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目的:研究Nrf2/ARE通路在1-甲基-4-苯基-1,2,3,6-四氢吡啶(1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine,MPTP)所致帕金森病(PD)动物模型发病机制中可能的作用,探讨导致多巴胺(DA)能神经元变性坏死的可能机制,以及丹参酮ⅡA对Nrf2/ARE通路的影响和对多巴胺能神经元的保护作用。方法:将60只C57BL/6N小鼠随机分为对照组、PD模型组和丹参酮ⅡA组,每组20只。PD模型组和丹参酮ⅡA组小鼠采用MPTP制备PD小鼠模型,丹参酮ⅡA组制备PD模型后腹腔注射丹参酮ⅡA。观察各组小鼠行为学表现,采用免疫组织化学、免疫蛋白印迹和免疫荧光双标法检测各组小鼠中脑黑质区酪氨酸羟化酶(TH)、核转录因子(Nrf2)、醌氧化还原酶(NQO1)和胶质纤维酸性蛋白(GFAP)阳性细胞数和蛋白表达水平。结果:与对照组比较,PD模型组小鼠出现典型的PD样症状。在MPTP最后一次注射后48小时,黑质区TH阳性神经元数量和蛋白表达水平明显下降,Nrf2、NQO1和GFAP阳性细胞数量和蛋白表达水平明显增加(P<0.01),在MPTP最后一次注射后7天,黑质区TH阳性神经元数量和蛋白表达水平较对照组减少约45%和50%(P<0.01)。与PD模型组比较,丹参酮ⅡA组小鼠PD样症状减轻,在MPTP最后一次注射后48小时,中脑黑质区Nrf2、NQO1和GFAP阳性细胞数量和蛋白表达水平进一步升高(P<0.01),在MPTP最后一次注射后7天,黑质区TH阳性神经元数量和蛋白表达水平明显升高(P<0.01)。结论:Nrf2/ARE通路在急性PD模型DA能神经元内可能被激活,其可能通过上调NQO1的表达对黑质DA能神经元起到保护作用,丹参酮ⅡA可能通过激活Nrf2/ARE通路对小鼠多巴胺能神经元起到一定保护作用。
Objective: To investigate the role of Nrf2 / ARE pathway in MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) Induced Parkinson’s disease (PD) in animal models and to explore the possible mechanism of degeneration and necrosis of dopaminergic neurons and the effect of tanshinone IIA on Nrf2 / ARE pathway and the protection of dopaminergic neurons effect. Methods: Sixty C57BL / 6N mice were randomly divided into control group, PD model group and tanshinone Ⅱ A group, with 20 rats in each group. PD model group and tanshinone Ⅱ A group mice using MPTP preparation of PD mouse model, tanshinone Ⅱ A group prepared PD model intraperitoneal injection of tanshinone Ⅱ A. The behavior of mice in each group was observed. Immunohistochemistry, Western blotting and immunofluorescence double-labeled method were used to detect the tyrosine hydroxylase (TH), nuclear transcription factor (Nrf2) Quinone oxidoreductase (NQO1) and glial fibrillary acidic protein (GFAP) positive cells and protein expression levels. Results: Compared with the control group, PD model mice showed typical symptoms of PD. At the 48th hour after the last injection of MPTP, the number of TH positive neurons and protein expression in the substantia nigra zone decreased significantly, while the number of Nrf2, NQO1 and GFAP positive cells and protein expression increased significantly (P <0.01). After the last injection of MPTP At 7 days, the number and protein expression of TH positive neurons in the substantia nigra zone decreased by about 45% and 50% (P <0.01) compared with the control group. Compared with PD model group, the symptoms of PD in mice of Tanshinone ⅡA group were alleviated. The number of Nrf2, NQO1 and GFAP positive cells and protein expression in substantia nigra were increased 48 h after the last MPTP injection (P <0.01) , The number of TH positive neurons and protein expression in substantia nigra zone increased significantly (P <0.01) 7 days after the last injection of MPTP. CONCLUSION: Nrf2 / ARE pathway may be activated in DA neurons of acute PD model, which may play a protective role on DA neurons in substantia nigra by up-regulating the expression of NQO1. Tanshinone IIA may activate the Nrf2 / ARE pathway in mice Dopaminergic neurons play a protective role.