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| DOI | Article adapted for Ch. 3 |
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Stem Cell Therapy: From Idea to Clinical PracticeRegenerative 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. 1. IntroductionDespite 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 level’s requirements, but also discuss how an idea is assessed during the transition from one level to the next and, finally, move on to marketing. 2. From Idea to Preclinical StudyIf 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 ]. 3. From Preclinical Study to Clinical TrialIn 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. 3.1. Stem Cell Source DeterminationAs 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. 3.1.1. Mesenchymal Stem Cells (MSCs)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 ]. 3.1.2. Hematopoietic Stem Cells (HSCs)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 ]. 3.1.3. Embryonic Stem Cells (ESCs)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 ]. 3.1.4. Induced Pluripotent Stem Cells (iPSCs)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. 3.2. Cell Dose SpecificationThe 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 × 10 5 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 × 10 6 /kg) resulted in poorer outcomes and a higher relapse rate than the lower dose of HSCs (<1 × 10 6 /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. 3.3. Route of AdministrationStem 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 administration—even within the same disease—based on many precedents [ 82 ]. In addition, researchers should explore the appropriate routes of administration for safer and more effective therapeutic effects. 3.4. Manipulation of Cell Transplantation for Safety and Efficiency Improvement of AdministrationAll 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, 25–100% 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 LVCAGs–transgenic 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 culture—including those derived from ESCs and iPSCs—both 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. 4. From Clinical Trial to Clinical PracticeBefore 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/P’s), 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 ( Figure 1 ). 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 ( Figure 2 ). 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 ( Figure 2 ). 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 patient’s 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 donor’s 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 Figure 3 . 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 ( Figure 4 ). 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 ( Figure 5 and Figure 6 ). 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 20–80, 100–300, 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., NCT02235844, NCT02383654, NCT03979898, and 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 Figure 7 . 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 ( Figure 8 ). 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 2014–2015 [ 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 ( Figure 9 ). 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. 5. Challenges and Future DirectionsOne 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, Wharton’s 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. 6. ConclusionsIn 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 I–IV 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. Author ContributionsAll 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. Conflicts of InterestThe author has no conflicts of interest to declare. Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. Home > Academic Units > College of Interdisciplinary Studies > Honors > Honors Scholars Collaborative Projects > 53 Honors Scholars Collaborative ProjectsA literary analysis of the origin of human embryonic stem cells, its advancements, philosophical, ethical, sociocultural, and political aspects; an investigation of the underlying attributes that affect one’s views on hesc research to resolve turkey and brazil’s hesc policy, religious, and cultural conflicts. Haleema Shamsuddin Follow Publication DateSpring 4-2021 Human embryonic stem cells (hESCs) are cells derived from 5-day human embryos and are self-renewing cell lines that change into any type of cell in the body, a trait called pluripotency. hESCs have almost unlimited clinical and medical research potential. Despite the great therapeutic promise of hESC research, it comes with a controversial ethical debate due to its involvement with the destruction of the human embryo. The central argument revolves around the question of whether or not these human embryos should be ascribed equal moral status to fully developed humans. This thesis aims to analyze the origin and advancements of human embryonic stem cells, as well as philosophical, ethical, sociocultural, and political aspects. The analysis will include an investigation of the many underlying attributes of an individual that affect their views on hESC research and two specific nations that have hESC policies that conflict with their religious and cultural views. Faculty AdvisorNick Ragsdale Document TypeHonors Thesis Recommended CitationShamsuddin, Haleema, "A Literary Analysis of the Origin Of Human Embryonic Stem Cells, its Advancements, Philosophical, Ethical, Sociocultural, and Political Aspects; An Investigation of the Underlying Attributes that Affect One’s Views on hESC Research to Resolve Turkey and Brazil’s hESC Policy, Religious, and Cultural Conflicts" (2021). 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- Published: 26 February 2019
Stem cells: past, present, and future- Wojciech Zakrzewski 1 ,
- Maciej Dobrzyński 2 ,
- Maria Szymonowicz 1 &
- Zbigniew Rybak 1
Stem Cell Research & Therapy volume 10 , Article number: 68 ( 2019 ) Cite this article 584k Accesses 937 Citations 53 Altmetric Metrics details In recent years, stem cell therapy has become a very promising and advanced scientific research topic. The development of treatment methods has evoked great expectations. This paper is a review focused on the discovery of different stem cells and the potential therapies based on these cells. The genesis of stem cells is followed by laboratory steps of controlled stem cell culturing and derivation. Quality control and teratoma formation assays are important procedures in assessing the properties of the stem cells tested. Derivation methods and the utilization of culturing media are crucial to set proper environmental conditions for controlled differentiation. Among many types of stem tissue applications, the use of graphene scaffolds and the potential of extracellular vesicle-based therapies require attention due to their versatility. The review is summarized by challenges that stem cell therapy must overcome to be accepted worldwide. A wide variety of possibilities makes this cutting edge therapy a turning point in modern medicine, providing hope for untreatable diseases. Stem cell classificationStem cells are unspecialized cells of the human body. They are able to differentiate into any cell of an organism and have the ability of self-renewal. Stem cells exist both in embryos and adult cells. There are several steps of specialization. Developmental potency is reduced with each step, which means that a unipotent stem cell is not able to differentiate into as many types of cells as a pluripotent one. This chapter will focus on stem cell classification to make it easier for the reader to comprehend the following chapters. Totipotent stem cells are able to divide and differentiate into cells of the whole organism. Totipotency has the highest differentiation potential and allows cells to form both embryo and extra-embryonic structures. One example of a totipotent cell is a zygote, which is formed after a sperm fertilizes an egg. These cells can later develop either into any of the three germ layers or form a placenta. After approximately 4 days, the blastocyst’s inner cell mass becomes pluripotent. This structure is the source of pluripotent cells. Pluripotent stem cells (PSCs) form cells of all germ layers but not extraembryonic structures, such as the placenta. Embryonic stem cells (ESCs) are an example. ESCs are derived from the inner cell mass of preimplantation embryos. Another example is induced pluripotent stem cells (iPSCs) derived from the epiblast layer of implanted embryos. Their pluripotency is a continuum, starting from completely pluripotent cells such as ESCs and iPSCs and ending on representatives with less potency—multi-, oligo- or unipotent cells. One of the methods to assess their activity and spectrum is the teratoma formation assay. iPSCs are artificially generated from somatic cells, and they function similarly to PSCs. Their culturing and utilization are very promising for present and future regenerative medicine. Multipotent stem cells have a narrower spectrum of differentiation than PSCs, but they can specialize in discrete cells of specific cell lineages. One example is a haematopoietic stem cell, which can develop into several types of blood cells. After differentiation, a haematopoietic stem cell becomes an oligopotent cell. Its differentiation abilities are then restricted to cells of its lineage. However, some multipotent cells are capable of conversion into unrelated cell types, which suggests naming them pluripotent cells. Oligopotent stem cells can differentiate into several cell types. A myeloid stem cell is an example that can divide into white blood cells but not red blood cells. Unipotent stem cells are characterized by the narrowest differentiation capabilities and a special property of dividing repeatedly. Their latter feature makes them a promising candidate for therapeutic use in regenerative medicine. These cells are only able to form one cell type, e.g. dermatocytes. Stem cell biologyA blastocyst is formed after the fusion of sperm and ovum fertilization. Its inner wall is lined with short-lived stem cells, namely, embryonic stem cells. Blastocysts are composed of two distinct cell types: the inner cell mass (ICM), which develops into epiblasts and induces the development of a foetus, and the trophectoderm (TE). Blastocysts are responsible for the regulation of the ICM microenvironment. The TE continues to develop and forms the extraembryonic support structures needed for the successful origin of the embryo, such as the placenta. As the TE begins to form a specialized support structure, the ICM cells remain undifferentiated, fully pluripotent and proliferative [ 1 ]. The pluripotency of stem cells allows them to form any cell of the organism. Human embryonic stem cells (hESCs) are derived from the ICM. During the process of embryogenesis, cells form aggregations called germ layers: endoderm, mesoderm and ectoderm (Fig. 1 ), each eventually giving rise to differentiated cells and tissues of the foetus and, later on, the adult organism [ 2 ]. After hESCs differentiate into one of the germ layers, they become multipotent stem cells, whose potency is limited to only the cells of the germ layer. This process is short in human development. After that, pluripotent stem cells occur all over the organism as undifferentiated cells, and their key abilities are proliferation by the formation of the next generation of stem cells and differentiation into specialized cells under certain physiological conditions. Oocyte development and formation of stem cells: the blastocoel, which is formed from oocytes, consists of embryonic stem cells that later differentiate into mesodermal, ectodermal, or endodermal cells. Blastocoel develops into the gastrula Signals that influence the stem cell specialization process can be divided into external, such as physical contact between cells or chemical secretion by surrounding tissue, and internal, which are signals controlled by genes in DNA. Stem cells also act as internal repair systems of the body. The replenishment and formation of new cells are unlimited as long as an organism is alive. Stem cell activity depends on the organ in which they are in; for example, in bone marrow, their division is constant, although in organs such as the pancreas, division only occurs under special physiological conditions. Stem cell functional divisionWhole-body development. During division, the presence of different stem cells depends on organism development. Somatic stem cell ESCs can be distinguished. Although the derivation of ESCs without separation from the TE is possible, such a combination has growth limits. Because proliferating actions are limited, co-culture of these is usually avoided. ESCs are derived from the inner cell mass of the blastocyst, which is a stage of pre-implantation embryo ca. 4 days after fertilization. After that, these cells are placed in a culture dish filled with culture medium. Passage is an inefficient but popular process of sub-culturing cells to other dishes. These cells can be described as pluripotent because they are able to eventually differentiate into every cell type in the organism. Since the beginning of their studies, there have been ethical restrictions connected to the medical use of ESCs in therapies. Most embryonic stem cells are developed from eggs that have been fertilized in an in vitro clinic, not from eggs fertilized in vivo. Somatic or adult stem cells are undifferentiated and found among differentiated cells in the whole body after development. The function of these cells is to enable the healing, growth, and replacement of cells that are lost each day. These cells have a restricted range of differentiation options. Among many types, there are the following: Mesenchymal stem cells are present in many tissues. In bone marrow, these cells differentiate mainly into the bone, cartilage, and fat cells. As stem cells, they are an exception because they act pluripotently and can specialize in the cells of any germ layer. Neural cells give rise to nerve cells and their supporting cells—oligodendrocytes and astrocytes. Haematopoietic stem cells form all kinds of blood cells: red, white, and platelets. Skin stem cells form, for example, keratinocytes, which form a protective layer of skin. The proliferation time of somatic stem cells is longer than that of ESCs. It is possible to reprogram adult stem cells back to their pluripotent state. This can be performed by transferring the adult nucleus into the cytoplasm of an oocyte or by fusion with the pluripotent cell. The same technique was used during cloning of the famous Dolly sheep. hESCs are involved in whole-body development. They can differentiate into pluripotent, totipotent, multipotent, and unipotent cells (Fig. 2 ) [ 2 ]. Changes in the potency of stem cells in human body development. Potency ranges from pluripotent cells of the blastocyst to unipotent cells of a specific tissue in a human body such as the skin, CNS, or bone marrow. Reversed pluripotency can be achieved by the formation of induced pluripotent stem cells using either octamer-binding transcription factor (Oct4), sex-determining region Y (Sox2), Kruppel-like factor 4 (Klf4), or the Myc gene Pluripotent cells can be named totipotent if they can additionally form extraembryonic tissues of the embryo. Multipotent cells are restricted in differentiating to each cell type of given tissue. When tissue contains only one lineage of cells, stem cells that form them are called either called oligo- or unipotent. iPSC quality control and recognition by morphological differencesThe comparability of stem cell lines from different individuals is needed for iPSC lines to be used in therapeutics [ 3 ]. Among critical quality procedures, the following can be distinguished: Short tandem repeat analysis—This is the comparison of specific loci on the DNA of the samples. It is used in measuring an exact number of repeating units. One unit consists of 2 to 13 nucleotides repeating many times on the DNA strand. A polymerase chain reaction is used to check the lengths of short tandem repeats. The genotyping procedure of source tissue, cells, and iPSC seed and master cell banks is recommended. Identity analysis—The unintentional switching of lines, resulting in other stem cell line contamination, requires rigorous assay for cell line identification. Residual vector testing—An appearance of reprogramming vectors integrated into the host genome is hazardous, and testing their presence is a mandatory procedure. It is a commonly used procedure for generating high-quality iPSC lines. An acceptable threshold in high-quality research-grade iPSC line collections is ≤ 1 plasmid copies per 100 cells. During the procedure, 2 different regions, common to all plasmids, should be used as specific targets, such as EBNA and CAG sequences [ 3 ]. To accurately represent the test reactions, a standard curve needs to be prepared in a carrier of gDNA from a well-characterized hPSC line. For calculations of plasmid copies per cell, it is crucial to incorporate internal reference gDNA sequences to allow the quantification of, for example, ribonuclease P (RNaseP) or human telomerase reverse transcriptase (hTERT). Karyotype—A long-term culture of hESCs can accumulate culture-driven mutations [ 4 ]. Because of that, it is crucial to pay additional attention to genomic integrity. Karyotype tests can be performed by resuscitating representative aliquots and culturing them for 48–72 h before harvesting cells for karyotypic analysis. If abnormalities are found within the first 20 karyotypes, the analysis must be repeated on a fresh sample. When this situation is repeated, the line is evaluated as abnormal. Repeated abnormalities must be recorded. Although karyology is a crucial procedure in stem cell quality control, the single nucleotide polymorphism (SNP) array, discussed later, has approximately 50 times higher resolution. Viral testing—When assessing the quality of stem cells, all tests for harmful human adventitious agents must be performed (e.g. hepatitis C or human immunodeficiency virus). This procedure must be performed in the case of non-xeno-free culture agents. Bacteriology—Bacterial or fungal sterility tests can be divided into culture- or broth-based tests. All the procedures must be recommended by pharmacopoeia for the jurisdiction in which the work is performed. Single nucleotide polymorphism arrays—This procedure is a type of DNA microarray that detects population polymorphisms by enabling the detection of subchromosomal changes and the copy-neutral loss of heterozygosity, as well as an indication of cellular transformation. The SNP assay consists of three components. The first is labelling fragmented nucleic acid sequences with fluorescent dyes. The second is an array that contains immobilized allele-specific oligonucleotide (ASO) probes. The last component detects, records, and eventually interprets the signal. Flow cytometry—This is a technique that utilizes light to count and profile cells in a heterogeneous fluid mixture. It allows researchers to accurately and rapidly collect data from heterogeneous fluid mixtures with live cells. Cells are passed through a narrow channel one by one. During light illumination, sensors detect light emitted or refracted from the cells. The last step is data analysis, compilation and integration into a comprehensive picture of the sample. Phenotypic pluripotency assays—Recognizing undifferentiated cells is crucial in successful stem cell therapy. Among other characteristics, stem cells appear to have a distinct morphology with a high nucleus to cytoplasm ratio and a prominent nucleolus. Cells appear to be flat with defined borders, in contrast to differentiating colonies, which appear as loosely located cells with rough borders [ 5 ]. It is important that images of ideal and poor quality colonies for each cell line are kept in laboratories, so whenever there is doubt about the quality of culture, it can always be checked according to the representative image. Embryoid body formation or directed differentiation of monolayer cultures to produce cell types representative of all three embryonic germ layers must be performed. It is important to note that colonies cultured under different conditions may have different morphologies [ 6 ]. Histone modification and DNA methylation—Quality control can be achieved by using epigenetic analysis tools such as histone modification or DNA methylation. When stem cells differentiate, the methylation process silences pluripotency genes, which reduces differentiation potential, although other genes may undergo demethylation to become expressed [ 7 ]. It is important to emphasize that stem cell identity, together with its morphological characteristics, is also related to its epigenetic profile [ 8 , 9 ]. According to Brindley [ 10 ], there is a relationship between epigenetic changes, pluripotency, and cell expansion conditions, which emphasizes that unmethylated regions appear to be serum-dependent. hESC derivation and mediahESCs can be derived using a variety of methods, from classic culturing to laser-assisted methodologies or microsurgery [ 11 ]. hESC differentiation must be specified to avoid teratoma formation (see Fig. 3 ). Spontaneous differentiation of hESCs causes the formation of a heterogeneous cell population. There is a different result, however, when commitment signals (in forms of soluble factors and culture conditions) are applied and enable the selection of progenitor cells hESCs spontaneously differentiate into embryonic bodies (EBs) [ 12 ]. EBs can be studied instead of embryos or animals to predict their effects on early human development. There are many different methods for acquiring EBs, such as bioreactor