Enhancement of differentiation efficiency of hESCs into vascular lineage cells in hypoxia via a paracrine mechanism
ABSTRACT Hypoxia is one way of inducing differentiation due to the activation of the key regulatory factor, Hypoxia-inducible factor 1 alpha (HIF-1α). However, the action of HIF-1α on the differentiation of hESCs was unclear until now. To investigate the effect of hypoxia on the differentiation of hESCs, we compared the differentiation efficacy into vascular lineage cells under normoxic and hypoxic conditions. We observed HIF-1α expression and the related expression of pro-angiogenic factors VEGF, bFGF, Ang-1 and PDGF in hEBs cultured under hypoxic conditions. Along with this, differentiation efficacy into vascular lineage cells was improved under hypoxic conditions. When HIF-1α was blocked by echinomycin, both angiogenic factors and the differentiation efficacy were down-regulated, suggesting that the enhancement of differentiation efficacy was caused by intrinsic up-regulation of HIF-1α and these pro-angiogenic factors under hypoxic condition. This response might be primarily regulated by the HIF-1α signal pathway, and hypoxia might be the key to improving the differentiation of hESCs into vascular lineage cells. Therefore, this study demonstrated that microenvironmental changes (i.e., hypoxia) can improve differentiation efficacy of hESCs into a vascular lineage without exogenous factors via cell-intrinsic up-regulation of angiogenic factors. These facts will contribute to the regulation of stem cell fate.
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- "In the same manner, low oxygen has been proven to facilitate the reprogramming of somatic cells to iPSC (Yoshida et al. 2009; Shimada et al. 2012). Moreover, differentiation of pluripotent stem cells under low oxygen tension has been shown to enhance the generation of a variety of cell phenotypes, including neuronal (Fernandes et al. 2010; Garita-Hern andez et al. 2013; Stacpoole et al. 2013; Binh et al. 2014), cardiac (Ng et al. 2010; Van Oorschot et al. 2011; Horton and Auguste 2012), endothelial (Han et al. 2010; Prado-Lopez et al. 2010; Shin et al. 2011), hematopoietic (Lesinski et al. 2012), and chondrogenic (Koay and Athanasiou 2008; Adesida et al. 2012) cells among others. Therefore, oxygen is becoming a key signaling molecule which has to be taken into account in the designing of efficient strategies to direct stem cell differentiation. "
ABSTRACT: Whole-organ decellularization technology has emerged as a new alternative for the fabrication of bioartificial lungs. Embryonic stem cells (ESC) and induced pluripotent stem cells (iPSC) are potentially useful for recellularization since they can be directed to express phenotypic marker genes of lung epithelial cells. Normal pulmonary development takes place in a low oxygen environment ranging from 1 to 5%. By contrast, in vitro ESC and iPSC differentiation protocols are usually carried out at room-air oxygen tension. Here, we sought to determine the role played by oxygen tension on the derivation of Nkx2.1+ lung/thyroid progenitor cells from mouse ESC and iPSC. A step-wise differentiation protocol was used to generate Nkx2.1+ lung/thyroid progenitors under 20% and 5% oxygen tension. On day 12, gene expression analysis revealed that Nkx2.1 and Foxa2 (endodermal and early lung epithelial cell marker) were significantly upregulated at 5% oxygen tension in ESC and iPSC differentiated cultures compared to 20% oxygen conditions. In addition, quantification of Foxa2+Nkx2.1+Pax8- cells corresponding to the lung field, with exclusion of the potential thyroid fate identified by Pax8 expression, confirmed that the low physiologic oxygen tension exerted a significant positive effect on early pulmonary differentiation of ESC and iPSC. In conclusion, we found that 5% oxygen tension enhanced the derivation of lung progenitors from mouse ESC and iPSC compared to 20% room-air oxygen tension.07/2014; 2(7). DOI:10.14814/phy2.12075
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ABSTRACT: Vasculopathy due to ischemia in damaged tissues is a major cause of morbidity and mortality. To treat these conditions, endothelial progenitor cells (EPCs) from various sources, such as umbilical cord or peripheral blood, have been the focus of the regenerative medicine field due to their proliferative and vasculogenic activities. However, the fundamental, molecular-level differences between EPCs obtained from different cellular sources have rarely been studied. In this study, we established endothelial progenitor cells derived from cord blood- and peripheral blood (CB- and PB-EPCs) and investigated their fundamental differences at the cellular and molecular levels through a combination of stem cell biology techniques and proteomic analysis. Our results suggest that specifically up-regulated factors such as STMIN 1, CFL 1, PARK 7, NME 1, GLO 1, HSP 27 and PRDX 2 in CB-EPCs as key elements that could be functionally active in ischemic regions. We also discussed functional behaviors important for inducing and maintaining long-lasting blood vessels under ischemic conditions. As a result, CB-EPCs retained a higher anti-oxidant and migration ability than PB-EPCs in vitro. Furthermore, CB-EPCs retained a higher therapeutic efficacy than PB-EPCs in a hindlimb ischemic disease model. The up-regulated expression pattern of STMIN 1, CFL 1, PARK 7, NME 1, GLO 1, HSP 27 and PRDX 2 was confirmed under several conditions in vitro and in vivo, indicating that the up-regulation of these molecules in CB-EPCs may be critical to the mechanism of healing in ischemic conditions and that CB-EPCs may be more appropriate for inducing neo-vessels. Thus, these results may aid in predetermining which cell sources will be of value for cell-based therapies of pathological conditions and identify several candidate molecules that may be involved in the therapeutic mechanism for ischemia.Biomaterials 12/2012; 34(6). DOI:10.1016/j.biomaterials.2012.11.017 · 8.56 Impact Factor
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ABSTRACT: Multipotent mesenchymal stem cells have recently emerged as an attractive cell type for the treatment of diabetes-associated wounds.The purpose of this study was to examine the potential biological function of human placenta-derived mesenchymal stem cells(PMSCs) in wound healing in diabetic Goto-Kakizaki(GK) rats. PMSCs were isolated from human placenta tissue and characterized by flow cytometry. A full-thickness circular excisional wound was created on the dorsum of each rat. Red fluorescent CM-DiI-labeled PMSCs were injected intradermally around the wound in the treatment group. After complete wound healing, full-thickness skin samples were taken from the wound sites for histological evaluation of the volume and density of vessels. Our data showed that the extent of wound closure was significantly enhanced in the PMSCs group compared with the no-graft controls. Microvessel density in wound bed biopsy sites was significantly higher in the PMSCs group compared with the no-graft controls. Most surprisingly, immunohistochemical studies confirmed that transplanted PMSCs localized to the wound tissue and were incorporated into recipient vasculature with improved angiogenesis. Notably, PMSCs secreted comparable amounts of proangiogenic molecules, such as VEGF, HGF, bFGF, TGF-β and IGF-1 at bioactive levels. This study demonstrated that PMSCs improved the wound healing rate in diabetic rats. It is speculated that this effect can be attributed to the PMSCs engraftment resulting in vascular regeneration via direct de novo differentiation and paracrine mechanisms. Thus, placenta-derived mesenchymal stem cells are implicated as a potential angiogenesis cell therapy for repair-resistant chronic wounds in diabetic patients.Biochemical and Biophysical Research Communications 07/2013; 438(2). DOI:10.1016/j.bbrc.2013.07.088 · 2.30 Impact Factor