摘要
目的:探讨成年豚鼠嗅球嗅神经鞘细胞(Olfactoryensheathingcells,OECs)取材、体外培养及纯化方法,为以豚鼠作为实验模型的神经移植实验及进一步研究其生物学特性打下基础。方法:在立体显微镜下仔细分离出嗅球的第一二层,经剪碎、消化及吹打后去除沉淀,并行培养前差速贴壁法初步纯化及培养后第7～10天用消化法行近一步纯化,用神经生长因子受体蛋白(p75NGFR)行免疫组织化学鉴定及估算OECs纯度。结果:体外培养成年豚鼠OECs形态与成年大鼠基本相似;未经纯化处理的嗅球组织中的OECs,在培养2周时被大量增殖的成纤维细胞所覆盖和取代。而经过精细取材、培养前纯化和培养后纯化的综合处理,可使OECs纯度达到70%。结论:精细取材和综合纯化可培养出纯度较高的OECs,为以豚鼠作为实验模型的神经移植实验及进一步研究其生物学特性打下基础。
AIM:To explore the methods o f dissociati on,culture in vitro and purification of olfactory ensheathing cell s(OECs )from adult guinea pig olfactory bulb s so as to lay foundations of fur ther rese arch on the biological characteristics of OECs and experiment of neura l regeneration based on experimental model o f adult guinea pig.METHODS:The ti ssue in the first two layers of o lfactory bulb was dissoci-ated carefully unde r anatomic micro scope,cut up and then incubated in 3mL /L collagenaseⅠfor 30min.After centrifuged,the ce ll pellet was dis-persed and the deposition was removed;the supernated fluid was initiall y purified with differential atta chm ent method before culture,and the fu rther purification was made with diges tiv e method from the 7th to 10th day after culture in vitro;and the purity of OECs was estimated and appraised by immunostaining positively for NGF r ecept or(p 75NGFR).RESULTS:The morphological features of OECs in vitro from adu lt guinea pig olfactory bulbs were basically s imilar to that of the cells from adult rats;and the OECs without purified were mo stly covered and replaced by p rolif-erated fibroblasts.While through r efined dissociation and synthetic p u rifica-tion both before and after culture,t he purity of OECs reached about 7 0%.Through refined dissociation and sy nthetic purification,a h igher purity of OECs from adult guin ea pig olfactory bulbs can be obtained,whi ch lays foundations of further re search on the biological characteristics of OE Cs and experiment of
出处
《中国临床康复》
CSCD
2004年第2期244-245,T004,共3页
Chinese Journal of Clinical Rehabilitation
