factors that regulate pancreatic development (Zhou et al. 2008). The idea of repairing tissue through a process of cellular reprogramming in situ is an attractive paradigm to be explored further.

A range of biomaterials are already in clinical use for tissue repair, in particular to repair defects in cartilage and bone (Kamitakahara et al. 2008). These can be considered as practical applications of our knowledge of the stem cell microenvironment. Advances in tissue engineering and materials science offer new opportunities to manipulate the stem niche and either facilitate expansion/differentiation of endogenous stem cells or deliver exogenous cells. Resorbable scaffolds can be exploited for controlled delivery and release of small molecules, growth factors and peptides. Conversely, scaffolds can be designed that are able to capture unwanted tissue debris that might impede repair. Hydrogels that can undergo controlled solgel transitions could be used to release stem cells once they have integrated within the target tissue.

Although most of the new clinical applications of stem cells have a long lead time, applications of stem cells in drug discovery are available immediately. Adult tissue stem cells, ES cells and iPS cells can all be used to screen for compounds that stimulate self-renewal or promote specific differentiation programmes. Finding drugs that selectively target cancer stem cells offers the potential to develop cancer treatments that are not only more effective, but also cause less collateral damage to the patient's normal tissues than drugs currently in use. In addition, patient-specific iPS cells provide a new tool to identify underlying disease mechanisms. Thus stem cell-based assays are already enhancing drug discovery efforts.

Amid all the hype surrounding stem cells, there are strong grounds for believing that over the next 50 years our understanding of stem cells will revolutionize medicine. One of the most exciting aspects of working in the stem cell field is that it is truly multidisciplinary and translational. It brings together biologists, clinicians and researchers across the physical sciences and mathematics, and it fosters partnerships between academics and the biotech and pharmaceutical industries. In contrast to the golden era of developmental biology, one of stem cell research's defining characteristics is the motivation to benefit human health.

We thank all members of our lab, past and present, for their energy, fearlessness and intellectual curiosity in the pursuit of stem cells. We are grateful to Cancer Research UK, the Wellcome Trust, MRC and European Union for financial support and to members of the Cambridge Stem Cell Initiative for sharing their ideas.

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The therapeutic potential of stem cells - PMC

Cystic fibrosis (CF) is a hereditary disorder characterized by mutations in the CFTR gene, leading to impaired chloride ion transport and subsequent thickening of mucus in various organs, particularly the lungs. Despite significant progress in CF management, current treatments focus mainly on symptom relief and do not address the underlying genetic defects. Stem cell and gene therapies present promising avenues for tackling CF at its root cause. Stem cells, including embryonic, induced pluripotent, mesenchymal, hematopoietic, and lung progenitor cells, offer regenerative potential by differentiating into specialized cells and modulating immune responses. Similarly, gene therapy aims to correct CFTR gene mutations by delivering functional copies of the gene into affected cells. Various approaches, such as viral and nonviral vectors, gene editing with CRISPR-Cas9, small interfering RNA (siRNA) therapy, and mRNA therapy, are being explored to achieve gene correction. Despite their potential, challenges such as safety concerns, ethical considerations, delivery system optimization, and long-term efficacy remain. This review provides a comprehensive overview of the current understanding of CF pathophysiology, the rationale for exploring stem cell and gene therapies, the types of therapies available, their mechanisms of action, and the challenges and future directions in the field. By addressing these challenges, stem cell and gene therapies hold promise for transforming CF management and improving the quality of life of affected individuals.

Keywords: CFTR gene; CRISPR-Cas9; cystic fibrosis; delivery system; gene therapy; genetic disorder; mesenchymal stem cells; stem cell therapy; vectors.

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Advancing the Battle against Cystic Fibrosis: Stem Cell and Gene ...

Scientists have observed that some genetic variations in microglia show a strong correlation with an increased risk of Alzheimers disease. One such correlation involves a gene called TREM2, which plays an essential role in in how microglia detect and address neurodegeneration. Certain genetic variants of TREM2 are among the strongest genetic risk factors for Alzheimers disease, Wernig said.

The data are convincing that microglial dysfunction can cause neurodegeneration in the brain, so it makes sense that restoring defective microglial function might be a way to fight neurodegeneration in Alzheimers disease, he added.

In the study, mice with a defective TREM2 gene received hematopoietic stem and progenitor cell transplants from mice with normal TREM2 function. The researchers found that the transplanted cells reconstituted the blood system and that some of them efficiently incorporated into the recipients brains and became cells that looked and behaved like microglia.

We showed that most of the brains original microglia were replaced by healthy cells, which led to a restoration of normal TREM2 activity, Wernig said.

Next, they investigated whether the restored TREM2 activity was enough to improve the brain health of the TREM2-deficient mice. Indeed, in the transplanted mice we saw a clear reduction in the deposits of amyloid plaques normally seen TREM2-deficient mice, Wernig said. They were also able to show a restoration of microglial function and reduction of other disease markers, indicating that functional restoration of this one gene had widespread positive effects.

Wernig and colleagues said they could transplant cells engineered to have supercharged TREM2 activity that may have an even greater effect.

They caution, however, that the microglia that formed from the transplanted cells were slightly different from the natural microglia in mouse brains. These differences might in some way have their own detrimental effect, Wernig said. We have to look at that very carefully.

In addition, the current procedure would be highly risky if it were developed for human therapy because transplantation of blood stem cells requires the recipient to undergo a highly toxic chemotherapy or radiation treatment to kill off native blood stem cells. However, many researchers, including some at the Institute for Stem Cell Biology and Regenerative Medicine, are developing less toxic methods of preconditioning patients for stem cell transplants. A brain cell therapy could then piggyback on such improved and safer transplantation methods.

The work was supported by the Kleberg Foundation, the Emerson Collective, a Howard Hughes Medical Institute faculty scholar award, a New York Stem Cell Foundation Druckenmiller award, a postdoctoral overseas training fellowship from the National Research Foundation of Korea and the German Research Foundation.

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Researchers find success with stem cell therapy in mice model of ...

May 23, 2024

Mayo Clinic researchers have demonstrated the safety and potential benefit of stem cell regenerative medicine therapy for patients with subacute and chronic spinal cord injury.

The results of the phase 1 Clinical Trial of Autologous Adipose-Derived Mesenchymal Stem Cells in the Treatment of Paralysis Due to Traumatic Spinal Cord Injury, known as CELLTOP, were published in Nature Communications.

Illustration shows the process of fat harvest via biopsy, adipose-derived mesenchymal stem cells (AD-MSC) preparation and administration of treatment.

All trial participants had experienced traumatic spinal injury classified as grade A or B on the American Spinal Injury Association Impairment Scale (AIS). Stem cell treatment was initiated on average 11 months after injury. Participants were evaluated over a two-year period.

Key findings:

As reported earlier in Mayo Clinic Proceedings, the first participant in the phase 1 trial was a superresponder who, after stem cell therapy, saw significant improvements in the function of his upper and lower extremities.

"Future research may show whether stem cells in combination with other therapies could be part of a new paradigm of treatment to improve outcomes for patients," says Mohamad Bydon, M.D., a neurosurgeon at Mayo Clinic in Rochester, Minnesota, and the first author of both studies. "Not every patient who receives stem cell treatment is going to be a superresponder. One objective in our future studies is to delineate the optimal treatment protocols and understand why patients respond differently."

Dr. Bydon notes that stem cells' mechanism of action isn't fully understood. The researchers are analyzing changes in participants' MRI and cerebrospinal fluid to identify avenues for potential regeneration. Work is also underway on a larger, controlled trial of stem cell regenerative therapy.

"For years, treatment of spinal cord injury has been limited to stabilization surgery and physical therapy," Dr. Bydon says. "Many historical textbooks state that this condition does not improve. We have seen findings in recent years that challenge prior assumptions. This research is a step forward toward the ultimate goal of improving treatments for patients."

Bydon M, et al. Intrathecal delivery of adipose-derived mesenchymal stem cells in traumatic spinal cord injury: Phase I trial. Nature Communications. 2024;15:2201.

Bydon M, et al. CELLTOP clinical trial: First report from a phase I trial of autologous adipose tissue-derived mesenchymal stem cells in the treatment of paralysis due to traumatic spinal cord injury. Mayo Clinic Proceedings. 2020;95:406.

Refer a patient to Mayo Clinic.

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Study finds stem cell therapy is safe and may benefit people with ...

Stem Cell Therapy Market Size to Hit USD 48.89 Billion by 2033  GlobeNewswire

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Stem Cell Therapy Market Size to Hit USD 48.89 Billion by 2033 - GlobeNewswire

Eves dream to walk: Family raising vital funds for two-year-olds stem cell therapy  Her.ie

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Eves dream to walk: Family raising vital funds for two-year-olds stem cell therapy - Her.ie

Chronic obstructive pulmonary diseases (COPD) is a heterogeneous lung disease that has high morbidity and mortality worldwide, and is an increasing economic and social burden [1, 2]. The recent burden of obstructive lung disease study and other large-scale epidemiological studies estimated that the global prevalence of COPD is 10.3% [3]. Another recent study estimated that the increasing prevalence of smoking in low- and middle-income countries and aging populations in high-income countries will lead to more than 5.4million annual deaths from COPD and related conditions by 2060 [4]. Long-term exposure to cigarette smoke, harmful chemicals, smoke from burning fuels, and 1-antitrypsin deficiency are the main causes of COPD [5] (Fig.1). As COPD progresses, patients experience irreversible obstruction of airflow, hypersecretion of mucus, destruction of alveolar wall, and proliferation of airway smooth muscle cells due to the chronic bronchitis and/or emphysema [6].

Risk factors and pathogenesis of COPD. By figdraw (www.figdraw.com)

Chronic bronchitis is characterized by non-specific inflammation of the mucosa and surrounding tissues of the trachea and bronchi, and is triggered by an infection or non-infectious factors (allergy, oxidative stress) in which the major pathological change is damage of epithelia in the central airway [7]. Prolonged inflammatory stimulation causes proliferation of bronchial mucosal cells and mucosal hypertrophy, resulting in obstruction of the airways and decreased ventilation. The excessive secretion of mucus and infiltration of inflammatory cells leads to the accumulation of mucus in the bronchial lumen, further aggravating the narrowing and obstruction of airways [8]. Emphysema is characterised by dilatation and destruction of lung tissue beyond the terminal fine bronchioles. Its manifestations are thinning of the alveolar walls; enlargement, rupture, or formation of large blisters in the alveolar cavities; reduced blood supply; and destruction of the elastic fibrous network [9].

COPD is, therefore, a multifactorial disease with a complex pathogenesis, and many studies have focused on the accumulation of inflammatory cells, the imbalance of protease/antiprotease activity, and oxidative stress [5, 10, 11] (Fig.1). There is evidence that cigarette smoke and other inhaled particulates stimulate epithelial cells to produce reactive oxygen species, and that this induces inflammatory cells to infiltrate the periphery of the airways, leading to an imbalance of protease/antiprotease activity [12,13,14]. Elastin is a major protein in the connective tissue of the lung parenchyma, and an imbalance between proteases and antiproteases can decrease the level of elastin, leading to lung hyperinflation, lung dilatation, and loss of lung elasticity, culminating emphysema [15,16,17]. During disease progression, COPD patients experience a gradual decline in lung capacity which initially limits their ability to exercise, and eventually becomes disabling [18]. Although chronic bronchitis and emphysema are the predominant clinical phenotypes of COPD, these patients may also present with several other complications, such as airway hyperresponsiveness, hyperimmune response, asthma, and other lung diseases [19]. The many symptoms of COPD are responsible for its huge personal, social, and economic burden. COPD is incurable, but the clinical management of symptoms includes smoking cessation, vaccinations for respiratory pathogens, various medications (especially bronchodilators and steroids), oxygen therapy, and pulmonary rehabilitation [8]. These treatments aim to control the symptoms, decrease inflammation, and improve functional capacity. The use of anti-inflammatory drugs and bronchodilators reduces the severity of symptoms and improves patient quality-of-life, but these treatments only relieve symptoms and do not block or reverse lung damage [20]. Although smoking cessation and long-term oxygen therapy are relatively effective, they do not halt the underlying pathology of increased inflammation, apoptosis, and oxidative stress [21,22,23]. There are currently no treatments for the irreversible loss of lung function and incompletely reversible limitation of expiratory airflow, so there is an urgent need to develop new treatments that can repair the damaged lung tissues of patients with COPD.

Stem cells are undifferentiated cells with the capacity for self-renewal and multispectral differentiation, and cell-based tissue reconstruction using stem cells is an important part of regenerative medicine [24]. Cellular therapeutic approaches may provide new treatment options for COPD in the future. The unique properties of stem cells are that they can promote tissue repair and regeneration by replacing damaged cells, modulate immune responses, reduce inflammation, and promote tissue homeostasis [25]. Thus, a promising general approach for using stem cells to treat COPD is to harness their capacity for differentiation by stimulating them to regenerate lung parenchymal cells and airways, and/or to promote stem/progenitor cell differentiation of epithelial cells to restore the balance between proliferation and apoptosis.

The most important function of stem cells is their maintenance of cellular regeneration, in that they can differentiate into at least one type of highly mature cell. This means that stem cells have great potential for use in tissue repair if they can be promoted to replace diseased and damaged tissues [26]. There are three general types of stem cells: embryonic stem cells (ESCs), induced pluripotent stem cells (iPSCs) and adult stem cells (MSCs and several others) [27]. ESCs and iPSCs have the potential for unlimited proliferation and differentiation into all germ layers, and therefore have great potential for treatment of refractory diseases and injuries [28]. Adult stem cells are immature cells in adult tissues that are in a resting state, which function as a reservoir for self-renewal because of their ability to differentiate into highly specialised cells and repair tissue following injury [29]. There are two types of adult stem cells that affect airway function in COPD: mesenchymal stem cells (MSCs) and lung progenitor/stem cells.

ESCs are pluripotent stem cells isolated from blastomeres that can differentiate into all types of cells, making them highly valuable in regenerative medicine [30, 31]. Since 1998, when hESCs were first cultured and differentiated, many studies of drug-based therapies using ESCs have examined their use for treatment of spinal cord injury, macular degeneration, type 1 diabetes, heart failure, and other conditions [32]. ESCs also have potential for treatment of lung diseases, and studies have demonstrated that hESCs and mouse ESCs can differentiate into type II alveolar epithelial cells (ATIICs) using controlled culture conditions [33, 34]. Previous studies have used established transfection and culture procedures in a laboratory setting to develop genetically screened and highly purified hESC-ATIICs that have characteristics typical of ATIICs, including formation of lamellipodia, expression of surfactant protein, and the ability to proliferate and differentiate into type I alveolar epithelial cells (ATICs) [35]. Previous research demonstrated that ATIICs derived from hESCs can be engrafted into the lungs of mice with acute lung injury, and that this treatment inhibited or reversed fibrotic changes induced by bleomycin [36]. Although hESC-ATIICs have potential as a cellular source for treatments of distal lung diseases, there are many concerns regarding the safety of cell transplantation from differentiated ESCs. Methods such as the formation of embryoid bodies or coculture of these cells with lung mesenchyme have only produced small numbers of ATIICs [33, 34, 37, 38]. Another important concern is that the presence of pluripotent cells within a mixed population of ESC-derived cells may lead to formation of a teratoma, making this approach unsuitable for transplantation into the lungs. There are also serious ethical concerns, because the acquisition of hESCs necessitates the destruction of embryos.

A disadvantage of allogeneic transplantation of hESCs is that they can elicit a strong immune reaction; this necessitates intensive immunosuppressive treatment to prevent rejection, which increases the risk of an opportunistic infection and tumor development [39, 40].Many studies have therefore attempted to produce personalized hESCs to prevent immune rejection. For example, the implementation of alternative targeted methods, such as b2-microglobulin-deficient hESCs, greatly decreases the susceptibility of stem cells to recognition by CD8+T cells. This approach can potentially provide a sustainable reservoir of cells for tissue regeneration without the need for matching of human leukocyte antigens (HLAs) [41]. A clinical case report showed that treatment with hESCs decreased symptoms in a patient with emphysema [42]. Although preclinical studies showed that ESCs can regenerate lung structures, the potential for teratoma formation, immune rejection, and ethical issues have prevented the development of clinical trials that use hESCs for treatment of COPD [43]. To address these ethical concerns and technical limitations, researchers have instead taken alternative approaches that use adult stem cells or iPSCs.

iPSCs, which are generated by reprogramming adult somatic cells (e.g., skin cells) into a pluripotent state, have great potential for use in regenerative medicine. In 2006, the Yamanaka team reported the successful reprogramming of differentiated somatic cells into a cell type resembling embryonic stem cells by introduction of specific transcription factors. Since then, many researchers have utilized these cells as a source for regeneration in studies of disease-specific cells in different animal models, drug screening, and development of cell-based therapies [44]. hiPSCs are a promising source of therapeutic cells because they have totipotency similar to hESCs, they can differentiate into a variety of cell types, they do not induce strong immune responses, and there are no serious ethical concerns regarding their use [45, 46]. Extracting somatic cells from a patient and reprogramming them into iPSCs greatly reduces the risk of immune rejection after transplantation and improves the feasibility and safety of cell transplantation. Controlled differentiation programs allow iPSCs to develop into specific cell lineages, so they can replace the damaged cells that are in the tissues or organs affected by specific diseases [47]. There is also evidence that hiPSCs undergo targeted differentiation to produce alveolar epithelial cell type 2 (iAT2) cells, and that hiPSCs derived from distal lung cells can be implanted into the lungs, where they contribute to the formation of functional distal lung units and slow the progression of emphysema [48, 49].

Although iPSCs appear to have great potential, several limitations need to be addressed. One concern is reprogramming efficiency, because the process is slow and inefficient, even though it is reproducible [50]. Although recent advances showed that iPSCs can be amplified in scalable suspension cultures, and that large quantities of human iPSCs can be produced in a fully controlled bioreactor [51], further technological improvements are needed for producing clinical-grade iPSCs. Another important consideration is the risk of genetic and epigenetic abnormalities during reprogramming, because reprogrammed iPSCs may have different genomic instabilities that lead to development of a malignancy. In particular, the random integration of multiple copies of a transgene into the host genome during the overexpression of reprogramming factors can lead to significant phenotypic variability, and to potential disruptions in the function of critical genes in the generated iPSCs [44]. An even greater concern is that genomic alterations caused by the integration of transcription factor genes into the genome could lead to tumorigenicity [31]. There are also immunological concerns associated with cell therapies that utilize iPSCs [52]. Given the continuing improvements in the protocols used for differentiation and the availability of better techniques for enrichment, purification, and analysis, it may soon be possible to eliminate undifferentiated cells from the differentiated cell products. The risk of teratoma formation appears to be very low when using highly enriched cell products [53].

iPSCs have several potential uses in regenerative medicine studies: directly for cell transplantation, as a source of differentiated cells, and in model systems to explore the role of epigenetic reprogramming in abnormally functioning cells. Most research has focused on the last two methods [54]. Although clinical trials are increasingly using hESC or hiPSC derivatives to repair organs and treat diseases, our search of ClinicalTrials.gov identified 96 clinical trials using iPSCs, but only 5 that were related to lung and respiratory diseases, and none that examined iPSCs as a treatment for lung diseases in [53]. The challenges associated with vector integration, suboptimal efficiency in generation of hiPSCs, and the demand for different types of transplantable cells led to the development of a novel vector incorporating a Tet-On inducible gene expression system, an ATII C-specific NEOR transgene, and loxP target sequences. This innovative approach enabled the successful generation of hiPSC-ATIICs whose ultrastructural characteristics and functional properties were similar to those of ATIICs [55]. Similar hESCs-ATIICs and hiPSCs-ATIICs have significant promise for in vivo transplantation, differentiation into ATICs for alveolar regeneration following bleomycin-induced injury, and improving the function of damaged lungs [44]. However, the process of obtaining tissue-specific cells from iPSCs requires labor-intensive reprogramming and directed differentiation. It is imperative to develop expedited approaches, such as direct reprogramming of one somatic cell type into another cell type, as a more effective strategy for generation of iPSCs [56].

The adult lung is a complex organ consisting of many types of cells that are distributed throughout the respiratory tract, and each region is characterized by its own distinct population of epithelial cells. Stem/progenitor cells from various epithelial lineages within the lung function in lung development, tissue maintenance, and repair following injury [57]. The maintenance of a precise equilibrium between each region-specific type of stem cell and its specific microenvironments is essential for the preservation of normal lung function and airway integrity during normal conditions and during the repair of lung damage. The region-specific epithelial stem/progenitor cells in the lungs of human adults and mice consist of basal cells, secretory cells, and mucous cells in the proximal airway submucosal glands; variant secretory cells in the small bronchi; bronchoalveolar duct junction stem cells; and a subset of ATIICs in the alveolar sac [58, 59]. Lung progenitor/stem cells have the capacity to undergo proliferation and differentiation following lung injury in order to restore damaged cell populations and uphold the typical physiological functioning of the lung [25]. Among these, basal cells are a key type of epithelial stem cell, because they play a crucial role in maintaining environmental homeostasis and promoting the repair of epithelial in the proximal airways. Bronchioalveolar stem cells are essential for repairing damage to the small bronchi and alveolar cells, and they play a crucial role in maintaining stability of the lung environment. Bronchoalveolar stem cells exhibit a diverse differentiation response to various forms of injury, giving rise to cell types including club cells and ciliated cells. Within the gas exchange region of the adult lung, a subset of ATIICs are stem/progenitor cells for ATICs, and ATIICs function as stem cells by maintaining epithelial homeostasis during normal conditions and in response to injury [57]. Through lineage tracing experiments, it has been demonstrated that AT2 cells, serving as stem/progenitor cells, possess the ability to self-renew and undergo transdifferentiation into AT1 cells [25]. The ability of the lung to self-repair following injury requires activation of stem cells and progenitor cells within each respiratory alveolus.

Individuals with COPD experience oxidative stress and disruption of the equilibrium between self-renewal and differentiation of stem/progenitor cells, and this prevents the regeneration of lung tissue [9]. Thus, there is an urgent need for more basic studies and clinical studies that examine the use of lung stem/progenitor cells for the management of COPD and other pulmonary ailments. We searched for clinical trials using bronchial basal cells for the treatment of COPD, of which only one had relevant results published. In this study, autologous P63+lung progenitor cells were transplanted into COPD subjects, and the results suggest that transplantation of cultured P63 lung progenitor cells is safe and may represent a potential therapeutic strategy for COPD (NCT03188627) [60]. There has been great interest in the potential use of exogenous stem/progenitor cells to regenerate or enhance lung repair in patients with COPD, but significant hurdles must be overcome. Firstly, it is difficult to obtain sufficient quantities of autologous stem cells. Secondly, there is insufficient information regarding the identification and characterization of defined lung stem/progenitor cell subpopulations. Lastly, most current cell implantation techniques are inefficient and lack long-term efficacy [61]. Although several clinical studies have examined the use of endothelial progenitor cells and basal epithelial cells from the bronchus for investigational interventions, direct implantation of cells required for airway function is a potential approach for regenerating damaged lung tissue, and further studies using animal models are needed.

MSCs are a subset of stem cells that exhibit morphological similarities to fibroblasts and are found within the connective tissues of nearly all organs. In 2006, the International Society for Cellular Therapy (ISCT) delineated specific criteria for the identification of MSCs [62]: (1) adherence to plastic surfaces under standard culture conditions, (2) expression of a defined set of surface antigens, and (3) the capacity to differentiate into osteoblasts, chondrocytes, and adipocytes. MSCs exhibit low immunogenicity due to their lack of MHC-II antigen expression. Additionally, MSCs possess immunosuppressive capabilities, forming the foundation for their application in cell therapy. Extensive research utilizing animal models has been undertaken to evaluate the efficacy of MSCs and to progressively elucidate the mechanisms underlying MSC-based tissue repair therapies. Consequently, MSCs are emerging as a potent therapeutic tool for a wide range of diseases. Further investigation into their biological properties is essential to establish the theoretical groundwork necessary for clinical applications. MSCs will be elaborated in more detail below.

Stem cells, with their unique characteristics of self-renewal and differentiation, are the focus of regenerative medicine. These different types of stem cells have distinct biological characteristics and differentiation potentials, and the best is still controversial. Advantages and disadvantages of different types of stem cells are briefly summarized in Fig.2. Embryonic stem cells are highly undifferentiated cells that can be induced to differentiate into almost all cell types of the organism, but cannot develop into complete individuals. ESCs have long been expected to be the most promising cell source in the regenerative medicine. Ethical controversies, tumour formation and immune rejection make the use of embryonic stem cells a huge obstacle. As a replacement for ESCs, iPSCs are not beset with any serious ethical issues, but oncogenicity and cellular deterioration due to incomplete reprogramming remain challenges. Lung progenitor/stem cells are adult stem cells with the advantages of having low tumourigenicity, being suitable for autologous transplantation, and being less ethically problematic. However, lung progenitor/stem cells are rare and difficult to isolate and characterize. All of these cells seem to have insurmountable difficulties to be used in therapy, but the discovery of mesenchymal stem cells offers new hope for the development of regenerative medicine. MSCs are not only readily available from a wide range of sources, with low immunogenicity to avoid rejection, but also perfectly avoid the major drawbacks of other stem cells through immune modulation and suppression of inflammatory responses, making them an ideal cell choice for use in regenerative medicine and therapeutic areas. Conversely, most small-scale clinical trials in regenerative medicine that used MSCs, which are considered non-pluripotent, have not reported significant safety concerns [63]. Despite these advantages, the heterogeneity of MSCs poses problems such as unstable differentiation efficiency and susceptibility to senescence. Extensive and ongoing research is underway to ensure that MSCs are safe and effective in the long term. The future strategies may include: pre-treating stem cells prior to transplantation, or combining stem cell therapies with other therapeutic modalities (e.g., pharmacological interventions or gene therapy) to enhance their therapeutic effects; and developing biomaterials and scaffolds that are structurally similar to tissues and provide structural support that promotes stem cell survival, differentiation, and integration into damaged lung tissues [47]. A number of other cellular approaches may lead to better treatments for chronic respiratory diseases, such as the use of stem/progenitor cells for lung repair, implantation of cells in small animal models, derivation of clinically relevant cell types from human iPSCs, and ex vivo lung generation using decellularized lung scaffolds [64].

Advantages and disadvantages of different types of stem cells. By figdraw (www.figdraw.com)

MSCs are multipotent stem cells that are widely used in regenerative medicine because they are easy to isolate and culture, and because they have immunomodulatory effects and immune privilege. Many preclinical and clinical trials have reported that MSCs have a favorable safety profile, making them suitable for therapeutic interventions that target chronic lung diseases because they can rapidly localize to the lungs after infusion. Compared to other types of stem cells, MSCs are easier to isolate and widely available, and their use is not prevented by ethical concerns.

MSCs are believed to reside within the connective tissue of numerous organs, such as adipose tissue, the placenta, umbilical cord blood, the umbilical cord, and dental pulp, and the most widely used types are bone marrow MSCs (BM-MSCs), adipose MSCs (AD-MSCs), and umbilical cord MSCs (UC-MSCs) [65,66,67,68,69]. A patient who receives MSCs that are obtained by isolating and culturing the patients own bone marrow or adipose tissue (autologous transplantation) have a greatly reduced the risk of immune rejection [47]. MSCs can also be manipulated in vitro and in vivo to differentiate into various cell types, including endodermal cells (pneumocytes, myoblasts, and intestinal epithelial cells) and ectodermal cells (epithelial cells and neurons), making them especially promising for treatment of lung diseases [70,71,72]. Moreover, MSCs have immunomodulatory properties, in that they can dampen inflammatory and autoimmune reactions, and they release growth factors and extracellular vesicles that can facilitate tissue regeneration and repair.

Studies of animal models of emphysema demonstrated that intravenous or intratracheal administration of lung-MSCs or BM-MSCs led to repair of lung injury; improved lung function; increased the levels of EGF, HGF, and VEGF; decreased airway inflammation; inhibited the release of proteases from inflammatory cells due to down-regulation of cyclooxygenase-2; and increased the proliferation of AT1 and AT2 cells [73,74,75]. Moreover, preclinical studies demonstrated that MSCs have significant therapeutic potential due to their anti-inflammatory, microbicidal, angiogenic, and antifibrotic properties, properties that lead to improved lung function and increased survival rates in individuals with chronic inflammatory lung diseases [76].

Despite the significant therapeutic potential of MSCs, their limitations must also be considered before they are used in patients. Firstly, because MSCs are adult stem cells they have a limited potential for differentiation, and therefore a limited ability to generate the specific types of lung cells required for complete tissue regeneration [47]. Secondly, MSCs are a heterogeneous group of cells whose therapeutic efficacy is highly variable, and there are differences in disease phenotypes and patient phenotypes. Different cell sources, therapeutic dosages, and routes of administration may all affect the function of MSCs; culture conditions, number of passages, and other factors can also affect their function. More notably, a growing number of studies showed that MSCs undergo rapid apoptosis, autophagy, or have cytotoxic effects after systemic and potentially after intratracheal administration [64]. Although a wealth of data from preclinical studies suggest that MSCs can decrease chronic inflammation, whether this also occurs in clinical settings and produces clinical benefits remains to be determined. Finally, aging and a decreased proliferative capacity can adversely affect the function and regenerative potential of MSCs. More specifically, multiple passaging of MSC cultures induces cellular senescence and decreases their potential therapeutic efficacy. Sorting and exclusion of CD26-positive MSCs from heterologous cell populations leads to enhanced cell attachment in vitro and reduces the secretion of senescence-associated cytokines, and CD26-negative MSCs had excellent efficacy in a mouse model of emphysema [77]. Therefore, strategies that rejuvenate or selectively remove senescent MSCs may increase the clinical efficacy of this approach.

To date, a relatively large number of clinical trials have demonstrated that MSCs are safe. However, despite the initial hope that implantation of MSCs would be an efficacious treatment for lung diseases, the therapeutic efficacy of MSCs in clinical settings has not yet been demonstrated. In 2011, a clinical trial of infusion of autologous bone marrow mononuclear cell for the treatment of patients with advanced emphysema was carried out in Brazil, and the follow-up results confirmed that bone marrow mononuclear cell infusion is safe (NCT01110252) [78]. Treatment of COPD with MSCs (Prochymal) showed a decrease in c-reactive protein levels, suggesting a possible improvement in the inflammatory process. However, no improvement in lung function or patient quality of life was found (NTC00683722) [79]. No adverse events were found in patients with severe emphysema who underwent lung volume reduction surgery followed by infusion of autologous MSC (NCT01306513) [80]. We found that individuals enrolled in clinical trials investigating MSC therapy for COPD primarily consisted of patients with moderate-to-severe COPD, potentially reflecting the progressive nature of the disease. While early-stage COPD symptoms can be managed with medication, individuals with end-stage respiratory failure from severe emphysema often require lung transplantation as the final treatment option. Numerous uncertainties and unresolved concerns persist regarding the effectiveness and safety of MSC therapies, underscoring the necessity for further clinical trials to address these gaps in knowledge.

MSCs of various origins, including bone marrow, adipose tissue, and the umbilical cord, are frequently used in clinical trials. Bone marrow is the primary and paramount reservoir of MSCs, and BM-MSCs have been extensively studied in preclinical studies and clinical stem cell therapy trials. Friedenstein and colleagues initially discovered MSCs in bone marrow stromal cells in the 1970s by using natural adhesion techniques. Since then, many preclinical studies have investigated the potential use of BM-MSCs for treating a range of diseases. Despite the perceived safety of bone marrow aspiration, the procedure is invasive, uncomfortable, and can lead to severe pain and infection [65]. Moreover, the restricted availability and high density of bone marrow lead to low yields of isolatable cells [81]. BM-MSCs from elderly patients have elevated expression of genes associated with aging, shorter telomeres, diminished proliferative capability, and decreased potential for differentiation [82].

There has been significant interest in AD-MSCs in recent years due to their high versatility and capability for differentiation. These cells are a distinct population of progenitor cells within adipose tissue stromal compartments that can differentiate into various types of cells, including neurons, skeletal muscle cells, and osteoblasts [65]. The pioneering work of Zuk et al. in 2001 described the isolation of AD-MSCs that had the potential for multilineage differentiation from liposuction-derived adipose tissue, demonstrating that AD-MSCs were a promising alternative to BM-MSCs [65]. Notably, adipose tissue has more MSCs than bone marrow, and the surge in clinical trials examining AD-MSCs may be because adipose tissue is more plentiful and easily obtained than bone marrow. AD-MSCs will likely play a prominent role in future research until superior alternatives are developed.

AD-MSCs are morphologically similar to BM-MSCs, they can differentiate into diverse mesodermal tissues, and they express analogous cell surface proteins. Adipose tissue contains significant number of primitive stromal stem cells, with up to 5000 AD-MSCs per gram of fat, in contrast, bone marrow contains 100 to 1000 stem cells per milliliter [65]. Collection of AD-MSCs is also more convenient than collection of BM-MSCs, because it can achieved by the minimally invasive procedure of liposuction under local anesthesia. Liposuction is a common cosmetic procedure in which the fat tissue is frequently discarded, but these tissues could be used as a valuable source of stem cells [83]. There is evidence that the proliferation of AD-MSCs is approximately two-fold greater than that of other types of stem cells. Thus, extracting a small quantity of fat allows the rapid collection of a large number of AD-MSCs, thereby mitigating the risks associated with cell differentiation and mutation during in vitro culture [84]. Although AD-MSCs and BM-MSCs have many biological similarities, they also have some differences in terms of immunophenotype, differentiation potential, transcriptome, proteome, and immunomodulatory activity. Despite these differences, AD-MSCs appear to be as effective as BM-MSCs in certain clinical applications and may be more suitable in some cases [85].

Perinatal mesenchymal stem cells can be obtained from various sources, such as the umbilical cord, umbilical cord blood, and Whartons jelly (muco-polysaccharides in the umbilical cord), and their collection is typically considered noninvasive and free from ethical concerns. Although UC-MSCs account for only 710% of all cells in the umbilical cord, their rapid proliferation allows for efficient expansion using in vitro culture [86].

Although MSCs from bone marrow, adipose tissue, and the umbilical cord have many characteristics in common, they differ in terms of immunophenotype, differentiation potential, and immunomodulatory properties. Clinical trials examining the efficacy of these different MSCs in treating neurodegenerative diseases, endocrine and reproductive disorders, skin regeneration, abnormalities in pulmonary development, and cardiovascular diseases have confirmed they have diverse functions and presumably different therapeutic potential. The process of selecting specific types of MSCs for treatment of different specific diseases is currently problematic. These cells may have similar effects on inhibiting disease progression in vitro, but the mechanisms differ, necessitating further preclinical research and clinical trials to indentify the mechanisms of MSCs that are from different sources [25].

MSCs, which are considered to be multipotent stem cells, can be obtained from diverse tissues. Previous studies have administered MSCs from different sources utilizing a range of techniques, dosages, and timing of administration. The therapeutic effects of MSC treatment on lung function were demonstrated by reductions in the mean linear intercept, decreased pulmonary epithelial cell apoptosis, and improvements in the structural integrity of injured lung tissue [87,88,89,90]. The cell-based approach to regenerative medicine posits that MSC transplantation promotes the repair of lung injury because it introduces healthy cells that function in the structural and functional regeneration of damaged or diseased lung tissue (i.e., cell replacement or implantation), or because the paracrine effects of MSCs promote endogenous regeneration and repair [64]. Studies of different types of lung injuries showed that systemic or direct injection of airways with MSCs successfully introduced these cells into rodent lungs, but most of these studies found that the population of cells implanted in the lungs was too low to be physiologically or functionally significant. Instead, the therapeutic effects of these cells were attributed to paracrine signaling. In fact, many studies found that MSCs can decrease systemic inflammation and stimulate the production of diverse anti-inflammatory molecules [75]. Additionally, MSCs can stimulate the proliferation of different cell types within the lungs, thereby facilitating endogenous repair of lung tissue [91, 92]. Some research showed that MSCs can attract native stem cells to the injury site and promote their differentiation, thereby initiating the regeneration of epithelial tissue [93]. Despite the many research findings supporting the potential efficacy of MSCs in the treatment of respiratory and degenerative disorders, there is still an incomplete understanding of the precise mechanisms by which they decrease lung inflammation and facilitate organ recovery. It seems likely that the type of stem cell and the nature of the injury or disease determines the mechanism, which may include direct differentiation into different cell types, immunomodulation, activation of paracrine pathways, and increasing antiapoptotic activity [74, 94,95,96,97] (Fig.3).

