化瘀祛痰方药对H2O2诱导人脐带静脉内皮细胞氧化应激保护作用的研究

目的探讨化瘀祛痰方药含药血清对H2O2诱导人脐带静脉内皮细胞（EA.hy926细胞）氧化应激的保护作用。方法体外培养EA.hy926细胞,随机分为5组。正常对照组：仅用DMEM完全培养基培养,不加任何药物;H2O2刺激组：培养液中加入终浓度为300μmol/L H2O2;含药血清低剂量组、含药血清中剂量组、含药血清高剂量组分别使用5%、10%、20%含药血清预处理24 h后加入终浓度为300μmol/L H2O2。各组细胞培养24 h后进行各项指标测定。采用MTT法检测细胞活性,黄嘌呤氧化酶法测定超氧化物歧化酶（SOD）活力,2,4-二硝基苯肼显色法检测乳酸脱氢酶（LDH）活性,硫代巴比妥酸法测定丙二醛（MDA）含量,硝酸还原酶法检测一氧化氮（NO）活性。采用细胞活性氧检测试剂盒,通过荧光显微镜观察细胞内活性氧水平的变化。结果与正常对照组相比,H2O2刺激后细胞活性明显下降,SOD、NO活性显著降低,而LDH活性、MDA含量则显著增加,活性氧水平明显升高。10%、20%化瘀祛痰方药含药血清可显著提高细胞存活率及SOD、NO活性,而显著降低LDH活性、MDA含量及活性氧水平。结论化瘀祛痰方药对H2O2诱导EA.hy926细胞氧化应激具有保护作用,这可能是其抗动脉粥样硬化的作用机制之一。

Effect of Huayuqutan Recipe on Oxidation Stress Induced by H2O2 in EA.hy926 Cells

Objective To observe the influence of Huayuqutan recipe on oxidation stress in cultured human umbilical veins endothelial cells EA.hy926 induced by H2O2. Methods EA.hy926 cells cultured in vitro were divided into five groups in random： the normal control group （cultured with DMEM complete medium without any drug）, H2O2 stimulating group （added with 300 μmol/L H2O2）, low, middle, high dose of Huayuqutan recipes drug-containing serum group （pretreated with 5%, 10%, 20% drug-containing serum for 24 h respectively and added with 300 μmol/L H2O2）. Then the levels of SOD, LDH, MDA, NO and ROS were detected. Results The concentration of LDH, MDA and ROS were increased, while those of SOD and NO decreased compared with normal group after stimulation of H2O2. Middle and high dose Huayuqutan recipes drug-containing serum could obviously inhibit the generation of LDH, MDA and ROS, while increase the levels of SOD and NO. Conclusion Huayuqutan recipe has protective effect on oxidation stress of EA.hy926 cells induced by H2O2, which may be one of the molecular mechanism of Huayqutan recipe against atherosclerosis.