The Effects Of Advanced Glycation End Products On Aging And Fibrosis Of Cardiac Fibroblasts And The Role Of Pitavastatin

With aging, aging-related biochemical changes of heart lead to pathophysiological conditions. Now there are tens of theories associated to mechanism for aging, advanced glycation end products(AGEs) based nonenzymatic glycation theory for the pathogenesis of cardiac fibrosis on which we can focus is widely recognized among theories of aging for the heart. As cardiac aging is a complex process which includes aging of cardiac fibroblasts and cardiomyocyte, abnormal accumulation of extracellular matrix in the myocardial interstitial can lead to cardiac fibrosis. Cardiac fibrosis is not only one of the important mechanisms on the myocardial remodeling of coronary heart disease, hypertension and heart failure, but also is an important feature of cardiac aging. Cardiac fibrosis could cause myocardial structure disordered, cardiac compliance decreased, stiffness increased, morbidity of arrhythmia and severe refractory heart failure increased. Now, it is urgent to develop studies on pathogenesis of cardiac fibrosis, even induced by AGEs. Statins were hypolipemia drugs applied widely in clinical practice. Some recent studies have indicated that statin may be benificial to CHF patients. Pitvastatin, which is a new generation of statins, is the first fully synthetic HMG-CoA reductase inhibitor. It is lack of the research about the effects of pitavastatin on the fibrosis of cardiac fibroblasts and its mechanism of action.The present study investigated the effects of advanced glycation end products on cardiac aging and myocardial fibrosis in cultured neonatal rat cardiac fibroblasts with Western blot and β-galactosidase staining and the potential molecular mechanism. Meanwhile, the current study investigated the effects of HMG-CoA reducase inhibitor pitavastatin on myocardial fibrosis in cultured neonatal cardiac fibroblasts induced by AGEs and the potential molecular mechanisms. The study provided new ideas and experimental evidences for anti-myocardial fibrosis therapy. The present study included two parts as following:Part1:The effects of Advanced Glycation End products on cardiac fibroblasts aging and fibrosisObjective:To investigate the mechanisms of cardiac fibroblasts aging and fibrosis induced by advanced glycation end productcs(AGEs).Methods:Neonatal rat cardiac fibroblasts were incubated for72h with AGEs(200μg/ml), anti-RAGE antibody (2μg/ml) and TGFβ/smad signaling pathway inhibitor (SB431542,10μM). Senescence-associated beta galactosidase activity and p16were observed; MMP-2, TGF-β1, p-smad2/3were measured by western blot.Results:1.After intervenced with AGEs for72h, senescence-associated beta galactosidase activity, and the level of p16significantly increased compared with control group(p<0.01), accompanied with evident elevation of the expressions of TGF-β1, p-smad2/3and MMP-2;2.Senescence-associated beta galactosidase activity and the expression of p16were remarkably down-regulated after pretreatment of anti-RAGE antibody and SB431542compared with group AGEs, meanwhile, the level of TGF-β1, p-smad2/3, MMP-2were also obviously decreased.Conclusion:We conclude that AGEs might induce cardiac fibroblasts aging through binding to its receptor RAGE, while cardiac fibrosis induced by TGF-β/smad pathway might be involved in this process.Part2:The role of Pitavastatin AGEs induced myocardial fibrosis on cardiac fibroblastsObjective:To investigate the effects of pitavastatin on AGEs induced myocardial fibrosis and its mechanism of action in cultured neonatal cardiac fibroblasts.Methods:Neonatal rat cardiac fibroblasts were incubated for72h with AGEs(200μg/ml), pitavastatin(600ng/ml) and anti-RAGE antibody (2μg/ml). The proinflammatory factor-NFκB(p105/p50) and MMP-2, TGF-β1, p-smad2/3were measured by western blot.Results:1. Comparing with control group, the level of NFκB(p105/p50) significantly increased in AGEs group (p<0.01), accompanied with evident elevation of the expressions of MMP-2, and TGF-β1, p-smad2/3; 2. The expression of NFicB(p105/p50) were remarkably down-regulated after pretreatment of pitavastatin and anti-RAGE antibody compared with group AGEs, meanwhile, the level of MMP-2and TGF-β1, p-smad2/3were also obviously decreased.Conclusion:Pitavastatin could inhibit AGEs stimulated cardiac fibroblasts myocardial fibrosis via inactivating the AGEs-RAGE/NFκB pathway, pitavastatin inhibited fibroblasts myocardial fibrosis by down TGF-β, p-smad2/3, MM-2regulating expression.