全反式维甲酸协同DAPT抑制胶质瘤干细胞的自我更新  

Synergistic Inhibition Effects of ATRA and DAPT on the Ability of Self-renewal of Glioma Stem Cells

作　　者：龚爱华[1,2] 熊二梦[1] 张严[1] 杜凤移[1] 彭琬昕[1] 邵根宝[1] 金洁[1] 程建军[1] 

机构地区：[1]江苏大学基础医学与医学技术学院,镇江212013 [2]癌基因及相关基因国家重点实验室,上海200032

出　　处：《中国细胞生物学学报》2013年第10期1477-1484,共8页Chinese Journal of Cell Biology

基　　金：江苏省高校自然科学基金(批准号:N07KJB310018);癌基因及相关基因国家重点实验室开放课题(批准号:90-13-05);国家自然科学基金(批准号:31100964;81372718)资助的课题~~

摘　　要：前期研究脑表明,脑胶质瘤干细胞(glioma stem cells,GSCs)是胶质瘤发生和发展的重要因素,探索靶向干预GSCs生长有可能成为脑胶质瘤治疗的有效途径之一。该研究旨在阐明两种药物ATRA和γ-分泌酶抑制剂DAPT协同抑制GSCs自我更新的生物学效应。通过用台盼蓝排染法、克隆球形成试验和免疫印迹分析了两种药物的单独使用或联用对GSC样细胞PGC1和PGC2生长、成球能力和自我更新以及干细胞标志物表达的影响。结果发现,单独使用ATRA对PGC1生长有一定的抑制作用,而对PGC2生长几乎没有影响;DAPT对PGCs的生长抑制作用明显强于ATRA。高浓度ATRA(80μmol/L)能诱导PGCs的分化,降低PGCs成球大小,且成球效率降至5%～8%,而正常对照组为32%～35%;同样,DAPT(40μmol/L)也能降低PGCs成球大小,且成球效率降至2%～3%;低浓度ATRA(20μmol/L)和DAPT(5μmol/L)对PGCs自我更新能力和干性没有明显影响,而联合使用后其明显降低PGCs的成球大小,且成球效率降至3%～5%,促进细胞凋亡,并且明显抑制了干细胞标志物Nestin、CD133、Sox2、Oct4的表达,提高了分化标志物GFAP的表达。该研究证明了低浓度的ATRA和DAPT能协同抑制脑胶质瘤干细胞PGCs的自我更新。研究结果将为脑胶质瘤的临床研究提供实验依据。Previous studies suggest that glioma stem cells （GSCs） play important roles in tumoragenesis and development of glioma. It is a promising therapeutic strategy to explore the approach to inhibition of GSCs growth. In this study, we assessed synergistic inhibition effects of ATRA and DAPT on GSCs through the ability of GSCs self-renwal. We examined the inhibition efficiency of GSC-like PGCs growth using trypan blue staining, the ability and efficiency of sphere formation by sphere formation assay under microscope, apoptosis and cell cycle by FACS, and the expression of markers of stem cells through Western blot. The results indicated that ATRA hadlimited inhibition of PGC 1 growth, whereas few effects on PGC2 growth. DAPT obviously inhibited the growth of PGCs compared with ATRA. Furthermore, ATRA （80 μmol/L） induced the differentiation of PGCs, and reduced sphere fromation in size and efficiency by 5%-8% compared with that of control group by 32%-35%. Similarily, DAPT （40 μmol/L） decreased sphere fromation in size and efficiency by 2%-3%. We also found that the ability of self-renwal and stemness was hardly affected in PGCs treated with ATRA （20 μmol/L） or DAPT （5 μmol/L）, respectively. On the contrary, combinition of both drugs resulted in decrease in sphere size and sphere formation efficiency by 3%-5%, promoted apoptosis and significantly downregulated the expression of markers of stem cells Nestin, CD133, Sox2, Oct4 and upregualted the expression of GFAP. Our finding con- firmed the synergistic inhibition effects of ATRA and DAPT on GSCs self-renwal, which might provide the evi- dence for further clinical research a^ainst ~lioma stem cells.

关 键 词：ATRA DAPT 脑胶质瘤干细胞 自我更新 

分 类 号：R739.4[医药卫生—肿瘤]