culture [ 13 ], hanging drop culture [ 12 ], or microwell technology [ 14 , 15 ]. These methods allow specific precursors to form in vitro [ 16 ]. The essential part of these culturing procedures is a separation of inner cell mass to culture future hESCs (Fig. 4 ) [ 17 ]. Rosowski et al. [ 18 ] emphasizes that particular attention must be taken in controlling spontaneous differentiation. When the colony reaches the appropriate size, cells must be separated. The occurrence of pluripotent cells lasts for 1–2 days. Because the classical utilization of hESCs caused ethical concerns about gastrulas used during procedures, Chung et al. [ 19 ] found out that it is also possible to obtain hESCs from four cell embryos, leaving a higher probability of embryo survival. Additionally, Zhang et al. [ 20 ] used only in vitro fertilization growth-arrested cells. Culturing of pluripotent stem cells in vitro. Three days after fertilization, totipotent cells are formed. Blastocysts with ICM are formed on the sixth day after fertilization. Pluripotent stem cells from ICM can then be successfully transmitted on a dish Cell passaging is used to form smaller clusters of cells on a new culture surface [ 21 ]. There are four important passaging procedures. Enzymatic dissociation is a cutting action of enzymes on proteins and adhesion domains that bind the colony. It is a gentler method than the manual passage. It is crucial to not leave hESCs alone after passaging. Solitary cells are more sensitive and can easily undergo cell death; collagenase type IV is an example [ 22 , 23 ]. Manual passage , on the other hand, focuses on using cell scratchers. The selection of certain cells is not necessary. This should be done in the early stages of cell line derivation [ 24 ]. Trypsin utilization allows a healthy, automated hESC passage. Good Manufacturing Practice (GMP)-grade recombinant trypsin is widely available in this procedure [ 24 ]. However, there is a risk of decreasing the pluripotency and viability of stem cells [ 25 ]. Trypsin utilization can be halted with an inhibitor of the protein rho-associated protein kinase (ROCK) [ 26 ]. Ethylenediaminetetraacetic acid ( EDTA ) indirectly suppresses cell-to-cell connections by chelating divalent cations. Their suppression promotes cell dissociation [ 27 ]. Stem cells require a mixture of growth factors and nutrients to differentiate and develop. The medium should be changed each day. Traditional culture methods used for hESCs are mouse embryonic fibroblasts (MEFs) as a feeder layer and bovine serum [ 28 ] as a medium. Martin et al. [ 29 ] demonstrated that hESCs cultured in the presence of animal products express the non-human sialic acid, N -glycolylneuraminic acid (NeuGc). Feeder layers prevent uncontrolled proliferation with factors such as leukaemia inhibitory factor (LIF) [ 30 ]. First feeder layer-free culture can be supplemented with serum replacement, combined with laminin [ 31 ]. This causes stable karyotypes of stem cells and pluripotency lasting for over a year. Initial culturing media can be serum (e.g. foetal calf serum FCS), artificial replacement such as synthetic serum substitute (SSS), knockout serum replacement (KOSR), or StemPro [ 32 ]. The simplest culture medium contains only eight essential elements: DMEM/F12 medium, selenium, NaHCO 3, l -ascorbic acid, transferrin, insulin, TGFβ1, and FGF2 [ 33 ]. It is not yet fully known whether culture systems developed for hESCs can be allowed without adaptation in iPSC cultures. Turning point in stem cell therapyThe turning point in stem cell therapy appeared in 2006, when scientists Shinya Yamanaka, together with Kazutoshi Takahashi, discovered that it is possible to reprogram multipotent adult stem cells to the pluripotent state. This process avoided endangering the foetus’ life in the process. Retrovirus-mediated transduction of mouse fibroblasts with four transcription factors (Oct-3/4, Sox2, KLF4, and c-Myc) [ 34 ] that are mainly expressed in embryonic stem cells could induce the fibroblasts to become pluripotent (Fig. 5 ) [ 35 ]. This new form of stem cells was named iPSCs. One year later, the experiment also succeeded with human cells [ 36 ]. After this success, the method opened a new field in stem cell research with a generation of iPSC lines that can be customized and biocompatible with the patient. Recently, studies have focused on reducing carcinogenesis and improving the conduction system. Retroviral-mediated transduction induces pluripotency in isolated patient somatic cells. Target cells lose their role as somatic cells and, once again, become pluripotent and can differentiate into any cell type of human body The turning point was influenced by former discoveries that happened in 1962 and 1987. The former discovery was about scientist John Gurdon successfully cloning frogs by transferring a nucleus from a frog’s somatic cells into an oocyte. This caused a complete reversion of somatic cell development [ 37 ]. The results of his experiment became an immense discovery since it was previously believed that cell differentiation is a one-way street only, but his experiment suggested the opposite and demonstrated that it is even possible for a somatic cell to again acquire pluripotency [ 38 ]. The latter was a discovery made by Davis R.L. that focused on fibroblast DNA subtraction. Three genes were found that originally appeared in myoblasts. The enforced expression of only one of the genes, named myogenic differentiation 1 (Myod1), caused the conversion of fibroblasts into myoblasts, showing that reprogramming cells is possible, and it can even be used to transform cells from one lineage to another [ 39 ]. Although pluripotency can occur naturally only in embryonic stem cells, it is possible to induce terminally differentiated cells to become pluripotent again. The process of direct reprogramming converts differentiated somatic cells into iPSC lines that can form all cell types of an organism. Reprogramming focuses on the expression of oncogenes such as Myc and Klf4 (Kruppel-like factor 4). This process is enhanced by a downregulation of genes promoting genome stability, such as p53. Additionally, cell reprogramming involves histone alteration. All these processes can cause potential mutagenic risk and later lead to an increased number of mutations. Quinlan et al. [ 40 ] checked fully pluripotent mouse iPSCs using whole genome DNA sequencing and structural variation (SV) detection algorithms. Based on those studies, it was confirmed that although there were single mutations in the non-genetic region, there were non-retrotransposon insertions. This led to the conclusion that current reprogramming methods can produce fully pluripotent iPSCs without severe genomic alterations. During the course of development from pluripotent hESCs to differentiated somatic cells, crucial changes appear in the epigenetic structure of these cells. There is a restriction or permission of the transcription of genes relevant to each cell type. When somatic cells are being reprogrammed using transcription factors, all the epigenetic architecture has to be reconditioned to achieve iPSCs with pluripotency [ 41 ]. However, cells of each tissue undergo specific somatic genomic methylation. This influences transcription, which can further cause alterations in induced pluripotency [ 42 ]. Source of iPSCsBecause pluripotent cells can propagate indefinitely and differentiate into any kind of cell, they can be an unlimited source, either for replacing lost or diseased tissues. iPSCs bypass the need for embryos in stem cell therapy. Because they are made from the patient’s own cells, they are autologous and no longer generate any risk of immune rejection. At first, fibroblasts were used as a source of iPSCs. Because a biopsy was needed to achieve these types of cells, the technique underwent further research. Researchers investigated whether more accessible cells could be used in the method. Further, other cells were used in the process: peripheral blood cells, keratinocytes, and renal epithelial cells found in urine. An alternative strategy to stem cell transplantation can be stimulating a patient’s endogenous stem cells to divide or differentiate, occurring naturally when skin wounds are healing. In 2008, pancreatic exocrine cells were shown to be reprogrammed to functional, insulin-producing beta cells [ 43 ]. The best stem cell source appears to be the fibroblasts, which is more tempting in the case of logistics since its stimulation can be fast and better controlled [ 44 ]. The self-renewal and differentiation capabilities of iPSCs have gained significant interest and attention in regenerative medicine sciences. To study their abilities, a quality-control assay is needed, of which one of the most important is the teratoma formation assay. Teratomas are benign tumours. Teratomas are capable of rapid growth in vivo and are characteristic because of their ability to develop into tissues of all three germ layers simultaneously. Because of the high pluripotency of teratomas, this formation assay is considered an assessment of iPSC’s abilities [ 45 ]. Teratoma formation rate, for instance, was observed to be elevated in human iPSCs compared to that in hESCs [ 46 ]. This difference may be connected to different differentiation methods and cell origins. Most commonly, the teratoma assay involves an injection of examined iPSCs subcutaneously or under the testis or kidney capsule in mice, which are immune-deficient [ 47 ]. After injection, an immature but recognizable tissue can be observed, such as the kidney tubules, bone, cartilage, or neuroepithelium [ 30 ]. The injection site may have an impact on the efficiency of teratoma formation [ 48 ]. There are three groups of markers used in this assay to differentiate the cells of germ layers. For endodermal tissue, there is insulin/C-peptide and alpha-1 antitrypsin [ 49 ]. For the mesoderm, derivatives can be used, e.g. cartilage matrix protein for the bone and alcian blue for the cartilage. As ectodermal markers, class III B botulin or keratin can be used for keratinocytes. Teratoma formation assays are considered the gold standard for demonstrating the pluripotency of human iPSCs, demonstrating their possibilities under physiological conditions. Due to their actual tissue formation, they could be used for the characterization of many cell lineages [ 50 ]. Directed differentiationTo be useful in therapy, stem cells must be converted into desired cell types as necessary or else the whole regenerative medicine process will be pointless. Differentiation of ESCs is crucial because undifferentiated ESCs can cause teratoma formation in vivo. Understanding and using signalling pathways for differentiation is an important method in successful regenerative medicine. In directed differentiation, it is likely to mimic signals that are received by cells when they undergo successive stages of development [ 51 ]. The extracellular microenvironment plays a significant role in controlling cell behaviour. By manipulating the culture conditions, it is possible to restrict specific differentiation pathways and generate cultures that are enriched in certain precursors in vitro. However, achieving a similar effect in vivo is challenging. It is crucial to develop culture conditions that will allow the promotion of homogenous and enhanced differentiation of ESCs into functional and desired tissues. Regarding the self-renewal of embryonic stem cells, Hwang et al. [ 52 ] noted that the ideal culture method for hESC-based cell and tissue therapy would be a defined culture free of either the feeder layer or animal components. This is because cell and tissue therapy requires the maintenance of large quantities of undifferentiated hESCs, which does not make feeder cells suitable for such tasks. Most directed differentiation protocols are formed to mimic the development of an inner cell mass during gastrulation. During this process, pluripotent stem cells differentiate into ectodermal, mesodermal, or endodermal progenitors. Mall molecules or growth factors induce the conversion of stem cells into appropriate progenitor cells, which will later give rise to the desired cell type. There is a variety of signal intensities and molecular families that may affect the establishment of germ layers in vivo, such as fibroblast growth factors (FGFs) [ 53 ]; the Wnt family [ 54 ] or superfamily of transforming growth factors—β(TGFβ); and bone morphogenic proteins (BMP) [ 55 ]. Each candidate factor must be tested on various concentrations and additionally applied to various durations because the precise concentrations and times during which developing cells in embryos are influenced during differentiation are unknown. For instance, molecular antagonists of endogenous BMP and Wnt signalling can be used for ESC formation of ectoderm [ 56 ]. However, transient Wnt and lower concentrations of the TGFβ family trigger mesodermal differentiation [ 57 ]. Regarding endoderm formation, a higher activin A concentration may be required [ 58 , 59 ]. There are numerous protocols about the methods of forming progenitors of cells of each of germ layers, such as cardiomyocytes [ 60 ], hepatocytes [ 61 ], renal cells [ 62 ], lung cells [ 63 , 64 ], motor neurons [ 65 ], intestinal cells [ 66 ], or chondrocytes [ 67 ]. Directed differentiation of either iPSCs or ESCs into, e.g. hepatocytes, could influence and develop the study of the molecular mechanisms in human liver development. In addition, it could also provide the possibility to form exogenous hepatocytes for drug toxicity testing [ 68 ]. Levels of concentration and duration of action with a specific signalling molecule can cause a variety of factors. Unfortunately, for now, a high cost of recombinant factors is likely to limit their use on a larger scale in medicine. The more promising technique focuses on the use of small molecules. These can be used for either activating or deactivating specific signalling pathways. They enhance reprogramming efficiency by creating cells that are compatible with the desired type of tissue. It is a cheaper and non-immunogenic method. One of the successful examples of small-molecule cell therapies is antagonists and agonists of the Hedgehog pathway. They show to be very useful in motor neuron regeneration [ 69 ]. Endogenous small molecules with their function in embryonic development can also be used in in vitro methods to induce the differentiation of cells; for example, retinoic acid, which is responsible for patterning the nervous system in vivo [ 70 ], surprisingly induced retinal cell formation when the laboratory procedure involved hESCs [ 71 ]. The efficacy of differentiation factors depends on functional maturity, efficiency, and, finally, introducing produced cells to their in vivo equivalent. Topography, shear stress, and substrate rigidity are factors influencing the phenotype of future cells [ 72 ]. The control of biophysical and biochemical signals, the biophysical environment, and a proper guide of hESC differentiation are important factors in appropriately cultured stem cells. Stem cell utilization and their manufacturing standards and culture systemsThe European Medicines Agency and the Food and Drug Administration have set Good Manufacturing Practice (GMP) guidelines for safe and appropriate stem cell transplantation. In the past, protocols used for stem cell transplantation required animal-derived products [ 73 ]. The risk of introducing animal antigens or pathogens caused a restriction in their use. Due to such limitations, the technique required an obvious update [ 74 ]. Now, it is essential to use xeno-free equivalents when establishing cell lines that are derived from fresh embryos and cultured from human feeder cell lines [ 75 ]. In this method, it is crucial to replace any non-human materials with xeno-free equivalents [ 76 ]. NutriStem with LN-511, TeSR2 with human recombinant laminin (LN-511), and RegES with human foreskin fibroblasts (HFFs) are commonly used xeno-free culture systems [ 33 ]. There are many organizations and international initiatives, such as the National Stem Cell Bank, that provide stem cell lines for treatment or medical research [ 77 ]. Stem cell use in medicineStem cells have great potential to become one of the most important aspects of medicine. In addition to the fact that they play a large role in developing restorative medicine, their study reveals much information about the complex events that happen during human development. The difference between a stem cell and a differentiated cell is reflected in the cells’ DNA. In the former cell, DNA is arranged loosely with working genes. When signals enter the cell and the differentiation process begins, genes that are no longer needed are shut down, but genes required for the specialized function will remain active. This process can be reversed, and it is known that such pluripotency can be achieved by interaction in gene sequences. Takahashi and Yamanaka [ 78 ] and Loh et al. [ 79 ] discovered that octamer-binding transcription factor 3 and 4 (Oct3/4), sex determining region Y (SRY)-box 2 and Nanog genes function as core transcription factors in maintaining pluripotency. Among them, Oct3/4 and Sox2 are essential for the generation of iPSCs. Many serious medical conditions, such as birth defects or cancer, are caused by improper differentiation or cell division. Currently, several stem cell therapies are possible, among which are treatments for spinal cord injury, heart failure [ 80 ], retinal and macular degeneration [ 81 ], tendon ruptures, and diabetes type 1 [ 82 ]. Stem cell research can further help in better understanding stem cell physiology. This may result in finding new ways of treating currently incurable diseases. Haematopoietic stem cell transplantationHaematopoietic stem cells are important because they are by far the most thoroughly characterized tissue-specific stem cell; after all, they have been experimentally studied for more than 50 years. These stem cells appear to provide an accurate paradigm model system to study tissue-specific stem cells, and they have potential in regenerative medicine. Multipotent haematopoietic stem cell (HSC) transplantation is currently the most popular stem cell therapy. Target cells are usually derived from the bone marrow, peripheral blood, or umbilical cord blood [ 83 ]. The procedure can be autologous (when the patient’s own cells are used), allogenic (when the stem cell comes from a donor), or syngeneic (from an identical twin). HSCs are responsible for the generation of all functional haematopoietic lineages in blood, including erythrocytes, leukocytes, and platelets. HSC transplantation solves problems that are caused by inappropriate functioning of the haematopoietic system, which includes diseases such as leukaemia and anaemia. However, when conventional sources of HSC are taken into consideration, there are some important limitations. First, there is a limited number of transplantable cells, and an efficient way of gathering them has not yet been found. There is also a problem with finding a fitting antigen-matched donor for transplantation, and viral contamination or any immunoreactions also cause a reduction in efficiency in conventional HSC transplantations. Haematopoietic transplantation should be reserved for patients with life-threatening diseases because it has a multifactorial character and can be a dangerous procedure. iPSC use is crucial in this procedure. The use of a patient’s own unspecialized somatic cells as stem cells provides the greatest immunological compatibility and significantly increases the success of the procedure. Stem cells as a target for pharmacological testingStem cells can be used in new drug tests. Each experiment on living tissue can be performed safely on specific differentiated cells from pluripotent cells. If any undesirable effect appears, drug formulas can be changed until they reach a sufficient level of effectiveness. The drug can enter the pharmacological market without harming any live testers. However, to test the drugs properly, the conditions must be equal when comparing the effects of two drugs. To achieve this goal, researchers need to gain full control of the differentiation process to generate pure populations of differentiated cells. Stem cells as an alternative for arthroplastyOne of the biggest fears of professional sportsmen is getting an injury, which most often signifies the end of their professional career. This applies especially to tendon injuries, which, due to current treatment options focusing either on conservative or surgical treatment, often do not provide acceptable outcomes. Problems with the tendons start with their regeneration capabilities. Instead of functionally regenerating after an injury, tendons merely heal by forming scar tissues that lack the functionality of healthy tissues. Factors that may cause this failed healing response include hypervascularization, deposition of calcific materials, pain, or swelling [ 84 ]. Additionally, in addition to problems with tendons, there is a high probability of acquiring a pathological condition of joints called osteoarthritis (OA) [ 85 ]. OA is common due to the avascular nature of articular cartilage and its low regenerative capabilities [ 86 ]. Although arthroplasty is currently a common procedure in treating OA, it is not ideal for younger patients because they can outlive the implant and will require several surgical procedures in the future. These are situations where stem cell therapy can help by stopping the onset of OA [ 87 ]. However, these procedures are not well developed, and the long-term maintenance of hyaline cartilage requires further research. Osteonecrosis of the femoral hip (ONFH) is a