CT引导脑内植入携带人类酪氨酸羟化酶基因腺相关病毒载体治疗实验性帕金森病猴  

A gene therapy of experimental Parkinsonism monkeys with intra-stristums implantation of AAV-TH with CT guiding

作　　者：王维[1] 毛俊[1] 于小平[1] 刘晟[1] 胡维新[2] 曾赵军[2] 蔡维军[2] 吴小兵[3] 

机构地区：[1]中南大学湘雅第三医院放射科,长沙410013 [2]中南大学湘雅医学院 [3]中国预防医学科学院病毒学研究所病毒基因工程国家重点实验室

出　　处：《中华放射学杂志》2003年第11期1029-1035,共7页Chinese Journal of Radiology

基　　金：国家自然科学基金资助项目 (3 9870 198)

摘　　要：目的　观察CT引导脑内注射人类酪氨酸羟化酶基因腺相关病毒载体 (AAV hTH)对神经细胞的转染能力 ,以及对帕金森病 (PD)猴的治疗效应。方法　经右侧颈内动脉注射 1 甲基 4 苯基 1,2 ,3,6 四氢吡啶 (MPTP)制作 6只偏侧帕金森病恒河猴模型 ,经CT引导将AAV hTH注入 5只帕金森病猴右侧尾状核内 ,观测猴模型的行为学改变 6个月以上 ,以高效液相色谱法 (HLPC)测定转染后尾状核内多巴胺 (DA)及其代谢产物含量 ,并以免疫组化检测法及逆转录 聚合酶链式反应 (RT PCR)方法检测酪氨酸羟化酶 (TH)基因的表达情况。结果　CT引导定位穿刺有实时操作、定位精确、微创的优点。所有注射AAV hTH的偏侧PD猴模型术后 2周即有帕金森病症状改善 ,持续 6个月以上 ,其中 1只动物阿朴吗啡旋转实验示完全停止了帕金森病旋转现象 ,其余动物旋转次数较术前减少约4 2 %～ 70 %。实验侧与健侧尾状核区多巴胺及其代谢产物含量比值较未治疗模型增高。免疫组化染色见穿刺针道部位有大量TH阳性细胞 ;RT PCR检测到实验侧尾状核区有TH mRNA存在 ;而对照侧尾状核区、其他部位脑组织与心、肝、肾组织及未注射模型实验侧尾状核区均未见阳性发现。结论　CT引导立体定向穿刺PD猴尾状核注入AAV hTH能使TH基因精确地在特定部位有效表达 。Objective To observe the transfection ability to neuron and the therapy effect of adeno-associated virus vector (AAV) mediated intracerebal expressing of human tyrosine hydroxylase (TH) in the Rhesus monkey of Parkinson disease with CT guiding. Methods Six Rhesus monkeys were induced into hemi-parkinsonism models by infusion of MPTP into right internal carotid artery. Under the guidance of CT and stereotactic apparatus, the recombinant adeno-associated virus vectors-human tyrosine hydroxylase were injected into the right caudate nucleus of 5 PD monkeys. The motor ability of the PD model was evaluated for 6 months after implantation. The content of DA, DOPAC, and HVA in the caudate nucleus was assayed with HPLC, and the expression of the tyrosine hydroxylase gene was detected with immunohistochemistry and RT-PCR. Results The method of stereotactic puncture guided with CT had the features of real-time operating, fine location, and mini-trauma. The motor ability of all five models was all ameliorated after stereotactical injection of AAV-hTH vectors from two weeks up to six months. One PD monkey did not show any twist in response to apomorphine test. The frequency of twirl in the other 4 PD monkeys in response to apomorphine test was decreased by 42%-70% compared with that preoperatively. The contents of DA and DA metabolites in the lesion caudate nucleus were increased. Many TH positive cells in the implantation sites of caudate nucleus were shown by immunohistochemistry, which was different from the contralateral caudate nucleus. TH-mRNA was found in the treated side of caudate nucleus with RT-PCR, but no positive detection in the untreated side of caudate nucleus or in other sites in the brain. The negative results of RT-PCR were also shown in samples of the heart, liver, kidney tissue, or right side caudate nucleus of the uninjected monkey. Conclusion Under the guidance of CT and stereotactic apparatus, the AAV-hTH vector can be injected accurately into the lesioned caudate nucleus of PD Rhesus monkey model.

关 键 词：尾状核 CT引导 帕金森病 PD 酪氨酸羟化酶 脑内 治疗 模型 实时 旋转 

分 类 号：R742.5[医药卫生—神经病学与精神病学] R749.13[医药卫生—临床医学]