| Objectives:After myocardial infarction(MI),myocardial ischemia and hypoxia,produce oxidative stress and inflammatory response,leading to cardiomyocyte apoptosis,cardiac function decline,meanwhile,skeletal muscle loss,and the quality of life decrease.Therefore,the treatment and rehabilitation of MI have become the top priority in protecting human health.Follistatin-like protein 1(FSTL1)is a secreted glycoprotein that inhibits cardiomyocyte apoptosis,improves cardiac function,promotes vascular regeneration in damaged skeletal muscle,and plays an important role in the protection of heart and skeletal muscle.Exercise can inhibit cardiomyocyte apoptosis and skeletal muscle loss,and up-regulate FSTL1 expression in myocardium and skeletal muscle of MI rats.Whether myocardium FSTL1 expression in MI rats up-regulated by resistance training comes from skeletal muscle?What is the specific signaling mechanism of it inhibits cardiomyocyte apoptosis?Whether skeletal muscle loss inhibited by resistance training is related to skeletal muscle FSTL1?What is the specific signal mechanism?Little literature has been reported.The aim of this study is to investigate the variation characteristics of circulating and follistatin-like protein 1(FSTLJ)in myocardium and skeletal muscle of rats with myocardial infarction(MI)by resistance training.In addtion to up-regulate FSTL1 by resistance training andthe relationship between expression and improvement of cardiac function and inhibition of MI caused skeletal muscle loss in rats,and explore its biological mechanism.Methods:The:ale Sprague-Dawley rats,weight about 180-220g were randomly divided into five groups(n=10):Sham-operated group(S),sedentary MI group(MI),MI with resistance training group(MR),MI with adeno-associated virus empty vector group(MV)and MI with FSTL1 adeno-associated virus group(MF)after the MI model established by left anterior descending(LAD)coronary artery ligation.S group underwent threading without ligation.1 week after surgery,rats in MR group subjected resistance training for 4 weeks,rats in MV and MF group were injected adeno-associated virus empty vector and FSTL1 adeno-associated virus in the tibialis anterior muscle of the left limb,respectively.The next day after training,rats were anesthetized and heart function was measured,the heart and left limb tibialis anterior muscle were extracted.Collagen volume fraction(%)of myocardium were observed and calculated by Masson staining;skeletal muscle cells proliferation,the FSTL1 expression in myocardium and skeletal muscle were measured by immunofluorescence;cardiomyocyte apoptosis was detected by TUNEL staining;skeletal muscle cell cross sectional area was determined by HE staining.mRNA expression of.fstl1 and dip2a in myocardium and skeletal muscle were detected by RT-qPCR.The protein content of FSTL1 in serum,the protein expression of FSTL1,DIP2A,p-Akt/Akt,p-mTOR/mTOR in myocardium and skeletal muscle,Bcl2/Bax in myocardium and p-P70 S6K/P70 S6K in skeletal muscle were measured by Western blotting.After Human recombinant FSTL1(rhFSTL1),AMPK activator AICAR,PI3K inhibitor LY294002 and LPS were intervented,H9C2 cells were divided into H9C2 cells control group,H9C2 with LPS group,H9C2 with LPS with LY294002 group,H9C2 with LPS with rhFSTL1 group,H9C2 with LPS with rhFSTL1 with LY294002 group,H9C2 with LPS with AICAR group,and H9C2 with LPS with AICAR with LY294002 group,H9C2 with LPS with rhFSTL1 with AICAR group.After rhFSTL1,AMPK activator AICAR,PI3K inhibitor LY294002 were intervented,C2C12 cells were divided into C2C12 cells control group,C2C12 with LY294002 group,C2C12 with AICAR group,C2C12 with LY294002 with AICAR group,C2C12 with rhFSTL1 group,C2C12 with LY294002 with rhFSTL1 group.H9C2 cell apoptosis was detected by TUNEL staining,H9C2 cell viability and C2C12 cell proliferation were determined