refractory disease associated with the collapse of the femoral head and risk of hip arthroplasty in younger populations [ 88 ]. Although total hip arthroplasty (THA) is clinically successful, it is not ideal for young patients, mostly due to the limited lifetime of the prosthesis. An increasing number of clinical studies have evaluated the therapeutic effect of stem cells on ONFH. Most of the authors demonstrated positive outcomes, with reduced pain, improved function, or avoidance of THA [ 89 , 90 , 91 ]. Rejuvenation by cell programmingAgeing is a reversible epigenetic process. The first cell rejuvenation study was published in 2011 [ 92 ]. Cells from aged individuals have different transcriptional signatures, high levels of oxidative stress, dysfunctional mitochondria, and shorter telomeres than in young cells [ 93 ]. There is a hypothesis that when human or mouse adult somatic cells are reprogrammed to iPSCs, their epigenetic age is virtually reset to zero [ 94 ]. This was based on an epigenetic model, which explains that at the time of fertilization, all marks of parenteral ageing are erased from the zygote’s genome and its ageing clock is reset to zero [ 95 ]. In their study, Ocampo et al. [ 96 ] used Oct4, Sox2, Klf4, and C-myc genes (OSKM genes) and affected pancreas and skeletal muscle cells, which have poor regenerative capacity. Their procedure revealed that these genes can also be used for effective regenerative treatment [ 97 ]. The main challenge of their method was the need to employ an approach that does not use transgenic animals and does not require an indefinitely long application. The first clinical approach would be preventive, focused on stopping or slowing the ageing rate. Later, progressive rejuvenation of old individuals can be attempted. In the future, this method may raise some ethical issues, such as overpopulation, leading to lower availability of food and energy. For now, it is important to learn how to implement cell reprogramming technology in non-transgenic elder animals and humans to erase marks of ageing without removing the epigenetic marks of cell identity. Cell-based therapiesStem cells can be induced to become a specific cell type that is required to repair damaged or destroyed tissues (Fig. 6 ). Currently, when the need for transplantable tissues and organs outweighs the possible supply, stem cells appear to be a perfect solution for the problem. The most common conditions that benefit from such therapy are macular degenerations [ 98 ], strokes [ 99 ], osteoarthritis [ 89 , 90 ], neurodegenerative diseases, and diabetes [ 100 ]. Due to this technique, it can become possible to generate healthy heart muscle cells and later transplant them to patients with heart disease. Stem cell experiments on animals. These experiments are one of the many procedures that proved stem cells to be a crucial factor in future regenerative medicine In the case of type 1 diabetes, insulin-producing cells in the pancreas are destroyed due to an autoimmunological reaction. As an alternative to transplantation therapy, it can be possible to induce stem cells to differentiate into insulin-producing cells [ 101 ]. Stem cells and tissue banksiPS cells with their theoretically unlimited propagation and differentiation abilities are attractive for the present and future sciences. They can be stored in a tissue bank to be an essential source of human tissue used for medical examination. The problem with conventional differentiated tissue cells held in the laboratory is that their propagation features diminish after time. This does not occur in iPSCs. The umbilical cord is known to be rich in mesenchymal stem cells. Due to its cryopreservation immediately after birth, its stem cells can be successfully stored and used in therapies to prevent the future life-threatening diseases of a given patient. Stem cells of human exfoliated deciduous teeth (SHED) found in exfoliated deciduous teeth has the ability to develop into more types of body tissues than other stem cells [ 102 ] (Table 1 ). Techniques of their collection, isolation, and storage are simple and non-invasive. Among the advantages of banking, SHED cells are: Guaranteed donor-match autologous transplant that causes no immune reaction and rejection of cells [ 103 ] Simple and painless for both child and parent Less than one third of the cost of cord blood storage Not subject to the same ethical concerns as embryonic stem cells [ 104 ] In contrast to cord blood stem cells, SHED cells are able to regenerate into solid tissues such as connective, neural, dental, or bone tissue [ 105 , 106 ] SHED can be useful for close relatives of the donor Fertility diseasesIn 2011, two researchers, Katsuhiko Hayashi et al. [ 107 ], showed in an experiment on mice that it is possible to form sperm from iPSCs. They succeeded in delivering healthy and fertile pups in infertile mice. The experiment was also successful for female mice, where iPSCs formed fully functional eggs . Young adults at risk of losing their spermatogonial stem cells (SSC), mostly cancer patients, are the main target group that can benefit from testicular tissue cryopreservation and autotransplantation. Effective freezing methods for adult and pre-pubertal testicular tissue are available [ 108 ]. Qiuwan et al. [ 109 ] provided important evidence that human amniotic epithelial cell (hAEC) transplantation could effectively improve ovarian function by inhibiting cell apoptosis and reducing inflammation in injured ovarian tissue of mice, and it could be a promising strategy for the management of premature ovarian failure or insufficiency in female cancer survivors. For now, reaching successful infertility treatments in humans appears to be only a matter of time, but there are several challenges to overcome. First, the process needs to have high efficiency; second, the chances of forming tumours instead of eggs or sperm must be maximally reduced. The last barrier is how to mature human sperm and eggs in the lab without transplanting them to in vivo conditions, which could cause either a tumour risk or an invasive procedure. Therapy for incurable neurodegenerative diseasesThanks to stem cell therapy, it is possible not only to delay the progression of incurable neurodegenerative diseases such as Parkinson’s disease, Alzheimer’s disease (AD), and Huntington disease, but also, most importantly, to remove the source of the problem. In neuroscience, the discovery of neural stem cells (NSCs) has nullified the previous idea that adult CNS were not capable of neurogenesis [ 110 , 111 ]. Neural stem cells are capable of improving cognitive function in preclinical rodent models of AD [ 112 , 113 , 114 ]. Awe et al. [ 115 ] clinically derived relevant human iPSCs from skin punch biopsies to develop a neural stem cell-based approach for treating AD. Neuronal degeneration in Parkinson’s disease (PD) is focal, and dopaminergic neurons can be efficiently generated from hESCs. PD is an ideal disease for iPSC-based cell therapy [ 116 ]. However, this therapy is still in an experimental phase ( https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4539501 /). Brain tissue from aborted foetuses was used on patients with Parkinson’s disease [ 117 ]. Although the results were not uniform, they showed that therapies with pure stem cells are an important and achievable therapy. Stem cell use in dentistryTeeth represent a very challenging material for regenerative medicine. They are difficult to recreate because of their function in aspects such as articulation, mastication, or aesthetics due to their complicated structure. Currently, there is a chance for stem cells to become more widely used than synthetic materials. Teeth have a large advantage of being the most natural and non-invasive source of stem cells. For now, without the use of stem cells, the most common periodontological treatments are either growth factors, grafts, or surgery. For example, there are stem cells in periodontal ligament [ 118 , 119 ], which are capable of differentiating into osteoblasts or cementoblasts, and their functions were also assessed in neural cells [ 120 ]. Tissue engineering is a successful method for treating periodontal diseases. Stem cells of the root apical areas are able to recreate periodontal ligament. One of the possible methods of tissue engineering in periodontology is gene therapy performed using adenoviruses-containing growth factors [ 121 ]. As a result of animal studies, dentin regeneration is an effective process that results in the formation of dentin bridges [ 122 ]. Enamel is more difficult to regenerate than dentin. After the differentiation of ameloblastoma cells into the enamel, the former is destroyed, and reparation is impossible. Medical studies have succeeded in differentiating bone marrow stem cells into ameloblastoma [ 123 ]. Healthy dental tissue has a high amount of regular stem cells, although this number is reduced when tissue is either traumatized or inflamed [ 124 ]. There are several dental stem cell groups that can be isolated (Fig. 7 ). Localization of stem cells in dental tissues. Dental pulp stem cells (DPSCs) and human deciduous teeth stem cells (SHED) are located in the dental pulp. Periodontal ligaments stem cells are located in the periodontal ligament. Apical papilla consists of stem cells from the apical papilla (SCAP) Dental pulp stem cell (DPSC)These were the first dental stem cells isolated from the human dental pulp, which were [ 125 ] located inside dental pulp (Table 2 ). They have osteogenic and chondrogenic potential. Mesenchymal stem cells (MSCs) of the dental pulp, when isolated, appear highly clonogenic; they can be isolated from adult tissue (e.g. bone marrow, adipose tissue) and foetal (e.g. umbilical cord) [ 126 ] tissue, and they are able to differentiate densely [ 127 ]. MSCs differentiate into odontoblast-like cells and osteoblasts to form dentin and bone. Their best source locations are the third molars [ 125 ]. DPSCs are the most useful dental source of tissue engineering due to their easy surgical accessibility, cryopreservation possibility, increased production of dentin tissues compared to non-dental stem cells, and their anti-inflammatory abilities. These cells have the potential to be a source for maxillofacial and orthopaedic reconstructions or reconstructions even beyond the oral cavity. DPSCs are able to generate all structures of the developed tooth [ 128 ]. In particular, beneficial results in the use of DPSCs may be achieved when combined with other new therapies, such as periodontal tissue photobiomodulation (laser stimulation), which is an efficient technique in the stimulation of proliferation and differentiation into distinct cell types [ 129 ]. DPSCs can be induced to form neural cells to help treat neurological deficits. Stem cells of human exfoliated deciduous teeth (SHED) have a faster rate of proliferation than DPSCs and differentiate into an even greater number of cells, e.g. other mesenchymal and non-mesenchymal stem cell derivatives, such as neural cells [ 130 ]. These cells possess one major disadvantage: they form a non-complete dentin/pulp-like complex in vivo. SHED do not undergo the same ethical concerns as embryonic stem cells. Both DPSCs and SHED are able to form bone-like tissues in vivo [ 131 ] and can be used for periodontal, dentin, or pulp regeneration. DPSCs and SHED can be used in treating, for example, neural deficits [ 132 ]. DPSCs alone were tested and successfully applied for alveolar bone and mandible reconstruction [ 133 ]. Periodontal ligament stem cells (PDLSCs)These cells are used in periodontal ligament or cementum tissue regeneration. They can differentiate into mesenchymal cell lineages to produce collagen-forming cells, adipocytes, cementum tissue, Sharpey’s fibres, and osteoblast-like cells in vitro. PDLSCs exist both on the root and alveolar bone surfaces; however, on the latter, these cells have better differentiation abilities than on the former [ 134 ]. PDLSCs have become the first treatment for periodontal regeneration therapy because of their safety and efficiency [ 135 , 136 ]. Stem cells from apical papilla (SCAP)These cells are mesenchymal structures located within immature roots. They are isolated from human immature permanent apical papilla. SCAP are the source of odontoblasts and cause apexogenesis. These stem cells can be induced in vitro to form odontoblast-like cells, neuron-like cells, or adipocytes. SCAP have a higher capacity of proliferation than DPSCs, which makes them a better choice for tissue regeneration [ 137 , 138 ]. Dental follicle stem cells (DFCs)These cells are loose connective tissues surrounding the developing tooth germ. DFCs contain cells that can differentiate into cementoblasts, osteoblasts, and periodontal ligament cells [ 139 , 140 ]. Additionally, these cells proliferate after even more than 30 passages [ 141 ]. DFCs are most commonly extracted from the sac of a third molar. When DFCs are combined with a treated dentin matrix, they can form a root-like tissue with a pulp-dentin complex and eventually form tooth roots [ 141 ]. When DFC sheets are induced by Hertwig’s epithelial root sheath cells, they can produce periodontal tissue; thus, DFCs represent a very promising material for tooth regeneration [ 142 ]. Pulp regeneration in endodonticsDental pulp stem cells can differentiate into odontoblasts. There are few methods that enable the regeneration of the pulp. The first is an ex vivo method. Proper stem cells are grown on a scaffold before they are implanted into the root channel [ 143 ]. The second is an in vivo method. This method focuses on injecting stem cells into disinfected root channels after the opening of the in vivo apex. Additionally, the use of a scaffold is necessary to prevent the movement of cells towards other tissues. For now, only pulp-like structures have been created successfully. Methods of placing stem cells into the root channel constitute are either soft scaffolding [ 144 ] or the application of stem cells in apexogenesis or apexification. Immature teeth are the best source [ 145 ]. Nerve and blood vessel network regeneration are extremely vital to keep pulp tissue healthy. The potential of dental stem cells is mainly regarding the regeneration of damaged dentin and pulp or the repair of any perforations; in the future, it appears to be even possible to generate the whole tooth. Such an immense success would lead to the gradual replacement of implant treatments. Mandibulary and maxillary defects can be one of the most complicated dental problems for stem cells to address. Acquiring non-dental tissue cells by dental stem cell differentiationIn 2013, it was reported that it is possible to grow teeth from stem cells obtained extra-orally, e.g. from urine [ 146 ]. Pluripotent stem cells derived from human urine were induced and generated tooth-like structures. The physical properties of the structures were similar to natural ones except for hardness [ 127 ]. Nonetheless, it appears to be a very promising technique because it is non-invasive and relatively low-cost, and somatic cells can be used instead of embryonic cells. More importantly, stem cells derived from urine did not form any tumours, and the use of autologous cells reduces the chances of rejection [ 147 ]. Use of graphene in stem cell therapyOver recent years, graphene and its derivatives have been increasingly used as scaffold materials to mediate stem cell growth and differentiation [ 148 ]. Both graphene and graphene oxide (GO) represent high in-plane stiffness [ 149 ]. Because graphene has carbon and aromatic network, it works either covalently or non-covalently with biomolecules; in addition to its superior mechanical properties, graphene offers versatile chemistry. Graphene exhibits biocompatibility with cells and their proper adhesion. It also tested positively for enhancing the proliferation or differentiation of stem cells [ 148 ]. After positive experiments, graphene revealed great potential as a scaffold and guide for specific lineages of stem cell differentiation [ 150 ]. Graphene has been successfully used in the transplantation of hMSCs and their guided differentiation to specific cells. The acceleration skills of graphene differentiation and division were also investigated. It was discovered that graphene can serve as a platform with increased adhesion for both growth factors and differentiation chemicals. It was also discovered that π-π binding was responsible for increased adhesion and played a crucial role in inducing hMSC differentiation [ 150 ]. Therapeutic potential of extracellular vesicle-based therapiesExtracellular vesicles (EVs) can be released by virtually every cell of an organism, including stem cells [ 151 ], and are involved in intercellular communication through the delivery of their mRNAs, lipids, and proteins. As Oh et al. [ 152 ] prove, stem cells, together with their paracrine factors—exosomes—can become potential therapeutics in the treatment of, e.g. skin ageing. Exosomes are small membrane vesicles secreted by most cells (30–120 nm in diameter) [ 153 ]. When endosomes fuse with the plasma membrane, they become exosomes that have messenger RNAs (mRNAs) and microRNAs (miRNAs), some classes of non-coding RNAs (IncRNAs) and several proteins that originate from the host cell [ 154 ]. IncRNAs can bind to specific loci and create epigenetic regulators, which leads to the formation of epigenetic modifications in recipient cells. Because of this feature, exosomes are believed to be implicated in cell-to-cell communication and the progression of diseases such as cancer [ 155 ]. Recently, many studies have also shown the therapeutic use of exosomes derived from stem cells, e.g. skin damage and renal or lung injuries [ 156 ]. In skin ageing, the most important factor is exposure to UV light, called “photoageing” [ 157 ], which causes extrinsic skin damage, characterized by dryness, roughness, irregular pigmentation, lesions, and skin cancers. In intrinsic skin ageing, on the other hand, the loss of elasticity is a characteristic feature. The skin dermis consists of fibroblasts, which are responsible for the synthesis of crucial skin elements, such as procollagen or elastic fibres. These elements form either basic framework extracellular matrix constituents of the skin dermis or play a major role in tissue elasticity. Fibroblast efficiency and abundance decrease with ageing [ 158 ]. Stem cells can promote the proliferation of dermal fibroblasts by secreting cytokines such as platelet-derived growth factor (PDGF), transforming growth factor β (TGF-β), and basic fibroblast growth factor. Huh et al. [ 159 ] mentioned that a medium of human amniotic fluid-derived stem cells (hAFSC) positively affected skin regeneration after longwave UV-induced (UVA, 315–400 nm) photoageing by increasing the proliferation and migration of dermal fibroblasts. It was discovered that, in addition to the induction of fibroblast physiology, hAFSC transplantation also improved diseases in cases of renal pathology, various cancers, or stroke [ 160 , 161 ]. Oh [ 162 ] also presented another option for the treatment of skin wounds, either caused by physical damage or due to diabetic ulcers. Induced pluripotent stem cell-conditioned medium (iPSC-CM) without any animal-derived components induced dermal fibroblast proliferation and migration. Natural cutaneous wound healing is divided into three steps: haemostasis/inflammation, proliferation, and remodelling. During the crucial step of proliferation, fibroblasts migrate and increase in number, indicating that it is a critical step in skin repair, and factors such as iPSC-CM that impact it can improve the whole cutaneous wound healing process. Paracrine actions performed by iPSCs are also important for this therapeutic effect [ 163 ]. These actions result in the secretion of cytokines such as TGF-β, interleukin (IL)-6, IL-8, monocyte chemotactic protein-1 (MCP-1), vascular endothelial growth factor (VEGF), platelet-derived growth factor-AA (PDGF-AA), and basic fibroblast growth factor (bFGF). Bae et al. [ 164 ] mentioned that TGF-β induced the migration of keratinocytes. It was also demonstrated that iPSC factors can enhance skin wound healing in vivo and in vitro when Zhou et al. [ 165 ] enhanced wound healing, even after carbon dioxide laser resurfacing in an in vivo study. Peng et al. [ 166 ] investigated the effects of EVs derived from hESCs on in vitro cultured retinal glial, progenitor Müller cells, which are known to differentiate into retinal neurons. EVs appear heterogeneous in size and can be internalized by cultured Müller cells, and their proteins are involved in the induction and maintenance of stem cell pluripotency. These stem cell-derived vesicles were responsible for the neuronal trans-differentiation of cultured Müller cells exposed to them. However, the research article points out that the procedure was accomplished only on in vitro acquired retina. Challenges concerning stem cell therapyAlthough stem cells appear to be an ideal solution for medicine, there are still many obstacles that need to be overcome in the future. One of the first problems is ethical concern. The most common pluripotent stem cells are ESCs. Therapies concerning their use at the beginning were, and still are, the source of ethical conflicts. The reason behind it started when, in 1998, scientists discovered the possibility of removing ESCs from human embryos. Stem cell therapy appeared to be very effective in treating many, even previously incurable, diseases. The problem was that when scientists isolated ESCs in the lab, the embryo, which had potential for becoming a human, was destroyed (Fig. 8 ). Because of this, scientists, seeing a large potential in this treatment