The stem cell niche in regenerative medicine

Stem cells are fundamental units for achieving regenerative therapies, which leads naturally to a theoretical and experimental focus on these cells for therapeutic screening and intervention. A growing body of data in many tissue systems indicates that stem cell function is critically influenced by extrinsic signals derived from the microenvironment, or "niche." In this vein, the stem cell niche represents a significant, and largely untapped, entry point for therapeutic modulation of stem cell behavior. This Perspective will discuss how the niche influences stem cells in homeostasis, in the progression of degenerative and malignant diseases, and in therapeutic strategies for tissue repair.     

脂肪干细胞定向分化的影响因素

脂肪干细胞是一类由脂肪组织分离出来的间充质干细胞,具有自我更新和多向分化潜能,可长期自我更新并在特定的条件下分化形成多种终末细胞。本文总结近年来关于脂肪干细胞定向分化的研究成果、介绍脂肪干细胞的分离培养技术、影响脂肪干细胞成脂成骨定向分化能力的主要因素,以及在各种终末分化细胞之间"转分化"的作用条件,从而为寻找治疗肥胖的新靶点以及骨组织工程学提供新思路。     

Brain tumour stem cells: implications for cancer therapy and regenerative medicine

The cancer relapse and mortality rate suggest that current therapies do not eradicate all malignant cells. Currently, it is accepted that tumorigenesis and organogenesis are similar in many respects, as for example, homeostasis is governed by a distinct sub-population of stem cells in both situations. There is increasing evidence that many types of cancer contain their own stem cells: cancer stem cells (CSC), which are characterized by their self-renewing capacity and differentiation ability. The investigation of solid tumour stem cells has gained momentum particularly in the area of brain tumours. Gliomas are the most common type of primary brain tumours. Nearly two-thirds of gliomas are highly malignant lesions with fast progression and unfortunate prognosis. Despite recent advances, two-year survival for glioblastoma (GBM) with optimal therapy is less than 30%. Even among patients with low-grade gliomas that confer a relatively good prognosis, treatment is almost never curative. Recent studies have demonstrated the existence of a small fraction of glioma cells endowed with features of primitive neural progenitor cells and a tumour-initiating function. In general, this fraction is characterized for forming neurospheres, being endowed with drug resistance properties and often, we can isolate some of them using sorting methods with specific antibodies. The molecular characterization of these stem populations will be critical to developing an effective therapy for these tumours with very dismal prognosis. To achieve this aim, the development of a mouse model which recapitulates the nature of these tumours is essential. This review will focus on glioma stem cell knowledge and discuss future implications in brain cancer therapy and regenerative medicine.     

Renal stem cell reprogramming: Prospects in regenerative medicine

Stem cell therapy is a promising future enterprise for renal replacement in patients with acute and chronic kidney disease, conditions which affect millions worldwide and currently require patients to undergo lifelong medical treatments through dialysis and/or organ transplant. Reprogramming differentiated renal cells harvested from the patient back into a pluripotent state would decrease the risk of tissue rejection and provide a virtually unlimited supply of cells for regenerative medicine treatments, making it an exciting area of current research in nephrology. Among the major hurdles that need to be overcome before stem cell therapy for the kidney can be applied in a clinical setting are ensuring the fidelity and relative safety of the reprogrammed cells, as well as achieving feasible efficiency in the reprogramming processes that are utilized. Further, improved knowledge about the genetic control of renal lineage development is vital to identifying predictable and efficient reprogramming approaches, such as the expression of key modulators or the regulation of geneactivity through small molecule mimetics. Here, we discuss several recent advances in induced pluripotent stem cell technologies. We also explore strategies that have been successful in renal progenitor generation, and explore what these methods might mean for the development of cell-based regenerative therapies for kidney disease.     

Derivation and application of pluripotent stem cells for regenerative medicine

Pluripotent stem cells (PSCs) are cells that can differentiate into any type of cells in the body, therefore have valuable promise in regenerative medicine of cell replacement therapies and tissue/organ engineering. PSCs can be derived either from early embryos or directly from somatic cells by epigenetic reprogramming that result in customized cells from patients. Here we summarize the methods of deriving PSCs, the various types of PSCs generated with different status, and their versatile applications in both clinical and embryonic development studies. We also discuss an intriguing potential application of PSCs in constructing tissues/organs in large animals by interspecies chimerism. All these emerging findings are likely to contribute to the breakthroughs in biological research and the prosperous prospects of regenerative medicine.     