by CCK8 assay;the protein expression of FSTL1,DIP2A,p-Akt/Akt,p-mTOR/mTOR,Bcl2/Bax,p-P70 S6K/P70 S6K were measured by Western blotting.Results:(1)Resistance training increased skeletal muscle fstl1 mRNA,there was no significant effecton myocardium.fstl1 mRNA in MI rats.Resistance training or fstl1 overexpression in skeletal muscle significantly increased skeletal muscle and myocardium FSTL1 expression and circulating FSTL1 level of MI rats,and there was a significant positive correlation between skeletal muscle FSTL1 and myocardium FSTL1 expression and circulating FSTL1 level.It is speculated that skeletal muscle may be an important source of myocardium FSTL1 protein.(2)Resistance training or fstl1 overexpression in skeletal muscle significantly decreased the number of TUNEL positive cells,increased the Bcl2/Bax ratio in MI myocardium and significantly inhibited cardiomyocyte apoptosis,there was a significantly negative correlation between the number of TUNEL positive cells and myocardium FSTL1 expression.(3)Resistance training or.fstl1 overexpression in skeletal muscle inhibited collagen fibrosis,decreased fibrosis area,improved cardiac function in MI rats,there was a significantly negative correlation between CVF%and myocardium FSTL1 expression.(4)Resistance training or fstll overexpression in skeletal muscle significantly increased myocardium FSTL1 receptor DIP2A expression and the p-Akt/Akt and p-mTOR/mTOR ratio in MI rats,and activated FSTL1/DIPZA-Akt/mTOR pathway.(5)AICAR significantly increased the expression of FSTL1 and its receptor DIP2A in LPS-induced H9C2 cells,significantly activated Akt/mTOR signaling pathway,inhibited LPS-induced H9C2 cells apoptosis and increased cell viability.(6)Exogenous rhFSTL1 significantly increased the expression of FSTL1 and its receptor DIP2A in H9C2 cells,significantly activated Akt/mTOR signaling pathway,inhibited LPS-induced H9C2 cells apoptosis and increased cell viability.(7)Resistance training or fstl1 overexpression in skeletal muscle significantly increased skeletal muscle FSTL1 expression,skeletal muscle relative weight,skeletal muscle cell cross sectional area and the number of skeletal muscle ki67-labeled cells,improved skeletal muscle 1oss in MI rats.There was a significantly positive correlation between FSTL1 protein and skeletal muscle weight/body weight ratio,weight/tibialis length ratio and cross sectional area.(8)Resistance training or fstl1 overexpression in skeletal muscle significantly increased DIP2A gene and protein expression,the p-Akt/Akt,p-mTOR/mTOR and p-P70 S6K/P70 S6K ratio in skeletal muscle of MI rats,and activated FSTL1/DIP2A-Akt/mTOR/P70 S6K signaling pathway.(9)AICAR intervention significantly increased the number of C2C12 cells per unit area,FSTL1 and its receptor DIP2A expression,exogenous rhFSTL1 significantly up-regulated C2C12 cells DIP2A expression.(10)Exogenous rhFSTL1 or AICAR intervention significantly increased the ratio of p-Akt/Akt,p-mTOR/mTOR and p-P70 S6K/P70 S6K,activated Akt/mTOR/P70 S6K signaling pathway,and promoted the proliferation of C2C12 cells.Conclusions:(1)Resistance training effectively stimulates the secretion of FSTL1 by skeletal muscle,increases circulating and cardiac FSTL1 levels,inhibits cardiomyocyte apoptosis,reduces reduce myocardium fibrosis,improves cardiac function and inhibits skeletal muscle loss in MI rats.(2)Activation of FSTL1/DIP2A-Akt/mTOR signaling pathway is an important mechanism that resistance training inhibits cardiomyocyte apoptosis,reduces myocardium fibrosis,improves cardiac function in MI rats and promotes skeletal muscle cells proliferation,inhibits skeletal muscle loss caused by MI.This study will provide experimental basis for the exercise rehabilitation methods and target screening of myocardial infarction. |
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