method, focused their efforts on making it possible to isolate stem cells without endangering their source—the embryo. Use of inner cell mass pluripotent stem cells and their stimulation to differentiate into desired cell types For now, while hESCs still remain an ethically debatable source of cells, they are potentially powerful tools to be used for therapeutic applications of tissue regeneration. Because of the complexity of stem cell control systems, there is still much to be learned through observations in vitro. For stem cells to become a popular and widely accessible procedure, tumour risk must be assessed. The second problem is to achieve successful immunological tolerance between stem cells and the patient’s body. For now, one of the best ideas is to use the patient’s own cells and devolve them into their pluripotent stage of development. New cells need to have the ability to fully replace lost or malfunctioning natural cells. Additionally, there is a concern about the possibility of obtaining stem cells without the risk of morbidity or pain for either the patient or the donor. Uncontrolled proliferation and differentiation of cells after implementation must also be assessed before its use in a wide variety of regenerative procedures on living patients [ 167 ]. One of the arguments that limit the use of iPSCs is their infamous role in tumourigenicity. There is a risk that the expression of oncogenes may increase when cells are being reprogrammed. In 2008, a technique was discovered that allowed scientists to remove oncogenes after a cell achieved pluripotency, although it is not efficient yet and takes a longer amount of time. The process of reprogramming may be enhanced by deletion of the tumour suppressor gene p53, but this gene also acts as a key regulator of cancer, which makes it impossible to remove in order to avoid more mutations in the reprogrammed cell. The low efficiency of the process is another problem, which is progressively becoming reduced with each year. At first, the rate of somatic cell reprogramming in Yamanaka’s study was up to 0.1%. The use of transcription factors creates a risk of genomic insertion and further mutation of the target cell genome. For now, the only ethically acceptable operation is an injection of hESCs into mouse embryos in the case of pluripotency evaluation [ 168 ]. Stem cell obstacles in the futurePioneering scientific and medical advances always have to be carefully policed in order to make sure they are both ethical and safe. Because stem cell therapy already has a large impact on many aspects of life, it should not be treated differently. Currently, there are several challenges concerning stem cells. First, the most important one is about fully understanding the mechanism by which stem cells function first in animal models. This step cannot be avoided. For the widespread, global acceptance of the procedure, fear of the unknown is the greatest challenge to overcome. The efficiency of stem cell-directed differentiation must be improved to make stem cells more reliable and trustworthy for a regular patient. The scale of the procedure is another challenge. Future stem cell therapies may be a significant obstacle. Transplanting new, fully functional organs made by stem cell therapy would require the creation of millions of working and biologically accurate cooperating cells. Bringing such complicated procedures into general, widespread regenerative medicine will require interdisciplinary and international collaboration. The identification and proper isolation of stem cells from a patient’s tissues is another challenge. Immunological rejection is a major barrier to successful stem cell transplantation. With certain types of stem cells and procedures, the immune system may recognize transplanted cells as foreign bodies, triggering an immune reaction resulting in transplant or cell rejection. One of the ideas that can make stem cells a “failsafe” is about implementing a self-destruct option if they become dangerous. Further development and versatility of stem cells may cause reduction of treatment costs for people suffering from currently incurable diseases. When facing certain organ failure, instead of undergoing extraordinarily expensive drug treatment, the patient would be able to utilize stem cell therapy. The effect of a successful operation would be immediate, and the patient would avoid chronic pharmacological treatment and its inevitable side effects. Although these challenges facing stem cell science can be overwhelming, the field is making great advances each day. Stem cell therapy is already available for treating several diseases and conditions. Their impact on future medicine appears to be significant. After several decades of experiments, stem cell therapy is becoming a magnificent game changer for medicine. With each experiment, the capabilities of stem cells are growing, although there are still many obstacles to overcome. Regardless, the influence of stem cells in regenerative medicine and transplantology is immense. Currently, untreatable neurodegenerative diseases have the possibility of becoming treatable with stem cell therapy. Induced pluripotency enables the use of a patient’s own cells. Tissue banks are becoming increasingly popular, as they gather cells that are the source of regenerative medicine in a struggle against present and future diseases. With stem cell therapy and all its regenerative benefits, we are better able to prolong human life than at any time in history. AbbreviationsBasic fibroblast growth factor Bone morphogenic proteins Dental follicle stem cells Dental pulp stem cells Embryonic bodies Embryonic stem cells Fibroblast growth factors Good Manufacturing Practice Graphene oxide Human amniotic fluid-derived stem cells Human embryonic stem cells Human foreskin fibroblasts Inner cell mass Non-coding RNA Induced pluripotent stem cells In vitro fertilization Knockout serum replacement Leukaemia inhibitory factor Monocyte chemotactic protein-1 Fibroblasts Messenger RNA Mesenchymal stem cells of dental pulp Myogenic differentiation Osteoarthritis Octamer-binding transcription factor 3 and 4 Platelet-derived growth factor Platelet-derived growth factor-AA Periodontal ligament stem cells Rho-associated protein kinase Stem cells from apical papilla Stem cells of human exfoliated deciduous teeth Synthetic Serum Substitute Trophectoderm Vascular endothelial growth factor Transforming growth factors Sukoyan MA, Vatolin SY, et al. Embryonic stem cells derived from morulae, inner cell mass, and blastocysts of mink: comparisons of their pluripotencies. Embryo Dev. 1993;36(2):148–58 Larijani B, Esfahani EN, Amini P, Nikbin B, Alimoghaddam K, Amiri S, Malekzadeh R, Yazdi NM, Ghodsi M, Dowlati Y, Sahraian MA, Ghavamzadeh A. Stem cell therapy in treatment of different diseases. Acta Medica Iranica. 2012:79–96 https://www.ncbi.nlm.nih.gov/pubmed/22359076 . Sullivan S, Stacey GN, Akazawa C, et al. Quality guidelines for clinical-grade human induced pluripotent stem cell lines. Regenerative Med. 2018; https://doi.org/10.2217/rme-2018-0095 . Amps K, Andrews PW, et al. Screening ethnically diverse human embryonic stem cells identifies a chromosome 20 minimal amplicon conferring growth advantage. Nat. Biotechnol. 2011; 29 (12):1121–44. Google Scholar Amit M, Itskovitz-Eldor J. Atlas of human pluripotent stem cells: derivation and culturing. New York: Humana Press; 2012. Ludwig TE, Bergendahl V, Levenstein ME, Yu J, Probasco MD, Thomson JA. Feeder-independent culture of human embryonic stem cells. Nat Methods. 2006;3:637–46. CAS PubMed Google Scholar Kang MI. Transitional CpG methylation between promoters and retroelements of tissue-specific genes during human mesenchymal cell differentiation. J. Cell Biochem. 2007;102:224–39. Vaes B, Craeye D, Pinxteren J. Quality control during manufacture of a stem cell therapeutic. BioProcess Int. 2012;10:50–5. Bloushtain-Qimron N. Epigenetic patterns of embryonic and adult stem cells. Cell Cycle. 2009;8:809–17. Brindley DA. Peak serum: implications of serum supply for cell therapy manufacturing. Regenerative Medicine. 2012;7:809–17. Solter D, Knowles BB. Immunosurgery of mouse blastocyst. Proc Natl Acad Sci U S A. 1975;72:5099–102. CAS PubMed PubMed Central Google Scholar Hoepfl G, Gassmann M, Desbaillets I. Differentiating embryonic stem cells into embryoid bodies. Methods Mole Biol. 2004;254:79–98 https://doi.org/10.1385/1-59259-741-6:079 . Lim WF, Inoue-Yokoo T, Tan KS, Lai MI, Sugiyama D. Hematopoietic cell differentiation from embryonic and induced pluripotent stem cells. Stem Cell Res Ther. 2013;4(3):71. https://doi.org/10.1186/scrt222 . Article CAS PubMed PubMed Central Google Scholar Mohr JC, de Pablo JJ, Palecek SP. 3-D microwell culture of human embryonic stem cells. Biomaterials. 2006;27(36):6032–42. https://doi.org/10.1016/j.biomaterials.2006.07.012 . Article CAS PubMed Google Scholar Doetschman TC, Eistetter H, Katz M, Schmidt W, Kemler R. The in vitro development of blastocyst-derived embryonic stem cell lines: formation of the visceral yolk sac, blood islands, and myocardium. J Embryol Exp Morphol. 1985;87:27–45. Kurosawa HY. Methods for inducing embryoid body formation: in vitro differentiation system of embryonic stem cells. J Biosci Bioeng. 2007;103:389–98. Heins N, Englund MC, Sjoblom C, Dahl U, Tonning A, Bergh C, Lindahl A, Hanson C, Semb H. Derivation, characterization, and differentiation of human embryonic stem cells. Stem Cells. 2004;22:367–76. Rosowski KA, Mertz AF, Norcross S, Dufresne ER, Horsley V. Edges of human embryonic stem cell colonies display distinct mechanical properties and differentiation potential. Sci Rep. 2015;5:Article number:14218. PubMed Google Scholar Chung Y, Klimanskaya I, Becker S, Li T, Maserati M, Lu SJ, Zdravkovic T, Ilic D, Genbacev O, Fisher S, Krtolica A, Lanza R. Human embryonic stem cell lines generated without embryo destruction. Cell Stem Cell. 2008;2:113–7. Zhang X, Stojkovic P, Przyborski S, Cooke M, Armstrong L, Lako M, Stojkovic M. Derivation of human embryonic stem cells from developing and arrested embryos. Stem Cells. 2006;24:2669–76. Beers J, Gulbranson DR, George N, Siniscalchi LI, Jones J, Thomson JA, Chen G. Passaging and colony expansion of human pluripotent stem cells by enzyme-free dissociation in chemically defined culture conditions. Nat Protoc. 2012;7:2029–40. Ellerström C, Hyllner J, Strehl R. single cell enzymatic dissociation of human embryonic stem cells: a straightforward, robust, and standardized culture method. In: Turksen K, editor. Human embryonic stem cell protocols. Methods in molecular biology: Humana Press; 2009. p. 584. Heng BC, Liu H, Ge Z, Cao T. Mechanical dissociation of human embryonic stem cell colonies by manual scraping after collagenase treatment is much more detrimental to cellular viability than is trypsinization with gentle pipetting. Biotechnol Appl Biochem. 2010;47(1):33–7. Ellerstrom C, Strehl R, Noaksson K, Hyllner J, Semb H. Facilitated expansion of human embryonic stem cells by single-cell enzymatic dissociation. Stem Cells. 2007;25:1690–6. Brimble SN, Zeng X, Weiler DA, Luo Y, Liu Y, Lyons IG, Freed WJ, Robins AJ, Rao MS, Schulz TC. Karyotypic stability, genotyping, differentiation, feeder-free maintenance, and gene expression sampling in three human embryonic stem cell lines deri. Stem Cells Dev. 2004;13:585–97. Watanabe K, Ueno M, Kamiya D, Nishiyama A, Matsumura M, Wataya T, Takahashi JB, Nishikawa S, Nishikawa S, Muguruma K, Sasai Y. A ROCK inhibitor permits survival of dissociated human embryonic stem cells. Nat Biotechnol. 2007;25:681–6. Nie Y, Walsh P, Clarke DL, Rowley JA, Fellner T. Scalable passaging of adherent human pluripotent stem cells. 2014. https://doi.org/10.1371/journal.pone.0088012 . Thomson JA, Itskovitz-Eldor J, Shapiro SS, Waknitz MA, Swiergiel JJ, Marshall VS, Jones JM. Embryonic stem cell lines derived from human blastocysts. Science. 1998;282:1145–7. Martin MJ, Muotri A, Gage F, Varki A. Human embryonic stem cellsexpress an immunogenic nonhuman sialic acid. Nat. Med. 2005;11:228–32. Smith AG, Heath JK, Donaldson DD, Wong GG, Moreau J, Stahl M, Rogers D. Inhibition of pluripotential embryonic stem cell differentiation by purified polypeptides. Nature. 1988;336(6200):688–90. https://doi.org/10.1038/336688a0 . Xu C, Inokuma MS, Denham J, Golds K, Kundu P, Gold JD, Carpenter MK. Feeder-free growth of undifferentiated human embryonic stem cells. Nature Biotechnol. 2001;19:971–4. https://doi.org/10.1038/nbt1001-971 . Article CAS Google Scholar Weathersbee PS, Pool TB, Ord T. Synthetic serum substitute (SSS): a globulin-enriched protein supplement for human embryo culture. J. Assist Reprod Genet. 1995;12:354–60. Chen G, Gulbranson DR, Hou Z, Bolin JM, Ruotti V, Probasco MD, Smuga-Otto K, Howden SE, Diol NR, Propson NE, Wagner R, Lee GO, Antosiewicz-Bourget J, Teng JM, Thomson JA. Chemically defined conditions for human iPSC derivation and culture. Nat. Methods. 2011;8:424–9. Sommer CA, Mostoslavsky G. Experimental approaches for the generation of induced pluripotent stem cells. Stem Cell Res Ther. 2010;1:26. PubMed PubMed Central Google Scholar Takahashi K, Yamanaka S. Induced pluripotent stem cells in medicine and biology. Development. 2013;140(12):2457–61 https://doi.org/10.1242/dev.092551 . Shi D, Lu F, Wei Y, et al. Buffalos ( Bubalus bubalis ) cloned by nuclear transfer of somatic cells. Biol. Reprod. 2007;77:285–91. https://doi.org/10.1095/biolreprod.107.060210 . Gurdon JB. The developmental capacity of nuclei taken from intestinal epithelium cells of feeding tadpoles. Development. 1962;10:622–40 http://dev.biologists.org/content/10/4/622 . CAS Google Scholar Kain K. The birth of cloning: an interview with John Gurdon. Dis Model Mech. 2009;2(1–2):9–10. https://doi.org/10.1242/dmm.002014 . Article PubMed Central Google Scholar Davis RL, Weintraub H, Lassar AB. Expression of a single transfected cDNA converts fibroblasts to myoblasts. Cell. 1987;24(51(6)):987–1000. Quinlan AR, Boland MJ, Leibowitz ML, et al. Genome sequencing of mouse induced pluripotent stem cells reveals retroelement stability and infrequent DNA rearrangement during reprogramming. Cell Stem Cell. 2011;9(4):366–73. Maherali N, Sridharan R, Xie W, Utika LJ, Eminli S, Arnold K, Stadtfeld M, Yachechko R, Tchieu J, Jaenisch R, Plath K, Hochedlinger K. Directly reprogrammed fibroblasts show global epigenetic remodeling and widespread tissue contribution. Cell Stem Cell. 2007;1:55–70. Ohi Y, Qin H, Hong C, Blouin L, Polo JM, Guo T, Qi Z, Downey SL, Manos PD, Rossi DJ, Yu J, Hebrok M, Hochedlinger K, Costello JF, Song JS, Ramalho-Santos M. Incomplete DNA methylation underlines a transcriptional memory of somatic cells in human IPS cells. Nat Cell Biol. 2011;13:541–9. Zhou Q, Brown J, Kanarek A, Rajagopal J, Melton DA. In vivo reprogramming of adult pancreatic exocrine cells to beta-cells. Nature. 2008;455:627–32 https://doi.org/10.1038/nature07314 . Hilfiker A, Kasper C, Hass R, Haverich A. Mesenchymal stem cells and progenitor cells in connective tissue engineering and regenerative medicine: is there a future for transplantation? Langenbecks Arch Surg. 2011;396:489–97. Zhang Wendy, Y., de Almeida Patricia, E., and Wu Joseph, C. Teratoma formation: a tool for monitoring pluripotency in stem cell research. StemBook, ed. The Stem Cell Research Community . June 12, 2012. https://doi.org/10.3824/stembook.1.53.1 . Narsinh KH, Sun N, Sanchez-Freire V, et al. Single cell transcriptional profiling reveals heterogeneity of human induced pluripotent stem cells. J Clin Invest. 2011;121(3):1217–21. Gertow K, Przyborski S, Loring JF, Auerbach JM, Epifano O, Otonkoski T, Damjanov I, AhrlundRichter L. Isolation of human embryonic stem cell-derived teratomas for the assessment of pluripotency. Curr Protoc Stem Cell Biol . 2007, Chapter 1, Unit 1B 4. 3: 1B.4.1-1B.4.29. Cooke MJ, Stojkovic M, Przyborski SA. Growth of teratomas derived from human pluripotent stem cells is influenced by the graft site. Stem Cells Dev. 2006;15(2):254–9. Przyborski SA. Differentiation of human embryonic stem cells after transplantation in immune-deficient mice. Stem Cells. 2005;23:1242–50. Tannenbaum SE, Turetsky TT, Singer O, Aizenman E, Kirshberg S, Ilouz N, Gil Y, Berman-Zaken Y, Perlman TS, Geva N, Levy O, Arbell D, Simon A, Ben-Meir A, Shufaro Y, Laufer N, Reubinoff BE. Derivation of xeno-free and GMP-grade human embryonic stem cells- platforms for future clinical applications. PLoS One. 2012;7:e35325. Cohen DE, Melton D. Turning straw into gold: directing cell fate for regenerative medicine. Nat Rev Genet. 2011;12:243–52. Hwang NS, Varghese S, Elisseeff J. Controlled differentiation of stem cells. Adv Drug Deliv Rev. 2007;60(2):199–214. https://doi.org/10.1016/j.addr.2007.08.036 . Turner N, Grose R. Fibroblast growth factor signalling: from development to cancer. Nat Rev Cancer. 2010;10:116–29. Rao TP, Kuhl M. An updated overview on Wnt signaling pathways: a prelude for more. Circ Res. 2010;106:1798–806. Moustakas A, Heldin CH. The regulation of TGFbeta signal transduction. Development. 2009;136:3699–714. Efthymiou AG, Chen G, Rao M, Chen G, Boehm M. Self-renewal and cell lineage differentiation strategies in human embryonic stem cells and induced pluripotent stem cells. Expert Opin Biol Ther. 2014;14:1333–44. Yang L, Soonpaa MH, Adler ED, Roepke TK, Kattman SJ, Kennedy M, Henckaerts E, Bonham K, Abbott GW, Linden RM, Field LJ, Keller GM. Human cardiovascular progenitor cells develop from a KDRþembryonic-stem-cell-derived population. Nature. 2008;453:524–8. Kroon E, Martinson LA, Kadoya K, Bang AG, Kelly OG, Eliazer S, Young H, Richardson M, Smart NG, Cunningham J, Agulnick AD, D’amour KA, Carpenter MK, Baetge EE. Pancreatic endoderm derived from human embryonic stem cells generates glucose-responsive insulin-secreting cells in vivo. Nat Biotechnol. 2008;26(4):443–52. https://doi.org/10.1038/nbt1393 . Vallier L, Reynolds D, Pedersen RA. Nodal inhibits differentiation of human embryonic stem cells along the neuroectodermal default pathway. Dev Biol. 2004;275:403–21. Burridge PW, Zambidis ET. Highly efficient directed differentiation of human induced pluripotent stem cells into cardiomyocytes. Methods Mol Biol. 2013;997:149–61. Cai J, Zhao Y, Liu Y, Ye F, Song Z, Qin H, Meng S, Chen Y, Zhou R, Song X, Guo Y, Ding M, Deng H. Directed differentiation of human embryonic stem cells into functional hepatic cells. Hepatology. 2007;45:1229–39. Takasato M, Er PX, Becroft M, Vanslambrouck JM, Stanley EG, Elefanty AG, Little MH. Directing human embryonic stem cell differentiation towards a renal lineage generates a selforganizing kidney. Nat Cell Biol. 2014;16:118–26. Huang SX, Islam MN, O’Neill J, Hu Z, Yang YG, Chen YW, Mumau M, Green MD, VunjakNovakovic G, Bhattacharya J, Snoeck HW. Efficient generation of lung and airway epithelial cells from human pluripotent stem cells. Nat Biotechnol. 2014;32:84–91. Kadzik RS, Morrisey EE. Directing lung endoderm differentiation in pluripotent stem cells. Cell Stem Cell. 2012;10:355–61. Wichterle H, Lieberam I, Porter JA, Jessell TM. Directed differentiation of embryonic stem cells into motor neurons. Cell. 2002;110:385–97. Spence JR, Mayhew CN, Rankin SA, Kuhar MF, Vallance JE, Tolle K, Hoskins EE, Kalinichenko VV, Wells SI, Zorn AM, Shroyer NF, Wells JM. Directed differentiation of human pluripotent stem cells into intestinal tissue in vitro. Nature. 2011;470:105–9. Oldershaw RA, Baxter MA, Lowe ET, Bates N, Grady LM, Soncin F, Brison DR, Hardingham TE, Kimber SJ. Directed differentiation of human embryonic stem cells toward chondrocytes. Nat Biotechnol. 2010;28:1187–94. Jun Cai, Ann DeLaForest, Joseph Fisher, Amanda Urick, Thomas Wagner, Kirk Twaroski, Max Cayo, Masato Nagaoka, Stephen A Duncan. Protocol for directed differentiation of human pluripotent stem cells toward a hepatocyte fate. 2012. DOI: https://doi.org/10.3824/stembook.1.52.1 . Frank-Kamenetsky M, Zhang XM, Bottega S, Guicherit O, Wichterle H, Dudek H, Bumcrot D, Wang FY, Jones S, Shulok J, Rubin LL, Porter JA. Small-molecule modulators of hedgehog signaling: identification and characterization of smoothened agonists and antagonists. J Biol. 2002;1:10. Oshima K, Shin K, Diensthuber M, Peng AW, Ricci AJ, Heller S. Mechanosensitive hair celllike cells from embryonic and induced pluripotent stem cells. Cell. 2010;141:704–16. Osakada F, Jin ZB, Hirami Y, Ikeda H, Danjyo T, Watanabe K, Sasai Y, Takahashi M. In vitro differentiation of retinal cells from human pluripotent stem cells by small-molecule induction. J Cell Sci. 2009;122:3169–79. Kshitiz PJ, Kim P, Helen W, Engler AJ, Levchenko A, Kim DH. Control of stem cell fate and function by engineering physical microenvironments. Intergr Biol (Camb). 2012;4:1008–18. Amps K, Andrews PW, Anyfantis G, Armstrong L, Avery S, Baharvand H, Baker J, Baker D, Munoz MB, Beil S, Benvenisty N, Ben-Yosef D, Biancotti JC, Bosman A, Brena RM, Brison D, Caisander G, Camarasa MV, Chen J, ChiaoE CYM, Choo AB, Collins D, et al. Screening ethnically diverse human embryonic stem cells identifies a chromosome 20 minimal amplicon conferring growth advantage. Nat Biotechnol. 2011;29:1132–44. Nukaya D, Minami K, Hoshikawa R, Yokoi N, Seino S. Preferential gene expression and epigenetic memory of induced pluripotent stem cells derived from mouse pancreas. Genes Cells. 