毛冬青皂苷E对H9C2心肌细胞缺氧/复氧损伤的影响

Effects of Ilexoside E on Hypoxia/Reoxygenation Injury of H9C2 Myocardial Cells and Its Mechanism

目的考察毛冬青皂苷E对H9C2心肌细胞缺氧/复氧损伤的保护作用,并探讨其作用机制。方法将H9C2心肌细胞随机分为正常对照组,模型组,毛冬青皂苷E低、中、高剂量对照组(10,50,250μg·m L-1)、毛冬青皂苷E低、中、高剂量治疗组(10,50,250μg·m L-1)。造模前24 h给予相应浓度的药物预处理,缺氧4 h,复氧4 h后,采用CCK-8法检测细胞存活率,DCFH-DA荧光探针检测细胞内活性氧(ROS)含量,并采用蛋白印迹法(Western Blot)检测Caspase-3和Cleaved caspase-3凋亡蛋白的表达。结果缺氧/复氧后,与正常对照组比较,模型组细胞存活率显著降低(P<0.05),细胞内ROS含量显著升高(P<0.01),毛冬青皂苷E各剂量治疗组预处理后,细胞存活率增加,ROS含量降低。缺氧/复氧损伤可使Caspase-3和Cleaved caspase-3蛋白表达增加。毛冬青皂苷E各剂量治疗组预处理后,Caspase-3和Cleaved caspase-3蛋白表达均下降。结论毛冬青皂苷E对H9C2心肌细胞缺氧/复氧损伤具有保护作用,其机制可能与减少ROS生成,抑制凋亡相关蛋白Caspase-3和Cleaved caspase-3的表达有关。

Objective To study the protective effects of ilexoside E on H9C2 myocardial cells after hypoxia/reoxygenation and to explore its mechanism. Methods H9C2 myocardial cells were randomly divided into normal control group, hypoxia/reoxygenation model group, and low-, middle- and high-dose ilexoside E （10, 50, 250 μg/mL） treatment groups. H9C2 myocardial cells were pretreated with ilexoside E for 24 h before hypoxia. After hypoxia for 4h and reoxygenation for 4h, cell viability was measured by CCK-8 method, intracellular reactive oxygen species（ROS） was detected by DCFH-DA fluorescent probe, and the expression levels of Caspase-3 and cleaved Caspase-3 were detected by Western blot. Results Compared with the normal control group, the viability of H9C2 myocardial cells decreased significantly （P 〈0.01）, and intracellular ROS level was elevated significantly （P 〈0.01） after hypoxia /reoxygeuation. After pretreatment with 10, 50, 250 μg/mL of ilexoside E, cell viability was significantly increased, the intracelluar ROS level was significantly decreased. Moreover, the expression levels of Caspase-3 and cleaved Caspase-3 were all increased in myocardial cells injured by hypoxia/reoxygenation. After pretreated with 10, 50, 250 μg·mL-1 of Ilexoside E, the expression levels of Caspase-3 and cleaved Caspase-3 were all reduced. Conclusion Ilexoside E has protective effects on H9C2 myocardial cells injured by H/R. And the mechanism may be related with the reduction of intracellular ROS level, with the inhibition of the expression of apoptosis-related protein Caspase-3 and cleaved Caspase-3.