【摘要】 目的探讨B7家族分子B7-H6在脑胶质瘤干细胞样细胞(GSLC)中的表达及其意义。方法使用亚克隆的方法并利用GSLC在体外形成神经细胞球的特性,从U87细胞株中筛选得到脑GSLC;通过实时定量PCR和流式细胞术检测干细胞标志分子c-myc、性别决定区基因Y盒2(Sox2)、CD133、神经上皮干细胞蛋白(nestin)、CXC趋化因子受体4(CXCR4)的表达;通过化疗药物多柔比星、顺铂、卡铂杀伤鉴定其是否具有耐药特性;通过实时定量PCR及流式细胞术获得脑GSLC中B7家族的表达;采用小干涉RNA(si RNA)干扰下调B7-H6表达,CCK-8法检测细胞增殖的变化。结果使用亚克隆方法分离获得的脑GSLC能在体外形成神经细胞球,上调表达多种干细胞标志分子CD133、nestin、CXCR4,具有显著的耐药特性。B7家族分子B7-H1、B7-H3、B7-H4和B7-H6在得到脑GSLC中均表达,与原代U87细胞相比,细胞膜上的B7-H6的表达发生显著上调,使用si RNA干扰B7-H6表达后,细胞的增殖受到抑制,且c-myc基因的表达也下调。结论 U87细胞来源的脑GSLCB7-H6表达上调,并且与GSLC生物学特性相关。更多还原

【Abstract】 Objective To investigate the expression and biological significance of costimulatory molecule B7-H6,a member of B7 family,in glioma stem like cells( GSLCs). Methods In virtue of the ability of forming neurospheres in vitro,GSLCs were isolated from U87 cells by cell sub-cloning. Real-time quantitative PCR and flow cytometry were performed to detect the expressions of stem cell related markers( c-myc,Sox2,CD133,nestin,and CXCR4),as well as the expressions of B7 family molecules. The different doses of adriamycin, carboplatin, cisplatin, were used to treat GSLCs for testing their chemotherapy-resistance. After the expression of B7-H6 in GSLCs was knockdown by si RNA,CCK-8 method was used to detect cell proliferation. Results GSLCs were successfully isolated from U87 cells,which formed neurospheres in vitro. The expressions of multiple stem cell markers were up-regulated and the GSLCs showed enhanced chemo therapy-resistance. B7 family members,B7-H1,B7-H3,B7-H4 and B7-H6 were expressed in GSLCs. Compared with primary U87 cells,GSLCs presented with a remarkably increased expression of B7-H6 on cell membrane. When B7-H6 was silenced by si RNA,cell proliferation was inhibited along with the decrease of c-myc expression. Conclusion The expression of B7-H6 is up-regulated in U87-derived GSLCs,which is associated with the biological characteristics of GSLCs.更多还原