2015;20:367–81. Murdoch A, Braude P, Courtney A, Brison D, Hunt C, Lawford-Davies J, Moore H, Stacey G, Sethe S, Procurement Working Group Of National Clinical H, E. S. C. F, National Clinical H, E. S. C. F. The procurement of cells for the derivation of human embryonic stem cell lines for therapeutic use: recommendations for good practice. Stem Cell Rev. 2012;8:91–9. Hewitson H, Wood V, Kadeva N, Cornwell G, Codognotto S, Stephenson E, Ilic D. Generation of KCL035 research grade human embryonic stem cell line carrying a mutation in HBB gene. Stem Cell Res. 2016;16:210–2. Daley GQ, Hyun I, Apperley JF, Barker RA, Benvenisty N, Bredenoord AL, Breuer CK, Caulfield T, Cedars MI, Frey-Vasconcells J, Heslop HE, Jin Y, Lee RT, Mccabe C, Munsie M, Murry CE, Piantadosi S, Rao M, Rooke HM, Sipp D, Studer L, Sugarman J, et al. Setting global standards for stem cell research and clinical translation: the 2016 ISSCR guidelines. Stem Cell Rep. 2016;6:787–97. Takahashi K, Yamanaka S. Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell. 2006;126(4):663–76. https://doi.org/10.1016/j.cell.2006.07.024 . Loh YH, Wu Q, Chew JL, Vega VB, Zhang W, Chen X, Bourque G, George J, Leong B, Liu J, et al. The Oct4 and Nanog transcription network regulates pluripotency in mouse embryonic stem cells. Nat Genet. 2006;38:431–40. Menasche P, Vanneaux V, Hagege A, Bel A, Cholley B, Cacciapuoti I, Parouchev A, Benhamouda N, Tachdjian G, Tosca L, Trouvin JH, Fabreguettes JR, Bellamy V, Guillemain R, SuberbielleBoissel C, Tartour E, Desnos M, Larghero J. Human embryonic stem cell-derived cardiac progenitors for severe heart failure treatment: first clinical case report. Eur Heart J. 2015;36:2011–7. Schwartz SD, Regillo CD, Lam BL, Eliott D, Rosenfeld PJ, Gregori NZ, Hubschman JP, Davis JL, Heilwell G, Spirn M, Maguire J, Gay R, Bateman J, Ostrick RM, Morris D, Vincent M, Anglade E, Del Priore LV, Lanza R. Human embryonic stem cell-derived retinal pigment epithelium in patients with age-related macular degeneration and Stargardt’s macular dystrophy: follow-up of two open-label phase 1/2 studies. Lancet. 2015;385:509–16. Ilic D, Ogilvie C. Concise review: human embryonic stem cells-what have we done? What are we doing? Where are we going? Stem Cells. 2017;35:17–25. Rocha V, et al. Clinical use of umbilical cord blood hematopoietic stem cells. Biol Blood Marrow Transplant. 2006;12(1):34–4. Longo UG, Ronga M, Maffulli N. Sports Med Arthrosc 17:112–126. Achilles tendinopathy. Sports Med Arthrosc. 2009;17:112–26. Tempfer H, Lehner C, Grütz M, Gehwolf R, Traweger A. Biological augmentation for tendon repair: lessons to be learned from development, disease, and tendon stem cell research. In: Gimble J, Marolt D, Oreffo R, Redl H, Wolbank S, editors. Cell engineering and regeneration. Reference Series in Biomedical Engineering. Cham: Springer; 2017. Goldring MB, Goldring SR. Osteoarthritis. J Cell Physiol. 2007;213:626–34. Widuchowski W, Widuchowski J, Trzaska T. Articular cartilage defects: study of 25,124 knee arthroscopies. Knee. 2007;14:177–82. Li R, Lin Q-X, Liang X-Z, Liu G-B, et al. Stem cell therapy for treating osteonecrosis of the femoral head: from clinical applications to related basic research. Stem Cell Res Therapy. 2018;9:291 https://doi.org/10.1186/s13287-018-1018-7 . Gangji V, De Maertelaer V, Hauzeur JP. Autologous bone marrow cell implantation in the treatment of non-traumatic osteonecrosis of the femoral head: five year follow-up of a prospective controlled study. Bone. 2011;49(5):1005–9. Zhao D, Cui D, Wang B, Tian F, Guo L, Yang L, et al. Treatment of early stage osteonecrosis of the femoral head with autologous implantation of bone marrow-derived and cultured mesenchymal stem cells. Bone. 2012;50(1):325–30. Sen RK, Tripathy SK, Aggarwal S, Marwaha N, Sharma RR, Khandelwal N. Early results of core decompression and autologous bone marrow mononuclear cells instillation in femoral head osteonecrosis: a randomized control study. J Arthroplast. 2012;27(5):679–86. Lapasset L, Milhavet O, Prieur A, Besnard E, Babled A, Aït-Hamou N, Leschik J, Pellestor F, Ramirez JM, De Vos J, Lehmann S, Lemaitre JM. Rejuvenating senescent and centenarian human cells by reprogramming through the pluripotent state. Genes Dev. 2011;25:2248–53. Sahin E, Depinho RA. Linking functional decline of telomeres, mitochondria and stem cells during ageing. Nature. 2010;464:520–8. Petkovich DA, Podolskiy DI, Lobanov AV, Lee SG, Miller RA, Gladyshev VN. Using DNA methylation profiling to evaluate biological age and longevity interventions. Cell Metab. 2017;25:954–60 https://doi.org/10.1016/j.cmet.2017.03.016 . Gerontology, Rejuvenation by cell reprogramming: a new horizon in. Rodolfo G. Goya, Marianne Lehmann, Priscila Chiavellini, Martina Canatelli-Mallat, Claudia B. Hereñú and Oscar A. Brown. Stem Cell Res Therapy . 2018, 9:349. https://doi.org/10.1186/s13287-018-1075-y . Ocampo A, Reddy P, Martinez-Redondo P, Platero-Luengo A, Hatanaka F, Hishida T, Li M, Lam D, Kurita M, Beyret E, Araoka T, Vazquez-Ferrer E, Donoso D, Roman JLXJ, Rodriguez-Esteban C, Nuñez G, Nuñez Delicado E, Campistol JM, Guillen I, Guillen P, Izpisua. In vivo amelioration of age-associated hallmarks by partial reprogramming. Cell. 2016;167:1719–33. de Lázaro I, Cossu G, Kostarelos K. Transient transcription factor (OSKM) expression is key towards clinical translation of in vivo cell reprogramming. EMBO Mol Med. 2017;9:733–6. Sun S, Li ZQ, Glencer P, Cai BC, Zhang XM, Yang J, Li XR. Bringing the age-related macular degeneration high-risk allele age-related maculopathy susceptibility 2 into focus with stem cell technology. Stem Cell Res Ther. 2017;8:135 https://doi.org/10.1186/s13287-017-0584-4 . Liu J. Induced pluripotent stem cell-derived neural stem cells: new hope for stroke? Stem Cell Res Ther. 2013;4:115 https://doi.org/10.1186/scrt326 . Shahjalal HM, Dayem AA, Lim KM, Jeon TI, Cho SG. Generation of pancreatic β cells for treatment of diabetes: advances and challenges. Stem Cell ResTher. 2018;9:355 https://doi.org/10.1186/s13287-018-1099-3 . Kroon E, Martinson LA, et al. Pancreatic endoderm derived from human embryonic stem cells generates glucose-responsive insulin-secreting cells in vivo. Nat Biotechnol. 2008;26;443–52. Arora V, Pooja A, Munshi AK. Banking stem cells from human exfoliated deciduous teeth. J Clin Pediatr Dent. 2009;33(4):289–94. Mao JJ. Stem cells and the future of dental care. New York State Dental J. 2008;74(2):21–4. Reznick, Jay B. Continuing education: stem cells: emerging medical and dental therapies for the dental Professional. Dentaltown Magazine . 2008, pp. 42–53. Arthur A, Rychkov G, Shi S, Koblar SA, Gronthos S. Adult human dental pulp stem cells differentiate toward functionally active neurons under appropriate environmental cues. Stem Cells. 2008;26(7):1787–95. Cordeiro MM, Dong Z, Kaneko T, Zhang Z, Miyazawa M, Shi S, Smith A. Dental pulp tissue engineering with stem cells from exfoliated. J Endod. 2008;34(8):962–9. Hayashi K, Ohta H, Kurimoto K, Aramaki S, Saitou M. Reconstitution of the mouse germ cell specification pathway in culture by pluripotent stem cells. Cell. 2011;146(4):519–32. https://doi.org/10.1016/j.cell.2011.06.052 . Sadri-Ardekani H, Atala A. Testicular tissue cryopreservation and spermatogonial stem cell transplantation to restore fertility: from bench to bedside. Stem Cell ResTher. 2014;5:68 https://doi.org/10.1186/scrt457 . Zhang Q, Xu M, Yao X, Li T, Wang Q, Lai D. Human amniotic epithelial cells inhibit granulosa cell apoptosis induced by chemotherapy and restore the fertility. Stem Cell Res Ther. 2015;6:152 https://doi.org/10.1186/s13287-015-0148-4 . Ma DK, Bonaguidi MA, Ming GL, Song H. Adult neural stem cells in the mammalian central nervous system. Cell Res. 2009;19:672–82. https://doi.org/10.1038/cr.2009.56 . Dantuma E, Merchant S, Sugaya K. Stem cells for the treatment of neurodegenerative diseases. Stem Cell ResTher. 2010;1:37 https://doi.org/10.1186/scrt37 . Wang Q, Matsumoto Y, Shindo T, Miyake K, Shindo A, Kawanishi M, Kawai N, Tamiya T, Nagao S. Neural stem cells transplantation in cortex in a mouse model of Alzheimer’s disease. J Med Invest. 2006;53:61–9. https://doi.org/10.2152/jmi.53.61 . Article PubMed Google Scholar Moghadam FH, Alaie H, Karbalaie K, Tanhaei S, Nasr Esfahani MH, Baharvand H. Transplantation of primed or unprimed mouse embryonic stem cell-derived neural precursor cells improves cognitive function in Alzheimerian rats. Differentiation. 2009;78:59–68. https://doi.org/10.1016/j.diff.2009.06.005 . Byrne JA. Developing neural stem cell-based treatments for neurodegenerative diseases. Stem Cell ResTher. 2014;5:72. https://doi.org/10.1186/scrt461 . Awe JP, Lee PC, Ramathal C, Vega-Crespo A, Durruthy-Durruthy J, Cooper A, Karumbayaram S, Lowry WE, Clark AT, Zack JA, Sebastiano V, Kohn DB, Pyle AD, Martin MG, Lipshutz GS, Phelps PE, Pera RA, Byrne JA. Generation and characterization of transgene-free human induced pluripotent stem cells and conversion to putative clinical-grade status. Stem Cell Res Ther. 2013;4:87. https://doi.org/10.1186/scrt246 . Peng J, Zeng X. The role of induced pluripotent stem cells in regenerative medicine: neurodegenerative diseases. Stem Cell ResTher. 2011;2:32. https://doi.org/10.1186/scrt73 . Wright BL, Barker RA. Established and emerging therapies for Huntington’s disease. 2007;7(6):579–87 https://www.ncbi.nlm.nih.gov/pubmed/17896994/579-87 . Lin NH, Gronthos S, Bartold PM. Stem cells and periodontal regeneration. Aust Dent J. 2008;53:108–21. Seo BM, Miura M, Gronthos S, Bartold PM, Batouli S, Brahim J, et al. Investigation of multipotent postnatal stem cells from human periodontal ligament. Lancet. 2004;364:149–55. Ramseier CA, Abramson ZR, Jin Q, Giannobile WV. Gene therapeutics for periodontal regenerative medicine. Dent Clin North Am. 2006;50:245–63. Shi S, Bartold PM, Miura M, Seo BM, Robey PG, Gronthos S. The efficacy of mesenchymal stem cells to regenerate and repair dental structures. OrthodCraniofac Res. 2005;8:191–9. Iohara K, Nakashima M, Ito M, Ishikawa M, Nakasima A, Akamine A. Dentin regeneration by dental pulp stem cell therapy with recombinant human bone morphogenetic protein. J Dent Res. 2004;83:590–5. Hu B, Unda F, Bopp-Kuchler S, Jimenez L, Wang XJ, Haikel Y, et al. Bone marrow cells can give rise to ameloblast-like cells. J Dent Res. 2006;85:416–21. Liu Y, Liu W, Hu C, Xue Z, Wang G, Ding B, Luo H, Tang L, Kong X, Chen X, Liu N, Ding Y, Jin Y. MiR-17 modulates osteogenic differentiation through a coherent feed-forward loop in mesenchymal stem cells isolated from periodontal ligaments of patients with periodontitis. Stem Cells. 2011;29(11):1804–16. https://doi.org/10.1002/stem.728 . Raspini G, Wolff J, Helminen M, Raspini G, Raspini M, Sándor GK. Dental stem cells harvested from third molars combined with bioactive glass can induce signs of bone formation in vitro. J Oral Maxillofac Res. 2018;9(1):e2. Published 2018 Mar 31. https://doi.org/10.5037/jomr.2018.9102 . Christodoulou I, Goulielmaki M, Devetzi M, Panagiotidis M, Koliakos G, Zoumpourlis V. Mesenchymal stem cells in preclinical cancer cytotherapy: a systematic review. Stem Cell Res Ther. 2018;9(1;336). https://doi.org/10.1186/s13287-018-1078-8 . Bansal R, Jain A. Current overview on dental stem cells applications in regenerative dentistry. J Nat Sci Biol Med. 2015;6(1):29–34. https://doi.org/10.4103/0976-9668.149074 . Article PubMed PubMed Central Google Scholar Edgar Ledesma-Martínez, Víctor Manuel Mendoza-Núñez, Edelmiro Santiago-Osorio. Mesenchymal stem cells derived from dental pulp: a review. Stem Cells Int . 2016, 4,709,572, p. doi: https://doi.org/10.1155/2016/4709572 ]. Grzech-Leśniak K. Making use of lasers in periodontal treatment: a new gold standard? Photomed Laser Surg. 2017;35(10):513–4. Miura M, Gronthos S, Zhao M, Lu B, Fisher LW, Robey PG, Shi S. SHED: stem cells from human exfoliated deciduous teeth. Proc Natl Acad Sci U S A. 2003;100(10):5807–12. https://doi.org/10.1073/pnas.0937635100 . Yasui T, Mabuchi Y, Toriumi H, Ebine T, Niibe K, Houlihan DD, Morikawa S, Onizawa K, Kawana H, Akazawa C, Suzuki N, Nakagawa T, Okano H, Matsuzaki Y. Purified human dental pulp stem cells promote osteogenic regeneration. J Dent Res. 2016;95(2):206–14. https://doi.org/10.1177/0022034515610748 . Yamamoto A, Sakai K, Matsubara K, Kano F, Ueda M. Multifaceted neuro-regenerative activities of human dental pulp stem cells for functional recovery after spinal cord injury. Neurosci Res. 2014;78:16–20. https://doi.org/10.1016/j.neures.2013.10.010 . d’Aquino R, De Rosa A, Lanza V, Tirino V, Laino L, Graziano A, Desiderio V, Laino G, Papaccio G. Human mandible bone defect repair by the grafting of dental pulp stem/progenitor cells and collagen sponge biocomplexes. Eur Cell Mater. 2009;12, PMID: 19908196:75–83. Wang L, Shen H, Zheng W, Tang L, Yang Z, Gao Y, Yang Q, Wang C, Duan Y, Jin Y. Characterization of stem cells from alveolar periodontal ligament. Tissue Eng. Part A. 2011;17(7–8):1015–26. https://doi.org/10.1089/ten.tea.2010.0140 . Iwata T, Yamato M, Zhang Z, Mukobata S, Washio K, Ando T, Feijen J, Okano T, Ishikawa I. Validation of human periodontal ligament-derived cells as a reliable source for cytotherapeutic use. J Clin Periodontol. 2010;37(12):1088–99. https://doi.org/10.1111/j.1600-051X.2010.01597.x . Chen F-M, Gao L-N, Tian B-M, Zhang X-Y, Zhang Y-J, Dong G-Y, Lu H, et al. Treatment of periodontal intrabony defects using autologous periodontal ligament stem cells: a randomized clinical trial. Stem Cell Res Ther. 2016;7:33. https://doi.org/10.1186/s13287-016-0288-1 . Bakopoulou A, Leyhausen G, Volk J, Tsiftsoglou A, Garefis P, Koidis P, Geurtsen W. Comparative analysis of in vitro osteo/odontogenic differentiation potential of human dental pulp stem cells (DPSCs) and stem cells from the apical papilla (SCAP). Arch Oral Biol. 2011;56(7):709–21. https://doi.org/10.1016/j.archoralbio.2010.12.008 . Han C, Yang Z, Zhou W, Jin F, Song Y, Wang Y, Huo N, Chen L, Qian H, Hou R, Duan Y, Jin Y. Periapical follicle stem cell: a promising candidate for cementum/periodontal ligament regeneration and bio-root engineering. Stem Cells Dev. 2010;19(9):1405–15. https://doi.org/10.1089/scd.2009.0277 . Luan X, Ito Y, Dangaria S, Diekwisch TG. Dental follicle progenitor cell heterogeneity in the developing mouse periodontium. Stem Cells Dev. 2006;15(4):595–608. https://doi.org/10.1089/scd.2006.15.595 . Handa K, Saito M, Tsunoda A, Yamauchi M, Hattori S, Sato S, Toyoda M, Teranaka T, Narayanan AS. Progenitor cells from dental follicle are able to form cementum matrix in vivo. Connect Tissue Res. 2002;43(2–3):406–8 PMID: 12489190. Guo W, Chen L, Gong K, Ding B, Duan Y, Jin Y. Heterogeneous dental follicle cells and the regeneration of complex periodontal tissues. Tissue Engineering. Part A. 2012;18(5–6):459–70 https://doi.org/10.1089/ten.tea.2011.0261 . Bai, Yudi et al. Cementum- and periodontal ligament-like tissue formation by dental follicle cell sheets co-cultured with Hertwig’s epithelial root sheath cells. Bone. 2011, 48, Issue 6, pp. 1417–1426, https://doi.org/10.1016/j.bone.2011.02.016 . Cordeiro MM, Dong Z, Kaneko T, Zhang Z, Miyazawa M, Shi S, et al. Dental pulp tissue engineering with stem cells from exfoliated deciduous teeth. 2008, 34, pp. 962–969. Dobie K, Smith G, Sloan AJ, Smith AJ. Effects of alginate, hydrogels and TGF-beta 1 on human dental pulp repair in vitro. Connect Tissue Res 2. 2002;43:387–90. Friedlander LT, Cullinan MP, Love RM. Dental stem cells and their potential role in apexogenesis and apexification. Int Endod J. 2009;42:955–62. Cai J, Zhang Y, Liu P, Chen S, Wu X, Sun Y, Li A, Huang K, Luo R, Wang L, Liu Y, Zhou T, Wei S, Pan G, Pei D, Generation of tooth-like structures from integration-free human urine induced pluripotent stem cells. Cell Regen (Lond). July 30, 2013, 2(1), pp. 6, doi: https://doi.org/10.1186/2045-9769-2-6 . Craig J. Taylor, Eleanor M. Bolton, and J. Andrew Bradley 2011 Aug 12 and https://doi.org/10.1098/rstb.2011.0030 ], 366(1575): 2312–2322. [doi:. Immunological considerations for embryonic and induced pluripotent stem cell banking,. Philos Trans R SocLond B Biol Sci. 2011, 366(1575), pp. 2312–2322, doi: https://doi.org/10.1098/rstb.2011.0030 . T.R. Nayak, H. Andersen, V.S. Makam, C. Khaw, S. Bae, X.F. Xu, P.L.R. Ee, J.H. Ahn, B.H. Hong, G. Pastorin, B. Ozyilmaz, ACS Nano, 5 (6) (2011), pp. 4. Graphene for controlled and accelerated osteogenic differentiation of human mesenchymal stem cells,. ACS Nano. 2011, pp. 4670–4678. Lee WC, Lim C, Shi H, Tang LAL, Wang Y, Lim CT, Loh KP. Origin of enhanced stem cell growth and differentiation on graphene and graphene oxide. ACS Nano. 2011;5(9):7334–41. Kenry LWC, Loh KP, Lim CT. When stem cells meet graphene: opportunities and challenges in regenerative medicine. Biomaterials. 2018;155:236–50. Yuan A, Farber EL, Rapoport AL, Tejada D, Deniskin R, Akhmedov NB, et al. Transfer of microRNAs by embryonic stem cell microvesicles. 2009. 2009, 4(3), p. https://doi.org/10.1371/journal.pone . 0004722. Oh, Myeongsik, et al. Exosomes derived from human induced pluripotent stem cells ameliorate the aging of skin fibroblasts. Int. J. Mol. Sci. 2018, 19(6), p. 1715. Ramirez MI. et al. Technical challenges of working with extracellular vesicles. Nanoscale. 2018;10:881–906. Valadi H, et al. Exosome-mediated transfer of mRNAs and microRNAs is a novel mechanism of genetic exchange between cells. Nat. Cell Biol. 2007;9:654–9. Mateescu B, et al. Obstacles and opportunities in the functional analysis of extracellular vesicle RNA—an ISEV position paper. J. Extracell. Vesicles. 2017;6(1). https://doi.org/10.1080/20013078.2017.1286095 . Nawaz M, et al. Extracellular vesicles: evolving factors in stem cell biology. Stem Cells Int. 2016;2016:17. Article ID 1073140. Helfrich, Y.R., Sachs, D.L. and Voorhees, J.J. Overview of skin aging and photoaging. Dermatol. Nurs. 20, pp. 177–183, https://www.ncbi.nlm.nih.gov/pubmed/18649702 . Julia Tigges, Jean Krutmann, Ellen Fritsche, Judith Haendeler, Heiner Schaal, Jens W. Fischer, Faiza Kalfalah, Hans Reinke, Guido Reifenberger, Kai Stühler, Natascia Ventura, Sabrina Gundermann, Petra Boukamp, Fritz Boege. The hallmarks of fibroblast ageing, mechanisms of ageing and development, 138, 2014, Pages 26–44. 2014, 138, pp. 26–44, ISSN 0047–6374, https://doi.org/10.1016/j.mad.2014.03.004 . Huh MI, Kim MS, Kim HK, et al. Effect of conditioned media collected from human amniotic fluid-derived stem cells (hAFSCs) on skin regeneration and photo-aging. Tissue Eng Regen Med. 2014;11:171 https://doi.org/10.1007/s13770-014-0412-1 . Togel F, Hu Z, Weiss K, et al. Administered mesenchymal stem cells protect against ischemic acute renal failure through differentiation-independent mechanisms. Am J Physiol Renal Physiol. 2005;289:F31. Liu J, Han G, Liu H, et al. Suppression of cholangiocarcinoma cell growth by human umbilical cord mesenchymal stem cells: a possible role of Wnt and Akt signaling. PLoS One. 2013;8:e62844. Oh M, et al. Promotive effects of human induced pluripotent stem cell-conditioned medium on the proliferation and migration of dermal fibroblasts. Biotechnol. Bioprocess Eng. 2017;22:561–8. Chen L, Tredget EE, Wu PY, Wu Y. Paracrine factors of mesenchymal stem cells recruit macrophages and endothelial lineage cells and enhance wound healing. PloS One. 2008;3:e1886. Bae J-S, Lee S-H, Kim J-E, Choi J-Y, Park R-W, Park JY, Park H-S, Sohn Y-S, Lee D-S, Lee EB. βig-h3 supports keratinocyte adhesion, migration, and proliferation through α3β1 integrin. Biochem. Biophys. Res. Commun. 2002;294:940–8. Zhou B-R, Xu Y, Guo S-L, Xu Y, Wang Y, Zhu F, Permatasari F, Wu D, Yin Z-Q, Luo D. The effect of conditioned media of adipose-derived stem cells on wound healing after ablative fractional carbon dioxide laser resurfacing. BioMed Res. Int. 2013;519:126. Peng Y, Baulier E, Ke Y, Young A, Ahmedli NB, Schwartz SD, et al. Human embryonic stem cells extracellular vesicles and their effects on immortalized human retinal Müller cells. PLoS ONE. 2018, 13(3), p. https://doi.org/10.1371/journal.pone.019400 . Harris MT, Butler DL, Boivin GP, Florer JB, Schantz EJ, Wenstrup RJ. Mesenchymal stem cells used for rabbit tendon repair can form ectopic bone and express alkaline phosphatase activity in constructs. J Orthop Res. 2004;22:998–1003. Mascetti VL, Pedersen RA. Human-mouse chimerism validates human stem cell pluripotency. Cell Stem Cell. 2016;18:67–72. Gandia C, Armiñan A, García-Verdugo JM, Lledó E, Ruiz A, Miñana MD, Sanchez-Torrijos J, Payá R, Mirabet V, Carbonell-Uberos F, Llop M, Montero JA, Sepúlveda P. Human dental pulp stem cells improve left ventricular function, induce angiogenesis, and reduce infarct size in rats with acute myocardial infarction. Stem Cells. 2007;26(3):638–45. Perry BC, Zhou D, Wu X, Yang FC, Byers MA, Chu TM, Hockema JJ, Woods EJ, Goebel WS. Collection, cryopreservation, and characterization of human dental pulp-derived mesenchymal stem cells for banking and clinical use. Tissue Eng Part C Methods. 2008;14(2):149–56. Garcia-Olmo D, Garcia-Arranz M, Herreros D, et al. A phase I clinical trial of the treatment of Crohn’s fistula by adipose mesenchymal stem cell transplantation. Dis Colon Rectum. 2005;48:1416–23. de Mendonça CA, Bueno DF, Martins MT, Kerkis I, Kerkis A, Fanganiello RD, Cerruti H, Alonso N, Passos-Bueno MR. Reconstruction of large cranial defects in nonimmunosuppressed experimental design with human dental pulp stem cells. J Craniofac Surg. 2008;19(1):204–10. Seo BM, Sonoyama W, Yamaza T, Coppe C, Kikuiri T, Akiyama K, Lee JS, Shi S. SHED repair critical-size calvarial defects in mice. Oral Dis. 2008;14(5):428–34. Abbas, Diakonov I., Sharpe P. Neural crest origin of dental stem cells. Pan European Federation of the International Association for Dental Research (PEF IADR). 2008, Vols. Seq #96 - Oral Stem Cells. Kerkis I, Ambrosio CE, Kerkis A, Martins DS, Gaiad TP, Morini AC, Vieira NM, Marina P, et al. Early transplantation of human immature dental pulp stem cells from baby teeth to golden retriever muscular dystrophy (GRMD) dogs. J Transl Med. 2008;6:35. Xianrui Yang, Li Li, Li Xiao, Donghui Zhang. Recycle the dental fairy’s package: overview of dental pulp stem cells. Stem Cell Res Ther . 