| 高英英.微 RNA?29b在心肌缺血再灌注损伤中的作用及机制研究[J].安徽医药,2019,23(12):2463-2467. |
| 微 RNA?29b在心肌缺血再灌注损伤中的作用及机制研究 |
| The role and mechanism of miR?29b in myocardial ischemia/reperfusion injury |
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| DOI：10.3969/j.issn.1009?6469.2019.12.033 |
| 中文关键词: 再灌注损伤 心肌缺血 蛋白质丝氨酸苏氨酸激酶 细胞凋亡 小鼠，近交系 微 RNA?29b Akt信号通路 |
| 英文关键词: Reperfusion injury Myocardial ischemia Protein?serine?threonine kinases Apoptosis Mice,inbred strains MicroRNA?29b Akt signaling pathway |
| 基金项目: |
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| 摘要点击次数: 2022 |
| 全文下载次数: 641 |
| 中文摘要: |
| 目的探讨微 RNA?29b(miR?29b)在心肌缺血 /再灌注(Ischemia/Reperfusion，I/R)损伤过程中是否发挥保护作用并探讨其可能的调控机制。方法通过 100 μmol/L叔丁基过氧化氢(TBHP)诱导 H9C2心肌细胞损伤构建 I/R细胞模型；分别设立正常对照组， TBHP处理组， NC miRNA处理组， miR?29b类似物(mimic)处理组。通过实时荧光定量多聚核苷酸链式反应(qPCR)检测各组心肌细胞内 miR?29b表达量；人胆囊收缩素 /缩胆囊素八肽(CCK?8)检测各组心肌细胞的存活率；蛋白质印迹法(West? ern Blot)检测心肌细胞中凋亡蛋白和蛋白激酶 B(Akt)蛋白磷酸化水平的改变。另一方面，将 30只 C57/B6小鼠(6~7周龄)逐个编号，通过随机数字表法抽样，随机分为三组，设立 I/R组、 NC miRNA处理组和 miR?29b激动剂(agomir)处理组，其中 miR? 29b agomir组小鼠预先给予 miR?29b agomir治疗。结扎冠状动脉左前降支 30 min，然后再灌注 4h建立心肌缺血再灌注动物模型。通过埃文斯蓝(Evans)和 2，3，5?氯化三苯基四氮唑(TTC)双重染色评估三组小鼠心肌梗死面积； qPCR检测各组小鼠心肌组织内 miR?29b表达量；蛋白质印迹法检测心肌组织内凋亡蛋白改变。结果细胞层面:与 TBHP处理组相比(65±3)%，上调 miR?29b表达可增加心肌细胞的存活率(85±3)%，下调心肌细胞凋亡蛋白表达(0.86±0.07)。进一步的蛋白质印迹法实验结果表明:与 TBHP处理组(0.81±0.05)相比，上调 miR?29b能够抑制 Akt蛋白磷酸化(0.41±0.02)。动物层面: miR?29b agomir处理组小鼠心肌梗死面积(24±3)%与 I/R组(62±2)%相比明显减少，且心肌组织凋亡蛋白表达明显减少(0.32±0.04)。结论上调 miR?29b表达可以减少缺血再灌注损伤引起的心肌细胞凋亡。 miR?29b可能是通过调控靶基因 Akt磷酸化水平，下调心肌细胞凋亡水平，从而保护心脏。 |
| 英文摘要: |
| Objective To investigate the changes on miR?29b expression in myocardial ischemia/reperfusion injury and to discusswhether there will be protective effects on myocardial tissue after up?regulating miR?29b expression，as well as to elucidate its pos? sible mechanism.Methods By inducing H9C2 cardiomyocyte injury by 100 μmol/L TBHP，the ischemia?reperfusion cell model was established.Then the normal control group，TBHP?treated group，NC miRNA?treated group and miR?29b mimic?treated group were established respectively.On the one hand，the expression of miR?29b in cardiomyocytes was detected by qPCR and the surviv?al rate of cardiomyocytes in each group was measured by CCK?8.Besides，the changes of apoptotic proteins as well as the phosphor? ylation of Akt protein in cardiomyocytes were detected by Western Blot.On the other hand，according to the method of random num? ber table，30 C57/B6 mice(6 to 7 weeks old)were randomly divided into three groups:an Ischemia/Reperfusion(I/R)group，a NC miRNA?treated group and a miR?29b agomir?treated group，mice in which were pretreated with miR?29b agomir.After anesthe? tizing the mice，the left anterior descending coronary artery was ligated for 30 minutes，and then reperfused for 4 hours to establish an animal model of myocardial ischemia?reperfusion.The myocardial infarction size of the three groups was evaluated by TTC and Evan’s blue staining.The expression of miR?29b in the myocardial tissue of each group was detected by qPCR.And the apoptosis protein in myocardial tissue was measured by Western Blot.Results Cell level:Compare with the TBHP?treated group(65±3)%， up?regulation of miR?29b expression could increase myocardial cell viability(85±3)% and down?regulate the expression cardio? myocyte apoptosis protein(0.86 ± 0.07).Further Western Blot results showed that compare with the TBHP?treated group(0.81 ± 0.05)，miR?29b can inhibit Akt protein phosphorylation(0.41±0.02).Animal level:Compared with I/R group(62±2)%，the myo? cardial infarction size of mice treated with miR?29b agomir(24±3)%，and the expression of myocardial apoptosis protein were sig? nificantly decreased(0.32±0.04).Conclusion Overexpression of miR?29b can reduce myocardial cell apoptosis induced by isch?emia?reperfusion injury.miR?29b may protect the heart by regulating the phosphorylation level of the target gene Akt and down?reg?ulating the level of myocardial apoptosis. |
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