Effect of Guasha on Regulation of TH Expression and Inflammatory Factors in Substantia Nigra and Nerve Function in Rats with Rotenone-Induced Parkinson's Disease
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摘要:
目的 观察刮痧对鱼藤酮诱导的帕金森病(PD)模型大鼠行为学及黑质酪氨酸羟化酶(TH)、核因子κB(NF-κB)、γ干扰素(IFN-γ)、白细胞介素-1β(IL-1β)的影响, 探究刮痧干预PD可能的作用机制。 方法 32只SD雄性大鼠随机分为假手术组、模型组、药物组和刮痧组, 每组8只。模型组、药物组和刮痧组大鼠采用颈背部皮下注射鱼藤酮(1 mg·kg-1)建立PD大鼠模型。药物组每天给予美多芭灌胃(50 mg·kg-1), 刮痧组隔日刮痧1次, 刮12次, 共计23 d。造模前、造模后、干预结束观察大鼠行为学改变; 干预结束, 采用Western blot和免疫荧光检测黑质区TH表达, 免疫组化法检测黑质TH、NF-κB表达, ELISA法检测中脑黑质NF-κB、IFN-γ、IL-1β含量, HE染色观察纹状体区细胞形态。 结果 与假手术组相比, 模型组大鼠行为学评分(3.42±1.248), 5 min内运动总距离缩短(P < 0.000 1), 起点停留时间延长(P < 0.05), 悬尾5 min内静止时间明显延长(P < 0.001), 中脑黑质TH表达减少(P < 0.001), 黑质NF-κB、IFN-γ、IL-1β含量均增加(P < 0.000 1), 纹状体区细胞肿胀明显(P < 0.05)。与模型组相比, 刮痧组大鼠5 min内运动总距离增加(P < 0.05)、起点停留时间缩短(P < 0.05)、悬尾5 min内静止时间缩短(P < 0.05), 黑质区TH表达增加(P < 0.05), 黑质NF-κB、IFN-γ、IL-1β含量均下降(P < 0.05), 纹状体区细胞肿胀情况改善(P < 0.000 1)。刮痧组与药物组相比, 各项指标差异均无统计学意义。 结论 刮痧可改善鱼藤酮诱导的PD大鼠的运动功能障碍, 其机制可能与下调炎性因子表达, 抑制神经炎性反应, 减少多巴胺能神经元丢失, 发挥神经保护作用有关。 Abstract:OBJECTIVE To observe the effect of Guasha on the behavior, the expression of tyrosine hydroxylase (TH), the level of nuclear factor κB (NF-κB), gamma interferon (IFN-γ) and interleukin-1β(IL-1β) in the substantia nigra(SN) of the midbrain of rotenone-induced rats with Parkinson's disease (PD), so as to explore the possible mechanism underlying improvement of PD. METHODS 32 SD male rats were randomly divided into the sham-operated group, model group, drug group and Guasha group, with 8 rats in each group. The rats in the model group, drug group and Guasha group were injected subcutaneously with rotenone (1 mg·kg-1) at the back of the neck to establish the PD rat model. The drug group was given metoba intragastric administration (50 mg·kg-1) every day, and the Guasha group was given 12 times of scraping every other day, a total of 23 d. The behavioral changes of rats were observed before modeling, after modeling and after intervention. After the intervention, the expression of TH in the SN was detected by Western blot and immunofluorescence. The expression of TH and NF-κB in the SN was detected by immunohistochemistry. The contents of NF-κB, IFN-γ and IL-1β in the SN were detected by enzyme-linked immunosorbent assay (ELISA). The cell morphology in the striatum was observed by HE staining. RESULTS In the comparison with the sham-operated group, the behavioral score of rats in the model group was (3.42±1.248), the total distance of movement within 5 minutes was shortened (P < 0.000 1), the residence time of the starting point was extended (P < 0.05), the rest time within 5 minutes of the tail suspension was significantly extended (P < 0.001), and the expression of TH in the SN was decreased (P < 0.001). The contents of NF-κB, IFN-γ and IL-1β in the SN were increased (P < 0.000 1), and the cell swelling in the striatum was significant (P < 0.05). In the comparison with the model group, the total distance of movement within 5 minutes in Guasha group was increased, the stay time at the starting point was shortened, the rest time within 5 minutes of suspension was shortened (P < 0.05), the expression of TH in SN was increased (P < 0.05), and the contents of NF-κB, IFN-γ and IL-1β in SN were all decreased (P < 0.05). Cell swelling in the striatum was improved (P < 0.000 1). There was no statistical significance in all indexes between the Guasha group and the drug group (P>0.05). CONCLUSIONS Guasha can improve the motor dysfunction of rotenone-induced PD rats, which may be associated with its role in down-regulating inflammatory factors, inhibiting neuroinflammatory response, reducing the loss of dopaminergic neurons, and protecting neuron. -
Key words:
- Guasha /
- Parkinson's disease /
- tyrosine hydroxylase /
- inflammatory factors /
- nuclear factor κB /
- rotenone
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图 1 鱼藤酮诱导的PD大鼠模型行为学评价
注: A.旷场实验5 min内运动总距离;B.旷场实验中大鼠在起始位置的停留时间;C. 大鼠悬尾5 min内静止时间;D.造模后, 旷场实验运动轨迹图, enter标注的是开始检测时大鼠脑袋的位置;E.造模后悬尾实验运动热图。与造模前相比, *P < 0.05, ***P < 0.001, ****P < 0.000 1;与假手术组相比, #P < 0.05, ###P < 0.001, ####P < 0.000 1。x±s, n=8。
Figure 1. Behavioral evaluation of rotenone induced PD model rats
图 3 刮痧减少PD大鼠黑质区TH阳性神经元丢失
注: A.黑质区TH阳性细胞的免疫组织化学图像, 比例尺为50 μm。B.免疫组化检测TH阳性细胞的定量统计; C.免疫荧光检测黑质区TH阳性神经元的形态,TH标记的黑质区多巴胺能神经元显示为绿色, 比例尺为200 μm, D.免疫荧光检测TH阳性细胞的定量统计;E.黑质区TH蛋白Western blot条带; F.黑质区TH蛋白水平定量。与假手术组相比, ***P < 0.001;与模型组相比, #P < 0.05, ##P < 0.01, ###P < 0.001。x±s, n=4。
Figure 3. Guasha alleviated the loss of TH neuron in the substantia nigra of PD model rats
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