2018, 9, 1, 1. https://doi.org/10.1186/s13287-018-1094-8 . Wang J, Wang X, Sun Z, Wang X, Yang H, Shi S, Wang S. Stem cells from human-exfoliated deciduous teeth can differentiate into dopaminergic neuron-like cells. Stem Cells Dev. 2010;19:1375–83. Wang J, et al. The odontogenic differentiation of human dental pulp stem cells on nanofibrous poly (L-lactic acid) scaffolds in vitro and in vivo. Acta Biomater. 2010;6(10):3856–63. Davies OG, Cooper PR, Shelton RM, Smith AJ, Scheven BA. A comparison of the in vitro mineralisation and dentinogenic potential of mesenchymal stem cells derived from adipose tissue, bone marrow and dental pulp. J Bone Miner Metab. 2015;33:371–82. Huang GT-J, Shagramanova K, Chan SW. Formation of odontoblast-like cells from cultured human dental pulp cells on dentin in vitro. J Endod. 2006;32:1066–73. Shi S, Robey PG, Gronthos S. Comparison of human dental pulp and bone marrow stromal stem cells by cDNA microarray analysis. Bone. 2001;29(6):532–9. Gronthos S, Mankani M, Brahim J, Robey PG, Shi S. Postnatal human dental pulp stem cells (DPSCs) in vitro and in vivo. Proc Natl Acad Sci U S A. 2000;97:13625–30. Nuti N, Corallo C, Chan BMF, Ferrari M, Gerami-Naini B. Multipotent differentiation of human dental pulp stem cells: a literature review. Stem Cell Rev Rep. 2016;12:511–23. Ferro F, et al. Dental pulp stem cells differentiation reveals new insights in Oct4A dynamics. PloS One. 2012;7(7):e41774. Conde MCM, Chisini LA, Grazioli G, Francia A, Carvalho RVd, Alcázar JCB, Tarquinio SBC, Demarco FF. Does cryopreservation affect the biological properties of stem cells from dental tissues? A systematic review. Braz Dent J. 2016;1210(6):633-40. https://doi.org/10.1590/0103-6440201600980 . Papaccio G, Graziano A, d’Aquino R, Graziano MF, Pirozzi G, Menditti D, De Rosa A, Carinci F, Laino G. Long-term cryopreservation of dental pulp stem cells (SBP-DPSCs) and their differentiated osteoblasts: a cell source for tissue repair. J Cell Physiol. 2006;208:319–25. Alge DL, Zhou D, Adams LL, et. al. Donor-matched comparison of dental pulp stem cells and bone marrow-derived mesenchymal stem cells in a rat model. J Tissue Eng Regen Med. 2010;4(1):73–81. Jo Y-Y, Lee H-J, Kook S-Y, Choung H-W, Park J-Y, Chung J-H, Choung Y-H, Kim E-S, Yang H-C, Choung P-H. Isolation and characterization of postnatal stem cells from human dental tissues. Tissue Eng. 2007;13:767–73. Gronthos S, Brahim J, Li W, Fisher LW, Cherman N, Boyde A, DenBesten P, Robey PG, Shi S. Stem cell properties of human dental pulp stem cells. J Dent Res. 2002;81:531–5. Laino G, d’Aquino R, Graziano A, Lanza V, Carinci F, Naro F, Pirozzi G, Papaccio G. A new population of human adult dental pulp stem cells: a useful source of living autologous fibrous bone tissue (LAB). J Bone Miner Res. 2005;20:1394–402. Zainal A, Shahrul H, et al. In vitro chondrogenesis transformation study of mouse dental pulp stem cells. Sci World J. 2012;2012:827149. Wei X, et al. Expression of mineralization markers in dental pulp cells. J Endod. 2007;33(6):703–8. Dai J, et al. The effect of co-culturing costal chondrocytes and dental pulp stem cells combined with exogenous FGF9 protein on chondrogenesis and ossification in engineered cartilage. Biomaterials. 2012;33(31):7699–711. Vasandan AB, et al. Functional differences in mesenchymal stromal cells from human dental pulp and periodontal ligament. J Cell Mol Med. 2014;18(2):344–54. Werle SB, et al. Carious deciduous teeth are a potential source for dental pulp stem cells. Clin Oral Investig. 2015;20:75–81. Nemeth CL, et al. Enhanced chondrogenic differentiation of dental pulp stem cells using nanopatterned PEG-GelMA-HA hydrogels. Tissue Eng A. 2014;20(21–22):2817–29. Paino F, Ricci G, De Rosa A, D’Aquino R, Laino L, Pirozzi G, et al. Ecto-mesenchymal stem cells from dental pulp are committed to differentiate into active melanocytes. Eur. Cell Mater. 2010;20:295–305. Ferro F, Spelat R, Baheney CS. Dental pulp stem cell (DPSC) isolation, characterization, and differentiation. In: Kioussi C, editor. Stem cells and tissue repair. Methods in molecular biology (methods and protocols): Humana Press. 2014;1210. Ishkitiev N, Yaegaki K, Imai T, Tanaka T, Nakahara T, Ishikawa H, Mitev V, Haapasalo M. High-purity hepatic lineage differentiated from dental pulp stem cells in serum-free medium. J Endod. 2012;38:475–80. Download references AcknowledgementsNot applicable. This work is supported by Wrocław Medical University in Poland. Availability of data and materialsPlease contact author for data requests. Author informationAuthors and affiliations. Department of Experimental Surgery and Biomaterials Research, Wroclaw Medical University, Bujwida 44, Wrocław, 50-345, Poland Wojciech Zakrzewski, Maria Szymonowicz & Zbigniew Rybak Department of Conservative Dentistry and Pedodontics, Krakowska 26, Wrocław, 50-425, Poland Maciej Dobrzyński You can also search for this author in PubMed Google Scholar ContributionsWZ is the principal author and was responsible for the first draft of the manuscript. WZ and ZR were responsible for the concept of the review. MS, MD, and ZR were responsible for revising the article and for data acquisition. All authors read and approved the final manuscript. Corresponding authorCorrespondence to Wojciech Zakrzewski . Ethics declarationsEthics approval and consent to participate, consent for publication, competing interests. The authors declare that they have no competing interests. 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Stem Cells and the Future of Regenerative Medicine (2002)Chapter: executive summary, executive summary. S tem cell research offers unprecedented opportunities for developing new medical therapies for debilitating diseases and a new way to explore fundamental questions of biology. Stem cells are unspecialized cells that can self-renew indefinitely and also differentiate into more mature cells with specialized functions. Research on human embryonic stem cells, however, is controversial, given the diverse views held in our society about the moral and legal status of the early embryo. The controversy has encouraged provocative and conflicting claims both inside and outside the scientific community about the biology and biomedical potential of both adult and embryonic stem cells. The National Research Council and Institute of Medicine formed the Committee on the Biological and Biomedical Applications of Stem Cell Research to address the potential of stem cell research. The committee organized a workshop that was held on June 22, 2001. At the workshop, the committee heard from many leading scientists who are engaged in stem cell research and from philosophers, ethicists, and legal scholars. (Audio files of the speakers’ presentations are available until December 31, 2002, at the workshop Web site, www.nationalacademies.org/stemcells .) The participants discussed the science of stem cells and a variety of ethical and other arguments relevant to public policy as it applies to stem cells. The committee considered the information presented, explored the literature on its own, and contemplated the substance and importance of the preliminary data from recent stem cell experiments. The committee’s deliberations on the issues led to the following conclusions and recommendations. Experiments in mice and other animals are necessary, but not sufficient, for realizing the potential of stem cells to develop tissue-replacement therapies that will restore lost function in damaged organs. Because of the substantial biological differences between nonhuman animal and human development and between animal and human stem cells, studies with human stem cells are essential to make progress in the development of treatments for human disease, and this research should continue. There are important biological differences between adult and embryonic stem cells and among adult stem cells found in different types of tissue. The implications of these biological differences for therapeutic uses are not yet clear, and additional data are needed on all stem cell types. Adult stem cells from bone marrow have so far provided most of the examples of successful therapies for replacement of diseased or destroyed cells. Despite the enthusiasm generated by recent reports, the potential of adult stem cells to differentiate fully into other cell types (such as brain, nerve, pancreas cells) is still poorly understood and remains to be clarified. In contrast, studies of human embryonic stem cells have shown that they can develop into multiple tissue types and exhibit long-term self-renewal in culture, features that have not yet been demonstrated with many human adult stem cells. The application of stem cell research to therapies for human disease will require much more knowledge about the biological properties of all types of stem cells. Although stem cell research is on the cutting edge of biological science today, it is still in its infancy. Studies of both embryonic and adult human stem cells will be required to most efficiently advance the scientific and therapeutic potential of regenerative medicine. Moreover, research on embryonic stem cells will be important to inform research on adult stem cells, and vice versa. Research on both adult and embryonic human stem cells should be pursued. Over time, all cell lines in tissue culture change, typically accumulating harmful genetic mutations. There is no reason to expect stem cell lines to behave differently. In addition, most existing stem cell lines have been cultured in the presence of non-human cells or serum that could lead to potential human health risks. Consequently, while there is much that can be learned using existing stem cell lines if they are made widely available for research, such concerns necessitate continued monitoring of these cells as well as the development of new stem cell lines in the future. High-quality, publicly funded research is the wellspring of medical breakthroughs. Although private, for-profit research plays a critical role in translating the fruits of basic research into medical advances that are broadly available to the public, stem cell research is far from the point of providing therapeutic products. Without public funding of basic research on stem cells, progress toward medical therapies is likely to be hindered. In addition, public funding offers greater opportunities for regulatory oversight and public scrutiny of stem cell research. Stem cell research that is publicly funded and conducted under established standards of open scientific exchange, peer review, and public oversight offers the most efficient and responsible means of fulfilling the promise of stem cells to meet the need for regenerative medical therapies. Conflicting ethical perspectives surround the use of embryonic stem cells in medical research, particularly where the moral and legal status of human embryos is concerned. The use of embryonic stem cells is not the first biomedical research activity to raise ethical and social issues among the public. Restrictions and guidelines for the conduct of controversial research have been developed to address such concerns in other instances. For example, when recombinant-DNA techniques raised questions and were subject to intense debate and public scrutiny, a national advisory body, the Recombinant DNA Advisory Committee, was established at the National Institutes of Health (NIH) to ensure that the research met the highest scientific and ethical standards. If the federal government chooses to fund research on human embryonic stem cells, a similar national advisory group composed of exceptional researchers, ethicists, and other stakeholders should be established at NIH to oversee it. Such a group should ensure that proposals to work on human embryonic stem cells are scientifically justified and should scrutinize such proposals for compliance with federally mandated ethical guidelines. Regenerative medicine is likely to involve the implantation of new tissue in patients with damaged or diseased organs. A substantial obstacle to the success of transplantation of any cells, including stem cells and their derivatives, is the immune-mediated rejection of foreign tissue by the recipient’s body. In current stem cell transplantation procedures with bone marrow and blood, success can hinge on obtaining a close match between donor and recipient tissues and on the use of immunosuppressive drugs, which often have severe and life-threatening side effects. To ensure that stem cell-based therapies can be broadly applicable for many conditions and individuals, new means to overcome the problem of tissue rejection must be found. Although ethically controversial, somatic cell nuclear transfer, a technique that produces a lineage of stem cells that are genetically identical to the donor, promises such an advantage. Other options for this purpose include genetic manipulation of the stem cells and the development of a very large bank of embryonic stem cell lines. In conjunction with research on stem cell biology and the development of stem cell therapies, research on approaches that prevent immune rejection of stem cells and stem cell-derived tissues should be actively pursued. The committee is aware of and respectful of the wide array of social, political, legal, ethical, and economic issues that must be considered in policy-making in a democracy. And it is impressed by the commitment of all parties in this debate to life and health, regardless of the different conclusions they draw. The committee hopes that this report, by clarifying what is known about the scientific potential of stem cells and how that potential can best be realized, will be a useful contribution to the debate and to the enhancement of treatments for disabling human diseases and injuries. On August 9, 2001, when President Bush announced a new federal policy permitting limited use of human embryonic stem cells for research, this report was already in review. Because this report presents the committee’s interpretation of the state of the science of stem cells independent of any specific policy, only minor modifications to refer to the new policy have been made in the report. stem cells are essential to make progress in the development of treatments for disease, and this research should continue. | This page in the original is blank. | Recent scientific breakthroughs, celebrity patient advocates, and conflicting religious beliefs have come together to bring the state of stem cell research—specifically embryonic stem cell research—into the political crosshairs. President Bush's watershed policy statement allows federal funding for embryonic stem cell research but only on a limited number of stem cell lines. Millions of Americans could be affected by the continuing political debate among policymakers and the public. Stem Cells and the Future of Regenerative Medicine provides a deeper exploration of the biological, ethical, and funding questions prompted by the therapeutic potential of undifferentiated human cells. In terms accessible to lay readers, the book summarizes what we know about adult and embryonic stem cells and discusses how to go about the transition from mouse studies to research that has therapeutic implications for people. Perhaps most important, Stem Cells and the Future of Regenerative Medicine also provides an overview of the moral and ethical problems that arise from the use of embryonic stem cells. This timely book compares the impact of public and private research funding and discusses approaches to appropriate research oversight. Based on the insights of leading scientists, ethicists, and other authorities, the book offers authoritative recommendations regarding the use of existing stem cell lines versus new lines in research, the important role of the federal government in this field of research, and other fundamental issues. READ FREE ONLINE Welcome to OpenBook!You're looking at OpenBook, NAP.edu's online reading room since 1999. Based on feedback from you, our users, we've made some improvements that make it easier than ever to read thousands of publications on our website. Do you want to take a quick tour of the OpenBook's features? Show this book's table of contents , where you can jump to any chapter by name. ...or use these buttons to go back to the previous chapter or skip to the next one. Jump up to the previous page or down to the next one. Also, you can type in a page number and press Enter to go directly to that page in the book. Switch between the Original Pages , where you can read the report as it appeared in print, and Text Pages for the web version, where you can highlight and search the text. To search the entire text of this book, type in your search term here and press Enter . Share a link to this book page on your preferred social network or via email. View our suggested citation for this chapter. Ready to take your reading offline? Click here to buy this book in print or download it as a free PDF, if available. Get Email UpdatesDo you enjoy reading reports from the Academies online for free ? Sign up for email notifications and we'll let you know about new publications in your areas of interest when they're released. Home > Dedman School of Law > Law Journals > scitech > Vol. 25 (2022) > No. 2 SMU Science and Technology Law ReviewCreating a united front: harmonizing the united states regulatory policies surrounding human embryonic stem cell research. Sydney Kossow , Southern Methodist University, Dedman School of Law Follow Stem cell therapy is an imperative development in science and medicine that is heavily regulated worldwide. With the potential to cure illnesses, help understand disease development, and advance regenerative medicine, a harmonized regulatory policy is crucial to capitalize on the benefits of stem cells. This article examines an important topic of discussion surrounding stem cell therapy and research: the political debate on how and when embryonic stem cells can be used. In addition to examining ethical challenges, this article discusses the legal challenges surrounding using embryonic stem cells to inform regenerative therapies. Specifically, this article will examine the National Institute of Health’s Guidelines for Human Stem Cell Research and the historic avenues of federal and state legislation to regulate the use of these cells in research. This article discusses the internal and external inconsistencies of the United States’ current regulation of embryonic stem cells and how the divide between states is problematic for the United States’ completive stance in developmental science and medicine. Finally, this article contemplates a cohesive regulatory system influenced by individual states and other countries that currently lead the medical field, to form a united front in approaching the use of stem cells. Recommended CitationSydney Kossow, Creating a United Front: Harmonizing the United States Regulatory Policies Surrounding Human Embryonic Stem Cell Research , 25 SMU Sci. & Tech. L. Rev. 295 (2022) Since December 20, 2022 Included inComputer Law Commons , Intellectual Property Law Commons , Internet Law Commons , Science and Technology Law Commons Digital Object Identifier (DOI)https://doi.org/10.25172/smustlr.25.2.7 - Journal Home
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Advanced Search About | My Account | Accessibility Statement Privacy Copyright An Ode to Stem CellsLeveraging the versatility of stem cells allows researchers to advance science across multiple disciplines.. Meenakshi is the Editor-in-Chief at The Scientist. Her diverse science communication experience includes journalism, podcasting, and corporate content strategy. Meenakshi earned her PhD in biophysics from the University of Goettingen, Germany. View full profile. Learn about our editorial policies. ABOVE: © istock.com, luismmolina U ntil a few hundred years ago, before scientific advances allowed researchers to peer inside the human body or extract cells to study them in the laboratory, I imagine that the mysteries of developmental biology—more than any other discipline—must have stumped humans. On one hand, one could visibly track a baby’s growth based on the size of the mother's growing bump; if everything went right, a healthy baby would emerge after nine months. On the other hand, sometimes a seemingly healthy mother lost her child mid-pregnancy or delivered an abnormal newborn. In the absence of scientific technologies to probe into the womb, the inner workings of embryonic development remained a black box and people had little to no way of knowing what to expect when someone was expecting. While several questions remain unanswered even today, scientists rapidly bridged most of the knowledge gaps once stem cell research entered the scene. You might recall that in our winter issue last year we covered how researchers used adult stem cells to better understand placental development . In one of the articles in this issue, we profile a biologist with expertise in endometrial research who found that stem cells were a powerful tool for solving long-standing mysteries about women's health. While access to adult stem cells has certainly helped answer some questions in developmental biology, embryonic stem cell studies, in my opinion, truly transformed the research area. Just like the newborn that the embryo eventually forms into, these early-stage cells have the potential to choose any path of maturation. By developing 3D embryonic stem cell models, researchers study the differentiation and growth of these cells without worrying about the restrictive rules on embryo research (read more about these advances in one of the feature stories in this issue). What also fascinates me is that researchers can now achieve single cell resolution in their quest to determine how life develops early on; one research team recently found that the initial two cells in an embryo take diverse developmental paths . 1 These studies are a stark reminder that scientists have indeed come a long way from not knowing what happens during the months-long gestation period to following the development of two individual cells to determine their eventual contribution to structure and function! In contrast to the early embryonic development studies inspired by visible signs, it must have been hard in those days to imagine problems within an individual’s brain when their symptoms did not match known physical disorders. Securing neurons from the brain is also not as easy as isolating most other cell types. It is no secret that the ability to transform normal cells into induced pluripotent stem cells revolutionized neuroscience, as researchers finally found a way to model brain disorders in these cells. Now with more knowledge about how sex affects disease, researchers are correcting the long-standing sex bias in the field to further refine our understanding of the human brain. For those interested in reading more about this topic, we dive into the measures that researchers are taking to include sex as a biological variable in their studies in a feature article in this issue. All in all, the applications of stem cells are as diverse as the cells’ differentiation abilities, and researchers have only scratched the surface so far. I hope you experience the same enthusiasm and excitement reading the stem cell stories in this issue that we had while crafting them. - Junyent S, et al. The first two blastomeres contribute unequally to the human embryo . Cell. 2024;187(11):2838-2854.
