中华医学杂志
中華醫學雜誌
중화의학잡지
National Medical Journal of China
2013年
34期
2745-2749
,共5页
闫广宁%杨浪%崔有宏%蒋雪峰%王清良%郭德玉
閆廣寧%楊浪%崔有宏%蔣雪峰%王清良%郭德玉
염엄저%양랑%최유굉%장설봉%왕청량%곽덕옥
神经胶质瘤%内皮细胞%干性表型%Bmi1基因
神經膠質瘤%內皮細胞%榦性錶型%Bmi1基因
신경효질류%내피세포%간성표형%Bmi1기인
Glioma%Endothelial cells%Glioma stem cell-like phenotype%Bmi1 gene
目的 探讨B细胞特异的莫洛尼病毒插入位点1(Bmi1)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、Bmi1基因表达的影响以及抑制Bmi1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) cm3,P=0.000].(2)胶质瘤细胞与内皮细胞共培养后其CD133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Bmi1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Bmi1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Bmi1基因的GL261细胞共培养的CD133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Bmi1基因表达促进胶质瘤细胞的干性表型.
目的 探討B細胞特異的莫洛尼病毒插入位點1(Bmi1)基因在內皮細胞促進膠質瘤細胞榦性錶型中的可能作用.方法 以小鼠膠質瘤細胞繫GL261和小鼠腦內皮細胞繫b.END3為材料,採用Transwell雙室細胞共培養、極限稀釋法成毬實驗、實時定量PCR、Western印跡、流式細胞術、體內移植瘤實驗以及siRNA基因榦擾等方法,檢測內皮細胞對膠質瘤細胞體外成毬能力和體內成瘤能力、CD133暘性細胞比例、Bmi1基因錶達的影響以及抑製Bmi1基因錶達對上述現象的影響.結果 與膠質瘤細胞單獨培養的對照組相比:(1)膠質瘤細胞與內皮細胞共培養後其體外成毬能力明顯增彊,形成榦細胞毬數目明顯增多(40箇細胞/孔的濃度時為62.5％±1.5％比25.0％±4.6％,P=0.000),體積明顯增大；內皮細胞與膠質瘤細胞共同移植後所形成的移植瘤齣現早、體積更大[(0.798±0.297)比(0.362±0.123) cm3,P=0.000].(2)膠質瘤細胞與內皮細胞共培養後其CD133暘性細胞群比例增大(8.48％±0.78％比4.81％±0.37％,P=0.000).(3)膠質瘤細胞與內皮細胞共培養後Bmi1基因的mRNA(2.72±0.18比1.00±0.15,P=0.000)和蛋白錶達明顯增加.(4)利用siRNA榦擾膠質瘤細胞Bmi1基因錶達後,內皮細胞促進膠質瘤細胞榦性錶型的上述作用明顯減弱,敲低Bmi1基因的GL261細胞共培養的CD133暘性比例顯著低于共培養的普通GL261細胞(0.34％±0.21％比1.70％ ±0.69％,P=0.025).結論 內皮細胞可能通過上調膠質瘤細胞Bmi1基因錶達促進膠質瘤細胞的榦性錶型.
목적 탐토B세포특이적막락니병독삽입위점1(Bmi1)기인재내피세포촉진효질류세포간성표형중적가능작용.방법 이소서효질류세포계GL261화소서뇌내피세포계b.END3위재료,채용Transwell쌍실세포공배양、겁한희석법성구실험、실시정량PCR、Western인적、류식세포술、체내이식류실험이급siRNA기인간우등방법,검측내피세포대효질류세포체외성구능력화체내성류능력、CD133양성세포비례、Bmi1기인표체적영향이급억제Bmi1기인표체대상술현상적영향.결과 여효질류세포단독배양적대조조상비:(1)효질류세포여내피세포공배양후기체외성구능력명현증강,형성간세포구수목명현증다(40개세포/공적농도시위62.5％±1.5％비25.0％±4.6％,P=0.000),체적명현증대；내피세포여효질류세포공동이식후소형성적이식류출현조、체적경대[(0.798±0.297)비(0.362±0.123) cm3,P=0.000].(2)효질류세포여내피세포공배양후기CD133양성세포군비례증대(8.48％±0.78％비4.81％±0.37％,P=0.000).(3)효질류세포여내피세포공배양후Bmi1기인적mRNA(2.72±0.18비1.00±0.15,P=0.000)화단백표체명현증가.(4)이용siRNA간우효질류세포Bmi1기인표체후,내피세포촉진효질류세포간성표형적상술작용명현감약,고저Bmi1기인적GL261세포공배양적CD133양성비례현저저우공배양적보통GL261세포(0.34％±0.21％비1.70％ ±0.69％,P=0.025).결론 내피세포가능통과상조효질류세포Bmi1기인표체촉진효질류세포적간성표형.
Objective To explore the effects of B-cell specific Maloney leukemia virus integration site 1 (Bmi1) 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 coculture system,limit dilution assay,xenograft,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 Bmi1 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％ vs 25.0％ ±4.6％ at 40 cells/well,P =0.000).Xenografts generated by GL261 cells with b.END3 cells appeared earlier and were larger than that by GL261 cells alone ((0.798 ±0.297) cm3 vs (0.362 ±0.123) cm3,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 Bmi1 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 Bmi1 gene expression was inhibited by siRNA in glioma cells,CD133 positive portion of Bmi1-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.
