Stem Cells and Cloning-Advances and Applications最新文献

Among the various strategies providing a cure for illness, cell-based therapies have caught the attention of the world with the advent of the "stem cell" era. Our inherent understanding indicates that stem cells have been in existence since the birth of multicellular organisms. However, the formal discovery of stem cells in the last century, followed by their intricate and extensive analysis, has led to clinical and translational efforts with the aim of using them in the treatment of conditions which don't have a definitive therapeutic strategy, has fueled our interest and expectations. Technological advances in our ability to study their cellular components in depth, along with surface markers and other finer constituents, that were unknown until last century, have improved our understanding, leading to several novel applications. This has created a need to establish guidelines, and in that process, there are expressed understandings and views which describe cell therapy along lines similar to that of biologic products, drugs, and devices. However, the age-old wisdom of using cells as tools for curing illness should not be misled by recent knowledge, to make cell therapy using highly complex stem cells equal to factory-synthesized and reproducible chemical compounds, drugs, or devices. This article analyses the differences between these two entities from various perspectives.

Stem cell-based therapies hold promise for regenerating the myocardium after injury. Recent data obtained from phase I clinical trials using endogenous cardiovascular progenitors isolated directly from the heart suggest that cell-based treatment for heart patients using stem cells that reside in the heart provides significant functional benefit and an improvement in patient outcome. Methods to achieve improved engraftment and regeneration may extend this therapeutic benefit. Endogenous cardiovascular progenitors have been tested extensively in small animals to identify cells that improve cardiac function after myocardial infarction. However, the relative lack of large animal models impedes translation into clinical practice. This review will exclusively focus on the latest research pertaining to humans and large animals, including both endogenous and induced sources of cardiovascular progenitors.

Throughout their lifetime, an individual may sustain many injuries and recover spontaneously over a period of time, without even realizing the injury in the first place. Wound healing occurs due to a proliferation of stem cells capable of restoring the injured tissue. The ability of adult stem cells to repair tissue is dependent upon the intrinsic ability of tissues to proliferate. The amazing capacity of embryonic stem cells to give rise to virtually any type of tissue has intensified the search for similar cell lineage in adults to treat various diseases including cardiovascular diseases. The ability to convert adult stem cells into pluripotent cells that resemble embryonic cells, and to transplant those in the desired organ for regenerative therapy is very attractive, and may offer the possibility of treating harmful disease-causing mutations. The race is on to find the best cells for treatment of cardiovascular disease. There is a need for the ideal stem cell, delivery strategies, myocardial retention, and time of administration in the ideal patient population. There are multiple modes of stem cell delivery to the heart with different cell retention rates that vary depending upon method and site of injection, such as intra coronary, intramyocardial or via coronary sinus. While there are crucial issues such as retention of stem cells, microvascular plugging, biodistribution, homing to myocardium, and various proapoptotic factors in the ischemic myocardium, the regenerative potential of stem cells offers an enormous impact on clinical applications in the management of cardiovascular diseases.

Stem cell transplantation for spinal cord injury (SCI) along with new pharmacotherapy research offers the potential to restore function and ease the associated social and economic burden in the years ahead. Various sources of stem cells have been used in the treatment of SCI, but the most convincing results have been obtained with neural progenitor cells in preclinical models. Although the use of cell-based transplantation strategies for the repair of chronic SCI remains the long sought after holy grail, these approaches have been to date the most successful when applied in the subacute phase of injury. Application of cell-based strategies for the repair and regeneration of the chronically injured spinal cord will require a combinational strategy that may need to include approaches to overcome the effects of the glial scar, inhibitory molecules, and use of tissue engineering strategies to bridge the lesion. Nonetheless, cell transplantation strategies are promising, and it is anticipated that the Phase I clinical trials of some form of neural stem cell-based approach in SCI will commence very soon.

