摘要
目的:探讨热休克转录因子1(HSF1)调控微小RNA-195a-3p (miR-195a-3p)对心肌微血管内皮细胞血管新生功能的影响,旨在阐明HSF1基因缺失加重压力超负荷下心脏重构的病理分子机制。方法:压力超负荷动物模型采用小鼠主动脉弓缩窄(TAC),辅以心脏超声功能评价。在体实验分组为HSF1基因敲除(HSF1^(-/-))小鼠假手术组、C57BL/6野生型(WT)小鼠假手术组、HSF1^(-/-)小鼠TAC模型组和C57BL/6 WT小鼠TAC组;细胞实验分组为对照组、miR-195a-3p模拟物干预组和阴性对照microRNA(miR-NC)干预组。TAC术后4周,通过病理组织切片检测各组小鼠心肌肥厚(HE染色)和血管新生(CD31染色),小鼠心超检查心脏主要功能指标变化;通过生物信息软件TargetScan 6.2结合萤光素酶(luciferase)报告基因检测,以及Western blot验证,测定miR-195a-3p调控血管新生的下游分子靶点;并用miR-195a-3p诱导微血管内皮细胞,观察其对细胞成管能力的影响。结果:HSF1缺失会导致TAC诱导的小鼠左心室重构加重。芯片筛查结果表明HSF1缺失可促进心脏12种microRNAs表达上调,5种microRNAs在心肌微血管内皮细胞中表达显著升高,其中miR-195a-3p的增高具有统计学显著性。miR-195a-3p过表达可以有效抑制微血管内皮细胞CD31和血管内皮生长因子(VEGF)的表达,阻滞细胞形成管腔样结构。TargetScan 6.2预测miR-195a-3p的靶点为AMP活化蛋白激酶α2(AMPKα2),在微血管内皮细胞用miR-195a-3p模拟物干预可以有效抑制AMPKα2的表达,同时miR-195a-3p模拟物也抑制了CD31和VEGF的表达。结论:HSF1基因缺失导致的miR-195a-3p表达上调是加速压力超负荷下心脏重构的重要诱因,其机制可能是通过抑制AMPKα2介导的血管新生信号通路的激活。
AIM: To investigate the effect of heat shock transcriptional factor 1(HSF1) on microRNA-195 a-3 p(miR-195 a-3 p) expression and angiogenesis in myocardial microvascular endothelial cells(MMECs), and to elucidate the mechanism of pressure overload-induced cardiac remodeling under the condition of HSF1 deficiency. METHODS: The animal model of pressure overload was achieved by aortic arch constriction(TAC) in mice and evaluated by echocardiography. The mice were divided into 4 groups: HSF1 gene knockout(HSF1-/-) mice and C57 BL/6 wild-type(WT) mice either with sham operation or TAC operation. The in vitro cultured MMECs were divided into control group, miR-195 a-3 p mimic group and negative control microRNA(miR-NC) group. At 4 weeks after TAC, cardiac hypertrophy was assessed by HE staining, cardiac angiogenesis was observed by CD31 staining, and cardiac function was evaluated by echocardiography. The downstream angiogenesis-related target gene that regulated by miR-195 a-3 p was analyzed by bioinformatics software TargetScan 6.2, which was further confirmed by luciferase reporter assay and Western blot. The effects of miR-195 a-3 p on apoptosis and ability of tube formation were also examined in MMECs. RESULTS: Deletion of HSF1 led to the exacerbation of left ventricular remodeling in the mice induced by TAC. The results of microRNA chip showed that 12 microRNAs were up-regulated in HSF1-/- mice as compared with WT mice. Among them, 5 microRNAs were up-regulated in the MMECs, and only the up-regulation of miR-195 a-3 p had statistical difference. miR-195 a-3 p over-expression in the MMECs effectively inhibited the expression of CD31 and VEGF, and inhibited capillary-like tube formation. TargetScan 6.2 predicted that the target of miR-195 a-3 p was AMP-activated protein kinase α2(AMPKα2). miR-195 a-3 p mimic treatment effectively inhibited the expression of AMPKα2, CD31 and VEGF in the MMECs. CONCLUSION: The up-regulation of miR-195 a-3 p in MMECs due to HSF1 deficiency accelerates pressure overload-induced cardi
作者
王时俊
徐磊
赵刚
杨继娥
马雷雷
崔兆强
邹云增
葛均波
WANG Shi-jun;XU Lei;ZHAO Gang;YANG Ji-e;MA Lei-lei;CUI Zhao-qiang;ZOU Yun-zeng;GE Jun-bo(Shanghai Institute of Cardiovascular Diseases,Zhongshan Hospital,Fudan University,Shanghai 200032,China)
出处
《中国病理生理杂志》
CAS
CSCD
北大核心
2019年第3期385-392,共8页
Chinese Journal of Pathophysiology
基金
国家自然科学基金资助项目(No.81770274
No.81700377
No.81300097)
上海市自然科学基金资助项目(No.13ZR1406800
No.16140901100)
