Abstract Purpose: 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. Methods: 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. Results: 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. Conclusion: Taken together, our findings indicated that citrate ameliorates H/R injury-induced cardiomyocytes autophagy by regulating miR-142-3p/Rac1 aix.

中文翻译：

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柠檬酸盐预处理通过调节 microRNA-142-3p/Rac1 aix 减轻缺氧/复氧诱导的心肌细胞损伤

摘要 目的：柠檬酸盐对改善心肌细胞的心力衰竭、耳房颤动等病理生理变化有积极作用。然而，其潜在机制仍不清楚。方法：采用大鼠心肌细胞建立缺氧/复氧（H/R）细胞模型。柠檬酸盐对心肌细胞进行预处理，microRNA-142-3p (miR-142-3p) 敲低和过表达用于确定其在心肌细胞中的功能的潜在机制。CCK-8和流式细胞术分别检测细胞活力和凋亡。通过蛋白质印迹和 qRT-PCR 确定蛋白质和 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 的调节。结果：H/R模型中miR-142-3p水平下调，但在柠檬酸盐处理后在心肌细胞中上调。柠檬酸盐减弱 H/R 损伤诱导的 miR-142-3p 水平和细胞活力，还抑制 H/R 损伤诱导的细胞凋亡、LD​​H、MDA 和自噬。细胞活力得到改善，自噬被 miR-142-3p 模拟物抑制，而抑制剂具有相反的结果。与H/R+miR-142-3p抑制剂组相比，Cit+H/R+miR-142-3p抑制剂组细胞活力较高，细胞凋亡和自噬较低。此外，与 H/R + miR-142-3p 抑制剂组相比，Rac1 是 miR-142-3p 的靶基因，并被柠檬酸盐降低。结论：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 aix 改善 H/R 损伤诱导的心肌细胞自噬。柠檬酸盐减弱 H/R 损伤诱导的 miR-142-3p 水平和细胞活力，还抑制 H/R 损伤诱导的细胞凋亡、LD​​H、MDA 和自噬。细胞活力得到改善，自噬被 miR-142-3p 模拟物抑制，而抑制剂具有相反的结果。与H/R+miR-142-3p抑制剂组相比，Cit+H/R+miR-142-3p抑制剂组细胞活力较高，细胞凋亡和自噬较低。此外，与 H/R + miR-142-3p 抑制剂组相比，Rac1 是 miR-142-3p 的靶基因，并被柠檬酸盐降低。结论：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 aix 改善 H/R 损伤诱导的心肌细胞自噬。柠檬酸盐减弱 H/R 损伤诱导的 miR-142-3p 水平和细胞活力，还抑制 H/R 损伤诱导的细胞凋亡、LD​​H、MDA 和自噬。细胞活力得到改善，自噬被 miR-142-3p 模拟物抑制，而抑制剂具有相反的结果。与H/R+miR-142-3p抑制剂组相比，Cit+H/R+miR-142-3p抑制剂组细胞活力较高，细胞凋亡和自噬较低。此外，与 H/R + miR-142-3p 抑制剂组相比，Rac1 是 miR-142-3p 的靶基因，并被柠檬酸盐降低。结论：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 aix 改善 H/R 损伤诱导的心肌细胞自噬。自噬被 miR-142-3p 模拟物抑制，而抑制剂具有相反的结果。与H/R+miR-142-3p抑制剂组相比，Cit+H/R+miR-142-3p抑制剂组细胞活力较高，细胞凋亡和自噬较低。此外，与 H/R + miR-142-3p 抑制剂组相比，Rac1 是 miR-142-3p 的靶基因，并被柠檬酸盐降低。结论：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 aix 改善 H/R 损伤诱导的心肌细胞自噬。自噬被 miR-142-3p 模拟物抑制，而抑制剂具有相反的结果。与H/R+miR-142-3p抑制剂组相比，Cit+H/R+miR-142-3p抑制剂组细胞活力较高，细胞凋亡和自噬较低。此外，与 H/R + miR-142-3p 抑制剂组相比，Rac1 是 miR-142-3p 的靶基因，并被柠檬酸盐降低。结论：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 aix 改善 H/R 损伤诱导的心肌细胞自噬。与 H/R + miR-142-3p 抑制剂组相比。结论：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 aix 改善 H/R 损伤诱导的心肌细胞自噬。与 H/R + miR-142-3p 抑制剂组相比。结论：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 aix 改善 H/R 损伤诱导的心肌细胞自噬。