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MicroRNA-18a通过HIF-1-alpha调控缺氧引起的人肺动脉平滑肌细胞增殖
引用本文:秦国伟,毛文君,王谦,陈静瑜,聂晓伟.MicroRNA-18a通过HIF-1-alpha调控缺氧引起的人肺动脉平滑肌细胞增殖[J].现代生物医学进展,2016,16(16):3023-3027.
作者姓名:秦国伟  毛文君  王谦  陈静瑜  聂晓伟
作者单位:南京医科大学附属无锡市人民医院麻醉科;南京医科大学附属无锡市人民医院胸外科;南京医科大学附属无锡市人民医院江苏省人体器官移植重点实验室
基金项目:国家自然科学基金项目(81500039);南京医科大学科技发展基金项目(2015NJMU148)
摘    要:目的:研究microRNA-18a (miR-18a) 对缺氧引起的人肺动脉平滑肌细胞(human pulmonary artery smooth muscle cells, hPASMCs)增殖的调控作用及其可能机制。方法:体外培养hPASMCs,分为未转染组、miR-18a 模拟物对照组、miR-18a 模拟物组、 miR-18a 抑制剂对照组、miR-18a 抑制剂组、siRNAcontrol组、siHIF-1-alpha组、miR-18a 抑制剂和siHIF-1-alpha共转染组。分别于常氧(21% O2)和低氧(3%O2)作用24小时。采用CCK-8 法检测细胞的增殖情况,萤光素酶报告基因系统验证缺氧诱导因子-1-alpha(HIF-1alpha)是 否为miR-18a 的靶基因,并通过western-blot 以及实时荧光定量PCR 技术检测相关蛋白和基因的表达。结果:缺氧可促进 hPASMCs 增殖,使miR-18a 表达减少;miR-18a 模拟物可抑制hPASMCs 增殖,而miR-18a 抑制剂可促进hPASMCs 增殖;抑制 miR-18a 可使HIF-1-alpha的表达上调。同时抑制miR-18a 和HIF-1-alpha,可使miR-18a 对hPASMCs 增殖调控的能力消失。结论:缺氧通 过抑制miR-18a,上调HIF-1-alpha的表达,促进hPASMCs增殖。

关 键 词:肺动脉平滑肌细胞  MicroRNA-18a  缺氧  缺氧诱导因子-1alpha

MicroRNA-18a Regulates the Proliferation Pulmonary Artery Smooth Muscle Cells Through HIF-1-alpha
Abstract:Objective:To study the effects of microRNA-18a (miR-18a) on human pulmonary artery smooth muscle cells (hPASMCs) proliferation and the mechanisms involved.Methods:hPASMCs were divided into eight groups, including: non-transfected group, miR-18a mimic control group, miR-18a mimic group, miR-18a inhibitor control group, miR-18a inhibitor group, siRNA control group, siHIF-1-alpgha group, miR-18a inhibitor and siHIF-1-alpha co-transfected group. Cells were subjected to normoxia (21%O2) and hypoxia (3%O2) for 24 h. Cell viability (CCK-8), miRNA-mRNA interactions (HIF-1-alpha3'' untranslated region Luciferase reporter assays), gene expression (quantitative PCR and western-blot) were assessed under normoxic or hypoxic conditions.Results:Hypoxia promoted hPASMCs proliferation but inhibited miR-18a expression. Ectopic expression of miR-18a in hPASMCs reduced proliferation, whereas miR-18a inhibition in hPASMCs promoted proliferation. We identified HIF-1-alpha as a direct target of miR-18a. Modulating miR-18a expression significantly affected HIF-1-alpha expression. The capacity of miR-18a on hPASMCs proliferation was attenuated by inhibiting miR-18a and HIF-1-alphatogether.Conclusion:Together, these data support a role of miR-18a in hypoxia-induced hPASMCs proliferation through a HIF-1-alpha -dependent pathway.
Keywords:Human pulmonary artery smooth muscle cells  Hypoxia  microRNA-18a  HIF-1alpha
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