Mechanisms of MSCs-based therapies for COPD. VEGF: vascular endothelial growth factor; HGF: hepatocyte-derived growth factor; Bcl-2: B cell lymphoma-2; Bax: Bcl-2-associated X protein; ROS: reactive oxygen species; TGF-: transforming growth factor-; MMPs: matrix metalloproteinases; ECM: extracellular matrix; IL: interleukin. By figdraw (www.figdraw.com)

MSCs have unique properties that make them suitable candidates for treatment of COPD. MSCs can differentiate into lung-specific cells and replace damaged or dead cells, increase the activity and regenerative potential of endogenous tissue-resident stem cells, promote regeneration of lung structures, improve the structural integrity of airways, and decrease airflow limitation and restore lung function [47]. Some in vitro studies shown that ESCs and adult stem cells can induce the expression of phenotype markers for airway and/or alveolar epithelial cells [98]. When administered intravenously, MSCs primarily target the lungs [99, 100]. Some research suggests that transplanted MSCs have an initial preference for the lungs before migrating to other organs, such as the liver [101]. Multiple studies have demonstrated that MSCs can engraft into mature differentiated airway and alveolar epithelial cells. AD-MSCs can differentiate into alveolar epithelial cells, thereby ameliorating lung injury in a murine model of elastase-induced emphysema [102]. Additionally, the implantation of BM-MSCs into the lungs of an animal model led to their differentiation into ATIICs and the inhibition of alveolar cell apoptosis, thereby preventing lung emphysema induced by radiation and papain protease [103]. The mechanisms underlying the recruitment of circulating or systemically administered stem cells or progenitor cells into the lungs have not yet been fully elucidated, but are likely to be influenced the age of both the donor and recipient, cell type, and route of administration. However, the current understanding suggests that exogenous stem cells have limited potential for structural repair or replacement of damaged lung epithelial cells, indicating the need for additional research to verify the potential of functional epithelial transplantation [104]. Recently, the concept of cell replacement or implantation has been revived. Implantation of basal-like airway epithelial progenitor cells, iPSC-derived lung epithelial cells, or embryonic stem cell-derived AT2 cells using alternative cell sources appears to provide potential therapeutic effect in regenerating damaged lung tissues [64].

In recent years, there has been growing interest in the paracrine effects of MSCs, and the secretome of MSCs has emerged as a potential alternative to cell therapy for various lung diseases. Schweitzer et al. demonstrated that AD-MSCs had therapeutic effects on lung and systemic injuries induced by cigarette smoke, such as lung airway dilation, weight loss, and bone marrow suppression, and proposed that paracrine factors released by AD-MSCs were responsible for these effects [105]. Shigemura et al. validated that the reparative potential of AD-MSCs in treating emphysema was mediated by secretion of hepatocyte growth factor, and this intervention led to improved gas exchange and enhanced exercise tolerance [91]. Hence, there is great interest in the MSC-mediated mechanism of lung tissue repair and the role of paracrine activity in this process. The secretome of MSCs, which includes conditioned medium (CM) and extracellular vesicles (EVs), has immunomodulatory effects and decreases inflammation in pulmonary airways [106].

MSC-CM is readily accessible and regarded as minimally manipulated cell-free material derived from MSCs. There is evidence that administration of MSC-CM leads to a significant reduction in the severity of lung injury, and had efficacy comparable to MSCs in various in vitro and in vivo animal models. For example, Shologu et al. studied CM from BM-MSCs and AD-MSCs, and examined its effect on low-oxygen-induced lung epithelial injury in alveolar epithelial cells. Their findings indicated that MSC-CM improved alveolar epithelial cell viability, decreased the secretion of proinflammatory mediators, and increased the production of the anti-inflammatory cytokine IL-10 [107]. Other studies evaluated the therapeutic efficacy of MSC-CM in a rodent model of COPD induced by cigarette smoke exposure, and the results suggested that MSC and MSC-CM lead to a notable decrease in emphysema and an increase in the quantity of pulmonary capillaries [108]. In addition to emphysema, cigarette smoke can also trigger apoptosis in lung fibroblasts. Thus, Kim et al. reported that MSC-CM mitigated the apoptosis of lung fibroblasts and promoted their proliferation in vivo and in vitro [109]. The findings of these many studies of lung injury models indicate a significant potential for utilization of MSC-CM for decreasing cell death and inflammatory reactions and for increasing tissue repair and endogenous regeneration.

EVs, which include proteins, mRNAs, miRNAs, long noncoding RNAs, and lipids, can regulate gene expression and modulate diverse pathways [110]. EVs are categorized as exosomes, microvesicles, or apoptotic bodies based on their origin, mechanism of secretion, size, and surface markers. A study in mice demonstrated that MSC-EVs decreased pulmonary fibrosis, restored lung structure, improved alveolar formation, and enhanced lung function [111]. MSCs-EVs presumably had these effects by transferring bioactive mediators to injured cells, thereby regulating pathological and physiological responses and promoting cell survival, while decreasing immune and inflammatory responses [112]. Thus, these recent preclinical studies of lung injury indicate that the secretome of MSCs, including CM and EVs, has promising therapeutic effects.

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Magellan Stem Cells has welcomed a $7 million grant from the federal governments Medical Research Future Fund (MRFF) to help fund a Phase 3 human trial of the companys donor stem cell treatment for osteoarthritis.

In announcing the grant, Health Minister Mark Butler said, We are living through a supercharged period of discovery in health and medical research, and the Albanese Government is proud to support Australias world class researchers.

Stem cell therapies could provide innovative treatments for many chronic and inherited diseases we cant yet treat effectively. Such therapies could also revolutionise how we test and develop new medications."

Magellans Phase 3 osteoarthritis trial is one of six projects sharing $34.5 million supported by the MRFF Stem Cell Therapies Research Grant Opportunity.

Osteoarthritis is a huge cause of pain and disability in Australia, said Minister Butler. Up until now the treatments only extended to pain relief and potentially replacement surgery, like knee replacements.

The trial of Magellans donor stem cell treatment for osteoarthritis is scheduled to begin next year.

The grant was announced following the publication of research by Magellan, which demonstrated the potential for significant therapeutic benefits of its MAG200, an off-the-shelf donor stem cell therapy for osteoarthritis.

The research findings are published in Osteoarthritis and Cartilage Open.

Lead researcher and Magellan chief medical officer, Associate Professor Julien Freitag, said, We are very grateful to the Australian Government and the

Medical Research Future Fund (MRFF) for their support for this potentially life-changing technology.

The grant is a vote of confidence in the future of the Australian biotech sector, stem cell technology and Magellans ground-breaking research.

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Magellan Stem Cells welcomes $7 million federal government grant

Stem Cell Therapy Research: Creative Biolabs Advances iPSC-Derived Macrophage Solutions  openPR

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Stem Cell Therapy Market Dynamics: Size, Share, and Growth - openPR

Researchers have been looking for something that can help the body heal itself. Although studies are ongoing, stem cell research brings this notion of regenerative medicine a step closer. However, many of its ideas and concepts remain controversial. So, what are stem cells, and why are they so important?

Stem cells are cells that can develop into other types of cells. For example, they can become muscle or brain cells. They can also renew themselves by dividing, even after they have been inactive for a long time.

Stem cell research is helping scientists understand how an organism develops from a single cell and how healthy cells could be useful in replacing cells that are not working correctly in people and animals.

Researchers are now studying stem cells to see if they could help treat a variety of conditions that impact different body systems and parts.

This article looks at types of stem cells, their potential uses, and some ethical concerns about their use.

The human body requires many different types of cells to function, but it does not produce every cell type fully formed and ready to use.

Scientists call a stem cell an undifferentiated cell because it can become any cell. In contrast, a blood cell, for example, is a differentiated cell because it has already formed into a specific kind of cell.

The sections below look at some types of stem cells in more detail.

Scientists extract embryonic stem cells from unused embryos left over from in vitro fertilization procedures. They do this by taking the cells from the embryos at the blastocyst stage, which is the phase in development before the embryo implants in the uterus.

These cells are undifferentiated cells that divide and replicate. However, they are also able to differentiate into specific types of cells.

There are two main types of adult stem cells: those in developed bodily tissues and induced pluripotent stem (iPS) cells.

Developed bodily tissues such as organs, muscles, skin, and bone include some stem cells. These cells can typically become differentiated cells based on where they exist. For example, a brain stem cell can only become a brain cell.

On the other hand, scientists manipulate iPS cells to make them behave more like embryonic stem cells for use in regenerative medicine. After collecting the stem cells, scientists usually store them in liquid nitrogen for future use. However, researchers have not yet been able to turn these cells into any kind of bodily cell.

Scientists are researching how to use stem cells to regenerate or treat the human body.

The list of conditions that stem cell therapy could help treat may be endless. Among other things, it could include conditions such as Alzheimers disease, heart disease, diabetes, and rheumatoid arthritis. Doctors may also be able to use stem cells to treat injuries in the spinal cord or other parts of the body.

They may do this in several ways, including the following.

In some tissues, stem cells play an essential role in regeneration, as they can divide easily to replace dead cells. Scientists believe that knowing how stem cells work can help treat damaged tissue.

For instance, if someones heart contains damaged tissue, doctors might be able to stimulate healthy tissue to grow by transplanting laboratory-grown stem cells into the persons heart. This could cause the heart tissue to renew itself.

One study suggested that people with heart failure showed some improvement 2 years after a single-dose administration of stem cell therapy. However, the effect of stem cell therapy on the heart is still not fully clear, and research is still ongoing.

Another investigation suggested that stem cell therapies could be the basis of personalized diabetes treatment. In mice and laboratory-grown cultures, researchers successfully produced insulin-secreting cells from stem cells derived from the skin of people with type 1 diabetes.

Study author Jeffrey R. Millman an assistant professor of medicine and biomedical engineering at the Washington University School of Medicine in St. Louis, MO said, What were envisioning is an outpatient procedure in which some sort of device filled with the cells would be placed just beneath the skin.

Millman hopes that these stem cell-derived beta cells could be ready for research in humans within 35 years.

Stem cells could also have vast potential in developing other new therapies.

Another way that scientists could use stem cells is in developing and testing new drugs.

The type of stem cell that scientists commonly use for this purpose is the iPS cell. These are cells that have already undergone differentiation but which scientists have genetically reprogrammed using genetic manipulation, sometimes using viruses.

In theory, this allows iPS cells to divide and become any cell. In this way, they could act like undifferentiated stem cells.

For example, scientists want to grow differentiated cells from iPS cells to resemble cancer cells and use them to test anticancer drugs. This could be possible because conditions such as cancer, as well as some congenital disabilities, happen because cells divide abnormally.

However, more research is taking place to determine whether or not scientists really can turn iPS cells into any kind of differentiated cell and how they can use this process to help treat these conditions.

In recent years, clinics have opened that offer different types of stem cell treatments. One 2016 study counted 570 of these clinics in the United States alone. They appear to offer stem cell-based therapies for conditions ranging from sports injuries to cancer.

However, most stem cell therapies are still theoretical rather than evidence-based. For example, researchers are studying how to use stem cells from amniotic fluid which experts can save after an amniocentesis test to treat various conditions.

The Food and Drug Administration (FDA) does allow clinics to inject people with their own stem cells as long as the cells are intended to perform only their normal function.

Aside from that, however, the FDA has only approved the use of blood-forming stem cells known as hematopoietic progenitor cells. Doctors derive these from umbilical cord blood and use them to treat conditions that affect the production of blood. Currently, for example, a doctor can preserve blood from an umbilical cord after a babys birth to save for this purpose in the future.

The FDA lists specific approved stem cell products, such as cord blood, and the medical facilities that use them on its website. It also warns people to be wary of undergoing any unproven treatments because very few stem cell treatments have actually reached the earliest phase of a clinical trial.

Historically, the use of stem cells in medical research has been controversial. This is because when the therapeutic use of stem cells first came to the publics attention in the late 1990s, scientists were only deriving human stem cells from embryos.

Many people disagree with using human embryonic cells for medical research because extracting them means destroying the embryo. This creates complex issues, as people have different beliefs about what constitutes the start of human life.

For some people, life starts when a baby is born, while for others, it starts when an embryo develops into a fetus. Meanwhile, other people believe that human life begins at conception, so an embryo has the same moral status and rights as a human child.

Former U.S. president George W. Bush had strong antiabortion views. He believed that an embryo should be considered a life and not be used for scientific experiments. Bush banned government funding for human stem cell research in 2001, but former U.S. president Barack Obama then revoked this order. Former U.S. president Donald Trump and current U.S. president Joe Biden have also gone back and forth with legislation on this.

However, by 2006, researchers had already started using iPS cells. Scientists do not derive these stem cells from embryonic stem cells. As a result, this technique does not have the same ethical concerns. With this and other recent advances in stem cell technology, attitudes toward stem cell research are slowly beginning to change.

However, other concerns related to using iPS cells still exist. This includes ensuring that donors of biological material give proper consent to have iPS cells extracted and carefully designing any clinical studies.

Researchers also have some concerns that manipulating these cells as part of stem cell therapy could lead to the growth of cancerous tumors.

Although scientists need to do much more research before stem cell therapies can become part of regular medical practice, the science around stem cells is developing all the time.

Scientists still conduct embryonic stem cell research, but research into iPS cells could help reduce some of the ethical concerns around regenerative medicine. This could lead to much more personalized treatment for many conditions and the ability to regenerate parts of the human body.

Learn more about stem cells, where they come from, and their possible uses here.

Link:
Stem cells: Therapy, controversy, and research - Medical News Today

Signal Transduct Target Ther. 2022; 7: 272.

1 Department of Research and Development, Vinmec Research Institute of Stem Cell and Gene Technology, Vinmec Healthcare System, Hanoi, Vietnam

2Department of Cellular Therapy, Vinmec High-Tech Center, Vinmec Healthcare System, Hanoi, Vietnam

1 Department of Research and Development, Vinmec Research Institute of Stem Cell and Gene Technology, Vinmec Healthcare System, Hanoi, Vietnam

2Department of Cellular Therapy, Vinmec High-Tech Center, Vinmec Healthcare System, Hanoi, Vietnam

1 Department of Research and Development, Vinmec Research Institute of Stem Cell and Gene Technology, Vinmec Healthcare System, Hanoi, Vietnam

1 Department of Research and Development, Vinmec Research Institute of Stem Cell and Gene Technology, Vinmec Healthcare System, Hanoi, Vietnam

1 Department of Research and Development, Vinmec Research Institute of Stem Cell and Gene Technology, Vinmec Healthcare System, Hanoi, Vietnam

1 Department of Research and Development, Vinmec Research Institute of Stem Cell and Gene Technology, Vinmec Healthcare System, Hanoi, Vietnam

1 Department of Research and Development, Vinmec Research Institute of Stem Cell and Gene Technology, Vinmec Healthcare System, Hanoi, Vietnam

3Institute for Science & Technology in Medicine, Keele University, Keele, UK

4Department of Biology, Stanford University, Stanford, CA USA

1 Department of Research and Development, Vinmec Research Institute of Stem Cell and Gene Technology, Vinmec Healthcare System, Hanoi, Vietnam

1 Department of Research and Development, Vinmec Research Institute of Stem Cell and Gene Technology, Vinmec Healthcare System, Hanoi, Vietnam

2Department of Cellular Therapy, Vinmec High-Tech Center, Vinmec Healthcare System, Hanoi, Vietnam

3Institute for Science & Technology in Medicine, Keele University, Keele, UK

4Department of Biology, Stanford University, Stanford, CA USA

Received 2022 Mar 15; Revised 2022 Jul 19; Accepted 2022 Jul 21.

Recent advancements in stem cell technology open a new door for patients suffering from diseases and disorders that have yet to be treated. Stem cell-based therapy, including human pluripotent stem cells (hPSCs) and multipotent mesenchymal stem cells (MSCs), has recently emerged as a key player in regenerative medicine. hPSCs are defined as self-renewable cell types conferring the ability to differentiate into various cellular phenotypes of the human body, including three germ layers. MSCs are multipotent progenitor cells possessing self-renewal ability (limited in vitro) and differentiation potential into mesenchymal lineages, according to the International Society for Cell and Gene Therapy (ISCT). This review provides an update on recent clinical applications using either hPSCs or MSCs derived from bone marrow (BM), adipose tissue (AT), or the umbilical cord (UC) for the treatment of human diseases, including neurological disorders, pulmonary dysfunctions, metabolic/endocrine-related diseases, reproductive disorders, skin burns, and cardiovascular conditions. Moreover, we discuss our own clinical trial experiences on targeted therapies using MSCs in a clinical setting, and we propose and discuss the MSC tissue origin concept and how MSC origin may contribute to the role of MSCs in downstream applications, with the ultimate objective of facilitating translational research in regenerative medicine into clinical applications. The mechanisms discussed here support the proposed hypothesis that BM-MSCs are potentially good candidates for brain and spinal cord injury treatment, AT-MSCs are potentially good candidates for reproductive disorder treatment and skin regeneration, and UC-MSCs are potentially good candidates for pulmonary disease and acute respiratory distress syndrome treatment.

Subject terms: Stem-cell research, Mesenchymal stem cells

The successful approval of cancer immunotherapies in the US and mesenchymal stem cell (MSC)-based therapies in Europe have turned the wheel of regenerative medicine to become prominent treatment modalities.13 Cell-based therapy, especially stem cells, provides new hope for patients suffering from incurable diseases where treatment approaches focus on management of the disease not treat it. Stem cell-based therapy is an important branch of regenerative medicine with the ultimate goal of enhancing the body repair machinery via stimulation, modulation, and regulation of the endogenous stem cell population and/or replenishing the cell pool toward tissue homeostasis and regeneration.4 Since the stem cell definition was introduced with their unique properties of self-renewal and differentiation, they have been subjected to numerous basic research and clinical studies and are defined as potential therapeutic agents. As the main agenda of regenerative medicine is related to tissue regeneration and cellular replacement and to achieve these targets, different types of stem cells have been used, including human pluripotent stem cells (hPSCs), multipotent stem cells and progenitor cells.5 However, the emergence of private and unproven clinics that claim the effectiveness of stem cell therapy as magic cells has raised highly publicized concerns about the safety of stem cell therapy. The most notable case involved the injection of a cell population derived from fractionated lipoaspirate into the eyes of three patients diagnosed with macular degeneration, resulting in the loss of vision for these patients.6 Thus, as regenerative medicine continues to progress and evolve and to clear the myth of the magic cells, this review provides a brief overview of stem cell-based therapy for the treatment of human diseases.

Stem cell therapy is a novel therapeutic approach that utilizes the unique properties of stem cells, including self-renewal and differentiation, to regenerate damaged cells and tissues in the human body or replace these cells with new, healthy and fully functional cells by delivering exogenous cells into a patient.7 Stem cells for cell-based therapy can be of (1) autologous, also known as self-to-self therapy, an approach using the patients own cells, and (2) allogeneic sources, which use cells from a healthy donor for the treatment.8 The term stem cell were first used by the eminent German biologist Ernst Haeckel to describe the properties of fertilized egg to give rise to all cells of the organism in 1868.9 The history of stem cell therapy started in 1888, when the definition of stem cell was first coined by two German zoologists Theodor Heinrich Boveri and Valentin Haecker,9 who set out to identify the distinct cell population in the embryo capable of differentiating to more specialized cells (Fig. ). In 1902, studies carried out by the histologist Franz Ernst Christian Neumann, who was working on bone marrow research, and Alexander Alexandrowitsch Maximov demonstrated the presence of common progenitor cells that give rise to mature blood cells, a process also known as haematopoiesis.10 From this study, Maximov proposed the concept of polyblasts, which later were named stem cells based on their proliferation and differentiation by Ernst Haeckel.11 Maximov described a hematopoietic population presented in the bone marrow. In 1939, the first case report described the transplantation of human bone marrow for a patient diagnosed with aplastic anemia. Twenty years later, in 1958, the first stem cell transplantation was performed by the French oncologist George Mathe to treat six nuclear researchers who were accidentally exposed to radioactive substances using bone marrow transplantation.12 Another study by George Mathe in 1963 shed light on the scientific community, as he successfully conducted bone marrow transplantation in a patient with leukemia. The first allogeneic hematopoietic stem cell transplantation (HSCT) was pioneered by Dr. E. Donnall Thomas in 1957.13 In this initial study, all six patients died, and only two patients showed evidence of transient engraftment due to the unknown quantities and potential hazards of bone marrow transplantation at that time. In 1969, Dr. E. Donnall Thomas conducted the first bone marrow transplantation in the US, although the success of the allogeneic treatment remained exclusive. In 1972, the year marked the discovery of cyclosporine (the immune suppressive drug),14 the first successes of allogeneic transplantation for aplastic anemia and acute myeloid leukemia were reported in a 16-year-old girl.15 From the 1960s to the 1970s, series of works conducted by Friendenstein and coworkers on bone marrow aspirates demonstrated the relationship between osteogenic differentiation and a minor subpopulation of cells derived from bone marrow.16 These cells were later proven to be distinguishable from the hematopoietic population and to be able to proliferate rapidly as adherent cells in tissue culture vessels. Another important breakthrough from Friendensteins team was the discovery that these cells could form the colony-forming unit when bone marrow was seeded as suspension culture following by differentiation into osteoblasts, adipocytes, and chondrocytes, suggesting that these cells confer the ability to proliferate and differentiate into different cell types.17 In 1991, combined with the discovery of human embryonic stem cells (hESCs), which will be discussed in the next section, the term mesenchymal stem cells, previously known as stromal stem cells or osteogenic stem cells, was first coined in Caplan and widely used to date.18 Starting with bone marrow transplantation 60 years ago, the journey of stem cell therapy has developed throughout the years to become a novel therapeutic agent of regenerative medicine to treat numerous incurable diseases, which will be reviewed and discussed in this review, including neurological disorders, pulmonary dysfunctions, metabolic/endocrine-related diseases, reproductive disorders, skin burns, and cardiovascular conditions).

Stem cell-based therapy: the history and cell source. a The timeline of major discoveries and advances in basic research and clinical applications of stem cell-based therapy. The term stem cells was first described in 1888, setting the first milestone in regenerative medicine. The hematopoietic progenitor cells were first discovered in 1902. In 1939, the first bone marrow transplantation was conducted in the treatment of aplasmic anemia. Since then, the translation of basic research to preclinical studies to clinical trials has driven the development of stem cell-based therapy by many discoveries and milestones. The isolations of mesenchymal stem cells in 1991 following by the discovery of human pluripotent stem cells have recently contributed to the progress of stem cell-based therapy in the treatment of human diseases. b Schematic of the different cell sources that can be used in stem cell-based therapy. (1) Human pluripotent stem cells, including embryonic stem cells (derived from inner cell mass of blastocyst) and induced pluripotent stem cells confer the ability to proliferate indefinitely in vitro and differentiate into numerous cell types of the human body, including three germ layers. (2) Mesenchymal stem cells are multipotent stem cells derived from mesoderm possessing self-renewal ability (limited in vitro) and differentiation potential into mesenchymal lineages. The differentiated/somatic cells can be reprogrammed back to the pluripotent stage using OSKM factors to generate induced pluripotent stem cells. It is important to note that stem cells show a relatively higher risk of tumor formation and lower risk of immune rejection (in the case of mesenchymal stem cells) when compared to that of somatic cells. The figure was created with BioRender.com

In this review, we described the different types of stem cell-based therapies (Fig. ), including hPSCs and MSCs, and provided an overview of their definition, history, and outstanding clinical applications. In addition, we further created the first literature portfolio for the targeted therapy of MSCs based on their origin, delineating their different tissue origins and downstream applications with an in-depth discussion of their mechanism of action. Finally, we provide our perspective on why the tissue origin of MSCs could contribute greatly to their downstream applications as a proposed hypothesis that needs to be proven or disproven in the future to further enhance the safety and effectiveness of stem cell-based therapy.

The clinical applications of stem cell-based therapies for heart diseases have been recently discussed comprehensively in the reviews19,20 and therefore will be elaborated in this study as the focus discussions related to hPSCs and MSCs in the following sections. In general, the safety profiles of stem cell-based therapies are supported by a large body of preclinical and clinical studies, especially adult stem cell therapy (such as MSC-based products). However, clinical trials have not yet yielded data supporting the efficacy of the treatment, as numerous studies have shown paradoxical results and no statistically significant differences in infarct size, cardiac function, or clinical outcomes, even in phase III trials.21 The results of a meta-analysis showed that stem cells derived from different sources did not exhibit any therapeutic effects on the improvement of myocardial contractility, cardiovascular remodeling, or clinical outcomes.22 The disappointing results obtained from the clinical trials thus far could be explained by the fact that the administered cells may exert their therapeutic effects via an immune modulation rather than regenerative function. Thus, well-designed, randomized and placebo-controlled phase III trials with appropriate cell-preparation methods, patient selection, follow-up schedules and suitable clinical measurements need to be conducted to determine the efficacy of the treatments. In addition, concerns related to optimum cell source and dose, delivery route and timing of administration, cell distribution post administration and the mechanism of action also need to be addressed. In the following section of this review, we present clinical trials related to MSC-based therapy in cardiovascular disease with the aim of discussing the contradictory results of these trials and analyzing the potential challenges underlying the current approaches.

Gastrointestinal diseases are among the most diagnosed conditions in the developed world, altering the life of one-third of individuals in Western countries. The gastrointestinal tract is protected from adverse substances in the gut environment by a single layer of epithelial cells that are known to have great regenerative ability in response to injuries and normal cell turnover.23 These epithelial cells have a rapid turnover rate of every 27 days under normal conditions and even more rapidly following tissue damage and inflammation. This rapid proliferation ability is possible owing to the presence of a specific stem cell population that is strictly compartmentalized in the intestinal crypts.24 The gastrointestinal tract is highly vulnerable to damage, tissue inflammation and diseases once the degradation of the mucosal lining layer occurs. The exposure of intestinal stem cells to the surrounding environment of the gut might result in the direct destruction of the stem cell layer or disruption of intestinal functions and lead to overt clinical symptoms.25 In addition, the accumulation of stem cell defects as well as the presence of chronic inflammation and stress also contributes to the reduction of intestinal stem cell quality.

In terms of digestive disorders, Crohns disease (CD) and ulcerative colitis are the two major forms of inflammatory bowel disease (IBD) and represent a significant burden on the healthcare system. The former is a chronic, uncontrolled inflammatory condition of the intestinal mucosa characterized by segmental transmural mucosal inflammation and granulomatous changes.26 The latter is a chronic inflammatory bowel disease affecting the colon and rectum, characterized by mucosal inflammation initiating in the rectum and extending proximal to the colon in a continuous fashion.27 Cellular therapy in the treatment of CD can be divided into haematopoietic stem cell-based therapy and MSC-based therapy. The indication and recommendation of using HSCs for the treatment of IBD were proposed in 1995 by an international committee with four important criteria: (1) refractory to immunosuppressive treatment; (2) persistence of the disease conditions indicated via endoscopy, colonoscopy or magnetic resonance enterography; (3) patients who underwent an imminent surgical procedure with a high risk of short bowel syndromes or refractory colonic disease; and (4) patients who refused to treat persistent perianal lesions using coloproctectomy with a definitive stroma implant.28 In the standard operation procedure, patents HSCs were recruited using cyclophosphamide, which is associated with granulocyte colony-stimulating factor (G-CSF), at two different administered concentrations (4g/m2 and 2g/m2). Recently, it was reported that high doses of cyclophosphamide do not improve the number of recruited HSCs but increase the risk of cardiac and bladder toxicity. An interest in using HSCTs in CD originated from case reports that autologous HSCTs can induce sustained disease remission in some29,30 but not all patients3133 with CD. The first phase I trial was conducted in Chicago and recruited 12 patients with active moderate to severe CD refractory to conventional therapies. Eleven of 12 patients demonstrated sustained remission after a median follow-up of 18.5 months, and one patient developed recurrence of active CD.31 The ASTIC trial (the Autologous Stem Cell Transplantation International Crohn Disease) was the first randomized clinical trial with the largest cohort of patients undergoing HSCT for refractory CD in 2015.34 The early report of the trial showed no statistically significant improvement in clinical outcomes of mobilization and autologous HSCT compared with mobilization followed by conventional therapy. In addition, the procedure was associated with significant toxicity, leading to the suggestion that HSCT for patients with refractory CD should not be widespread. Interestingly, by using conventional assessments for clinical trials for CD, a group reassessed the outcomes of patients enrolled in the ASTIC trial showing clinical and endoscopic benefits, although a high number of adverse events were also detected.35 A recent systematic review evaluated 18 human studies including 360 patients diagnosed with CD and showed that eleven studies confirmed the improvement of Crohns disease activity index between HSCT groups compared to the control group.36 Towards the cell sources, HSCs are the better sources as they afforded more stable outcomes when compared to that of MSC-based therapy.37 Moreover, autologous stem cells were better than their allogeneic counterparts.36 The safety of stem cell-based therapy in the treatment of CD has attracted our attention, as the risk of infection in patients with CD was relatively higher than that in those undergoing administration to treat cancer or other diseases. During the stem cell mobilization process, patient immunity is significantly compromised, leading to a high risk of infection, and requires carefully nursed and suitable antibiotic treatment to reduce the development of adverse events. Taken together, stem cell-based therapy for digestive disease reduced inflammation and improved the patients quality of life as well as bowel functions, although the high risk of adverse events needs to be carefully monitored to further improve patient safety and treatment outcomes.

The liver is the largest vital organ in the human body and performs essential biological functions, including detoxification of the organism, metabolism, supporting digestion, vitamin storage, and other functions.38 The disruption of liver homeostasis and function might lead to the development of pathological conditions such as liver failure, cirrhosis, cancer, alcoholic liver disease, nonalcoholic fatty liver disease (NAFLD), and autoimmune liver disease (ALD). Orthotropic liver transplantation is the only effective treatment for severe liver diseases, but the number of available and suitable donor organs is very limited. Currently, stem cell-based therapies in the treatment of liver disease are associated with HSCs, MSCs, hPSCs, and liver progenitor cells.

Liver failure is a critical condition characterized by severe liver dysfunctions or decompensation caused by numerous factors with a relatively high mortality rate. Stem cell-based therapy is a novel alternative approach in the treatment of liver failure, as it is believed to participate in the enhancement of liver regeneration and recovery. The results of a meta-analysis including four randomized controlled trials and six nonrandomized controlled trials in the treatment of acute-on-chronic liver failure (ACLF) demonstrated that clinical outcomes of stem cell therapy were achieved in the short term, requiring multiple doses of stem cells to prolong the therapeutic effects.39,40 Interestingly, although MSC-based therapies improved liver functions, including the model of end-stage liver disease score, albumin level, total bilirubin, and coagulation, beneficial effects on survival rate and aminotransferase level were not observed.41 A randomized controlled trial illustrated the improvement of liver functions and reduction of severe infections in patients with hepatitis B virus-related ACLF receiving allogeneic bone marrow-derived MSCs (BM-MSCs) via peripheral infusion.42 HSCs from peripheral blood after the G-CSF mobilization process were used in a phase I clinical trial and exhibited an improvement in serum bilirubin and albumin in patients with chronic liver failure without any specific adverse events related to the administration.43 Taken together, an overview of stem cell-based therapy in the treatment of liver failure indicates the potential therapeutic effects on liver functions with a strong safety profile, although larger randomized controlled trials are still needed to assure the conclusions.

Liver cirrhosis is one of the major causes of morbidity and mortality worldwide and is characterized by diffuse nodular regeneration with dense fibrotic septa and subsequent parenchymal extinction leading to the collapse of liver vascular structure.44 In fact, liver cirrhosis is considered the end-stage of liver disease that eventually leads to death unless liver transplantation is performed. Stem cell-based therapy, especially MSCs, currently emerges as a potential treatment with encouraging results for treating liver cirrhosis. In a clinical trial using umbilical cord-derived MSCs (UC-MSCs), 45 chronic hepatitis B patients with decompensated liver cirrhosis were divided into two groups: the MSC group (n=30) and the control group (n=15).45 The results showed a significant reduction in ascites volume in the MSC group compared with the control. Liver function was also significantly improved in the MSC groups, as indicated by the increase in serum albumin concentration, reduction in total serum bilirubin levels, and decrease in the sodium model for end-stage liver disease score.45 Similar results were also reported from a phase II trial using BM-MSCs in 25 patients with HCV-induced liver cirrhosis.46 Consistent with these studies, three other clinical trials targeting liver cirrhosis caused by hepatitis B and alcoholic cirrhosis were conducted and confirmed that MSC administration enhanced and recovered liver functions.4749 With the large cohort study as the clinical trial conducted by Fang, the safety and potential therapeutic effects of MSC-based therapies could be further strengthened and confirmed the feasibility of the treatment in virus-related liver cirrhosis.49 In terms of delivery route, a randomized controlled phase 2 trial suggested that systemic delivery of BM-MSCs does not show therapeutic effects on patients with liver cirrhosis.50 MSCs are not the only cell source for liver cirrhosis. Recently, an open-label clinical trial conducted in 19 children with liver cirrhosis due to biliary atresia after the Kasai operation illustrated the safety and feasibility of the approach by showing the improvement of liver function after bone marrow mononuclear cell (BMNC) administration assessed by biochemical tests and pediatric end-stage liver disease (PELD) scores.51 Another study using BMNCs in 32 decompensated liver cirrhosis patients illustrated the safety and effectiveness of BMNC administration in comparison with the control group.52 Recently, a long-term analysis of patients receiving peripheral blood-derived stem cells indicated a significant improvement in the long-term survival rate when compared to the control group, and the risk of hepatocellular carcinoma formation did not increase.53 CD133+ HSC infusion was performed in a multicentre, open, randomized controlled phase 2 trial in patients with liver cirrhosis; the results did not support the improvement of liver conditions, and cirrhosis persisted.54 Notably, these results are in line with a previous randomized controlled study, which also reported that G-CSF and bone marrow-derived stem cells delivered via the hepatic artery did not introduce therapeutic potential as expected.55 Thus, stem cell-based therapy for liver cirrhosis is still in its immature stage and requires larger trials with well-designed experiments to confirm the efficacy of the treatment.

Nonalcoholic fatty liver disease (NAFLD) is the most common medical condition caused by genetic and lifestyle factors and results in a severe liver condition and increased cardiovascular risk.56 NAFLD is the hidden enemy, as most patients are asymptomatic for a long time, and their routine life is unaffected. Thus, the detection, identification, and management of NAFLD conditions are challenging tasks, as patients diagnosed with NAFLD often develop nonalcoholic steatohepatitis, cirrhosis, and hepatocellular carcinoma.57 Although preclinical studies have shown that stem cell administration could enhance liver function in NAFLD models, a limited number of clinical trials were performed in human subjects. Recently, a multi-institutional clinical trial using freshly isolated autologous adipose tissue-derived regenerative cells was performed in Japan to treat seven NAFLD patients.58 The results illustrated the improvement in the serum albumin level of six patients and prothrombin activity of five patients, and no treatment-related adverse events or severe adverse events were observed. This study illustrates the therapeutic potential of stem cell-based therapy in the treatment of NAFLD.

