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1.
Journal of Sun Yat-sen University(Medical Sciences) ; (6): 642-650, 2023.
Artigo em Chinês | WPRIM | ID: wpr-979218

RESUMO

ObjectiveTo investigate the effect of myosin heavy chain 7 gene-derived miRNA-208b-3p on the fibrotic phenotype of cardiac fibroblasts. MethodsmiRNA chip array was performed to detect the dysregulated miRNAs in the myocardium of diabetic db/db mice and db/m control mice. Neonatal mouse ventricular cardiomyocytes (NMVCs) and cardiac fibroblasts (CFs) were isolated from C57BL/6 mice and cultured. Real-time quantitative PCR (RT-qPCR) was conducted to determine the expression of miR-208b-3p in mouse CFs and NMVCs subjected to angiotensinⅡ(AngⅡ) and high glucose plus glucose oxidase (G/Go) treatment, respectively. Cell counting kit 8(CCk8) assay, flow cytometry and determination of fibrosis-related protein, including COL1A1, COL3A1and α-SMA, were performed in mCFs transfected with miR-208b-3p. Dual luciferase reporter assay was performed to confirm the interaction between miR-208b-3p and the 3'-UTR of metal response element binding transcription factor 2 (Mtf2) and progesterone receptor membrane component 1(Pgrmc1), respectively. The expressions of Mtf2 and Pgrmc1 at the mRNA and protein levels in mCFs after miR-208b-3p mimic transfection were determined using RT-qPCR and Western blot assay, respectively. The small interfering RNA (siRNA) was used to inhibit Mtf2 and Pgrmc1 expression in mCFs, and the effects of Mtf2 siRNA, Pgrmc1 siRNA and miR-208b-3p on fibrosis-related protein expression in mCFs were investigated. ResultsResults of miRNA chip array and RT-qPCR assay showed that miR-208b-3p was up-regulated in the myocardium of the diabetic db/db mice. miR-208b precursor and the host gene of Myh7 were consistently increased in db/db mice. miR-208b-3p and Myh7 mRNA were expressed in mCFs and NMVCs, but the levels of miR-208b-3p and Myh7 mRNA in NMVCs were much higher than those in mCFs. miR-208b-3p was up-regulated in mCFs and NMVCs subjected to Ang Ⅱ and G/Go treatment, respectively. miR-208b-3p could significantly enhance fibrosis-related protein, including COL1A1, COL3A1 and α-SMA, in mCFs, without affecting the proliferation activity and cell cycle distribution of mCFs. Dual luciferase reporter assay revealed the interactions of miR-208b-3p with the 3'-UTR of Mtf2 and Pgrmc1. The results of RT-qPCR and Western blotting confirmed that miR-208b-3p inhibited Mtf2 and Pgrmc1 expression at the post- transcriptional level. Transfection with miR-208b-3p mimic, Mtf2 siRNA and Pgrmc1 siRNA could consistently enhance the fibrosis-related protein expression in the cardiac fibroblasts. ConclusionsmiR-208b-3p enhances fibrosis-related gene expression by targeting Mtf2 and Pgrmc1in mCFs.

2.
Chongqing Medicine ; (36): 153-155, 2018.
Artigo em Chinês | WPRIM | ID: wpr-691756

RESUMO

Objective To investigate the role of miR-208b-3p in dexmedetomidine(DEX) alleviating rat myocardial ischemia reperfusion injury(MIRI).Methods Eighty adult male Wister rats were divided into the sham operation(Sham) group,I/R group,DEX group and miR-208b-3p+ DEX group.In the Sham group,a 6-0 silk suture was only placed without ligation,and other 3 groups were treated by I/R model.Before ischemia reperfusion,the DEX group received a loading dose of DEX(5 μg/kg) via the femoral vein followed by a continuous infusion of 5 μg · kg-1 · h-1 for 1 h.In the miR-208b-3p+ DEX group,rats received intramuscular injection of miR-208b-3p analogue at 24 h before ischemia reperfusion.The cardiac function indexes were monitored at 120 min after reperfusion,including heart rate(HR),left ventricular systolic blood pressure(LVSP),left ventricular end diastolic pressure(LVEDP) and left ventricular rate maximum(dp/dtmax).The serum levels of cTn-Ⅰ and CK-MB were detected and the apoptosis rate(AI) was measured by in situ apoptosis assay.The levels of oxidative stress related indicators were detected and Western blot was used to detect the level of myocardial apoptosis protein.Results Compared with the Sham group,the cardiac function in the I/R group was decreased,but the levels of CTn-Ⅰ,CK-MB,AI,MDA,Bax and Caspase-3 were increased(P<0.05);compared with the I/R group,the above indexes in the DEX group were improved(P<0.05);compared with the DEX group,the above indexes in the miR-208b-3p+DEX group were deteriorated(P<0.05).Conclusion Overexpression of miR-208b-3p can eliminate the protective effect of DEX on MIRI.

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