基础医学与临床 ›› 2018, Vol. 38 ›› Issue (5): 604-609.

• 研究论文 • 上一篇    下一篇

肺高压iPSC-ECs的内皮间质转化和成管缺陷机制

赵双,周芳,杨隽   

  1. 中国医学科学院基础医学研究所 北京协和医学院基础学院
  • 收稿日期:2018-01-16 修回日期:2018-03-20 出版日期:2018-05-05 发布日期:2018-04-28
  • 通讯作者: 杨隽 E-mail:jy270@ibms.pumc.edu.cn
  • 基金资助:
    中国医学科学院中央级公益性科研院所基本科研项目;国家自然科学基金

Cellular molecular mechanism of EndoMT and tube formation dysfunction in iPSC-ECs of pulmonary arterial hypertension

  • Received:2018-01-16 Revised:2018-03-20 Online:2018-05-05 Published:2018-04-28
  • Contact: Jun YANG E-mail:jy270@ibms.pumc.edu.cn

摘要: 目的 利用遗传性肺动脉高压(HPAH)患者和对照志愿者(CON)来源的诱导多能干细胞,研究其定向分化得到的内皮细胞(iPSC-ECs)表型及功能的差异,阐明BMPRII突变引起的PAH特征性内皮紊乱病理机制。方法 将CON和HPAH患者的肺动脉平滑肌细胞诱导得到的iPSCs分化为ECs,通过荧光定量PCR分析iPSC-ECs分化过程中多能性和内皮相关基因的mRNA表达水平;免疫荧光鉴定iPSC-ECs表面特征分子;检测参与调控内皮-间充质转分化(EndoMT)过程的相关基因α-SMA,HMGA1,Slug和Snail1表达;比较HPAH和CON来源iPSC-ECs成管功能,检测血管内皮细胞生长因子受体2(VEGFR2)的mRNA和蛋白表达。结果 HPAH与CON多能性基因表达趋势基本一致,而ECs相关基因的表达变化存在差异;HPAH患者来源的iPSC-ECs的α-SMA和EndoMT过程的相关基因HMGA1,Slug和Snail1表达均高于CON ECs(P<0.05);与CON相比,HPAH患者来源的iPSC-ECs成管功能存在缺陷,VEGFR2的mRNA和蛋白水平均较低,VEGF165可使其得到恢复。 结论 HPAH来源的iPSC–ECs存在EndoMT现象和成管功能缺陷,HMGA1、Snail家族转录因子和VEGF通路可能参与对PAH血管重构的调控。

关键词: 肺动脉高压, 内皮细胞, EndoMT, 血管内皮细胞生长因子受体2

Abstract: Objective To investigate the phenotypic and functional differences of endothelial cells derived from induced pluripotent stem cells (iPSC-ECs) between HPAH patient (HPAH) and Control donor (CON), and clarify the molecular mechanism of phenotypic changes in BMPRII deficient ECs which is a pathological characteristic of PAH. Methods Differentiate the iPSCs derived from human pulmonary arterial smooth muscle cells (hPASMCs) to ECs. The expression of several genes related to stem cell and endothelial marker were analyzed at different time points during the differentiation. Immunofluorescence staining showed the expression of surface markers of iPSC-ECs. The transcription factors involved in the EndoMT process were detected by real-time quantitative PCR (qPCR). HPAH and CON-derived iPSC-ECs were compared for tube formation. VEGFR2 mRNA and protein expression were detected by qPCR and immunofluorescence staining. Results The expression of several endothelial cell related genes of HPAH were different from CON during differentiationthrough they both expressed pluripotency genes. The expression of α-SMA, HMGA1, Slug and Snail1 in HPAH iPSC-ECs were significantly higher compared with CON ECs (P<0.05). The tube formation of HPAH-derived ECs reduced which could be rescued by VEGF165. Their VEGFR2 mRNA and protein expression were lower compared with CON ECs. Conclusions It suggests that enhanced HMGA1, Slug and Snail1 genes involve in the EndoMT of iPSC-ECs from HAPH, reduced VEGFR2 may contribute to tube formation dysfunctionin pulmonary vascular remodeling of PAH.

Key words: Key words PAH, endothelial cells, EndoMT, VEGFR2

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