小鼠多潜能干细胞来源的内皮细胞的基因表达及功能特点
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863项目(2006AA02A102);973项目(2007CB947901)


Gene Expression and Functional Characteristics of Mouse iPS cell-derived Endothelial Cells
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    摘要:

    目的探讨小鼠成纤维细胞来源的多潜能干细胞向内皮细胞诱导分化的能力。方法采用单层贴壁诱导及流式细胞术进行Flk1+细胞分化与分选,进而与OP9基质细胞共培养,并用VE-cadherin为标志纯化内皮细胞(O9-EC);采用免疫荧光检测分化细胞的内皮细胞特异蛋白表达,检测分化的内皮细胞体外成血管能力,吞噬DiI-Ac标记的低密度脂蛋白和结合荆豆凝集素的功能;采用定量实时PCR检测分化过程中的基因表达。结果流式细胞仪分选得到的分化细胞能表达CD31、VE-cadherin、vWF等表面标志,吞噬DiI-Ac-LDL,结合荆豆凝集素,并能在matrigel上形成血管样结构。分化过程中内皮细胞特异性基因如CD31、Tie2、eNOS等表达上调,而Oct4、Sox2、Klf4和c-Myc这四种诱导多潜能干细胞形成的转录因子的表达显著下调。结论多潜能干细胞能向内皮细胞诱导分化。

    Abstract:

    AimTo investigate the potential of mouse fibroblast cell-derived embryonic-like induced pluripotent stem cells (iPS cells) to differentiate into endothelial cells.MethodsO9 iPS cells were induced to differentiate into endothelial cells by monolayer culture, FACS sorting Flk1+ cells, followed by OP9 stromal cell co-culture, and purified by FACS sorting VE-cadherin+ cells(O9-EC).The endothelial specific protein expression were examined by immunofluorescence.The functions of angiogenesis in vitro, uptaking DiI-Ac-LDL and binding UEA1 were also tested.Gene expression during differentiation process were tested by real-time PCR.ResultsMouse O9 iPS cells derived Flk1+cells could differentiate into endothelial-like cells on OP9 stromal cells.VE-cadherin could be used as a marker to purify O9-EC.O9-EC could express CD31, VE-cadherin and vWF, uptake DiI-Ac-LDL, bind UEA1 and form tube-like structures on matrigel.The endothelial specific genes such as CD31, Tie2 and eNOS were up-regulated in the endothelial differentiation process.Four reprogramming factors (Oct4, Sox2, Klf4 and c-Myc) that induce the generation of iPS cells were significantly down-regulated in this process.ConclusionThis study supports that iPS cells can differentiate into endothelial cells in vitro.This procedure may contribute to clinic application of patient-specific iPS cells derived endothelial cells in vascular degeneration diseases.

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孙璇,卢光琇.小鼠多潜能干细胞来源的内皮细胞的基因表达及功能特点[J].中国动脉硬化杂志,2010,18(1):15~19.

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  • 收稿日期:2009-11-20
  • 最后修改日期:2009-01-12
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