^(99m)Tc-Annexin V检测早期凋亡多巴胺能神经元的实验研究  

Early Detection of Apoptotic Dopaminergic Neurons Using ^(99m)Tc-Annexin V

作　　者：黄金莎[1] 曹卫[2] 孙圣刚[1] 兰晓莉[2] 王涛[1] 张永学[2] 

机构地区：[1]华中科技大学同济医学院附属协和医院神经科,湖北武汉430022 [2]华中科技大学同济医学院附属协和医院核医学科,湖北武汉430022

出　　处：《中国神经免疫学和神经病学杂志》2007年第4期211-215,共5页Chinese Journal of Neuroimmunology and Neurology

基　　金：国家自然科学青年基金资助项目(30400516);武汉青年科技晨光计划基金资助项目(20045006071-2)

摘　　要：目的研究放射性核素标记的膜联蛋白V(Annexin V)与凋亡的多巴胺能神经元的结合特性,探讨使用凋亡显像剂99mTc-Annexin V早期活体显像诊断帕金森病的可行性。方法采用不同浓度的1-甲基-4-苯基吡啶离子(1-methyl-4-phenylpyridinium,MPP+)处理大鼠肾上腺嗜铬细胞瘤细胞(PC12)和人神经母细胞瘤细胞(SH-SY5Y)以诱导其凋亡(PC120~200μm/L,SH-SY5Y0~500μm/L),FITC-Annexin V及碘化吡啶(pro-pidiumiodide,PI)双染进行流式细胞仪凋亡检测。以99mTc-Annexin V与凋亡的细胞进行饱和结合实验及细胞摄取实验,研究凋亡细胞与Annexin V的亲和力及其摄取99mTc-Annexin V的动力学。结果MPP+可诱导PC12细胞及SH-SY5Y细胞发生凋亡,并有明显的量效关系。凋亡细胞可特异性与99mTc-Annexin V结合,亲和力可达(7.16±1.78)nmol/L,每个凋亡的多巴胺能神经元表面的结合位点可达到(179±33)f mol/106cells(PC12)及(220±26)f mol/106cells(SH-SY5Y),且神经元的凋亡水平与其膜结合的99mTc-Annexin V放射性强度有相关性(P<0.001)。结论凋亡的多巴胺能神经元与99mTc-Annexin V有高度亲和力,其所结合的99mTc-Annexin V放射性强度与细胞的凋亡水平相关,99mTc-Annexin V可用于检测多巴胺能神经元的早期凋亡。Objective To investigate the binding characteristics of radiolabeled Annexin V to apoptotic dopaminergic neurons, as to seek the feasibility of ^99mTc-Annexin V imaging for in vivo early detection apoptosis in Parkinson disease. Methods PC12 and SH-SY5Y cell lines were induced apoptosis by 1-methyl-4- phenylpyridinium （MPP＋） in different concentrations, then analyzed by flow cytometry with fluorescein isothiocyanate-conjugated Annexin V （FITC-Annexin V） and propidium iodide （PI） staining. Meanwhile, Annexin V was radiolabeled with ^99myc. The saturable binding and cell uptake test were carried out. The correlation of apoptotic rate detected by flow cytometry and cell uptake was analyzed. Results MPP＋ at different concentrations can induce cell apoptosis in a dose-dependent manner. ^99mTc-Annexin V can bind to apoptotic cells with a high affinity of 7.16±1.78 nmol/L, and the binding sites were 179±33 fmol/106cells （PC12） and 220±26 fmol/106cells （SH-SY5Y）. The membrane-bound radiolabeled Annexin V activity linearly correlated to total fluorescence as observed by FITC-Annexin V flow cytometry （r= 0. 924 for PC12 and r= 0. 937 for SH-SY5Y, P〈0.01）. Conclusions ^99mTc-Annexin V has a high affinity to apoptotic dopaminergic neurons, and the radioactivity correlated well with cell apoptosis rate, thus we can detect neuron apoptosis using ^99mTc-Annexin V in vivo.

关 键 词：帕金森病 膜联蛋白V 凋亡 放射性示踪剂 

分 类 号：R742.5[医药卫生—神经病学与精神病学]