Orchestrating stem cell fate: Novel tools for regenerative medicine

Mesenchymal stem cells are undifferentiated cells able to acquire different phenotypes under specific stimuli. In vitro manipulation of these cells is focused on understanding stem cell behavior, proliferation and pluripotency. Latest advances in the field of stem cells concern epigenetics and its role in maintaining self-renewal and differentiation capabilities. Chemical and physical stimuli can modulate cell commitment, acting on gene expression of Oct-4, Sox-2 and Nanog,the main stemness markers, and tissue-lineage specific genes. This activation or repression is related to the activity of chromatin-remodeling factors and epigenetic regulators, new targets of many cell therapies. The aim of this review is to afford a view of the current state of in vitro and in vivo stem cell applications,highlighting the strategies used to influence stem cell commitment for current and future cell therapies. Identifying the molecular mechanisms controlling stem cell fate could open up novel strategies for tissue repairing processes and other clinical applications.     

Embryonic stem cells: hepatic differentiation and regenerative medicine for the treatment of liver disease

Hepatocyte transplantation is considered a potential treatment for liver diseases and a bridge for patients awaiting liver transplantation, but its application has been hampered by a limited supply of hepatocytes. Embryonic stem (ES) cells established from early mouse and human embryos are pluripotent, and proliferate indefinitely in an undifferentiated state in vitro. Since differentiation from ES cells seems to recapitulate early embryonic development, if hepatocytes could be efficiently generated in vitro, ES cells might become a source of transplantable hepatocytes for cell replacement therapy. Hepatocytes have been generated from ES cells in vitro, and the hepatocytes differentiated from ES cells have been found to express many hepatocyte-related genes and perform hepatic functions. However, it remains unclear whether the hepatocytes differentiated from ES cells are derived from definitive endoderm or primitive endoderm. Because visceral endoderm, which expresses many hepatocyte-related genes, is derived from primitive endoderm and is fated to form extraembryonic yolk sac tissues, not to form hepatocytes, ES cells must be directed to a definitive endoderm lineage in vitro. This article discusses the differentiation of ES cells into hepatocytes in vitro in comparison with early embryogenesis, and describes the efficacy of ES cell-derived hepatocyte transplantation.     

The effect of quercetin on hepatic differentiation of human adipose-derived mesenchymal stem cells

Human adipose-derived mesenchymal stem cells (MSCs) can be stimulated to differentiate into hepatic cells. MSC differentiation was induced by fibroblast growth factor-4, hepatocyte growth factor, oncostatin M, and dexamethasone. The influence of quercetin on MSC hepatic differentiation in culture was assayed, and 1 or 10 μmole/L quercetin added into the induction medium enhanced the manifestation of MSC hepatic differentiation. Urea secretion, cytokeratin 19 expression, and α-fetoprotein synthesis were increased. Quercetin modulated CYP1A–cytochrome P450 activity in the differentiated cells. MSCs differentiated in the presence of quercetin exhibited higher viability and resistance to oxidative stress.     

Surface functionalization of nanobiomaterials for application in stem cell culture,tissue engineering,and regenerative medicine

Stem cell‐based approaches offer great application potential in tissue engineering and regenerative medicine owing to their ability of sensing the microenvironment and respond accordingly (dynamic behavior). Recently, the combination of nanobiomaterials with stem cells has paved a great way for further exploration. Nanobiomaterials with engineered surfaces could mimic the native microenvironment to which the seeded stem cells could adhere and migrate. Surface functionalized nanobiomaterial‐based scaffolds could then be used to regulate or control the cellular functions to culture stem cells and regenerate damaged tissues or organs. Therefore, controlling the interactions between nanobiomaterials and stem cells is a critical factor. However, surface functionalization or modification techniques has provided an alternative approach for tailoring the nanobiomaterials surface in accordance to the physiological surrounding of a living cells; thereby, enhancing the structural and functional properties of the engineered tissues and organs. Currently, there are a variety of methods and technologies available to modify the surface of biomaterials according to the specific cell or tissue properties to be regenerated. This review highlights the trends in surface modification techniques for nanobiomaterials and the biological relevance in stem cell‐based tissue engineering and regenerative medicine. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:554–567, 2016     

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.     