Inadequate blood supply to tissues caused by obstruction of arterioles and/or capillaries results in ischemic injuries - these injuries can range from mild (eg, leg ischemia) to severe conditions (eg, myocardial infarction, stroke). Surgical and/or endovascular procedures provide cutting-edge treatment for patients with vascular disorders; however, a high percentage of patients are currently not treatable, owing to high operative risk or unfavorable vascular involvement. Therapeutic angiogenesis has recently emerged as a promising new therapy, promoting the formation of new blood vessels by the introduction of bone marrow-derived stem and progenitor cells. These cells participate in the development of new blood vessels, the enlargement of existing blood vessels, and sprouting new capillaries from existing blood vessels, providing evidence of the therapeutic utility of these cells in ischemic tissues. In this review, the authors describe peripheral arterial disease, an ischemic condition affecting the lower extremities, summarizing different aspects of vascular regeneration and discussing which and how stem cells restore the blood flow. The authors also present an overview of encouraging results from early-phase clinical trials using stem cells to treat peripheral arterial disease. The authors believe that additional research initiatives should be undertaken to better identify the nature of stem cells and that an intensive cooperation between laboratory and clinical investigators is needed to optimize the design of cell therapy trials and to maximize their scientific rigor. Only this will allow the results of these investigations to develop best clinical practices. Additionally, although a number of stem cell therapies exist, many treatments are performed outside international and national regulations and many clinical trials have been not registered on databases such as ClinicalTrials.gov or EudraCT. Therefore, more rigorous clinical trials are required to confirm the first hopeful results and to address the challenging issues.

A number of clinical observations have indicated that the regenerative potential and overall function of the epidermis is modified with age. The epidermis becomes thinner, repairs itself less efficiently after wounding, and presents modified barrier function recovery. In addition, the dermal papillae fatten out with increasing age, suggesting a modification in the interaction between epidermal and dermal compartments. As the epidermal regenerative capacity is dependent upon stem and progenitor cell function, it is naturally of interest to identify and understand age-related changes in these particular keratinocyte populations. Previous studies have indicated that the number of stem cells does not decrease with age in mouse models but little solid evidence is currently available concerning human skin. The objective of this study was to evaluate the clonogenic potential of keratinocyte populations isolated from the epidermis of over 50 human donors ranging from 18 to 71 years old. The data indicate that the number of epidermal cells presenting high regenerative potential does not dramatically decline with age in human skin. The authors believe that changes in the microenvironment controlling epidermal basal cell activity are more likely to explain the differences in epidermal function observed with increasing age.

Somatic tissue engraftment was studied in BXSB mice treated with mesenchymal stem cell transplantation. Hosts were conditioned with nonlethal radiation prior to introducing donor cells from major histocompatibility complex-matched green fluorescent protein transgenic mice. Transplant protocols differed for route of injection, ie, intravenous (i.v.) versus intraperitoneal (i.p.), and source of mesenchymal stem cells, ie, unfractionated bone marrow cells, ex vivo expanded mesenchymal stem cells, or bone chips. Tissue chimerism was determined after short (10-12 weeks) or long (62 weeks) posttransplant follow-up by immunohistochemistry for green fluorescent protein. Engraftment of endothelial cells was seen in several organs including liver sinusoidal cells in i.v. treated mice with ex vivo expanded mesenchymal stem cells or with unfractionated bone marrow cells. Periportal engraftment of liver hepatocytes, but not engraftment of endothelial cells, was found in mice injected i.p. with bone chips. Engraftment of adipocytes was a common denominator in both i.v. and i.p. routes and occurred during early phases post-transplant. Disease control was more robust in mice that received both i.v. bone marrow and i.p. bone chips compared to mice that received i.v. bone marrow alone. Thus, the data support potential use of mesenchymal stem cell transplant for treatment of severe lupus. Future studies are needed to optimize transplant conditions and tailor protocols that may in part be guided by fat and endothelial biomarkers. Furthermore, the role of liver chimerism in disease control and the nature of cellular communication among donor hematopoietic and mesenchymal stem cells in a chimeric host merit further investigation.

