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目的研究6-羟基多巴胺(6-OHDA)所致帕金森病(PD)模型大鼠黑质膜铁转运蛋白1(FP1)基因表达及其启动子甲基化状态的变化。方法采用6-OHDA单侧损毁大鼠内侧前脑束(MFB),免疫组化方法观察脑黑质区酪氨酸羟化酶(TH)阳性神经元细胞存活情况,半定量逆转录聚合酶链反应(RT-PCR)技术检测黑质区FP1mRNA表达的变化,甲基化特异性PCR(MSP)检测FP1基因启动子区甲基化状态的变化。结果 6-OHDA单侧损毁大鼠MFB后,与未损毁侧及对照组相比,损毁侧中脑黑质区TH阳性细胞显著减少(t=20.619、18.404,P<0.01),FP1表达下调(t=8.123、7.609,P<0.01);与对照组相比,实验组注射侧FP1基因启动子区甲基化阳性率增高,差异有显著性(χ2=20.0,P<0.01)。结论 6-OHDA所致PD大鼠模型中黑质FP1基因表达降低,该基因启动子区甲基化可能参与FP1的表达下调。
Objective To study the changes of gene expression of iron transporter 1 (FP1) and its methylation status in Parkinson’s disease (PD) induced by 6-hydroxydopamine (6-OHDA). Methods The medial forebrain bundle (MFB) was unilaterally damaged by 6-OHDA. The survival of tyrosine hydroxylase (TH) positive neurons in the substantia nigra was observed by immunohistochemistry. The semi-quantitative reverse transcription polymerase chain (RT-PCR) technique was used to detect the expression of FP1mRNA in substantia nigra. Methylation-specific PCR (MSP) was used to detect the methylation status of FP1 promoter. Results Compared with the uninjured group and the control group, the number of TH-positive cells in the substantia nigra zone was significantly decreased (t = 20.619,18.404, P <0.01) and the expression of FP1 was down-regulated t = 8.123,7.609, P <0.01). Compared with the control group, the positive rate of FP1 gene promoter methylation in experimental group increased significantly (χ2 = 20.0, P <0.01). Conclusions The expression of FP1 in substantia nigra in PD rat model induced by 6-OHDA is reduced. The methylation of this gene may be involved in the down-regulation of FP1 expression.