转染HACE1基因过表达或敲降病毒质粒对急性氧化应激损伤心肌细胞活性、凋亡及HO-1表达的影响OACSTPCD
Effects of transfection with HACE1 gene overexpression or knockdown viral plasmids on viability,apoptosis,and HO-1 expression in cardiomyocytes with acute oxidative stress injury
目的 观察转染HACE1基因过表达或敲降病毒质粒对急性氧化应激损伤心肌细胞H9C2细胞活性、凋亡及急性氧化应激过程中血红素加氧酶(HO-1)表达的影响.方法 取H9C2细胞分成对照组、HACE1过表达组、HACE1敲降组,HACE1过表达组、HACE1敲降组H9C2细胞分别转染HACE1基因过表达病毒质粒HACE1-OE、HACE1基因敲降慢病毒质粒HACE1 shRNA.三组细胞继续培养72 h后,应用缺氧小室将细胞缺氧培养4 h后复氧2 h模拟心肌细胞急性氧化应激损伤过程,构建急性氧化应激损伤心肌细胞模型.采用CCK-8法测算各组细胞活性,采用Hoechest33342染色法测算各组细胞凋亡率,采用Western blotting法检测各组细胞中氧化应激指标HO-1蛋白.结果 对照组、HACE1过表达组、HACE1敲降组细胞活性分别为67.0%±4.4%、75.2%±4.2%、56.7%±3.0%,组间相比,P均<0.05.对照组、HACE1过表达组、HACE1敲降组细胞凋亡率分别为18.90%±1.02%、13.30%±0.86%、28.70%±0.67%,组间相比,P均<0.05.对照组、HACE1过表达组、HACE1敲降组细胞中HO-1蛋白相对表达量分别为1.00±0.17、0.71±0.19、1.96±0.28,组间相比,P均<0.05.结论 转染HACE1基因过表达病毒质粒可提高急性氧化应激损伤心肌细胞H9C2的细胞活性,抑制其凋亡及氧化应激指标HO-1蛋白的表达;转染HACE1基因敲降病毒质粒可降低急性氧化应激损伤心肌细胞H9C2的细胞活性,促进其凋亡及氧化应激指标HO-1蛋白的表达.
Objective To investigate the effects of transfection with HACE1 gene overexpression or knockdown viral plasmids on cell viability,apoptosis,and heme oxygenase-1(HO-1)expression in cardiomyocytes H9C2 with acute oxi-dative stress injury.Methods H9C2 cardiomyocytes were divided into three groups:control group,HACE1 overex-pression(HACE1-OE)group,and HACE1 knockdown(shRNA-HACE1)group.The cells in the HACE1-OE group were transfected with the HACE1 overexpression viral plasmid,and cells in the shRNA-HACE1 group were transfected with HACE1 knockdown viral plasmid(shRNA-HACE1),respectively.After cells were all cultured for 72 h under nor-moxic condition,cells were then cultured for 4 h in a hypoxic chamber,followed by reoxygenation for 2 h to simulate acute oxidative stress injury in H9C2 cardiomyocytes.Cell viability was assessed using CCK-8,the apoptosis rate in each group was measured via Hoechst 33342 staining,and HO-1 protein expression level as an indicator of oxidative stress in-jury was detected by Western blotting.Results The cell viability in the control group,HACE1-OE group,and shRNA-HACE1 group was 67.0%±4.4%,75.2%±4.2%,and 56.7%±3.0%,respectively,with statistically significant dif-ferences between these groups(all P<0.05).Apoptosis rates of the control group,HACE1-OE group,and shRNA-HACE1 group were 18.90%±1.02%,13.30%±0.86%,and 28.70%±0.67%,and there were statistically signifi-cant differences between these groups(all P<0.05).The relative expression levels of HO-1 protein in the control group,HACE1-OE group,and shRNA-HACE1 group were 1.00±0.17,0.71±0.19,and 1.96±0.28,and there were statistically significant differences between these groups(all P<0.05).Conclusion Transfection with HACE1 gene overexpression viral plasmid in H9C2 cardiomyocytes with acute oxidative stress injury significantly promotes cell viability,inhibits apoptosis,and reduces HO-1 protein expression;conversely,transfection with HACE1 gene knock-down viral plasmid leads to decreased cell viability accompanied by increased apoptosis rate and elevated HO-1 protein expression.
王梦;宋辉;姚恒臣
山东第一医科大学附属中心医院心血管内科,济南 250013山东第一医科大学附属聊城市人民医院心内科
基础医学
HACE1基因心肌细胞急性氧化应激心肌损伤血红素加氧酶-1
HACE1 genecardiomyocytesacute oxidative stressmyocardial damageheme oxygenase-1
《山东医药》 2024 (033)
7-10 / 4
山东省自然科学基金面上项目(ZR2016HM49).
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