Autoimmune liver disease (ALD) is a severe liver condition affecting children and adults worldwide, with a female predominance.59 The condition occurs in genetically predisposed patients when a stimulator, such as virus infection, leads to a T-cell-mediated autoimmune response directed against liver autoantigens. As a result, patients with ALD might develop liver cirrhosis, hepatocellular carcinoma, and, in severe cases, death. To date, HSCT and bone marrow transplantation are the two common stem cell-based therapies exhibiting therapeutic potential for ALD in clinical trials. An interesting report illustrated that haploidentical HSCTs could cure ALD in patients with sickle cells.60 This report is particularly important, as it illustrates the potential therapeutic approach of using haploidentical HSCTs to treat patients with both sickle cells and ALD. Another case report described a 19-year-old man with a 4-year history of ALD who developed acute lymphoblastic leukemia and required allogeneic bone marrow transplantation from this wholesome brother.61 The clinical data showed that immunosuppressive therapy for transplantation generated ALD remission in the patient.62 However, the data also provided valid information related to the sustained remission and the normalization of ASGPR-specific suppressor-inducer T-cell activity following bone marrow transplantation, suggesting that these suppressor functions originated from donor T cells.61 Thus, it was suggested that if standard immunosuppressive treatment fails, alternative cellular immunotherapy would be a viable option for patients with ALD. Primary biliary cholangitis (PBC), usually known as primary biliary cirrhosis, is a type of ALD characterized by a slow, progressive destruction of small bile ducts of the liver leading to the formation of cirrhosis and accumulation of bile and other toxins in the liver. A pilot, single-arm trial from China recruited seven patents with PBC who had a suboptimal response to ursodeoxycholic acid (UDCA) treatment.63 These patients received UDCA treatment in combination with three rounds of allogeneic UC-MSCs at 4-week intervals with a dose of 0.5106 cells/kg of patient body weight via the peripheral vein. No treatment-related adverse events or severe adverse events were observed throughout the course of the study. The clinical data indicated significant improvement in liver function, including reduction of serum ALP and gamma-glutamyltransferase (GGT) at 48 weeks post administration. The common symptoms of PBC, including fatigue, pruritus, and hypogastric ascites volume, were also reduced, supporting the feasibility of MSC-based therapy in the treatment of PBC, although major limitations of the study were nonrandomized, no control group and small sample size. Another study was conducted in China with ten PBC patients who underwent incompetent UDCA treatment for more than 1 year. These patients received a range of 35 allogeneic BM-MSCs/kg body weight by intravenous infusion.64 Although these early studies have several limitations, such as small sample size, nonrandomization, and no control group, their preliminary data related to safety and efficacy herald the prospects and support the feasibility of stem cell-based therapy in the treatment of ALD.

In summary, the current number of trials for liver disease using stem cell-based therapy has provided fundamental data supporting the safety and potential therapeutic effects in various liver diseases. Unfortunately, due to the small number of trials, several obstacles need to be overcome to prove the effectiveness of the treatments, including (1) stem cell source and dose, (2) administration route, (3) time of intervention, and (4) clinical assessments during the follow-up period. Only by addressing these challenges we will be able to prove, facilitate and promote stem cell-based therapy as a mainstream treatment for liver diseases.

Arthritis is a general term describing cartilage conditions that cause pain and inflammation of the joints. Osteoarthritis (OA) is the most common form of arthritis caused by persistent degeneration and poor recovery of articular cartilage.65 OA affects one or several diarthrodial joints, such as small joints at the hand and large joints at the knee and hips, leading to severe pain and subsequent reduction in the mobility of patients. There are two types of OA: (1) primary OA or idiopathic OA and secondary OA caused by causative factors such as trauma, surgery, and abnormal joint development at birth.66 As conventional treatments for OA are not consistent in their effectiveness and might cause unbearable pain as well as long-term rehabilitation (in the case of joint replacement), there is a need for a more reliable, less painful, and curative therapy targeting the root of OA.67 Thus, stem cell therapy has recently emerged as an alternative approach for OA and has drawn great attention in the regenerative field.

The administration of HSCs has been proven to reduce bone lesions, enhance bone regeneration and stimulate the vascularization process in degenerative cartilage. Attempts were made to evaluate the efficacy of peripheral blood stem cells in ten OA patients by three intraarticular injections. Post-administration analysis indicated a reduction in the WOMAC index with a significant reduction in all parameters. All patients completed 6-min walk tests with an increase of more than 54 meters. MRI analysis indicated an improvement in cartilage thickness, suggesting that cartilage degeneration was reduced post administration. To further enhance the therapeutic potential of HSCT, CD34+ stem cells were proposed to be used in combination with the rehabilitation algorithm, which included three stages: preoperative, hospitalization and outpatient periods.68 Currently, a large wave of studies has been directed to MSC-based therapy for the treatment of OA due to their immunoregulatory functions and anti-inflammatory characteristics. MSCs have been used as the main cell source in several multiple and small-scale trials, proving their safety profile and potential effectiveness in alleviating pain, reducing cartilage degeneration, and enhancing the regeneration of cartilage structure and morphology in some cases. However, the best source of MSCs, whether from bone marrow, adipose tissue, or umbilical cord, for the management of OA is still a great question to be answered. A systematic review investigating over sixty-one of 3172 articles with approximately 2390 OA patients supported the positive effects of MSC-based therapy on OA patients, although the study also pointed out the fact that these therapeutic potentials were based on limited high-quality evidence and long-term follow-up.69 Moreover, the study found no obvious evidence supporting the most effective source of MSCs for treating OA. Another systematic review covering 36 clinical trials, of which 14 studies were randomized trials, provides an interesting view in terms of the efficacy of autologous MSC-based therapy in the treatment of OA.70 In terms of BM-MSCs, 14 clinical trials reported the clinical outcomes at the 1-year follow-up, in which 57% of trials reported clinical outcomes that were significantly better in comparison with the control group. However, strength analysis of the data set showed that outcomes from six trials were low, whereas the outcomes of the remaining eight trials were extremely low. Moreover, the positive evidence obtained from MRI analysis was low to very low strength of evidence after 1-year post administration.70 Similar results were also found in the outcome analysis of autologous adipose tissue-derived MSCs (AT-MSCs). Thus, the review indicated low quality of evidence for the therapeutic potential of MSC therapy on clinical outcomes and MRI analysis. The low quality of clinical outcomes could be explained by the differences in interventions (including cell sources, cell doses, and administration routes), combination treatments (with hyaluronic acid,71 peripheral blood plasma,72 etc.), control treatments and clinical outcome measurements between randomized clinical trials.73 In addition, the data of the systematic analysis could not prove the better source of MSCs for OA treatment. Taken together, although stem cell-based therapy has been shown to be safe and feasible in the management of OA, the authors support the notion that stem cell-based therapy could be considered an alternative treatment for OA when first-line treatments, such as education, exercise, and body weight management, have failed.

Stem cell therapy in the treatment of cancer is a sensitive term and needs to be used and discussed with caution. Clinicians and researchers should protect patients with cancer from expensive and potentially dangerous or ineffective stem cell-based therapy and patients without a cancer diagnosis from the risk of malignancy development. In general, unproven stem cell clinics employed three cell-based therapies for cancer management, including autologous HSCTs, stromal vascular fraction (SVF), and multipotent stem cells, such as MSCs. Allogeneic HSCTs confer the ability to generate donor lymphocytes that contribute to the suppression and regression of hematological malignancies and select solid tumors, a specific condition known as graft-versus-tumor effects.74 However, stem cell clinics provide allogeneic cell-based therapy for the treatment of solid malignancies despite limited scientific evidence supporting the safety and efficacy of the treatment. High-quality evidence from the Cochrane library shows that marrow transplantation via autologous HSCTs in combination with high-dose chemotherapy does not improve the overall survival of women with metastatic breast cancer. In addition, a study including more than 41,000 breast cancer patients demonstrated no significant difference in survival benefits between patients who received HSCTs following high-dose chemotherapy and patients who underwent conventional treatment.75 Thus, the use of autologous T-cell transplants as monotherapy and advertising stem cell-based therapies as if they are medically approved or preferred treatment of solid tumors is considered untrue statements and needs to be alerted to cancer patients.76

Over the past decades, many preclinical studies have demonstrated the potential of MSC-based therapy in cancer treatment due to their unique properties. They confer the ability to migrate toward damaged sites via inherent tropism controlled by growth factors, chemokines, and cytokines. MSCs express specific CXC chemokine receptor type 4 (CXCR4) and other chemokine receptors (including CCR1, CCR2, CCR4, CCR7, etc.) that are essential to respond to the surrounding signals.77 In addition, specific adherent proteins, including CD49d, CD44, CD54, CD102, and CD106, are also expressed on the MSC surface, allowing them to attach, rotate, migrate, and penetrate the blood vessel lumen to infiltrate the damaged tissue.78 Similar to damaged tissues, tumors secrete a wide range of chemoattractant that also attract MSC migration via the CXCL12/CXCR4 axis. Previous studies also found that MSC migration toward the cancer site is tightly controlled by diffusible cytokines such as interleukin 8 (IL-8) and growth factors including transforming growth factor-beta 1 (TGF-1),79 platelet-derived growth factor (PDGF),80 fibroblast growth factor 2 (FGF-2),81 vascular endothelial growth factor (VEGF),81 and extracellular matrix molecules such as matrix metalloproteinase-2 (MMP-2).82 Once MSCs migrate successfully to cancerous tissue, accumulating evidence demonstrates the interaction between MSCs and cancer cells to exhibit their protumour and antitumour effects, which are the major concerns of MSC-based therapy. MSCs are well-known for their regenerative effects that regulate tissue repair and recovery. This unique ability is also attributed to the protumour functions of these cells. A previous study reported that breast cancer cells induce MSC secretion of chemokine (CC motif) ligand 5 (CCL-5), which regulates the tumor invasion process.83,84 Other studies also found that MSCs secrete a wide range of growth factors (VEGF, basic FGF, HGF, PDGF, etc.) that inhibits apoptosis of cancer cells.85 Moreover, MSCs also respond to signals released from cancer cells, such as TGF-,86 to transform into cancer-associated fibroblasts, a specific cell type residing within the tumor microenvironment capable of promoting tumorigenesis.87 Although MSCs have been proven to be involved in protumour activities, they also have potent tumor suppression abilities that have been used to develop cancer treatments. It has been suggested that MSCs exhibit their tumor inhibitory effects by inhibiting the Wnt and AKT signaling pathways,88 reducing the angiogenesis process,89 stimulating inflammatory cell infiltration,90 and inducing tumor cell cycle arrest and apoptosis.91 To date, the exact functions of MSCs in both protumour and antitumor activities are still a controversial issue across the stem cell field. Other approaches exploit gene editing and tissue engineering to convert MSCs into a Trojan horse that could exhibit antitumor functions. In addition, MSCs can also be modified to express specific anticancer miRNAs exhibiting tumor-suppressive behaviors.92 However, genetically modified MSCs are still underdeveloped and require intensive investigation in the clinical setting.

To date, ~25 clinical trials have been registered on ClinicalTrials.gov aimed at using MSCs as a therapeutic treatment for cancer.93 These trials are mostly phase 1 and 2 studies focusing on evaluating the safety and efficacy of the treatment. Studies exploiting MSC-based therapy have combined MSCs with an oncolytic virus approach. Oncolytic viruses are specific types of viruses that can be genetically engineered or naturally present, conferring the ability to selectively infect cancer cells and kill them without damaging the surrounding healthy cells.94 A completed phase I/II study using BM-MSCs infected with the oncolytic adenovirus ICOVIR5 in the treatment of metastatic and refractory solid tumors in children and adult patients demonstrated the safety of the treatment and provided preliminary data supporting their therapeutic potential.95 The same group also reported a complete disappearance of all signs of cancer in response to MSC-based therapy in one pediatric case three years post administration.96 A reported study in 2019 claimed that adipose-derived MSCs infected with vaccinia virus have the potential to eradicate resistant tumor cells via the combination of potent virus amplification and senitization of the tumor cells to virus infection.97 However, in a recently published review, a valid question was posed regarding the 2019 study that do these reported data merit inclusion in the publication record when they were collected by such groups using a dubious therapeutic that was eventually confiscated by US Marshals?76

Taken together, cancer research and therapy have entered an innovative and fascinating era with advancements in traditional therapies such as chemotherapy, radiotherapy, and surgery on one hand and stem cell-based therapy on the other hand. Although stem cell-based therapy has been considered a novel and attractive therapeutic approach for cancer treatment, it has been hampered by contradictory results describing the protumour and antitumour effects in preclinical studies. Despite this contradictory reality, the use of stem cell-based therapy, especially MSCs, offers new hope to cancer patients by providing a new and more effective tool in personalized medicine. The authors support the use of MSC-based therapy as a Trojan horse to deliver specific anticancer functions toward cancer cells to suppress their proliferation, eradicate cancer cells, or limit the vascularization process of cancerous tissue to improve the clinical safety and efficacy of the treatment.

The discovery of hPSCs, including human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs), has revolutionized stem cell research and cell-based therapy.98 hESCs were first isolated from blastocyst-stage embryos in 1998,99 followed by breakthrough reprogramming research that converted somatic cells into hiPSCs using just four genetic factors.100,101 Methods have been developed to maintain these cells long-term in vitro and initiate their differentiation into a wide variety of cell types, opening a new era in regenerative medicine, particularly cell therapy to replace lost or damaged tissues.

hPSCs are defined as self-renewable cell types that confer the ability to differentiate into various cellular phenotypes of the human body, including three germ layers.102 Historically, the first pluripotent cell lines to be generated were embryonic carcinoma (EC) cell lines established from human germ cell tumors103 and murine undifferentiated compartments.104 Although EC cells are a powerful tool in vitro, these cells are not suitable for clinical applications due to their cancer-derived origin and aneuploidy genotype.105 The first murine ESCs were established in 1981 based on the culture techniques obtained from EC research.106 Murine ESCs are derived from the inner cell mass (ICM) of the pre-implantation blastocyst, a unique biological structure that contains outer trophoblast layers that give rise to the placenta and ICM.107 In vivo ESCs only exist for a short period during the embryos development, and they can be isolated and maintained indefinitely in vitro in an undifferentiated state. The discovery of murine ESCs has dramatically changed the field of biomedical research and regenerative medicine over the last 40 years. Since then, enormous investigations have been made to isolate and culture ESCs from other species, including hESCs, in 1998.99 The success of Thomson et al. in 1998 triggered the great controversy in media and ethical research boards across the globe, with particularly strong objections being raised to the use of human embryos for research purposes.108 Several studies using hESCs have been conducted demonstrating their therapeutic potential in the clinical setting. However, the use of hESCs is limited due to (1) the ethical barrier related to the destruction of human embryos and (2) the potential risk of immunological rejection, as hESCs are isolated from pre-implantation blastocysts, which are not autologous in origin. To overcome these two great obstacles, several research groups have been trying to develop technology to generate hESCs, including nuclear transfer technology, the well-known strategy that creates Dolly sheep, although the generation of human nuclear transfer ESCs remains technically challenging.109 Taking a different approach, in 2006, Yamanaka and Takahashi generated artificial PSCs from adult and embryonic mouse somatic cells using four transcription factors (Oct-3/4, Sox2, Klf4, and c-Myc, called OSKM) reduced from 24 factors.100 Thereafter, in 2007, Takahashi and colleagues successfully generated the first hiPSCs exhibiting molecular and biological features similar to those of hESCs using the same OSKM factors.101 Since then, hiPSCs have been widely studied to expand our knowledge of the pathogenesis of numerous diseases and aid in developing new cell-based therapies as well as personalized medicine.

Since its beginning 24 years ago, hPSC research has evolved momentously toward applications in regenerative medicine, disease modeling, drug screening and discovery, and stem cell-based therapy. In clinical trial settings, the uses of hESCs are restricted by ethical concerns and tight regulation, and the limited preclinical data support their therapeutic potential. However, it is important to acknowledge several successful outcomes of hESC-based therapies in treating human diseases. In 2012, Steven Schwartz and his team reported the first clinical evidence of using hESC-derived retinal pigment epithelium (RPE) in the treatment of Stargardts macular dystrophy, the most common pediatric macular degeneration, and an individual with dry age-related macular degeneration.110,111 With a differentiation efficiency of RPE greater than 99%, 5104 RPEs were injected into the subretinal space of one eye in each patient. As the hESC source of RPE differentiation was exposed to mouse embryonic stem cells, it was considered a xenotransplantation product and required a lower dose of immunosuppression treatment. This study showed that hESCs improved the vision of patients by differentiating into functional RPE without any severe adverse events. The trial was then expanded into two open-label, phase I/II studies with the published results in 2015 supporting the primary findings.112 In these trials, patients were divided into three groups receiving three different doses of hESC-derived RPE, including 10104, 15104 and 50104 RPE cells per eye. After 22 months of follow-up, 19 patients showed improvement in eyesight, seven patients exhibited no improvement, and one patient experienced a further loss of eyesight. The technical challenge of hESC-derived RPE engraftment was an unbalanced proliferation of RPE post administration, which was observed in 72% of treated patients. A similar approach was also conducted in two South Korean patients diagnosed with age-induced macular degeneration and two patients with Stargardt macular dystrophy.113 The results supported the safety of hESC-derived RPE cells and illustrated an improvement in visual acuity in three patients. Recently, clinical-graded hESC-derived RPE cells were also developed by Chinese researchers under xeno-free culture conditions to treat patients with wet age-related degeneration.114 As hESC development is still associated with ethical concerns and immunological complications related to allogeneic administration, hiPSC-derived RPE cells have emerged as a potential cell source for macular degeneration. Although RPE differentiation protocols have been developed and optimized to improve the efficacy of hiPSC-derived RPE cells, they are still insufficient, time-consuming and labor intensive.115,116 For clinical application, an efficient differentiation of primed to nave state hiPSCs toward the RPE was developed using feeder-free culture conditions utilizing the transient inhibition of the FGF/MAPK signaling pathway.117 Overexpression of specific transcription factors in hiPSCs throughout the differentiation process is also an interesting approach to generate a large number of RPE cells for clinical use. In a recent study, overexpression of three eye-field transcription factors, including OTX2, PAX6, and MITF, stimulated RPE differentiation in hiPSCs and generated functional RPE cells suitable for transplantation.118 To date, although reported data from phase I/II clinical trials have been produced enough to support the safety of hESC-derived RPE cells, the treatment is still in its immature stage. Thus, future studies should focus on the development of the cellular manufacturing process of RPE and the subretinal administration route to further improve the outcomes of RPE fabrication and engraftment into the patients retina (recommended review119).

Numerous studies have demonstrated that hESC-derived cardiomyocytes exhibit cardiac transcription factors and display a cardiomyocyte phenotype and immature electrical phenotype. In addition, using hPSC-derived cardiomyocytes could provide a large number of cells required for true remuscularization and transplantation. Thus, these cells can be a promising novel therapeutic approach for the treatment of human cardiovascular diseases. In a case report, hESC-derived cardiomyocytes showed potential therapeutic effects in patients with severe heart failure without any subsequent complications.120 This study was a phase I trial (ESCORT [Transplantation of Human Embryonic Stem Cell-derived Progenitors in Severe Heart Failure] trial) to evaluate the safety of cardiomyocyte progenitor cells derived from hESCs seeded in fibrin gel scaffolds for 10 patients with severe heart failure (NCT02057900). The encouraging results from this study demonstrated the feasibility of producing hESC-derived cardiomyocyte progenitor cells toward clinical-grade standards and combining them with a tissue-engineered scaffold to treat severe heart disease (the first patient of this trial has already reached the 7-year follow-up in October 2021).121 Currently, the two ongoing clinical trials using hPSC-derived cardiomyocytes have drawn great attention, as their results would pave the way to lift the bar for approving therapies for commercial use. The first trial was conducted by a team led by cardiac surgeon Yoshiki Sawa at Osaka University using hiPSC-derived cardiomyocytes embedded in a cell sheet for engraftment (jRCT2052190081). The trials started first with three patients followed by ten patients to assess the safety of the approach. Once safety is met, the treatment can be sold commercially under Japans fast-track system for regenerative medicine.122 Another trial used a collagen-based construct called BioVAT-HF to contain hiPSC-derived cardiomyocytes. The trial was divided into two parts to evaluate the cell dose: (Part A) recruiting 18 patients and (Part B) recruiting 35 patients to test a broad range of engineered human myocardium (EHM) doses. The expected results from this study will provide the proof-of-concept for the use of EHM in the stimulation of heart remuscularization in humans. To date, no adverse events or severe adverse events have been reported from these trials, supporting the safety of the procedure. However, as the number of treated patients was relatively small, limitations in drawing conclusions regarding efficacy are not yet possible.21,123

One of the first clinical trials using hPSC-based therapy was conducted by Geron Corporation in 2010 using hESC-derived oligodendrocyte progenitor cells (OPC1) to treat spinal cord injury (SCI). The results confirmed the safety one year post administration in five participants, and magnetic resonance imaging demonstrated improvement of spinal cord deterioration in four participants.124 Asterias Biotherapeutic (AST) continued the Geron study by conducting the SCiStar Phase I/IIa study to evaluate the therapeutic effects of AST-OPC1 (NCT02302157). The trials results published in clinicaltrials.gov demonstrated significant improvement in running speed, forelimb stride length, forelimb longitudinal deviations, and rear stride frequency. Interestingly, the recently published data of a phase 1, multicentre, nonrandomized, single-group assignment, interventional trial illustrated no evidence of neurological decline, enlarging masses, further spinal cord damage, or syrinx formation in patients 10 years post administration of the OPC1 product.125 This data set provides solid evidence supporting the safety of OPC1 with an event-free period of up to 10 years, which strengthens the safety profile of the SCiStar trial.

Analysis of the global trends in clinical trials using hPSC-based therapy showed that 77.1% of studies were observational (no cells were administered into patient), and only 22.9% of studies used hPSC-derived cells as interventional treatment.126 The number of studies using hiPSCs was relatively higher than that using hESCs, which was 74.8% compared to 25.2%, respectively. The majority of observational studies were performed in developed countries, including the USA (41.6%) and France (16.8%), whereas interventional studies were conducted in Asian countries, including China (36.7%), Japan (13.3%), and South Korea (10%). The trends in therapeutic studies were also clear in terms of targeted diseases. The three most studied diseases were ophthalmological conditions, circulatory disorders, and nervous systems.127 However, it is surprising that the clinical applications of hPSCs have achieved little progress since the first hESCs were discovered worldwide. The relatively low number of clinical trials focusing on using iPSCs as therapeutic agents to administer into patients could be ascribed to the unstable genome of hiPSCs,128 immunological rejection,129 and the potential for tumor formation.130

Approximately 55 years ago, fibroblast-like, plastic-adherent cells, later named mesenchymal stem cells (MSCs) by Arnold L. Caplan,18 were discovered for the first time in mouse bone marrow (BM) and were later demonstrated to be able to form colony-like structures, proliferate, and differentiate into bone/reticular tissue, cartilage, and fat.131 Protocols were subsequently established to directly culture this subpopulation of stromal cells from BM in vitro and to stimulate their differentiation into adipocytes, chondroblasts, and osteoblasts.132 Since then, MSCs have been found in and derived from different human tissue sources, including adipose tissue (AT), the umbilical cord (UC), UC blood, the placenta, dental pulp, amniotic fluid, etc.133 To standardize and define MSCs, the International Society for Cell and Gene Therapy (ISCT) set minimal identification criteria for MSCs derived from multiple tissue sources.134 Among them, MSCs derived from AT, BM, and UC are the most commonly studied MSCs in human clinical trials,135 and they constitute the three major tissue sources of MSCs that will be discussed in this review.

The discovery of MSCs opened an era during which preclinical studies and clinical trials have been performed to assess the safety and efficacy of MSCs in the treatment of various diseases. The major conclusion of these studies and trials is that MSC-based therapy is safe, although the outcomes have usually been either neutral or at best marginally positive in terms of the clinically relevant endpoints regardless of MSC tissue origin, route of infusion, dose, administration duration, and preconditioning.136 It is important to note that a solid background of knowledge has been generated from all these studies that has fueled the recent translational research in MSC-based therapy. As MSCs have been intensively studied over the last 55 years and have become the subject of multiple reviews, systematic reviews, and meta-analyses, the objective of this paper is not to duplicate these publications. Rather, we will discuss the questions that both clinicians and researchers are currently exploring with regard to MSC-based therapy, diligently seeking answers to the following:

With a solid body of data supporting their safety profiles derived from both preclinical and clinical studies, does the tissue origin of MSCs also play a role in their downstream clinical applications in the treatment of different human diseases?

Do MSCs derived from AT, BM, and UC exhibit similar efficacy in the treatment of neurological diseases, metabolic/endocrine-related disorders, reproductive dysfunction, skin burns, lung fibrosis, pulmonary disease, and cardiovascular conditions?

To answer these questions, we will first focus on the most recently published clinical data regarding these targeted conditions, including neurological disorders, pulmonary dysfunctions, metabolic/endocrine-related diseases, reproductive disorders, skin burns, and heart-related diseases, to analyze the potential efficacy of MSCs derived from AT, BM, and UC. Based on the level of clinical improvement observed in each trial, we analyzed the potential efficacy of MSCs derived from each source to visualize the correlation between patient improvement and MSC sources. We will then address recent trends in the exclusive use of MSC-based products, focusing on the efficacy of treatment with MSCs from each of the abovementioned sources, and we will analyze the relationship between the respective efficacies of MSCs from these sources in relation to the targeted disease conditions. Finally, we propose a hypothesis and mechanism to achieve the currently still unmet objective of evaluating the use of MSCs from AT, BM, and UC in regenerative medicine.

In general, MSCs are reported to be isolated from numerous tissue types, but all of these types can be organized into two major sources: adult137 and perinatal sources138 (Fig. ). Adult sources of MSCs are defined as tissues that can be harvested or obtained from an individual, such as dental pulp,139 BM, peripheral blood,140 AT,141 lungs,142 hair,143 or the heart.144 Adult MSCs usually reside in specialized structures called stem cell niches, which provide the microenvironment, growth factors, cell-to-cell contacts and external signals necessary for maintaining stemness and differentiation ability.145 BM was the first adult source of MSCs discovered by Friedenstein131 and has become one of the most documented and largely used MSC sources to date, followed by AT. BM-MSCs are isolated and cultured in vitro from BM aspirates using a Ficoll gradient-centrifugation method146 or a red blood cell lysate buffer to collect BM mononuclear cell populations, whereas AT-MSCs are obtained from stromal vascular fractions of enzymatically digested AT obtained through liposuction,141 lipoplasty, or lipectomy procedures.147 These tissue collection procedures are invasive and painful for the patient and are accompanied by a risk of infection, although BM aspiration and adipose liposuction are considered safe procedures for BM and AT biopsies. The number of MSCs that can be isolated from these adult tissues varies significantly in a tissue-dependent manner. The percentage of MSCs in BM mononuclear cells ranges from 0.001 to 0.01% following gradient centrifugation.132 The number of MSCs in AT is at least 500 times higher than that in BM, with approximately 5,000 MSCs per 1g of AT. Perinatal sources of MSCs consist of UC-derived components, such as UC, Whartons jelly, and UC blood, and placental structures, such as the placental membrane, amnion, chorion membrane, and amniotic fluid.138 The collection of perinatal MSCs, such as UC-MSCs, is noninvasive, as the placenta, UC, UC blood, and amnion are considered waste products that are usually discarded after birth (with no ethical barriers).148 Although MSCs represent only 107% the cells found in UC, their higher proliferation rate and rapid population doubling time allow these cells to rapidly replicate and increase in number during in vitro culture.149 Under standardized xeno-free and serum-free culture platforms, AT-MSCs show a faster proliferation rate and a higher number of colony-forming units than BM-MSCs.149 UC-MSCs have the fastest population doubling time compared to AT-MSCs and BM-MSCs in both conventional culture conditions and xeno- and serum-free environments.149 MSCs extracted from AT, BM and UC exhibit all minimal criteria listed by the ISCT, including morphology (plastic adherence and spindle shape), MSC surface markers (95% positive for CD73, CD90 and CD105; less than 2% negative for CD11, CD13, CD19, CD34, CD45, and HLR-DR) and differentiation ability into chondrocytes, osteocytes, and adipocytes.150

The two major sources of MSCs: adult and perinatal sources. The adult sources of MSCs are specific tissue in human body where MSCs could be isolated, including bone marrow, adipose tissue, dental pulp, peripheral blood, menstrual blood, muscle, etc. The perinatal sources of MSCs consist of umbilical cord-derived components, such as umbilical cord, Whartons jelly, umbilical cord blood, and placental structures, such as placental membrane, amnion, chorion membrane, amniotic fluid, etc. The figure was created with BioRender.com

In fact, although MSCs derived from either adult or perinatal sources exhibit similar morphology and the basic characteristics of MSCs, studies have demonstrated that these cells also differ from each other. Regarding immunophenotyping, AT-MSCs express high levels of CD49d and low levels of Stro-1. An analysis of the expression of CD49d and CD106 showed that the former is strongly expressed in AT-MSCs, in contrast to BM-MSCs, whereas CD106 is expressed in BM-MSCs but not in AT-MSCs.151 Increased expression of CD133, which is associated with stem cell regeneration, differentiation, and metabolic functions,152 was observed in BM-MSCs compared to MSCs from other sources.153 A recent study showed that CD146 expression in UC-MSCs was higher than that in AT- and BM-MSCs,153 supporting the observation that UC-MSCs have a stronger attachment and a higher proliferation rate than MSCs from other sources, as CD146 is a key cell adhesion protein in vascular and endothelial cell types.154 In terms of differentiation ability, donor-matched BM-MSCs exhibit a higher ability to differentiate into chondrogenic and osteogenic cell types than AT-MSCs, whereas AT-MSCs show a stronger capacity toward the adipogenic lineage.150 The findings from an in vitro differentiation study indicated that BM-MSCs are prone to osteogenic differentiation, whereas AT-MSCs possess stronger adipogenic differentiation ability, which can be explained by the fact that the epigenetic memory obtained from either BM or AT drives the favored MSC differentiation along an osteoblastic or adipocytic lineage.155 Interestingly, although UC-MSCs have the ability to differentiate into adipocytes, osteocytes, or chondrocytes, their osteogenic differentiation ability has been proven to be stronger than that of BM-MSCs.156 The most interesting characteristic of MSCs is their immunoregulatory functions, which are speculated to be related to either cell-to-cell contact or growth factor and cytokine secretion in response to environmental/microenvironmental stimuli. MSCs from different sources almost completely inhibit the proliferation of peripheral blood mononuclear cells (PBMCs) at PBMC:MSC ratios of 1:1 and 10:1.149 At a higher ratio, BM-MSCs showed a significantly higher inhibitory effect than AT- or UC-MSCs.153 Direct analysis of the immunosuppressive effects of BM- and UC-MSCs has revealed that these cells exert similar inhibitory effects in vitro with different mechanisms involved.157 With these conflicting data, the mechanism of action related to the immune response of MSCs from different sources is still poorly understood, and long-term investigations both in preclinical studies and in clinical trial settings are needed to shed light on this complex immunomodulation function.

The great concern in MSC-based therapy is the fate of these cells post administration, especially through different delivery routes, including systemic administration via an intravenous (IV) route or tissue-specific administration, such as dorsal pancreatic administration. It is important to understand the distribution of these cells after injection to expand our understanding of the underlying mechanisms of action of treatments; in addition, this knowledge is required by authorized bodies (the Food and Drug Administration (FDA) in the United States or the regulation of advanced-therapy medicinal products in Europe, No. 1394/2007) prior to using these cells in clinical trials. The preclinical data using various labeling techniques provide important information demonstrating that MSCs do not have unwanted homing that could lead to the incorrect differentiation of the cells or inappropriate tumor formation. In a mouse model, human BM-MSCs and AT-MSCs delivered via an IV route are rapidly trapped in the lungs and then recirculate through the body after the first embolization process, with a small number of infused cells found mainly in the liver after the second embolization.158 Using the technetium-99 m labeling method, intravenously infused human cells showed long-term persistence up to 13 months in the bone, BM compartment, spleen, muscle, and cartilage.159 A similar result was reported in baboons, confirming the long-term homing of human MSCs in various tissues post administration.160 Although the retainment of MSCs in the lungs might potentially reduce their systemic therapeutic effects,161 it provides a strong advantage when these cells are used in the treatment of respiratory diseases. Local injection of MSCs also revealed their tissue-specific homing, as an injection of MSCs via the renal artery route resulted in the majority of the injected cells being found in the renal cortex.162 Numerous studies have been conducted to track the migration of administered MSCs in human subjects. Henriksson and his team used MSCs labeled with iron sucrose in the treatment of intervertebral disc degeneration.163 Their study showed that chondrocytes differentiated from infused MSCs could be detected at the injured intervertebral discs at 8 months but not at 28 months. A study conducted in a patient with hemophilia A using In-oxine-labeled MSCs showed that the majority of the cells were trapped in the lungs and liver 1h post administration, followed by a reduction in the lungs and an increase in the number of cells in the liver after 6 days.164 Interestingly, a small proportion of infused MSCs were found in the hemarthrosis site at the right ankle after 24h, suggesting that MSCs are attracted and migrate to the injured site. The distribution of MSCs was also reported in the treatment of 21 patients diagnosed with type 2 diabetes using 18-FDG-tagged MSCs and visualized using positron emission tomography (PET).165 The results illustrated that local delivery of MSCs via an intraarterial route is more effective than delivery via an IV route, as MSCs home to the pancreatic head (pancreaticoduodenal artery) or body (splenic artery). Therefore, although the available data related to the biodistribution of infused MSCs are still limited, the results obtained from both preclinical and clinical studies illustrate a comparable set of data supporting results on homing, migration to the injured site, and the major organs where infused MSCs are located. The following comprehensive and interesting reviews are highly recommended.166168

To date, 1426 registered clinical trials spanning different trial phases have used MSCs for therapeutic purposes, which is four times the number reported in 2013.169,170 As supported by a large body of preclinical studies and advancements in conducting clinical trials, MSCs have been proven to be effective in the treatment of numerous diseases, including nervous system and brain disorders, pulmonary diseases,171 cardiovascular conditions,172 wound healing, etc. The outcomes of MSC-based therapy have been the subject of many intensive reviews and systematic analyses with the solid conclusion that these cells exhibit strong safety profiles and positive outcomes in most tested conditions.173175 In addition, the available data have revealed several potential mechanisms that could explain the beneficial effects of MSCs, including their homing efficiency, differentiation potential, production of trophic factors (including cytokines, chemokines, and growth factors), and immunomodulatory abilities. However, it is still not known which MSC types should be used for which diseases, as it seems to be that MSCs exhibit beneficial effects regardless of their sources.169

The theory that brain cells can never regenerate has been challenged by the discovery of newly formed neurons in the human adult hippocampus or the migration of stem cells in the brain in animal models.176 These observations have triggered hope for regeneration in the context of neuronal diseases by using exogenous stem cell sources to replenish or boost the stem cell population in the brain. Moreover, the limited regenerative capacity of the brain and spinal cord is an obstacle for traditional treatments of neurodegenerative diseases, such as autism, cerebral palsy, stroke, and spinal cord injury (SCI). As current treatments cannot halt the progression of these diseases, studies throughout the world have sought to exploit cell-based therapies to treat neurodegenerative diseases on the basis of advances in the understanding and development of stem cell technology, including the use of MSCs. Successful stem cell therapy for treating brain disease requires therapeutic cells to reach the injured sites, where they can repair, replace, or at least prevent the deteriorative effects of neuronal damage.177 Hence, the gold standard of cell-based therapy is to deliver the cells to the target site, stimulate the tissue repair machinery, and regulate immunological responses via either cell-to-cell contact or paracrine effects.178 Among 315 registered clinical trials using stem cells for the treatment of brain diseases, MSCs and hematopoietic stem cells (HSCs; CD34+ cells isolated from either BM aspirate or UC blood) are the two main cell types investigated, whereas approximately 102 clinical trials used MSCs and 62 trials used HSCs for the treatment of brain disease (main search data from clinicaltrial.gov). MSCs are widely used in almost all clinical trials targeting different neuronal diseases, including multiple sclerosis,179 stroke,180 SCI,181 cerebral palsy,182 hypoxic-ischemic encephalopathy,183 autism,184 Parkinsons disease,185 Alzheimers disease185 and ataxia. Among these trials in which MSCs were the major cells used, nearly two-thirds were for stroke, SCI, or multiple sclerosis. MSCs have been widely used in 29 registered clinical trials for stroke, with BM-MSCs being used in 16 of these trials. With 26 registered clinical trials, SCI is the second most common indication for using MSCs, with 16 of these trials using mainly expanded BM-MSCs. For multiple sclerosis, 15 trials employed BM-MSCs among a total of 23 trials conducted for the treatment of this disease. Hence, it is important to note that in neuronal diseases and disorders, BM-MSCs have emerged as the most commonly used therapeutic cells among other MSCs, such as AT-MSCs and UC-MSCs.

