木犀草素通过调节circ917/RBM4/miR-146靶向SERCA2a改善心肌缺血/再灌注损伤的机制研究

Sarcoplasmic reticulum Ca2+ ATPase 2a (SERCA2a) plays a protective role in myocardial ischemia/reperfusion (I/R) injury. Our previous study found that luteolin improved myocardial I/R injury through upregulation SERCA2a mediated by miR-146, however, the regulation mechanism is not elucidated. Circular RNA (circRNA) can influence on the miRs inhibition on target gene translation. Our results of circRNA gene chip suggested that luteolin could reduce the elevation of circ917 induced by myocardial I/R injury. It was reported that knockdown of circular intronic RNAs (ciRNAs) led to the decreased transcript of their parental genes. Circ917 belongs to ciRNA which indicates that circ917 could regulate the transcription of its parental gene in cis. The parental gene of circ917 is RNA binding motif protein 4 (Rbm4). Indeed, the expression of Rbm4 mRNA increased simultaneously in myocardial I/R injury, which could be reduced by luteolin and positively correlated with the level of circ917 in our pre-experiment. RBM4 protein was reported to enhance inhibition effect of miR-146 on targets. Based on above-mentioned issue, we propose a hypothesis that circ917 can promote the inhibition effection of miR-146 on SERCA2a in the translational level through facilitating the transcript of its parental gene Rbm4 in the nucleus and enhancing expression of RBM4 in the cytoplasm, which will be a new tartget for luteolin on protecting I/R injured myocardium mediating by upregulating SERCA2a expression in intervention on miR-146. This study will use SERCA2a knockout mice and Nuclear Run-on assay, RNA immunoprecipitation (RIP) assay and other techniques in both cell and animal levels to provide the experimental basis for the prevention and treatment of myocardium I/R injury.

肌浆网钙ATP酶2a（SERCA2a）在心肌缺血/再灌注（I/R）损伤发挥保护作用。我们在前期研究中发现：木犀草素（Lut）可调控miR-146上调SERCA2a改善心肌I/R损伤，但机制不明。环状RNA(circRNA)可影响miRs对其靶标表达的调控。circRNA芯片结果显示：Lut可下调心肌I/R引起的circ917的升高；预实验也显示其亲本基因Rbm4转录和circ917表达同向。文献提示RBM4蛋白可增强miR-146对靶标的抑制作用。综上，我们提出假设：circ917通过促进Rbm4转录，增强RBM4促进miR-146抑制SERCA2a表达的作用，成为Lut干预miR-146介导SERCA2a表达而保护I/R损伤心肌的新靶点。本课题将从细胞和动物两水平，使用SERCA2a基因敲除鼠和Nuclear Run-on、RIP等技术，探讨以上假设并为心肌I/R损伤的防治提供实验依据。

冠状动脉血运重建后术的技术和手段不断改进和发展，缺血心肌因得到有效复灌而引起缺血/再灌注(ischemia /reperfusion, I/R)损伤导致的后果日益引起临床关注。为深入研究环状RNA(circular RNA，circRNA)是否参与早期心肌I/R损伤的分子机制，为探究冠心病心肌I/R损伤机制并提供有效的干预方法。我们针对筛选出的circRNA展开功能和机制研究。circ917可以促进RBM4基因的转录，并促进RBM4蛋白从胞核向胞浆转移；在胞浆中可能以细胞骨架的作用连接并增强了与RBM4蛋白在胞浆中作用密切相关的AGO2蛋白的活性和功能，从而增强RBM4下调SERCA2a的作用。