Preservation of stem cells

Adult stem cells (hematopoietic and mesenchymal) have demonstrated tremendous human therapeutic potential. Currently, human embryonic stem cells are used principally for understanding development and disease progression but also hold tremendous therapeutic potential. The ability to preserve stem cells is critical for their use in clinical and research applications. Preservation of cells permits the transportation of cells between sites, as well as completion of safety and quality control testing. Preservation also permits the development of a ‘manufacturing paradigm’ for cell therapies, thereby maximizing the number of products that can be produced at a given facility. in this article, we will review modes of preservation and the current status of preservation of hematopoietic, mesenchymal and human embryonic stem cells. Current and emerging issues in the area of stem cell preservation will also be described.     

Stem cells: properties and perspectives of therapeutic use

In the present work, we review the properties of some stem cell types, namely embryonic, hematopoietic and mesenchymal stem cells, which present the most significant interest for use in medicine. Stem cells are undifferentiated cells capable of both self-maintenance and differentiation into mature specialized cells. According to their origin, stem cells can be classified as embryonic and somatic ones. The first ones can be indefinitely maintained in culture, and possess the ability to differentiate into all cells of the adult organism. The second ones possess the limited capacity to differentiate and, probably, a limited proliferative potential. For therapeutic use, important but hotly debated is the plasticity of somatic stem cells, i.e. context-dependent differentiation into "non-related" cell types. It is assumed that the differentiation of the majority of stem cell types proceeds according to the principle of stepwise hierarchical maturation through the stage of intermediate rapidly proliferating progenitor cells. The use of stem cells in medicine is mostly at the preclinical stage now. Despite the fact that embryonic stem cells are highly promising as therapeutic agents, a number of circumstances substantially limits their therapeutic use in the near future. At the same time, approaches involving autotransplantation of hematopoietic or mesenchymal stem cells are beginning to be applied successfully in the clinical trials for treatment of limb ischaemia and myocardial infarction. It is clear that despite a large number of problems and unsolved questions, the use of stem cells in medicine promises a dramatic progress in the treatment of many severe diseases.     

胚胎干细胞向血液干细胞定向分化的研究进展

骨髓移植是目前治疗恶性白血病以及遗传性血液病最有效的方法之一。但是HLA相匹配的骨髓捐献者严重短缺,骨髓造血干细胞（hematopoietic stem cells,HSCs）体外培养困难,在体外修复患者骨髓造血干细胞技术不成熟,这些都大大限制了骨髓移植在临床上的应用。多能性胚胎干细胞（embryonic stem cells,ESCs）具有自我更新能力,在合适的培养条件下分化形成各种血系细胞,是造血干细胞的另一来源。在过去的二十多年里,血发生的研究是干细胞生物学中最为活跃的领域之一。小鼠及人的胚胎干细胞方面的研究最近取得了重大进展。这篇综述总结了近年来从胚胎干细胞获得造血干细胞的成就,以及在安全和技术上的障碍。胚胎干细胞诱导生成可移植性血干细胞的研究能够使我们更好地了解正常和异常造血发生的机制,同时也为造血干细胞的临床应用提供理论和实验依据。     

Mesenchymal progenitor cells localize within hematopoietic sites throughout ontogeny

Mesenchymal stem cells (MSCs) have great clinical potential for the replacement and regeneration of diseased or damaged tissue. They are especially important in the production of the hematopoietic microenvironment, which regulates the maintenance and differentiation of hematopoietic stem cells (HSCs). In the adult, MSCs and their differentiating progeny are found predominantly in the bone marrow (BM). However, it is as yet unknown in which embryonic tissues MSCs reside and whether there is a localized association of these cells within hematopoietic sites during development. To investigate the embryonic origins of these cells, we performed anatomical mapping and frequency analysis of mesenchymal progenitors at several stages of mouse ontogeny. We report here the presence of mesenchymal progenitors, with the potential to differentiate into cells of the osteogenic, adipogenic and chondrogenic lineages, in most of the sites harboring hematopoietic cells. They first appear in the aorta-gonad-mesonephros (AGM) region at the time of HSC emergence. However, at this developmental stage, their presence is independent of HSC activity. They increase numerically during development to a plateau level found in adult BM. Additionally, mesenchymal progenitors are found in the embryonic circulation. Taken together, these data show a co-localization of mesenchymal progenitor/stem cells to the major hematopoietic territories, suggesting that, as development proceeds, mesenchymal progenitors expand within these potent hematopoietic sites.     

