| Myocardial infarction(MI)is the main cause of cardiac dysfunction and sudden death.Myocardial infarction-induced ventricular remodeling and neural remodeling lead to cardiac morphology,structural and functional abnormalities,which even resulte in heart failure,malignant arrhythmia and sudden death.Therefore,inhibition of myocardial infarction-induced ventricular remodeling and neural remodeling may improve the severe complications after MI.ActivinA is a homodimer protein and belongs to a member family of the transforming growth factor-beta(TGF-?)superfamily.A large number of studies have reported that serum concentrations of ActivinA and creatinine as well as myocardial infarct size are consistent with left ventricular ejection fraction with acute myocardial infarction(AMI)patients[1,2].In patients and animals exhibiting heart failure post-MI,ActivinA is as a proinflammatory and fibrogenic factor to involve in the pathological process of post-MI heart failure,which promotes myocardial infarction-induced ventricular remodeling and ultimately lead to deterioration of cardiac function[3].In patients with ST-elevation MI,high levels of ActivinA 6 months after MI were associated with low left ventricular ejection fraction and high left ventricular end-diastolic volume,and ActivinA level is more greater than 129 pg/ml to be able to predicte left ventricular remodeling and all-cause mortality of ST-elevation MI patients at post-MI 3 years[1].Studies have also shown that serum high concentrations of ActivinA are important damage factors in diabetic cardiomyopathy,dilated cardiomyopathy,and valvular disease patients[4,5,6,7,8,9].These studies strongly demonstrate that ActivinA participated in the pathological process of cardiovascular diseases.But there are no reports that how ActivinA participates in MI-induced interstitial fibrosis and whether to participate in MI-induced nerve remodeling,and what mechanisms are involved in these pathologies is little known.In this study,we induced rat MI model via ligating the anterior descending of coronary artery,at the same time,we cultured the SD adult rat CF with different drug treatment,for example,recombinant rat ActivinA,ActivinA inhibitor follistatin and MAPKs signaling pathways inhibitors.We observeed that whether ActivinA influenced in post-MI ventricular remodeling(interstitial fibrosis and nerve remodeling)and myocardial function,and further explored its possible mechanisms.Part I:Effects of ActivinA on ventricular remodeling and cardiac function after rat MIObjective:In the current clinical and experimental studies,the relationship between ActivinA and post-MI cardiac function mainly focused on related studies.Therefore,we block the endogenous ActivinA to further observe whether it can regulate the pathology process of post-MI ventricular remodeling and cardiac function.Methods:SD Rat MI model was established by left anterior descending coronary artery ligation.Our experiment all the rats were divided into three groups and treated with drugs.The sham group(the anterior descending coronary artery was not ligated),MI group(intraperitoneal injection of sodium carboxymethyl cellulose)and follistatin group(intraperitoneal injection of ActivinA specific inhibitor follistatin).The survival rate of the rats was calculated at different time points.At 4 weeks postoperatively,the basal indexes of the three groups were measured:body weight,heart rate,relative lung weight ratio and relative cardiac weight ratio;cardiac struction were assessed by transthoracic echocardiography and cardiac function were measured by hemodynamics;HE staining observed the changes of myocardial tissue.Results:(1)It found no difference in survival rate,body weight,heart rate and mean arterial pressure among the different groups(p>0.05).(2)Compared with the relative lung weight ratio and relative cardiac weight ratio of the sham group,it significantly increased in the MI group(p<0.05).However,the follistatin group was lower than that of the MI group(p<0.05).(3)The transthoracic echocardiography and hemodynamic indexes of the rats in the MI group obviously deteriorated compared to the sham group(p<0.05),but the indexes of follistatin group were obviously perfected than those in the MI group(p<0.05).(4)HE staining results displayed that the infarct area were significantly thinner,showing a small amount of residual cardiomyocytes,and compensatory hypertrophy of cardiomyocytes in the peri-infarcted area in the MI group.However,follistatin group was significantly improved compared with the MI group(p<0.05),and the infarct size obviously decreased(p<0.05).Conclusion:Inhibition of endogenous ActivinA can improve ventricular remodeling and cardiac dysfunction after MI in rats.Part 2:Effect of ActivinA on myocardial interstitial fibrosis after rat MIObjective:Post-MI activated CF proliferate and differentiate into myofibroblasts,and which synthesize and release large amounts of extracellular matrix,such as collagen I and collagen III,leading to myocardial interstitial fibrosis after MI[10,11,12].Studies have reported that ActivinA can induce the proliferation and differentiation of human lung fibroblasts,kidney fibroblasts and NRK-49F cells[13,14].Moreover,blocking endogenous ActivinA can reduce CCL4-induced liver fibrosis and bleomycin-induced rat pulmonary fibrosis[15,16].Therefore,We speculate that blocking the endogenous ActivinA may inhibit post-MI the activation of cardiac fibroblasts,thereby reduce post-MI myocardial interstitial fibrosis,and then improve the ventricular structure remodeling,finally reverse cardiac dysfunction.The aim of this study was to investigate whether endogenous ActivinA can regulate the post-MI myocardial interstitial fibrosis and its possible mechanism.Methods:SD Rat MI model was established by left anterior descending coronary artery ligation.Our experiment all the rats were divided into three groups and treated with drugs.The sham group(the anterior descending coronary artery was not ligated),MI group(intraperitoneal injection of sodium carboxymethyl cellulose)and follistatin group(intraperitoneal injection of ActivinA specific inhibitor follistatin).At 4 weeks postoperatively,after surviving SD rats were subjected to echocardiography and hemodynamics.After the rats were quickly anesthetized,immediately collected heart blood and heart samples.