ABSTRACT
Aims To study the effects of clenbuterol on anoxia/reoxygenation( A/R) injury in neonatal Wistar rat cardiomyocytes and to explore whether its mecha-nism is related to reperfusion injury salvage kinase ( RISK) or not. Methods The cultured primary neo-natal cardiomyocytes were randomly divided into eight groups: ①normal culture group; ②anoxia/reoxygen-ation( A/R) group;③ clenbuterol ( 1 μmol · L-1 ) +A/R;④ICI118,551(10 μmol·L-1) + clenbuterol ( 1 μmol · L-1 ) + A/R; ⑤Metoprolol ( 10μmol · L-1 ) + clenbuterol(1μmol·L-1 ) + A/R group;⑥Metoprolol ( 10 μmol · L-1 ) + A/R group; ⑦PD98059 ( 20 μmol · L-1 ) + clenbuterol ( 1 μmol · L-1 ) + A/R group;⑧ LY294002(10 μmol·L-1 ) +clenbuterol(1 μmol · L-1 ) + A/R group. Cell via-bility was determined by the conventional MTT reduc-tion assay. The content of LDH in cultured medium was measured with colorimetry. Cardiomyocyte apopto-sis was determined by Hoechst33342 . Intracellular re-active species( ROS) were monitored by the fluorescent DCFH-DA. Total ERK2 and phosphorylated ERK were detected by western blot. Results Compared with A/R group, clenbuterol significantly increased vaibility of cells, reduced LDH release, lowered the rate of apop-tosis and ROS production. When addedβ2 receptor an-tagonist ICI118 , 551 , PI3 K inhibitor LY294002 and ERK inhibitor PD98059 , the effects of clenbuterol a-bove were inhibited; but β1 receptor antagonist Meto-prolol protected the cardiomyocytes from A/R injury, as evidenced by decreased LDH release and increased cell viability. There were no synergistic effects in the combined use of clenbuterol and Metoprolol. Conclu-sion clenbuterol exerts cardioprotective effects against A/R injury by inhibiting oxidative stress and apopto-sis. The protection of clenbuterol is inhibited by ICI118 , 551 , LY294002 and PD98059 . clenbuterol protects cardiomyocytes against A/R injury via RISK pathway by activation of β2 receptor.
ABSTRACT
Aim To study the mechanisms of the pro-tective effect of procyanidin B2 ( PCB2 ) on the myocar-dial cell apoptosis induced by lipopolysaccharide ( LPS) . Methods Using the primary culture rat myo-cardial cells, myocardial cell injury model was induced by LPS. PCB2 low, medium and high dose groups, were cultured with 6. 25 , 12. 5 , 25. 0 μmol · L-1 PCB2 respectively in DMEM medium for 24 h continu-ously. Myocardial cell survival rate was determined by MTT colorimetric method. Cardiacmyocyte NOX activi-ty was determined by lucigen chemiluminescence meth-od . Western blot analysis was used to detect myocardi-al NADPH oxidase p47phox expression. TUNEL method was used to detect apoptosis and flow cytometry was used to determine the content of myocardial cells ROS. Results Compared with control group, the cell dam-age induced by LPS group myocardial cell survival rate significantly decreased ( P <0. 01 ) , and myocardial cell NOX activity, p47phox expression, apoptotic cell number and ROS content were significantly increased (P<0. 01). PCB2 low, medium and high dose groups cell survival rates were significantly elevated, myocar-dial cell NOX activity and p47phox expression, apoptotic cell number and the ROS content decreased significant-ly in a dose-dependent manner ( P <0. 01 ) . Conclu-sion PCB2 protects myocardial cell apoptosis induced by LPS via inhibiting the expression of NADPH oxidase activation, p47phox expression and reactive oxygen spe-cies generation.