The outcomes of the use of BM-MSCs in the treatment of neuronal diseases have been widely reported in various clinical trial types. A review by Zheng et al. indicated that although the treatments appeared to be safe in patients diagnosed with stroke, there is a need for well-designed phase II multicentre studies to confirm the outcomes.173 One of the earliest trials using autologous BM-MSCs was conducted by Bang et al. in five patients diagnosed with stroke in 2005. The results supported the safety and showed an improved Barthel index (BI) in MSC-treated patients.186 In a 2-year follow-up clinical trial, 16 patients with stroke received BM-MSC infusions, and the results showed that the treatment was safe and improved clinical outcomes, such as motor impairment scale scores.187 A study conducted in 12 patients with ischemic stroke showed that autologous BM-MSCs expanded in vitro using autologous serum improved the patients modified Rankin Scale (mRS), with a mean lesion volume reduced by 20% at 1 week post cell infusion.188 In 2011, a modest increase in the Fugl Meyer and modified BI scores was observed after autologous administration of BM-MSCs in patients with chronic stroke.189 More recently, a prospective, open-label, randomized controlled trial with blinded outcome evaluation was conducted, with 39 patients and 15 patients in the BM-MSC administration and control groups, respectively. The results of this study indicated that autologous BM-MSCs with autologous serum administration were safe, but the treatment led to no improvements at 3 months in modified Rankin Scale (mRS) scores, although leg motor improvement was observed.180 Researchers explored whether the efficacy of BM-MSC administration was maintained over time in a 5-year follow-up clinical trial. Patients (85) were randomly assigned to either the MSC group or the control group, and follow-ups on safety and efficacy were performed for 5 years, with 52 patients being examined at the end of the study. The MSC group exhibited a significant improvement in terms of decreased mRS scores, whereas the number of patients with an mRS score increase of 03 was statistically significant.187 Although autologous BM-MSCs did not improve the Basel index, mRS, or National Institutes of Health Stroke Scale (NIHSS) score 2 years post infusion, patients who received BM-MSC therapy showed improvement in their motor function score.190 In addition, a prospective, open-label, randomized controlled trial by Lee et al. showed that autologous BM-MSCs primed with autologous ischemic serum significantly improved motor functions in the MSC-treated group. Neuroimaging analysis also illustrated a significant increase in interhemispheric connectivity and ipsilesional connectivity in the MSC group.191 Recently, a single intravenous infection of allogeneic BM-MSCs has been proven to be safe and feasible in patients with chronic stroke with a significant improvement in BI score and NIHSS score.192

In two systematic reviews using MSCs for the treatment of SCI, BM-MSCs (n=16) and UC-MSCs (n=5) were reported to be safe and well-tolerated.193,194 The results indicated significant improvements in the stem cell administration groups compared with the control groups in terms of a composite of the American Spinal Injury Association (ASIA) impairment scale (AIS) grade, AIS grade A, and ASIA sensory scores and bladder function (Table ). However, larger experimental groups with a randomized and multicentre design are needed for further confirmation of these findings. For multiple sclerosis, several early-phase (phase I/II) registered clinical studies have used BM-MSCs. A study compared the potential efficacy of BM-MSC and BM mononuclear cell (BMMNC) transplantation in 105 patients with spastic cerebral palsy.195 The results showed that the GMFM (gross motor function measure) and the FMFM (fine motor function measure) scores of the BM-MSC transplant group were higher than those of the BMNNC transplant group at 3, 6, and 12 months of assessment. In terms of autism spectrum disorder, a review of 254 children after BMMNC transplantation found that over 90% of patients ISAA (Indian Scale for Assessment of Autism) and CARS (Childhood Autism Rating Scale) scores improved. Young patients and those in whom autism spectrum disorder was detected early generally showed better improvement.196

The reported clinical trials using MSCs from AT, BM, and UC in the treatment of brain-related injuries and neurological disorders

One of the biggest limitations when using BM-MSCs is the bone marrow aspiration process, as it is an invasive procedure that can introduce a risk of complications, especially in pediatric and elderly patients.197 Therefore, UC-MSCs have been suggested as an alternative to BM-MSCs and are being studied in clinical trials for the treatment of neurological diseases in approximately 1550 patients throughout the world; however, only three studies have been completed, with data published recently.198 A recent study showed that UC-MSC administration improved both gross motor function and cognitive skills, assessed using the Activities of Daily Living (ADL), Comprehensive Function Assessment (CFA), and GMFM, in patients diagnosed with cerebral palsy. The improvements peaked 6 months post administration and lasted for 12 months after the first transplantation.199 In a single-targeted phase I/II clinical trial using UC-MSCs for the treatment of autism, Riordan et al. reported decreases in Autism Treatment Evaluation Checklist (ATEC) and CARS scores for eight patients, but the paper has been retracted due to a violation of the journals guidelines.200 In an open-label, phase I study, UC-MSCs were used as the main cells to treat 12 patients with autism spectrum disorder via IV infusions. It is important to note that five participants developed new class I anti-human leukocyte antigen in response to the specific lot of manufactured UC-MSCs, although these responses did not exhibit any immunological response or clinical manifestations. Only 50% of participants showed improvements in at least two autism-specific measurements.201 Although not as widely used as BM-MSCs, these trials have demonstrated the efficacy of using UC-MSCs in the treatment of SCIs. In a pilot clinical study, Yang et al. showed that the use of UC-MSCs has the potential to improve disease status through an increase in total ASIA and SCI Functional Rating Scale of the International Association of Neurorestoratology (IANR-SCIFRS) scores, as well as an improvement in pinprick, light touch, motor and sphincter scores.202 A study of 22 patients with SCIs showed a potential therapeutic effect in 13 patients post UC-MSC infusion.203 AT-MSCs were also used to treat SCI, with a single case report indicating an improvement in neurological and motor functions in a domestic ferret patient.204 However, a result obtained from another phase I trial using AT-MSCs showed mild improvements in neurological function in a small number of patients.205 A phase II, randomized, double-blind, placebo-controlled, single-center, pilot clinical trial using AT-MSCs in the treatment of acute ischemic stroke published a data set that supports the safety of the therapy, although patients who received AT-MSCs showed a nonsignificant improvement after 24 months of follow-up.206 In all of the above studies, the safety of using either AT-MSCs or UC-MSCs was evaluated, and no significant reactions were reported after infusion.

Therefore, based on the number of recovered patients post-transplantation and the number of recruited patients in large-scale trials using BM-MSCs, it seems that BM-MSCs are the prominent cells in regard to treating neurodegenerative disease with potentially good outcomes (Table ). It is important to note that we do not negate the fact that AT- and UC-MSCs also show positive outcomes in the treatment of neuronal diseases, with numerous ongoing large-scale, multicentre, randomized, and placebo-control trials,207,208 but we suggest alternative and thoughtful decisions regarding which sources of MSCs are best for the treatment of neuronal diseases and degenerative disorders.

In the last decade, significant increases in respiratory disease incidence due to air pollution, smoking behavior, population aging, and recently, respiratory virus infections such as coronavirus disease 2019 (COVID-19)209 have been observed, leading to substantial burdens on public health and healthcare systems worldwide. Respiratory inflammatory diseases, including bronchopulmonary dysplasia (BPD), chronic obstructive pulmonary disease (COPD), and acute respiratory distress syndrome (ARDS), have recently emerged as three prevalent pulmonary diseases in children and adults. These conditions are usually associated with inflammatory cell infiltration, a disruption of alveolar structural integrity, a reduction in alveolar fluid clearance ability, cytokine release and associated cytokine storms, airway remodeling, and the development of pulmonary fibrosis. Traditional treatments are focused on relieving symptoms and preventing disease progression using surfactants, artificial respiratory support, mechanical ventilation, and antibiotic/anti-inflammatory drugs, with limited effects on the damaged airway, alveolar fluid clearance, and other detrimental effects caused by the inflammatory response. MSCs are known for their immunomodulatory abilities, showing potential in injury reduction and aiding lung recovery after injury. According to ClinicalTrials.gov, from 2017 to date, there have been 159 studies testing the application of MSCs in the treatment of pulmonary diseases, including but not limited to BPD, COPD, and ARDS, suggesting a trend in the use of MSCs as an alternative approach for the treatment of respiratory diseases, especially MSCs from UC as an off-the-shelf and allogeneic source.

Extremely premature infants are born with arrested lung development at the canalicular-saccular phases prior to alveolarization and before pulmonary maturation occurs, which results in the development of BPD.210 These infants require intensive care during the first three months of life using postnatal interventions, including positive pressure mechanical ventilation, external oxygen support, and surfactant infusions, and the newborns have recurrent infections that further compromise normal lung development.211 To date, 13 clinical trials have been proposed to use UC-MSCs in the treatment of BPD, recruiting ~566 premature infants throughout the world, including Vietnam, Korea, the United States, Spain, Australia, and China. The majority of these trials use UC-derived stem cells for phases I and II, focusing on evaluating the safety and efficacy of stem cell-based therapy.212 Human UC tissue and its derivative components are considered the most attractive cell sources for MSCs in the treatment of BPD due to the ease of obtaining them, being readily available, with no ethical concerns, low antigenicity, a high cell proliferation rate, and superior regenerative potential. Chang et al. used MSCs derived from UC blood in a phase I dose-escalation clinical trial to treat 9 preterm infants via intratracheal administration to prevent the development of BPD.213 All 9 preterm infants survived, and only three developed BPD; these infants had significantly decreased BPD severity compared with the historically matched control group. A follow-up study of the same patients after 24 months indicated that only one infant had an E. cloacae infection after discharge at 4 months, with subsequent disseminated intravascular coagulation, which was later proven to be unrelated to the intervention. The remaining eight patients survived with normal pulmonary development and function, suggesting that the therapy was safe. MSCs from UC blood were also used for the treatment of 12 extremely low birthweight preterm patients using the same administration route, which further confirmed the safety of the therapy in the treatment of BPD, although ten of 12 infants still developed severe BPD at 36 weeks.214 Our group also reported the safety and potential efficacy of using UC-MSCs in the treatment of four preterm infants, and the results supported the safety of UC-MSCs and demonstrated that patients could be weaned from oxygen supply and develop normal lung structure and function.215 A phase II clinical trial of 66 infants born at 2328 weeks with a birthweight of 5001250g who were recruited and randomized into an MSC-administration group and a control group was conducted. Although the results supported the safety of MSC administration in preterm infants, the efficacy of the treatment was not supported by statistical analysis, potentially due to the small sample size. Subgroup analysis showed that patients with severe BPD born at 2324 weeks showed a significant improvement in BPD severity, but those born at 2528 weeks did not.216 Hence, it is important to conduct controlled phase II clinical trials with larger cohort sizes to further substantiate the efficacy of UC blood-derived MSCs in the treatment of infants with BPD.

With more than 65 million patients worldwide, COPD was the third-leading cause of death in 2020, according to World Health Organization records. COPD is classified as a chronic inflammatory and destructive pulmonary disease characterized by a progressive reduction in lung function. Averyanov et al. performed a randomized, placebo-controlled phase I/IIa study in 20 patients with mild to moderate idiopathic pulmonary fibrosis (IPF). Treatment group patients received two IV doses of allogeneic MSCs (2108cells) every 3 months, and the second group received a placebo.217 Evaluation tests were performed at weeks 13, 26, 39, and 52. The 6-min walking test distance (6MWTD) results showed that patient fitness improved from week 13 onwards and was maintained until up to the 52nd week. Pulmonary function indicators improved markedly before and after treatment in the treated group but did not change significantly in the placebo group. The goal of MSC therapy in the treatment of COPD is to promote the regeneration of parenchymal cells and alveolar structure and the restoration of lung function. Based on the results of a phase I trial of a commercial BM-MSC product, ProchymalTM, which led to improvements in pulmonary function in treated patients, a multicentre, double-blind, placebo-controlled phase II trial was conducted in 62 patients diagnosed with COPD to determine the safety and potential efficacy of the product. Although the results supported the safety of BM-MSCs, their effectiveness in the treatment of COPD was not assured. No statistically significant differences in FEV1 or FEV1%, total lung capacity, or carbon monoxide diffusing capacity were detected after 2 years of follow-up between the two treatment groups. To date, there have been five clinical trials using BM-MSCs as the main stem cells for the treatment of COPD, but the overall clinical outcomes did not demonstrate the potential therapeutic effects of the treatment.218222 In clinical trial NCT001110252, the results showed that there was an overall reduction in the process of COPD pathological development 3 years after the administration of BM-MSCs, although the trial had a phase I design, with no control group, and evaluated only a small cohort (four patients).219 To alleviate local inflammatory progression in COPD, Oliveira et al. studied the combination treatment of one-way endobronchial valve (EBV) and BM-MSC intubation.223 Ten GOLD (Global Initiative for Obstructive Lung Disease) stage C or D patients were equally divided into 2 groups: one group received a dose of 108 cells before valve insertion, and the other group received a normal saline infusion. The follow-up time was 90 days. Inflammation was significantly improved as assessed by the CRP (C-reactive protein) index at 30 and 90 days after infusion. In addition, improvements in St. Georges Respiratory Questionnaire (SGRQ) scores indicated improved patient quality of life. Furthermore, an investigation into the homing ability of MSCs in vivo was performed on 9 GOLD patients, from stage A to stage D. Each patient received two 2106 BM-MSC/kg IV infusions 1-week apart.224 The marking of MSCs with indium-111 showed that MSCs were retained in the pulmonary vasculature longer in patients with mild COPD and that the levels of inflammatory mediators improved after 7 days of treatment. The results of the evaluation survey conducted after 1 year showed that the number of COPD exacerbations decreased to six times/year compared to 11 times/year before treatment. In addition, AT-MSCs present in the stromal vascular fraction were used to treat patients with COPD, and no adverse events were observed after 12 months of follow-up, but the clinical improvements post administration were not clear.225 The results from a phase I clinical trial using AT-MSCs in eight patients with COPD also reported no significant change in pulmonary function test parameters.226 A study evaluating the use of AT-MSCs as adjunctive therapy for COPD in 12 patients was performed.227 AT was obtained using standard liposuction, MSCs were isolated, and 150300 million cells were intravenously infused. The patients showed improvements in quality of life, with improved SGRQ scores after 3 and 6 months of treatment. Recently, UC-MSCs have emerged as potential allogeneic stem cell candidates for the treatment of COPD.228 In a pilot clinical study, it was demonstrated that allogeneic administration of UC-MSCs in the treatment of COPD was safe and potentially effective.229 In one study, 20 patients, including 9 at stage C and 11 at stage D per the GOLD classification, with histories of smoking were recruited and received cell-based therapy. The patients who received UC-MSC treatment showed significant reductions in Modified Medical Research Council scores, COPD assessment test scores, and the number of pulmonary exacerbations 6 months post administration. The results of the second trial using UC-MSCs showed that the mean FEV1/FVC ratios were increased along with improvements in SGRQ scores and 6MWTDs at three months post administration.230 Although thorough assessments of the effectiveness of UC-MSCs are still in the early stages, the number of trials using UC-MSCs for the treatment of COPD is increasing steadily, with larger sample sizes and stronger designs (randomized or matched casecontrol studies), providing a data set strongly supporting the future applications of UC-MSCs.231

The ongoing pandemic of the 21st century, the COVID-19 pandemic, emerged as a major pulmonary health problem worldwide, with a relatively high mortality rate. Numerous studies, reviews, and systematic analyses have been conducted to discuss and expand our knowledge of the virus and propose different mechanisms by which the virus could alter the immune system.232 One of the most critical mechanisms is the generation of cytokine storms, which result from the initiation of hyperreactions of the adaptive immune response to viral infection.233 These cytokine storms are formed by the establishment of waves of hypercytokinaemia generated from overreactive immune cells, which enhance their expression of TNF-, IL-6, and IL-10, preventing T-lymphocyte recruitment and proliferation and culminating in T-lymphocyte apoptosis and T-cell exhaustion. In COVID-19, once a cytokine storm is formed, it spreads from an initial focal area through the body via circulation, which has been discussed in a comprehensive review by Jamilloux et al.234 At the time of writing this review, there were 74 clinical trials using MSCs from UC (29 trials; including WJ-derived MSCs (WJ-MSCs) and placenta-derived MSCs (PL-MSCs)), AT (15 trials), and BM (11 trials) (comprehensive review171,235). Hence, UC-MSCs have emerged as the most common MSCs for the treatment of COVID-19, with a total of 1047 patients participating in these trials. Among these trials, 15 completed trials using UC-MSCs (including WJ- and PL-MSCs) have been reported, with clinical data from approximately 600 recruited patients.232 Eight of these 15 studies used allogenic UC-MSC transplantation to treat critically ill patients.236 A list of case reports using UC-MSCs showed that the treatments were safe and well-tolerated in 14 patients with COVID-19, with the primary outcomes including increased percentages and numbers of T cells,237,238 improved respiratory and renal functions,239 reductions in inflammatory biomarker levels,240 and positive outcomes in the PaO2/FiO2 ratio.240 In a pilot study conducted in ten patients with severe COVID-19, a single dose of UC-MSCs was safe and improved clinical outcomes, although the study did not investigate whether multiple doses of UC-MSCs could further improve the outcomes.241 Two trials without a control group were conducted in 47 patients, and the results indicated that UC-MSCs were safe and feasible for the treatment of patients with COVID-19.235,242 A single-center, open-label, individually randomized, standard treatment-controlled trial was performed in 41 patients (12 patients assigned to the UC-MSC group), and the results showed that significant improvements in C-reactive protein levels, IL-6 levels, oxygen indices, and lymphocyte numbers were found in the MSC groups. Chest computed tomography (CT) illustrated significant reductions in lung inflammatory responses as reflected by CT findings, the number of lobes involved, and pulmonary consolidation.238 In a phase I trial conducted in 18 hospitalized patients with COVID-19, UC-MSCs were administered via an IV route in nine patients (five patients with moderate COVID-19 and 4 patients with severe COVID-19) at days 0, 3, and 6, with no treatment-related adverse events or severe adverse events.243 Only one patient in the UC-MSC group required mechanical ventilation, compared to four patients in the control group. However, the clinical outcomes, such as COVID-19 symptoms, laboratory test results, CT findings of lung damage, and pulmonary function test parameters, were improved in both groups. Interestingly, a 1-year follow-up of the same sample revealed that the patients who received UC-MSC administration improved in terms of whole-lung lesion volume compared to the control group.244 Moreover, chest CT at 12 months showed significant regeneration of lung tissue in the MSC-administered groups, whereas lung fibrosis was found in all patients in the control group. This finding is of interest because it indicates that a long time is needed to detect the regenerative functions of MSC-based therapy, as the biological process to enhance lung tissue regeneration occurs relatively slowly and requires multiple steps. The effects of UC-MSCs in the attenuation and prevention of the development of cytokine storms were illustrated in an interventional, prospective, three-parallel arm study with two control arms conducted in 30 patients in moderate and critical clinical conditions.245 The results indicated a significant decrease in proinflammatory cytokines (IFN, IL-6, IL-17A, IL-2, and IL-12) and an increase in anti-inflammatory cytokines (IL-10, IL-13, and IL-1ra), suggesting that UC-MSCs might participate in the prevention of cytokine storm development. Lanzoni et al. performed a double-blind, randomized, controlled trial and found that UC-MSC infusions significantly decreased cytokine levels at day 6 and improved survival in patients with COVID-19 with ARDS. In this trial, 24 patients were randomized and assigned 1:1 to receive either MSCs or placebo.246 MSC treatment was associated with a significant improvement in the survival rate without serious adverse events. To date, other trials conducted using UC-MSCs as the main MSCs provide a solid data set on their safety and efficacy in preventing the development of cytokine storms, reducing the inflammatory response, improving pulmonary function, reducing intensive care unit (ICU) stay duration, enhancing lung tissue regeneration, and reducing lung fibrosis progression.240,247249 In two large cohort studies (phase I with 210 patients and phase II with 100 patients), the volume of lung lesions and solid component injuries of patients lungs were reduced significantly after the administration of UC-MSCs,250 and clinical symptoms and inflammatory levels were improved.251 Of the 26 reported clinical trials for the treatment of COVID-19 with MSCs, 1 study used AT-MSCs as the main MSCs.236 Thirteen COVID-19 adult patients under invasive mechanical ventilation who had received previous antiviral and/or anti-inflammatory treatments (including steroids, lopinavir/ritonavir, hydroxychloroquine, and/or tocilizumab, among others) were treated with allogeneic AT-MSCs. With a mean follow-up time of 16 days after infusion, 9/13 patients clinical symptoms improved, and 7/13 patients were intubated. A decrease in inflammatory cytokines and an increase in immunoregulatory cells were also observed in patients, especially in the group of patients with overall clinical improvement. Although there is a lack of clinical efficacy data supporting the use of AT-MSCs in the treatment of patients with COVID-19, AT-MSCs are still potential candidates for inhibiting COVID-19 due to their high secretory activity, strong immune-modulatory effects, and homing ability.252254

For ARDS, in a phase IIa trial, 60 patients with moderate to severe disease were randomized into 2 groups. A group of 40 patients received a single infusion of BM-MSCs at a dose of 1106 cells/kg body weight, and another 20 patients received a placebo.255 After 6 and 24h of infusion, the decrease in plasma inflammatory cytokine levels in the MSC group was significantly greater than that in the placebo group. For severe pulmonary hypertension (PH) associated with BPD (BPD-PH), in a small trial, two preterm infants born at 2627 weeks of age were intravenously administered heterologous BM-MSCs at a dose of 5106 cells per kg of body weight; the treatment reduced oxygen requirements and supported respiration in the infants.256 The administration of allogeneic AT-MSCs in the treatment of ARDS appeared to be safe and well-tolerated in 12 adult patients, but clinical outcomes were not observed.257 The results of two patients who received BM-MSCs showed that both patients had improved respiratory function and hemodynamic function and a reduction in multiorgan failure.258 Although the safety of BM-MSCs was confirmed in a multicentre, open-label, dose-escalation, phase I clinical trial (The Stem cells for ARDS treatmentSTART trial),259 no significant improvements were found in a phase II trial, including in respiratory function and ARDS conditions.260 The safety profile of UC-MSCs is also supported by the findings of a previous phase I clinical trial conducted in 9 patients, which showed that a single IV administration of UC-MSCs was safe and led to positive outcomes in terms of respiratory function and a reduction in the inflammatory response.261 The findings of this study were also supported by those of the REALIST (Repair of Acute Respiratory Distress with Stromal Cell Administration) trial, which further confirmed the maximum tolerated dose of allogeneic UC-MSCs in patients with moderate to severe ARDS.262

Although AT- and BM-MSCs have demonstrated therapeutic potential with similar mechanisms of action, UC-MSCs have emerged as potential candidates in the treatment of pulmonary diseases due to their ease of production as off-the-shelf products, rapid proliferation, noninvasive isolation methods, and supreme immunological regulation as well as anti-inflammatory effects.263 However, it is important to note that there is a need to conduct phase III clinical trials with larger cohorts and trials with at least two sources of MSCs in the treatment of pulmonary conditions to further confirm this speculation.264 Table summarizes several clinical trials with published results discussed in this review.

The reported clinical trials using MSCs from AT, BM, and UC in the treatment of respiratory diseases

The human body maintains function and homeostatic regulation via a complex network of endocrine glands that synthesize and release a wide range of hormones. The endocrine system regulates body functions, including heartbeat, bone regeneration, sexual function, and metabolic activity. Endocrine system dysregulation plays a vital role in the development of diabetes, thyroid disease, growth disorder, sexual dysfunction, reproductive malfunction, and other metabolic disorders. The central dogma of regenerative medicine is the use of adult stem cells as a footprint for tissue regeneration and organ renewal. The functions of these stem cells are tightly regulated by microenvironmental stimuli from the nervous system (rapid response) and endocrine signals via hormones, growth factors, and cytokines. This harmonized and orchestrated system creates a symphony of signals that directly regulate tissue homeostasis and repair after injury. The disruption of these complex networks results in an imbalance of tissue homeostasis and regeneration that can lead to the development of endocrine disorders in humans, such as diabetes, sexual hormone deficiency, premature ovarian failure (POF), and Asherman syndrome.

In recent years, obesity and diabetes (type 1 diabetes mellitus (T1DM) and type 2 diabetes mellitus (T2DM)) have been the two biggest challenges in endocrinology research, and the application of MSCs has emerged as a novel approach for therapeutic consideration. T1DM is characterized by the autoimmune destruction of pancreatic -cells, whereas T2DM is defined as a combination of insulin resistance and pancreatic insulin-producing cell dysfunction. Regenerative medicine seeks to provide an exogenous cell source for replacing damaged or lost -cells to achieve the goal of stabilizing patients blood glucose levels. To date, there are 28 clinical trials using MSCs in the treatment of T1DM (http://www.clinicaltrials.gov, searched in October 2021), among which three trials were completed using autologous BM-MSCs (NCT01068951), allogeneic BM-MSCs (NCT00690066), and allogeneic AT-MSCs (NCT03920397). Interestingly, UC-MSCs were the most favored MSCs for the remaining trials. All published studies confirmed the safety of MSC therapy in the treatment of T1DM with no adverse events. The first study using autologous BM-MSCs showed that patients who were randomized into the MSC-administration group showed an increase in C-peptide levels in response to a mixed-meal tolerance test (MMTT) in comparison to the control group.265 Unfortunately, there was no significant improvement in C-peptide levels, HbA1C or insulin requirements. The use of autologous AT-MSCs in combination with vitamin D was safe and improved HbA1C levels 6 months post administration.266 WJ-MSCs were used as the main MSCs for the treatment of new-onset T1DM, which showed a significant improvement in both HbA1C and C-peptide levels when compared to those of the control group at three and six months post administration.267,268 The combination of allogeneic WJ-MSCs with autologous BM-derived mononuclear cells improved insulin secretion and reduced insulin requirements in patients with T1DM.269 In terms of T2DM, 23 studies were registered on clinicaltrials.gov (searched in October 2021), with six completed studies (three studies used BM-MSCs and three studies used allogeneic UC-MSCs). Although the number of studies using MSCs for the treatment of T2DM is small, their findings support the safety of MSCs, with no severe adverse events observed during the course of these studies.270 It was confirmed that MSC therapy potentially reduced fasting blood glucose and HbA1C levels and increased C-peptide levels. However, these effects were short-term, and multiple doses were required to maintain the MSC effects. Interestingly, the autologous MSC approach in the treatment of patients with diabetes in general is hampered, as both BM-MSCs and AT-MSCs isolated from patients with diabetes showed reduced stemness and functional characteristics.271,272 In addition, the durations of diabetes and obesity are strongly associated with autologous BM-MSC metabolic function, especially mitochondrial respiration, and the accumulation of mitochondrial DNA, which directly interfere with the functions of BM-MSCs and reduce the effectiveness of the therapy.271 Therefore, the allogeneic approach using MSCs from healthy donors provides an alternative approach for stem cell therapy in the treatment of patients with diabetes.

Modern society is increasingly facing the problem of infertility, which is defined as the inability to become pregnant after more than 1 year of unprotected intercourse.273 This problem has emerged as an important worldwide health issue and social burden. Assisted reproductive techniques and in vitro fertilization technology have recently become the most effective methods for the treatment of infertility in humans, but the use of these approaches is limited, as they cannot be applied in patients with no sperm or those who are unable to support implantation during pregnancy, they are associated with complications, they are time-consuming and expensive, and they are associated with ethical issues in certain territories.274 Numerous conditions are related to infertility, including POF, nonobstructive azoospermia, endometrial dysfunction, and Asherman syndrome. Recent progress has been illustrated in preclinical studies for the potential applications of stem cell-based therapy for reproductive function recovery, especially recent studies in the field of MSCs, which provide new hope for patients with infertility and reproductive disorders.275

POF is characterized by a loss of ovarian activity during middle age (before 40 years old) and affects 12% of women of reproductive age.276 Patients diagnosed with POF exhibit oligo-/amenorrhea for at least 4 months, with increased levels of follicle-stimulating hormone (FSH) (>25IU/L) on two occasions more than 1 month apart.277 Diverse factors, such as genetic backgrounds, autoimmune disorders, environmental conditions, and iatrogenic and idiopathic situations, have been reported to be the cause of POF.278 POF can be treated with limited effectiveness via psychosocial support, hormone replacement intervention, and fertility management.279 MSCs from AT, BM, and UC have been used in the treatment of POF, with improvements in ovarian function in preclinical studies using chemotherapy-induced POF animal models. The early published POF study using BM-MSCs as the main cell source is a single case report in which a perimenopausal woman showed an improvement in follicular regeneration, and increased AMH levels resulted in a successful pregnancy followed by delivery of a healthy infant.280 A report using autologous BM-MSCs in two women with POF illustrated an increase in baseline estrogen levels and the volume of the treated ovaries along with amelioration of menopausal symptoms.281 The clinical procedures used in this early trial were invasive, as patients underwent two operations: (1) BM aspiration and (2) laparoscopy. A similar approach was used in two trials conducted in 10 women with POF (age range from 2633 years old) and 30 patients (age from 18 to 40 years old).282 A later study investigated two different routes of cell delivery, including laparoscopy and the ovarian artery, but the results have not been reported at this time.282 Based on the positive outcomes of the mouse model, an autologous stem cell ovarian transplantation (ASCOT) trial was deployed using BM-derived stem cells with encouraging observations of improved ovarian function, as determined by elevated levels of AMH and AFC in 81.3% of participants, six pregnancies, and the successful delivery of three healthy babies.283 A randomized trial (NCT03535480) was conducted in 20 patients with POF aged less than 39 years to further elaborate on the results of the ASCOT trial.284 To date, there are no completed trials using AT-MSCs or UC-MSCs in the treatment of patients with POF, limiting the evaluation of these MSCs in the treatment of POF. The speculated reason is that POF is a rare disease, affecting 1% of women younger than 40 years, and with improvements in assisted productive technology, patients have several alternative options to enhance the recovery of reproductive function.285

Burns are the fourth most common injury worldwide, affecting ~11 million people, and are a major cause of death (180,000 patients annually). The severity of burns is defined based on the percentage of surface area burned, burn depth, burn location and patient age, and burns are usually classified into first-, second-, third-, and fourth-degree burns on the basis of their severity.286 Postburn recovery depends on the severity of the burn and the effectiveness of treatment. Rapid healing may occur over weeks, while alternatively, healing can take months, with the ultimate result being scar formation and disability in patients with severe burns. Different from mechanical injury, burn injury is an invasive progression of damage to tissue at the burn site, including both mechanical damage to the skin surface and biological damage caused by natural apoptosis that prolongs excessive inflammation, oxidative stress, and impaired tissue perfusion.287 To date, completely reversing the devastating damage of severe burns remains unachievable in medicine, and stem cell therapy provides an alternative option for patients with burn injury. The first case report of the use of BM-MSCs to treat a 45-year-old patient with burns on 40% of their body demonstrated the safety of the therapy and showed partial improvements in vascularization at the wound site and reduced coarse cicatrices.288,289 Later, patients with second- and third-degree burns as well as deep burns were treated using either autologous BM-MSCs or allogeneic BM-MSCs by spraying the MSCs onto the burn sites or adding MSCs over a dermal matrix sheet to cover the wound. The results in these case reports revealed the potential efficacy of MSC-based therapy, which not only enhanced the speed of wound recovery but also reduced pain and improved blood supply without introducing infection.288,290,291 In 2017, a study conducted in 60 patients with 1025% of their total body surface areas burned treated with either autologous BM-MSCs or UC-MSCs showed that both MSC types improved the rate of healing and reduced the hospitalization period.292 The drawback of BM-MSCs in the treatment of burns is the invasive harvesting method, which causes pain and possible complications in patients. Hence, treatment with allogeneic MSCs obtained from healthy donors is the method of choice, and AT- and UC-MSCs are two suitable candidates for this option. To date, a limited number of clinical trials have been conducted using MSC therapy. These trials have several limitations in trial design, such as a lack of a negative control group and blinding, small sample sizes, and the use of standardized measurement tools for burn injury and wound healing. Currently, AT-MSCs are being used in seven ongoing phase I and II trials in the treatment of burns. Hence, it is important to note that among the most widely studied MSCs, AT-MSCs have advantages over BM-MSCs when obtained from an allogeneic source, while their abilities in burn treatment remain to be determined. The main MSCs that should be used in the regeneration of burn tissue remain undefined (Table ), and we observed the trend that AT-MSCs are more suitable candidates due to their biological nature, which contributes to the generation of keratinocytes and secretion profiles that strongly enhance the skin regeneration process.293296

The reported clinical trials using MSCs from AT, BM, and UC in the treatment of the endocrinological disorder, reproductive disease, and skin healing

In the last two decades, great advancements have been achieved in the development of novel regenerative medicine and cardiovascular research, especially stem cell technology.297 The discovery of human embryonic stem cells and human induced pluripotent stem cells (hiPSCs) opened a new door for basic research and therapeutic investigation of the use of these cells to treat different diseases.298 However, the clinical path of hiPSCs and hiPSC-derived cardiomyocytes in the treatment of cardiovascular diseases is limited due to the potential for teratoma formation with hiPSCs and the immaturity of hiPSC-derived cardiomyocytes, which might pose a risk of cancer formation,299 arrhythmia, and cardiac arrest to patients.300 A recently emerged stem cell type is adult stem cells/progenitor cells, including MSCs, which can stimulate myocardial repair post administration due to their paracrine effects. Promising results of MSC-based therapy obtained from preclinical studies of cardiac diseases enhance the knowledge and strengthen the clinical research to investigate the safety and efficacy in a clinical trial setting. There are papers that discuss the importance of MSC therapy in the treatment of cardiovascular diseases, with the following references being highly recommended.301306 To date, 36 trials have evaluated the therapeutic potential of MSCs in different pathological conditions, with the most prevalent types being BM-MSCs (25 trials), followed by UC-MSCs (7 trials) and AT-MSCs (4 trials).303 However, the reported results are contradictory and create controversy about the efficacy of the treatments.

One of the first trials using MSCs in the treatment of chronic heart failure was the Cardiopoietic Stem Cell Therapy in Heart Failure (C-CURE) trial, a multicentre, randomized clinical trial that recruited 47 patients. The trial findings supported the safety of BM-MSC therapy and provided a data set that demonstrated improvements in cardiovascular scores along with New York Heart Association functional class, quality of life, and general physical health.307 Despite these encouraging results in the phase I trial, the treatment failed to achieve the primary outcomes in the phase II/III trial (CHART-1 trial), including no significant improvements in cardiac structure or function or patient quality of life.308 A positive outcome was also found in a phase I/II, randomized pilot study called the POSEIDON trial, which was the first trial to demonstrate the superior effectiveness of the administration of allogeneic BM-MSCs compared to allogeneic MSCs from other sources.309,310 Published results from the MSC-HF study, with 4 years of follow-up results,311,312 and the TRIDENT study313 illustrated the positive outcomes of BM-MSCs in the treatment of heart failure. However, a contradictory result from the recently published CONCERT-HF trial demonstrated that the administration of autologous BM-MSCs to patients diagnosed with chronic ischemic heart failure did not improve left ventricular function or reduce scar size at 12 months post administration, but the patients quality of life was improved.314 This observation is similar to that of the TAC-HFT trial315 but completely different from the reported results of the MSC-HF trial. A comprehensive investigation is still needed to determine the reasons behind these contradictory results. The largest clinical trial to date using BM-MSCs is the DREAM-HF study, which was a randomized, double-blind, placebo-controlled, phase III trial that was conducted at 55 sites across North America and recruited a total of 565 patients with ischemic and nonischaemic heart failure.172 Although recent reports from the sponsor confirmed that the trial missed its primary endpoint (a reduction in recurrent heart failure-related hospitalization), other prespecified endpoints were met, such as a reduction in overall major adverse cardiac events (including death, myocardial infarction, and stroke).306 Thus, a complete report from the DREAM-HF trial will provide pivotal data supporting the therapeutic potential of BM-MSCs in the treatment of heart failure and open a new path for the FDA to approve cell-based therapy for cardiovascular diseases.

