CRAMP Protects Against Myocardial Ischemia-reperfusion Injury And Its Underlying Mechanism

Objective: Cathelicidin related antimicrobial peptide(CRAMP)controls immune and inflammatory reactions,and is associated with cell apoptosis.Myocardial ischemia-reperfusion injury(IRI)induces oxidative stress and inflammatory infiltration,thus leading to cardiomyocyte apoptosis and necrosis.However,the role of CRAMP in myocardial IRI remains unclear.Methods: In this project,we used the enzyme-linked immunosorbent assay(ELISA)to determine the level of CRAMP in myocardium and serum of murine model of myocardial IRI induced by coronary artery constriction,and in serum from acute myocardial infarction patients.In vitro,we performed gain-and loss-of function assays by CRAMP preconditioning and si RNA to CRAMP,and used tunel staining to investigate oxygen-glucose deprivation reperfusion(OGDR)-induced apoptosis in neonatal rat cardiac myocytes(NRCM).In vivo,we used CRAMP intraperitoneal injection and CRAMP knockout mouse models to measure the myocardial infarct size of IRI based on triphenyltetrazolium chloride(TTC)staining and myocardial apoptosis by tunel staining.We then examined the ERK1/2-AKT-Fox O3 a signaling pathway by immunoblotting.Finally,we used the inhibitor to MEK-ERK(PD98059)and AKT(MK-2206)to rescue the function of CRAMP.Results: Our ELISA data indicated that CRAMP was downregulated in infarct area(23.96%)and serum(54.57%)of mouse myocardial IRI,and the serum(53.81%)of human acute myocardial infarction patients.In addition,tunel staining demonstrated that CRAMP could reduce OGDR-induced apoptosis in NRCM by 35.61%.In contrast,the si RNA to CRAMP increased apoptosis by 30.73%.Consistently,myocardial infarct size and myocardial apoptosis was reduced by 34.33% and 52.97% respectively with CRAMP injection preconditioning.Inversely,CRAMP knockout mice increased infarct size by 37.34% and myocardial apoptosis by 77.37%.Moreover,we found that CRAMP activated phosphorylation of ERK1/2 and AKT and reduced the ratio of Bax/Bcl-2 and Cleaved Caspase-3/ Caspase-3.Functional rescue expriments showed that ERK1/2 and AKT phosphorylation mediated CRAMP protective effect in myocardial IRI.Finally,we found that CRAMP could activate phosphorylation of Fox O3 a and increased nuclear export of Fox O3 a in the condition of OGDR.Conclusion: CRAMP protects against myocardial IRI and apoptosis.CRAMP might be a novel target for the treatment of myocardial IRI.