miRNA-182通过FGF9调节肺动脉平滑肌细胞表型的研究  

miRNA-182 regulates pulmonary artery smooth muscle cells phenotype via FGF9

作　　者：李广华[1] 朱瑞冬 邓海龙 王秋实[1] 徐广全[1] 

机构地区：[1]哈尔滨医科大学附属第二医院胸外科,黑龙江哈尔滨150081 [2]海伦市人民医院胸外科,黑龙江海伦152300

出　　处：《哈尔滨医科大学学报》2017年第5期391-395,共5页Journal of Harbin Medical University

基　　金：黑龙江省自然科学基金资助项目(C201204;C2015004)

摘　　要：目的研究miRNA-182及其靶蛋白成纤维生长因子9(FGF9)对肺动脉平滑肌细胞表型转换的调节作用。方法体外培养大鼠肺动脉平滑肌细胞(PASMC),用miRNA-182模拟体转染的方法使其过表达,FGF9siRNA转染使其沉默;使用CCK8方法检测细胞增殖能力,分别用RT-q PCR检测miRNA-182的表达,PCR方法检测FGF9及收缩型表型标记物(SMA,SM22α及myosin)的基因水平,Western blot检测其蛋白水平。结果过表达miRNA-182,收缩表型标记物SMA,SM22α及myosin基因表达水平明显升高,SMA蛋白表达量明显增加,转染miRNA-182抑制物之后,SMA蛋白表达量降低;另外,细胞相对吸光度降低,增殖被抑制,并且转染模拟体80 nmol/L较转染40 nmol/L吸光度更低;而转染miRNA-182抑制物之后,细胞相对吸光度明显增强,细胞增殖增加。过表达miR-182后,FGF9mRNA的表达明显降低,并且与模拟体的转染浓度直接相关。沉默FGF9表达之后SMA、SM22α和myosin表达增加,并抑制PASMC的增殖能力,同时沉默miR-182和FGF9表达之后,SMA、SM22α和myosin表达增高。结论miRNA-182通过靶向抑制FGF9蛋白的表达,抑制PASMC收缩型向合成型转化,同时抑制其增殖能力。Objective To study the important roles of miRNA-182 and fibroblast growth factor 9 （FGF9） in rat pulmonary artery smooth muscle ceils （PASMCs） phenotypic modulation. Methods Rat PASMCs were cultured in vitro, miRNA-182 mimics and FGF9 siRNA were transfected into PASMCs. miRNA-182 were examined by RT-qPCR; the gene levels of FGFg, SMA, SM22α and myosin were detected by using PCR, and the protein levels of them were detected by Western blot. The proliferation of PASMCs was tested by CCK8 assay. Results The gene levels of SMC specific markers, including SMA, SM22α and myosin, were distinctly increased by transfection with miR-182 mimics. The Western blot analysis showed that the transfection of SMCs with miR-182 mimic or inhibitor altered the protein levels of SMA. miR-182 overexpression markedly suppressed SMC proliferation, and transfection with miR-182 mimics at 80 nmol/L significantly inhibited cell proliferation to a greater extent than at 40 nmol/L. Transfection with miR-182 mimic slightly inhibited the FGF9 mRNA level depending on the concentrations of mimics. The expression of SMA, SM22α and myosin increased after transfection of FGF9 siRNA into PASMC, the expression of SMA, SM22α and myosin also increased after transfection of FGF9 siRNA and miR-182 inhibitor into PASMC. Conclusion miRNA-182 prevents phenotypic switch and proliferation of PASMCs by regulating the expression of the targeted FGF9.

关 键 词：miRNA-182 肺动脉平滑肌细胞表型转换 纤维母细胞生长因子9 肺动脉高压 肺动脉平滑肌细胞 

分 类 号：R543.2[医药卫生—心血管疾病]