CAJ | 학술논문

拟建立一种不仅操作简便而且能较好模拟帕金森病(Parkinson's disease,PD)临床症状和病理进程的造模方法.选取健康老龄雌性恒河猴12只,随机分为实验组9只(进而按临床症状及行为学评分又分为3个亚组)和对照组3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-methyl-4-phenyl-1,2,3,6-tetrahydropyridine,MPTP)来建立慢性 PD 恒河猴动物模型,对照组注射同等剂量的0.9%氯化钠溶液.每日于MPTP 给药前后观察记录各组动物行为学表现各30 min.实验结束后,全部处死动物,采用免疫组织化学法检测黑质酪氨酸羟化酶(tyrosine hydroxylase,TH)和α-突触核蛋白(α-synuclein,α-syn)的分布与表达变化对模型进行验证.行为学结果显示,实验组动物出现3种不同程度的临床表现,根据其轻重,将其分为3个亚组；TH 免疫组织化学结果显示,阳性物质主要分布于神经元胞质和突起内,与对照组相比,实验组中的第1、第2、第3亚组阳性总面积分别减少71.90%,61.90%,45.74%,差异有统计学意义(P <0.01)；α-syn 免疫组织化学结果显示,阳性物质主要分布于神经突起内,其次为细胞间质,并在濒临死亡动物的黑质致密部检测到路易小体,实验各组动物黑质阳性总面积较对照组分别增加170.29%,137.82%,47.88%,差异有统计学意义(P <0.01).上述结果表明,采用小剂量、多次重复肌内注射 MPTP 方式能够建立老龄恒河猴 PD 动物模型,该模型能够较好地模拟 PD 病人的临床症状和病理进程,是研究 PD 病因、发病机制、药物治疗及基因治疗较可靠的实验动物模型.
의건립일충불부조작간편이차능교호모의파금삼병(Parkinson's disease,PD)림상증상화병리진정적조모방법.선취건강노령자성항하후12지,수궤분위실험조9지(진이안림상증상급행위학평분우분위3개아조)화대조조3지,이0.2 mg/(kg?d)소제량、다차중복기내주사1-갑기-4-분기-1,2,3,6-사경필정(1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine,MPTP)래건립만성 PD 항하후동물모형,대조조주사동등제량적0.9%록화납용액.매일우MPTP 급약전후관찰기록각조동물행위학표현각30 min.실험결속후,전부처사동물,채용면역조직화학법검측흑질락안산간화매(tyrosine hydroxylase,TH)화α-돌촉핵단백(α-synuclein,α-syn)적분포여표체변화대모형진행험증.행위학결과현시,실험조동물출현3충불동정도적림상표현,근거기경중,장기분위3개아조；TH 면역조직화학결과현시,양성물질주요분포우신경원포질화돌기내,여대조조상비,실험조중적제1、제2、제3아조양성총면적분별감소71.90%,61.90%,45.74%,차이유통계학의의(P <0.01)；α-syn 면역조직화학결과현시,양성물질주요분포우신경돌기내,기차위세포간질,병재빈림사망동물적흑질치밀부검측도로역소체,실험각조동물흑질양성총면적교대조조분별증가170.29%,137.82%,47.88%,차이유통계학의의(P <0.01).상술결과표명,채용소제량、다차중복기내주사 MPTP 방식능구건립노령항하후 PD 동물모형,해모형능구교호지모의 PD 병인적림상증상화병리진정,시연구 PD 병인、발병궤제、약물치료급기인치료교가고적실험동물모형.
Summary Parkinson's disease(PD) is a common and age-related progressive neurodegenerative disorder,and its pathogenesis is not yet entirely clear.The present study is to establish a model easy to control and better simulate the clinical symptoms,processes and pathological changes of PD patients,expecting to provide a foundation and platform service for pathogenesis and treatment research of Parkinson's disease. <br> Twelve healthy aging female rhesus monkeys,divided into experimental group (nine rhesus monkeys) and control group (three rhesus monkeys) randomly,were respectively daily injected a small dose(0.2 mg/(kg?d))of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine(MPTP) and the same dose of saline by intramuscular injection for 45 days repeatedly.The behavioral manifestations of all monkeys were evaluated before and after the injection for 30 min,respectively.After sacrificed,the expression and distribution of tyroxine hydroxylase (TH) and α-synuclein (α-syn) in substantia nigra were studied by immunohistochemical method,and the immunopositive total area and average optical density were used to validate the model. <br> The results of behavioral manifestations indicated that,experimental animals showed three different degrees of clinical manifestations and could be divided into three subgroups (three animals each subgroup) according to the behavioral and clinical manifestations.The first subgroup showed typical behavioral manifestations of Parkinson's disease and one animal was in the moribund period.The activity,posture and bradypraxia of the third subgroup were slightly abnormal,and behavioral manifestations of the second subgroup were between the first and the third subgroup.However the control group did not show any abnormality.The immunopositive productions of TH were mainly distributed at the cytoplasm and neurites.Compared with the control group,the positive cytoplasm and neurites of the first subgroup were severely reduced,even no positive production was observed in partial substantia nigra pars compacta;the neuron structure of the third subgroup was mild blurred,and neurites were slightly shorter and fewer;the positive productions and structure of the second subgroup were between the first and the third subgroup.Also compared with the control group,the positive total area of each group decreased by 71.90%, 61.90% and 45.74%,showing statistical significance(P <0.01).The immunopositive productions ofα-syn mainly distributed at neuritis,and also distributed at the intercellular substance,and Lewy bodies were detected at the substantia nigra pars compacta of the moribund animal.Compared with the control group,positive total area of each group increased by 1 70.29%,137.82% and 47.88%,showing statistical significance(P <0.01). <br> The above results suggest that the aging rhesus PD animal model can be established via small doses and repeated intramuscular injection of MPTP, and the PD model can better simulate the clinical behavioral manifestations,disease processes and pathological changes of PD patients,so the model is more reliable for etiology,pathogenesis,drug therapy and gene therapy of Parkinson's disease.