ARPE-19 conditioned medium promotes neural differentiation of adipose-derived mesenchymal stem cells

BACKGROUNDAdipose-derived stem cells (ASCs) have been increasingly explored for cell-based medicine because of their numerous advantages in terms of easy availability, high proliferation rate, multipotent differentiation ability and low immunogenicity. In this respect, they have been widely investigated in the last two decades to develop therapeutic strategies for a variety of human pathologies including eye disease. In ocular diseases involving the retina, various cell types may be affected, such as Müller cells, astrocytes, photoreceptors and retinal pigment epithelium (RPE), which plays a fundamental role in the homeostasis of retinal tissue, by secreting a variety of growth factors that support retinal cells.AIMTo test ASC neural differentiation using conditioned medium (CM) from an RPE cell line (ARPE-19).METHODSASCs were isolated from adipose tissue, harvested from the subcutaneous region of healthy donors undergoing liposuction procedures. Four ASC culture conditions were investigated: ASCs cultured in basal Dulbecco''s Modified Eagle Medium (DMEM); ASCs cultured in serum-free DMEM; ASCs cultured in serum-free DMEM/F12; and ASCs cultured in a CM from ARPE-19, a spontaneously arising cell line with a normal karyotype derived from a human RPE. Cell proliferation rate and viability were assessed by crystal violet and MTT assays at 1, 4 and 8 d of culture. At the same time points, ASC neural differentiation was evaluated by immunocytochemistry and western blot analysis for typical neuronal and glial markers: Nestin, neuronal specific enolase (NSE), protein gene product (PGP) 9.5, and glial fibrillary acidic protein (GFAP).RESULTSDepending on the culture medium, ASC proliferation rate and viability showed some significant differences. Overall, less dense populations were observed in serum-free cultures, except for ASCs cultured in ARPE-19 serum-free CM. Moreover, a different cell morphology was seen in these cultures after 8 d of treatment, with more elongated cells, often showing cytoplasmic ramifications. Immunofluorescence results and western blot analysis were indicative of ASC neural differentiation. In fact, basal levels of neural markers detected under control conditions significantly increased when cells were cultured in ARPE-19 CM. Specifically, neural marker overexpression was more marked at 8 d. The most evident increase was observed for NSE and GFAP, a modest increase was observed for nestin, and less relevant changes were observed for PGP9.5. CONCLUSIONThe presence of growth factors produced by ARPE-19 cells in tissue culture induces ASCs to express neural differentiation markers typical of the neuronal and glial cells of the retina.     

The effect of exogenous histone H1 on rat adipose-derived stem cell proliferation, migration, and osteogenic differentiation in vitro

Adipose-derived stem cells (ASCs) are of great interest for the development of novel cell therapies due to their ease of isolation and expansion, immunosuppressive activity, and multilineage differentiation potential. However, the mechanisms underlying the therapeutic potential of ASCs remain to be elucidated. Others and we have shown that nuclear proteins such as histone H1 and high mobility group box 1 (HMGB1) play important roles in the maturation of dendritic cells (DCs). Furthermore, we previously demonstrated translocation of histone H1 from the nucleus to the cytoplasm and activation of mitogen-activated protein kinases (MAPKs) in DCs. In the present study, we confirmed that histone H1 does not alter the immunophenotype and immunosuppression potential of ASCs, but that histone H1 enhanced wound healing and increased interleukin (IL)-6 expression. Moreover, histone H1 treated-ASCs showed up-regulation of MAPKs extracellular-regulated kinase 1/2 (ERK1/2) and sequential NF-κB translocation. Finally, we found that culture in differentiation media supplemented with histone H1 enhanced ASC osteogenesis. In contrast, inhibition of histone H1 by small interfering RNA (siRNA) reduced osteogenic differentiation markers including ALP. These results suggest that histone H1 may be useful for induction of mesenchymal stem cells in tissue engineering and future potential ASC therapies.     

