Journal of Stem Cells & Regenerative Medicine最新文献

All kinds of cells from all kinds of organisms (i.e., animals, plants, fungi and bacteria) are covered by a dense layer of glycans. The origin of glycans or carbohydrates is not known^[1], however, the above fact implies that they are widely and closely associated with various biological phenomena based on cellular communications, which include development, differentiation, morphogenesis, carcinogenesis, immunity and infection. It is also notable that glycoproteins, one of existing forms of glycans (i.e., glycoconjugates) are generally synthesized in lumen sites of endoplasmic reticulum and the following Golgi apparatus, distinct from cytoplasmic proteins, which are not subjected to glycosylation, a major event of posttranslational modifications. In fact, glycan structures largely depend on a series of (e.g., >200 in human) glycol-genes, which are defined as genes involved in glycan synthesis (e.g., glycosyltransferases, sulfotransferases, nucleotide sugar transporters), of which expressions differ under different conditions. Because expression of each glycol-gene differs in different cell types (e.g., biological origin, tissue) and states (e.g., developmental stage, malignancy), glycans can be a good marker for cell typing (e.g., SSEA-1) and serum diagnosis (e.g., cancer biomarker such as CA19-9). However, glycan preparation as well as its analysis and total understanding are much more difficult compared with other major disciplines like genomics and proteomics. As a result, most of nonglycoscientists tend to hesitate glycomics, i.e., "glycophobia". Nevertheless, glycoscience is a very important field of life science, particularly in the future, without which many remaining issues will not be solved. In this plenary lecture, a novel approach to glycan profiling^[2] and its applications to biomarker investigation and regenerative medicine^[3] will be described.

Thrombolysis (rt-PA) is the only United States Food and Drug Administration (FDA) approved drug currently available. Unfortunately, its effect has been limited by the narrow therapeutic time window. Human cord blood mononuclear cells (cbMNC) is a promising treatment for ischemic stroke by forming collateral and neo-vascularization where it is one of the important factors that contribute to cell repair. Therefore, evaluation of neo-vascularization in sub-acute stroke may be beneficial for recovery. One group for healthy rat and three groups (n=6 per group) of male wistar rats have undergone permanent middle cerebral artery occlusion (MCAO). Transplantation 1x10⁶ cells/kg of human cbMNC intra-arterially (IA) and intra-venously (IV) were administered after 7 days. Behavioural tests were performed before MCAO, 1 week after MCAO and at 3,9 and 14 days after cbMNC transplantation. Beta III tubulin protein (TUJ1), glial fibrillary acidic protein (GFAP) and vascular endothelial growth factor (VEGF) antibody marker were evaluated. Spontaneous activity of transplanted rats by cbMNC have significantly improved compared to placebo group (p<0.05). Angiogenesis in IA group showed significant difference (P<0.001) when compared to IV and placebo respectively. The existence of neovascularization in the transplanted rats of cbMNC provide hope in accelerating repairment of the neuronal cells and functional outcome.

We have evaluated the cardiomyogenic potential of clonal populations of human bone marrow mesenchymal stem cells (BM-MSC). Four rapidly proliferating clones of BM-MSC were obtained from the BM of a healthy donor which were then treated with 5-azacytidine and evaluated for the expression of GATA-4, NKx-2.5, FOG-2, TDGF-1, β-MHC, MEF2D and NPPA genes and cTnT, Desmin and β-MHC proteins. Of the four clones (i) Clone-1 had high expression of GATA-4 (1.89 fold (p<0.05), Nkx2.5 (2.29 fold; p<0.05), FOG2 (2.76 fold; p<0.05), TDGF1 (6.97 fold, p<0.005), βMHC (10.22 fold; p<0.005), MEF-2D (1.91 fold; p<0.005) and NPPA (1.65 fold; p<0.005); (ii) clone-2 had up-regulation of Nkx2.5 (1.98 fold; p<0.05) but down-regulation of rest of the genes; (iii) clone-3 had up-regulation of Nkx2.5 (2.11 fold; p<0.05), TDGF1 (1.88 fold; p<0.05), MEF-2D (1.30 fold; p<0.05) and NPPA (1.21 fold; p<0.05), down regulation of GATA-4 and Fog-2 but no change in βMHC gene; and (iv) clone-4 had up-regulation of MEF-2D (1.17 fold; p<0.05) and down regulation of GATA-4, Nkx2.5 but no change in other genes compared to untreated cells of the clones. At the protein level, clone-1 expressed cTnT, Desmin, and βMHC; clone-2 Desmin only while clones-3 and 4 each expressed cTnT, Desmin, and βMHC. Our data shows that BM-MSC are a heterogenous population of stem cells with sub-populations exhibiting a marked difference in the expression of cardiac markers both at gene and protein levels. This highlights that administering selected sub-populations of BM-MSC with a cardiomyogenic potential may be more efficacious than whole population of cells for cardiac regeneration.

Research on stem cells is one of the fastest growing areas of regenerative medicine that paves the way for a comprehensive solution to cell therapy. Today, stem cells are precious assets for generating different types of cells derived from either natural embryonic stem (ES) cells or induced pluripotent stem (iPS) cells. The iPS technology can revolutionize the future of clinics by offering personalized medicine, which will provide the future treatment for curing untreatable diseases. Although iPS cell therapy is now at its infancy, promising research has motivated scientists to pursue this therapeutic approach. In this article, we provide information regarding similarities and differences between ES and iPS cells, and focus on the non-integrating methods of iPS generation via RNA molecules, especially microRNAs with an emphasis on the elucidation of their role and importance in pluripotency.