(1)The myocardial interstitial fibrosis was observed by Masson staining.(2)collengenI,ERK1/2 and p38-MAPK signal pathway activation related protein expression in the peri-infarcted zone was detected by western blot.In addition,we cultured the SD adult rat CF with different drug treatment,for example,ActivinA,ERK1/2 and p38-MAPK inhibitor,as well as recombinant rat ActivinA(rActivinA),we aim to further observe this phenomenon,experimental method:CCK8 and EdU detected CF proliferation,Western blot detected the protein expression of a-smooth muscle actin(a-SMA),collengenl,p-ERK1/2 and p-p38-MAPK.Results:(1)For animal model,compared with the sham group,the protein expression of collengenl,p-ERK1/2 and p-p38-MAPK in the peri-infarcted zone were significantly higher than in MI group(p<0.05).While,above factors were lower in follistatin group than in MI group(p<0.05).For cell model,(2)ActivinA was expressed and secreted in normal adult rats CF,and the expression and secretion of ActivinA significantly increased in the presence of Ang? compared with the sham group(p<0.05).The above effect can be reversed by follistatin(p<0.05).At the same time,compared with the Ang? group,follistatin could down-regulated the differentiation,differentiation,collengen I expression,ERK1/2 and p38-MAPK pathway activation of Ang?-induced CF(p<0.05).Further research found that,in basal or Ang?-induced CF,recombinant ActivinA was able to upregulate the CF differentiation,differentiation,collengenI expression as well as activation of ERK1/2 and p38-MAPK pathways(p<0.05).Interestingly,SB203580(p38-MAPK inhibitor)and SB431542(ALK4 inhibitor,ALK4 is a ActivinA-specific receptor)significantly reverse these effects(p<0.05);PD98059(ERK1/2 inhibitor)can only be attenuated CF proliferation and collengenI expression(p<0.05)and it had no effect on CF differentiation(p>0.05).Importantly,the model treated with Ang? was more pronounced related to model treated normal cell(p<0.05).Conclusion:Blocking the endogenous ActivinA can improve the myocardial interstitial fibrosis after myocardial infarction,its potential mechanism may inhibit post-MI activation of ERK1/2 and p38-MAPK signaling pathways and then weaken the post-MI CF proliferation and differentiation.Part 3:Effects of ActivinA on sympathetic remodeling after rat MIObjective:Inflammatory response plays an important role in the process of sympathetic remodeling after MI,and sympathetic remodeling is the main mechanism of post-MI arrhythmia and sudden death[17].ActivinA is a dimeric protein that plays an important role in cell inflammation and immune regulation[18].ActivinA activates the NF-?B pathway[19]and NF-kB pathway also promotes the production of ActivinA[20].Importantly,inhibition of NF-kB pathway can reverse inflammation-mediated left ventricular remodeling and cardiac dysfunction after MI[21].Studies have also found that exogenous ActivinA can promote the expression of glutamate hydroxylase(TH)in striatum cells[22],but also can induce dopamine gene transcription and secretion of norepinephrine[23].Therefore,we speculated that ActivinA may promote the inflammatory response through the NF-?B pathway and lead to sympathetic remodeling after MI.The purpose of this study is to investigate whether endogenous ActivinA can regulate the infiltration of macrophages(ED1)and a-SMA-induced myofibroblasts,the tumor necrosis factor-a(TNF-a)and interleukin-1?(IL-1?)protein expression and NF-?B inflammation signaling pathway activity in the peri-infarcted zone.Methods:SD Rat MI model was established by left anterior descending coronary artery ligation.Our experiment all the rats were divided into three groups and treated with drugs.The sham group(the anterior descending coronary artery was not ligated),MI group(intraperitoneal injection of sodium carboxymethyl cellulose)and follistatin group(intraperitoneal injection of ActivinA specific inhibitor follistatin).At 4 weeks postoperatively,after surviving SD rats were subjected to echocardiography and hemodynamics.After the rats were quickly anesthetized,immediately collected heart blood and heart samples.(1)Immunohistochemistry was used to detect the expression of ED1,?-SMA,tyrosine hydroxylase(TH)and growth associated protein 43(GAP43)positive cells in the peri-infarcted zone.(2)Western blot was used to detect the protein expression of IL-1?,TNF-a,p-p 65 and p-IkBa in the peri-infarcted zone(P<0.05).(3)Western blot was also used to detect the protein expression of nerve growth factor(NGF),TH and GAP43 in the peri-infarcted zone.Results:(1)The infiltration of ED1 positive cells and a-SMA positive cells in the peri-infarcted zone was increased significantly in sham group compared to the MI group(p<0.05),while the infiltration of ED1 positive cells and a-SMA positive cells in the follistatin group was lower than that in the MI group(p<0.05).Compared with the sham group,the levels of IL-1p,TNF-a,p-p 65 and p-IkBa in the myocardium of the peri-infarcted zone in MI group were significantly higher(p<0.05),while the above indexes in follistatin group were lower than in the MI group(p<0.05).(3)Compared with the sham group,the levels of TH,GAP43 and NGF were increased in the MI group,while the expression of TH,GAP43 and NGF was significantly decreased in the follistatin group compared with the MI group(p<0.05).Conclusion:Blocking endogenous ActivinA can reduce the post-MI inflammatory response by inhibiting NF-?B signaling pathway,and further reversed post-MI the sympathetic remodeling. |
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