In vitro, in vivo animal, and human clinical data show a broad field of application for mesenchymal stem cells (MSCs). There is overwhelming evidence of the usefulness of MSCs in regenerative medicine, tissue engineering, and immune therapy. At present, there are a significant number of clinical trials exploring the use of MSCs for the treatment of various diseases, including myocardial infarction and stroke, in which oxygen suppression causes widespread cell death, and others with clear involvement of the immune system, such as graft-versus-host disease, Crohn's disease, and diabetes. With no less impact, MSCs have been used as cell therapy to treat defects in bone and cartilage and to help in wound healing, or in combination with biomaterials in tissue engineering development. Among the MSCs, allogeneic MSCs have been associated with a regenerative capacity due to their unique immune modulatory properties. Their immunosuppressive capability without evidence of immunosuppressive toxicity at a global level define their application in the treatment of diseases with a pathogenesis involving uncontrolled activity of the immune system. Until now, the limitation in the number of totally characterized autologous MSCs available represents a major obstacle to their use for adult stem cell therapy. The use of premanufactured allogeneic MSCs from controlled donors under optimal conditions and their application in highly standardized clinical trials would lead to a better understanding of their real applications and reduce the time to clinical translation.

The identification of normal and cancerous stem cells and the recent advances made in isolation and culture of stem cells have rapidly gained attention in the field of drug discovery and regenerative medicine. The prospect of performing screens aimed at proliferation, directed differentiation, and toxicity and efficacy studies using stem cells offers a reliable platform for the drug discovery process. Advances made in the generation of induced pluripotent stem cells from normal or diseased tissue serves as a platform to perform drug screens aimed at developing cell-based therapies against conditions like Parkinson's disease and diabetes. This review discusses the application of stem cells and cancer stem cells in drug screening and their role in complementing, reducing, and replacing animal testing. In addition to this, target identification and major advances in the field of personalized medicine using induced pluripotent cells are also discussed.

Mesenchymal stem cells (MSCs) have become an attractive tool for tissue engineering and targets in clinical transplantation due to their regeneration potential and immuno-suppressive capacity. Although MSCs derived from bone marrow are the most widely used, their harvest requires an invasive procedure. The umbilical cord, which is discarded at birth, can provide an inexhaustible source of stem cells for therapy. The Wharton's jelly-derived MSCs (WJ-MSCs), from the umbilical cord, have been shown to have faster proliferation rates and greater expansion capability compared with adult MSCs. The standard isolation and in vitro culture protocol for WJ-MSCs utilizes fetal bovine serum (FBS) or calf serum as a nutrient supplement. However, FBS raises potential safety concerns such as transmission of viral/prion disease and may initiate xenogeneic immune reactions against bovine antigens. Therefore, for therapeutic applications, there is an urgent requirement to establish an alternative nutrient supplement which would favor cell proliferation, retain MSC characteristics, and prove safe in human subjects. In the present study, we isolated and expanded WJ-MSCs in 5% pooled, allogeneic human serum (HS) supplemented with 2 ng/mL of basic fibroblast growth factor. For cell dissociation, porcine trypsin was replaced with TrypLE, a recombinant enzyme, and a protease-free protocol was adapted for isolation of MSCs from WJ. We determined their growth kinetics, in vitro differentiation potential, surface marker expression, and colony-forming unit potential and compared them against standard WJ-MSC cultures expanded in 10% FBS. All these parameters matched quite well between the two MSC populations. To test whether there is any alteration in gene expression on switching from FBS to HS, we analyzed a panel of stem cell and early lineage markers using Taqman® low density array. No significant deviation in gene expression was observed between the two populations. Thus we established an efficient, complete xeno-free protocol for propagation of human WJ-MSCs.