芍药苷对MPP^+所致大鼠黑质脑片多巴胺能神经元损伤的保护作用

Protective effects of paeoniflorin on dopaminergic neurons in brain slice of substantia nigra treated with 1-methyl-4-phenylpytidinium

目的研究芍药苷(paeoniflorin,PF)对MPP+所致大鼠黑质脑片多巴胺能神经元损伤的保护作用,并观察PF对α-synuclein(α-syn)mRNA表达水平的影响。方法离体培养SD大鼠黑质器官型脑片,培养的d10给予不同浓度的MPP+(0.1、0.5、1.0mmol·L-1)处理24h,并在0.5mmol.L-1MPP+处理的基础上,给予1μmol·L-1或10μmol·L-1的PF进行干预。免疫组化法计数黑质致密部酪氨酸羟化酶(tyrosine hydroxylase,TH)阳性细胞数,定量RT-PCR法检测α-syn mRNA的表达丰度。结果MPP+(0.1、0.5、1.0mmol.L-1)处理导致黑质致密部TH+细胞数较正常对照组减少,差异均有统计学意义(P<0.05或P<0.01);MPP+(0.5mmol.L-1)使α-syn mRNA的表达水平明显升高(P<0.01)。PF(10μmol·L-1)有效增加了TH+细胞的数量(P<0.01),并明显下调了α-syn mRNA的表达(P<0.05)。结论PF能有效拮抗MPP+导致的多巴胺能神经元受损死亡,其机制可能与下调α-syn的表达有关。

Abstract：Aim To study the protective effects of Paeoniflorin （PF） on dopaminergic neurons in brain sliceof substantia nigra treated with MPP^＋ and to investigate the transcription of alpha - synuclei （ α - syn ） mRNA.Methods The organatypic brain slice culture of substantia nigra prepared from neonatal SD rats was placed on Millicell-CM porous membranes and cultured to day-10. Then the cuhrues of slice were treated with different concentrations （0. 1, 0.5,1.0 mmol · L^-1 ） of MPP^＋ for 24 h. Some of the cuhrues treated with 0. 5 mmol · L^-1 MPP^＋ also received PF （1 or 10 μmol · L^-1）. Slices cultured in normal medium were used as vehicle control. The tyrosine hydroxylase （TH） immunohistochemical staining with the cell counting was used to determine the dopaminergic neruons. The transcription of α-syn mRNA was examined by real-time quantitative RT-PCR. Results MPP ^＋ （ 0. 1,0. 5,1.0 mmol · L^-1） exposure markedly decreased the numberof TH^＋ cells in a dose-dependent manner （P 〈 0. 05 or P 〈0. 01 ） and sharply induced the transcription of α- syn mRNA （ P 〈 0. 01 ） in slices treated with 0. 5 mmol · L^-1 MPP^＋. The addition of PF （ 10 μmol · L^-1 ） to MPP ^＋ -treated slices significantly increased dopaminergic neurons survival （ P 〈 0. 01 ） and downregulated the transcription of α-syn mRNA significantly （P 〈 0. 05）. Conclusion PF can effectively inhibit the injury of dopaminergic neurons induced by MPP^＋ on brain slice of substantia nigra and downregulate the transcription of alpha-synuclein.