大蒜素对氧糖剥夺再灌注致PC12细胞损伤的保护作用及抗氧化应激机制

目的探讨大蒜素(AL)保护氧糖剥夺再灌注(OGD/RP)致PC12细胞损伤的抗氧化应激机制。方法采用无糖的Earle’s平衡盐溶液加缺氧法复制氧糖剥夺再灌注致PC12细胞损伤模型,同时给予不同浓度的大蒜素进行干预。采用MTT法检测细胞存活率;化学比色法测定细胞乳酸脱氢酶(LDH)漏出率;Hoechst 33342染色法检测大蒜素对细胞凋亡的影响,并计算凋亡率;生化法检测细胞超氧化物歧化酶(SOD)、丙二醛(MDA)、谷胱甘肽过氧化物酶(GSH-Px)的活性或含量;RT-PCR和Western blotting分子生物学方法检测氧化应激相关因子SOD、肿瘤坏死因子(TNF-α)、过氧化氢酶(CAT)mRNA和蛋白的表达。结果与正常对照组比较,OGD/RP模型组PC12细胞存活率下降,LDH漏出率、细胞凋亡率明显升高;细胞SOD、GSH-Px活性明显降低,MDA含量明显升高;细胞SOD、CAT mRNA及蛋白表达明显下降,TNF-αmRNA及蛋白表达明显增加。与模型组比较,大蒜素(60、30mg/L)可显著升高OGD/RP PC12细胞的存活率,降低凋亡率和LDH漏出率;大蒜素(60、30mg/L)可显著升高OGD/RP PC12细胞SOD、GSH-Px活性,减少MDA含量;大蒜素(60mg/L)可明显升高OGD/RP PC12细胞SOD、CAT mRNA及蛋白表达,降低TNF-αmRNA及蛋白表达。结论大蒜素对OGD/RP致PC12细胞损伤具有保护作用,其机制与抗氧化应激有关。

Effects of allicin on oxidative stress in PC12 cells following oxygen-glucose deprivation and reperfusion injury

Objective To determine the anti-oxidative stress effects of allicin in PC12 cell following oxygen-glucose deprivation and reperfusion (OGD/RP) injury and its related mechanisms. Methods The PC12 cells were pretreated with allicin 24 hours before exposed to sugar-free Earle’s solution and hypoxia then reperfusion injury was used as an in vitro model of OGD/RP. The OGD/RP PC12 cell survival rate was detected by MTT and apoptosis rate was determined by Hoechst 33342 staining. The PC12 cell lactate dehydrogenase (LDH) leakage rate was detected by chemical colorimetry. The superoxide dismutase (SOD), malonaldehyde(MDA) and glutathione pexoxidase(GSH-Px) content or activity in OGD/RP PC12 cell was measured by biochemical methods. The expression levels of SOD, TNF-α and catalase(CAT) protein and mRNA were detected by Western blotting and RT-PCR respectively. Results Compared with the control group, OGD/RP group could reduce the cell survival rate, SOD and GSH-Px activity, the LDH leakage rate increased, MDA content and apoptosis rate. Compared with OGD/RP model group, allicin (60, 30 mg/L) could increase OGD/RP PC12 cell survival rate, SOD and GSH-Px activity, decreased apoptosis rate, LDH leakage rate and MDA content; allicin (60 mg/L) could increase OGD/RP PC12 cell SOD, CAT mRNA and protein express levels, reduced TNF-α mRNA and protein express levels. Conclusion The Allicin have protective effects on OGD/RP injury in PC12 cell via anti-oxidative stress.