The Pro-hypertrophic Role And Mechanism Of MiR-214in Cardiac Hypertrophy

Background and aims:Cardiac hypertrophy is a late clinical stage of various cardiac diseases. In China, the incidence of cardiac hypertrophy increases year by year and ranks first among the reasons for the occurrence of cardiac failure. Nevertheless, we are still lack of comprehensive and in-depth understanding of the pathogenesis of cardiac hypertrophy. But it is certain that the development of cardiac hypertrophy is a multi-gene involved and multi-channel process. The process has many genes changes. MicroRNAs are a class of small non-coding RNA capable of negatively regulating the gene expression. In cardiac hypertrophy, there are also abnormal expression of microRNAs. MicroRNAs may play an important role in pathogenesis of cardiac hypertrophy.This study used quantitative PCR method to verify the miR-214expression in cardiomyocytes and tissues of cardiac hypertrophy model. We utilized miR-214transgenic mice to identify the biological function of miR-214in cardiac hypertrophy. Meanwhile, we predicted the miR-214target gene and signal pathway referring to cardiac hypertrophy process.Methods and Results:Our research focused on discussing the pro-hypertrophic role and mechanism of miR-214in cardiac hypertrophy. The main investigations and results are as follows: 1. The cardiomyocytes area of PE inciting hypertrophic model are bigger than normal cardiomyocytes. MiR-214expression was quantified in PE inciting hypertrophic model by qPCR. Its expression upregulated while comparing with normal cardiomyocytes. Meanwhile, we use TAC rats model to identify morphologic features, HW/BW parameter, hypertrophic genes:Actal, Myh7, Nppa expression. We found all of the above results of TAC models demonstrating the occurrence of cardiac hypertrophy. Moreover, the miR-214expression level is also found upregulation in TAC model.2. We also constructed miR-214transgenetic mice model. The miR-214expression level increased as5folds high as littermate controls. Further we found cardiac dysfunction:LVIDs, LVIDd increment, LVPWs decrement, EF%, FS%increment and LVM/BW increment by M-mode echocardiography. Meanwhile, morphologic features, HW/BW and HW/TL parameters dramatically increased comparing with littermate controls. Finally we used immunohistochemistry to detect the cardiomyocytes area of miR-214transgenetic mice. They were also significant enlargement.3. We explored the possible target of miR-214by bioinformatical method. We predicted that EZH2could be downregulated by miR-214. Further qPCR analysis, Westernblot and luciferase assay confirmed that miR-214transcriptionally repressed EZH2by interacting with the essential binding sequence located in3’-UTR of EZH2.4. By lentiviral transfection of miR-214plasmid and sponge, we revealed the mechanism of miR-214regulated cardiac hypertrophy via EZH2/Sixl signal pathway. Lenti-miR-214enlarged the cardiomyocytes area and upregulated Actal, Myh7, Nppa expression. Meanwhile, in this process EZH2downregulated and Sixl upregulated. Lenti-miR-214spg repressed the PE inciting cardiac hypertrophy according to decrement of the cardiomyocytes area and Actal, Myh7, Nppa downregulation. Moreover EZH2upregulated and Sixl downregulated by Lenti-miR-214spg transfection.5. We observed the antagomir-214role in TAC mice model to identify whether it can rescue the cardiac hypertrophy. We demonstrated that antagomir-214relieved the level of cardiac hypertrophy according to morphologic features, HW/BW and HW/TL parameters, HE stained and immunohistochemistrical results and Actal, Myh7, Nppa downregulation. These results all suggested significant improvement of cardiac hypertrophy by antagomir-214. Further cardiac dysfunction also was recoverd on the basis of M-mode echocardiography results.Conclusions:1. MiR-214is novel stimulating microRNA of cardiac hypertrophy.2. EZH2is a direct target of miR-214, and it played an important role in the pro-hypertrophic mechanism of miR-214.3. MiR-214provokes cardiac hypertrophy and dysfunction via EZH2/Sixl signal pathway. It could be a novel therapeutic target in cardiac hypertrophy.