Nuclear reprogramming: a key to stem cell function in regenerative medicine

The goal of regenerative medicine is to restore form and function to damaged tissues. One potential therapeutic approach involves the use of autologous cells derived from the bone marrow (bone marrow-derived cells, BMDCs). Advances in nuclear transplantation, experimental heterokaryon formation and the observed plasticity of gene expression and phenotype reported in multiple phyla provide evidence for nuclear plasticity. Recent observations have extended these findings to show that endogenous cells within the bone marrow have the capacity to incorporate into defective tissues and be reprogrammed. Irrespective of the mechanism, the potential for new gene expression patterns by BMDCs in recipient tissues holds promise for developing cellular therapies for both proliferative and post-mitotic tissues.     

Accelerated and safe expansion of human mesenchymal stromal cells in animal serum-free medium for transplantation and regenerative medicine

Human bone marrow mesenchymal stromal cells (hMSC) are currently investigated for a variety of therapeutic applications. However, most expansion protocols still use fetal calf serum (FCS) as growth factor supplement which is a potential source of undesired xenogeneic pathogens. We established an expansion protocol for hMSC based on the use of GMP-produced basic medium LP02 supplemented with 5% of platelet lysate (PL) obtained from human thrombocyte concentrates. Compared to FCS-supplemented culture conditions, we found a significant increase in both colony forming unit-fibroblast (CFU-F) as well as cumulative cell numbers after expansion. This accelerated growth is optimized by pooling of at least 10 thrombocyte concentrates. A minimal requirement is the use of 5% of PL with an optimal platelet concentration of 1.5 x 10(9)/ml, and centrifugation of thawed lysate at high speed. Cells expanded by this protocol meet all criteria for mesenchymal stromal cells (MSCs), e.g. plastic adherence, spindle-shaped morphology, surface marker expression, lack of hematopoietic markers, and differentiation capability into three mesenchymal lineages. MSC at passage 6 were cytogenetically normal and retained their immune-privileged potential by suppressing allogeneic reaction of T-cells. Additionally, gene expression profiles show increased mRNA levels of genes involved in cell cycle and DNA replication and downregulation of developmental and differentiation genes, supporting the observation of increased MSC-expansion in PL-supplemented medium. In summary, we have established a GMP-compatible protocol for safe and accelerated expansion of hMSC to be used in cell and tissue therapy.     

Effects of GSK3 inhibitors on in vitro expansion and differentiation of human adipose-derived stem cells into adipocytes

Background  

Multipotent stem cells exist within adipose tissue throughout life. An abnormal recruitment of these adipose precursor cells could participate to hyperplasia of adipose tissue observed in severe obesity or to hypoplasia of adipose tissue observed in lipodystrophy. Therefore, pharmacological molecules that control the pool of stem cells in adipose tissue are of great interest. Glycogen Synthase Kinase (GSK) 3 has been previously described as involved in differentiation of preadipose cells and might be a potential therapeutic target to modulate proliferation and differentiation of adipocyte precursors. However, the impact of GSK3 inhibition on human adipose-derived stem cells remained to be investigated. The aim of this study was to investigate GSK3 as a possible target for pharmacological inhibition of stem cell adipogenesis. To reach this goal, we studied the effects of pharmacological inhibitors of GSK3, i.e. lithium chloride (LiCl) and BIO on proliferation and adipocyte differentiation of multipotent stem cells derived from human adipose tissue.     

The umbilical cord: a rich and ethical stem cell source to advance regenerative medicine

Science and medicine place a lot of hope in the development of stem cell research and regenerative medicine. This review will define the concept of regenerative medicine and focus on an abundant stem cell source - neonatal tissues such as the umbilical cord. Umbilical cord blood has been used clinically for over 20 years as a cell source for haematopoietic stem cell transplantation. Beyond this, cord blood and umbilical cord-derived stem cells have demonstrated potential for pluripotent lineage differentiation (liver, pancreatic, neural tissues and more) in vitro and in vivo. This promising research has opened up a new era for utilization of neonatal stem cells, now used beyond haematology in clinical trials for autoimmune disorders, cerebral palsy or type I diabetes.