NIH Stem Cell InformationNih guidelines for human stem cell research. SUMMARY: The National Institutes of Health (NIH) is hereby publishing final "National Institutes of Health Guidelines for Human Stem Cell Research" (Guidelines). On March 9, 2009, President Barack H. Obama issued Executive Order 13505: Removing Barriers to Responsible Scientific Research Involving Human Stem Cells. The Executive Order states that the Secretary of Health and Human Services, through the Director of NIH, may support and conduct responsible, scientifically worthy human stem cell research, including human embryonic stem cell (hESC) research, to the extent permitted by law. These Guidelines implement Executive Order 13505, as it pertains to extramural NIH-funded stem cell research, establish policy and procedures under which the NIH will fund such research, and helps ensure that NIH-funded research in this area is ethically responsible, scientifically worthy, and conducted in accordance with applicable law. Internal NIH policies and procedures, consistent with Executive Order 13505 and these Guidelines, will govern the conduct of intramural NIH stem cell research. EFFECTIVE DATE: These Guidelines are effective on July 7, 2009. SUMMARY OF PUBLIC COMMENTS ON DRAFT GUIDELINES: On April 23, 2009 the NIH published draft Guidelines for research involving hESCs in the Federal Register for public comment, 74 Fed. Reg. 18578 (April 23, 2009). The comment period ended on May 26, 2009. The NIH received approximately 49,000 comments from patient advocacy groups, scientists and scientific societies, academic institutions, medical organizations, religious organizations, and private citizens. The NIH also received comments from members of Congress. This Notice presents the final Guidelines together with the NIH response to public comments that addressed provisions of the Guidelines. Title of the Guidelines, Terminology, and Background : Respondents felt the title of the NIH draft guidelines was misleading, in that it is entitled "National Institutes of Health Guidelines for Human Stem Cell Research," yet addresses only one type of human stem cell. The NIH notes that although the Guidelines pertain primarily to the donation of embryos for the derivation of hESCs, one Section also applies to certain uses of both hESCs and human induced pluripotent stem cells. Also, the Guidelines discuss applicable regulatory standards when research involving human adult stem cells or induced pluripotent stem cells constitutes human subject research. Therefore, the title of the Guidelines was not changed. Respondents also disagreed with the definition of human embryonic stem cells in the draft Guidelines, and asked that the NIH define them as originating from the inner cell mass of the blastocyst. The NIH modified the definition to say that human embryonic stem cells "are cells that are derived from the inner cell mass of blastocyst stage human embryos, are capable of dividing without differentiating for a prolonged period in culture, and are known to develop into cells and tissues of the three primary germ layers." Financial Gain Respondents expressed concern that derivers of stem cells might profit from the development of hESCs. Others noted that because the stem cells eligible for use in research using NIH funding under the draft Guidelines are those cells that are subject to existing patents, there will be insufficient competition in the licensing of such rights. These respondents suggested that this could inhibit research, as well as increase the cost of any future clinical benefits. The Guidelines do not address the distribution of stem cell research material. It is, however, the NIH's expectation that stem cell research materials developed with NIH funds, as well as associated intellectual property and data, will be distributed in accordance with the NIH’s existing policies and guidance, including "Sharing Biomedical Research Resources, Principles and Guidelines for Recipients of NIH Grants and Contracts" and "Best Practices for the Licensing of Genomic Inventions." http://www.ott.nih.gov/policy/policies_and_guidelines.aspx Even where such policies are not directly applicable, the NIH encourages others to refrain from imposing on the transfer of research tools, such as stem cells, any conditions that hinder further biomedical research. In addition, the Guidelines were revised to state that there should be documentation that "no payments, cash or in kind, were offered for the donated embryos." Respondents were concerned that donor(s) be clearly "apprised up front by any researchers that financial gain may come from the donation and that the donor(s) should know up front if he/she will share in the financial gain." The Guidelines address this concern by asking that donor(s) was/were informed during the consent process that the donation was made without any restriction or direction regarding the individual(s) who may receive medical benefit from the use of the stem cells, such as who may be the recipients of cell transplants. The Guidelines also require that the donor(s) receive(s) information that the research was not intended to provide direct medical benefit to the donor(s); that the results of research using the hESCs may have commercial potential, and that the donor(s) would not receive financial or any other benefits from any such commercial development. IRB Review under the Common Rule Respondents suggested that the current regulatory structure of IRB review under the Common Rule (45 C.F.R. Part 46, Subpart A) addresses the core ethical principles needed for appropriate oversight of hESC derivation. They noted that IRB review includes a full review of the informed consent process, as well as a determination of whether individuals were coerced to participate in the research and whether any undue inducements were offered to secure their participation. These respondents urged the NIH to replace the specific standards to assure voluntary and informed consent in the draft Guidelines with a requirement that hESC research be reviewed and approved by an IRB, in conformance with 45 C.F.R. Part 46, Subpart A, as a prerequisite to NIH funding. Respondents also requested that the NIH create a registry of eligible hESC lines to avoid burdensome and repetitive assurances from multiple funding applicants. The NIH agrees that the IRB system of review under the Common Rule provides a comprehensive framework for the review of the donation of identifiable human biological materials for research. However, in the last several years, guidelines on hESC research have been issued by a number of different organizations and governments, and different practices have arisen around the country and worldwide, resulting in a patchwork of standards. The NIH concluded that employing the IRB review system for the donation of embryos would not ameliorate stated concerns about variations in standards for hESC research and would preclude the establishment of an NIH registry of hESCs eligible for NIH funding, because there would be no NIH approval of particular hESCs. To this end and response to comments, these Guidelines articulate policies and procedures that will allow the NIH to create a Registry. These Guidelines also provide scientists who apply for NIH funding with a specific set of standards reflecting currently recognized ethical principles and practices specific to embryo donation that took place on or after the issuance of the Guidelines, while also establishing procedures for the review of donations that took place before the effective date of the Guidelines. Federal Funding Eligibility of Human Pluripotent Cells from Other Sources Respondents suggested that the allowable sources of hESCs potentially available for federal funding be expanded to include hESC lines from embryos created expressly for research purposes, and lines created, or pluripotent cells derived, following parthenogenesis or somatic cell nuclear transfer (SCNT). The Guidelines allow for funding of research using hESCs derived from embryos created using in vitro fertilization (IVF) for reproductive purposes and no longer needed for these purposes, assuming the research has scientific merit and the embryos were donated after proper informed consent was obtained from the donor(s). The Guidelines reflect the broad public support for federal funding of research using hESCs created from such embryos based on wide and diverse debate on the topic in Congress and elsewhere. The use of additional sources of human pluripotent stem cells proposed by the respondents involve complex ethical and scientific issues on which a similar consensus has not emerged. For example, the embryo-like entities created by parthenogenesis and SCNT require women to donate oocytes, a procedure that has health and ethical implications, including the health risk to the donor from the course of hormonal treatments needed to induce oocyte production. Respondents noted that many embryos undergo Pre-implantation Genetic Diagnosis (PGD). This may result in the identification of chromosomal abnormalities that would make the embryos medically unsuitable for clinical use. In addition, the IVF process may also produce embryos that are not transferred into the uterus of a woman because they are determined to be not appropriate for clinical use. Respondents suggested that hESCs derived from such embryos may be extremely valuable for scientific study, and should be considered embryos that were created for reproductive purposes and were no longer needed for this purpose . The NIH agrees with these comments. As in the draft, the final Guidelines allow for the donation of embryos that have undergone PGD. Donation and Informed Consent Respondents commented in numerous ways that the draft Guidelines are too procedurally proscriptive in articulating the elements of appropriate informed consent documentation. This over-reliance on the specific details and format of the informed consent document, respondents argued, coupled with the retroactive application of the Guidelines to embryos already donated for research, would result in a framework that fails to appreciate the full range of factors contributing to the complexity of the informed consent process. For example, respondents pointed to several factors that were precluded from consideration by the proposed Guidelines, such as contextual evidence of the consent process, other established governmental frameworks (representing local and community influences), and the changing standards for informed consent in this area of research over time. Respondents argued that the Guidelines should be revised to allow for a fuller array of factors to be considered in determining whether the underlying ethical principle of voluntary informed consent had been met. In addition to these general issues, many respondents made the specific recommendation that all hESCs derived before the final Guidelines were issued be automatically eligible for Federal funding without further review, especially those eligible under prior Presidential policy, i.e., "grandfathered." The final Guidelines seek to implement the Executive Order by issuing clear guidance to assist this field of science to advance and reach its full potential while ensuring adherence to strict ethical standards. To this end, the NIH is establishing a set of conditions that will maximize ethical oversight, while ensuring that the greatest number of ethically derived hESCs are eligible for federal funding. Specifically, for embryos donated in the U.S. on or after the effective date of the Guidelines, the only way to establish eligibility will be to either use hESCs listed on the NIH Registry, or demonstrate compliance with the specific procedural requirements of the Guidelines by submitting an assurance with supporting information for administrative review by the NIH. Thus, for future embryo donations in the United States, the Guidelines articulate one set of procedural requirements. This responds to concerns regarding the patchwork of requirements and guidelines that currently exist. However, the NIH is also cognizant that in the more than a decade between the discovery of hESCs and today, many lines were derived consistent with ethical standards and/or guidelines developed by various states, countries, and other entities such as the International Society for Stem Cell Research (ISSCR) and the National Academy of Sciences (NAS). These various policies have many common features, rely on a consistent ethical base, and require an informed consent process, but they differ in details of implementation. For example, some require specific wording in a written informed consent document, while others do not. It is important to recognize that the principles of ethical research, e.g., voluntary informed consent to participation, have not varied in this time period, but the requirements for implementation and procedural safeguards employed to demonstrate compliance have evolved. In response to these concerns, the Guidelines state that applicant institutions wishing to use hESCs derived from embryos donated prior to the effective date of the Guidelines may either comply with Section II (A) of the Guidelines or undergo review by a Working Group of the Advisory Committee to the Director (ACD). The ACD, which is a chartered Federal Advisory Committee Act (FACA) committee, will advise NIH on whether the core ethical principles and procedures used in the process for obtaining informed consent for the donation of the embryo were such that the cell line should be eligible for NIH funding. This Working Group will not undertake a de novo evaluation of ethical standards, but will consider the materials submitted in light of the principles and points to consider in the Guidelines, as well as 45 C.F.R. Part 46 Subpart A. Rather than “grandfathering,” ACD Working Group review will enable pre-existing hESCs derived in a responsible manner to be eligible for use in NIH funded research. In addition, for embryos donated outside the United States prior to the effective date of these Guidelines, applicants may comply with either Section II (A) or (B). For embryos donated outside of the United States on or after the effective date of the Guidelines, applicants seeking to determine eligibility for NIH research funding may submit an assurance that the hESCs fully comply with Section II (A) or submit an assurance along with supporting information, that the alternative procedural standards of the foreign country where the embryo was donated provide protections at least equivalent to those provided by Section II (A) of these Guidelines. These materials will be reviewed by the NIH ACD Working Group, which will recommend to the ACD whether such equivalence exists. Final decisions will be made by the NIH Director. This special consideration for embryos donated outside the United States is needed because donation of embryos in foreign countries is governed by the laws and policies of the respective governments of those nations. Although such donations may be responsibly conducted, such governments may not or cannot change their national donation requirements to precisely comply with the NIH Guidelines. The NIH believes it is reasonable to provide a means for reviewing such hESCs because ethically derived foreign hESCs constitute an important scientific asset for the U.S. Respondents expressed concern that it might be difficult in some cases to provide assurance that there was a "clear separation" between the prospective donor(s)’ decision to create embryos for reproductive purposes and the donor(s)’ decision to donate the embryos for research purposes. These respondents noted that policies vary at IVF clinics, especially with respect to the degree to which connections with researchers exist. Respondents noted that a particular clinic’s role may be limited to the provision of contact information for researchers. A clinic that does not have any particular connection with research would not necessarily have in place a written policy articulating the separation contemplated by the Guidelines. Other respondents noted that embryos that are determined not to be suitable for medical purposes, either because of genetic defects or other concerns, may be donated prior to being frozen. In these cases, it is possible that the informed consent process for the donation might be concurrent with the consent process for IVF treatment. Respondents also noted that the initial consent for IVF may contain a general authorization for donating embryos in excess of clinical need, even though a more detailed consent is provided at the actual time of donation. The NIH notes that the Guidelines specifically state that consent should have been obtained at the time of donation, even if the potential donor(s) had given prior indication of a general intent to donate embryos in excess of clinical need for the purposes of research. Accordingly, a general authorization for research donation when consenting for reproductive treatment would comply with the Guidelines, so long as specific consent for the donation is obtained at the time of donation. In response to comments regarding documentation necessary to establish a separation between clinical and research decisions, the NIH has changed the language of the Guidelines to permit applicant institutions to submit consent forms, written policies or other documentation to demonstrate compliance with the provisions of the Guidelines. This change should provide the flexibility to accommodate a range of practices, while adhering to the ethical principles intended. Some respondents want to require that the IVF physician and the hESC researcher should be different individuals, to prevent conflict of interest. Others say they should be the same person, because people in both roles need to have detailed knowledge of both areas (IVF treatment and hESC research). There is also a concern that the IVF doctor will create extra embryos if he/she is also the researcher. As a general matter, the NIH believes that the doctor and the researcher seeking donation should be different individuals. However, this is not always possible, nor is it required, in the NIH's view, for ethical donation. Some respondents want explicit language (in the Guidelines and/or in the consent) stating that the embryo will be destroyed when the inner cell mass is removed. In the process of developing guidelines, the NIH reviewed a variety of consent forms that have been used in responsible derivations. Several had extensive descriptions of the process and the research to be done, going well beyond the minimum expected, yet they did not use these exact words. Given the wide variety and diversity of forms, as well as the various policy, statutory and regulatory obligations individual institutions face, the NIH declines to provide exact wording for consent forms, and instead endorses a robust informed consent process where all necessary details are explained and understood in an ongoing, trusting relationship between the clinic and the donor(s). Respondents asked for clarification regarding the people who must give informed consent for the donation of embryos for research. Some commenters suggested that NIH should require consent from the gamete donors, in cases where those individuals may be different than the individuals seeking reproductive treatment. The NIH requests consent from “the individual(s) who sought reproductive treatment” because this/these individual(s) is/are responsible for the creation of the embryo(s) and, therefore, its/their disposition. With regard to gamete donation, the risks are associated with privacy and, as such, are governed by requirements of the Common Rule, where applicable. Respondents also requested clarification on the statement in the draft Guidelines noting that "although human embryonic stem cells are derived from embryos, such stem cells are not themselves human embryos." For the purpose of NIH funding, an embryo is defined by Section 509, Omnibus Appropriations Act, 2009, Pub. L. 111-8, 3/11/09, otherwise known as the Dickey Amendment, as any organism not protected as a human subject under 45 