School of Basic Medicine, Youjiang Medical University for Nationalities.
Int Heart J. 2021 May 29;62(3):647-657. doi: 10.1536/ihj.20-600. Epub 2021 May 15.
Bioinformatics analysis showed that miR-448-5p expression in the myocardial tissue of rats with myocardial infarction significantly increased, suggesting that it may participate in myocardial cell apoptosis in myocardial infarction. This study aimed to explore the protective effects of miR-448-5p on hypoxic myocardial cells.H9C2 cells were cultured and subjected to anoxia for 2, 4, and 8 hours to establish a hypoxia model. MiR-448-5p mimic and inhibitor were transfected into the cells; then, a dual-luciferase experiment was conducted to verify the targeting relationship between miR-448-5p and VEGFA. Cell viability and apoptosis was detected by cell counting kit-8 and flow cytometry, respectively. The expressions of apoptosis-related proteins, miR-448-5p, FAS, and FAS-L were measured using western blotting and quantitative reverse transcription-polymerase chain reaction (qRT-PCR).Hypoxia-reduced H9C2 cell viability and promoted apoptosis. MiR-448-5p expression was increased after H9C2 cell hypoxia. MiR-448-5p mimic significantly inhibited the viability and promoted the apoptosis of hypoxia-induced model cells. Hypoxia promoted the expression of apoptosis-related protein B-cell lymphoma-2 (Bcl-2) and inhibited the expressions of Bcl-2-associated x protein (Bax), cleaved caspase-3, and caspase-3, whereas the effect of inhibitor on hypoxia-reduced H9C2 cell and apoptotic protein expression were opposite to miR-448-5p mimic. MiR-448-5p targeted VEGFA and regulated its expression. Silenced VEGFA expression significantly inhibited inhibitor effect on increasing cell viability and promoted apoptosis. In addition, miR-448-5p mimic inhibited the effect of hypoxia on promoting the expressions of FAS and FAS-L of H9C2 cells. Inhibitors had the opposite effect on cell hypoxia model.The miR-448-5p/VEGFA axis could protect cardiomyocytes from hypoxia through inhibiting the FAS/FAS-L signaling pathway.
生物信息学分析显示，心肌梗死后大鼠心肌组织中 miR-448-5p 的表达明显增加，提示其可能参与心肌梗死后心肌细胞凋亡。本研究旨在探讨 miR-448-5p 对缺氧心肌细胞的保护作用。培养 H9C2 细胞并进行缺氧 2、4 和 8 小时，建立缺氧模型。将 miR-448-5p 模拟物和抑制剂转染到细胞中；然后，进行双荧光素酶实验验证 miR-448-5p 与 VEGFA 的靶向关系。通过细胞计数试剂盒-8 和流式细胞术分别检测细胞活力和细胞凋亡。使用 Western blot 和定量逆转录聚合酶链反应（qRT-PCR）检测凋亡相关蛋白、miR-448-5p、FAS 和 FAS-L 的表达。缺氧降低 H9C2 细胞活力并促进细胞凋亡。H9C2 细胞缺氧后 miR-448-5p 表达增加。miR-448-5p 模拟物显著抑制缺氧诱导模型细胞的活力并促进细胞凋亡。缺氧促进凋亡相关蛋白 B 细胞淋巴瘤-2（Bcl-2）的表达，并抑制 Bcl-2 相关 X 蛋白（Bax）、裂解半胱氨酸天冬氨酸蛋白酶-3 和半胱氨酸天冬氨酸蛋白酶-3 的表达，而抑制剂对缺氧降低 H9C2 细胞和凋亡蛋白表达的影响与 miR-448-5p 模拟物相反。miR-448-5p 靶向 VEGFA 并调节其表达。沉默 VEGFA 表达显著抑制抑制剂对增加细胞活力和促进凋亡的作用。此外，miR-448-5p 模拟物抑制缺氧对 H9C2 细胞 FAS 和 FAS-L 表达的促进作用。抑制剂对细胞缺氧模型有相反的作用。miR-448-5p/VEGFA 轴可通过抑制 FAS/FAS-L 信号通路保护心肌细胞免受缺氧。
