摘要
目的:探讨Ⅰ型神经纤维瘤病(type 1 neurofibromatosis,NF-1)肿瘤组织中成纤维细胞(fibroblast,FB)体外分离、培养及鉴定的方法。方法:以4例NF-1患者的神经纤维瘤组织为材料,采用组织块法分离培养FB,细胞铺满培养瓶底后传代,差速贴壁法(15min/次)纯化P1、P2代FB。倒置显微镜观察细胞形态学变化,并用免疫细胞化学方法检测成纤维细胞特异蛋白-1、波形蛋白和S-100蛋白表达情况,对培养细胞进行鉴定。结果:组织块法原代培养约5d可见大量细胞从组织块周围迁出,20d左右达80%～90%融合。经纯化后P3代FB成纤维细胞特异蛋白-1和波形蛋白免疫组化染色阳性,阳性率分别为99.4%和99.6%;S-100染色阴性。结论:组织块培养法结合差速贴壁法可成功分离出高纯度NF-1肿瘤组织内FB,该培养方法是一种较理想的NF-1肿瘤组织FB培养方法。
Objective：To explore the method of isolation,culture and identification for type 1 neurofibromatosis（NF-1）neuro fibroma-derived fibroblasts in vitro.Method：The neurofibroma was harvested from four patients with NF-1,and the chipped explants were cultured to obtain the fibroblasts.Differential attachment method（15 minutes per time）were used to purify the P1 and P2 fibroblasts after the cell overgrew the bottom of the culture flask.The morphology of cultured cells was observed under inverted microscope,immunochemistry was used to test staining of the expression of fibroblast-specific protein-1,vimentin and S-100,and the fibroblast phenotype was identified either.Result：Large number of cultured cells emigrated from the chipped explants 5 days after the primary culture in tissue mass culture method,and 80%-90%confluence of cultured cells was evidenced about 20 days later.In the purified P3 fibroblasts,positive staining of fibroblast-specific protein-1 and vimentin and negative staining of S-100 was observed,with the rate of positive staining of 99.4%and 99.6%respectively.Conclusion：The high purified of NF-1 neurofibroma-derived fibroblasts can be isolated and cultured successfully by tissue mass culture method and differential attachment method.It is an ideal culture system.
出处
《中国脊柱脊髓杂志》
CAS
CSCD
北大核心
2010年第6期494-497,共4页
Chinese Journal of Spine and Spinal Cord
基金
国家自然科学基金项目(编号:30672131)
