【摘要】 目的:探讨一氧化氮合酶(NOS)抑制剂-非对称性二甲基精氨酸(ADMA)对谷氨酸(Glu)兴奋性毒性损伤PC12细胞的影响及其机制。方法:用不同浓度的谷氨酸处理PC12细胞,建立谷氨酸兴奋性神经毒性损伤细胞的实验模型;应用四甲基偶氮唑蓝(MTT)比色法检测细胞存活率;乳酸脱氢酶(LDH)释放试验评价细胞的损伤程度;双氢罗丹明123(DHR)染色后流式细胞仪(FCM)检测细胞内活性氧(ROS)水平;应用试剂盒及分光光度计测定NOS活性和NO水平。结果:谷氨酸(1-6mmol/L)处理PC12细胞24h,可呈剂量依赖性地降低PC12细胞的存活率;在谷氨酸作用PC12细胞前30min给予ADMA,可明显地抑制谷氨酸引起的细胞存活率降低及LDH释放增加,减少谷氨酸引起的细胞内ROS堆积,抑制谷氨酸过度激活NOS和增加NO的生成。结论:ADMA能显著地减弱谷氨酸对PC12细胞的兴奋性毒性损伤作用;其作用机制可能与抑制NOS活性,减少NO生成,进而减轻细胞内ROS的堆积有关。更多还原

【Abstract】 AIM:To investigate the effects of asymmetric dimethylaoyoinine (ADMA) on glutamate-induced PC12 cell damage and its mechanisms. METHODS: PC12 cells were treated with different concentrations of glutamate as an in vitro excitotoxic trauma model. The cell viability was measured by MTT assay. Glutamate cytotoxicity was evaluated by lactate dehydrogenase (LDH) release assay. Intercellular reactive oxygen species (ROS) was detected by dihydrorhodamine123 (DHR) staining and flow cytometric (FCM) analysis. Nitric oxide synthase (NOS) activity and nitric oxide (NO) production were detected by using commercial kits with a spectrophotometer. RESULTS: Glutamate at concentrations of 1 mmol/L to 6 mmol/L dose-dependently decreased PC12 cell viability. Pretreatment 30 min with ADMA prior to administration of glutamate significantly attenuated the inhibition of cell viability, LDH release and ROS accumulation induced by glutamate. Pretreatment with ADMA significantly inhibited the increases in NOS activity and NO production caused by glutamate. CONCLUSION: ADMA obviously protects PC12 cells against glutamate-induced excitotoxicity by inhibiting NOS activity, overproduction of NO and accumulation of intracellular ROS.更多还原