9L脑胶质瘤干细胞模型建立及MRI成像表现

Flow sytometry isolation of glioma stem cells in 9L and establishment of rat intracerebral glioma model

目的：建立9L脑胶质瘤干细胞F344大鼠载瘤模型,采用3.0T磁共振成像评价肿瘤生长。方法：采用流式细胞仪分选CD133表达阳性细胞群,免疫荧光检测分选后细胞群的干细胞标记CD133、Nestin的表达情况。利用三维立体定向技术建立脑胶质瘤干细胞大鼠载瘤模型,定期行MR检查,观察颅内肿瘤生长情况。HE染色和免疫组织化学染色法观察肿瘤组织细胞形态和GFAP、S-100蛋白表达情况。结果：分选后细胞群CD133、Nestin蛋白表达阳性;干细胞荷瘤大鼠中位生存时间为（21.00±0.33）天;接种后1~3周MR扫描示肿瘤持续生长,增强扫描瘤体显像清晰,第3周末瘤周水肿显著;病理结果示肿瘤浸润生长,局部见出血、坏死,免疫组化示肿瘤组织胶质纤维GFAP阳性,S-100蛋白弱阳性。结论：流式技术分选9L脑胶质瘤干细胞建立的F344大鼠脑胶质瘤模型稳定可靠,符合恶性胶质瘤生物学特点;3.0T MR扫描仪能够动态观察脑胶质瘤生长状态。

Objective： To isolate glioma stem cells in 9L cells by flow sytometry and develop a reliable animal intracerebral glioma model. Methods： The cell subtypes expressed CD133^＋were isolated by flow cytometry technique.To characterize the separated cells expressed the stem cell- related makers（ CD133,Nestin） immunofluorescent staining experiments were performed. The 9L stem cells were injected stereotactiocally into the left caudate nucleus of 24 syngeneic Fischer 344 rats. Each rat received 1 × 10^6 cells in a volume of 10μl. Observated after implantation and MRI scan on every continuous first to third weekend. Hematoxylinand eosin- stained histology was performed to examine tumors directly for the presence of cellular organizational patterns within tumors. Tumors were sectioned after the rats natural death,the tumor samples were stained by HE,GFAP and S- 100 protein. Results： The isolated cells were characterized as a phenotype of CD133,Nestin positive. The median survival time of tumor- bearing rats was（ 21. 00± 0. 33） days. On the first to third weekend after injection,enhancement MRI showing intracerebral tumors were observed clearly with Gd- DTPA,and continued growing. Peritumoral edema was found at the third weekend. This tumor forms invasive,vascular tumors with histological features in F344 rat model. Immunohistochemistry GFAP protein stains of glial fibrillary between tumor cells was positive,S- 100,weakly positive. Conclusion： The method isolate stem cells in 9L cells by flow sytometry is successfully,and the F344 rat intracerebral glioma model is stable and reliable. Histopathological analysis of tumors resembled as malignant gliomas.