Angiotensin Ii-stimulated Increase In Cell Growth And Collagen Secretion Of Cardiac Fibroblasts Is Inhibited By Simvastatin :the Molecular Signalling Mechanisms

Background & Objective:The main pathological characteristics of myocardial fibrosis were proliferation of cardiac fibroblasts, increased collagen synthesis and irregular deposition of extracellular matrix components. Cardiac fibroblasts play a pivotal role in the process of formation of myocardial fibrosis. AngII has multiple effects on cardiac fibroblasts: It induces cardiac fibroblast proliferation, differentiation and synthesis multifunctional cytokines such as osteopontin and ?3integrin,activation of nuclear transcription factors NF B and AP-1 .In this study, we evaluated AngII effects on rat cardiac fibroblasts in order to explore the pathophysiological and biochemical mechanisms involved in the development of cardiac fibrosis.Hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase inhibitors (statins) are well-established therapies in the prevention and treatment of atherosclerotic cardiovascular disease.Increasing evidence has indicated that statins ameliorated Ang II–induced cardiac hypertrophy, fibrosis, and cardiac remodeling independently of cholesterol reduction. In viro study , atorvastatin directly inhibit proliferation and collagen production of cardiac fibroblasts. But the molecular mechanisms for its effect are far from being elucidated. In the present study ,using cultured rat cardiac fibroblasts, we tested whether simvastatin 1) inhibits cardiac fibroblast proliferation and differentiation 2) blocks NF B and AP-1 activation by cardiac fibroblasts stimulated with Ang II 3) affects the expression of osteopontin and ?3 integrin of cardiac fibroblasts,in an attempt to elucidate a possible benefic effect of statins for the inhibition of collagen production and provide theoretical bases of preventing and treating cardiac myofibrosis.Methods: Third to fifth passage of neonatal rat cardiac fibroblasts were used in the experiment and divided into 5 groups. (A) CFs treated with Ang II (1×10-7 mol/L) contatins vehicle (B) CFs treated with vehicle (C) CFs co-treated with simvastatin (1×10-7 mol/L) and Ang II (1×10-7 mol/L)(D)CF co-treated with simvastatin (1×10-6 mol/L) and Ang II (1×10-7 mol/L) (E) CF co-treated with simvastatin (1×10-5 mol/L) and Ang II (1×10-7 mol/L)Cardiac fibroblast proliferation was measured by MTT assay. Immunocytochemistry was used to measure the production ofα-smooth muscle actin, ?3 integrin ,collagen type I and III .The expression of osteopontin ,NF B P50 Subunit and AP-1 c-Jun Subunit protein levers were evaluated by western blot and immunocytochemistry .The nuclear translocation of transcription factors(NF- B and AP-1) was determined by immunocytochemistry .Results:1 Simvastatin (10-7mol/L-10-5mol/L) inhibited the Ang II-induced cardiac fibroblast proliferation, differentiation and production of collagen type I and III (P<0.01) .2 After 48 hours stimulation, Ang II(10-7mol/L) induced significant upregulation of osteopontin(P<0.05) and ?3 integrins(P<0.01). Simvastatin(10-6 mol/L-10-5 mol/L)attenuated Ang II-induced upregulation of osteopontin(P<0.01). Simvastatin ( 10-7 mol/L-10-5 mol/L)attenuated Ang II-induced upregulation of ?3 integrins(P<0.01).3 AngII up-regulated AP-1, NF- B activity by increasing nuclear translocation of c-Jun and P50 this effect was almost completely abolitied by simvastatin (10-5 mol/L).4 Ang II treatment (10-7 mol/L) markedly up-regulated production of NF B P50 subunit and AP-1 c-Jun subunit proteins . This effect was blocked by simvastatin (10-7 mol/L-10-5 mol/L) (P<0.05)Conclusion:Simvastatin suppressed proliferation ,differentiation and collagen type I and III secretion of cardiac fibroblasts. The effect is possibly mediated by inhibition of a molecular signalling pathway, including ?3 integrin, osteopontin and nuclear transcription factors (NF- B and AP-1).