The early trial using AT-derived cells was the PRECISE trial, which was a phase I, randomized, placebo-controlled, double-blind study that examined the safety and efficacy of adipose-derived regenerative cells (ADRCs) in the treatment of chronic ischemic cardiomyopathy.316 ADRCs are a homogenous population of cells obtained from the vascular stromal fraction of AT, which contains a small proportion of AT-MSCs.317 Although the study supported the safety of ADRC administration and illustrated a preserved functional capacity (peak VO2) in the treated group and improvements in heart wall motion, neither poor left ventricle (LV) volume nor poor left ventricular ejection fraction (LVEF) was ameliorated. The follow-up trial of the PRECISE trial, called the ATHENA trial, was conducted in 31 patients, although the study was terminated prematurely because two cerebrovascular events occurred, which were not related to the cell product itself.318 The results of the study illustrated increases in functional capacity, hospitalization rate, and MLHFQ scores, but the LV volume and LVEF were not significantly different between the two groups. Kastrup and colleagues conducted the first in vitro expanded AT-MSC trial in ten patients with ischemic heart disease and ischemic heart failure in 2017. The results confirmed that ready-to-use AT-MSCs were well-tolerated and potentially effective in the treatment of ischemic heart disease and heart failure.319 Comparable results of AT-MSCs were also reported from the MyStromalCell Trial, which was a randomized placebo-controlled study. In this trial, 61 patients were randomized at a 2:1 ratio into two groups, with the results showing no significant difference in the primary endpoint, which was a change in the maximal bicycle exercise tolerance test (ETT) score from baseline to 6 months post administration.320 A 3-year follow-up report from the MyStromalCell Trial confirmed that patients who received AT-MSC administration maintained their preserved exercise capacity and their cardiac symptoms improved, whereas the control group experienced a significant reduction in exercise performance and a worsened cardiovascular condition.321

UC-MSCs are potential allogeneic cells for the treatment of cardiovascular disease, as they are ready to use and easy to isolate, they rapidly proliferate, and they secrete hepatocyte growth factors,322 which are involved in cardioprotection and cardiovascular regeneration.323 The pilot study using UC-MSCs in 30 patients with heart failure, called the RIMECARD trial, was the first reported trial for which the results supported the effectiveness of UC-MSCs, as seen in the improved ejection fraction, left ventricular function, functional status, and quality of life in patients administered UC-MSCs.324 Encouraging results reported from a phase I/II HUC-HEART trial325 showed improvements in LVEF and reductions in the size of the injured area of the myocardium. However, the opposite observations were also reported from a recently published phase I randomized trial using a combination of UC-MSCs and a collagen scaffold in patients with ischemic heart conditions, in which the size of fibrotic scar tissue was not significantly reduced.326

Although MSCs from AT, BM, and UC have proven to be safe and feasible in the treatment of cardiovascular diseases, the correlation between the MSC types and their therapeutic potentials is still uncertain because different results have been reported from different clinical trials (Table ). The mechanisms by which MSCs participate in recovery and enhance myocardial regeneration have been discussed comprehensively in a recently published review;305,327 therefore, they will not be discussed in this review. In fact, the challenges of MSC-based therapy in cardiovascular diseases have been clearly described previously,328 including (1) the lack of an in vitro evaluation of the transdifferentiation potential of MSCs to functional cardiac and endothelial cells,329 (2) the uncontrollable differentiation of MSCs to undesirable cell types post administration,330 and (3) the undistinguishable nature of MSCs derived from different sources with various levels of differentiation potential.331 Therefore, the applications of MSC-based therapy in cardiovascular disease are still in their immature stage, with potential benefits to patients. Thus, there is a need to conduct large-scale, well-designed randomized clinical trials not only to confirm the therapeutic potential of MSCs from various sources but also to enhance our knowledge of cardiovascular regeneration post administration.

The reported clinical trials using MSCs from AT, BM, and UC in the treatment of cardiovascular diseases

Bones are complex structures constituting a part of the vertebrate skeleton, and they play a vital role in the production of blood cells from HSCs. Similar to the functions of most vertebrate organs, bone function is tightly regulated by its constituents and by long-range signaling from AT and the adrenal glands, parathyroid glands, and nervous system.332 The central nervous system (CNS) orchestrates the voluntary and involuntary input transmitted by a network of peripheral nerves, which act as the bridge between the nervous system and target organs. The CNS controls involuntary responses via the autonomic nervous system (ANS), consisting of the sympathetic nervous system and the parasympathetic nervous system, and voluntary responses via the somatic nervous system. The ANS penetrates deep into the BM cavity, reaching the regions of hematopoietic activity to deliver neurotransmitters that tightly regulate BM stem cell niches.333 The BM microenvironment consists of various cell types that participate in the maintenance of HSC niches, which are composed of specialized cells, including BM-MSCs (Fig. ). The release of a specific neurotransmitter, circadian norepinephrine, from the sympathetic nervous system at nerve terminals leads to a reduction in the circadian expression of CX-C chemokine ligand 12 (CXCL12, which is also known as stromal cell-derived factor-1 (SDF-1)) by Nestin+/NG22+ BM-MSCs, resulting in the secretion of HSCs into the peripheral bloodstream.334,335 In fact, BM-MSCs play a significant role in the regulation of HSC quiescence and are closely associated with arterioles and sympathetic nervous system nerve fibers. Nestin-expressing BM-MSCs have been shown to express high levels of SDF-1, stem cell factor (SCF), angiopoietin-1 (Ang-1), interleukin-7, vascular cell adhesion molecule 1 (VCAM-1), and osteopontin (OPN), which are directly involved in the regulation and maintenance of HSC quiescence.336 The depletion of BM-MSCs in BM leads to the mobilization of HSCs into the peripheral bloodstream and spleen. The findings from a previous study demonstrated that reduced SDF-1 expression in norepinephrine-treated BM-MSCs resulted in the mobilization of CXCR4+ HSCs into circulation.337 The ability of BM-MSCs to produce SDF-1 is tightly related to their neuronal protective functions.338 SDF-1 is a member of a chemokine subfamily that orchestrates an enormous diversity of pathways and functions in the CNS, such as neuronal survival and proliferation. The chemokine has two receptors, CXCR4 and CXCR7, that are involved in the pathogenic development of neurodegenerative and neuroinflammatory diseases.339 In the damaged brain, SDF-1 functions as a stem cell homing signal, and in acquired immune deficiency syndrome (AIDS), SDF-1 has been reported to be involved in the protection of damaged neurons by preventing apoptosis. In a traumatic brain injury model, SDF-1 was found to function as an inhibitor of the caspase-3 pathway by upregulating the Bcl-2/Bax ratio, which in turn protects neurons from apoptosis.340 Moreover, the release of SDF-1 also facilitates cell recruitment, cell migration, and the homing of neuronal precursor cells in the adult CNS by activating the CXCR4 receptor.341,342 Existing data support that SDF-1 acts as the guiding signal for the regeneration of axon growth in damaged neurons and enhances spinal nerve regeneration.343,344 Hence, the ability of BM-MSCs to express SDF-1 in response to the neuronal environment provides a unique neuronal protective effect that could explain the potential therapeutic efficacy of BM-MSCs in the treatment of neurodegenerative diseases (Fig. ).

The nature of the stem niche of bone marrow-derived mesenchymal stem cells (BM-MSCs) supports their therapeutic potential in neuron-related diseases. a Bone marrow is a complex stem cell niche regulated directly by the central nervous system to maintain bone marrow homeostasis and haematopoietic stem cell (HSC) functions. MSCs in bone marrow respond to the environmental changes through the release of norepinephrine (NE) from the sympathetic nerves that regulate the synthesis of SDF-1 and the migration of HSCs through the sinusoids. The secretion of stem cell factors (SCFs), VCAM-1 and angiotensin-1 from MSCs also plays a significant role in the maintenance of HSCs. b BM-MSCs have the ability to produce and release SDF-1, which directly contributes to neuroprotective functions at the damaged site through interaction with its receptors CXCR4/7, located on the neuronal membrane. c Neuronal protection and the functional remyelination induced by BM-MSCs are also modulated by the release of a wide range of growth factors, including VEGF, BDNF, and NGF, by the BM-MSCs. d BM-MSCs also have the ability to regulate neuronal immune responses by direct interaction or paracrine communication with microglia. Figure was created with BioRender.com

The migration of exogenous MSCs after systemic administration to the brain is limited by the physical bloodbrain barrier (BBB), which is a selective barrier formed by CNS endothelial cells to restrict the passage of molecules and cells. The mechanism of molecular movement across the BBB is well established, but how stem cells can bypass the BBB and home to the brain remains unclear. Recent studies have reported that MSCs are able to migrate through endothelial cell sheets by paracellular or transcellular transport followed by migration to the injured or inflammatory site of the brain.345,346 During certain injuries or ischemic events, such as brain injury, stroke, or cerebral palsy, the integrity and efficiency of BBB protection is compromised, which allows MSC migration across the BBB via paracellular transport through the transient formation of interendothelial gaps.347 CD24 expression has been detected in human BM-MSCs, which are regulated by TGF-3,348 allowing them to interact with activated endothelial cells via P-selectin and initiate the tethering and rolling steps of MSCs.349 Additionally, BM-MSCs express high levels of CXCR4 or CXCR7,350,351 which bind to integrin receptors, such as VLA-4, to activate the integrin-binding process and allow the cells to anchor to endothelial cells, followed by the migration of MSCs through the endothelial cell layer and basement membrane in a process called transmigration.352 This process is facilitated by the secretion of matrix metalloproteinases (MMPs), which degrade the endothelial basement membrane, allowing BM-MSCs to enter the brain environment.353,354 BM-MSCs can also regulate the integrity of the BBB via the secretion of tissue inhibitor of matrix metalloproteinase-3 (TIMP3), which has been shown to ameliorate the effects of a compromised BBB in traumatic brain injury.355 The secretion of TIMP3 from MSCs directly blocked vascular endothelial growth factor a (VEGF-a)-induced breakdown of endothelial cell adherent junctions, demonstrating the potential mechanism of BM-MSCs in the regulation of BBB integrity.

The therapeutic applications of BM-MSCs in neurodegenerative conditions have been significantly increased by the demonstration of BM-MSC involvement in axonal and functional remyelination processes. Remyelination is a spontaneous regenerative process occurring in the human CNS to protect oligodendrocytes, neurons, and myelin sheaths from neuronal degenerative diseases.356 Remyelination is considered a neuroprotective process that limits axonal degeneration by demyelination and neuronal damage. The first mechanism of action of BM-MSCs related to remyelination is the activation of the JAK/STAT3 pathway to regulate dorsal root ganglia development.357 It was reported that BM-MSCs secrete vascular endothelial growth factor-A (VEGF-A),358 brain-derived neurotrophic factor (BDNF), interleukin-6, and leukemia inhibitor factor (LIF), which directly function in neurogenesis and neurite growth.357 VEGF-A is a key regulator of hemangiogenesis during development and bone homeostasis. Postnatally, osteoblast- and MSC-derived VEGF plays a critical role in maintaining and regulating bone homeostasis by stimulating MSC differentiation into osteoblasts and suppressing their adipogenic differentiation.359361 To balance osteoblast and adipogenic differentiation, VEGF forms a functional link with the nuclear envelope protein laminin A, which in turn directly regulates the osteoblast and adipocyte transcription factors Runx2 and PPAR, respectively.361,362 In the brain, VEGF is a potent growth factor mediating angiogenesis, neural migration, and neuroprotection. VEGF-A, secreted from BM-MSCs under in vitro xeno- and serum-free culture conditions, is the most studied member of the VEGF family and is suggested to play a protective role against cognitive impairment, such as in the context of Alzheimers disease pathology or stroke.363365 Recently, it was reported that the neurotrophic and neuroprotective function of VEGF is mediated through VEGFR2/Flk-1 receptors, which are expressed in the neuroproliferative zones and extend to astroglia and endothelial cells.366 In animal models of intracerebral hemorrhage and cerebral ischemia, the transfusion of Flk-1-positive BM-MSCs promotes behavioral recovery and anti-inflammatory and angiogenic effects.367,368 Moreover, supplementation with VEGF-A in neuronal disorders enhances intraneural angiogenesis, improves nerve regeneration, and promotes neurotrophic capacities, which in turn increase myelin thickness via the activation of the prosurvival transcription factor nuclear factor-kappa B (NF-kB). This activation, together with the downregulation of Mdm2 and increased expression of the pro-apoptotic transcription factor p53, is considered to be the neuroprotective process associated with an increased VEGF-A level.369371 An analysis of microRNA (miRNA) in extracellular vesicles (EVs) secreted from BM-MSCs revealed that BM-MSCs release substantial amounts of miRNA133b, which suppresses the expression of connective tissue growth factor (CTGF) and protects hippocampal neurons from apoptosis and inflammatory injury372374 (Fig. ).

In terms of immunoregulatory functions, the administration of human BM-MSCs into immunocompetent mice subjected to SCI or brain ischemia showed that BM-MSCs exhibited a short-term neuronal protective function against neurological damage (Fig. ). Further investigation demonstrated the ability of BM-MSCs to directly communicate with host microglia/macrophages and convert them from phenotypic polarization into alternative activated microglia/macrophages (AAMs), which are key players in axonal extension and the reconstruction of neuronal networks.375 Other studies have also illustrated that the administration of AAMs directly to the injured spinal cord induced axonal regrowth and functional improvement.376 The mechanism by which BM-MSCs activate the conversion of microglia/macrophages occurs through two representative macrophage-related chemokine axes, CCL2/CCR2 and CCL-5/CCR5, both of which exhibit acute or chronic elevation following brain injury or SCI.377 The CCL2/CCR2 axis contributed to the enhancement of inflammatory function, and BM-MSC-mediated induction of CCL2 did not alter the total granulocyte number (Fig. ). Although the chemokine-mediated mechanism of BM-MSCs in the activation of AAMs and enhanced axonal regeneration at the damage sites is evident, the direct mechanism by which the communication between BM-MSCs and the target cells results in these phenomena remains unclear, and further investigation is needed.

BM-MSCs also confer the ability to regulate the inflammatory regulation of the immune cells present in the brain by (1) promoting the polarization of macrophages toward the M2 type, (2) suppressing T-lymphocyte activities, (3) stimulating the proliferation and differentiation of regulatory T cells (Tregs), and (4) inhibiting the activation of natural killer (NK) cells. BM-MSCs secrete glial cell line-derived neurotrophic factor (GDNF), a specific growth factor that contributes directly to the transition of the microglial destructive M1 phenotype into the regenerative M2 phenotype during the neuroinflammatory process.378 A similar result was also found in AT-379 and UC-MSCs380 under neuroinflammation-associated conditions, suggesting that AT-, BM-, and UC-MSCs share the same mechanism in promoting macrophage polarization. In terms of T-lymphocyte suppression, compared to MSCs from AT and BM, UC-MSCs show the strongest potential to inhibit the proliferation of T-lymphocytes by promoting cell cycle arrest (G0/G1 phase) and apoptosis.381 In addition, UC-MSCs have been proven to be more effective in promoting the proliferation of Tregs382 and inhibiting NK activation.383 Although MSCs are well-known for their inflammatory regulatory ability, the mechanism is not exclusive to BM-MSCs, especially in neurological disorders.384

In contrast to AT-MSCs and BM-MSCs, UC-MSCs have lower expression of major histocompatibility complex I (MHC I) and no expression of MHC II, which prevents the complications of immune rejection.385 Moreover, as UC is considered a waste product after birth, with the option of noninvasive collection, UC-MSCs are easier to obtain and culture than AD- and BM-MSCs.386 These advantages of UC-MSCs have contributed to their use in the treatment of pulmonary diseases, especially during the rampant COVID-19 pandemic, as off-the-shelf products. Numerous pulmonary diseases have been the subject of applications of UC-MSCs, including BPD, COPD, ARDS, and COVID-19-induced ARDS. In BPD, premature infants are born before the alveolarization process, resulting in arrested lung development and alveolar maturation. Upon administration via an IV route, the majority of exogenous UC-MSCs reach the immature lung and directly interact with immune cells to exert their immunomodulatory properties via cell-to-cell interaction mechanisms (Fig. ). UC-MSCs interact with T cells via the PD-L1 ligand, which binds to the PD-1 inhibitory molecule on T cells, resulting in the suppression of CD3+ T-cell proliferation and effector T-cell responses.387 In addition, UC-MSCs also express CD54 (ICAM-1), which plays a crucial role in the immunomodulatory functions of T cells.388 Direct contact between UC-MSCs and macrophages via CD54 expression on UC-MSCs promotes the immune regulation of UC-MSCs via the regulation of phagocytosis by monocytes.389 Moreover, the contact of UC-MSCs with macrophages during proinflammatory responses increases the secretion of TSG-6 by UC-MSCs, which in turn promotes the inhibitory regulation of CD3+ T cells, macrophages, and monocytes by MSCs.390 Recently, upregulation of SDF-1 was described in neonatal lung injury, especially in layers of the respiratory epithelium.391 SDF-1 has been shown to participate in the migration and initiation of the homing process of MSCs via the CXCR4 receptors on their surface.392 It was reported that UC-MSCs express low levels of CXCR4, allowing them to induce SDF-1-associated migration processes via the Akt, ERK, and p38 signal transduction pathways.393 Hence, in BPD, the upregulation of SDF-1 together with the homing ability of UC-MSCs strongly supports the therapeutic effects of UC-MSCs in the treatment of BPD. Furthermore, UC-MSCs have the ability to communicate with immune cells via cell-to-cell contact to reduce proinflammatory responses and the production of proinflammatory cytokines (such as TGF-, INF-, macrophage MIF, and TNF-). The modulation of the human innate immune system by UC-MSCs is mediated by cellcell interactions via CD54-LFA-1 that switch macrophage polarization processes, promoting the proliferation of M2 macrophages, which in turn reduce inflammatory responses in the immature lung.394 Moreover, UC-MSCs also have the ability to produce VEGF and hepatocyte growth factors (HGFs), promoting angiogenesis and enhancing lung maturation.395

Adipose tissue-derived mesenchymal stem cells (AT-MSCs) and the nature of their tissue of origin support their use in therapeutic applications. a Adipose tissue is considered an endocrine organ, supporting and regulating various functions, including appetite regulation, immune regulation, sex hormone and glucocorticoid metabolism, energy production, the orchestration of reproduction, the control of vascularization, and blood flow, the regulation of coagulation, and angiogenesis and skin regeneration. b In terms of metabolic disorders, such as type 2 diabetes mellitus (T2DM), as adipose tissue is directly involved in the metabolism of glucose and lipids and the regulation of appetite, the detrimental effects of T2DM also alter the functions of AT-MSCs, which in turn, hampers their therapeutic effects. Hence, the use of autologous AT-MSCs is not recommended for the treatment of metabolic disorders, including T2DM, suggesting that allogeneic AT-MSCs from healthy donors could be a better alternative approach. c AT-MSCs are suitable for the treatment of reproductive disorders due to their unique ability to mobilize and home to the thecal layer of the injured ovary, enhance the regeneration and maturation of thecal cells, increase the structure and function of damaged ovaries via exosome-activated SMAD, decrease oxidative stress and autophagy, and increase the proliferation of granulosa cells via PI3K/AKT pathways. These functions are regulated specifically by growth hormones produced by AT-MSCs in response to the surrounding environment, including HGF, TGF-, IGF-1, and EGF. d AT-MSCs are also good candidates for skin healing and regeneration as their growth factors strongly support neovascularization and angiogenesis by reducing PLL4, increase anti-apoptosis via the activation of PI3K/AKT pathways, regulate inflammation by downregulating NADPH oxidase isoform 1, and increase immunoregulation through the inhibition of NF-B activation. The figure was created with BioRender.com

COPD is characterized by an increase in hyperinflammatory reactions in the lung, compromising lung function and increasing the development of lung fibrosis. The mechanism by which UC-MSCs contribute to the response to COPD is inflammatory regulation (Fig. ). The administration of UC-MSCs prevented the infiltration of inflammatory cells in peribronchiolar, perivascular, and alveolar septa and switched macrophage polarization to M2.396 A significant reduction in proinflammatory cytokines, including IL-1, TNF-, and IL-8, was also observed following UC-MSC administration.224 MSCs, including UC-MSCs, have been reported to trigger the production of secretory leukocyte protease inhibitors in epithelial cells through the secretion of HGF and epidermal growth factor (EGF), which is believed to have beneficial effects on COPD.397,398 In addition to their inflammatory regulation ability, UC-MSCs exhibit antimicrobial effects through the inhibition of bacterial growth and the alleviation of antibiotic resistance during Pseudomonas aeruginosa infection.399 The combination of the regulation of the host immune response and the antimicrobial effects of UC-MSCs may be relevant for the prevention and treatment of COPD exacerbations, as inflammation and bacterial infections are important risk factors that significantly contribute to the morbidity and mortality of patients with COPD. In terms of regenerative functions, UC-MSCs were reported to be able to differentiate into type 2 alveolar epithelial cells in vitro and alleviate the development of pulmonary fibrosis via -catenin-regulated cell apoptosis.400 Furthermore, UC-MSCs enhanced alveolar epithelial cell migration and proliferation by increasing matrix metalloproteinase-2 levels and reduced their endogenous inhibitors, tissue inhibitors of matrix metalloproteinases, providing a potential mechanism underlying their anti-pulmonary-fibrosis effects.401,402

In ARDS, especially that associated with COVID-19, the proinflammatory state is initiated by increases in plasma concentrations of proinflammatory cytokines, such as IL-1 beta, IL-7, IL-8, IL-9, IL-10, bFGF, granulocyte colony-stimulating factor (G-CSF), GM-CSF, IFN-, and TNF-. The significant increases in the concentrations of these cytokines in patient plasma suggest the development of a cytokine storm, which is a leading cause of COVID-induced mortality. In addition to the immunomodulatory functions regulated via cell-to-cell interactions between UC-MSCs and immune cells, such as macrophages, monocytes, and T cells, UC-MSCs exert their functions via paracrine effects through the secretion of growth factors, cytokines, and exosomes (Fig. ). The most relevant immunomodulatory function of UC-MSCs is considered to be their inhibition of effector T cells via the induction of T-cell apoptosis and cell cycle arrest by the production of indoleamine 2,3- dioxygenase (IDO), prostaglandin E2 (PGE-2), and TGF-. Elevated levels of PGE-2 in patients with COVID-19 are reported to be a crucial factor in the initiation of inflammatory regulation by UC-MSCs post administration and prevent the development of cytokine storms by direct inhibition of T- and B lymphocytes.403 UC-MSCs exert these inhibitory activities through a PGE-2-dependent mechanism.404 It was reported that UC-MSCs confer the ability to secrete tolerogenic mediators, including TGF-1, PGE-2, nitric oxide (NO), and TNF-, which are directly involved in their immunoregulatory mechanism. The secretion of NO from UC-MSCs is reported to be associated with the desensitization of T cells via the IFN-inducible nitric oxide synthase (iNOS) pathways and to stimulate the migration of T cells in close proximity to MSCs that subsequently suppress T-cell sensitivities via NO.405 Lung infection with viruses usually leads to impairments in alveolar fluid clearance and protein permeability. The administration of UC-MSCs enhances alveolar protection and restores fluid clearance in patients with COVID-19. UC-MSCs secrete growth factors associated with angiogenesis and the regeneration of pulmonary blood vessels and micronetworks, including angiotensin-1, VEGF, and HGF, which also reduce oxidative stress and prevent fibrosis formation in the lungs. These trophic factors have been identified as key players in the modulation of the microenvironment and promote pulmonary repair. Additionally, UC-MSCs are more effective than BM-MSCs in the restoration of impaired alveolar fluid clearance and the permeability of airways in vitro, supporting the use of UC-MSCs in the treatment of patients with pulmonary pneumonia.406 In the context of pulmonary regeneration, UC-MSCs were shown to inhibit apoptosis and fibrosis in pulmonary tissue by activating the PI3K/AKT/mTOR pathways via the secretion of HGF, which also acts as an inhibitory stimulus that blocks alveolar epithelial-to-mesenchymal transition.407,408 Moreover, UC-MSCs can reverse the process of fibrosis via enhanced expression of macrophage matrix-metallopeptidase-9 for collagen degradation and facilitate alveolar regeneration via Toll-like receptor-4 signaling pathways.409 UC-MSCs were shown to communicate with CD4+ T cells through HGF induction not only to inhibit their differentiation into Th17 cells, reducing the secretion of IL-17 and IL-22 but also to switch their differentiation into regulatory T cells.410,411 In addition, UC-MSCs conferred the ability to facilitate the number of M2 macrophages and reduce M1 cells via the control of the macrophage polarization process.412

There are several potential mechanisms of UC-MSCs in the treatment of patients with pulmonary diseases and pneumonia, including the regulation of immune cell function, immunomodulation, the enhancement of alveolar fluid clearance and protein permeability, the modulation of endoplasmic reticulum stress, and the attenuation of pulmonary fibrosis. Hence, based on these discussions, UC-MSCs are recommended as suitable candidates for the treatment of pulmonary disease both in pediatric and adult patients.

Human AT was first viewed as a passive reservoir for energy storage and later as a major site for sex hormone metabolism, the production of endocrine factors (such as adipsin and leptin), and a secretion source of bioactive peptides known as adipokines.413 It is now clear that AT functions as a complex and highly active metabolic and endocrine organ, orchestrating numerous different biological features414 (Fig. ). In addition to adipocytes, AT contains hematopoietic-derived progenitor cells, connective tissue, nerve tissue, stromal cells, endothelial cells, MSCs, and pericytes. AT-MSCs and pericytes mobilize from their perivascular locations to aid in healing and tissue regeneration throughout the body. As AT is involved directly in energy storage and metabolism, AT-MSCs are also mediated and regulated by growth factors related to these pathways. In particular, interleukin-6 (IL-6), IL-33, and leptin regulate the maintenance of metabolic activities by increasing insulin sensitivity and preserving homeostasis related to AT. Nevertheless, in the development of obesity and diabetes, omental and subcutaneous AT maintains a low-grade state of inflammation, resulting in the impairment of glucose metabolism and potentially contributing to the development of insulin resistance.415 In normal AT, direct regulation of Pre-B-cell leukemia homeobox (Pbx)-regulating protein-1 (PREP1) by leptin and thyroid growth factor-beta 1 (TGF-1) in AT-MSCs and mature adipocytes is involved in the protective function and maintenance of AT homeostasis. However, under diabetic conditions, the balance between the expression of leptin and the secretion of TGF-1 is compromised, resulting in the malfunction of AT-MSC metabolic activity and the proliferation, differentiation, and maturation of adipocytes. Therefore, the use of autologous AT-MSCs in the treatment of diabetic conditions is not a suitable option, as the functions of AT-MSCs are directly altered by diabetic conditions, which reduces their effectiveness in cell-based therapy (Fig. ).

Umbilical cord-derived mesenchymal stem cells (UC-MSCs) are good candidates for the treatment of pulmonary diseases. a Lung immaturity and fibrosis are the major problems of patients with bronchopulmonary dysplasia and lead to increased levels of SDF-1, the development of fibrosis, the induction of the inflammatory response, and the impairment of alveolarization. UC-MSCs are attracted to the damaged lung via the chemoattractant SDF-1, which is constantly released from the immature lung via SDF-1 and CXCR4 communication. Moreover, UC-MSCs reduce the level of proinflammatory cytokines (TGF-, INF-, macrophage MIF, and TNF-) via a cell-to-cell contact mechanism. The ability of UC-MSCs to produce and secrete VEGF also involves in the regeneration of the immature lung through enhanced angiogenesis. b Upon an exacerbation of chronic obstructive pulmonary disease (COPD), UC-MSCs respond to the surrounding stimuli by reducing IL-8 and TNF- levels, resulting in the inhibition of the inflammatory response but an increase in the secretion of growth factors participating in the protection of alveoli, fluid clearance and reduced oxidative stress and lung fibrosis, including HGF, TGF-, IGF-1, and exosomes. c In a similar manner, UC-MSCs prevent the formation of cytokine storms in coronavirus disease 2019 (COVID-19) by inhibiting CD34+ T-cell differentiation into Th17 cells and enhancing the number of regulatory T cells. Moreover, UC-MSCs also have antibacterial activity by secreting LL-3717 and lipocalin. Figure was created with BioRender.com

Preclinical studies and clinical trials have revealed the therapeutic effects of MSCs, in general, and AT-MSCs, in particular, in the management of POF, with relatively high efficacy and enhanced regeneration of the ovaries. Understanding the molecular and cellular mechanisms underlying these effects is the first step in the development of suitable MSC-based therapies for POF. One of the mechanisms by which MSCs exert their therapeutic effects is their ability to migrate to sites of injury, a process known as homing. Studies have shown that MSCs from different sources have the ability to migrate to different compartments of the injured ovary. For example, BM-MSCs administered through IV routes migrated mostly to the ovarian hilum and medulla,416 whereas a significant number of UC-MSCs were found in the medulla.417 Interestingly, AT-MSCs were found to be engrafted in the theca layers of the ovary but not in the follicles, where they acted as supportive cells to promote follicular growth and the regeneration of thecal layers.418 The structure and function of the thecal layer have a great impact on fertility, which has been reviewed elsewhere.419 In brief, the thecal layer consists of two distinct parts, the theca interna, which contains endocrine cells, and the theca externa, which is an outer fibrous layer. The thecal layer contains not only endocrine-derived cells but also vascular- and immune-derived cells, whose functions are to maintain the structural integrity of the follicles, transport nutrients to the inner compartment of the ovary and produce key reproductive hormones such as androgens (testosterone and dihydrotestosterone) and growth factors (morphogenic proteins, e.g., BMPs and TGF-).420 As AT-MSCs originate from an endocrine organ, their ability to sense signals and migrate to the thecal layer is anticipated. Additionally, secretome analysis of AT-MSCs showed a wide range of growth factors, including HGF, TBG-, VEGF, insulin-like growth factor-1 (IGF-1), and EGF,421 that are directly involved in the restoration of the structure and function of damaged ovaries by stimulating cell proliferation and reducing the aging process of oocytes via the activation of the SIRT1/FOXO1 pathway, a key regulator of vascular endothelial homeostasis.422,423 In POF pathology, autophagy and its correlated oxidative stress contribute to the development of POF throughout a patients life. Recently, AT-MSCs were shown to be able to improve the structure and function of mouse ovaries by reducing oxidative stress and inflammation, providing essential data supporting the mechanism of AT-MSCs in the treatment of POF.424 Several studies have illustrated that AT-MSCs secrete biologically active EVs that regulate the proliferation of ovarian granulosa cells via the PI3K/AKT pathway, resulting in the enhancement of ovarian function.425 Direct regulation of ovarian cell proliferation modulates the state of these cells, which in turn restores the ovarian reserve.426 Other mechanisms supporting the effectiveness of MSCs have been carefully reviewed, confirming the therapeutic potential of MSCs derived from different sources426 (Fig. ).

In the last decade, the number of clinical trials using AT-MSCs in the treatment of chronic skin wounds and skin regeneration has exponentially increased, with data supporting the enhancement of the skin healing processes, the reduction of scar formation, and improvements in skin structure and quality. Several mechanisms are directly linked to the origin of AT-MSCs, including differentiation ability, neovascularization, anti-apoptosis, and immunological regulation. AT is a connective and supportive tissue positioned just beneath the skin layers. AT-MSCs have a strong ability to differentiate into adipocytes, endothelial cells,427 epithelial cells428 and muscle cells.429 The adipogenic differentiation of AT-MSCs is one of the three mesoderm lineages that defines MSC features, and AT-MSCs are likely to be the best MSC type harboring this ability compared to BM- and UC-MSCs. Recent reports detailed that AT-MSCs accelerated diabetic wound tissue closure through the recruitment and differentiation of endothelial cell progenitor cells into endothelial cells mediated by the VEGF-PLC-ERK1/ERK2 pathway.430 Upon injury, the skin must be healed as quickly as possible to prevent inflammation and excessive blood loss. The reparation process occurs through distinct overlapping phases and involves various cell types and processes, including endothelial cells, keratinocyte proliferation, stem cell differentiation, and the restoration of skin homeostasis.431 Hence, the differentiation ability of AT-MSCs plays a critical role in their therapeutic effect on skin wound regeneration and healing processes. AT-MSCs accelerate wound healing via the production of exosomes that serve as paracrine factors. It was reported that AT-MSCs responded to skin wound injury stimuli by increasing their expression of the lncRNA H19 exosome, which upregulated SOX9 expression via miR-19b, resulting in the acceleration of human skin fibroblast proliferation, migration, and invasion.432 In addition, the engraftment of AT-MSCs supported wound bed blood flow and epithelialization processes.433 Anti-apoptosis plays a critical role in AT-MSC-based therapy, as without a microvascular supply network established within 4 days post injury, adipocytes undergo apoptosis and degenerate. Exogenous sources of AT-MSCs mediate anti-apoptosis via IGF-1 and exosome secretion by triggering the activation of PI3K signaling pathways.434 Another mechanism supporting the therapeutic potential of AT-MSCs is their anti-inflammatory function, which results in the reduction of proinflammatory factors, such as tumor necrosis factor (TNF) and interferon- (IFN-), and increases the production of the anti-inflammatory factors IL-10 and IL-4. Exosomes from AT-MSCs in response to a wound environment were found to contain high levels of Nrf2, which downregulated wound NADPH oxidase isoform 1 (NOX1), NADPH oxidase isoform 4 (NOX4), IL-1, IL-6, and TNF- expression. The anti-inflammatory functions of AT-MSCs are also regulated by their immunomodulatory ability, partially through the inhibition of NF-B activation in T cells via the PD-L1/PD-1 and Gal-9/TIM-3 pathways, providing a novel target for the acceleration of wound healing435 (Fig. ).

Therefore, as an endocrine organ in the human body, AT and its derivative stem cells, including AT-MSCs, have shown great potential in the treatment of reproductive disorders and skin diseases. Their potential is supported by mechanisms that are directly related to the nature of AT-MSCs in the maintenance of tissue homeostasis, angiogenesis, anti-apoptosis, and the regulation of inflammatory responses.

Over the past decades, MSC-based research and therapy have made tremendous advancements due to their advantages, including immune evasion, diverse tissue sources for harvesting, ease of isolation, rapid expansion, and cryopreservation as off-the-shelf products. However, several important challenges have to be addressed to further enhance the safety profile and efficacy of MSC-based therapy. In our opinion, the most important challenge of MSC-based therapy is the fate of these cells post administration, especially the long-term survival of allogeneic cells in the treatment of certain diseases. Although reported data confirm that the majority of MSCs are trapped in the lung and rapidly removed from the circulation, caution has been raised related to the occurrence of embolism events post infusion, which was proven to be related to MSC-induced innate immune attack (called instant blood-mediated inflammatory reaction).436 Another related challenge is the homing ability of infused cells, as successful homing at targeted tissue might result in long-term benefits to patients. Other concerns related to MSC-based therapy are the number of dead cells infused into the patients. An interesting study reported that dead MSCs alone still exerted the same immunomodulatory property as live MSCs by releasing phosphatidylserine.437 This is an interesting observation, as there is always a certain number of dead cells present in the cell-based product, and concerns are always raised related to their effects on the patients health. Finally, the hypothesis presented in this review is also a great challenge of the field, which has been proposed for future studies to answer the question: What is the impact of MSC sources on their downstream application?. Tables and illustrate the comparative studies that were conducted in preclinical and clinical settings to address the MSC source challenge. Other challenges of MSC-based therapies have been discussed in several reviews and systematic studies,135,185,438,439 which are highly recommended.