Potential application for mesenchymal stem cells in the treatment of cardiovascular diseases

Stem cells isolated from various sources have been shown to vary in their differentiation capacity or pluripotentiality. Two groups of stem cells, embryonic and adult stem cells, may be capable of differentiating into any desired tissue or cell type, which offers hope for the development of therapeutic applications for a large number of disorders. However, major limitations with the use of embryonic stem cells for human disease have led researchers to focus on adult stem cells as therapeutic agents. Investigators have begun to examine postnatal sources of pluripotent stem cells, such as bone marrow stroma or adipose tissue, as sources of mesenchymal stem cells. The following review focuses on recent research on the use of stem cells for the treatment of cardiovascular and pulmonary diseases and the future application of mesenchymal stem cells for the treatment of a variety of cardiovascular disorders.     

Anthropological and ethical reflections on the production and use of embryonic stem cells

Abstract.  Stem cells and their potential therapeutic application have generated tremendous public interest, great enthusiasm among researchers and intense commercial interest. There are diverse sources of stem cells. According to their origin and their biological characteristics, they are classified as embryonic stem cells, germline stem cells and tissue stem cells. Until now, the most concrete therapeutic results have come from some adult tissue stem cells, with promising prospects also being offered by umbilical cord stem cells. Regarding embryonic stem cells, there is concern that they would be difficult to control in vivo . Nonetheless, many researchers are still pursuing their potential uses, convinced that they will be useful not only for study, but also for therapy, especially as a result of their high capacity for self-renewal as well as their broad potential for differentiation. This discussion which is eminently scientific in nature, and not lacking in ethical and political repercussions, will not be entered into above all regarding the allocation of available intellectual and economic resources.     

Diploid,but not haploid,human embryonic stem cells can be derived from microsurgically repaired tripronuclear human zygotes

Human embryonic stem cells have shown tremendous potential in regenerative medicine, and the recent progress in haploid embryonic stem cells provides new insights for future applications of embryonic stem cells. Disruption of normal fertilized embryos remains controversial; thus, the development of a new source for human embryonic stem cells is important for their usefulness. Here, we investigated the feasibility of haploid and diploid embryo reconstruction and embryonic stem cell derivation using microsurgically repaired tripronuclear human zygotes. Diploid and haploid zygotes were successfully reconstructed, but a large proportion of them still had a tripolar spindle assembly. The reconstructed embryos developed to the blastocyst stage, although the loss of chromosomes was observed in these zygotes. Finally, triploid and diploid human embryonic stem cells were derived from tripronuclear and reconstructed zygotes (from which only one pronucleus was removed), but haploid human embryonic stem cells were not successfully derived from the reconstructed zygotes when two pronuclei were removed. Both triploid and diploid human embryonic stem cells showed the general characteristics of human embryonic stem cells. These results indicate that the lower embryo quality resulting from abnormal spindle assembly contributed to the failure of the haploid embryonic stem cell derivation. However, the successful derivation of diploid embryonic stem cells demonstrated that microsurgical tripronuclear zygotes are an alternative source of human embryonic stem cells. In the future, improving spindle assembly will facilitate the application of triploid zygotes to the field of haploid embryonic stem cells.     

Deriving respiratory cell types from stem cells

The reported pluripotential capabilities of many human stem cell types has made them an attractive area of research, given the belief they may hold considerable therapeutic potential for treating a wide range of human diseases and injuries. Although the bulk of stem cell based research has focused on developing procedures for the treatment of pancreatic, neural, cardiovascular and haematopoietic diseases, the potential for deriving respiratory cell types from stem cells for treatment of respiratory specific diseases has also been explored. It is suggested that stem cell derivatives may be used for lung replacement/regeneration therapeutics and high though-put pharmacological screening strategies for a variety of respiratory injuries and diseases including: cystic fibrosis, chronic obstructive pulmonary disease, respiratory distress syndrome, pulmonary fibrosis and pulmonary edema. This review will explore recent progress in characterizing adult respiratory and bone marrow derived stem cells with respiratory potential as well as the endogenous mechanisms directing the homing of these cells to the diseased and injured lung. In addition, the potential for embryonic stem cell based therapies in this domain as well as the histological, anatomical and molecular aspects of respiratory development will be summarized.     

哺乳动物体细胞核移植研究进展

体细胞核移植技术已经在基础研究领域与产业化应用领域体现出了重要的价值,因而体细胞核移植技术及其相关研究已经成为了生物领域的持续性研究热点,但是围绕体细胞核移植技术仍然存在许多质疑,其中最主要的就是体细胞核移植的效率较低。尽管如此,体细胞核移植研究仍然在近年来取得了令人瞩目的成就,包括小鼠与恒河猴核移植胚胎干细胞系的建立。该文就体细胞核移植的研究历史与进展进行简要的论述,同时针对体细胞核移植研究中的细胞重编程与治疗性克隆研究中的发展与问题进行剖析,希望能够积极推动治疗性克隆的研究进展,加速核移植与干细胞技术在产业化领域中的应用。     

Comparative proteomic analysis of human mesenchymal and embryonic stem cells: Towards the definition of a mesenchymal stem cell proteomic signature

