参芎葡萄糖注射液对H2O2诱导的HUVEC细胞氧化损伤的保护作用

目的:探讨参芎葡萄糖注射液(Shenxiong glucose injection,SGI)对H_2O_2诱导的人脐静脉上皮细胞(HUVEC)细胞氧化模型损伤的保护作用及其机制。方法:体外培养HUVEC人脐静脉内皮细胞,用H_2O_2(130 mmol·L~(-1))处理0.5 h,建立HUVEC细胞H_2O_2氧化损伤模型。SGI组HUVEC细胞用(6%,8%,10%)SGI预处理6 h后,再用H_2O_2处理0.5 h。用MTS法检测细胞存活率;用ELISA法检测乳酸脱氢酶(LDH)漏出量、丙二醛(MDA)含量、超氧化物歧化酶(SOD)活性、过氧化氢酶(CAT)和谷胱甘肽过氧化物酶(GSH-Px)活力;用实时荧光定量聚合酶链式反应(Real-time PCR)和蛋白质免疫印迹(Western blot)技术检测凋亡相关基因及蛋白B细胞淋巴瘤/白血病-2(Bcl-2),Bcl-2相关X蛋白(Bax)和半胱氨酸天冬氨酸蛋白酶-3(Caspase-3)的mRNA和蛋白表达情况。结果:在130 mmol·L~(-1)H_2O_2作用细胞0.5 h的情况下,细胞存活率降低至50%左右,降低的程度合适,且实验结果重复性好,因此后续实验用该条件建立氧化损伤模型。与H_2O_2组比较,SGI预处理6 h能显著升高细胞存活率(P0.05,P0.01),减少LDH的外漏和MDA的生成(P0.05,P0.01),显著增加SOD,GSH-Px和CAT的活性(P0.05,P0.01)。RT-PCR和Western blot结果表明,SGI能显著上调Bcl-2的表达(P0.05,P0.01),下调Caspase-3,Bax的表达(P0.05,P0.01)。结论:参芎葡萄糖注射液能保护HUVEC细胞对抗H_2O_2诱导的氧化损伤,具有一定的剂量依赖关系,其作用机制可能与抑制细胞凋亡有关。

Protective Effect of Shenxiong Glucose Injection on H2O2-induced Oxidative Damage of HUVEC Cells

Objective: To investigate the protective effect and mechanism of Shenxiong Glucose injection (SGI) on H₂O₂-induced oxidative damage of HUVEC cells. Method: The in vitro models of oxidative stress damage based on HUVEC cells were established by treatment with H₂O₂(130 mmol·L^-1) for 0.5 h. SGI pretreated group:HUVEC cells were pretreated with SGI (6%, 8%, 10%) for 6 h, and then treated with H₂O₂ for 0.5 h. The cell viability was detected by MTS assay; ELISA method was used to determine lactate dehydrogenase(LDH) release, malondialdehyde(MDA) content as well as superoxide dismutase(SOD), catalase(CAT), glutathione peroxidase(GSH-Px) activities; gene and protein expression levels of Bcl-2, Bax and Caspase-3 in HUVEC cells were detected by Real-time PCR and Western blot. Result: After HUVEC cells were treated with H₂O₂(130 mmol·L^-1) for 0.5 h, the survival rate of the cells decreased to about 50%with a proper range and good repeatability, so that oxidative damage models were established under this condition. As compared with the H₂O₂ group, the cells survival was significantly increased (P < 0.05, P < 0.01) after treatment with SGI for 6 h. SGI not only decreased the release of LDH, MDA (P < 0.05, P < 0.01), but also significantly increased the activities of SOD, GSH-Px, and CAT (P < 0.05, P < 0.01). In addition, Real-time PCR and Western blot showed SGI can significantly up-regulate the expression level of Bcl-2 (P < 0.05, P < 0.01) and down-regulate the expression level of Caspase-3, Bax (P < 0.05, P < 0.01). Conclusions: SGI could protect the HUVEC cells against H₂O₂-induced oxidative damage in a certain range. The mechanism of action seemed to be relevant with inhibiting apoptosis.