文章摘要

载脂蛋白M在心肌细胞缺氧/复氧损伤中的表达差异及其作用

作者: 1程港丽, 1姚霜, 1施媛萍, 1张俊, 1罗光华, 1郑璐
1 苏州大学附属第三医院临床医学研究中心,江苏 常州 213003
通讯: 郑璐 Email: qz.zhenglu@163.com
DOI: 10.3978/j.issn.2095-6959.2022.04.001
基金: 江苏省自然科学基金(BK20191158);江苏省333人才项目(B 2020158);常州市国际合作项目(CZ20190022)。

摘要

目的:探讨载脂蛋白M(apolipoprotein M,ApoM)在大鼠H9C2心肌细胞缺氧/复氧(hypoxia/reoxygenation,H/R)损伤中的表达差异及作用。方法:利用安宁包缺氧体系构建大鼠H9C2心肌细胞H/R损伤模型,采用CCK-8、细胞凋亡检测试剂盒、caspase-3活性检测试剂盒检测H9C2细胞H/R后细胞增殖、凋亡的差异,同时检测ApoM和1-磷酸鞘氨醇受体1(sphingosine-1-phosphate receptor 1,S1PR1)在H/R条件下的表达情况。建立体外过表达ApoM(apoM overexpression,ApoM-OE)的H9C2细胞,采用CCK-8检测心肌细胞存活率,比色法检测细胞培养上清中乳酸脱氢酶(lactate dehydrogenase,LDH)、超氧化物歧化酶(superoxide dismutase,SOD)活性,蛋白质印迹法检测心肌细胞cleaved caspase-3、caspase-3表达情况,分析ApoM在H/R诱导的心肌细胞损伤中的可能作用。结果:在H/R处理后,H9C2心肌细胞活力逐渐降低,细胞凋亡率、caspase-3活性升高,证实大鼠H9C2心肌细胞H/R损伤模型构建成功;在H/R处理后,心肌细胞中ApoM和S1PR1mRNA表达水平显著上调(均P<0.01)。与对照组相比,ApoM-OE组心肌细胞存活率、LDH活性、SOD活性、及cleaved caspase-3表达水平均无明显变化(P>0.05)。结论:ApoM和S1PR1在H9C2细胞H/R损伤中表达水平升高,但ApoM对H/R诱导的H9C2细胞损伤无明显的保护作用。
关键词: 载脂蛋白M;缺氧/复氧损伤;1-磷酸鞘氨醇

Expression difference and role of apolipoprotein M in hypoxia/reoxygenation-induced cardiomyocyte injury

Authors: 1CHENG Gangli, 1YAO Shuang, 1SHI Yuanping, 1ZHANG Jun, 1LUO Guanghua, 1ZHENG Lu
1 Clinical Medical Research Center, Third Affiliated Hospital of Soochow University, Changzhou Jiangsu 213003, China

CorrespondingAuthor: ZHENG Lu Email: qz.zhenglu@163.com

DOI: 10.3978/j.issn.2095-6959.2022.04.001

Foundation: This work was supported by the Natural Science Foundation of Jiangsu Province (BK20191158), “333” Talent Project of Jiangsu Province (B 2020158), and the International Cooperation Foundation of Changzhou City (CZ20190022), China.

Abstract

Objective: To investigate the differential expression and the effect of apolipoprotein M (ApoM) on H9C2 rat cardiomyocyte injury induced by hypoxia/reoxygenation (H/R). Methods: Using the AnaeroPack System, we established the H/R injury model of H9C2 rat cardiomyocytes. CCK-8 cell viability assay, cell apoptosis and caspase-3 activity assay were used to detect the difference of cell proliferation and apoptosis after H/R injury. Furthermore, the expression levels of ApoM and sphingosine-1-phosphate receptor 1 (S1PR1) were detected. H9C2 cardiomyocytes overexpressing ApoM (ApoM-OE) were established. CCK-8 assay was used to detect the survival rate of cardiomyocytes, the colorimetric method was applied to detect the activities of lactate dehydrogenase (LDH) and superoxide dismutase (SOD) in the cell culture supernatant, immunoblotting was used to determine the expression levels of cleaved caspase-3 and caspase-3, and the role of ApoM in H/R-induced cardiomyocyte injury was analyzed. Results: The H9C2 cardiomyocyte viability decreased gradually after H/R, while apoptosis rate and caspase-3 activity increased after H/R. It confirmed that the H/R injury model of H9C2 rat cardiomyocytes was successfully established through AnaeroPack System. The mRNA expression levels of ApoM and S1PR1 were upregulated in cardiomyocytes after H/R (both P<0.01). Compared with the NC group, the cell survival rate, HDL activity, SOD activity and relative expression level of cleaved caspase-3 in the ApoM-OE group were no significant difference (P>0.05). Conclusion: The expression levels of ApoM and S1PR1 were upregulated in H9C2 after H/R, but ApoM had no significant protective effect on H9C2 rat cardiomyocytes injured by H/R.
Keywords: apolipoprotein M; hypoxia/reoxygenation injury; sphingosine-1-phosphate