转化生长因子-β1促血管内皮细胞向间质细胞转化的作用研究.docVIP

转化生长因子-β1促血管内皮细胞向间质细胞转化的作用研究 简宇1，简华刚1，黄宏2，徐祥2，郭敏1，代卉2，崔文慧2，蒋建新2，朱佩芳2，王正国2 (400016重庆，重庆医科大学第二附属医院创伤烧伤科1；400042重庆，第三军医大学第三附属医院野战外科研究所，创伤、烧伤与复合伤国家重点实验室2) [摘要] 目的 观察转化生长因子-β1（transforming growth factor-β1，TGF-β1）对促进内皮细胞向间质细胞转化（endothelial-mesenchymal (myofibroblast) transition，EnMT）的效应，探讨肌成纤维细胞的来源以及纤维化疾病发生的新机制。方法 分离培养人脐静脉内皮细胞，采用不同浓度的TGF-β1（10、25、和50 ng/ml）刺激细胞 72 h后，倒置显微镜观察诱导后细胞形态变化，免疫荧光方法检测第Ⅷ因子相关抗原和α-SMA表达变化，RT-PCR检测VE-钙粘蛋白（VE-Cadherin），α-SMA和Ⅰ型胶原基因表达变化。 结果 正常对照组内皮细胞呈铺路石样生长，TGF-β1刺激组内皮细胞由铺路石样形态向梭形转化。免疫荧光检测可见对照组含有少量FⅧ，α-SMA双阳性细胞。TGF-β1组FⅧ，α-SMA双阳性细胞明显增多，并且具有明显的浓度依赖性（P0.05,P0.01）。RT-PCR检测结果显示，TGF-β1刺激导致内皮细胞特异性标记物VE-钙粘蛋白（VE-cadherin）基因的表达显著减少（P0.05），随着TGF-β1刺激浓度的增加，VE-钙粘蛋白基因表达呈明显的下降趋势；相反，内皮细胞间质性标记物α-SMA和I型胶原基因的表达显著增加（P0.05,P0.01）,并且具有明显的浓度依赖性。结论TGF-β1刺激内皮细胞后，细胞表型和功能均表现为间质细胞特性，提示TGF-β1具有促进内皮细胞向间质细胞转化（EnMT）的作用，并且其促EnMT效应具有明显的浓度依赖性。 [关键词] 内皮细胞；肌成纤维细胞；转分化；转化生长因子-β1 TGF-β1-induced transformation of endothelial into myofibroblast : a study of role Jian Yu1, Huang Hong2, Jian Hua-gang1, Guo Min1, Dai Hui2, Cui Wen-hui2, Zhang Bo2, Jiang Jian-xin2, Zhu Pei-fang2, Wang Zheng-guo2(1Department of burn and trauma, The Second Affiliated Hospital, Chongqing Medical University, Chongqing 400016; 2State Key Laboratory of Trauma, Burns and Combined Injury, Institute of Surgery Research, The Third Affiliated Hospital, Third Military Medical University, Chongqing, 400042, China) Abstract: Objective To explore the effect of endothelial-mesenchymal (myofibroblast）transition of human umbilical vein endothelial cells induced by different concentration TGF-β1, and the changes of cell phenotype and function. Methods Human umbilical vein endothelial cells (HUVECs) were isolated and cultured and then prepared as experiment cells after 3 passages. HUVECs were divided into four groups randomly: PBS was added to culture medium in control group; 10, 25 and 50 ng/mL TGF-β1 were added to culture medium in TGF-β1 groups respectively. At culturing for 72 hours.The changes of cell morpholgy were observed w