Effect and mechanism of salidroside for 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine induced Parkinson's disease in mice
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摘要:目的
探讨红景天苷对1-甲基-4-苯基-1-2-3-6-四氢吡啶（MPTP）诱导帕金森病（PD）小鼠的作用及机制。
方法通过MPTP诱导C57BL/6J小鼠建立PD小鼠模型，通过旷场实验记录小鼠行动轨迹；采用免疫组织化学法检测小鼠脑组织酪氨酸羟化酶（TH）、α-突触核蛋白（α-syn）、核因子E2相关因子2（Nrf2）和醌氧化还原酶1（NQO1）表达变化。通过MPTP刺激SN4741细胞建立PD体外细胞模型，红景天苷预处理后，采用TUNEL实验检测各组细胞凋亡情况；采用免疫荧光法检测各组细胞TH、α-syn、Nrf2和NQO1表达变化情况；转染si-RNA敲减Nrf2表达，采用TUNEL实验检测各组细胞凋亡情况。通过酶联免疫吸附试验（ELISA）检测脑组织提取液和细胞培养液中白细胞介素-1β（IL-1β）、白细胞介素-18（IL-18）蛋白表达水平。
结果旷场实验结果显示，红景天苷能够改善PD小鼠3 min活动总距离、平均速度和活动轨迹，差异有统计学意义（P < 0.05）；与PD模型小鼠相比，红景天苷处理的PD小鼠大脑TH、Nrf2、NQO1表达增加，α-syn、IL-1β、IL-18表达减少，差异有统计学意义（P < 0.05）；与MPTP刺激细胞相比，红景天苷预处理组细胞凋亡减少，TH、Nrf2和NQO1表达增高，α-syn、IL-1β和IL-18表达降低，差异有统计学意义（P < 0.05）；敲减Nrf2表达后，红景天苷对MPTP刺激细胞的保护作用减弱甚至消失。
结论红景天苷可能通过激活Nrf2-ARE信号通路而减轻神经元细胞凋亡情况，有望成为一种新的PD治疗药物。
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关键词:
- 红景天苷 /
- 核因子E2相关因子2 /
- 抗氧化反应元件 /
- 帕金森病 /
- 细胞凋亡
Abstract:ObjectiveTo investigate the effect and mechanism of salidroside on1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP)-induced Parkinson's disease (PD) in mice.
MethodsC57BL/6J mice were induced by MPTP to establish a PD mouse model and the movement track of mice was recorded by open field experiment. The expression of tyrosine hydroxylase (TH), α-synuclein (α-syn), nuclear factor E2 associated factor 2 (Nrf2) and quinone oxidoreductase 1 (NQO1) were detected by immunohistochemistry in brain tissue of mice. SN4741 cells were stimulated by MPTP to establish a PD cell model in vitro. MPTP stimulation of SN4741 cells to establish an in vitro cell model of PD. After pretreatment with salidroside, TUNEL was used to detect cell apoptosis in each group. The expressions of TH, α-syn, Nrf2 and NQO1 were detected by immunofluorescence. Nrf2 expression was knocked down by being transfected with si-RNA, and apoptosis condition was detected by TUNEL assay. The expression levels of interleukin-1β (IL-1β) and interleukin-18 (IL-18) protein in brain tissue extract and cell culture were detected by enzyme-linked immunosorbent assay (ELISA).
ResultsThe open field experiment results showed that salidroside could improve 3 min total distance of activity, average speed and activity track of PD mice (P < 0.05). Compared with the PD model mice, the salidroside treated mice showed significantly increased expression of TH, Nrf2 and NQO1 in the brain, and reduced expression of IL-18, α-Syn, IL-1β. Compared with the MPTP stimulated cells, the salidroside pretreatment group showed a decrease in cell apoptosis and an increase in the expressions of TH, Nrf2 and NQO1, but reduced expressions in α-Syn, IL-1β and IL-18. After si-RNA knockdown of Nrf2 expression, the protective effect of salidroside on MPTP stimulated cells weakened or even disappeared.
ConclusionSalidroside may alleviate neuronal apoptosis by activating Nrf2-ARE signaling pathway, which is expected to be a new drug for PD treatment.
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图 1 各组小鼠的旷场实验结果
A: 旷场实验活动轨迹图; B、C: 旷场内3 min总距离、平均速度(与对照组比较, *P < 0.05; 与MPTP组比较, #P < 0.05)。
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图 2 各组小鼠脑组织TH、α-syn、Nrf2、NQO1、IL-1β和IL-18表达情况
A: TH、α-syn、Nrf2、NQO1表达情况(免疫组化染色法，标尺20 μm); B、C: IL-1β、IL-18表达情况
(与对照组比较, **P < 0.01; 与MPTP组比较, #P < 0.05, ##P < 0.01)。![]()
图 3 各组SN4741细胞TUNEL实验结果和ELISA实验结果
A: 细胞凋亡情况(TUNEL染色法，标尺20 μm); B: IL-1β、IL-18表达(与对照组比较, **P < 0.01; 与MPTP组比较, #P < 0.05, ##P < 0.01)。
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