缺氧预处理人华通胶间充质干细胞条件上清对心肌缺血再灌注损伤的作用及机制  

Effect and mechanism of conditioned medium from hypoxia preconditioned human Wharton’s Jelly derived mesenchymal stem cells on myocardial ischemia/reperfusion injury

作　　者：李艳[1,2] 张燕[2] 张宁坤[2] 陈宇 Li Yan;Zhang Yan;Zhang Ningkun;Chen Yu(Naval Clinical College,Anhui Medical University;The Fifth Clinical Medicine,Anhui Medical University,Hefei 230032,Anhui Province,China)

机构地区：[1]安徽医科大学海军临床学院,安徽医科大学第五临床医学院,安徽省合肥市230032 [2]解放军总医院第六医学中心,北京市100048

出　　处：《中国组织工程研究》2022年第31期5008-5013,共6页Chinese Journal of Tissue Engineering Research

基　　金：国家自然科学基金项目(81370238),项目负责人:陈宇。

摘　　要：背景:间充质干细胞主要通过旁分泌作用改善组织损伤,低氧预处理后其旁分泌作用增强,但关于人华通胶间充质干细胞低氧预处理后的条件上清液对心肌缺血再灌注损伤的作用及其机制仍需深入研究。目的:探讨缺氧预处理人华通胶间充质干细胞条件上清对心肌缺血再灌注损伤H9C2细胞凋亡的影响及机制。方法:常氧、低氧处理人华通胶间充质干细胞,获得条件上清;大鼠H9C2细胞通过缺氧复氧和无血清培养6 h模拟体内心肌缺血再灌注损伤,试剂盒检测H9C2细胞培养上清中乳酸脱氢酶活力。将大鼠H9C2细胞分为4组:常氧培养组、模型组、常氧条件上清组和低氧条件上清组,造模后给予常氧条件上清和低氧条件上清干预24 h后,流式细胞仪检测细胞凋亡率,Western blot检测各组H9C2细胞中凋亡相关蛋白Bax、Bcl-2和自噬相关蛋白Beclin-1、p62、LC3的表达。结果与结论:①经缺氧复氧无血清处理及加入人华通胶间充质干细胞条件上清后,H9C2细胞形态无明显变化;②与常氧培养组相比,模型组H9C2细胞上清液中乳酸脱氢酶活力明显升高,加入两种人华通胶间充质干细胞条件上清后乳酸脱氢酶活力均下降;③与模型组相比,常氧条件上清组和低氧条件上清组H9C2细胞凋亡减少,Bax、Beclin-1、LC3蛋白表达降低,p62蛋白表达升高,且低氧条件上清组改变较常氧条件上清组显著;④结果表明,缺氧预处理人华通胶间充质干细胞条件上清可减少心肌缺血再灌注损伤导致的H9C2细胞凋亡,且该保护作用可能与其抑制自噬有关。BACKGROUND:Mesenchymal stem cells improve tissue injury mainly through their paracrine action,which is enhanced by hypoxic preconditioning.However,the effect of conditioned medium from hypoxia preconditioned human Wharton’s Jelly derived mesenchymal stem cells on myocardial ischemia/reperfusion injury and its mechanism still need to be further studied.OBJECTIVE:To investigate the effect and mechanism of conditioned medium from hypoxia preconditioned human Wharton’s Jelly derived mesenchymal stem cells on the apoptosis of H9C2 cells after ischemia/reperfusion injury.METHODS:Human Wharton’s Jelly derived mesenchymal stem cells were treated with normoxia or hypoxia to obtain conditioned medium.The H9C2 cells were subjected to hypoxia/reoxygenation and serum-free culture for 6 hours to mimic the myocardial ischemia/reperfusion injury model in vitro.Lactate dehydrogenase activity in H9C2 conditioned medium was detected using lactate dehydrogenase assay kit.Rat myocardial H9C2 cells were divided into four groups:normoxic culture group,model group,normoxic conditioned medium group and hypoxic conditioned medium group.After treatment with normoxic or hypoxic conditioned medium for 24 hours after hypoxia,the apoptosis rate of H9C2 cells was detected by flow cytometry.Western blot assay was applied to detect the expression of apoptotic markers(Bax and Bcl-2)and autophagic markers(Beclin-1,P62 and LC3)in each group.RESULTS AND CONCLUSION:(1)The morphology of H9C2 cells did not change significantly after treatment with ischemia/reperfusion or human Wharton’s Jelly derived mesenchymal stem cells-conditioned medium.(2)Compared with the normoxic culture group,lactate dehydrogenase activity in supernatant of H9C2 cells in the model group was significantly increased,but decreased in both conditioned medium groups.(3)Compared with the model group,the apoptosis rate of H9C2 and the expression of Bax,Beclin-1,and LC3 protein were downregulated and the expression of p62 protein was upregulated in H9C2 cells in the normoxic co

关 键 词：华通胶间充质干细胞 缺血再灌注损伤 自噬 缺氧预处理 心肌梗死 

分 类 号：R459.9[医药卫生—治疗学] R319[医药卫生—临床医学]