丹参多酚酸盐对过氧化氢所致乳鼠心肌细胞凋亡的影响

Effects of Salvianolate on neonatal rat cadiocytes apoptosis induced by H_2O_2

[目的]研究丹参多酚酸盐(Salvianolate)对过氧化氢(H_2O_2)所致乳鼠心肌细胞凋亡的影响,并探讨其可能的作用机制。[方法]于无菌环境下分离并培养乳鼠心肌细胞72 h后随机分为空白对照组、H_2O_2组、丹参多酚酸盐(50、100和200 mg/L)+H_2O_2组,每组设8个复孔;给药干预24 h后,检测培养液中谷草转氨酶(AST)、磷酸肌酸激酶(CPK)、乳酸脱氢酶(LDH)含量,四甲基噻唑蓝(MTT)法检测细胞存活率,通过流氏细胞术(Flow Cytometry)检测细胞凋亡状况并计算凋亡率;逆转录-聚合酶链反应(RT-PCR)法检测AKT m RNA和bcl-2 m RNA、Bax m RNA表达并计算bcl-2/Bax表达比值,Western blotting法检测细胞中半胱氨酸天冬氨酸蛋白酶-3(caspase-3)、核转录因子-κB(NF-κB)表达并进行半定量分析,测定细胞中超氧化物歧化酶(SOD)、过氧化氢酶(CAT)活性和丙二醛(MDA)含量。[结果]与H_2O_2组比较,丹参多酚酸盐(100、200 mg/L)+H_2O_2组培养液中AST、CPK、LDH含量显著降低,细胞存活率显著升高,细胞凋亡率显著降低。细胞中细胞中AKT m RNA、bcl-2 m RNA表达显著上调,Bax m RNA表达显著下调,bcl-2/Bax表达比值显著升高,caspase-3、NF-κB表达显著下调,SOD、CAT活性显著升高且MDA含量显著降低,上述差异均具有统计学意义(P<0.05或P<0.01)。[结论]丹参多酚酸盐对H_2O_2所致乳鼠心肌细胞凋亡具有抑制作用。

[Objective] To investigate the effects of Salvianolate on neonatal rat cadiocytes apoptosis induced by H_2O_2. [Methods]Cadiocytes of neonate rat was cultivated for 72 hours and divided into six groups: normal control group, H_2O_2 group, Salvianolate(50, 100,200 mg/L)+ H_2O_2groups(n=8). The 12 hours after the drugs were given. The content of AST, CPK, LDH in culture medium were detected;the survival rate was detected by MTT; the cardiomyocytes apoptosis was observed by flow cytometry and the apoptosis rate was calculated; the expression of AKT m RNA, bcl-2 m RNA, Bax m RNA were detected by RT-PCR and the ratio of bcl-2/Bax was calculated;the expression of caspase-3, NF-κB protein was detected by Western blot and Semi-quantitative analysized; the activity of SOD, CAT and the content of MDA in cardiomyocytes were determinted. [Results] Compared with the H_2O_2 group, the content of AST, CPK, LDH in culture medium of Salvianolate(100, 200 mg/L)+ H_2O_2 groups were significantly decreased; the survival rate were significantly increased;the cardiomyocytes apoptosis were improved and the apoptosis rate were significantly decreased; the expression of AKT, bcl-2 m RNA were significantly up-regulated, while the expression of Bax m RNA was significantly down-regulated, the ratio of bcl-2/Bax were significantly increased; the expression of caspase-3, NF-κB protein were significantly down-regulated; the activity of SOD, CAT in cardiomyocytes were significantly increased and the content of MDA was significantly decreased, all of the difference above were significant(P<0.05 or P<0.01). [Conclusion] Salvianolate had inhibitive effects on neonatal rat cadiocytes apoptosis induced by H_2O_2.