Tang Lili, Chen Chang, Xia Baomei, Wu Wei, Wei Ruide, Zhu Guoxue, Tang Juanjuan, Zhou Xin, Liang Yan, Zhang Zhen-Nian, Lu Yan, Yang Ye, Zhao Yang
Nanjing University of Chinese Medicine, Nanjing, Jiangsu, China.
Department of Neurology, Nanjing Hospital of Chinese Medicine Affiliated to Nanjing University of Chinese Medicine, Nanjing, Jiangsu, China.
Evid Based Complement Alternat Med. 2020 Jun 21;2020:9838295. doi: 10.1155/2020/9838295. eCollection 2020.
This study aimed to explore the protective effects of decoction on dopaminergic (DA) neuron injury in a rotenone-induced mouse model with chronic Parkinson's disease (PD) and explore its mechanism of action. Ultraperformance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) was used to measure the content of six main components in the decoction. The chronic PD mouse model was established by treating 10-month-old healthy wild C57BL/6 male mice with rotenone 30 mg/kg/day for 28 days in succession. The pole test and rotarod test were applied to detect the rescue effect of decoction in high, medium, and low dosages, respectively, on PD-like behaviors in mice with chronic PD. The protective effect of decoction on the mesencephalic nigrostriatal DA neuron injury was determined employing tyrosine hydroxylase (TH) immunofluorescence staining. Enzyme-linked immunosorbent assay (ELISA) was adopted to measure the inflammatory cytokines in serum, including TNF- (tumor necrosis factor-alpha), IFN- (interferon gamma), NF-B (nuclear factor kappa-B), and IL-1 (interleukin-1 beta). Western blotting was performed to quantify the expression of phosphorylated -Jun N-terminal kinase (-JNK), cleaved caspase-3, B-cell lymphoma-2 (Bcl-2), and NF-B in the brain. Our results showed that the decoction in high, medium, and low dosages reduced the turning time of mice ( < 0.01, < 0.01, and < 0.05). The high and medium dosages shortened the total climbing time of PD mice in the pole test ( < 0.01 and < 0.05). Meanwhile, the high, medium, and low dosages increased the rod-standing time of PD mice in the rotarod test ( < 0.01, < 0.05, and < 0.05). Besides, the decoction reversed the decrease in TH-positive neurons induced by rotenone, upregulated TH protein expression, and downregulated the -syn expression in the PD model. Moreover, the decoction in high dosage significantly inhibited the expression of -JNK, cleaved caspase-3, and NF-B in the midbrain of PD mice ( < 0.05, < 0.05, and < 0.01), upregulated the expression of Bcl-2 ( < 0.05), and decreased the content of TNF-, IFN-, NF-B, and IL-1 in the serum ( < 0.001, < 0.001, < 0.001, and < 0.001). Taken together, the decoction could protect mice from rotenone-induced chronic PD, which might be related to the reduction of the DA neuron apoptosis via suppressing the inflammatory reaction and the neuronal apoptosis pathway.
本研究旨在探讨[中药名称]水煎剂对鱼藤酮诱导的慢性帕金森病（PD）小鼠模型中多巴胺能（DA）神经元损伤的保护作用，并探讨其作用机制。采用超高效液相色谱-串联质谱法（UPLC-MS/MS）测定水煎剂中六种主要成分的含量。通过对10月龄健康野生型C57BL/6雄性小鼠连续28天给予30 mg/kg/天的鱼藤酮建立慢性PD小鼠模型。分别应用杆试验和转棒试验检测高、中、低剂量水煎剂对慢性PD小鼠帕金森样行为的挽救作用。采用酪氨酸羟化酶（TH）免疫荧光染色法测定水煎剂对中脑黑质纹状体DA神经元损伤的保护作用。采用酶联免疫吸附测定法（ELISA）检测血清中的炎性细胞因子，包括肿瘤坏死因子-α（TNF-α）、干扰素-γ（IFN-γ）、核因子κB（NF-κB）和白细胞介素-1β（IL-1β）。采用蛋白质免疫印迹法（Western blotting）定量检测大脑中磷酸化的应激活化蛋白激酶（p-JNK）、裂解的半胱天冬酶-3（cleaved caspase-3）、B细胞淋巴瘤-2（Bcl-2）和NF-κB的表达。结果显示，高、中、低剂量的水煎剂均能减少小鼠的旋转时间（P<0.01、P<0.01、P<0.05）。高、中剂量缩短了PD小鼠在杆试验中的总攀爬时间（P<0.01和P<0.05）。同时，高、中、低剂量均增加了PD小鼠在转棒试验中的站立时间（P<0.01、P<0.05、P<0.05）。此外，水煎剂可逆转鱼藤酮诱导的TH阳性神经元减少，上调TH蛋白表达，并下调PD模型中α-突触核蛋白（α-syn）的表达。而且，高剂量水煎剂显著抑制PD小鼠中脑p-JNK、裂解的caspase-3和NF-κB的表达（P<0.05、P<0.05、P<0.01），上调Bcl-2的表达（P<0.05），并降低血清中TNF-α、IFN-γ、NF-κB和IL-1的含量（P<0.001、P<0.001、P<0.001、P<0.001）。综上所述，水煎剂可保护小鼠免受鱼藤酮诱导的慢性PD，这可能与通过抑制炎症反应和神经元凋亡途径减少DA神经元凋亡有关。
