Bmil基因在内皮细胞促进胶质瘤细胞干性表型中的作用  

Effects of Bmil gene on endothelial cells promoting glioma stem cell-like phenotype

作　　者：闫广宁[1] 杨浪[1] 崔有宏[1] 蒋雪峰[1] 王清良[1] 郭德玉[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：《中华医学杂志》2013年第34期2745-2749,共5页National Medical Journal of China

基　　金：基金项目：国家自然科学基金（30870965）

摘　　要：目的探讨B细胞特异的莫洛尼病毒插入位点1（Bmil）基因在内皮细胞促进胶质瘤细胞干性表型中的可能作用。方法以小鼠胶质瘤细胞系GL261和小鼠脑内皮细胞系b．END3为材料，采用Transwell双室细胞共培养、极限稀释法成球实验、实时定量PCR、Western印迹、流式细胞术、体内移植瘤实验以及siRNA基因干扰等方法，检测内皮细胞对胶质瘤细胞体外成球能力和体内成瘤能力、CD133阳性细胞比例、Bmil基因表达的影响以及抑制Bmil基因表达对上述现象的影响。结果与胶质瘤细胞单独培养的对照组相比：（1）胶质瘤细胞与内皮细胞共培养后其体外成球能力明显增强，形成干细胞球数目明显增多（40个细胞／孔的浓度时为62．5％±1．5％比25．0％±4．6％，P=0．000），体积明显增大；内皮细胞与胶质瘤细胞共同移植后所形成的移植瘤出现早、体积更大[（0．798±0．297）比（0．362±0．123）cm2，P=0．000]。（2）胶质瘤细胞与内皮细胞共培养后其CDl33阳性细胞群比例增大（8．48％±0．78％比4．81％±0．37％，P=0．000）。（3）胶质瘤细胞与内皮细胞共培养后Broil基因的mRNA（2．72±0．18比1．00±0．15，P=0．000）和蛋白表达明显增加。（4）利用siRNA干扰胶质瘤细胞Bmil基因表达后，内皮细胞促进胶质瘤细胞干性表型的上述作用明显减弱，敲低Bmil基因的GL261细胞共培养的CDl33阳性比例显著低于共培养的普通GL261细胞（0．34％±0．21％比1．70％±0．69％，P=0．025）。结论内皮细胞可能通过上调胶质瘤细胞Bmil基因表达促进胶质瘤细胞的干性表型。Objective To explore the effects of B-cell specific Maloney leukemia virus integration site 1 （Broil） gene on endothelial cells promoting glioma stem cell （GSC）-like phenotype. Methods Glioblastoma cell line GL261 and brain micro-vessel endothelial cell line b. END3 were used. Transwell co- culture system, limit dilution assay, xenografl, real-time polymerase chain reaction （PCR） , Western blot, fluorescence activating cell sorter （FACS） and gene knock-down assay were used to determine the GSC-like phenotype and Broil gene expression in glioma cells. Results Compared with the control of GL261 cell alone, （1） more and larger tumor spheres formed after co-culturing with endothelial cells （62.5%±1.5%比25.0%±4.6% at 40 cells/well, P =0. 000）. Xenografis generated by GL261 cells with b. END3 cells appeared earlier and were larger than that by GL261 cells alone （ （0.798±0.297） vs （0.362±0.123）cm2, P = 0. 000 ）; （2） CD133 positive glioma cells increased after co-culturing with endothelial cells （8.48%± 0. 78% vs 4. 81%± 0. 37%, P = 0. 000） ; （3） the expression of ~Bmil in co-cultured glioma cells was up-regulated at mRNA level （2.72 ±0. 18 vs 1. 00±＋0. 15, P =0. 000） and at protein level; （4） the above phenomenon was attenuated when Broil gene expression was inhibited by siRNA in glioma cells, CD133 positive portion of Broil-knockdown GL261 cells co-culturing with b. END3 cells decreased than that ofwildtype GL261 cells （0.34% ±0.21% vs 1.70% ±0.69%, P=0.025）. Conclusion Endothelial cells promote GSC-like phenotype by up-regulating the expression of Bmil in glioma cells.

关 键 词：神经胶质瘤 内皮细胞 干性表型 Broil基因 

分 类 号：R739.4[医药卫生—肿瘤]