褪黑素对H9c2心肌细胞缺氧/复氧损伤的保护作用及其机制  

Protective effects and mechanism of melatonin against H9c2 cardiomyocyte injury induced by hypoxia/reoxygenation

作　　者：杨吉平[1] 费琳[2] 钟钰西 余蕾[1] 苟兴春[1] YANG Jiping;FEI Lin;ZHONG Yuxi;YU Lei;GOU Xingchun(Institute of Basic Medical Sciences,Shaanxi Key Laboratory of Ischemic Cardiovascular Disease,Xi’an Medical University,Xi’an 710021,China;Departments of Psychiatry,First Affiliated Hospital of Xi’an Jiaotong University)

机构地区：[1]西安医学院基础医学研究所,陕西省缺血性心血管疾病重点实验室,西安710021 [2]西安交通大学第一附属医院精神科

出　　处：《山西医科大学学报》2020年第8期761-765,共5页Journal of Shanxi Medical University

基　　金：国家自然科学基金资助项目(81971330);陕西省教育厅重点实验室项目(19JS061);西安医学院大学生创新创业项目(2018DC-26)。

摘　　要：目的研究褪黑素(MLT)对H9c2心肌细胞缺氧/复氧(H/R)损伤的保护作用,并探讨其机制。方法将H9c2心肌细胞随机分为对照组、H/R组、MLT组和受体相互作用蛋白激酶3抑制剂(GSK-872)组。对照组用不含药物的DMEM处理,H/R组对H9c2细胞进行缺氧4 h/复氧6 h处理,MLT组和GSK-872组细胞在H/R前2 h分别给予100μmol/L MLT和5μmol/L GSK-872预处理。采用CCK-8法检测细胞活力,试剂盒测定培养液中乳酸脱氢酶(LDH)的活性,碘化丙啶(PI)染色观察细胞坏死情况,Western blot检测RIPK3和磷酸化钙离子/钙调蛋白依赖蛋白激酶(p-CaMK)Ⅱ的表达。结果与对照组比较,H/R组细胞存活率显著下降(P<0.01),培养液中LDH含量明显升高(P<0.01),坏死细胞数量显著增多(P<0.01),RIPK3和p-CaMKⅡ蛋白表达水平显著升高(P<0.01)。与H/R组比较,MLT组和GSK-872组细胞存活率明显提高(P<0.01),LDH含量显著降低(P<0.01),坏死细胞明显减少(P<0.01),RIPK3和p-CaMKⅡ蛋白的表达明显下调(P<0.01)。而MLT组与GSK-872组上述指标比较差异均无统计学意义(P>0.05)。结论褪黑素可减轻H9c2心肌细胞的H/R损伤,其机制可能与抑制RIPK3/CaMKⅡ信号通路有关。Objective To investigate the effects and mechanism of melatonin(MLT)on H9c2 cardiomyocyte injury induced by hypoxia/reoxygenation(H/R).Methods H9c2 cardiomyocytes were randomly divided into control group,H/R group,MLT group and receptor interaction protein kinase 3(RIPK3)inhibitor group(GSK-872).H9c2 cardiomyocytes in control group were treated with drug-free DMEM.H9c2 cardiomyocytes were treated with hypoxia for 4 h and reoxygenation for 6 h in H/R group,and the cells were pretreated with 100μmol/L MLT and 5μmol/L GSK-8722 h before H/R treatment in MLT group and GSK-872 group,respectively.CCK-8 assay was used to detect the cell vitality.The activity of lactate dehydrogenase(LDH)in medium was determined by LDH kit.Cell necrosis was observed with propidiumiodide(PI)staining.The expression levels of RIPK3 and phosphorylated Ca 2+/calmodulin-dependent protein kinase(p-CaMK)Ⅱwere detected by Western blot.Results Compared with control group,the cell viability dropped significantly in H/R group(P<0.01),LDH content in culture medium was raised significantly(P<0.01),the number of necrotic cells was increased significantly(P<0.01),and the expression levels of RIPK3 and p-CaMKⅡwere increased significantly(P<0.01).Compared with H/R group,the cell viability were obviously improved in MLT group and GSK-872 group(P<0.01),LDH content was decreased significantly(P<0.01),the number of necrotic cells was reduced significantly(P<0.01),and the expression levels of RIPK3 and p-CaMKⅡwere significantly decreased(P<0.01).However,there was no significant difference in the above indexes between MLT group and GSK-872 group(P>0.05).Conclusion MLT can protect against H9c2 cardiomyocyte injury induced by H/R,which may be related to the inhibition of RIPK3/CaMKⅡpathways.

关 键 词：褪黑素 H9C2心肌细胞 缺氧/复氧损伤 受体相互作用蛋白激酶3 钙离子/钙调蛋白依赖蛋白激酶Ⅱ 

分 类 号：R363[医药卫生—病理学]