Mesenchymal stem cells (MSC) are adult multipotential progenitors which have a high potential in regenerative medicine. They can be isolated from different tissues throughout the body and their homogeneity in terms of phenotype and differentiation capacities is a real concern. To address this issue, we conducted a 2‐DE gel analysis of mesenchymal stem cells isolated from bone marrow (BM), adipose tissue, synovial membrane and umbilical vein wall. We confirmed that BM and adipose tissue derived cells were very similar, which argue for their interchangeable use for cell therapy. We also compared human mesenchymal to embryonic stem cells and showed that umbilical vein wall stem cells, a neo‐natal cell type, were closer to BM cells than to embryonic stem cells. Based on these proteomic data, we could propose a panel of proteins which were the basis for the definition of a mesenchymal stem cell proteomic signature.     

The Cannabinoid Receptor Type 2 as Mediator of Mesenchymal Stromal Cell Immunosuppressive Properties

Mesenchymal stromal cells are non-hematopoietic, multipotent progenitor cells producing cytokines, chemokines, and extracellular matrix proteins that support hematopoietic stem cell survival and engraftment, influence immune effector cell development, maturation, and function, and inhibit alloreactive T-cell responses. The immunosuppressive properties of human mesenchymal stromal cells have attracted much attention from immunologists, stem cell biologists and clinicians.Recently, the presence of the endocannabinoid system in hematopoietic and neural stem cells has been demonstrated. Endocannabinoids, mainly acting through the cannabinoid receptor subtype 2, are able to modulate cytokine release and to act as immunosuppressant when added to activated T lymphocytes.In the present study, we have investigated, through a multidisciplinary approach, the involvement of the endocannabinoids in migration, viability and cytokine release of human mesenchymal stromal cells.We show, for the first time, that cultures of human mesenchymal stromal cells express all of the components of the endocannabinoid system, suggesting a potential role for the cannabinoid CB2 receptor as a mediator of anti-inflammatory properties of human mesenchymal stromal cells, as well as of their survival pathways and their capability to home and migrate towards endocannabinoid sources.     

Bioreactor development for stem cell expansion and controlled differentiation

Widespread use of embryonic and adult stem cells for therapeutic applications will require reproducible production of large numbers of well-characterized cells under well-controlled conditions in bioreactors. During the past two years, substantial progress has been made towards this goal. Human mesenchymal stem cells expanded in perfused scaffolds retained multi-lineage potential. Mouse neural stem cells were expanded as aggregates in serum-free medium for 44 days in stirred bioreactors. Mouse embryonic stem cells expanded as aggregates and on microcarriers in stirred vessels retained expression of stem cell markers and could form embryoid bodies. Embryoid body formation from dissociated mouse embryonic stem cells, followed by embryoid body expansion and directed differentiation, was scaled up to gas-sparged, 2-l instrumented bioreactors with pH and oxygen control.     

核移植胚胎干细胞的研究及其应用前景

随着核移植技术和干细胞技术的逐渐成熟,目前已获得牛、小鼠核移植胚胎干细胞,以及人 - 兔异种间核移植胚胎干细胞,这些细胞在体外可分化成多种细胞形态 . 已经进行的实验性动物克隆性治疗,显示了诱人的潜力,但人核移植胚胎干细胞研究还面临着许多问题,如建系效率低、卵母细胞来源有限以及伦理学和安全性问题等 . 长远地看,随着克隆效率的提高,在道德与法律之间达成共识,核移植胚胎干细胞必将造福人类 .     

Membrane glycolipids in stem cells

Stem cells, such as embryonic stem cells, hematopoietic stem cells, neural stem cells, mesenchymal stem cells, and very small embryonic-like stem cells, are undifferentiated cells that are endowed with a high potential for proliferation and the capacity for self-renewal with retention of pluri/multipotency to differentiate into their progenies. Recently, studies regarding the biological functions of glycolipids and cell surface microdomains (caveolae, lipid rafts, or glycolipid-enriched microdomains) in stem cells are emerging. In this review, we introduce the expression patterns of glycolipids and the functional roles of cell surface microdomains in stem cells.     

Epigenetic stability of embryonic stem cells and developmental potential

Recent studies highlight the tremendous potential of human embryonic stem (ES) cells and their derivatives as therapeutic tools for degenerative diseases. However, derivation and culture of ES cells can induce epigenetic alterations, which can have long lasting effects on gene expression and phenotype. Research on human and mouse stem cells indicates that developmental, cancer-related genes, and genes regulated by genomic imprinting are particularly susceptible to changes in DNA methylation. Together with the occurrence of genetic alterations, epigenetic instability needs to be monitored when considering human stem cells for therapeutic and technological purposes. Here, we discuss the maintenance of epigenetic information in cultured stem cells and embryos and how this influences their developmental potential.