一种新的神经胶质瘤原位移植模型的建立  

Establishment of a novel neuroglioma transplantation model in rats

作　　者：裘玮[1] 黄倩[1] 李惠明[1] 王丰[1] 伍瑛[2] 陈霞芳[1] 易苗英[1] 

机构地区：[1]上海交通大学附属第一人民医院中心实验室,上海200080 [2]上海交通大学附属第一人民医院B超室,上海200080

出　　处：《中国肿瘤生物治疗杂志》2007年第3期259-263,共5页Chinese Journal of Cancer Biotherapy

基　　金：国家重点基础研究发展计划("973"计划)项目(2004GB518800);国家自然科学基金杰出青年项目(30325043)~~

摘　　要：目的:建立一种新的可动态观察肿瘤细胞生长的大鼠神经胶质瘤原位移植瘤模型。方法:利用脂质体将含有强化绿色荧光蛋白(enhanced green fluorescence protein,EGFP)基因的真核表达质粒pEGFP-N1转染大鼠神经胶质瘤C6细胞,经G418筛选及亚克隆,获取稳定表达EGFP的细胞系C6-gfp。采用CCK-8试剂盒检测C6及C6-gfp细胞增殖情况;将C6-gfp细胞接种于Wistar大鼠颅内,通过B超、大体标本、荧光体视镜、H-E染色和免疫组化观察颅内成瘤情况及瘤组织中绿色荧光蛋白的表达。结果:流式细胞术分析体外培养的C6-gfp细胞97.7%产生绿色荧光;C6-gfp细胞与亲代C6细胞相比,生长曲线无明显不同(P>0.05)。C6-gfp接种于颅内3周后成瘤率达70%;利用B超可动态观察肿瘤生长,利用荧光体视镜可见肿瘤组织发出绿色荧光,可与周围正常组织区别。H-E染色可见肿瘤细胞核大,染色深,核分裂明显,瘤内有很多新生血管。免疫组化可见GFP阳性细胞染色深浅不一。结论:成功构建了能稳定高水平表达EGFP的大鼠神经胶质瘤C6-gfp细胞株,其生物学行为未发生改变,能在同系大鼠颅内成瘤。该模型为神经胶质瘤防治研究提供了良好的基础。Objective： To establish a novel neuroglioma transplantation model in rats to demonstrate the dynamic growth of tumor. Methods： pEGFP-N1, a eukaryotic plasmid containing enhanced green fluorescent protein （EGFP） gene, was transfected into the rats glioma cell line C6 with the help of liposome lipofectamine. Then the cell clones （ C6 - gfp） stably expressing EGFP was obtained through neomycin G418 screening and subcloning. The growth of C6 and C6-gfp cells were observed using Cell Counting Kit-8 （CCK-8）. Then the C6-gfp cells were transplanted into Wistar rat＇s cranium. The tumor formation was observed using B ultrasound and H-E staining. The EGFP expression in the tumor was detected using fluorescence stereomicroscope and anti-GFP immunohistochemistry. Results：Flow cytometry showed that 97. 7% of the screened C6-gfp cells were positive of EGFP. There was no obvious difference in the growth curves of C6 and C6-gfp cells （ P 〉 0.05 ）. The tumor formation rate in the rat＇s cranium was 70% after C6-gfp transplantation. The dynamic growth of tumor could be observed using B ultrasound and the expression of green fluorescence could be observed under the fluorescence stereomicroscope. H-E staining showed large nuclei in glioma ceils, deep dying with more nuclear divisions. New vessels could be seen in the tumor. Anti-GFP immunohistochemistry showed GFP-positive cells with distinct intensity. Conclusion：A C6-gfp cell line stably expressing GFP has been successfully established in the present study and it can readily form tumors in the cranium, which lays a foundation for further study of prevention and treatment of glioma.

关 键 词：神经胶质瘤 移植瘤模型 C6-gfp细胞系 瞬时转染 稳定转染 绿色荧光蛋白 

分 类 号：R739.41[医药卫生—肿瘤]