Protective Effect of miR-200a on Myocardium of Elderly Diabetic Cardiomyopathy Rats by Up-regulating Nrf2
YANG Xiaojing
XIAO Ting
CAO Chen
DENG Xiangqun
Abstract：Objective:To investigate the protective effect of microRNA-200a(miR-200a)on myocardium of elderly diabetic rats and its mechanism.Methods:The rats diabetic cardiomyopathy model was induced by high-fat diet+streptococcin(STZ)and divided into control group and STZ group.miR-200a was overexpressed in myocardium by injecting miR-200a-carrying adeno-associated virus 9(AAV9),which was divided into control+AAV9-control group,control+AAV9-miR-200a group,STZ+AAV9-control group,and STZ+AAV9-miR-200a group,with 8 rats in each group.The cardiomyocytes H9c2 of rats were cultured in vitro and divided into control group(5.5 mmol/L glucose for 24 h),high glucose+AAV9-control+si RNA group(transfected with AAV9-control and si RNA for 48 h and then cultured with 33 mmol/L glucose for 24 h),high glucose+AAV9-control+si nuclear factor E2-related factor 2(Nrf2)group(transfected with AAV9-control and si Nrf2 for 48 h and incubated with 33 mmol/L glucose for 24 h),high glucose+AAV9-miR-200a+si RNA group(transfected with AAV9-miR-200a and si RNA for 48 h and incubated with 33 mmol/L glucose for 24 h),high glucose+AAV9-miR-200a+si Nrf2 group(transfected with AAV9-miR-200a and si Nrf2 for 48 h and then incubated with 33 mmol/L glucose for 24 h).The levels of superoxide dismutase(SOD),creatine kinase isoenzyme(CK-MB),lactate dehydrogenase(LDH),and malondialdehyde(MDA)were detected using enzyme-linked immunosorbent assay(ELISA).miR-200a expression was detected by real-time fluorescence quantitative polymerase chain reaction(PCR).The protein expression was detected by immunoblotting.The level of reactive oxygen species(ROS)was detected by staining with 2',7'-dichlorodihydrofluorescein diacetate(DCFH-DA),and Nrf2 expression was detected by immunofluorescence.Results:Compared with the control group,miR-200a expression of myocardial tissue downregulated in the STZ group(P<0.05).Compared with the control+AAV9-control group,levels of LDH,CK-MB,and MDA in the STZ+AAV9-control group were significantly higher,and SOD,heme oxidase-1(HO-1),cytosolic Nrf2 protein,cytoplasmic Nrf2 protein expression,and Nrf2 fluorescence density were significantly lower(P<0.05).Compared with the STZ+AAV9-control group,levels of LDH,CK-MB,MDA and TUNEL-positive cell rate were significantly lower in the STZ+AAV9-miR-200a group,and SOD,HO-1,cytosolic Nrf2 protein,cytoplasmic Nrf2 protein expression and Nrf2 fluorescence density were significantly higher(P<0.05).High-glucose treatment could decrease the survival rate of H9c2 cells and increase the levels of ROS,MDA,and LDH(P<0.05).The levels of the above indexes were opposite expressed after transfection of miR-200a.The simultaneous transfection of miR-200a and si Nrf2 resulted in the elevation of ROS,MDA,and LDH,and the decrease of cell survival rate(P<0.05).Conclusion:miR-200a enhanced Nrf2 expression to attenuate oxidative stress and apoptosis induced by high glucose,which was related to the promotion of Nrf2 nuclear translocation and its downstream anti-oxidative stress signal pathway.
Keywords：diabetic cardiomyopathymicroRNA-200anuclear factor E2 related factor 2oxidative stressratsexperimental study
Publication Date：2024-05-16
Online Publishing Date：2025-08-15(First online date of this platform, not the publication date of the document)
Pages：6( 1777-1782 )