ABSTRACT
AIM:To investigate the role of microRNA (miR)-199a-5p in myocardial fibrosis and the potential target of miR-199a-5p.METHODS:C57BL/6 mouse cardiac fibroblasts were isolated and cultured for cellular experimen-tal study.Dual-luciferase reporter assay was performed to confirm the interaction between miR-199a-5p and the 3'-untrans-lated region (3'-UTR) of silent information regulator 1 (SIRT1).The expression of SIRT1 and fibrosis markers collagen (Col) 1a1, Col3a1 and α-smooth muscle actin (α-SMA) at mRNA and protein levels was determined by RT-qPCR and Western blot, respectively.RESULTS:The expression levels of miR-199a-5p, Col1a1, Col3a1 andα-SMA were marked-ly increased in cardiac fibroblasts after treatment with angiotensin Ⅱ(AngⅡ).The over-expression of miR-199a-5p signif-icantly increased the expression of Col1a1, Col3a1 andα-SMA in cardiac fibroblasts.Moreover, the results of dual-lucifer-ase reporter assay revealed that miR-199a-5p interacted with the 3'-UTR of SIRT1.miR-199a-5p inhibited SIRT1 expres-sion at post-transcriptional level.Meanwhile, miR-199a-5p mimic, in parallel to SIRT1 siRNA, inhibited SIRT1 expres-sion, increased the expression of Col1a1, Col3a1 and α-SMA in cardiac fibroblasts.Inactivation of NF-κB signaling con-tributed to the decrease in miR-199a-5p in Ang II-treated cardiac fibroblasts .CONCLUSION:SIRT1 is a target gene of miR-199a-5p, which mediates the pro-fibrotic effect of miR-199a-5p on cardiac fibroblasts .
ABSTRACT
AIM:To investigate the role of microRNA (miR)-199a-5p in myocardial fibrosis and the potential target of miR-199a-5p.METHODS:C57BL/6 mouse cardiac fibroblasts were isolated and cultured for cellular experimen-tal study.Dual-luciferase reporter assay was performed to confirm the interaction between miR-199a-5p and the 3'-untrans-lated region (3'-UTR) of silent information regulator 1 (SIRT1).The expression of SIRT1 and fibrosis markers collagen (Col) 1a1, Col3a1 and α-smooth muscle actin (α-SMA) at mRNA and protein levels was determined by RT-qPCR and Western blot, respectively.RESULTS:The expression levels of miR-199a-5p, Col1a1, Col3a1 andα-SMA were marked-ly increased in cardiac fibroblasts after treatment with angiotensin Ⅱ(AngⅡ).The over-expression of miR-199a-5p signif-icantly increased the expression of Col1a1, Col3a1 andα-SMA in cardiac fibroblasts.Moreover, the results of dual-lucifer-ase reporter assay revealed that miR-199a-5p interacted with the 3'-UTR of SIRT1.miR-199a-5p inhibited SIRT1 expres-sion at post-transcriptional level.Meanwhile, miR-199a-5p mimic, in parallel to SIRT1 siRNA, inhibited SIRT1 expres-sion, increased the expression of Col1a1, Col3a1 and α-SMA in cardiac fibroblasts.Inactivation of NF-κB signaling con-tributed to the decrease in miR-199a-5p in Ang II-treated cardiac fibroblasts .CONCLUSION:SIRT1 is a target gene of miR-199a-5p, which mediates the pro-fibrotic effect of miR-199a-5p on cardiac fibroblasts .
ABSTRACT
[Objective]To investigate the role and the potential target of miR-92b-3p in angiotensin Ⅱ(Ang-Ⅱ)-induced mouse cardiac hypertrophy.[Methods]Ang-Ⅱ-induced cardiac hypertrophy models were established in adult C57BL/6 mice. AgomiR-92b-3p,the cholesterol-modified miR-92b-3p mimic,was delivered to increase the level of miR-92b-3p in mouse myocar?dium via tail vein injection. In the present study,three groups of mice were used in the animal experiment as follows,the agomiR-negative control(agomiR-NC)+saline group,the agomiR-NC+Ang-Ⅱgroup and the agomiR-92b-3p+Ang-Ⅱgroup. A cell model of cardiac hypertrophy was also established in Ang-Ⅱ-induced neonatal mouse cardiomyocytes in this study Luciferase activity was assayed after transfection using a luciferase reporter assay system. The expression of Myocyte-specific enhancer factor 2D( MEF2D) and hypertrophy-related genes atrial natriuretic peptide (ANP),cardiac muscle α-actin (ACTA1) and β-myosin heavy chain (MHC)at mRNA and protein levels in Ang-Ⅱ-induced hypertrophic myocardium and cardiomyocytes were detected by qRT-PCR and Western blot,respectively.[Results]The expression of ANP,ACTA1,β-MHC were markedly increased in Ang-Ⅱ-induced hypertrophic myocardium and cardiomyocytes. Dual luciferase reporter assay revealed that MEF2D is a potential target gene of miR-92b-3p. And miR-92b-3p can reduce the expression of MEF2D at the post-transcriptional level. Functionally,miR-92b-3p mimic, consistent with MEF2D siRNA,inhibited cell size increase and protein expression of ANP,ACTA1 andβ-MHC in Ang-II-treated mouse cardiomyocytes.[Conclusions]MEF2D is a novel target of miR-92b-3p,a target gene of miR-92b-3,which mediates the ef?fect of miR-92b-3p on attenuating cardiomyocyte hypertrophy.