MicroRNA-181b诱导血管内皮细胞衰老和功能异常的机制研究  

Micro RNA-181b Induces Senescence and Dysfunction of Human Vascular Endothelial Cells

作　　者：米雪楠[1] 杨淑均 陈宇[1] 惠汝太 张伟丽[1] 

机构地区：[1]北京协和医学院中国医学科学院 国家心血管病中心阜外医院心血管疾病国家重点实验室,北京市100037

出　　处：《中国分子心脏病学杂志》2014年第6期1157-1161,共5页Molecular Cardiology of China

基　　金：国家自然科学基金重大研究计划项目(91339101);高血压病研究北京市重点实验室2014年开放课题

摘　　要：目的研究micro RNA-181b(mi R-181b)和胰岛素样生长因子1受体(Insulin-like growth factor 1 receptor,IGF1R)参与血管内皮细胞衰老和血管新生的调节机制。方法体外培养人脐静脉内皮细胞(Human umbilical vein endothelial cells,HUVECs),传代培养计算细胞群体倍增水平(Population doublings,PDL),将PDL≤8 HUVECs定义为年轻细胞,PDL≥44 HUVECs定义为衰老细胞,并检测mi R-181b和IGF1R在年轻(PDL8)和年老(PDL44)HUVECs中的表达。PDL8 HUVECs过表达或抑制mi R-181b后,分别用MTS比色法、划痕(Wound healing)和成管(Tube formation)实验检测内皮细胞的增殖、迁移、成管等血管新生能力。采用双荧光素酶报告系统法检测mi R-181b对IGF1R转录后水平的调控。此外,给予缺氧刺激,观察缺氧应激条件下mi R-181b对IGF1R表达的调控作用。结果过表达mi R-181b可抑制PDL8内皮细胞的增殖能力22%(P<0.001)、迁移能力23%(P<0.001),对成管能力没有显著影响(P>0.05)。与PDL8HUVECs比较,mi R-181b在PDL44衰老内皮细胞中上调64%(P=0.046),IGF1R的m RNA和蛋白水平分别下调39%和45%(P=0.004,P=0.014)。荧光素酶活性实验显示mi R-181b可与IGF1R的3’UTR结合,但过表达mi R-181b对IGF1R的表达水平无显著影响(P>0.05)。在缺氧应激条件下,mi R-181b可上调IGF1R的表达(P=0.005)。结论 Mi R-181b抑制血管内皮细胞增殖、迁移等血管新生能力,这一作用与mi R-181b在缺氧应激条件下上调血管发育相关基因IGF1R的表达相关,其机制仍需进一步研究。Objective To study the role of micro RNA-181b（mi R-181b） and insulin-like growth factor 1 receptor（IGF1R） in vascular endothelial cells senescence and angiogenesis. Methods Human umbilical vein endothelial cells（HUVECs） were cultured in vitro and population-doubling levels（PDLs） were calculated during passages. PDL8 and PDL44 were respectively identified as young and senescent HUVECs, in which the differential expression of mi R-181 b and IGF1 R were detected. Mi R-181 b mimics and inhibitor were separately transfected into PDL8 HUVECs, and then the abilities of cell proliferation, migration and tube formation were assayed by MTS test, wound healing test, and tube formation test, respectively. The dual luciferase reporter assay was used to analyze the post-transcriptional regulation of IGF1 R by mi R-181 b. Mi R-181 b mimics was transfected into PDL8 HUVECs to detect the expression change of IGF1 R both on m RNA and protein level, furthermore, the regulation of IGF1 R by mi R-181 b was observed under hypoxia condition. Results In vitro analysis of PDL8 HUVECs, overexpression of mi R-181 b inhibited endothelial proliferation by 22%（P〈0.001） and migration by 23%（P〈0.001）, and had no significant effect on endothelial tube formation（P〉0.05）. Compared with PDL8 HUVECs, mi R-181 b was increased by 64%（P=0.046） in PDL44 HUVECs, and the m RNA and protein expression of IGF1 R were decreased by 39%（P=0.004） and 45%（P=0.014）, respectively. The luciferase assay showed that IGF1 R was the putative target of mi R-181 b, but, overexpression of mi R-181 b in PDL8 HUVECs did not change the expression of IGF1 R both on m RNA and protein level（P〉0.05）. However, IGF1 R was surprisingly up-regulated by mi R-181 b under hypoxia condition（P=0.005）. Conclusions Mi R-181 b can inhibit proliferation and migration of endothelial cells, and the role was associated with up-regulation of the angiogenesis-related IGF1 R gene under hypoxia condition, of which mechanisms need a further stud

关 键 词：MICRO RNAS 内皮细胞衰老 血管新生 胰岛素样生长因子1受体 

分 类 号：R363[医药卫生—病理学]