Comparative analysis of the effectiveness of MSC sources in a preclinical setting

Increase BDNF levels in the injured spinal cord, reduce lesion cavity volume and microglia/macrophage infiltration

Induce angiogenesis, axonal regeneration

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Stem cell-based therapy for human diseases - PMC

Abstract

Peripheral nerve injury (PNI) is a common neurological disorder and complete functional recovery is difficult to achieve. In recent years, bone marrow mesenchymal stem cells (BMSCs) have emerged as ideal seed cells for PNI treatment due to their strong differentiation potential and autologous transplantation ability. This review aims to summarize the molecular mechanisms by which BMSCs mediate nerve repair in PNI. The key mechanisms discussed include the differentiation of BMSCs into multiple types of nerve cells to promote repair of nerve injury. BMSCs also create a microenvironment suitable for neuronal survival and regeneration through the secretion of neurotrophic factors, extracellular matrix molecules, and adhesion molecules. Additionally, BMSCs release pro-angiogenic factors to promote the formation of new blood vessels. They modulate cytokine expression and regulate macrophage polarization, leading to immunomodulation. Furthermore, BMSCs synthesize and release proteins related to myelin sheath formation and axonal regeneration, thereby promoting neuronal repair and regeneration. Moreover, this review explores methods of applying BMSCs in PNI treatment, including direct cell transplantation into the injured neural tissue, implantation of BMSCs into nerve conduits providing support, and the application of genetically modified BMSCs, among others. These findings confirm the potential of BMSCs in treating PNI. However, with the development of this field, it is crucial to address issues related to BMSC therapy, including establishing standards for extracting, identifying, and cultivating BMSCs, as well as selecting application methods for BMSCs in PNI such as direct transplantation, tissue engineering, and genetic engineering. Addressing these issues will help translate current preclinical research results into clinical practice, providing new and effective treatment strategies for patients with PNI.

Keywords: Bone marrow mesenchymal stem cells, Peripheral nerve injury, Schwann cells, Myelin sheath, Tissue engineering

Core Tip: Bone marrow mesenchymal stem cells (BMSCs) have become ideal seed cells for the treatment of peripheral nerve injury (PNI) due to their strong differentiation potential and the possibility of autologous transplantation. In this review, we introduce the biological characteristics of BMSCs related to PNI, outline the current mechanisms by which BMSCs promote the regeneration and repair of PNI, and summarize the various application methods of BMSCs in PNI, confirming the potential of BMSCs in the treatment of PNI and providing great support for the development of new treatment strategies for nerve regeneration and repair in PNI.

Peripheral nerve injury (PNI) refers to damage that occurs to the peripheral nerve trunk or its branches due to direct or indirect trauma from external sources. It is characterized by sensory, motor, and autonomic dysfunction in the trunk or limbs, representing one of common neurological disorders in clinical practice[1]. PNI is a global issue, with an annual incidence rate of approximately 13/100000 to 23/100000 in developed countries[2-5]. While peripheral nerve axons can regenerate after injury, achieving complete functional recovery is often challenging in cases of proximal nerve injuries or large nerve defects[6]. Currently, autologous nerve transplantation is considered the gold standard for PNI repair[7]. However, even under ideal conditions, this approach does not fully restore impaired motor and sensory functions[8]. Additionally, it has significant drawbacks, such as prolonged surgical time, high economic costs, insufficient donor areas for reconstruction of long or multiple nerve defects, and potential donor site damage (painful neuroma, scarring, and sensory deficits)[9]. In recent years, several new methods for PNI repair have emerged, showing positive effects on restoring the continuity of injured neuroanatomy. However, their ability to restore nerve function is not ideal, and they all have varying degrees of limitations[10].

Tissue engineering is an emerging discipline in the field of biotechnology and has gained significant attention in PNI research. Previous studies have demonstrated that transplantation of Schwann cells (SCs) can promote nerve regeneration and accelerate nerve function recovery[11]. However, obtaining a large number of SCs in a short period is challenging, and it may cause irreversible damage to the donor area, thus limiting the clinical application of SCs transplantation[12]. Recent research has found that adult mesenchymal stem cells (MSCs) can also promote nerve regeneration and show potential for treating PNI, making them a more ideal alternative to SCs. Bone marrow MSCs (BMSCs) are one type of adult MSC with strong differentiation potential and advantages in autologous transplantation. Numerous studies have indicated that BMSCs can differentiate into nerve-like cells during the PNI treatment process and play a crucial role in nerve growth factor (NGF) secretion, endogenous stem cell migration and differentiation, and neovascularization[13-15]. These findings suggest that BMSCs effectively promote the repair of neurological deficits, which makes them ideal seed cells for PNI repair. Researchers are also striving to translate preclinical research findings into practical clinical applications for PNI patients. BMSCs can be applied to PNI therapy through a variety of techniques, such as cell transplantation, tissue engineering, gene engineering, and cell therapy, including the use of BMSC-derived exosomes. These approaches have the potential to improve the effectiveness of PNI regeneration and offer new hope for PNI patients.

Through literature search and analysis (Figure ), in this review, we present the biological properties of BMSCs associated with PNI. We summarize the current mechanisms by which BMSCs promote nerve regeneration and repair in PNI, as well as various application methods in PNI. Moreover, based on these findings, we identify the existing problems and limitations in order to deepen our understanding of BMSCs, optimize treatment strategies, address their shortcomings in clinical application in PNI, and promote their use in PNI clinical practice.

Flow chart of literature search and selection criteria. The initial search resulted in 344 articles. Out of 344 full-texts assessed, 251 articles were excluded. Thus, 93 articles that met the eligibility criteria were included.

BMSCs are a type of pluripotent stem cell that, under specific conditions, can differentiate not only into tissue cells from the mesodermal lineage, such as osteocytes, chondrocytes, and cardiomyocytes[16,17], but also undergo transdifferentiation across germ layers to form neurons, glial-like cells from the ectoderm, and hepatocytes, among others[18]. Silva et al[19] discovered that BMSCs express genes associated with both epithelial tissues and mesenchymal tissues, providing a theoretical basis for their multi-lineage differentiation potential at the gene level. Additionally, BMSCs possess self-renewal capacity. Tamir et al[20] found that approximately 90% of BMSCs are in the G0/G1 phase, which confirms their robust self-renewal capabilities.

BMSCs have no specific surface markers and generally exhibit low expression of major histocompatibility complex (MHC)-I molecules and do not express MHC-II molecules. They also do not express molecules required for T lymphocyte activation, such as Fas ligand and co-stimulatory molecules like B7-1, B7-2, and CD40 L[21]. This characteristic gives BMSCs low immunogenicity and strong immune-suppressive properties. Therefore, studies have shown that when co-cultured with allogeneic and xenogeneic T lymphocytes, BMSCs do not induce significant T cell proliferation but rather inhibit T cell proliferation[22]. In addition to being non-immunogenic, BMSCs are not targeted by CD8+ T cells, which allows them to evade cytotoxic T cell and natural killer cell killing, making them beneficial for successful autologous and allogeneic transplantations[23]. Furthermore, the antigenicity of BMSCs does not increase with their differentiation[24].

Indeed, it is evident that BMSCs possess the potential for multi-lineage differentiation and robust self-renewal capacity. Moreover, when transplanted into the body, they do not trigger significant rejection responses and can be allografted without causing immune rejection reactions[25,26]. The fact that BMSCs do not require the use of immunosuppressive drugs further adds to their appeal as seed cells for treating PNI, making them a promising candidate for potential applications in PNI therapy.

After PNI, if neurons have not died, their axons can undergo regeneration. SCs play a critical role in the repair of the peripheral nervous system. Following Wallerian degeneration of the peripheral nerve, SCs rapidly and massively proliferate, forming Bngner bands. They are involved not only in the formation, synthesis, and secretion of various NGFs but also in the synthesis and secretion of various extracellular matrix (ECM) components and other cell adhesion molecules. The above-mentioned NGFs, ECM, and cell adhesion molecules form gaps or tight junctions with adjacent axons, creating direct channels for the transfer of small molecules and information. These play an essential role in nerve injury regeneration and repair. Under specific conditions, BMSCs can differentiate into neural cells, including SC-like cells, and exert corresponding effects. In this section, we will explore the various functions of BMSCs in PNI repair and list the involved molecular mechanisms.

BMSCs are one of the most widely used sources of cells for nerve regeneration. After transplantation, they can differentiate into different cells, such as neurons, astrocytes, and SC-like cells, under the influence of different physiological microenvironments and express corresponding antigen markers. In vitro studies have found that BMSCs can be induced to differentiate into neural-like cells by antioxidants (such as dimethyl sulfoxide and -mercaptoethanol), cytokines [retinoic acid, basic fibroblast growth factor (bFGF), and epidermal growth factor], traditional Chinese medicine preparations (tetramethylpyrazine and baicalin), gene transfection, and other methods[27,28]. However, whether these induced neural-like cells possess the functional characteristics of normal neurons remains controversial. For instance, Hofstetter et al[29] successfully induced rat BMSCs to differentiate into neural cells using butylated hydroxyanisole but did not record the electrophysiological activity of mature neuronal cells. Some researchers believe that this phenomenon is not related to cell differentiation but rather cytotoxic changes[27]. On the other hand, other studies have shown successful induction of rat BMSCs into neural-like cells using a combination of bFGF, dimethyl sulfoxide, and butylated hydroxyanisole, with the capture of excitatory electrophysiological characteristics[27,28]. Wislet-Gendebien et al[30], through co-culturing, induced rat BMSCs to differentiate into neural cells that produced single action potentials and responded to neurotransmitters such as -aminobutyric acid, glycine, and glutamate. These findings suggest that BMSCs can differentiate into excitable neural-like cells in vitro.

In in vivo studies, Cuevas et al[31] injected 50000 bone MSCs (pre-labelled with bromodeoxyuridine BrdU) in 5 L of culture medium solution into the distal stump of transected sciatic nerve of the rats, and found that after 33 d of implantation, almost 5% of BrdU cells express Schwann cell-like phenotype. Dezawa et al[32] obtained GFP-expressing BMSCs (GFP-MSCs) by retroviral vectors, adjusted the concentration of GFP-MSCs to (1-2) 107 cells/mL, and then injected them into hollow fibres to make an artificial graft. The artificial graft was anastomosed to the cut end of the proximal nerve segment of the sciatic nerve in rats, and a large number of newly formed fibers were observed after 3 wk. They found that BMSCs had a myelination effect in regenerating nerve fibers through immunoelectron microscopy and confocal microscopy, indicating that BMSCs can differentiate into neuron-like cells and secrete a large amount of NGFs to induce axon growth. Additionally, BMSCs can directly transform into SCs to repair injured nerves, which has attracted considerable attention[33]. Furthermore, inflammatory cytokines such as tumor necrosis factor-alpha (TNF-) and interleukin (IL)-1 have been reported to affect the differentiation of MSCs, possibly driving MSCs toward specific cell phenotypes, such as astrocytes. Elevated levels of such pro-inflammatory cytokines can inhibit neuronal differentiation and promote the differentiation of BMSCs into astrocytes[34]. In conclusion, under specific conditions, BMSCs can differentiate into SCs and neural-like cells both in vitro and in vivo, facilitating nerve repair through cell replacement.

Neurotrophic factors have the function of promoting nerve growth and inducing cell differentiation into neural cells, and they can be used to induce the differentiation of BMSCs into neural cells. BMSCs can secrete a variety of neurotrophic factors, such as brain-derived neurotrophic factor (BDNF), NGF, vascular endothelial growth factor (VEGF), bFGF, and insulin-like growth factor (IGF)[14]. They upregulate the expression of VEGF receptor (VEGFR) and IGF1 receptor (IGF-1R) and promote the secretion of endogenous neurotrophic factors in the central nervous system. These neurotrophic factors are synthesized and retrogradely transported to nerve cells, transmitting information or paracrine signals to proximal and distal nerves. They bind to their specific receptors, such as NGF with NGF receptor A, BDNF with tyrosine receptor kinase B, and neurotrophin-3 (NT-3) and neurotrophin-4/5 with neurotrophic tyrosine receptor kinase 3. Activation or inhibition of signaling pathways such as PI3K/Akt, Ras-ERK, cAMP/PKA, and PLC--dependent pathways occurs, thereby promoting neuron survival, accelerating axonal and vascular growth, stimulating nerve fiber regeneration, preventing cell apoptosis, inducing SCs migration, proliferation, and myelination formation, and slowing down muscle atrophy, thus reversing the negative effects of PNI (such as preventing cell death caused by axonal injury)[5,35-37]. This improves the supportive microenvironment for neuron survival and regeneration[38] and exerts a neuroprotective effect on nerve cells[39]. Neuhuber et al[40] suggested that the neurotrophic factors produced by human BMSCs are essential for mediating axonal growth and functional recovery after spinal cord injury.

Wang et al[41] conducted a study and reported that using BMSCs transplantation in rats with PNI achieved results similar to autologous nerve transplantation, possibly due to the release of a large number of neurotrophic factors by BMSCs. Isele et al[42] found that the growth condition medium of BMSCs significantly reduced cross-cell-induced apoptosis in fetal rat hippocampal neurons, demonstrating a significant neuroprotective effect. During this process, they observed an increase in phosphorylation of MAPK/ERK and Akt. Blocking this protective effect occurred when using MAPK/ERK and PI3K/Akt specific inhibitors, suggesting that the neurotrophic factors secreted by BMSCs counteracted apoptosis stress response by activating these survival pathways and exerting a neuroprotective effect. They also discovered that stressed neuronal cells stimulated BMSCs to increase the secretion of trophic factors. In another study by Yang et al[43], they used BMSCs as support cells and injected them into a silk fibroin-based nerve conduit. This approach increased the expression of the SCs marker molecule S100 and enhanced the secretion of various neurotrophic growth factors such as BDNF, bFGF, and ciliary neurotrophic factor (CNTF). This, in turn, facilitated histological and functional recovery in rats with sciatic nerve injuries.

The ECM is a complex reticular structure composed of large molecules such as proteins and polysaccharides secreted by cells. It includes laminin, fibronectin, collagen, and other components. The ECM plays a crucial role in promoting cell proliferation and differentiation, supporting the transmission of important signals in the peripheral nervous system[44], which, together with neurotrophic factors and cell adhesion molecules, provides a favorable microenvironment for the survival of nerve cells and the formation of nerve connections[45-47]. Chen et al[48] mixed BMSCs cultured in vitro with gelatin and transplanted them into a 15 mm defect model of the rat sciatic nerve using silicone conduits. Compared to the gelatin-only control group, the experimental group showed improved walking behavior in rats, reduced atrophy of the gastrocnemius muscle, and decreased reduction in compound motor action potential amplitude, with a significant amount of regenerated axons observed. Both in vitro and in vivo, BMSCs synthesize and secrete various ECM components, including NGF, CNTF, BDNF, glial cell-derived neurotrophic factor (GDNF), as well as type I and type IV collagen, fibronectin, laminin, and other ECM molecules. After transplantation, both early and late stages of nerve regeneration are accompanied by high expression of neurotrophic factors. Wright et al[49] reported that BMSCs can stimulate neuronal development and mediate nerve regeneration by modulating the expression of ECM components such as chondroitin sulfate proteoglycans, myelin-associated glycoproteins, and Nogo-A.

Cell adhesion molecules are also critical for axon guidance, including integrins, neural cell adhesion molecules, and calcium-binding proteins such as N-cadherin. Among them, neural cell adhesion molecules may preferentially promote the growth of sensory axons[50]. BMSCs can express various factors related to cell adhesion, such as Ninjurins 1 and 2, Netrin 4, Robo 1, and Robo 4[51-53]. These factors are recognized as neuroregenerative factors and effectively promote axonal growth and cell migration. In summary, BMSCs improve the microenvironment for neuron survival and regeneration through paracrine secretion of neurotrophic factors, ECM factors, adhesion molecules, and various other mechanisms. By promoting the regeneration of damaged neurons, BMSCs contribute to the repair of neural functions.

After PNI occurs, the blood vessels within the nerves are damaged. Therefore, promoting vascular regeneration and restoring blood circulation are essential for the recovery of the normal neural tissue environment. Peripheral nerve regeneration is closely related to angiogenesis, which is a crucial process in the repair of peripheral nerves. VEGF is considered an effective factor for both angiogenesis and neuron generation, and it has long been recognized for its importance in promoting neuron survival and SCs proliferation. Popovich et al[54] reported that BMSCs can secrete various neuroprotective trophic factors such as BDNF, NGF, and VEGF in an autocrine and/or paracrine manner, which can upregulate the expression of these factors, thereby promoting local microvascular regeneration, nerve regeneration, and reconstruction, and ultimately facilitating the repair of injured cells. Induced SCs-like cells from BMSCs have been found to exhibit enhanced immunostaining for VEGF, suggesting that BMSCs may also promote blood vessel formation[55]. BMSCs can also increase the expression levels of endogenous VEGF and its receptor VEGFR2 in the ischemic penumbra, thereby promoting neovascularization[15]. Zurita and Vaquero[56] also observed that blood vessel wall cells in newly regenerated neural tissue at the site of spinal cord injury were differentiated from injected BMSCs. These studies indicate that BMSCs can promote angiogenesis through paracrine secretion of VEGF, and the newly formed blood vessels can, in turn, facilitate the repair of peripheral nerve injuries.

Myelination is another essential process in the regeneration of PNI, determining the quality and functional recovery of nerve regeneration[5,35,47]. Typically, myelination can be achieved by promoting endogenous repair mechanisms or providing an exogenous source of myelinating cells, leading to subsequent nerve function restoration[47]. In a study conducted by Kizilay et al[57], the systemic application of BMSCs was explored in a PNI compression model. Wistar albino rats were used, and the sciatic nerve was compressed for 5 min to create the model. Approximately 5 105 BMSCs were injected intravenously. The results showed that animals treated with BMSCs exhibited higher nerve conduction velocity, compound action potential, and axon numbers compared to the control group. In addition, myelin damage was less severe in the BMSC-treated group, suggesting that systemic application of BMSCs has a positive impact on both myelination and axon survival in the peripheral nerve compression model.

SCs and various types of adult stem cells (in the form of SCs-like cells) have the ability to form myelinating neuronal cells and regenerate nerves. During the regeneration process after PNI, intracellular cAMP levels are elevated when SCs or SCs-like cells further differentiate into myelin-forming cells. This leads to the synthesis and secretion of abundant myelin proteins, such as myelin basic protein, myelin protein zero, peripheral myelin protein 22 (PMP22), and other proteins that are crucial for myelin structure and function. This promotes remyelination during and after regeneration[5,47] and increased expression of IGF-1R and neurofilament type 1 and type 3 enhances axon alignment and myelination gene expression, resulting in increased myelin thickness and internodal length[35,50]. BMSCs also provide various cytokines and growth factors for nerve regeneration[58], including NGF, NT-3, VEGF, PMP22[59-62], and more. Zhao et al[63] also demonstrated that exosomes from BMSCs upregulate the expression of PMP22, VEGF, NGF receptors, and S100 protein, promoting increased neuronal length and axon diameter in the dorsal root ganglion. These protein factors play crucial roles in peripheral nerve regeneration. During the repair process, BMSCs not only directly affect SCs through their neurotrophic functions[64] but may also differentiate towards SCs directionally.

BMSCs, in addition to their ability to differentiate into neuron-like cells[65], also stimulate and induce axonal growth[66], and play an important role in maintaining the normal structure and function of myelin sheaths[67,68]. BMSCs can promote the repair of damaged nerves by regulating the expression of myelination-related genes. For instance, differentiation of BMSCs into SC-like cells can enhance the mRNA expression of myelin-associated factors, significantly increasing the number of myelinated axons, thereby promoting the functional recovery of the facial nerve[69]. In conclusion, MSCs promote myelination and axonal regeneration through various mechanisms, including the secretion of neurotrophic factors, direct interactions with neurons, and upregulation of genes involved in myelination. These combined effects contribute to enhanced axonal growth and improved functional recovery after PNI.

After PNI, various immune cells and cytokines are present, and the coordination of local inflammatory response is essential for the recovery of PNI. BMSCs possess significant immunomodulatory properties, which can promote neural tissue regeneration and alleviate inflammation, therefore making them valuable in PNI treatment. BMSCs can exert immunomodulatory effects by regulating the expression of various cytokines. IL-6 is a multifunctional cytokine produced by macrophages and fibroblasts during PNI[70]. IL-17 is produced by activated CD4+ T cells and can increase the production of pro-inflammatory cytokines and neutrophil chemoattractants, showing elevated levels after PNI[71]. Studies by Ge et al[72] found that BMSCs can secrete high levels of IL-6 to modulate the balance of CD4+ T cell subgroups, promote the proliferation and differentiation of T helper type 17 (Th17) cells that secrete IL-17, and subsequently stimulate prostaglandin E2 secretion. Elevated prostaglandin E2 levels then inhibit Th17 cell secretion of IL-17, achieving therapeutic effects for facial nerve injury. The increased expression of IL-10 protein is associated with regeneration of myelin protein. Research by Cui et al[73] revealed that IL-10-stimulated BMSCs can inhibit the expression of the pro-inflammatory cytokines TNF- and IL-1. Fan et al[74] suggested that this may be achieved by reducing the release of the pro-inflammatory cytokines IL-2, interferon-, and TNF- and increasing the secretion of IL-10 in lymphocyte supernatant and serum, thereby promoting neural regeneration.

BMSCs can modulate the polarization of macrophages, promoting their transition from the pro-inflammatory M1 phenotype to the anti-inflammatory M2 phenotype. This shift in macrophage polarization is crucial for controlling inflammation and establishing an environment for tissue repair and regeneration. Zhong et al[75] reported that BMSCs secrete GDNF, which converts the damaging M1 phenotype in microglia to the regenerative M2 phenotype, thereby suppressing neural inflammation. This process may be related to inhibiting the nuclear factor-kappaB signaling pathway and promoting the PI3K/AKT signaling pathway.

Another important aspect of MSC-mediated immune regulation is the release of extracellular vesicles (EVs), including apoptotic bodies, exosomes, microvesicles, etc.[76], which contain bioactive components. These EVs are considered an intriguing non-cellular therapy due to their low immunogenicity and ability to mediate cell-to-cell communication and modulate the function of recipient immune cells, contributing to the overall immunomodulatory effects of BMSCs. BMSCs EVs may exhibit similar anti-inflammatory functions as BMSCs themselves by decreasing the levels of inflammatory cytokines and enhancing anti-inflammatory responses. For instance, Schfer et al[77] found that BMSCs can release soluble mediators such as TNF- and IL-1 to alleviate inflammation after PNI. It is evident that BMSCs can exert their immunomodulatory effects through various mechanisms, including regulating the expression of various cytokines, regulating macrophage polarization, releasing EVs, and secreting soluble factors. These effects can help control inflammation, prevent autoimmune reactions, and create a more favorable environment for nerve repair and regeneration following PNI.

In summary, BMSCs play a crucial role in promoting PNI repair and regeneration through various mechanisms (Table ). First, BMSCs are able to differentiate into nerve cells (such as neurons and SCs) to replace damaged nerve cells and facilitate nerve regeneration. Second, they secrete neurotrophic factors, ECM molecules, and adhesion molecules, while also exerting immunomodulatory effects, creating a supportive microenvironment for the growth, differentiation, and survival of nerve cells. Third, BMSCs promote the formation of new blood vessels to ensure the necessary blood supply for the repair and accelerated regeneration of damaged nerves. Lastly, by synthesizing and releasing of proteins related to myelination and axon regeneration, BMSCs enhance the growth of myelinated axons and ultimately promote neuron regeneration. BMSCs utilize these different mechanisms to promote the repair and regeneration of damaged nerve cells and enhance the functional recovery after PNI. Utilizing these pathways can significantly enhance the therapeutic potential of BMSCs in PNI treatment.

Mechanisms of bone marrow mesenchymal stem cell therapy for peripheral nerve injury

The unique mechanisms of action of BMSC make them promising candidates for the treatment of PNI. In this section, we will explore the various application methods of MSCs in PNI treatment (Figure ), analyzing the advantages and disadvantages of each approach in order to comprehensively explore their potential in PNI treatment.

Application of bone marrow-derived mesenchymal stem cells in the treatment of peripheral nerve injury. Bone marrow-derived mesenchymal stem cells can be isolated from bone marrow, expanded in vitro, and directly transplanted into damaged nerve tissue. They can be loaded onto nerve conduits, which provide structural support, using tissue engineering techniques. Additionally, bone marrow-derived mesenchymal stem cells can be genetically modified with neurotrophic factors before being applied to the treatment of peripheral nerve injury to promote neuronal repair and regeneration. PNI: Peripheral nerve injury; MSC: Mesenchymal stem cell.

BMSCs have self-renewal and multi-lineage differentiation capabilities that make neuronal regeneration and nerve function recovery possible, rendering them one of the best choices for stem cell therapy in PNI treatment. Apart from their regenerative potential, BMSCs have been shown to migrate to the injury site and home to the injured area, exhibiting potential for targeted therapy[78,79]. Furthermore, BMSCs do not significantly stimulate the proliferation of T cells nor serve as a target for CD8+ T cells. Thus, when applied in autologous or allogeneic transplantation, they can evade the killing and clearance by immune cells in the body, further exerting their reparative effects. Cuevas et al[31] and Cuevas et al[80] cultured BMSCs from adult rats, labeled them with BrdU, and then injected them into the distal stump of the 5 mm-deficient sciatic nerve in rats. At 18 d and 33 d post-surgery, footprint analysis showed significant improvement in the motor function of the rat limbs compared to the control group injected with only culture medium. Immunofluorescence double-labeling showed that BrdU-labeled cells survived for at least 33 d after surgery, and nearly 5% of the cells expressed the S100 phenotype of SCs. In March 2004, they conducted a similar study on the long-term recovery of rat limbs 180 d after BMSC transplantation, finding that BMSCs continued to have a promoting effect on long-term recovery after surgery[80]. This experiment proves the great potential of BMSCs in peripheral nerve regeneration and lays the foundation for their application in the field of peripheral nerve regeneration. Wang et al[41] investigated the reparative effects of BMSCs by injecting them into the muscles after sciatic nerve injury in rats, and the results showed that the number of regenerating nerve fibers and spinal cord ventral horn neurons increased significantly, as well as a significant increase in regenerated myelin sheath thickness, which indicated that transplantation of BMSCs in PNI rats can achieve similar results as autologous nerve transplantation. Hu et al[81] transplanted BMSCs to repair a 50 mm midline nerve injury in monkeys and found that the healing process was similar to that of autologous transplantation, showing good functional and morphological outcomes. Another study found that when BMSCs were directly transplanted around the sciatic nerve stump, they induced axonal growth by differentiating into neuron-like cells and secreting neurotrophic factors[32]. They also differentiated into SCs to repair the injured nerves[33] and promoted remyelination of regenerating nerve fibers. From this, it can be seen that direct transplantation of BMSCs has played a positive role in repairing various PNI-damaged nerves. However, the invasive procedures required for obtaining BMSCs and the limited quantity of cells obtained, as well as the reduced proliferative and differentiation abilities with increasing patient age, have restricted the research and application of BMSCs in clinical settings.

Scaffold technology has become a hot topic in tissue engineering research in recent years, and nerve conduits are a type of artificial tubular scaffold. BMSCs can simulate the structure and function of the human nervous system when loaded onto nerve conduits and connecting on both sides of the nerve stump. Nerve conduits can be made from natural materials such as chitosan and collagen or synthetic materials such as polyglycolic acid and polylactic acid. Each material has its own characteristics, generally inducing nerve axon regeneration and preventing infiltration of surrounding tissues to interfere with nerve repair. By loading BMSCs onto nerve conduits, not only does it achieve the neurotrophic guidance function of the nerve conduit, but it also provides a space for BMSCs and nerve axon regeneration induction, which helps to promote the effects of BMSCs in promoting nerve growth and regulating the microenvironment of the injury site[82]. In the process of repairing injured nerves using tissue engineering methods, comparing the transplantation effects of nerve conduits with and without BMSCs, it was found that the number and diameter of nerve axons in the experimental group significantly increased, and the improvement of nerve function was significantly better than that in the control group[83].

Costa et al[84] inplanted BMSCs into poly(L-lactic acid) nerve conduit scaffolds for repairing facial nerve defects in rats. The results showed that BMSCs could successfully integrate into the conduit, survive within the nerve tissue, and maintain their phenotype for up to 6 wk. In another study, researchers loaded BMSCs into chitosan nerve conduits and observed cell survival and proliferation within the conduit for 8-16 wk, which effectively promoted the repair of an 8 mm nerve defect[85]. Subsequent research by this team demonstrated that BMSC-loaded chitosan nerve conduits not only accelerated the efficiency of nerve repair but also improved the quantity and quality of regenerated nerve fibers, achieving therapeutic effects comparable to autologous nerve transplantation[86]. The degradation products of nerve conduit materials often trigger local immune reactions, leading to an inflammatory state at the site of injury, which can affect the repair outcome. However, in a study by Hsu et al[87], researchers modified chitosan nerve conduits with laminin to enhance the adhesion capability of BMSCs within the conduit. They observed that BMSCs successfully inhibited the local inflammatory response caused by chitosan degradation, resulting in improved promotion of nerve repair. Other experimental studies have also used BMSCs implanted in nerve conduits made of different materials, such as fibroin gel conduits[88], polylactic-co-glycolic acid conduits with ECM gel[89], and polyglycolic acid conduits[90], to intervene in PNI animal models, and all achieved favorable results.

Although encouraging results have been obtained in animal experiments, further research is still needed to optimize the design of nerve conduits, determine the optimal combination of BMSCs and biomaterials[91], and assess the long-term safety and efficacy of nerve conduits in clinical settings[92]. By addressing these issues, the use of BMSCs in tissue engineering approaches may have a more significant impact on PNI treatment, providing new strategies to promote neural functional recovery and improve the quality of life for patients.

Gene-modified BMSCs have also gained increasing attention in tissue engineering research. In the field of neural repair tissue engineering, the main purpose of gene modification is to design target cells to overexpress growth factors, migration molecules, and adhesion molecules, as well as to inhibit the expression of defective genes. NT-3, NT-4, BDNF, NGF, CNTF, bFGF, and others are major neural growth factors suitable for peripheral nerve gene delivery, as they can provide a suitable microenvironment for the survival and axonal growth of BMSCs. In a study by Zhang et al[93] in 2015, BMSCs transfected with BDNF and CNTF were used for the treatment of rat sciatic nerve injuries. The results showed that BDNF- and CNTF-transfected BMSCs combined with nerve transplantation significantly improved the sciatic nerve function index, promoted the recovery of muscle activity, and increased the thickness of regenerating nerve myelin sheaths. This indicates that this approach is effective in promoting axonal growth and facilitates nerve repair in PNI. In another study[94], BDNF was successfully transfected into BMSCs using gene engineering technology, and the transfected BMSCs were combined with decellularized allogeneic nerve grafts to repair peripheral nerve defects. The results showed a significant improvement in the repair effectiveness of the nerve grafts and the morphology of the injured nerves. Gene-modified MSCs have multiple potentials in the treatment of PNI. However, since gene therapy is still in the experimental stage, its application in clinical settings requires addressing numerous challenges, such as the selection of diverse target genes, stable expression of target genes in the host, combination therapy with multiple genes, and ethical considerations.

Unlike the central nervous system, the peripheral nervous system has the ability for self-regeneration and repair after injury. However, this endogenous repair is limited, and extensive nerve damage cannot be fully repaired. Cell therapy is considered to be an important direction for future medical development, and in recent years, the field of PNI neural regeneration and repair has made vigorous progress, with enormous market potential and clinical application value. BMSCs have the advantages of abundant sources, easy and simple procurement, being easy to isolate and cultivate, and the potential for rapid expansion under certain conditions. Additionally, autologous BMSCs transplantation avoids ethical issues and immune rejection, offering broad prospects for PNI treatment. In this paper, we have reviewed the current biological characteristics of BMSCs related to PNI, summarized the mechanisms by which BMSCs promote PNI neural regeneration and repair, and explored various application methods of BMSCs in PNI, confirming the potential of BMSCs in treating PNI.

However, most research on BMSCs transplantation for PNI intervention is still in the pre-clinical stage and has not yet had significant implications for clinical practice, and there are also certain limitations, such as the lack of specific surface markers on BMSCs[21], which poses some difficulties in identifying cultured BMSCs, and the lack of standardized treatment regimens, where many times after BMSC transplantation, the survival rate is not high, and the proportion of differentiation into neurons is low, resulting in unsatisfactory nerve repair effects. There are also safety issues with BMSC transplantation, where inducers transplanted into the human body along with BMSCs can cause varying degrees of damage to the human body, and there is a possibility of BMSCs transforming into malignant tumors[95]. These issues that need to be resolved point to a certain direction for future research, such as establishing standardized procedures for the extraction, identification, and cultivation of BMSCs; further clarifying the therapeutic mechanisms of BMSCs; and observing the safety of BMSCs applications. The choice of BMSCs application methods in PNI, such as direct transplantation, tissue engineering, and gene engineering, also requires further investigation. In conclusion, BMSCs transplantation offers broad prospects for PNI treatment, but significant theoretical and experimental research are needed before its clinical application can be fully developed and perfected.

Xiong-Fei Zou, Department of Orthopedic Surgery, Peking Union Medical College Hospital, Beijing 100730, China.

Bao-Zhong Zhang, Department of Orthopedic Surgery, Peking Union Medical College Hospital, Beijing 100730, China. nc.hcmup@hzbgnahz.

Wen-Wei Qian, Department of Orthopedic Surgery, Peking Union Medical College Hospital, Beijing 100730, China.

Florence Mei Cheng, College of Nursing, The Ohio State University, Ohio, OH 43210, United States.

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Stem Cell Therapy Mexico: R3 Stem Cell Unveils Innovative and Affordable Non-Invasive Solutions  openPR

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Stem Cell Therapy Mexico: R3 Stem Cell Unveils Innovative and Affordable Non-Invasive Solutions - openPR

'Didn't know this would be possible': Autistic teen's mom on stem cell therapy benefits  WZTV

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'Didn't know this would be possible': Autistic teen's mom on stem cell therapy benefits - WZTV

November 16, 2022

Karen M. Wai, MD, Theodore Leng, MD, MS, and Jeffrey Goldberg, MD, PhD, Byers Eye Institute at Stanford, Stanford University School of Medicine, Palo Alto, CA

In recent years, the potential of stem cell-based therapies to treat a wide range of medical conditions has given hope to patients in search of novel treatments or cures. At the same time, thousands of rogue clinics have sprung up across the U.S and around the world, offering stem cell-based therapies before being tested for safety and efficacy. When communicating to the public about stem cell-based therapies, it is important to put any treatment claims in context.

Stem cell-based therapies include any treatment that uses human stem cells. These cellshave the potential to develop into many different types of cells in the body. They offer a theoretically unlimited source of repair cells and/or tissues. (For more about stem cells, seehttps://stemcells.nih.gov.)

Over the past three decades, the Food and Drug Administration (FDA) has approved several stem cell-based products. These include bone marrow transplants, which have been transformational for many cancer patients, and therapies for blood and immune system disorders.1 Other approved treatments include dental uses for gum and tissue growth and in skin for burns. Since the early 2000s, stem cell-based therapies have been explored in many eye diseases, including age-related macular degeneration and glaucoma.2 Stem cell-based therapies are also being explored for neurodegenerative diseases such as stroke and Alzheimers disease, and for countless other conditions.

Over time, we expect that breakthroughs will continue with stem cell-based therapies for many conditions. However, at this time, rogue clinics, driven by profits, are taking advantage of patients desperate for cures and are claiming dramatic results, often exaggerated in sensational media testimonials. The clinics may mimic legitimate practices. They may extract a patients own stem cells, concentrate or modify the cells, and then re-inject them. Some manufacturers offer stem cell-based derived products, such as biologic eye drops made with placenta extract or amniotic fluid to treat dry eye. Clinics may provide misleading information and advertise their practice as running clinical trials. However, these clinics almost always work without FDA regulatory approval and outside of legitimate clinical trial approaches.

