MiR-219a-2 relieves myocardial ischemia-reperfusion injury by reducing calcium overload and cell apoptosis through HIF1α/ NMDAR pathway
MiR-219a-2 relieves myocardial ischemia-reperfusion injury by reducing calcium overload and cell apoptosis through HIF1α/ NMDAR pathway
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DOI：
10.1016/j.yexcr.2020.112172
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发表时间：
2020-10-01
影响因子：
3.7
通讯作者：
Chen, Kui
中科院分区：
文献类型：
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作者：
Hu, Fudong;Zhang, Shengye;Chen, Kui
目的：心肌缺血再灌注损伤（MIRI）过程中，细胞内Ca2+浓度（[Ca2+](i)）持续升高，导致心肌细胞凋亡，最终造成心肌损伤，但钙超载的上游调控机制尚不清楚。本研究重点探讨miR-219a-2在MIRI中的作用，旨在阐明其对MIRI过程中发生的钙超载的调节机制。方法：通过qRT-PCR测定MIRI小鼠心脏组织中miR-219a-2的表达。使用荧光酶标仪通过 Fluo-3 测量 [Ca2+](i)。通过 qRT-PCR 和蛋白质印迹法测量缺氧诱导因子 1 α (HIF1 α) 和 NR1（N-甲基-D-天冬氨酸受体 1 (NMDAR) 的必需亚基）的表达。荧光素酶报告基因测定用于确认 miR-219a-2 和 HIF1 α 之间的相互作用以及 HIF1 α 和 NR1 之间的相互作用。通过 B 细胞淋巴瘤 2 相互作用细胞死亡介质 (Bim) 的表达水平和 TUNEL 阳性细胞的数量来测量细胞凋亡。采用TTC/伊文思蓝染色法测定小鼠心肌梗死面积。结果：MIRI小鼠心脏组织中miR-219a-2表达下调。 miR-219a-2 过表达降低缺氧/复氧 (H/R) 处理的 HL-1 细胞中的 [Ca2+](i) 以及 HIF1 α 和 NR1 的表达。然后，荧光素酶报告基因检测显示miR-219a-2抑制HIF1α的转录，而HIF1α促进NR1的转录。 HIF1 α过表达和 NMDAR 功能增强均消除了 miR-219a-2 对 H/R 处理的 HL-1 细胞中钙超载和细胞凋亡的抑制作用。最后，miR-219a-2的过表达降低了MIRI小鼠的Ca2+浓度、细胞凋亡和心肌梗死面积，而NMDAR功能增强剂逆转了miR-219a-2的治疗效果。结论：miR-219a-2通过HIF1α/NR1轴减少NMDAR介导的钙超载，从而减轻MIRI中的细胞凋亡。
Objective: During the process of myocardial ischemia-reperfusion injury (MIRI), the intracellular Ca2+ concentration ([Ca2+](i)) continues to increase, leads to the cardiomyocyte apoptosis and eventually causes myocardial damage, while the upstream regulation mechanism of calcium overload is still unknown. This study focuses on the role of miR-219a-2 in MIRI and aims to elaborate its regulatory mechanism on calcium overload that occurs during MIRI.Methods: The expression of miR-219a-2 was determined in the heart tissues of MIRI mice by qRT-PCR. The [Ca2+](i) was measured by fluo-3 using a fluorescence microplate reader. The expression of hypoxiainducible factor 1 alpha (HIF1 alpha) and NR1, the obligatory subunit of N-methyl-D-aspartate receptor 1 (NMDAR), were measured by qRT-PCR and western blot. The luciferase reporter assay was used to confirm the interplay between miR-219a-2 and HIF1 alpha and the interplay between HIF1 alpha and NR1. The cell apoptosis was measured by the expression level of B-cell lymphoma 2 interacting mediator of cell death (Bim) and the number of TUNEL-positive cells. The myocardial infarct size of mice was measured by TTC/Evans Blue staining.Results: MiR-219a-2 was down-regulated in the heart tissues of MIRI mice. miR-219a-2 overexpression decreased [Ca2+](i) and the expression of HIF1 alpha and NR1 in hypoxia/reoxygenation (H/R)-treated HL-1 cells. Then, the luciferase reporter assay showed that miR-219a-2 inhibited the transcription of HIF1 alpha and HIF1 alpha promoted the transcription of NR1. Both HIF1 alpha overexpression and NMDAR function enhancement removed the inhibitory effect of miR-219a-2 on calcium overload and cell apoptosis in H/R-treated HL-1 cells. Finally, the overexpression of miR-219a-2 decreased Ca2+ concentration, cell apoptosis, and myocardial infarction size in MIRI mice, while the NMDAR function enhancer reversed the therapeutic effect of miR-219a-2.Conclusion: MiR-219a-2 reducing NMDAR-mediated calcium overload via HIF1 alpha/NR1 axis, thus alleviating cell apoptosis in MIRI.