帕金森恒河猴血液P53、Bax、Bcl-2、Caspase-3 mRNA的进程性变化

发布时间：2019-03-30 08:01

【摘要】：帕金森(Parkinson's disease,PD)是一种以运动功能障碍为主要特征的神经退行性疾病,大量研究表明细胞凋亡参与了PD的发病机制。PD动物模型对于深入研究PD病因及发病机理、进行有效预防和提高治疗效果将起到重要作用。为探讨PD恒河猴(Macaca mulatta)外周血中肿瘤抑制基因P53、B细胞淋巴瘤-2(B cell lymphoma-2,Bcl-2)、B细胞淋巴瘤-2相关X蛋白(BCL-2-associated X protein,Bax)和半胱天冬氨酸蛋白酶-3(cysteinyl aspartate-specific proteinase-3,Caspase-3)mRNA表达水平与行为学变化之间的关系,本研究选取健康青年恒河猴6只,以0.2 mg/(kg·d)小剂量、多次重复肌内注射1-甲基-4-苯基-1,2,3,6-四氢吡啶(1-methy-4-pheny-l,2,3,6-tetrahydropyridine,MPTP)建立了PD恒河猴模型。于每次MPTP给药前后观察记录动物行为学表现30 min。分别于未注射MPTP(第0周),注射MPTP后第4、8、12、16、20和24周采集恒河猴前臂静脉血,采用qRT-PCR检测外周血P53、Bax、Bcl-2和Caspase-3 mRNA表达水平。行为学观察结果显示,第4周时,实验动物表现出温和的运动迟缓,随后症状逐渐加重,运动能力持续下降,到12周时实验动物均表现出活动量显著减少,姿势僵硬和严重的运动障碍。13周后临床症状基本稳定,实验动物表现出运动迟缓,姿势异常,肌强直和静止性震颤等。qRT-PCR结果显示,与0周相比,注射MPTP后第4、8、12周P53和Caspase-3 mRNA表达显著升高(P0.05),而第16、20、24周差异不显著;与0周相比,Bcl-2/Bax比值在第4、8、12周显著降低(P0.05),而第16、20、24周差异不显著。0~12周行为学评分与外周血P53和Caspase-3 mRNA表达水平呈正相关(r=0.975,r=0.956;P0.05)。结果表明,当外周血P53和Caspase-3 mRNA表达水平显著升高时临床症状也处于逐渐加重的阶段,因此通过检测血液中凋亡调控基因的表达,能反应出PD恒河猴的疾病进程。研究结果可为PD诊断和疾病进程监测提供辅助参考。
[Abstract]:Parkinson's disease (Parkinson's disease,PD) is a neurodegenerative disease characterized by motor dysfunction. A large number of studies have shown that apoptosis is involved in the pathogenesis of PD. PD animal model for in-depth study of the etiology and pathogenesis of PD, Effective prevention and improvement of therapeutic effects will play an important role. To investigate the tumor suppressor gene P53, B cell lymphoma-2 (B cell lymphoma-2,Bcl-2), B cell lymphoma-2-associated X protein (BCL-2-associated X protein,) in peripheral blood of PD rhesus monkey (Macaca mulatta) The relationship between the expression level of Bax and caspase-3 (cysteinyl aspartate-specific proteinase-3,Caspase-3) mRNA and behavioral changes was studied in this study. Six healthy young rhesus monkeys were selected and given a low dose of 0.2 mg/ (kg 路d). PD rhesus monkey model was established by repeated intramuscular injection of 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP). The behavior of animals was observed and recorded for 30 min. before and after each MPTP administration. The blood samples of forearm vein of rhesus monkeys were collected at 4,8,12,16,20 and 24 weeks after injection of MPTP (0 weeks), respectively. The expression levels of P53, Bax, Bcl-2 and Caspase-3 mRNA in peripheral blood were detected by qRT-PCR. Behavioral observation showed that at the 4th week, the animals showed mild slow movement, then the symptoms became more severe and the motor ability continued to decrease, and at the 12th week, all the animals showed a significant decrease in the amount of activity, and the experimental animals showed a significant decrease in activity at the end of the 12th week. The clinical symptoms were basically stable after 13 weeks, the experimental animals showed slow movement, abnormal posture, muscular rigidity and quivering tremor, etc. The results of qRT-PCR showed that compared with 0 weeks, 4,8,8 after MPTP injection, the clinical symptoms were basically stable, and the experimental animals showed slow movement, abnormal posture, muscular stiffness and quivering tremor. The expression of p53 and Caspase-3 mRNA increased significantly at the 12th week (P0.05), but there was no significant difference at the 16th, 20th, 24th week. Compared with 0 weeks, the ratio of Bcl-2/Bax decreased significantly at 4,8,12 weeks (P0.05), but there was no significant difference at 16,20,24 weeks (P 0.05). There was a positive correlation between the behavioral score and the expression of p53 and Caspase-3 mRNA in peripheral blood (r = 0.975, r = 0.956) at week 16, week 20 and week 24 (r = 0.975, r = 0.956). (P0.05). The results showed that when the expression level of p53 and Caspase-3 mRNA in peripheral blood was significantly increased, the clinical symptoms were in the stage of gradual aggravation. Therefore, the expression of apoptosis-regulated genes could reflect the disease progression of PD rhesus monkeys by detecting the expression of apoptosis-regulated genes in blood. The results can be used as a reference for the diagnosis of PD and the monitoring of disease progression.
【作者单位】：四川农业大学动物医学院/动物疾病模型实验室;四川农业大学预防兽医研究所;四川普莱美生物科技有限公司/国家实验猕猴种源基地;
【基金】：国家科技支撑计划课题(No.2014BAI03B01) 国家重大科学仪器设备开发专项(No.2013YQ49085906)
【分类号】：R742.5

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