microRNA-1和热休克蛋白90在心肌缺氧复氧中的关系

The Relation Between MicroRNA-1 and HSP90 in Cardiac Hypoxia/reoxygenation

目的初步探讨心肌细胞缺氧复氧(H/R)时miR-1和热休克蛋白90(HSP90)的关系。方法建立乳大鼠心肌细胞H/R模型,Western-Blot检测HSP90蛋白表达。生物学信息分析miR-1和HSP90亚型的结合情况。重组miR-1慢病毒感染乳大鼠心肌细胞,Western-Blot检测HSP90aa1和HSP90b1蛋白表达,实时荧光定量PCR检测HSP90aa1和HSP90b1的mRNA表达。重组miR-1慢病毒感染乳大鼠心肌细胞后H/R处理,Western-Blot检测HSP90、HSP90aa1、HSP90b1蛋白的表达。结果心肌细胞H/R后,HSP90蛋白表达水平下降。TargetScan分析显示HSP90蛋白的两个亚型HSP90aa1和HSP90b1的3’-UTR与miR-1结合。与对照组相比,HSP90aa1和HSP90b1蛋白在感染LV-rno-miR-1组表达水平降低,而在感染LV-rno-miR-1 sponge组中表达水平增加。感染LV-rno-miR-1组和LV-rno-miR-1 sponge组较对照组的HSP90aa1和HSP90b1的mRNA表达无明显变化。相对于感染LV-rno-miR-1组和缺氧复氧组,感染LV-rno-miR-1病毒后缺氧4小时复氧12小时,HSP90、HSP90aa1、HSP90b1蛋白表达水平进一步降低。结论心肌缺氧复氧时,HSP90蛋白及其亚型aa1和b1表达下降。miR-1可能在心肌缺氧复氧中直接或间接调控HSP90。

Aim The study was aimed to preliminarily explore the relation between miR-1 and HSP90 in a cell model of cardiac hypoxia/reoxygenation(H/R).Methods The myocardial hypoxia/reoxygenation(H/R)model was established using neonatal rat ventricular myocytes.The expression of HSP90 was detected by Western-Blot.TargetScan were applied to predict the potential relation between miRNA-1 and HSP90.After the infection of recombinant miR-1 lentivirus,Western-Blot and qPCR was applied to analyze the level of HSP90 aa1 and HSP90 b1.After the infection of recombinant miR-1 lentivirus,the cardiomyocytes with or without H/R treatment were collected,and Western-Blot was used to analyze the level of HSP90/HSP90 aa1/HSP90 b1.Results The expression level of HSP90 was downregulated in H/R.Using TargetScan software,HSP90 aa1 and HSP90 b1 were identified as the target gene of miR-1.Overexpression of miR-1 by infecting LV-rno-miR-1 downregulated the expression level of HSP90 aa1 and HSP90 b1 in rat myocardial cells,while inhibition of miR-1 by infecting LV-rno-miR-1 sponge increased the HSP90 aa1 and HSP90 b1 expression.However,there is no difference in the mRNA of HSP90 aa1/b1.Compared with overexpression of miR-1 and cell model of H/R,upregulated miR-1 by infecting LV-rno-miR-1 in H/R decreased the level of HSP90,HSP90 aa1 and HSP90 b1.Conclusion Our data demonstrate that HSP90/90 aa1/b1 was decreased in H/R.miR-1 may regulate HSP90 in H/R in a direct or indirect way.