Xiang Haiyan, Yang Juesheng, Li Jin, Yuan Linhui, Lu Fei, Liu Chen, Tang Yanhua
Department of Cardiac Surgery, The Second Affiliated Hospital of Nanchang University, Nanchang, China.
J Recept Signal Transduct Res. 2020 Dec;40(6):560-569. doi: 10.1080/10799893.2020.1768548. Epub 2020 May 26.
Citrate has a positive effect on improving the pathophysiological changes of cardiomyocytes such as cardiac failure and auricular fibrillation. However, the underlying mechanism remains still unclear. Rat cardiomyocytes were used to establish hypoxia/reoxygenation (H/R) cell model. Citrate was conduct to pretreat with cardiomyocytes, and microRNA-142-3p (miR-142-3p) knockdown and overexpression were used to determine the underlying mechanism of their functions in cardiomyocytes. Cell viability and apoptosis were respectively detected by CCK-8 and flow cytometry. Protein and mRNA levels were determined by Western blot and qRT-PCR. Luciferase reporter assay and Targetscan were performed to study the regulation of miR-142-3p and Rac1. The level of miR-142-3p was down-regulated in H/R model, but up-regulated in cardiomyocytes following citrate treatment. Citrates attenuated H/R injury induced miR-142-3p level and cell viability, and also inhibited H/R injury induced apoptosis, LDH, MDA and autophagy. Cell viability was improved, and autophagy was suppressed by miR-142-3p mimic, while inhibitor had opposite results. Compared with H/R + miR-142-3p inhibitor group, cell viability was higher, and apoptosis and autophagy were lower in Cit + H/R + miR-142-3p inhibitor group. Furthermore, Rac1 was target gene of miR-142-3p, and decreased by citrate, in comparison with H/R + miR-142-3p inhibitor group. Taken together, our findings indicated that citrate ameliorates H/R injury-induced cardiomyocytes autophagy by regulating miR-142-3p/Rac1 aix.
柠檬酸盐对改善心肌细胞的病理生理变化（如心力衰竭和心房颤动）具有积极作用。然而，其潜在机制仍不清楚。使用大鼠心肌细胞建立缺氧/复氧（H/R）细胞模型。用柠檬酸盐对心肌细胞进行预处理，并采用微小RNA-142-3p（miR-142-3p）敲低和过表达来确定其在心肌细胞中功能的潜在机制。分别通过CCK-8和流式细胞术检测细胞活力和凋亡情况。通过蛋白质印迹法和qRT-PCR测定蛋白质和mRNA水平。进行荧光素酶报告基因检测和Targetscan以研究miR-142-3p和Rac1的调控。在H/R模型中miR-142-3p水平下调，但在柠檬酸盐处理后的心肌细胞中上调。柠檬酸盐减轻了H/R损伤诱导的miR-142-3p水平和细胞活力，还抑制了H/R损伤诱导的凋亡、乳酸脱氢酶、丙二醛和自噬。miR-142-3p模拟物可提高细胞活力并抑制自噬，而抑制剂则产生相反的结果。与H/R + miR-142-3p抑制剂组相比，Cit + H/R + miR-142-3p抑制剂组的细胞活力更高，凋亡和自噬更低。此外，Rac1是miR-142-3p的靶基因，与H/R + miR-142-3p抑制剂组相比，柠檬酸盐使其水平降低。综上所述，我们的研究结果表明，柠檬酸盐通过调节miR-142-3p/Rac1轴改善H/R损伤诱导的心肌细胞自噬。