These unproven, unregulated stem cell treatments carry significant risk. The risks range from administration site reactions to dangerous adverse events. For example, injected cells can multiply into inappropriate cell types or even dangerous tumors. A 2017 report described one Florida clinic that blinded patients with stem cell eye injections.3

The Pew Charitable Trusts gathered 360 reports of adverse events related to unapproved stem cell therapies, including 20 cases that caused death.4 Further, adverse events are likely underreported because these products are not FDA approved or regulated. Many unproven stem cell-based therapies cost thousands of dollars to patients and are not covered by insurance. Further, even if patients avoid adverse events from these therapies, they may suffer consequences from delaying evidence-based treatments.

The FDA has made substantial progress toward regulation of stem cell-based therapies. In 2017, it released guidance under the 21st Century Cures Act that clarifies which stem-cell based therapies fall under FDA regulation. It also better defined how the agency will act against unsafe or unregulated products.5 As of May 2021, the FDA has more strongly enforced compliance for clinics that continue to market unproven treatments.6

Despite this increased regulation, rogue clinics are still relatively commonplace. A 2021 study estimated that there are over 2,500 U.S. clinics selling unproven stem cell treatments.7Patients at these clinics are often led to believe that treatments are either approved by the FDA, registered with the FDA, or do not require FDA approval. It is important to recognize that there are limits to the FDAs expanded reach, especially when it is targeting hundreds of clinics at once. Our clinic at Stanford recently cared for a patient who had received stem cell injections behind his eyes, where he developed tumors that ultimately ruined vision in both eyes.

Progress in stem cell science is rapidly translating to the clinic, but it is not yet the miracle answer we envision. With time, stem cell-based therapies will likely expand treatment options. People considering a stem-cell based therapy should find out if a treatment is FDA-approved or being studied under an FDA-approved clinical investigation plan. This is called an Investigational New Drug Application. Importantly, being registered with ClinicalTrials.gov does not mean that a therapy or clinical study has been authorized or reviewed by the FDA. For more information about stem cell therapies, visit http://www.closerlookatstemcells.org, a resource from the International Society for Stem Cell Research.

As we look hopefully to the future, we need greater awareness of the current limitations of stem cell therapy and the dangers posed by unregulated stem cell clinics. Strong FDA regulation and oversight are important for ensuring that stem cell-based therapies are safe and effective for patients. Accurate communication to the public, careful advocacy by physicians, and education of patients all continue to be crucial.

References:

1 U.S. Food and Drug Administration, Approved Cellular and Gene Therapy Products, Sept. 9, 2022,https://www.fda.gov/vaccines-blood-biologics/cellular-gene-therapy-products/approved-cellular-and-gene-therapy-products.

2 Stern JH, Tian Y, Funderburgh J, Pellegrini G, Zhang K, Goldberg JL, Ali RR, Young M, Xie Y, Temple S. Regenerating Eye Tissues to Preserve and Restore Vision. Cell Stem Cell. 2018 Sep 6;23(3):453. doi: 10.1016/j.stem.2018.08.014. Erratum for: Cell Stem Cell. 2018 Jun 1;22(6):834-849. PMID: 30193132.

3 Kuriyan AE, Albini TA, Townsend JH, Rodriguez M, Pandya HK, Leonard RE 2nd, Parrott MB, Rosenfeld PJ, Flynn HW Jr, Goldberg JL. Vision Loss after Intravitreal Injection of Autologous "Stem Cells" for AMD. N Engl J Med. 2017 Mar 16;376(11):1047-1053. doi: 10.1056/NEJMoa1609583. PMID: 28296617; PMCID: PMC5551890.

4 The Pew Charitable Trusts, Harms Linked to Unapproved Stem Cell Interventions Highlight Need for Greater FDA Enforcement, June 1, 2021,https://www.pewtrusts.org/en/research-and-analysis/issue-briefs/2021/06/harms-linked-to-unapproved-stem-cell-interventions-highlight-need-for-greater-fda-enforcement.

5 U.S. Food and Drug Administration, FDA announces comprehensive regenerative medicine policy framework, Feb. 2, 2022,https://www.fda.gov/news-events/press-announcements/fda-announces-comprehensive-regenerative-medicine-policy-framework.

6 U.S. Food and Drug Administration, FDA Extends Enforcement Discretion Policy for Certain Regenerative Medicine Products, July 7, 2020,https://www.fda.gov/news-events/press-announcements/fda-extends-enforcement-discretion-policy-certain-regenerative-medicine-products.

7Turner L. The American stem cell sell in 2021: U.S. businesses selling unlicensed and unproven stem cell interventions. Cell Stem Cell. 2021 Nov 4;28(11):1891-1895. doi: 10.1016/j.stem.2021.10.008. PMID: 34739831.

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Putting Stem Cell-Based Therapies in Context | National Institutes of ...

Eggs from men, sperm from women: Stem cell therapy may just turn reproduction upside down!  The Economic Times

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Eggs from men, sperm from women: Stem cell therapy may just turn reproduction upside down! - The Economic Times

Int J Mol Sci. 2022 Mar; 23(5): 2850.

Akiko Maeda, Academic Editor and Ali Gorji, Academic Editor

Received 2022 Jan 25; Accepted 2022 Mar 3.

Regenerative medicine is a new and promising mode of therapy for patients who have limited or no other options for the treatment of their illness. Due to their pleotropic therapeutic potential through the inhibition of inflammation or apoptosis, cell recruitment, stimulation of angiogenesis, and differentiation, stem cells present a novel and effective approach to several challenging human diseases. In recent years, encouraging findings in preclinical studies have paved the way for many clinical trials using stem cells for the treatment of various diseases. The translation of these new therapeutic products from the laboratory to the market is conducted under highly defined regulations and directives provided by competent regulatory authorities. This review seeks to familiarize the reader with the process of translation from an idea to clinical practice, in the context of stem cell products. We address some required guidelines for clinical trial approval, including regulations and directives presented by the Food and Drug Administration (FDA) of the United States, as well as those of the European Medicine Agency (EMA). Moreover, we review, summarize, and discuss regenerative medicine clinical trial studies registered on the Clinicaltrials.gov website.

Keywords: regenerative medicine, stem cell therapy, mesenchymal stem cell, clinical trial

Despite the progress in medical science, there still exist various diseases in the world for which there is no suitable treatment. People affected by incurable disorders typically use treatment methods intended to decrease the somatic and psychological symptoms and, in these situations, the physician offers treatment methods only to manage the disease, not treat it. Therefore, researchers are attempting to develop new treatment methods to not only control the symptoms of, but also to treat those diseases for which no cure is available at present.

Regenerative medicine is considered a promising new source of treatment for untreatable diseases in modern science [1]. Regenerative medicine is a multidisciplinary field including cell biology, genetic, biomechanics, material science, and computer science [2,3], the ultimate target of which is returning normal function to defective cells and tissues [4]. Since the discovery of stem cells and the spread of awareness regarding their unique properties, they have been defined as therapeutic agents for organ and tissue repair, and so are widely considered good candidates for regenerative medicine, due to their many potential applications [5]. Regenerative medicine is now regarded as an alternative to traditional drug-based treatments by researchers who study its potential applications in various diseases, including degenerative diseases, among others [6,7,8,9,10]. The main concept of regenerative medicine is implied tissue/organ regeneration using cells and, to reach this target, different kinds of cells have been used. However, various studies have indicated that cell therapy is restricted by a few limitations. In recent years, different alternatives have been introduced for cell therapy in order to resolve these limitations, including the improved application of stem cells for the restoration of tissue, such as the combination of cells with scaffolds, cell cultures with suitable biochemical properties, gene editing, and the immunomodulation of stem cells, as well as the use of stem cell derivatives [11,12,13,14,15]; however, the use of these alternatives clinically may be postponed, as more preclinical studies are required due to their status as newer technologies [16].

Stem cells are a group of immature cells that have the potential to build and recover every tissue/organ in the body due to their unique proliferative, differentiation, and self-renewal abilities [17]. Stem cells provide therapeutic effects which improve physical development by regenerating damaged cells to assist in organ recovery. Relying on the natural abilities of stem cells, researchers have used their biological mechanisms for stem-cell-based therapy. The mechanisms of action through which stem cells can promote the regeneration of tissue are diverse, including (1) inhibition of inflammation cascades [18,19], (2) reduction of apoptosis [20,21], (3) cell recruitment [22,23], (4) stimulation of angiogenesis [24,25], and (5) differentiation [26]. The cause of a disease is a vital consideration in selecting the proper stem cell mechanism and in the regeneration of tissue/organs using stem cells. Many examinations must be carried out to determine the main mechanisms involved in treatment when these cells are to be used in clinical practice, and the convergence of stem cell therapeutic mechanisms and disease mechanisms is expected to increase the chance of developing cures through stem cell applications.

From 1971 to 2021, 40,183 research papers were published regarding stem-cell-based therapies. All of these studies were conducted around discoveries and for the goal of Stem Cell Therapy based on the therapeutic efficacy of stem cells [27]. As basic stem cell research has soared over the past few years, translation research, a relatively new field of research, has recently greatly developed, making use of basic research results to develop new treatments. Although many articles on stem-cell-based therapies are published annually and their number increases every year, the number of clinical trial studies has not increased rapidly. Furthermore, among these studies, only a small portion of them can receive full regulatory approval for verification as treatment methods. Although one reason for this difference is due to the need for various prerequisite preclinical studies before carrying out a clinical trial study, the main reason is due to the sharply defined guidelines which prevent the translation of many preclinical studies to clinical trials.

In this review, we provide a general overview regarding the translation of stem cell therapies from idea to clinical service. Understanding the step-by-step knowledge underlying the translation of ideas to medical services is the first step in introducing a new treatment method. In this review, we divide this pathway into four levels, including idea evaluation, preclinical studies, clinical trial studies, and clinical practice. We focus not only on understanding each levels requirements, but also discuss how an idea is assessed during the transition from one level to the next and, finally, move on to marketing.

If a researcher has an idea regarding regenerative medicine using stem cells that inspires their use in a study, it must first be evaluated. During the evaluation step, it is important to select the target disease and make sure that the mechanism causing the disease is understood. Disease-related mechanisms refer to the cellular and molecular processes by which a particular disorder is caused [28,29], and stem-cell-based therapies are considered a treatment method intended to compensate for the disruption caused by such mechanisms in order to finally restore the defective tissue. Multiple mechanisms cause diseases [30,31,32]; however, stem cells, with their tremendous differentiation, self-renewal, angiogenesis, anti-inflammation, anti-apoptotic, and immunomodulatory potentials, as well as their capacity for induction of growth factor secretion and cell signaling, can affect these mechanisms [33,34,35,36,37].

After subject evaluation, preclinical studies should be carried out to determine whether the idea has any potential to treat the disease, and the safety of the final product should be assessed in an animal model of the target disease [38,39,40]. Preclinical studies are composed of in vitro and in vivo studies. In vitro experiments are performed with biological molecules and cells based on various hypotheses and, during the in vitro evaluation, a new treatment method is assayed in this controlled environment [39]. In contrast, during in vivo studies, as controlling all biological entities is impossible, the new product may be affected by various factors and thus present different effects. The general purpose of a preclinical study is to present scientific evidence supporting the performance of a clinical study, and the following are required for a decision to move forward to clinical study: (i) the feasibility and establishment of the rationale (e.g., validation, separation of active ingredients in vitro, and determination of its mechanism in vivo), (ii) establishment of a pharmacologically effective capacity (e.g., secure initial dose verification), (iii) optimization of administration route and usage (e.g., safe administration method, repeated administration, and interval verification), (iv) identification and verification of the potential activity and toxicity (e.g., toxicity analysis according to single and repetitive testing), (v) identification of the potential for special toxicity (e.g., genetic, carcinogenic, immunological, and neurotoxic analyses), and (vi) determination of whether to continue or discontinue development of the treatment [41,42].

In principle, any idea regarding stem cell therapy should be assessed using comprehensive studies (i.e., in vitro and in vivo) before a clinical trial is considered, and the results of these studies should be proved by competent authorities. It can be easy during an in vitro study to create manipulative biological environments such as through the use of genetic mutation, drug testing, and pharmaceuticals, and it is easy to observe changes through the application of manipulated variables through living cells [43,44,45]. However, given the many associated variables, such as molecular transport through circulating blood and organ interactions, it is hard to say whether such a study can completely mimic the in vivo environment [43,44,45]. Before application in patients, in vivo experiments are conducted after in vitro experiments in order to overcome these weaknesses.

Many researchers use rodents for in vivo studies, due to their anatomical, physiological, and genetic similarities to humans, as well as their other unique advantages including small size, ease of maintenance, short life cycle, and abundant genetic resources [46]. The strength of in vivo studies is that they can supplement the limitations of in vitro studies, and the outcomes of their applications can be inferred in humans through the use of human-like biological environments. To establish in vivo experiments for stem cell therapies, the most correlated animal model should be selected depending on the specific safety aspects to be evaluated. Where possible, cell-derived drugs made for humans should be used for proof-of-concept and safety studies [47]. Homogeneous animal models can also be utilized as the most correlated systems in proof-of-concept studies [48].

Furthermore, in vivo studies require ethical responsibilities and obligations to be upheld according to experimental animal ethics. In other words, unnecessary and unethical experiments must be avoided. Summing up the above, we can see that both in vitro and in vivo approaches are used in preclinical studies, which should be carried out before clinical trial applications based on various interests.

Several factors must be considered in different in vitro and in vivo studies, including cell type determination, cell dose specification, route of administration, and safety and efficiency.

As expectations rise for regenerative treatment through the application of stem cell therapies, the number of applications of various types and stem cell sources has increased, and stem cell therapies have diversified from autologous to allogenic to iPSCs. These stem cell treatments can vary in risk, depending on the cell manufacturing process [49], among other factors, and in clinical experience, such that all types of stem cell treatments must be evaluated on the same basis [50]. Therefore, the strengths and weaknesses of each type of stem cell should be identified in order to determine the maximum therapeutic effect of stem cells in various diseases. This will enable us to build disease-targeted stem cells by applying the appropriate stem cells to the appropriate diseases. Below, we briefly discuss the characteristics of various stem cells.

MSCs are lineage-committed cells that divide into mesenchymal systems, primarily fatty cells, chondrocytes, and osteocytes [51]. It is well known that MSCs can be differentiated into dry cells, nerve cells, glioma cells, and skeletal muscle cells under proper in vitro culture conditions [52,53,54,55,56,57]. MSCs are primarily derived from myeloid and adipose tissues [58,59]. At present, MSCs are also isolated from many other tissues, such as the retina, liver, gastric mucosa, tendon, cartilage, placenta, cord blood, and blood [60,61,62,63]. The biggest characteristics of MSCs are their immunosuppressive functions, which prevent the proliferation of activated T cells through immunosuppressive cytokine secretion and suppression of programmed cell death signaling [64,65]. Due to this role, they have been spotlighted as a potential treatment for immune-related inflammation and disease. The initial clinical application of MSCs was in a case of patients with severe graft versus host disease (GVHD), and these cells have since been well applied in clinical practice, as evidenced through various studies [66,67,68].

MSCs have a variety of characteristics according to their organ of origin [69]. BM-MSCs, which are isolated from bone marrow, are useable in both autologous and allogenic contexts, and can perform stromal functions. However, the process of cell isolation from bone marrow is not only accompanied by the risk of pain and infection, but also has a lower efficiency of collection than other MSC sources. Furthermore, these cells have a longer doubling time (DT) in comparison to MSCs derived from other sources (approximately 60 h) [70]. Compared to BM-MSCs, AD-MSCs are not only easy to collect, but are also 100 to 500 times more efficient to harvest and have a shorter DT (approximately 20 h) [71]. However, these are adipose-derived stem cells that have a strong characteristic of adipogenic differentiation, such that they can be suggested as a valid alternative to BM-MSCs, but their nature must be considered regarding proper culture and body environment. Furthermore, there are concerns that these factors may affect the efficacy of treatment, as the amount of cytokines secreted is significantly lower when compared to BM-MSCs [72]. MSCs extracted from the umbilical cord (UC-MSCs) have come into the spotlight to compensate for these issues: UC-MSCs not only have the advantage of being easily collected compared to other stem cells, but also avoid ethical or donor age issues. They have superior proliferation and differentiation capabilities compared to BM-MSCs and AD-MSCs, and their DT has been reported as 24 h [69,73]. UC-MSCs are currently a subject of concern, as although they are easy to store frozen for a long time (e.g., in a cord blood bank), the cell survival rate and success rate during extraction are not high, due to exposure to cryogenic protectors during cryogenic storage [73]. Furthermore, as the cells are isolated from other organs, they have limited self-renewal capacity, and their senescence is faster than in other stem cells in long-term cultivation [66,74].

HSCs can be differentiated into cells from all hematopoietic systems present in the bone marrow and chest glands, namely myeloid cells and lymphocytes. HSCs can be obtained at good levels from adult bone marrow, the placenta, and cord blood. They can cause immunological problems such as transplant rejection. Nevertheless, they have been shown to be an effective treatment method in various diseases, including leukemia, malignant lymphoma, and regenerative anemia, as well as congenital metabolism, congenital immunodeficiency, nonresponsive autoimmune disease, and solid cancer to date. Furthermore, HSCs are the only stem cell type approved for stem cell treatment by the Food and Drug Administration (FDA) [75,76].

ESCs have established cell lines that can be maintained through in vitro culture. They are pluripotent cells that can be differentiated into almost any type of cell present in the body, and can be differentiated in vitro by adding external factors to the culture medium or by genetic modification. However, they may form teratomas, which are composed of various forms of cells derived from the endoderm, mesoderm, and exoderm, when transplanted into an acceptable host [77].

iPSCs are artificially created stem cells. These cells are made by reprogramming adult somatic cells such as fibroblast cells. They share many of the characteristics of ESCs, including self-renewability, pluripotent differentiation, and malformed species performance. Unfortunately, these cells have little scientific evidence regarding changes in cell-specific regulatory pathways, gene expression, and epigenetic regulation. These characteristics pose a risk of tissue chimerism or cell dysfunction [78].

In summary, although the FDA-approved stem cell type is HSCs from healthy donors, a variety of issues have been raised, including a lack of donors and immune rejection. Therefore, we need to understand the characteristics of stem cells in order to handle them accordingly and overcome their disadvantages while maximizing their advantages. As stem cells derived from various sources have different characteristics, capabilities, potential, and efficiency, selecting the right source of stem cells that is appropriate for the target can be effective in assuring treatment efficiency.

The effective range of administration (i.e., dosage) of stem cells or stem-cell-derived products used in treatment should be determined through in vivo and in vitro studies. The safe and effective treatment capacity must be identified and, where possible, the minimum effective capacity must also be determined. When administered to vulnerable areas such as the central nervous system and myocardium, it has been reported that conducting normal dosage determination tests is unlikely. Thus, if the results of nonclinical studies can safety demonstrate treatment validity, it may be appropriate to conduct early human clinical trials with doses that may indicate therapeutic effects [79].

Will a high cell dose have better effects, considering only the effectiveness of stem cells? We answer this question below. An increasing dose of CD34+ cells (0.5 105 per mouse) has been shown to have positive effects, stimulating multilineage hematopoiesis at early stages and increasing the magnitude of reconstitution at post-transplant stages. Furthermore, improved T-cell reconstitution was correlated with higher cell doses of stem cells, compared to lower cell doses [80]. However, a few studies related to acute myeloblastic leukemia (AML) have reported that high doses of HSCs were correlated with restored function and rapid hematological and immunological recovery, but these results were not unconditional. In this study, a higher dose of HSCs (7 106/kg) resulted in poorer outcomes and a higher relapse rate than the lower dose of HSCs (<1 106/kg) [81]. In preclinical studies on heart disease, Golpanian et al. have demonstrated, through comparison of some preclinical studies for optimized cell dose, the therapeutic effects of stem cell types (i.e., allogenic and autologous MSCs), as well as the proper cell dose of stem cells and route of administration (direct epicardial and intravenous) in heart disease. Their results showed that the total number of cells used was different, but were inconsistent with the hypothesis that a higher number of cells would have higher therapeutic efficacy [82]. Therefore, these conclusions suggest that the currently reported data do not provide a decisive answer, such that sufficient and detailed early-stage studies may be needed before proceeding with clinical trials.

Stem cells have been extensively studied under various disease conditions, depending on their type and characteristics. At this time, the route of administration should not be overlooked in favor of the number of stem cells transplanted. Several reports have shown that engraftment ability typically has a lower rate of reaching target organs relative to the number of transplanted cells, and does not have a temporary longer duration [83,84].

The methods of stem cell administration can largely be divided into local and systemic transmission. Local transmission involves specific injections through various manipulations and direct intra-organ injections, such as intraperitoneal (IP), intramuscular, and intracardiac injections. Systemic transmission uses vascular pathways, such as intravenous (IV) and intra-arterial (IA) methods. According to the publications in the literature, IV is the most common method, followed by intrasplenic and IP [85,86,87]. In a liver disease model, IV was shown to be not only suitable for targeting the liver, but also showed better liver regeneration effects than other routes of administration [85,88]. Intracardial injection showed better cell retention in heart disease, while intradermal injection showed better treatment in skin diseases [89,90]. Hence, we can determine that, in the context of these various diseases, the routes of administration should be different depending on the target organ. Many researchers have suggested that intravascular injection is a minimally invasive procedure, but it also poses a risk of clogged blood vessels, such that direct intravascular injection increases the risk of requiring open-air operations [91]. Clinical trials have reported that the number of cells and treatment efficacy under the same conditions, as in preclinical studies, are not significant, but also differ in significance depending on the route of administration [92,93]. Therefore, researchers should continue to study which cells are appropriate for a given route of administrationeven within the same diseasebased on many precedents [82]. In addition, researchers should explore the appropriate routes of administration for safer and more effective therapeutic effects.

All medical treatments have benefits and risks. It is not particularly safe to apply these unproven stem cell treatments to patients. As expectations for regenerative treatment through stem cell therapies increase, the application of various administration pathways, including through the spinal cord, subcutaneous, and intramuscular, as well as the stem cell therapies themselves, have been diversifying, from autologous to homogenous to iPS. These stem cell treatments can vary in risk, depending on the cell type manufacturing process among other factors, and they differ in clinical experience, such that all types of stem cell treatments must be evaluate on the same basis. Furthermore, it should only be in limited and justified contexts that stem cells which can proliferate and have all-purpose differentiation remain in a final product.

Unfortunately, the only safe stem cells that have been employed in regenerative medicine so far are omnipotent stem cells, such as HSCs and MSCs, which are isolated from their self-origin [94]. Unfortunately, potential clinical applications using iPSCs and ESCs face many hurdles, as they present higher risk, including the possibility of rejection, teratoma formation, and genomic instability [95]. Hence, many researchers have attempted to overcome stem cell tracking for safety assessment. To check the engraftment and the remaining amount of stem cells, they have been labeled using BrdU, CM-Dil, and iron oxide nanoparticles, and visualized using Magnetic resonance imaging (MRI) [84,96,97].

A close analysis of the distribution patterns of administrative sites and target organs is required, as well as whether a distribution across the body is expected, and the organ that the cells are predicted to be distributed through should undergo a full-term analysis, including evaluation at administrative sites. To date, studies have reported assessments in the brain, lungs, heart, spleen, testicles, ovaries, kidneys, pancreas, bone marrow, blood, and lymph nodes, including areas of administration [98].

Some researchers have carried out the detection of transplanted UC-MSCs delivered by IV injection in the lung, heart, spleen, kidney, and liver. According to their results, the transplanted cells were not detected in other organs, except the lung and liver, for 7 days. In the lung and liver, the detected cells persisted at least 7 days after the transplant [99]. Furthermore, in a study comparing BM-MSCs and UC-MSCs in terms of cell tracking, they reported on the persistence of stem cells according to the route of administration used. In the results of the comparison of intracardiac and intravenous routes, the transplanted stem cells were detected in the lung for 10 days, but the signal disappeared after 21 days [100]. In other research, the stem cells were transplanted with using a biomaterial scaffold. The AD-MSCs were transplanted with hyaluronic acid/alginate hydrogel through intradermal injection, and could be detected by CM-Dil staining for 30 days [101]. These studies may show that the transplanted cells localized to the damaged organs through their homing ability, but the results of these previous studies seem to indicate that the residual volume and the residual date vary significantly depending on the target disease, organs, and type of stem cells. The cell residual means the survival of the cell, which represents the risk of formation of tumors. To overcome the problem of teratoma formation, the following results have been reported: According to one study, ESCs showed the following rates of teratoma formation: 100% under the kidney capsule, 60% intratesticular, 25100% subcutaneous, and 12.5% intramuscular. To overcome this problem, the investigators performed a co-injection with Matrigel into an animal model. According to their results, subcutaneous implantation of ESCs in the presence of Matrigel appeared to be the most efficient, reproducible, and easiest approach for preventing teratoma formation, other than only using ESCs [102]. Moreover, cellular products derived from iPSCs have higher potential as potential cell sources in personalized medicine [103]. Their applicability is currently limited due to concerns regarding the potential risk of serious transplant-related side effects, such as tumor formation due to residual pluripotent cells [104]. Hence, a recent study reported the establishment of an optimized tool for therapeutic intervention that allows for controlled specific and selective ablation of iPSCs through the use of LVCAGstransgenic iPSCs [104].

Unlike MSCs, which are generally considered immune-tolerant as an immunomodulator, transplantation of ESCs and HSCs requires close examination of the matching of histocompatibility antigen (HLA) between the donor and beneficiary [105,106]. Although homogeneous mesenchymal stem cells are known to have immunogenicity in immune-active rodent models and are quickly removed from the peripheral blood, studies have shown that a few MSCs remain for weeks to months. Therefore, it is recommended to conduct a study to assess the persistence of MSCs in the cell preparations administered, in order to assess the risk of stem cell removal. Therefore, for stem cell therapies that have undergone extensive in vitro manipulation such as long-term cell cultureincluding those derived from ESCs and iPSCsboth oncogenicity and genetic stability must be evaluated before clinical research begins. Furthermore, we must constantly review and study the latest research on safety, as well as the effects of regeneration using stem cells, and discuss and study the potential of regenerative medicine [107,108,109,110,111].

As discussed earlier, in vitro and in vivo preclinical studies are the direction of current research, and encompass the tasks that need to be completed. If we reinforce the current strengths and weaknesses based on the preceding content, we are already a step closer to developing stem cell treatments.

Before a treatment is applied in humans (i.e., patients), preclinical study must involve checking whether the effect of treatment will be positive or negative and, if there are any negative effects, the researcher must check the safety possibilities at every step. Due to concerns relating to treatment using stem-cell-based products, deciding whether preclinical studies are sufficient for translating to clinical trials raises several issues that must be assessed by competent authorities. An application for a clinical trial should be submitted to the Food and Drug Administration (FDA), the European Medicine Agency (EMA), or another organization, based on the country [112].

The FDA is responsible for certifying clinical trial studies for stem-cell-based products in the United States [113]. If a new drug is introduced to a clinical investigator which has not been approved by the FDA, an Investigational New Drug (IND) application may need to be submitted [114]. The IND application includes data from animal pharmacology and toxicology studies, clinical protocols, and investigator information [115]. A lack of preclinical support (e.g., in vitro and in vivo studies) can lead to required modification or disapproval. If the FDA has announced that an IND requires modifications (meaning that the application is intended to secure approval but has not yet been approved), the results of the preclinical studies were deemed insufficient or inadequate for translation to clinical trial study, such that further study must be completed, after which an amended IND should be submitted.

The FDA has published guidelines for the submission of an IND in the Code of Federal Regulations (CFR). These regulations are presented in 21 CFR part 210, 211 (Current Good Manufacturing Practice (cGMP)), 21 CFR part 312 (Investigational New Drug Application), 21 CFR 610 (General Biological Product Standards), and 21 CFR 1271 (Human Cells, Tissues, and Cellular and Tissue-Based Products) [116,117,118]. These guidelines have been issued for the development of stem cell products with the highest standards of safety and potential effective translation to clinical trial studies.

The FDA issued 21 CFR parts 210 and 211to ensure the quality of the final products [119]. The 21 CFR part 210 contains the minimum current good manufacturing practice (cGMP) considered at the stages of manufacturing, processing, packing, or holding of a drug, while the 21 CFR part 211 contains the cGMP for producing final products. The 21 CFR 211 includes FDA guidelines for personnel, buildings and facilities, equipment, and control of components, process, packaging, labeling, holding, and so on, all of which are critical for pharmaceutical production [116,117,118,119,120,121]. The requirements for IND submission and conducting clinical trial studies, reviewed by the FDA in the 21 CFR part 312 (Investigational New Drug Applications), includes exemptions that are described in detail in 312.2 (general provisions). Such exemptions do not require an IND to be submitted, but other studies must present an IND based on 21 CFR part 312. The section, 21 CFR part 312, provides different information, including the requirements for an IND, its content and format, protocols, general principles of IND submission, and so on. In addition, the FDA describes the administrative actions of IND submission, the responsibilities of sponsors and investigators, and so on, in this section [116,117,122]. The 21 CFR part 610 contains general biological product standards for final product characterization. The master cell bank (MCB) or working cell bank (WCB) used as a source for stem-cell-based final products must be tested before the release or use of the product in humans. The MCB and WCB should be tested for sterility, mycoplasma, purity, identity, and potency, among other tests based on the final products (e.g., viability, stability, phenotypes), before use at the clinical level. The FDA provides all required information regarding general biological product standards in this section, including release requirements, testing requirements, labeling standards, and so on [116,117,123,124]. The 21 CFR part 1271 focuses on introducing the regulations for human cells, tissues, and cellular and tissue-based products (HCT/Ps), in order to ensure adequate control for preventing the transmission of communicable disease from cell/tissue products. Current Good Tissue Practice (GTP) is a part of 21 CFR part 1271, where the purpose of GTP is to present regulations for the establishment and maintenance of quality control for prevention of introduction, transmission, or spread of communicable diseases, including regulations for personnel, procedures, facilities, environmental control, equipment, and so on [125,126,127,128].

The EMA is an agency in the European Union (EU) which is responsible for evaluating any investigational medical products (IMPs) in order to make sure that the final product is safe and efficient for public use. When planning to introduce a new drug for a clinical trial in Europe, one may be required to submit clinical trial applications to the EMA for IMPs. Clinical trial applications for IMPs include summaries of chemical, pharmacological, and biological preclinical data (e.g., from in vivo and in vitro studies) [129]. The EMA has presented different regulations to support the development of safe and efficient products for public usage, including Regulation (EC) No. 1394/2007, Directive 2004/23/EC, Directive 2006/17/EC, Directive 2006/86/EC, Directive 2001/83/EC, Directive 2001/20/EC, and Directive 2003/94/EC.

Regulation (EC) No. 1394/2007 defines the criteria for regulation regarding ATMPs. Advanced therapy products (ATMPs) are focused on gene therapy medicinal products (GTMP), somatic cell therapy medicinal products (sCTMP), tissue-engineered products (TEP), and combined ATMPs, which refers to a combination of two different medical technologies. Regulation (EC) No. 1394/2007 includes the requirements to be used in development, manufacturing, or administration of ATMPS [130,131,132]. Directive 2004/23/EC, Directive 2006/17/EC, and Directive 2006/86/EC define standards for safety and quality, as well as technical requirements for donation, procurement, testing, preservation, storage, and distribution of tissue and cells intended for human applications [133,134,135]. Directive 2001/83/EC applies to medicinal products for human use [136]. Directive 2001/20/EC presents the regulations for the implantation of products in clinical trials in the EU [137]; however, this directive will be replaced by regulation (EU) No. 536/2014. Regulation (EU) No. 536/2014 was adapted by the European Parliament in 2014, and provides regulation for clinical trials on medical products intended for human use. The new EU regulation comes into effect on 31 January 2022 and aims to coordinate all clinical trials performed throughout the EU, using clinical trials submitted into CTIS (Clinical Trials Information System). The definition of regulation (EU) No. 536/2014 as a homogeneous regulation serves an important role in the EU, as all member states of the EU can be involved in multicenter clinical trials using international coordination, thus allowing larger patient populations [138]. Directive 2003/94/EC provides Good Manufacturing Practice (GMP) Guidelines in relation to medicinal products or IMPs intended for human use [139]. All process and application requirements for the IMP application are present in the regulations and directives of the EMA. After presenting an IND/IMP to the regulatory authority responsible for clinical trial oversight (FDA or EMA), the application will be reviewed in accordance with the FDA/EMA criteria and, if assured of the protection of humans enrolled in the clinical study, the application will be approved by the investigational review boards (IRBs) in the United States or Ethics Committees (ECs) in the European Union. Clinical trial studies are composed of different steps where, at each step, products are assessed using different quality and quantity measurements by the responsible agency. An efficient clinical trial study should address the safety and efficiency of new stem cell products in each of the different steps, and it is important to complete each step based on defined instructions and regulations, as the results of previous steps are needed to move forward.

Almost all clinical trial studies that have been approved for testing in humans have been registered online (https://www.clinicaltrials.gov/ accessed 12 December 2021). Our search on this website revealed more than 6500 records for interventional studies registered using Stem Cells up to December 2021. The recorded clinical trials can be analyzed from different aspects.

Recruiting status: The recruiting status of these studies indicated that 18% of these studies were ongoing (recruitment) and 42% were completed (). Although completed, suspended, terminated, and withdrawn studies are all terms used for studies that have ended, each is used to describe a different status. Completed studies are those that have ended normally and the participants were completely enrolled in the study. Suspended, terminated, and withdrawn studies are studies that stopped early; however, the participant enrolment status differs between them. A suspended study may start again, but nobody can continue to participate in terminated or withdrawn studies [140,141].

Status of clinical trials using stem cells.

Type of disease: Stem-cell-based therapy is a new approach for the treatment of various diseases in different clinical trial studies. Blood and lymph diseases are the most common diseases that have benefited from this new approach (). Blood and lymph diseases refer to any type of disorder related to blood and lymph deficiency or abnormality, such as anemia, blood protein disorder, bone marrow disease, leukemia, hemophilia, thalassemia, thrombophilia, lymphatic disease, lymphoproliferative disease, thymoma, and so on. In addition, various clinical trial studies have been performed using stem cells to treat immune system disease; neoplasm, heart, and blood disease; and gland- and hormone-related disease (). However, this does not mean that all of these studies had great results, nor does it mean that all of these studies introduced a new treatment method; some of these clinical trial studies were only intended to increase treatment efficiency, compare different types of treatment methods, or analyze various parameters after the administration of stem cells into the body.

Diseases considered in clinical trials using stem cells.

Autologous vs. Allogenic: Stem-cell-based products for use in clinical trial studies can be divided into two categories: autologous and allogeneic stem cells. In autologous stem cell therapy, the stem cells are collected from the patients own body. Culture-expanded autologous stem cells are autologous stem cells that are expanded before transplantation, and can be divided into two groups: modified and unmodified expanded autologous stem cells. If autologous stem cells were transplanted to the donor immediately after collection, this is a nonexpanded autologous stem cell treatment. The use of these cells usually has fewer restrictions for receiving clinical trial authorization. The classification of allogenic stem cells is similar to that of autologous stem cells, except that allogeneic stem cells are collected from a healthy donor. The use of these cells requires more prerequisite tests, in order to check the donors health. Allogenic stem cells have been used more than autologous stem cells in the clinical trial studies (46.34% vs. 44.51%), as shown in .

Applied stem cell types in clinical trials using stem cells.

Phase: Clinical trial studies are conducted in different phases. In each phase, the purpose of study, the number of participants, and the follow-up duration may differ. A new phase of clinical trials should not be started unless the results of the completed phase(s) have been reviewed by competent authorities, in order to that certify the results of the completed phase(s) are valid for authorization of the start a new phase of the clinical trial. For this purpose, at the end of each phase of a clinical trial study, competent authorities evaluate whether the new drug is safe, efficient, and effective for the treatment of the target disease ().

