Experimental Study Of The Apoptosis Of Cardiac Fibroblasts Induced By Hydrazine And The Protection Of NADH

Objective: To observe the apoptosis of cardiac fibroblasts cultured in vitro induced by hydrazine and investigate the apoptosis mechanism of cardiac fibroblasts induced by hydrazine; detect the protection of NADH to cardiac fibroblast injured by hadrazine.Methods: Cardiac fibroblasts from neonatal rat were separated, cultured and divided in groups.each group incubated for 72 hours by 1,2,3,4,8,16 mM (mmolA) hydrazine, NADH cured in 400umol/L.Then, MTT assayed the proliferation of cardiac fibroblast affected by hydrazine .The neucleus changes of apoptosis of cardiac fibroblast stained by Hoechst33258 was observed through fluorescence microscope. The rates of apoptosis and necrosis of cardiac fibroblast stained by Annexin V-FITC \PI Apoptosis Detect Kit I were detected by flowcytometry.The mitochondria membrane potential of cells stained by rhodamine123 was assayed by flowcytometry.The expression of caspase-3,Bax,Bcl-2 were detected by western-blot.Result: Each group incubated for 72 hours by 1,2, 3, 4, 8, 16 mM (mmol/l) hydrazine compared with the control:Hydrazine inhibited the proliferation of cardiac fibroblasts and could induced cardiac fibroblasts apoptosis . The neucleus of cardiac fibroblasts appeared neucleus aggregation phenomenon. The difference of the rate of apoptosis and necrosis compared with the control was significant. The mitochondria membrane potential of cells decreased compared with the control. The expression of caspase-3 appeared,the Bax expression increaseed,the Bcl-2 expression decreased.NADH cured group compared with the hydrazine injured group: NADH could alleviate the Hydrazine's proliferation inhabition role of cardiac fibroblasts and cardiac fibroblasts apoptosis also decreased. The neucleus aggregation of cardiac fibroblasts decreased. The difference of the rate of apoptosis and necrosis decreased. The mitochondria membrane potential of cells increased. The expression of caspase-3 decreased.the Bax expression decreased.the Bcl-2 expression increased.Conclusion: Hydrazine might inhibit the cardiac fibroblast proliferation and induced CBF apoptosis, Hydrazine might injure the membrane of mitochondria anddecreased the mitochondria membrane potential.Which made the permeability Transition pore open ,and cytochrome C was released into cytosol where it interacted with apoptotic protease-activating factor- l(Apaf-l),ATP\dATP,and pro-caspase-9 formed the apoptosome.and then activated pro-caspase-3 .result in CBF apoptosis.NADH could ameliorate the function of mitochondria and increase Bcl-2 expression to decrease cytochrome C released into cytosol.inhibited activation of pro-caspase-3 .reduced cell apoptosis.