Stem cell research for treating or curing ischemic heart disease has, till date, culminated in three basic approaches: the use of induced pluripotent stem cell (iPSC) technology; reprogramming cardiac fibroblasts; and cardiovascular progenitor cell regeneration. As each approach has been shown to have its advantages and disadvantages, exploiting the advantages while minimizing the disadvantages has been a challenge. Using human germline pluripotent stem cells (hgPSCs) along with a modified version of a relatively novel cell-expansion culture methodology to induce quick, indefinite expansion of normally slow growing hgPSCs, it was possible to emphasize the advantages of all three approaches. We consistently found that unipotent germline stem cells, when removed from their niche and cultured in the correct medium, expressed endogenously, pluripotency genes, which induced them to become hgPSCs. These cells are then capable of producing cell types from all three germ layers. Upon differentiation into cardiac lineages, our data consistently showed that they not only expressed cardiac genes, but also expressed cardiac-promoting paracrine factors. Taking these data a step further, we found that hgPSC-derived cardiac cells could integrate into cardiac tissue in vivo. Note, while the work presented here was based on testes-derived hgPSCs, data from other laboratories have shown that ovaries contain very similar types of stem cells that can give rise to hgPSCs. As a result, hgPSCs should be considered a viable option for eventual use in patients, male or female, with ischemic heart disease.

Reprogramming technology holds great promise for the study and treatment of Parkinson's disease (PD) as patient-specific ventral midbrain dopamine (vmDA) neurons can be generated. This should facilitate the investigation of early changes occurring during PD pathogenesis, permitting the identification of new drug targets and providing a platform for drug screening. To date, most studies using reprogramming technology to study PD have employed induced pluripotent stem cells. Research into PD using direct reprogramming has been limited due to an inability to generate high yields of authentic human vmDA neurons. Nevertheless, direct reprogramming offers a number of advantages, and development of this technology is warranted. Previous reports have indicated that induced neural precursors (iNPs) derived from adult human fibroblasts by lineage factor-mediated direct reprogramming can give rise to dopamine neurons expressing tyrosine hydroxylase (TH+). Using normal adult human fibroblasts, the present study aimed to extend these findings and determine the capacity of iNPs for generating vmDA neurons, with the aim of utilising this technology for the future study of PD. While iNPs expressed late vmDA fate markers such as NURR1 and PITX3, critical early regional markers LMX1A, FOXA2 and EN1 were not expressed. Upon differentiation, iNPs gave rise to dopamine neuronal-like cells expressing TUJ1, TH, AADC, DAT, VMAT2 and GIRK2. To induce an authentic A9 phenotype, a series of experiments investigated temporal exposure to patterning factors. Exposure to SHH-C24II, purmorphamine, CHIR99021 and/or FGF8b during or after reprogramming was insufficient to induce expression of early vmDA regional markers. Addition of LMX1A/FOXA2 to the transfection cocktail did not induce a sustained vmDA iNP phenotype. This study reports for the first time that iNPs derived from healthy adult human cells by non-viral expression of lineage factors can give rise to dopamine neuronal-like cells. Direct-to-iNP reprogramming could be a suitable strategy for modelling PD in vitro using aged donor-derived cells.

Background: For regenerative therapies in the orthopedic field, one prerequisite for therapeutic success in the treatment of cartilage defects is the potential of body's own cells to migrate, proliferate and differentiate into functional cells. While this has been demonstrated for mesenchymal stem and progenitor cells (MPC) from healthy tissue sources, the potential of cells from degenerative conditions is unclear. In this study the regenerative potential of MPC derived from subchondral cancellous bone with diagnosed osteoarthritis is evaluated in vitro. Methods: OaMPC isolated from bone chips of three individual patients with Kellgren grade 3 osteoarthritis were characterized by analysis of cell surface antigen pattern. Cell proliferation was evaluated by doubling time and population doubling rate. Cell migration was assessed using a multi-well migration assay. Multi-lineage potential was evaluated by histological staining of adipogenic, osteogenic and chondrogenic markers. In addition, chondrogenic differentiation was verified by qPCR. Results: OaMPC showed a stable proliferation and a typical surface antigen pattern known from mesenchymal stem cells. Cell migration of oaMPC can be induced by human blood serum. OaMPC were capable of adipogenic, osteogenic and chondrogenic differentiation comparable to MPC derived from healthy conditions. Conclusion: OaMPC derived from knee joints affected by osteoarthritic conditions showed regeneration potential regarding migration, proliferation and chondrogenic differentiation. This suggests that oaMPC are able to contribute to cartilage repair tissue formation.

Objective: Multipotential cells are mobilized into peripheral blood in response to trauma, in particular in severe burns. These cells migrate to the site of injury in response to chemotactic signals to modulate inflammation, repair damaged tissue and facilitate tissue regeneration. We evaluated the possibility of isolating and in vitro expand mesenchymal stromal cells (MSCs) from granulation tissue (GT) during debridement of a burn wound, as a persective strategy to improve skin regeneration. Methods: GT obtained from a 12-month-old burn patient was in vitro cultured. Expanded MCSs were characterized for morphology, immunophenotype, differentiation capacity and proliferative growth. Antifibrotic features were also evaluated. Results: It was possible to isolate and in vitro expand cells from GT with the morphology, phenotype, proliferative and differentiation capacity typical of MSC, these cells were defined as GT-MSC. GT-MSCs exhibited antifibrotic features by releasing soluble factors, this activity was superior to that observed in BM-MSC. Conclusions: Successful isolation and expansion of MSCs from GT is reported. Considering their functional characteristics, GT-MSCs could be considered a good candidate adjuvant therapy to improve burn wound healing, particularly in pediatrics.