摘要
目的探讨长链非编码RNA(lncRNA)FGD5-AS1与微小RNA-421(miR-421)的靶向调控关系及其对缺氧/复氧(H/R)诱导的心肌细胞损伤的影响。方法利用H/R诱导心肌细胞H9C2损伤,采用实时荧光定量PCR检测lncRNAFGD5-AS1和miR-421表达。在心肌细胞中转染pcDNA-FGD5-AS1、anti-miR-421,或共转染pcDNAFGD5-AS1和miR-421,之后进行H/R损伤。采用试剂盒评估丙二醛(MDA)含量和超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-Px)活性,流式细胞术分析凋亡率,Westernblotting法测定B细胞淋巴瘤/白血病-2(Bcl-2)和Bcl-2相关X蛋白(Bax)蛋白表达。生物信息学预测与双荧光素酶报告实验验证lncRNAFGD5-AS1对miR-421的靶向调控。结果缺氧/复氧使心肌细胞中lncRNAFGD5-AS1表达、SOD活性、GSH-Px活性、Bcl-2蛋白表达减少,并使miR-421表达、MDA含量、凋亡率、Bax蛋白表达增多(P均<0.05)。lncRNAFGD5-AS1过表达降低缺氧/复氧诱导的心肌细胞的MDA含量、凋亡率、Bax蛋白表达,提高SOD活性、GSH-Px活性、Bcl-2蛋白表达(P均<0.05),抑制miR-421表达与其效果相同。lncRNAFGD5-AS1靶向调控miR-421的表达。共转染pcDNA-FGD5-AS1和miR-421提高缺氧/复氧诱导的心肌细胞的MDA含量、凋亡率、Bax蛋白表达,降低SOD活性、GSH-Px活性、Bcl-2蛋白表达(P均<0.05)。结论在缺氧/复氧诱导的心肌细胞中,lncRNAFGD5-AS1通过靶向miR-421,减轻细胞氧化应激,并抑制其凋亡,从而减轻缺氧/复氧诱导的心肌细胞损伤。
Objective To investigate the targeted regulation of long noncoding RNA(lncRNA)FGD5-AS1 on miRNA-421(miR-421)and its effect on cardiomyocyte injury induced by hypoxia/reoxygenation(H/R).Methods H/R was used to induce the damage of cardiomyocytes H9C2.Real-time fluorescence quantitative PCR(qRT-PCR)was used to detect the expression of lncRNA FGD5-AS1 and miR-421.We transfected pcDNA-FGD5-AS1,anti-miR-421,or co-transfected pcDNA-FGD5-AS1 and miR-421 into cardiomyocytes,and then performed H/R injury.The kits were used to evaluat content of malondialdehyde(MDA)and activity of superoxide dismutase(SOD)and glutathione peroxidase(GSH-Px).Flow cytometry was used to measure the apoptosis rate of cells,and Western blotting was applied to determine the protein expression of B cell lymphoma/lewkmia-2(Bcl-2)and Bcl-2 associated X protein(Bax).Bioinformatics prediction and dual luciferase report experiment verified the targeted regulation of lncRNA FGD5-AS1 on miR-421.Results Hypoxia/reoxygenation reduced the expression of lncRNA FGD5-AS1,activity of SOD and GSH-Px and expression of Bcl-2 protein,while increased the expression of miR-421,MDA content,apoptosis rate,expression of Bax protein in the cardiomyocyte,and the difference was statistically significant(all P<0.05).Overexpression of lncRNA FGD5-AS1 decreased the MDA content,apoptosis rate,and Bax protein expression of cardiomyocytes induced by hypoxia/reoxygenation,while increased activity of SOD,GSH-Px,and expression of Bcl-2 protein,and the difference was statistically significant(all P<0.05),and it was the same as the result of inhibiting the expression of miR-421.lncRNA FGD5-AS1 targeted and regulated the expression of miR-421.Co-transfection of pcDNA-FGD5-AS1 and miR-421 increased the MDA content,apoptosis rate,Bax protein expression of cardiomyocytes induced by hypoxia/reoxygenation,and decreased SOD,GSH-Px activity,and Bcl-2 protein expression.The differences were statistically significant(all P<0.05).Conclusion In cardiomyocytes induced by hypoxia/reoxygenation,lncRNA FGD5-AS1 can reduce oxidative stress and inhibit apoptosis of cardiomyocytes by targeting miR-421,thereby reducing injury of cardiomyocytes induced by hypoxia/reoxygenation.
作者
孟晓京
刘亚军
项宁
MENG Xiaojing;LIU Yajun;XIANG Ning(Qinhuangdao First Hospital,Qinhuangdao 066000,China)
出处
《山东医药》
CAS
2021年第16期30-34,共5页
Shandong Medical Journal
