目的研究凋亡显像剂^(99)Tc^m-人膜联蛋白V(Annexin V)的制备及其与多巴胺能神经元凋亡模型的体外结合特性。方法运用双功能螯合剂联肼尼克酰胺(HYNIC)进行^(99)Tc^m 标记 Annexin V,形成^(99)Tc^m-HYNIC-Annexin V,经 Sephadex G-25柱层析分离纯化,采用快速薄层层析(ITLC)法检测放化纯。将^(99)Tc^m-HYNIC-Annexin V 与经过1-甲基-4-苯基吡啶离子(MPP^+)处理制模的大鼠肾上腺嗜铬细胞瘤PC12细胞进行体外细胞量及时间-温度结合实验、饱和结合实验和竞争结合实验.得到^(99)Tc^m-HYNIC-Annexin V 与多巴胺能神经元凋亡模型的结合特性,并比较其与正常细胞及不同凋亡水平下细胞模型的结合特性。结果①^(99)Tc^m-YNIC—Annexin V 标记率(64.56±6.23)%,比活度(3.7～74)×10~5kBq/mg,放化纯(93.6±2.48)%,室温放置4 h 后放化纯仍大于90%。②^(99)Tc^m-HYNIC-Annexin V 与多巴胺能神经元凋亡模型的结合具有特异性、可饱和性,Scatchard 图示平衡解离常数(K_d)为(7.16±1.78)nmol/L,最大结合容量(B_(max))为(178.73±32.62)fmol/10~6细胞。结论多巴胺能神经元凋亡模型对^(99)Tc^m-HYNIC-Annexin V 有良好的摄取,可进一步行动物体内研究。
Objective: To investigate the apoptosis of dopaminergic neurons and the protective effect of nicotine in 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine ( MPTP) -induced mouse model of Parkinson′s disease. Methods: The mouse model of Parkin-son's disease were formed by MPTP ( 30 mg /kg /d×7, i.p.) ; and the loss and apoptosis of dopaminergic neurons was ob-served by Tyrosine Hydroxylase( TH) and TUNEL stains.In "Nicotime plus MPTP"group, mice were pretreated with nicotinebefore MPTP injection. The putative protective effect of nicotine was analyzed. Results: The number of TH-positive cells de-creased during MPTP treatment. Apoptotic neurons began to appear after three injections of MPTP and peaked on the 8th day.In the MPTP-intoxicated mice treated with nicotine, the loss of TH-positive cells was significantly less than that of MPTP-treated group ( 30 mg /kg /d ×7) ( P < 0.05) . Conclusion: The chronic treatment of MPTP can induce the apoptosis ofdopaminergic neurons in substantia nigra, and nicotine might have a neuroprotecitve effect on dopaminergic neurons againstMPTP toxicity.
