The Molecular Mechanisms By Which RAGE New Ligand CTRP1 And HMGB2 Promote The Progression Of Coronary Artery Disease

ObjectiveCoronary artery disease(CAD)is a dynamic process characterized by endothelial dysfunction,atherosclerosis formation,myocardial ischemia,and finally ends up with plaque rupture or chronic heart failure.Recently,increasing studies have revealed that innate immunity and chronic inflammation play pivotal roles in the progression of CAD The pattern recognition receptor RAGE has been reported to play multiple roles in modulating oxidative stress and inflammatory responses.By using proteomic screening analysis,we for the first time identified CTRP1 and HMGB2 as novel ligands for RAGE Several studies have indicated that CTRP1 and HMGB2 are involved in the regulation of oxidative stress and inflammation.However,the role of CTRP1 and HMGB2 in cardiovascular diseases is still unclear.The present study is aimed to determine the effects and corresponding molecular mechanisms of CTRP1 and HMGB2 in the pathogenesis of CAD and myocardial infarction.Our findings should aid in the improvement of our understandings of the mechanisms underlying CAD and identification of novel targets for treating CAD patientsMethodsEffects and mechanisms of RAGE new ligand CTRP1 on CAD were investigated from below 3 aspects1)CTRP1 promotes lipid accumulation and inflammation in macrophagesPBMC were isolated from healthy adults,we investigated the changes in CTRP expression levels during vital biological processes in macrophages and their relation to inflammatory responses2)CTRP1 impaires endothelial diastolic function via arginase-1 C57BL/6 or CTRP1 knockout mice were administered intraperitoneally with recombinant mouse CTRP1 or normal saline,then we evaluated endothelial-dependent vasodilation by microcirculatory studies and aortic ring assay.Mechanisms by which CTRP1 impairs endothelial diastolic function were tested in HUVEC.3)CTRP1 promotes atherosclerosis:We measured CTRP1 levels in sera,atherosclerotic plaques and peripheral blood mononuclear cells from CAD patients and health controls.Expression of inflammatory factors was assessed by quantitative RT-PCR and western blot analysis.Mechanisms that CTRP1 promotes leucocyte-endothelium interactions were tested in vitro and in vivo.Apolipoprotein E-deficient(apoE-/-)male mice were administered intraperitoneally with recombinant mouse CTRP1 or normal saline.After 6 mouths,the area of atherosclerotic lesions was quantified in apoE-/-and CTRP1-/-/apoE-/-mice by Oil Red O staining,respectively.On the other hand,we investigated the effects and mechanisms of HMGB2 on CAD.1)HMGB2 serum levels were measured in STEMI patients and health controls.We analyzed the correlation between HMGB2 levels and the severity of myocardial infarction and major adverse cardiac events(MACE)after 1 month.2)The effect of HMGB2 on myocardial hypoxia injury was studied on rat MI models and hypoxic H9C2 cell lines.3)H9C2 was treated with HMGB2 in combination with or without NADPH oxidase inhibitor apocynin.Reactive oxygen species(ROS)production was determined by fluorescence microscopy.We evaluated the effects of HMGB2 on inflammation,apoptosis and the impairment of autophagosome clearance by molecular biology methods.Results1)CTRP1 serum level was related to the severity of atherosclerosis.2)CTRP1 was up-regulated upon exposure to oxidized low-density lipoprotein(oxLDL)in a peroxisome proliferator-activated receptor(PPAR)-dependent manner.Stimulation of CTRP1 led to enhanced lipid accumulation and inflammation factors expression in cultured human macrophages.3)CTRP1 increased arginase-1 expression in a RAGE-dependent way,which causing NO synthesis decrease and endothelial dependent diastolic dysfunction4)In vivo and in vitro studies confirmed that CTRP1 promoted eNOS uncoupling and endothelial ROS production5)CTRP1 promotes atherosclerosis and vascular inflammation6)Increased serum HMGB2 levels are associated with MI severity and MACE at 1 month7)HMGB2 administration enhanced ROS production in ischemic myocardium and hypoxic H9C2 cells,with aggravation of apoptosis,inflammation,and impairment of autophagosome clearanceConclusion1)The novel RAGE ligand CTRP1 promotes atherosclerosis by increasing macrophage lipid accumulation,impairing vascular functions,and disturbing vascular homeostasis2)The novel RAGE ligand HMGB2 promotes myocardial ischemic injury in rats and hypoxic H9C2 cells through enhancing inflammation,apoptosis and impairment of autophagosome clearance.