神经干细胞移植治疗大鼠无神经节细胞症的初步研究

摘要

目的 观察神经干细胞(Neural Stem cells,NSCs)在无神经节细胞症大鼠直肠壁内移植的存活和分化情况,以及对肠道运动和反射功能的改善,探讨神经干细胞移植对肠无神经节细胞症的治疗作用.方法 用0.5%苯扎氯铵溶液浸湿造模,造模成功后,随机分为两组,每组各30只.模型组用免疫荧光标记的提纯好的脐带间充质神经干细胞150 μL(浓度107/mL)分3处注入大鼠无神经节细胞肠壁内,对照组用生理盐水注射.并分别于注射后2,4,8周取材,观察大体表现,免疫荧光显微镜观察神经干细胞在病变肠管中的成活情况,免疫组化检测神经元特异性烯醇化酶( NSE)和S-100蛋白表达情况,钡灌肠检查肠道功能情况.结果 本组造模成功60例,无试验过程中死亡.模型组移植后2周取材,其中22只移植后可检测到肠壁内移植细胞存活,证明神经干细胞移植成功;4周取材可见分化为神经节细胞和星型胶质细胞,免疫组化可见肠壁内NSE和S-100蛋白表达;8周取材与4周结果 相同.注射生理盐水组无一只有神经节细胞存活.免疫组化检测,移植组病变肠段肠壁内神经细胞表达明显高于对照组.钡灌肠结果 显示,神经干细胞移植前后结肠形态上有明显不同.结论 脐带间充质干细胞来源的NSCs在无神经节细胞的肠壁内可以存活和分化,并在一定程度上恢复肠神经功能,从而为NSCs移植治疗无神经节细胞性巨结肠病提供了实验依据.%Objetive To elucidate the potential and the biological significance of grafting of central nervous system-derived NSCs( CNS-NSCs)as a therapeutic strategy for neuronal replacement in disorders of the enteric nervous system (ENS) such as aganglionic megacolon. Methods To induce chemical ablation of the enteric plexus in rat to produce an aperistaltic narrow segment simulating to Hirschsprung' s disease, the cat-ionic surfactant BAC was applied to a 2 cm serosal surface of the rectum. CNE-NSCs were transplanted into the denervated segment of rectum. After 2、4 and 8 weeks grafted cells were visualized in rectum tissue sections by fluorescent staining for neuronal and astrocytic markers. Results 60 rats arrived after chemical induce animal model. We separated into 2 groups (n = 30 for each group). We can see the alive NSCs in the enteric wall (22/30) after being transplanted into the denervated gut after 2 weeks injection. 4 weeks later, NSCs could survive and could differentiate into neurons and glial cells in vivo. At the 8-week assessment, some transplanted NSCs exhibited immunophenotypes of neuronal and glail cell. Barium enema show us there are enteric function improvement after NSCs transplantation. Conclusions Grafted CNS - NSCs can survive and differentiation , And restore the denervated rectum of rat in vivo nerve function to a certain extent, which indicates that NSCs provide a promising cellular replacement candidate for treatment of aganglionosis and some other disorder of the enteric nervous system.