人胶质瘤干细胞趋化因子受体CXCR4活化促进血管生成的作用  

Activation of CXCR4 in human glioma stem cells promotes tumor angiogenesis

作　　者：平轶芳[1] 姚小红[1] 卞修武[1] 陈剑鸿[1] 章容[1] 易良[1] 周志华[1] 

机构地区：[1]第三军医大学西南医院病理学研究所,重庆400038

出　　处：《中华病理学杂志》2007年第3期179-183,共5页Chinese Journal of Pathology

基　　金：国家重点基础研究发展计划(973计划)基金(2006CB708503);国家自然科学基金(30670804)

摘　　要：目的从人恶性胶质瘤细胞系 U87中分离、培养和鉴定胶质瘤干细胞,观测其 CXCR4表达情况及其活化后促血管生成因子分泌的变化。方法通过流式细胞术检测 U87细胞中 CD133阳性细胞的比例。使用 CD133免疫磁珠分离试剂盒通过磁性细胞分选系统分离胶质瘤干细胞。采用间接免疫荧光标记、激光共聚焦扫描显微术观测胶质瘤干细胞中神经巢蛋白(nestin)、胶质纤维酸性蛋白(GFAP)、趋化因子受体 CXCR4的表达;以 CXCR4配体刺激通过钙流试验检测受体功能,采用酶联免疫吸附试验(ELISA)检测培养上清中血管内皮生长因子(VEGF)和白细胞介素-8(IL-8)的含量。建立裸鼠皮下移植瘤模型,观察胶质瘤干细胞成瘤情况及瘤体内 VEGF 表达情况。结果U87细胞系中 CD133阳性细胞的比例为0.5%,这些细胞具有干细胞增殖和生长特性;它们表达CXCR4,用其相应配体激活后导致胞内钙流增加、分泌 VEGF 和 IL-8增多。与 CD133阴性细胞相比,CD133阳性细胞在体外分泌 VEGF、IL-8多,在体内成瘤率高,形成的移植瘤生长迅速,表达更多VEGF。结论人恶性胶质细胞瘤细胞系 U87中含有极少量胶质瘤干细胞,表达功能性 CXCR4、分泌更多促血管生成因子,提示这些干细胞也直接参与胶质瘤血管生成。Objective To isolate, culture and identify glioma stem cells from human malignant glioma cell line U87, and investigate the changes of pro-angiogenic factors production by glioma stem cells followed by activation of CXCR4 and observe their tumorigenesis as well as the expression of vascular endothelial growth factor when implanted into nude mice. Methods The ratio of CD133 positive cells was detected by flow cytometry. Magnetic separation of CD133 positive cells was carried out on the magnetic cell sorting system （MACS）. Expression of nestin, glial fibrillary acidic protein （GFAP） and CXCR4 on tumorspheres was detected by indirect immunofluorescenco under confocal laser scanning microscopy. The functional activation of CXCR4 was assessed by calcium mobilization experiments. ELISA was used to detect the production of vascular endothelial growth factor （VEGF） and interleukin-8 （IL-8） in conditioned medium. Glioma stem cells were implanted into nude mice to assess their tumorigenesis ability and the expression of VEGF. Results The ratio of CD133 positive cells with stem cell property was 0. 5% in U87 cells. Activation of CXCR4 on glioma stem cells induced calcium mobilization and increased VEGF and IL-8 protein secretion. CD133 positive cells secreted more VEGF and IL-8 than their negative counterparts in vitro. Tumors derived from CD133 positive cells grew more rapidly and expressed elevated level of VEGF than their negative counterparts. Conclusions There are a small fraction of glioma stem cells in human glioblastoma cell line U87. Expressing functional CXCR4 and secreting more pro-angiogenic factors may be involved in tumor angiogenesis mediated by glioma stem cells.

关 键 词：神经胶质瘤 肿瘤干细胞 受体 CXCR4 新生血管化 病理性 

分 类 号：R739.41[医药卫生—肿瘤]