Effect of RORα overexpression viral vector on doxorubicin-induced cardiotoxicity in mice and its mechanism
Objective To investigate the effect of retinoid-related orphan receptor α(RORα)overexpression viral vector on doxorubicin-induced cardiotoxicity(DIC)in mice and to explore its potential mechanism.Methods Eighty male C57BL/6 mice were randomly divided into four groups:Con group,DIC group,DIC+RORα group and DIC+RORα+EX527 group,with 20 mice in each group.Mice in the DIC+RORα group and DIC+RORα+EX527 group were injected with ade-no-associated virus type 9(AAV9)vector that overexpressed RORα through the tail vein,while mice in the DIC group were injected with the blank control vector of AAV9.After three weeks of AAV9 vector injection,the DIC mouse model was established by DOX injection through tail vein except for the Con group.DIC+RORα+EX527 group was intraperitone-ally injected with SIRT1 specific inhibitor EX527 after each DOX injection.After 4 weeks,left ventricular ejection frac-tion(LVEF)and left ventricular fractional shortening(LVFS)were measured using cardiac ultrasonography.Orbital blood and myocardial tissues were taken after death.HE staining was used to observe the pathological changes of myocardial tis-sues.Serum creatine kinase-MB(CK-MB),lactate dehydrogenase(LDH),cardiac troponin I(cTnI),interleukin-1β(IL-1β),interleukin-6(IL-6),and tumor necrosis factor-α(TNF-α)were detected by ELISA.The apoptosis rate was calculat-ed after TUNEL staining,the expression of reactive oxygen species(ROS)in myocardial tissue was detected by DHE stain-ing,the expression levels of superoxide dismutase(SOD),glutathione peroxidase(GSH-Px)and malondialdehyde(MDA)in the myocardial tissues were detected by colorimetry method,and the macrophage infiltration and NF-κB p65 expression in the myocardial tissues were observed by immunofluorescence staining.The protein expression levels of cleaved Caspase-3,RORα,and SIRT1 were detected by Western blotting,and Ac-NF-κB p65/NF-κB p65 and Ac-foxo1/foxo1 were calcu-lated.The mRNA expression levels of RORα and SIRT1 in the myocardial tissues were detected by real-time fluorescent quantitative PCR.Results Compared with the Con group,myocardial fibers in the DIC group were disordered and bro-ken,and the degree of vacuolation of cardiomyocyte was significant.Compared with the DIC group and DIC+RORα+EX527 group,the degree of myocardial fiber breakage and cardiomyocyte vacuolation were relieved in the DIC+RORα group.LVEF,LVFS,SOD,and GSH-Px were as follows:Con group>DIC+RORα group>DIC group,DIC+RORα+EX527 group(all P<0.05).Serum CK-MB,LDH,cTnI,IL-1β,IL-6,TNF-α,myocardial apoptosis rate,cleaved Caspase-3 protein,ROS,MDA,macrophage infiltration,and NF-κB were as follows:Con group<DIC+RORα group<DIC group,DIC+RORα+EX527 group(all P<0.05).RORα mRNA and protein were in the following order:Con group,DIC+RORα group,DIC+RORα+EX527 group>DIC group(all P<0.05).SIRT1 mRNA and protein were as follows:Con group,DIC+RORα group>DIC group,DIC+RORα+EX527 group(all P<0.05).Ac-foxo1/foxo1 and Ac-NF-κB p65/NF-κB p65 were as fol-lows:Con group,DIC+RORα group<DIC group,DIC+RORα+EX527 group(all P<0.05).Conclusion RORα overex-pression viral vector can alleviate DOX-induced myocardial injury,and its mechanism may be related to inhibiting myocar-dial oxidative stress and inflammatory response through activation of SIRT1 signaling pathway.
万方数据
维普
