miR-101-3p通过MAPK信号通路保护心肌缺血再灌注损伤的机制研究-现代医学

miR-101-3p通过MAPK信号通路保护心肌缺血再灌注损伤的机制研究

单位：商丘医学高等专科学校基础医学教学部, 河南商丘 476100

关键词：微小RNA-101-3p 心肌缺血/再灌注小鼠缺氧/复氧心肌细胞丝裂原活化激酶信号通路

分类号：R542.2

出版年·卷·期（页码）：2022·50·第六期（665-671）

摘要：

目的：观察miRNA (miR)-101-3p对小鼠心肌缺血/再灌注(I/R)损伤心脏的作用及缺氧/复氧(H/R) H9C2细胞存活的影响，并探究其作用机制。方法：建立I/R小鼠模型和H/R H9C2细胞模型。miR-NC、miR-101-3p注射入I/R小鼠再进行I/R模型制作；脂质体法将agomiR-NC、agomiR-101-3p转入H9C2细胞再进行H/R模型制作，部分细胞用二甲基亚砜(DMSO)、丝裂原活化激酶(MAPK)信号通路抑制剂SB203580预处理30 min；心脏超声检测小鼠左室舒张末压(LVDP)、左室射血分数(LVEF)、左室短轴缩短率(LVFS)等指标；细胞计数试剂盒(CCK8)、膜联蛋白V-异硫氰酸荧光素-碘化丙锭(Annexin V-FITC/PI)双染法检测细胞增殖率、凋亡率；双荧光素酶报告实验检测细胞荧光活性；蛋白质印迹法(WB)检测细胞MAPK1、MAPK6、MAPK8蛋白表达。结果：与对照组小鼠心脏或H9C2组细胞相比，I/R组小鼠和H/R H9C2细胞中miR-101-3p的表达均显著降低，小鼠心脏LVDP、LVEF、LVFS均显著下降，细胞增殖率显著降低，凋亡率显著升高(P＜0.05)。过表达miR-101-3p后，小鼠心脏LVDP、LVEF、LVFS均显著上升，细胞增殖率显著升高，凋亡率显著降低(P＜0.05)；miR-101-3p显著抑制野生型MAPK1、MAPK6、MAPK8细胞的荧光活性，并负向调控其蛋白表达；MAPK信号通路抑制剂加强了miR-101-3p过表达对H/R H9C2细胞的促进增殖和抑制凋亡的作用。结论：miR-101-3p可保护缺血再灌注心肌损伤，促进心肌细胞增殖，抑制凋亡，其机制与直接靶向抑制MAPK信号通路活性有关。

Objective: To observe the cardiac effect of miR-101-3p on myocardial ischemia/reperfusion injury(I/R) mice and the survival of hypoxia/reoxygenation(H/R) H9C2 cells, and to explore its mechanism. Methods: I/R mouse model and H/R H9C2 cell model were established. AgomiR-NC and agomiR-101-3p were injected into mice, and then the I/R model was made; agomiR-NC and agomiR-101-3p were transferred into H9C2 cells by liposome method, and then the H/R model was made. Some cells were pretreated with dimethyl sulfoxide(DMSO) and mitogen activated kinase(MAPK) signal pathway inhibitor SB203580 for 30 min. Left ventricular end diastolic pressure(LVDP), left ventricular ejection fraction(LVEF) and left ventricular short axis shortening(LVFS) were measured by echocardiography; Cell Counting Kit 8(CCK8) and annexin v-fluorescein isothiocyanate propidium iodide(annexin V-FITC/PI) double staining were used to detect the cell proliferation rate and apoptosis rate; Double luciferase assay was used to detect the fluorescence activity of cells; The protein expressions of MAPK1, MAPK6 and MAPK8 were detected by Western blot(WB). Results: Compared with the hearts of control mice or H9C2 cells, the expression of miR-101-3p in I/R mice and H/R H9c2 cells significantly decreased. LVDP, LVEF and LVFS significantly decreased, cell proliferation rate decreased and apoptosis rate significantly increased(P<0.05). After overexpression of miR-101-3p, LVDP, LVEF and LVFS significantly increased, cell proliferation rate increased and apoptosis rate obviously decreased(P<0.05). MiR-101-3p significantly inhibited the fluorescence activity of wild-type MAPK1, MAPK6 and MAPK8 cells and negatively regulated their protein expression; MAPK signaling pathway inhibitor enhanced the proliferation promotion and apoptosis inhibition of H/R H9C2 cells after overexpression of miR-101-3p. Conclusions: miR-101-3p could protect ischemia-reperfusion myocardial injury, promote cardiomyocyte proliferation and inhibit apoptosis. Its mechanism may be related to the direct targeted inhibition of MAPK signal pathway activity.

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