The effect of microRNA-21 on myocardial fibrosis in mice with chronic viral myocarditis
10.3760/cma.j.issn.0253-3758.2018.06.008
- VernacularTitle: 微小RNA-21在小鼠慢性病毒性心肌炎心肌纤维化中的作用
- Author:
Yimin XUE
1
;
Mingguang CHEN
;
Dewei CHEN
;
Weifeng WU
;
Yanli LIU
;
Fenghui LIN
Author Information
1. Fourth Department of Critical Care Medicine, Fujian Provincial Hospital, Provincial Clinical College of Fujian Medical University, Fuzhou 350001, China
- Publication Type:Journal Article
- Keywords:
Myocarditis;
Myocardial fibrosis;
MicroRNA-21
- From:
Chinese Journal of Cardiology
2018;46(6):450-457
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To explore the effect of microRNA-21 (miR-21) on myocardial fibrosis in mice with chronic viral myocarditis (CVMC) and related mechanisms.
Methods:Forty 4-week-old Balb/c male mice were randomly divided into 4 groups (n=10 each): phosphate buffer saline (PBS) group, CVMC group, CVMC+miR-21 inhibitor group, CVMC+isotype control group. The first injection of Coxsackie virus B3 (CVB3) or PBS was performed on day 0, and the total study time was 42 days. Each mouse in CVMC group, CVMC+miR-21 inhibitor group and CVMC+isotype control group was intraperitoneally (i.p) injected with 100TCID50 CVB3 0.1, 0.15, and 0.2 ml on day 0, 14, and 28, respectively. The mice in PBS group were i.p injected with the same dose of PBS at the same time point. After the initial infection, each mouse in CVMC+miR-21 inhibitor group and CVMC+isotype control group was intravenously injected with 0.1 ml miR-21 inhibitor or 0.1 ml isotype control, on day 14 and 28. Cardiac function was measured on surviving mice of 4 groups by echocardiography on day 42. Then, the hearts were removed aseptically to observe the expressions of green fluorescence protein (GFP). The myocardial pathological changes were examined with HE, Masson staining and the myocardial pathological scores (PS), the collagen volume fraction (CVF) were calculated respectively. The levels of miR-21, collagen typeⅠ-A1 (COL1-A1) and collagen type Ⅲ-A1 (COL3-A1) mRNA in heart were detected by quantitative real-time polymerase chain reaction (RT-qPCR). Furthermore, the expressions of transforming growth factor-β1 (TGF-β1) and mothers against decapentaplegic homolog 7(Smad7) in heart were determined with Western blot assay.
Results:(1) Cardiac function in 4 groups: Compared with PBS group, left ventricular end systolic diameter (LVESD) and left ventricular end diastolic diameter (LVEDD) were markedly increased in CVMC group and CVMC+isotype control group (all P<0.05), whereas the left ventricular ejection fraction (LVEF) was decreased (P<0.05). LVESD and LVEDD were significantly decreased, and LVEF was increased in CVMC+miR-21 inhibitor group compared with those in CVMC group and CVMC+isotype control group (all P<0.05). (2) Myocardial pathological changes: The expressions of GFP in CVMC+miR-21 inhibitor group and CVMC+isotype control group were visible in heart tissues frozen sections. The hearts in CVMC group and CVMC+isotype control group were enlarged and stiff, inflammatory cells were visible and significantly increased myocardial fibrosis was evidenced in mice of these two groups. Higher PS and CVF were evidenced in CVMC group (PS: 1.14±0.69 vs. 0, CVF: (17.86±2.61)% vs. (5.70±1.42)%, all P<0.05) and CVMC+isotype control group(PS: 1.00±0.63 vs. 0, CVF: (16.78±2.58)% vs. (5.70±1.42)%, all P<0.05) compared to PBS group. Compared with CVMC group and CVMC+isotype control group, degree of cardiac fibrosis was reduced in mice of CVMC+miR-21 inhibitor group (CVF: (11.01±2.55)% vs. (17.86±2.61)%, (11.01±2.55)% vs. (16.78±2.58)%, all P<0.05), whereas PS were similar between them (PS: 0.89±0.60 vs. 1.14±0.69, 0.89±0.60 vs. 1.00±0.63, all P>0.05). (3) Cardiac expressions of miR-21, COL1-A1 and COL3-A1 mRNA: The cardiac expressions of miR-21, COL1-A1 mRNA, COL3-A1mRNA in CVMC group and CVMC+isotype control group were markedly higher than those in PBS group (all P<0.05), which were significantly downregulated in CVMC+miR-21 inhibitor group (all P<0.05 vs. CVMC group and CVMC+isotype control group). (4) The cardiac expressions of TGF-β1 and Smad7 protein: The cardiac expressions of TGF-β1 protein in CVMC group and CVMC+isotype control group were markedly higher, whereas the cardiac Smad7 protein expressions were significantly lower (all P<0.05) than those in PBS group (all P<0.05), these changes could be reversed in CVMC+miR-21 inhibitor group (P<0.05 vs. CVMC group and CVMC+isotype control group).
Conclusions:Our results suggest that miR-21 contributes to the myocardial fibrosis in CVMC mice through modulating TGF-β1/Smad7 signaling pathway.