Status of clinical phase within clinical trials using stem cells.

Early Phase I emphasizes the effects of the drug on the human body and how the drug is processed in the body.

Phase I of a clinical trial is carried out to ensure that a new treatment is safe and to determine how the new medicine works in humans. The FDA has estimated that about 70% of the studies pass this phase.

In Phase II, the accurate dose is determined and initial data on the efficiency and possible side effects are collected. The FDA has estimated that roughly 33% of the studies move to the next phase.

Phase III evaluates the safety and effectiveness of products. The result of this phase is submitted to the FDA/EMA for new product approval, which allows manufacturing and marketing of the drug. The FDA has estimated that 25%30% of the drugs pass at this phase.

Phase IV take place after the approval of new products and is carried out to determine the public safety of the new product [142,143,144].

The number of participants and the duration: A new stem cell product is eligible for marketing after completing successful clinical trial phases. As the new product has been used on volunteers and the effects/side effects of the drug have also been followed for a long time throughout the different phases, it is now possible to make a decision regarding its introduction to the market for public use. The number of participants and the duration of long-term follow-up in each study and each phase differ ( and ). The number of volunteers that participate in each phase of a clinical trial study varies, as each phase has a different target. The FDA has recommended 2080, 100300, and several hundred to thousands of volunteers for Phase I, Phase II, and Phase III, respectively [144,145]. Although the FDA has defined a range for enrolments per phase, the number of participants can vary depending on the type of disease. The number of participants for clinical studies in rare diseases will be lower than when studying common diseases. Searching for stem cells in clinicaltrial.gov, studies can be found with only one participant (e.g., {"type":"clinical-trial","attrs":{"text":"NCT02235844","term_id":"NCT02235844"}}NCT02235844, {"type":"clinical-trial","attrs":{"text":"NCT02383654","term_id":"NCT02383654"}}NCT02383654, {"type":"clinical-trial","attrs":{"text":"NCT03979898","term_id":"NCT03979898"}}NCT03979898, and {"type":"clinical-trial","attrs":{"text":"NCT01142856","term_id":"NCT01142856"}}NCT01142856). The sponsor/investigator must provide the FDA with strong documentation regarding the selection of such a number of volunteers. The volunteers for each clinical trial study, before attending, should be informed about the enrolment criteria of each study, possible side effects, and the advantages of the study.

Enrolment of clinical trials using stem cells.

The duration of each clinical trial study using stem cells.

Age of participants: Roughly 190,000 people participated in all the completed clinical trial studies using stem cells that had been registered. Each clinical study was performed in different age groups, which differed among the various studies depending on the type of drug, type of disease, and sponsor decision, as shown in .

The age of patients participating in clinical trials using stem cells.

Number of clinical trial studies: The number of clinical trial studies increased gradually from 2000 to 2014, although it fluctuated after 2014 but did not change significantly (). The reason for this increase in 2014 is not clear, but it may have been related to the introduction of the first advanced medicinal therapy product containing stem cells (Holoclar) by the EMA in 20142015 [146].

The proportion of clinical trials using stem cells by year: (A) the proportion of new clinical trial studies using stem cells by year (green bar) and the proportion of registration results accordingly (orange color line); (B) the proportion of completed registered clinical trial studies using stem cells by year (blue bar) and the updated results of completed clinical trial studies using stem cells by year (orange line).

Place of study: According to economic website reports, the cell therapy market has grown significantly in recent years, and it is expected to grow more in the coming years; therefore, many countries have begun research in this field. Our data from clinicaltrial.gov showed that the United States has conducted the most clinical trials using stem cells (). Government agencies, industry, individuals, universities, and private organizations have all invested in stem-cell-based therapy. The number of stem-cell-based companies has rapidly increased in recent years, and a brief overview of the submitted clinical trial studies indicated that the studies were mostly aimed at introducing therapeutic products for clinical applications. Therefore, we can expect the introduction of stem-cell-based products to the market.

The registered and completed clinical trial studies using stem cells according to participating countries: (A) top 10 participating countries with registered clinical trials using stem cells; and (B) top 10 countries based on the completion of registered clinical trials using stem cells.

As indicated above, translational research from the laboratory to clinical services has many layers which must be passed through, each with its own requirements and measurements. Therefore, the only way to introduce a new stem-cell-based product onto the market is for competent authorities to make sure that the discovery is safe and effective for its intended human use, and that the product has successfully passed all of the clinical trial stages.

One of the most important issues regarding the introduction of a new product for use in humans through a clinical trial is evaluation of its safety. Although many clinical trials have been performed using stem cells for the treatment of various diseases, as stem-cell-based therapies are one of the newest groups of therapeutic products in medicine, it is very hard to introduce new products based on stem cells onto the market, as many different parameters must be evaluated. There are several concerns regarding stem-cell-based therapies, including genetic instability after long-term expansion, stem cell migration to inappropriate regions of the body, immunological reaction, and so on. However, all challenges depend on the type of stem cell (e.g., embryonic stem cell, adult stem cell, iPS), type of disease, route of administration, and many other factors. Almost all researchers in the field of stem cell therapy believe that despite stem cells having great potential to treat disease through their intrinsic potential, unproven stem-cell-based therapies that have not been shown to be safe or effective may be accompanied by very serious health risks. In order to receive clinical trial approval from a competent regulatory authority, different tests must be performed for each study phase, and the results of one study should not be generalized to another study. The FDA and EMA have defined different regulations to ensure that stem-cell-based products are consistently controlled through the use of different preclinical studies (in vitro and in vivo). Based on these preclinical data, the FDA and EMA have the authority to approve a clinical trial study, as discussed in this review.

Another challenge that researchers and companies face is the duration of a clinical trial study before a stem-cell-based product can be introduced onto the market. At present, hematopoietic progenitor cells are the only FDA-approved product for use in patients with defects in blood production, while other stem-cell-based products used in clinical trials have not yet been introduced to the market.

In the past few years, several clinical trials have been conducted using stem cells, most of which have indicated the safety and high efficiency of stem-cell-based therapies. An attractive future option for regenerative medicine is the use of cell derivatives, including exosomes, amniotic fluid, Whartons jelly, and so on, for the treatment of diseases. Recently, the safety and efficiency of these products have been evaluated and optimized in preclinical studies. In addition, regenerative medicine using modified stem cells and combinations of stem cells with scaffolds and chemicals to overcome stem cell therapy challenges and increase the associated efficiency are two important future directions of research. However, establishing a safe method for stem cell modification and moving this technology toward clinical trial studies requires many preclinical studies.

The regenerative medicine market is developing and, due to encouraging findings in preclinical studies and predictable economic benefits, competition has increased between companies focused on the development of cell products. Therefore, government agencies, industries, individuals, universities, and private organizations have invested heavily into the development of the regenerative medicine market in recent years, such that we can be more hopeful about the future of stem-cell-based therapies.

In recent years, regenerative medicine has become a promising treatment option for various diseases. Due to their therapeutic potential, including the inhibition of inflammation or apoptosis, cell recruitment, stimulation of angiogenesis, and differentiation, stem cells can been seen as good candidates for regenerative medicine. In the last 50 years, more than 40,000 research papers have focused on stem-cell-based therapies. In this review study, we present a general overview of the translation of stem cell therapy from scientific ideas to clinical applications. Multiple mechanisms causing disease could be reversed by stem cells, due to their tremendous therapeutic potential. However, preclinical studies including in vitro and in vivo experiments are necessary to evaluate the potential of stem-cell-based treatments. Through preclinical research, it is possible to present scientific evidence and optimal treatment options for subsequent clinical studies. Before starting a clinical trial based on preclinical data, the application must be approved by a relevant regulatory administration, such as the FDA, EMA, or another organization. If the application is for the use of a new drug (including stem cells) which has never been tested before, the submission of an IND is required for FDA approval. Approximately 50% of clinical trials using stem cells take 2 to 5 years to complete. To minimize possible side effects, every new stem cell product should be approved for clinical marketing only after completing Phase IIV clinical trials successfully. Interestingly, the number of stem-cell-based companies aimed at introducing clinical applications has rapidly increased in recent years. Therefore, it may be possible to find stem-cell-based products on the clinical market in the near future. As described in this paper, there are several steps that should be carried out on the path from the laboratory to the clinical setting. To develop new stem-cell-based medicine for the clinical market, researchers should follow the guidelines suggested by the relevant authorities. Through these well-controlled development processes, researchers can achieve safe and effective stem-cell-based therapies, thus brings their research ideas into the clinical field.

All authors have read and agreed to the published version of the manuscript.

This review funded by National Institutes of Health grant: R01HD087417-01A1, R01HD094378-01, R01HD094380-01, R01HD100367-01, R01HD100563, R01HD100563.

The author has no conflicts of interest to declare.

Publishers Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations.

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Stem Cell Therapy: From Idea to Clinical Practice - PMC

Stem Cell Investig. 2020; 7: 8.

1Department of Basic Dental Science, National Research Centre, Cairo, Egypt;

2Stem Cell Laboratory, Center of Excellence for Advanced Sciences, National Research Centre, Cairo, Egypt

1Department of Basic Dental Science, National Research Centre, Cairo, Egypt;

2Stem Cell Laboratory, Center of Excellence for Advanced Sciences, National Research Centre, Cairo, Egypt

Received 2020 Jan 3; Accepted 2020 Apr 30.

Recent research reporting successful translation of stem cell therapies to patients have enriched the hope that such regenerative strategies may one day become a treatment for a wide range of vexing diseases. In fact, the past few years witnessed, a rather exponential advancement in clinical trials revolving around stem cell-based therapies. Some of these trials resulted in remarkable impact on various diseases. In this review, the advances and challenges for the development of stem-cell-based therapies are described, with focus on the use of stem cells in dentistry in addition to the advances reached in regenerative treatment modalities in several diseases. The limitations of these treatments and ongoing challenges in the field are also discussed while shedding light on the ethical and regulatory challenges in translating autologous stem cell-based interventions, into safe and effective therapies.

Keywords: Stem cells, therapies, clinical trials, translation

Cell-based therapy as a modality of regenerative medicine is considered one of the most promising disciplines in the fields of modern science & medicine. Such an advanced technology offers endless possibilities for transformative and potentially curative treatments for some of humanities most life threatening diseases. Regenerative medicine is rapidly becoming the next big thing in health care with the particular aim of repairing and possibly replacing diseased cells, tissues or organs and eventually retrieving normal function. Fortunately, the prospect of regenerative medicine as an alternative to conventional drug-based therapies is becoming a tangible reality by the day owing to the vigorous commitment of the research communities in studying the potential applications across a wide range of diseases like neurodegenerative diseases and diabetes, among many others (1).

Recent research reporting successful translation of stem cell therapies to patients have enriched the hope that such regenerative strategies may one day become a treatment for a wide range of vexing diseases (2). In fact, the past few years witnessed, a rather exponential advancement in clinical trials revolving around stem cell-based therapies. Some of these trials resulted in remarkable impact on various diseases (3). For example, a case of Epidermolysis Bullosa manifested signs of skin recovery after treatment with keratinocyte cultures of epidermal stem cells (4). Also, a major improvement in eyesight of patients suffering from macular degeneration was reported after transplantation of patient-derived induced pluripotent stem cells (iPSCs) that were induced to differentiate into pigment epithelial cells of the retina (5).

However, in spite of the increased amount of publications reporting successful cases of stem cell-based therapies, a major number of clinical trials have not yet acquired full regulatory approvals for validation as stem cell therapies. To date, the most established stem cell treatment is bone marrow transplants to treat blood and immune system disorders (1,6,7).

In this review, the advances and challenges for the development of stem-cell-based therapies are described, with focus on the use of stem cells in dentistry in addition to the advances reached in regenerative treatment modalities in several diseases. The limitations of these treatments and ongoing challenges in the field are also discussed while shedding light on the ethical and regulatory challenges in translating autologous stem cell-based interventions, into safe and effective therapies.

Stem cell-based therapies are defined as any treatment for a disease or a medical condition that fundamentally involves the use of any type of viable human stem cells including embryonic stem cells (ESCs), iPSCs and adult stem cells for autologous and allogeneic therapies (8). Stem cells offer the perfect solution when there is a need for tissue and organ transplantation through their ability to differentiate into the specific cell types that are required for repair of diseased tissues.

However, the complexity of stem cell-based therapies often leads researchers to search for stable, safe and easily accessible stem cells source that has the potential to differentiate into several lineages. Thus, it is of utmost importance to carefully select the type of stem cells that is suitable for clinical application (7,9).

There are mainly three types of stem cells. All three of them share the significant property of self-renewal in addition to a unique ability to differentiate. However, it should be noted that stem cells are not homogeneous, but rather exist in a developmental hierarchy (10). The most basic and undeveloped of stem cells are the totipotent stem cells. These cells are capable of developing into a complete embryo while forming the extra-embryonic tissue at the same time. This unique property is brief and starts with the fertilization of the ovum and ends when the embryo reaches the four to eight cells stage. Following that cells undergo subsequent divisions until reaching the blastocyst stage where they lose their totipotency property and assume a pluripotent identity where cells are only capable of differentiating into every embryonic germ layer (ectoderm, mesoderm and endoderm). Cells of this stage are termed embryonic stem cells and are obtained by isolation from the inner cell mass of the blastocyst in a process that involves the destruction of the forming embryo. After consecutive divisions, the property of pluripotency is lost and the differentiation capability becomes more lineage restricted where the cells become multipotent meaning that they can only differentiate into limited types of cells related to the tissue of origin. This is the property of adult stem cells, which helps create a state of homeostasis throughout the lifetime of the organism. Adult stem cells are present in a metabolically quiescent state in almost all specialized tissues of the body, which includes bone marrow and oral and dental tissues among many others (11).

Many authors consider adult stem cells the gold standard in stem cell-based therapies (12,13). Adult stem cells demonstrated signs of clinical success especially in hematopoietic transplants (14,15). In contrast to ESCs, adult stem cells are not subjected to controversial views regarding their origin. The fact that ESCs derivation involves destruction of human embryos renders them unacceptable for a significant proportion of the population for ethical and religious convictions (16-18).

It was in 2006 when Shinya Yamanka achieved a scientific breakthrough in stem cell research by succeeding in generating cells that have the same properties and genetic profile of ESCs. This was achieved via the transient over-expression of a cocktail of four transcription factors; OCT4, SOX2, KLF4 and MYC in, fully differentiated somatic cells, namely fibroblasts (19,20). These cells were called iPSCs and has transformed the field of stem cell research ever since (21). The most important feature of these cells is their ability to differentiate into any of the germ layers just like ESCs precluding the ethical debate surrounding their use. The development of iPSCs technology has created an innovative way to both identify and treat diseases. Since they can be generated from the patients own cells, iPSCs thus present a promising potential for the production of pluripotent derived patient-matched cells that could be used for autologous transplantation. True these cells symbolize a paradigm shift since they enable researchers to directly observe and treat relevant patient cells; nevertheless, a number of challenges still need to be addressed before iPSCs-derived cells can be applied in cell therapies. Such challenges include; the detection and removal of incompletely differentiated cells, addressing the genomic and epigenetic alterations in the generated cells and overcoming the tumorigenicity of these cells that could arise on transplantation (22).

With the rapid increase witnessed in stem cell basic research over the past years, the relatively new research discipline Translational Research has evolved significantly building up on the outcomes of basic research in order to develop new therapies. The clinical translation pathway starts after acquiring the suitable regulatory approvals. The importance of translational research lies in its a role as a filter to ensure that only safe and effective therapies reach the clinic (23). It bridges the gap from bench to bed. Currently, some stem cell-based therapies utilizing adult stem cells are clinically available and mainly include bone marrow transplants of hematopoietic stem cells and skin grafts for severe burns (23). To date, there are more than 3,000 trials involving the use of adult stem cells registered in WHO International Clinical Trials Registry. Additionally, initial trials involving the new and appealing iPSCs based therapies are also registered. In fact, the first clinical attempt employing iPSCs reported successful results in treating macular degeneration (24). Given the relative immaturity in the field of cellular therapy, the outcomes of such trials shall facilitate the understanding of the timeframes needed to achieve successful therapies and help in better understanding of the diseases. However, it is noteworthy that evaluation of stem cell-based therapies is not an easy task since transplantation of cells is ectopic and may result in tumor formation and other complications. This accounts for the variations in the results reported from previous reports. The following section discusses the published data of some of the most important clinical trials involving the use of different types of stem cells both in medicine and in dentistry.

The successful generation of neural cells from stem cells in vitro paved the way for the current stem cell-based clinical trials targeting neurodegenerative diseases (25,26). These therapies do not just target detaining the progression of irrecoverable neuro-degenerative diseases like Parkinsons, Alzheimers, amyotrophic lateral sclerosis (ALS), and multiple sclerosis (MS), but are also focused on completely treating such disorders.

PD is characterized by a rapid loss of midbrain dopaminergic neurons. The first attempt for using human ESC cells to treat PD was via the generation of dopaminergic-like neurons, later human iPSCs was proposed as an alternative to overcome ESCs controversies (27). Both cells presented hope for obtaining an endless source of dopaminergic neurons instead of the previously used fetal brain tissues. Subsequently, protocols that mimicked the development of dopaminergic neurons succeeded in generating dopaminergic neurons similar to that of the midbrain which were able to survive, integrate and functionally mature in animal models of PD preclinically (28). Based on the research presented by different groups; the Parkinsons Global Force was formed which aimed at guiding researchers to optimize their cell characterization and help promote the clinical progress toward successful therapy. Recently, In August 2018, Shinya Yamanka initiated the first approved clinical trial to treat PD using iPSCs. Seven patients suffering from moderate PD were recruited (29). Donor matched allogeneic cells were used to avoid any genetic influence of the disease. The strategy behind the trial involved the generation of dopaminergic progenitors followed by surgical transplantation into the brains of patients by a special device. In addition, immunosuppressant medications were given to avoid any adverse reaction. Preliminary results so far revealed the safety of the treatment.

MS is an inflammatory and neurodegenerative autoimmune disease of the central nervous system. Stem cell-based therapies are now exploring the possibility of halting the disease progression and reverse the neural damage. A registered phase 1 clinical trial was conducted by the company CelgeneTM in 2014 using placental-derived mesenchymal stem cells (MSCs) infusion to treat patients suffering from MS (30). This trial was performed at 6 centers in the United States and 2 centers in Canada and included 16 patients. Results demonstrated that cellular infusions were safe with no signs of paradoxical aggravation. However, clinical responses from patients indicated that the cellular treatment did not improve the MS condition (31). For the last decade immunoablative therapy demonstrated accumulative evidence of inducing long-term remission and improvement of disability caused by MS. This approach involves the replacement of the diseased immune system through administration of high-dose immunosuppressive therapy followed by hematopoietic stem cells infusion (32). However, immunoablation strategies demonstrated several complications such as infertility and neurological disabilities. A number of randomized controlled trials are planned to address these concerns (32). Currently, new and innovative stem cell-based therapies for MS are only in the initial stages, and are based on different mechanisms exploring the possibility of replacing damaged neuronal tissue with neural cells derived from iPSCs however, the therapeutic potential of iPSCs is still under research (33).

ALS is a neurodegenerative disease that causes degeneration of the motor neurons which results in disturbance in muscle performance. The first attempt to treat ALS was through the transplantation of MSCs in a mouse model. The outcomes of this experiment were promising and resulted in a decrease of the disease manifestations and thus providing proof of principal (34). Based on these results, several planned/ongoing clinical trials are on the way. These trials mainly assess the safety of the proposed concept and have not proved clinical success to date. Notably, while pre-clinical studies have reported that cells derived from un-diseased individuals are superior to cells from ALS patients; most of the clinical trials attempted have employed autologous transplantation. This information may account for the absence of therapeutic improvement reported (35).

Other neurologic indications for the use of stem cells are spinal cord injuries. Though the transplantation of different forms of neural stem cells and oligo-dendrocyte progenitors has led to growth in the axons in addition to neural connectivity which presents a possibility for repair (36), proof of recovered function has yet to be established in stringent clinical trials. Nevertheless, Japan has recently given approval to stem-cell treatment for spinal-cord injuries. This approval was based on clinical trials that are yet to be published and involves 13 patients, who are suffering from recent spinal-cord injury. The Japanese team discovered that injection of stem cells isolated from the patients bone marrow aided in regaining some lost sensation and mobility. This is the first stem cell-based therapy targeting spinal-cord injuries to gain governmental approval to offer to patients (37).

A huge number of the currently registered clinical trials for stem cell-based therapies target ocular diseases. This is mainly due to the fact that the eye is an immune privileged site. Most of these trials span various countries including Japan, China, Israel, Korea, UK, and USA and implement allogeneic ESC lines (35,36). Notably, the first clinical trial to implement the use autologous iPSCs-derived retinal cells was in Japan which followed the new regulatory laws issued in 2014 by Japans government to regulate regenerative medicine applications. Two patients were recruited in this trial, the first one received treatment for macular degeneration using iPSCs-generated retinal cell sheet (37). After 1 year of follow-up, there were no signs of serious complications including abnormal proliferation and systemic malignancy. Moreover, there were no signs of rejection of the transplanted retinal epithelial sheet in the second year follow-up. Most importantly, the signs of corrected visual acuity of the treated eye were reported. These results were enough to conclude that iPSCs-based autologous transplantation was safe and feasible (38). It is worthy to mention that the second patient was withdrawn from the study due to detectable genetic variations the patients iPSCs lines which was not originally present in the patients original fibroblasts. Such alterations may jeopardize the overall safety of the treatment. The fact that this decision was taken, even though the performed safety assays did not demonstrate tumorgenicity in the iPSCs-derived retinal pigment epithelium (RPE) cells, indicates that researchers in the field of iPSCs have full awareness of the importance of safety issues (39).

Pancreatic beta cells are destructed in type 1 diabetes mellitus, because of disorders in the immune system while in type 2 insulin insufficiency is caused by failure of the beta-cell to normally produce insulin. In both cases the affected cell is the beta cell, and since the pancreas does not efficiently regenerate islets from endogenous adult stem cells, other cell sources were tested (38). Pluripotent stem cells (PSCs) are considered the cells of choice for beta cell replacement strategies (39). Currently, there are a few industry-sponsored clinical trials that are registered targeting beta cell replacement using ESCs. These trials revolve around the engraftment of insulin-producing beta cells in an encapsulating device subcutaneously to protect the cells from autoimmunity in patients with type 1 diabetes (40). The company ViaCyteTM in California recently initiated a phase I/II trial ({"type":"clinical-trial","attrs":{"text":"NCT02239354","term_id":"NCT02239354"}}NCT02239354) in 2014 in collaboration with Harvard University. This trial involves 40 patients and employs two subcutaneous capsules of insulin producing beta cells generated from ESCs. The results shall be interesting due to the ease of monitoring and recovery of the transplanted cells. The preclinical studies preceding this trial demonstrated successful glycemic correction and the devices were successfully retrieved after 174 days and contained viable insulin-producing cells (41).

Stem cells have been successfully isolated from human teeth and were studied to test their ability to regenerate dental structures and periodontal tissues. MSCs were reported to be successfully isolated from dental tissues like dental pulp of permanent and deciduous teeth, periodontal ligament, apical papilla and dental follicle (42-44). These cells were described as an excellent cell source owing to their ease of accessibility, their ability to differentiate into osteoblasts and odontoblasts and lack of ethical controversies (45). Moreover, dental stem cells demonstrated superior abilities in immunomodulation properties either through cell to cell interaction or via a paracrine effect (46). Stem cells of non-dental origin were also suggested for dental tissue and bone regeneration. Different approaches were investigated for achieving dental and periodontal regeneration (47); however, assessments of stem cells after transplantation still require extensive studying. Clinical trials have only recently begun and their results are yet to be fully evaluated. However, by carefully applying the knowledge acquired from the extensive basic research in dental and periodontal regeneration, stem cell-based dental and periodontal regeneration may soon be a readily available treatment. To date, there are more than 6,000 clinical trials involving the use of with stem cells, however only a total of 44 registered clinical trials address oral diseases worldwide (48). Stem cell-based clinical trials with reported results targeting the treatment of oral disease are discussed below.

The first human clinical study using autologous dental pulp stem cells (DPSCs) for complete pulp regeneration was reported by Nakashima et al. in 2017 (49). This pilot study was based on extensive preclinical studies conducted by the same group (50). Patients with irreversible pulpitis were recruited and followed up for 6 months following DPSCs transplantation. Granulocyte colony-stimulating factor was administered to induce stem cell mobilization to enrich the stem cell populations. The research team reported that the use of DPSCs seeded on collagen scaffold in molars and premolars undergoing pulpectomy was safe. No adverse events or toxicity were demonstrated in the clinical and laboratory evaluations. Positive electric pulp testing was obtained after cell transplantation in all patients. Moreover, magnetic resonance imaging of the de-novo tissues formed in the root canal demonstrated similar results to normal pulp, which indicated successful pulp regeneration. A different group conducted a clinical trial that recruited patients diagnosed with necrotic pulp. Autologous stem cells from deciduous teeth were employed to induce pulp regeneration (51). Follow-up of the cases after a year from the intervention reported evidence of pulp regeneration with vascular supply and innervation. In addition, no signs of adverse effects were observed in patients receiving DPSCs transplantation. Both trials are proceeding with the next phases, however the results obtained are promising.

Aimetti et al. performed a study which included eleven patients suffering from chronic periodontitis and have one deep intra bony defect in addition to the presence of one vital tooth that needs extraction (52). Pulp tissue was passed through 50-m filters in presence of collagen sponge scaffold and was followed by transplantation in the bony defects caused by periodontal disease. Both clinical and radiographic evaluations confirmed the efficacy of this therapeutic intervention. Periodontal examination, attachment level, and probe depth showed improved results in addition to significant stability of the gingival margin. Moreover, radiographic analysis demonstrated bone regeneration.

The first clinical study using DPSCs for oro-maxillo-facial bone regeneration was conducted in 2009 (53). Patients in this study suffered from extreme bone loss following extraction of third molars. A bio-complex composed of DPSCs cultured on collagen sponge scaffolds was applied to the affected sites. Vertical repair of the damaged area with complete restoration of the periodontal tissue was demonstrated six months after the treatment. Three years later, the same group published a report evaluating the stability and quality of the regenerated bone after DPSCs transplantation (54). Histological and advanced holotomography demonstrated that newly formed bone was uniformly vascularized. However, it was of compact type, rather than a cancellous type which is usually the type of bone in this region.

Sjgrens syndrome (SS) is a systemic autoimmune disease marked by dry mouth and eyes. A novel therapeutic approach for SS. utilizing the infusion of MSCs in 24 patients was reported by Xu et al. in 2012 (55). The strategy behind this treatment was based on the immunologic regulatory functions of MSCs. Infused MSCs migrated toward the inflammatory sites in a stromal cell-derived factor-1-dependent manner. Results reported from this clinical trial demonstrated suppressed autoimmunity with subsequent restoration of salivary gland secretion in SS patients.

The ability to bank autologous stem cells at their most potent state for later use is an essential adjuvant to stem cell-based therapies. In order to be considered valid, any novel stem cell-based therapy should be as effective as the routine treatment. Thus, when appraising a type of stem cells for application in cellular therapies, issues like immune rejection must be avoided and at the same time large numbers of stem cells must be readily available before clinical implementation. iPSCs theoretically possess the ability to proliferate unlimitedly which pose them as an attractive source for use in cell-based therapies. Unlike, adult stem cells iPSCs ability to propagate does not decrease with time (22). Recently, California Institute for Regenerative Medicine (CIRM) has inaugurated an iPSCs repository to provide researchers with versatile iPSCs cell lines in order to accelerate stem cell treatments through studying genetic variation and disease modeling. Another important source for stem cells banking is the umbilical cord. Umbilical cord is immediately cryopreserved after birth; which permits stem cells to be successfully stored and ready for use in cell-based therapies for incurable diseases of a given individuals. However, stem cells of human exfoliated deciduous teeth (SHEDs) are more attractive as a source for stem cell banking. These cells have the capacity to differentiate into further cell types than the rest of the adult stem cells (56). Moreover, procedures involving the isolation and cryopreservation of these cells are un-complicated and not aggressive. The most important advantage of banking SHEDs is the insured autologous transplant which avoids the possibility of immune rejection (57). Contrary to cord blood stem cells, SHEDs have the ability to differentiate into connective tissues, neural and dental tissues (58) Finally, the ultimate goal of stem cell banking, is to establish a repository of high-quality stem cell lines derived from many individuals for future use in therapy.

With the increased number of clinical trials employing stem cells as therapeutic approaches, the need for developing regulatory guidelines and standards to ensure patients safety is becoming more and more essential. However, the fact that stem cell therapy is rather a new domain makes it subject to scientific, ethical and legal controversies that are yet to be regulated. Leading countries in the field have devised guidelines serving that purpose. Recently, the Food and Drug Administration (FDA) has released regulatory guidelines to ensure that these treatments are safe and effective (59). These guidelines state that; treatments involving stem cells that have been minimally manipulated and are intended for homogeneous use do not require premarket approval to come into action and shall only be subjected to regulatory guidelines against disease transmission. In 2014, a radical regulatory reform in Japan occurred with the passing of two new laws that permitted conditional approval of cell-based treatments following early phase clinical trials on the condition that clinical safety data are provided from at least ten patients. These laws allow skipping most of the traditional criteria of clinical trials in what was described as fast track approvals and treatments were classified according to risk (60). To date, the treatments that acquired conditional approval include those targeting; spinal-cord injury, cardiac disease and limb ischemia (61). Finally, regulatory authorities are now demanding application of standardization and safety regulations protocols for cellular products, which include the use of Xeno-free culture media, recombinant growth factors in addition to Good Manufacturing Practice (GMP) culture supplies.

Stem cell-based therapies face many obstacles that need to be urgently addressed. The most persistent concern is the ethical conflict regarding the use of ESCs. As previously mentioned, ESCs are far superior regarding their potency; however, their derivation requires destruction human embryos. True, the discovery of iPSCs overcame this concern; nevertheless, iPSCs themselves currently face another ethical controversy of their own which addresses their unlimited capacity of differentiation with concerns that these cells could one day be applied in human cloning. The use of iPSCs in therapy is still considered a high-risk treatment modality, since transplantation of these cells could induce tumor formation. Such challenge is currently addressed through developing optimized protocols to ensure their safety in addition to developing global clinical-grade iPSCs cell lines before these cells are available for clinical use (61). As for MSCs, these cells have been universally considered safe, however continuous monitoring and prolonged follow-up should be the focus of future research to avoid the possibility of tumor formation after treatments (62). Finally, it could be postulated that one of the most challenging ethical issues faced in the field of stem cell-based therapies at the moment, is the increasing number of clinics offering unproven stem cell-based treatments. Researchers are thus morally obligated to ensure that ethical considerations are not undermined in pursuit of progress in clinical translation.

Stem cell therapy is becoming a tangible reality by the day, thanks to the mounting research conducted over the past decade. With every research conducted the possibilities of stem cells applications increased in spite of the many challenges faced. Currently, progress in the field of stem cells is very promising with reports of clinical success in treating various diseases like; neurodegenerative diseases and macular degeneration progressing rapidly. iPSCs are conquering the field of stem cells research with endless possibilities of treating diseases using patients own cells. Regeneration of dental and periodontal tissues using MSCs has made its way to the clinic and soon enough will become a valid treatment. Although, challenges might seem daunting, stem cell research is advancing rapidly and cellular therapeutics is soon to be applicable. Fortunately, there are currently tremendous efforts exerted globally towards setting up regulatory guidelines and standards to ensure patients safety. In the near future, stem cell-based therapies shall significantly impact human health.

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Current state of stem cell-based therapies: an overview - PMC

Few of us know what they are or exactly how they work. But many of us have heard about the healing powers of stem cells, as well as the controversy surrounding them. Stem cells are well-debated and highly complex with promises ranging from fixing damaged knees to regenerating receding hairlines.

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But what are stem cells? And, whats all the fuss all about?

Director of the Center for Regenerative Medicine and Surgery, Amy Lightner, MD, shares the differences between stem cell types, how stem cells can be used and when to be cautious of claims that might be too good to be true.

When most of us think of stem cells, we probably recall images of Dolly the cloned sheep. While its true that Dolly was born of stem cells, her place in science history is just one of many advancements in the field.

In fact, there are many different types of stem cells, each of which has different responsibilities and abilities. What unifies them is their ability to regenerate into new cells.

Regenerative medicine is an emerging field that uses innovative treatments to help regenerate or heal cell function thats lost due to aging, disease or injury, Dr. Lightner explains. The way we achieve this is by using stem cells in large quantities, targeted to a certain area, that the body uses to promote healing.

Adult stem cells are the only type of stem cells that are currently approved for medical use in the United States by the U.S. Food and Drug Administration (FDA). The term adult stem cells is a little confusing because theyre actually found in infants, children and adults. These cells live in a variety of tissue in our bodies including bone marrow, muscles, your brain, your intestines and more.

Think of adult stem cells as a little army of cells that can regenerate themselves into new cells to maintain and repair the tissue or muscle where theyre found. The catch with adult stem cells is that they cant become different types of cells (for example, blood stem cells can only become new blood cells, not skin or brain cells).

Unlike adult stem cells, embryonic stem cells have many more possibilities. Harvested during an embryos blastocyst stage (about five or six days after an embryo has been fertilized in a lab), embryonic stem cells have the potential to become any type of cell (called pluripotent cells). For these reasons, embryonic stem cells are the type of stem cells that generate the controversy most people associate with the topic.

Stem cell therapy has been around since the 1970s, when the first adult bone marrow cells were used to treat blood disease. A bone marrow transplant allows a recipient whose bone marrow cells have been damaged by chemotherapy or disease to receive healthy bone marrow stem cells from a donor.

Those stem cells have the potential to mature within the blood system into different immune cells that recognize and fight off different types of blood cancer. And they also have the ability to heal, says Betty Hamilton, MD, Department of Hematology and Medical Oncology.

Bone marrow transplants are currently used to treat diseases including:

While you may have heard about the use of stem cell therapy for knees, back pain, arthritis, hair loss, diabetes and more, no other types of stem cell therapy beyond bone marrow transplants have yet been approved by the FDA. But thousands of clinical trials are available ranging from treatments for Crohns disease to multiple sclerosis and more. The common link between all these trials is the ability of the stem cells to reduce inflammation and repair damage to your body.

Dr. Hamilton and Dr. Lightner agree that were only just beginning to scratch the surface of stem cell therapy. In recent years, during the height of the COVID-19 pandemic, many clinical trials were underway to explore whether stem cells could be used to help treat the damaged lungs in people severely affected by the disease.

I think potential is the perfect word to describe stem cells, says Dr. Hamilton. We know they have these anti-inflammatory and regenerative properties where they can provide a significant improvement to someone dealing with a certain disease. There are so many diseases where inflammation happens, and something needs to be repaired, and so any help the immune system can get provides a lot of potential.

Scientists are also researching whether adult stem cells can turn into pluripotent stem cells, which would allow the cells to change into any cell type without involving the use of embryonic stem cells.

While the potential for stem cell therapy is great, doctors caution that were not quite there yet.

I always tell patients that ask about stem cell therapy clinics or traveling overseas for stem cell therapy treatment that if its not something that is a clinical trial with FDA oversight, then they have no real way of knowing whats being given to them, advises Dr. Lightner.

This means more harm can come than good if you dont know exactly whats being given to you. Or, in some cases, youre just spending thousands of dollars for what ends up being saline, Dr. Lightner says.

The best way to know that youre receiving sound medical treatment is to make sure the one youre considering is approved by the FDA on its Clinical Trials database.

Dr. Lightner cautions against treatments that sound too good to be true. While stem cell therapy has helped improve and save millions of lives, its best to know what exactly youre signing up for by seeking out a qualified medical provider offering an FDA-approved clinical trial.

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Stem Cell Therapy Is It Right for You? Cleveland Clinic