Isor hamnetin Reduces H2 O2－Induced Intrinsic Pathway of Apoptosis in H9c2 Cardiacmuscle Cells：The Role of ROS Scavenging and ERK／p53 Pathway

Gui Bosun，Jing Xiao，Sun Bing，Rong Changchen，Xin Wang，Xiao Bosun

Institute of Medicinal Plant Development（IMPLAD），Chinese Academy of Medical Sciences ＆ Peking Union Medical College，No.151，Malianwa Nort h Road，Hai Dian District，Beijing 100193，PR China；＊Correspondence

Myocardial apoptosis during reperfusion after ischemia is believed to be caused by reactive oxygen species（ROS）.Isorhamnetin，a flavone and potent antioxidant，was investigated with respected to prevent against ROS－induced myocardial apoptosis.In the present study，an in vitro model of H2O2－induced apoptosis in H9c2 cardiomyocytes was used to mimicmyocardial ischemia／reperfusion injury.Isorhamnetin attenuated H2O2－induced decreased cell viability in a dose－dependenTManner.Qualitative immunofluorescent and quantitative laser scanning cytometry approaches demonstrated that H9c2 cardiomyocytes exposed to H2O2displayed increased DNA fragment（TUNEL staining），increased ROS generation（DHF staining）and lowered mitochondia membrane potential（JC－1 stainning），whereas isorhamnetin pretreaTMent（12.5μg／ml）attenuated all the action caused by H2O2.We also found that isorhamnetin reduced caspase 3 and 9 activity，whereas has no obvious effecton caspase 8.Qualitative real－ti me RT－PCR demonstrated that isor hamnetin suppressed H2O2－induced down－regulation in Bcl－2 antiapoptotic genes and up－regulation in Bax－like proapoptotic genes.Further more，H2O2exposure caused ERK1／2 MAPK phosphorylation which can be attenuated by is orhamnetin，but has no obvious effecton p38 MAPK.In conclusion，our present study indicates that isorhamnetin protected H9c2 cardiomyocyte against H2O2－induced apoptosis partially through scavenging ROS and ERK／p53－dependent intrinsic pathway of apoptosis.So isorhamnetin is a promising agent which may be used to treaTMyocardial ischemia／reperf usion injury.