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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。
AIM: To investigate the effects and possible mechanisms of nitric oxide synthase (NOS) inhibitor-asymmetric dimethylarginine (ADMA) on PC12 cells injured by glutamate (Glu) excitotoxicity. METHODS: PC12 cells were treated with different concentrations of glutamic acid to establish an experimental model of glutamate excitotoxicity injury cells. Cell viability was measured by MTT colorimetric assay. Lactate dehydrogenase (LDH) release assay was used to evaluate the degree of cell injury. Intracellular reactive oxygen species (ROS) levels were detected by flow cytometry (FCM) after DHR staining. The activity of NOS and NO were measured by kit and spectrophotometer Level. Results: Glutamic acid (1-6 mmol / L) treatment of PC12 cells for 24 h could reduce the survival rate of PC12 cells in a dose-dependent manner; ADMA 30 min before glutamate-induced PC12 cells inhibited glutamate-induced Decreased cell viability and increased release of LDH, reduced glutamate-induced intracellular ROS accumulation, inhibited glutamate over-activation of NOS and increased NO production. CONCLUSION: ADMA can significantly attenuate the excitotoxicity of glutamate on PC12 cells. Its mechanism may be related to the inhibition of NOS activity, the reduction of NO production, and the reduction of intracellular ROS accumulation.