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多西环素对脂多糖-帕金森病模型大鼠多巴胺能神经元的保护作用

Protective effects of doxycycline upon dopaminergic neuron in LPS-induced rat model of Parkinson′s disease

摘要:

目的 探讨多西环素对脂多糖(LPS)-帕金森病(PD)模型大鼠多巴胺能神经元的保护作用及其机制.方法 动物随机分为3组:正常对照组、LPS组和多西环素干预组;采用LPS黑质内立体定向注射建立PD模型;免疫组化染色法观察多西环素干预前后黑质多巴胺能神经元和MHCⅡ阳性小胶质细胞的变化;高效液相色谱-电化学检测仪检测纹状体多巴胺(DA)、DOPAC(二羟苯乙酸)含量的变化;Western 印迹检测黑质小胶质细胞MHCⅡ(主要组织相容性复合物Ⅱ)蛋白的表达.结果 多西环素干预后,黑质残存多巴胺神经元由LPS组的38%±5%上升到79%±4%(P<0.01);纹状体DA及DOPAC含量分别由LPS组的4.89±0.27和0.70±0.07上升到7.00±0.34和1.10±0.10(P<0.01);腹腔注射阿朴吗啡诱导动物旋转的平均圈数由LPS组的(208±14)次/30 min减少到(80±12)次/30 min(P<0.01);而黑质致密部MHCⅡ阳性细胞数量由LPS组的835±82减少到354±59(P<0.01);Western 印迹检测MHCⅡ蛋白的表达也明显减少.结论 多西环素能够抑制LPS诱导的黑质多巴胺能神经元变性,它可以通过下调小胶质细胞MHCⅡ的表达来实现其神经保护作用.

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Objective To explore the protective effect of doxycycline (DC) upon dopaminergic neuron in lipopolysaccharide (LPS)-induce rat model of Parkinson′s disease (PD).Methods Sixty SD rats were randomly divided into three groups: control, LPS and doxycycline treatment. LPS was stereostatically injected into unilateral substantia nigra (SNc) of rats to establish the PD models. The damage to the substantia nigra DA neurons was observed by using tyrosine-hydroxylase (TH) immunohistochemical staining. Specific antibody OX6 (MHCⅡ marker) was used to detect the changes in morphology and the numbers of microglia. The contents of dopamine and DOPAC in striatum were measured by high performance liquid chromatography (HPLC). Western blot were used to detect the expression of MHCⅡ (Major histocompatibility complex classⅡ) protein.Results After doxycycline treatment,the number of TH-positive cells remaining in the SNc increased from 38%±5% to 79%±4%(P<0.01). The contents of dopamine and DOPAC in striatum increased from 4.89±0.27 and 0.70±0.07 to 7.00±0.34 and 1.10±0.10 respectively(P<0.01);there was a significant decrease in rotational asymmetry in the doxycycline treatment group[ (80±12) turns/30 min] when compared to the LPS group [(208±14) turns/30 min] (P<0.01).However, the number of MHCⅡ-positive microglia decreased significantly (LPS group: 835±82 vs doxycycline treatment group: 354±59, P<0.01) after doxycycline treatment. Western blot were used to detect the expression of MHCⅡprotein. The results showed that the expression of MHCⅡ protein on microglia of LPS group increased significantly compared to the control group,but the expression of MHCⅡ protein were inhibited significantly after doxycycline treatment in the doxycycline treatment group, as compared to the LPS group.Conclusions Doxycycline might inhibit dopaminergic neuron degeneration by down-regulating the MHCⅡexpression on microglia.

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