The Roles And Mechanisms Of EphrinB2,IL-17A In Cardiac Remodeling After Myocardial Injury

Part Ⅰ.EphrinB2 Regulates Cardiac Fibrosis Through Modulating the Interaction of Stat3 and TGF-β/Smad3 SignalingPurpose:Cardiac fibrosis is a common feature in left ventricular remodeling that leads to heart failure,regardless of the cause.EphrinB2(erythropoietin-producing hepatoma interactor B2),a pivotal bidirectional signaling molecule ubiquitously expressed in mammals,is crucial in development and angiogenesis during disease progression.Recently,EphrinB2 was reported to protect kidneys from injury-induced fibrogenesis but promote skin and lung fibrosis after injury.However,its role in cardiac fibrosis remains to be clarified.We sought to determine the role of EphrinB2 in cardiac fibrosis and the underlying mechanisms during the pathological remodeling process.Methods and Results:EphrinB2 was highly expressed in the myocardium of patients with advanced heart failure,as well as in mouse models of myocardial infarction and cardiac hypertrophy induced by angiotensin Ⅱ infusion,which was accompanied by myofibroblast activation and collagen fiber deposition.In contrast,intramyocardial injection of lentiviruses carrying EphrinB2.shRNA ameliorated cardiac fibrosis and improved cardiac function in mouse model of myocardial infarction.Furthermore,in vitro studies in cultured cardiac fibroblasts demonstrated that EphrinB2 promoted the differentiation of cardiac fibroblasts into myofibroblasts in normoxic and hypoxic conditions.Mechanistically,the profibrotic effect of EphrinB2 on cardiac fibroblast was determined via activating the Stat3(signal transducer and activator of transcription 3)and TGF-β(transforming growth factor-β)/Smad3(mothers against decapentaplegic homolog 3)signaling.We further determined that EphrinB2 modulated the interaction between Stat3 and Smad3 and identified that the MAD homology 2 domain of Smad3 and the coil-coil domain and DNA-binding domain of Stat3 mediated the interaction.Conlusion:This study uncovered a previously unrecognized profibrotic role of EphrinB2 in cardiac fibrosis,which is achieved through the interaction of Stat3 with TGF-β/Smad3 signaling,clarifying the further mechanism of cardiac remodeling and implying a promising therapeutic target in heart failure.Part Ⅱ.The Roles and Mechanisms of Interleukin-17A and Stat3 in Cardiomyocyte Apoptosis after Myocardial Ischemia/Reperfusion Injury in MicePurpose:Cardiomyocyte apoptosis is a key pathological mechanism of myocardial ischemia/reperfusion(I/R)injury.Interleukin-17A,a pro-inflammatory cytokine,has a direct proapoptotic effect on cardiomyocytes.However,the specific mechanism has not been clarified.Methods and Results:In the present study,an in-vitro model of cardiomyocyte apoptosis induced by IL-17A stimulation was employed and the roles of iNOS and Stat3 involved were investigated.Our data showed that the neonatalmouse cardiomyocytes express IL-17 receptors:IL-17RA and IL-17RC,but did not express IL-17A.Exogenous IL-17A significantly induces iNOS expression and hence the cardiomyocyte apoptosis.Moreover,IL-17A-induced cardiomyocyte apoptosis can be achieved directly via iNOS activation.We further showed that exogenous IL-17A simultaneously triggers Stat3 activation,which in turn inhibits IL-17A-induced iNOS expression in cardiomyocytes.Both ChIP and dual-luciferase results confirmed that Stat3 directly inhibits transcriptional activities of iNOS via binding to its specific promoter region.Consistent with these data,silencing of Stat3 in fact can aggravate IL-17A-triggered cardiomyocyte apoptosis.Finally,using an in vivo myocardial I/R injury mouse model,we verified that Stat3 inhibition increased iNOS expression and exacerbated cardiomyocyte apoptosis.Conlusion:Our study strongly support the notion that Stat3 plays a compensatory anti-apoptotic role in IL-17A/iNOS-mediated cardiomyocyte apoptosis via inhibiting iNOS transcription,providing a novel molecular mechanism of apoptosis regulation and complicated interactions between IL-17A/iNOS and IL-17A/Stat3 signalings.