大果木姜子油调控miR-328基因抗心律失常H9c2细胞的机制研究  

Study on Mechanism of Cinnamomi Oleum Anti-arrhythmia of H9c2 Cells by Regulating miR-328 Gene

作　　者：孙恒灿 张丽艳 李军 汪尊冬 汤小芳 刘维琴 SUN Heng-can;ZHANG Li-yan;LI Jun;WANG Zun-dong;TANG Xiao-fang;LIU Wei-qin(Guizhou University of Traditional Chinese Medicine,Guiyang 550025,China;Chongqing Shapingba District Hospital of Tradi-tional Chinese Medicine,Chongqing 400030,China)

机构地区：[1]贵州中医药大学,贵州贵阳550025 [2]重庆市沙坪坝区中医院,重庆400030

出　　处：《中药材》2024年第1期168-175,共8页Journal of Chinese Medicinal Materials

基　　金：国家自然科学基金项目(81760910,82060770);贵州省科技计划项目(黔科合后补助[2020]3003);贵州省科技支撑计划(黔科合支撑[2018]2770)。

摘　　要：目的:探讨大果木姜子油通过调控miR-328基因抗心律失常H9c2细胞的可能机制。方法:慢病毒转染构建miR-328过表达H9c2细胞株,再运用生物机能实验系统构建H9c2细胞和miR-328过表达H9c2细胞心律失常模型。将H9c2细胞分为正常对照组、模型组、大果木姜子油组、miR-328过表达组和miR-328过表达+大果木姜子油组。根据分组条件培养结束后,qPCR检测H9c2细胞中miR-328相对表达量;ELISA检测炎性因子IL-1β、IL-6、TGF-β含量;比色法检测SOD活性和MDA浓度;DCFH荧光探针检测ROS水平;流式细胞术检测细胞凋亡率;Western Blot检测H9c2细胞L型钙通道相关蛋白表达。结果:成功构建H9c2细胞和miR-328过表达H9c2细胞心律失常模型。与正常对照组比较,模型组和miR-328过表达组细胞TGF-β、SOD活性及CaM、CaMKⅡ、p-CaMKⅡ蛋白表达显著降低,细胞凋亡率、miR-328及CaV1.2蛋白表达和IL-1β、IL-6、MDA、ROS水平显著升高(P<0.01)。经大果木姜子油干预后,心律失常模型细胞形态改善,上述指标均被显著逆转(P<0.01)。结论:大果木姜子油可改善心律失常H9c2细胞,其作用机制可能与调控miR-328基因降低炎症因子水平、调节氧化应激、抑制细胞凋亡及调节L型钙通道相关蛋白表达有关。Objective:To investigate the possible mechanism of Cinnamomi Oleum anti-arrhythmia of H9c2 cells by regulating miR-328 gene.Methods:The miR-328 over-expression H9c2 cell line was constructed by lentivirus transfection,and the arrhythmia model of H9c2 cells and miR-328 over-expression H9c2 cells was constructed by biological function experiment system.H9c2 cells were divided into normal control group,model group,Cinnamomi Oleum group,miR-328 over-expression group and miR-328 over-expression with Cinnamomi Oleum group.After the culture was finished according to the group conditions,the relative expression of miR-328 in H9c2 cells was detected by qPCR.The contents of IL-1β,IL-6 and TGF-βwere detected by ELISA.SOD activity and MDA concent were detected by colorimetry.DCFH fluorescence was used to detect the level of ROS,the apoptosis rate was detected by flow cytometry.Western Blot was used to detect the expression of L-type calcium channel-related proteins in H9c2 cells.Results:The arrhythmia model of H9c2 cells and miR-328 over-expression H9c2 cells was successfully constructed.Compared with the normal control group,the TGF-β,SOD activity and CaM,CaMKⅡ,p-CaMKⅡprotein expressions in the model group and miR-328 over-expression group were significantly decreased,the apoptosis rate,protein expressions of miR-328 and CaV1.2 and the levels of IL-1β,IL-6,MDA and ROS were significantly increased(P<0.01).After Cinnamomi Oleum intervention,the morphology of cells in arrhythmia model group was improved,and the above indexes were significantly reversed(P<0.01).Conclusion:Cinnamomi Oleum can improve arrhythmia H9c2 cells,and its mechanism may be related to the regulation of miR-328 gene to reduce the level of inflammatory factors,regulate oxidative stress,inhibit apoptosis and regulate L-type calcium channel protein expressions.

关 键 词：心律失常 大果木姜子油 miR-328基因 炎症因子 氧化应激 细胞凋亡 L型钙通道 

分 类 号：R285.5[医药卫生—中药学]