C.F.R. Part 46 that is derived by fertilization, parthenogenesis, cloning or any other means from one or more human gametes or human diploid cells. Since 1999, the Department of Health and Human Services (HHS) has consistently interpreted this provision as not applicable to research using hESCs, because hESCs are not embryos as defined by Section 509. This long-standing interpretation has been left unchanged by Congress, which has annually reenacted the Dickey Amendment with full knowledge that HHS has been funding hESC research since 2001. These guidelines therefore recognize the distinction, accepted by Congress, between the derivation of stem cells from an embryo that results in the embryo’s destruction, for which federal funding is prohibited, and research involving hESCs that does not involve an embryo nor result in an embryo’s destruction, for which federal funding is permitted. Some respondents wanted to ensure that potential donor(s) are either required to put their "extra" embryos up for adoption before donating them for research, or are at least offered this option. The Guidelines require that all the options available in the health care facility where treatment was sought pertaining to the use of embryos no longer needed for reproductive purposes were explained to the potential donor(s). Since not all IVF clinics offer the same services, the healthcare facility is only required to explain the options available to the donor(s) at that particular facility. Commenters asked that donor(s) be made aware of the point at which their donation decision becomes irrevocable. This is necessary because if the embryo is de-identified, it may be impossible to stop its use beyond a certain point. The NIH agrees with these comments and revised the Guidelines to require that donor(s) should have been informed that they retained the right to withdraw consent for the donation of the embryo until the embryos were actually used to derive embryonic stem cells or until information which could link the identity of the donor(s) with the embryo was no longer retained, if applicable. Medical Benefits of Donation Regarding medical benefit, respondents were concerned that the language of the Guidelines should not somehow eliminate a donor's chances of benefitting from results of stem cell research. Respondents noted that although hESCs are not currently being used clinically, it is possible that in the future such cells might be used for the medical benefit of the person donating them. The Guidelines are meant to preclude individuals from donating embryos strictly for use in treating themselves only or from donating but identifying individuals or groups they do or do not want to potentially benefit from medical intervention using their donated cells. While treatment with hESCs is one of the goals of this research, in practice, years of experimental work must still be done before such treatment might become routinely available. The Guidelines are designed to make it clear that immediate medical benefit from a donation is highly unlikely at this time. Importantly, it is critical to note that the Guidelines in no way disqualify a donor from benefitting from the medical outcomes of stem cell research and treatments that may be developed in the future. Monitoring and Enforcement Actions Respondents have expressed concern about the monitoring of funded research and the invocation of possible penalties for researchers who do not follow the Guidelines. A grantee's failure to comply with the terms and conditions of award, including confirmed instances of research misconduct, may cause the NIH to take one or more enforcement actions, depending on the severity and duration of the non-compliance. For example, the following actions may be taken by the NIH when there is a failure to comply with the terms and conditions of any award: (1) Under 45 CFR 74.14, the NIH can impose special conditions on an award, including but not limited to increased oversight/monitoring/reporting requirements for an institution, project, or investigator; and (2) under 45 CFR 74.62 the NIH may impose enforcement actions, including but not limited to withholding funds pending correction of the problem, disallowing all or part of the costs of the activity that was not in compliance, withholding further awards for the project, or suspending or terminating all or part of the funding for the project. Individuals and institutions may be debarred from eligibility for all Federal financial assistance and contracts under 2 CFR Part 376 and 48 CFR Subpart 9.4, respectively. The NIH will undertake all enforcement actions in accordance with applicable statutes, regulations, and policies. National Institutes of Health Guidelines for Research Using Human Stem CellsThese Guidelines apply to the expenditure of National Institutes of Health (NIH) funds for research using human embryonic stem cells (hESCs) and certain uses of induced pluripotent stem cells (See Section IV). The Guidelines implement Executive Order 13505. Long-standing HHS regulations for Protection of Human Subjects, 45 C.F.R. 46, Subpart A establish safeguards for individuals who are the sources of many human tissues used in research, including non-embryonic human adult stem cells and human induced pluripotent stem cells. When research involving human adult stem cells or induced pluripotent stem cells constitutes human subject research, Institutional Review Board review may be required and informed consent may need to be obtained per the requirements detailed in 45 C.F.R. 46, Subpart A. Applicants should consult http://www.hhs.gov/ohrp/humansubjects/guidance/45cfr46.html . It is also important to note that the HHS regulation, Protection of Human Subjects , 45 C.F.R. Part 46, Subpart A, may apply to certain research using hESCs. This regulation applies, among other things, to research involving individually identifiable private information about a living individual, 45 C.F.R. § 46.102(f). The HHS Office for Human Research Protections (OHRP) considers biological material, such as cells derived from human embryos, to be individually identifiable when they can be linked to specific living individuals by the investigators either directly or indirectly through coding systems. Thus, in certain circumstances, IRB review may be required, in addition to compliance with these Guidelines. Applicant institutions are urged to consult OHRP guidances at http://www.hhs.gov/ohrp/humansubjects/guidance/45cfr46.html To ensure that the greatest number of responsibly derived hESCs are eligible for research using NIH funding, these Guidelines are divided into several sections, which apply specifically to embryos donated in the U.S. and foreign countries, both before and on or after the effective date of these Guidelines. Section II (A) and (B) describe the conditions and review processes for determining hESC eligibility for NIH funds. Further information on these review processes may be found at www.NIH.gov . Sections IV and V describe research that is not eligible for NIH funding. These guidelines are based on the following principles: - Responsible research with hESCs has the potential to improve our understanding of human health and illness and discover new ways to prevent and/or treat illness.
- Individuals donating embryos for research purposes should do so freely, with voluntary and informed consent.
As directed by Executive Order 13505, the NIH shall review and update these Guidelines periodically, as appropriate. For the purpose of these Guidelines, "human embryonic stem cells (hESCs)" are cells that are derived from the inner cell mass of blastocyst stage human embryos, are capable of dividing without differentiating for a prolonged period in culture, and are known to develop into cells and tissues of the three primary germ layers. Although hESCs are derived from embryos, such stem cells are not themselves human embryos. All of the processes and procedures for review of the eligibility of hESCs will be centralized at the NIH as follows: - that were created using in vitro fertilization for reproductive purposes and were no longer needed for this purpose;
- that were donated by individuals who sought reproductive treatment (hereafter referred to as "donor(s)") and who gave voluntary written consent for the human embryos to be used for research purposes; and
- All options available in the health care facility where treatment was sought pertaining to the embryos no longer needed for reproductive purposes were explained to the individual(s) who sought reproductive treatment.
- No payments, cash or in kind, were offered for the donated embryos.
- Policies and/or procedures were in place at the health care facility where the embryos were donated that neither consenting nor refusing to donate embryos for research would affect the quality of care provided to potential donor(s).
- Decisions related to the creation of human embryos for reproductive purposes should have been made free from the influence of researchers proposing to derive or utilize hESCs in research. The attending physician responsible for reproductive clinical care and the researcher deriving and/or proposing to utilize hESCs should not have been the same person unless separation was not practicable.
- At the time of donation, consent for that donation should have been obtained from the individual(s) who had sought reproductive treatment. That is, even if potential donor(s) had given prior indication of their intent to donate to research any embryos that remained after reproductive treatment, consent for the donation for research purposes should have been given at the time of the donation.
- Donor(s) should have been informed that they retained the right to withdraw consent for the donation of the embryo until the embryos were actually used to derive embryonic stem cells or until information which could link the identity of the donor(s) with the embryo was no longer retained, if applicable.
- that the embryos would be used to derive hESCs for research;
- what would happen to the embryos in the derivation of hESCs for research;
- that hESCs derived from the embryos might be kept for many years;
- that the donation was made without any restriction or direction regarding the individual(s) who may receive medical benefit from the use of the hESCs, such as who may be the recipients of cell transplants.;
- that the research was not intended to provide direct medical benefit to the donor(s);
- that the results of research using the hESCs may have commercial potential, and that the donor(s) would not receive financial or any other benefits from any such commercial development;
- whether information that could identify the donor(s) would be available to researchers.
- By complying with Section II (A) of the Guidelines; or
The materials submitted must demonstrate that the hESCs were derived from human embryos: 1) that were created using in vitro fertilization for reproductive purposes and were no longer needed for this purpose; and 2) that were donated by donor(s) who gave voluntary written consent for the human embryos to be used for research purposes. The Working Group will review submitted materials, e.g., consent forms, written policies or other documentation, taking into account the principles articulated in Section II (A), 45 C.F.R. Part 46, Subpart A, and the following additional points to consider. That is, during the informed consent process, including written or oral communications, whether the donor(s) were: (1) informed of other available options pertaining to the use of the embryos; (2) offered any inducements for the donation of the embryos; and (3) informed about what would happen to the embryos after the donation for research. - For embryos donated outside the United States before the effective date of these Guidelines, applicants may comply with either Section II (A) or (B). For embryos donated outside of the United States on or after the effective date of the Guidelines, applicants seeking to determine eligibility for NIH research funding may submit an assurance that the hESCs fully comply with Section II (A) or submit an assurance along with supporting information, that the alternative procedural standards of the foreign country where the embryo was donated provide protections at least equivalent to those provided by Section II (A) of these Guidelines. These materials will be reviewed by the NIH ACD Working Group , which will recommend to the ACD whether such equivalence exists. Final decisions will be made by the NIH Director.
- NIH will establish a new Registry listing hESCs eligible for use in NIH funded research. All hESCs that have been reviewed and deemed eligible by the NIH in accordance with these Guidelines will be posted on the new NIH Registry.
Prior to the use of NIH funds, funding recipients should provide assurances, when endorsing applications and progress reports submitted to NIH for projects using hESCs, that the hESCs are listed on the NIH registry. This section governs research using hESCs and human induced pluripotent stem cells, i.e., human cells that are capable of dividing without differentiating for a prolonged period in culture, and are known to develop into cells and tissues of the three primary germ layers. Although the cells may come from eligible sources, the following uses of these cells are nevertheless ineligible for NIH funding, as follows: - Research in which hESCs (even if derived from embryos donated in accordance with these Guidelines) or human induced pluripotent stem cells are introduced into non-human primate blastocysts.
- Research involving the breeding of animals where the introduction of hESCs (even if derived from embryos donated in accordance with these Guidelines) or human induced pluripotent stem cells may contribute to the germ line.
- NIH funding of the derivation of stem cells from human embryos is prohibited by the annual appropriations ban on funding of human embryo research (Section 509, Omnibus Appropriations Act, 2009, Pub. L. 111-8, 3/11/09), otherwise known as the Dickey Amendment.
- Research using hESCs derived from other sources, including somatic cell nuclear transfer, parthenogenesis, and/or IVF embryos created for research purposes, is not eligible for NIH funding.
Raynard S Kington, M.D., Ph.D. Acting Director, NIH |
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Since then, research that utilizes human embryonic cells has been a widely debated, controversial ethical issue. Human embryonic cells possess the ability to become stem cells, which are used in medical research due to two significant features. First, they are unspecialized cells, meaning they can undergo cell division and renew themselves even ...
Introduction. The field of stem cell research has undergone a significant transformation with the advent of human embryonic stem cells (hESCs). Since their pioneering isolation in 1998, hESCs have been at the forefront of scientific inquiry due to their unique ability for self-renewal and pluripotency [1, 2].This comprehensive review article delves into the advancements, challenges, and ...
MS: Proponents argue that embryonic stem cell research holds great promise for understanding and curing diabetes, Parkinson's disease, spinal cord injury, and other debilitating conditions. Opponents argue that the research is unethical, because deriving the stem cells destroys the blastocyst, an unimplanted human embryo at the sixth to ...
Embryonic stem cells have immense medical potential. While both their acquisition for and use in research are fraught with controversy, arguments against their usage are rebutted by showing that embryonic stem cells are not equivalent to human lives. It is then argued that not using human embryos is unethical. Finally, an alternative to embryonic stem cells is presented.
The Ethics of Embryonic Stem Cell Research. KATRIEN DEVOLDER, 2015. Oxford, Oxford University Press 167 pp., £30 (hb) Human embryonic stem cell (hESC) research promises to enhance the way that we understand, prevent, and treat disease, potentially alleviating human suffering on a global scale. However it also involves the destruction of the ...
Sozen, B. et al. Self-assembly of embryonic and two extra-embryonic stem cell types into gastrulating embryo-like structures. Nat. Cell Biol. 20 , 979-989 (2018).
Abstract. The use of human embryos for research on embryonic stem (ES) cells is currently high on the ethical and political agenda in many countries. Despite the potential benefit of using human ES cells in the treatment of disease, their use remains controversial because of their derivation from early embryos.
regarding abortion are long and highly sophisticated; the proposed alternatives to. Embryonic Stem cell research as a whole hold promising results, but they have. drawbacks of their own as well, and these are part of the reason why other. countries outside of the U.S. are moving ahead in alternative forms of stem cell.
The Stem-Cell Debate At first glance, the case for federal funding of embryonic stem-cell research seems too obvious to need defending. Why should the government refuse to support research that hol...
This thesis is concerned with two different aspects of human embryonic stem cell research: legal and moral. These are not two distinct areas: the law cannot regulate this controversial area of science without the input of morality. There is not one moral viewpoint on the use of human embryos in scientific research and as such this thesis discusses several different moral viewpoints before ...
There is great ethical debate regarding the complex area of human embryonic stem cell research (hESCR) because in order to access the cells, destruction of the embryo is required. Therefore many different opinions regarding the moral status of the human embryo have developed. The environment of hESCR is highly politicised and one of the few scientific fields that is prohibited in some ...
Part of Nature Outlook: Stem cells. But ethical guidelines on human embryo use have halted most research into these phases of development — until now. This May, the International Society for ...
In the context of my mouse model, I combine not one but three distinct cell types to generate blastocyst-like structures: 1) wildtype embryonic stem cells (ESCs) to form the epiblast, 2) trophoblast stem cells (TSCs) to form the trophectoderm, and 3) Gata4-inducible ESCs to form the primitive endoderm.
Embryonic stem cells (ESCs) are found in the inner cell mass of the human blastocyst, an early stage of the developing embryo lasting from the 4th to 7th day after fertilization. In normal embryonic development, they disappear after the 7th day, and begin to form the three embryonic tissue layers. ESCs extracted from the inner cell mass during the blastocyst stage, however, can be cultured in ...
The Ethics Of Embryonic Stem Cell Research. Howard J. Curzer. Texas Tech University, Lubbock, TX, USA. In this article I rebut conservative objections to five phases of embryonic stem cell research. I argue that researchers using existing embryonic stem cell lines are not complicit in the past destruction of embryos because beneficiaries of ...
Embryonic Stem Cells (ESCs) ... 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 ...
Human embryonic stem cells (hESCs) are cells derived from 5-day human embryos and are self-renewing cell lines that change into any type of cell in the body, a trait called pluripotency. hESCs have almost unlimited clinical and medical research potential. Despite the great therapeutic promise of hESC research, it comes with a controversial ethical debate due to its involvement with the ...
In recent years, stem cell therapy has become a very promising and advanced scientific research topic. The development of treatment methods has evoked great expectations. This paper is a review focused on the discovery of different stem cells and the potential therapies based on these cells. The genesis of stem cells is followed by laboratory steps of controlled stem cell culturing and derivation.
Research on human embryonic stem cells, however, is controversial, given the diverse views held in our society about the moral and legal status of the early embryo. The controversy has encouraged provocative and conflicting claims both inside and outside the scientific community about the biology and biomedical potential of both adult and ...
Sydney Kossow, Creating a United Front: Harmonizing the United States Regulatory Policies Surrounding Human Embryonic Stem Cell Research, 25 SMU Sci. & Tech. L. Rev. 295 (2022) Stem cell therapy is an imperative development in science and medicine that is heavily regulated worldwide. With the potential to cure illnesses, help understand disease ...
Human embryonic stem (hES) cells are unique in their demonstrated potential to differentiate into all cell lineages. Reports by T. Wakayama et al. ("Differentiation of embryonic stem cell lines generated from adult somatic cells by nuclear transfer," 27 Apr., p. 740) and N. Lumelsky et al. ("Differentiation of embryonic stem cells to insulin-secreting structures similar to pancreatic ...
While access to adult stem cells has certainly helped answer some questions in developmental biology, embryonic stem cell studies, in my opinion, truly transformed the research area. Just like the newborn that the embryo eventually forms into, these early-stage cells have the potential to choose any path of maturation.
The Executive Order states that the Secretary of Health and Human Services, through the Director of NIH, may support and conduct responsible, scientifically worthy human stem cell research, including human embryonic stem cell (hESC) research, to the extent permitted by law. These Guidelines implement Executive Order 13505, as it pertains to ...