Oct4通过类泛素修饰途径维持胶质瘤干细胞增殖潜能

Oct4 maintains the proliferation potential of glioma stem cells by modifying its ubiquitin

目的研究类泛素蛋白(SUMO)与肿瘤干细胞"干性"诱导基因Oct4共同维持胶质瘤干细胞于持续增殖状态的分子机制。方法采用极低密度细胞接种法培养C6胶质瘤肿瘤干细胞克隆球,血清还原+免疫荧光方法验证肿瘤干细胞分化能力。实验设对照组、无义组和SUMO1组。持续测量克隆球直径,免疫荧光方法检测Ki67、CD133、巢蛋白和胶质纤维酸性蛋白(GFAP)表达情况,Western blot法检测Oct4和SUMO1的表达水平。结果肿瘤细胞克隆球被血清诱导分化后高表达胶质细胞标记蛋白GFAP。与对照组和无义组比较,SUMO1组细胞克隆球直径增长速度、Ki67、CD133、巢蛋白阳性率明显降低(P<0.05),GFAP阳性率明显升高[(57.35±7.87)%vs(1.09±0.27)%,(0.87±0.21)%,P<0.05],SUMO1组游离及共价结合状态的SUMO1及Oct4明显降低(P<0.05)。结论 Oct4通过与SUMO1共价结合共同维持胶质瘤干细胞持续增殖潜能。

Objective To study the molecular mechanism of SUMO and ＂sternness＂ gene Oct4 in maintaining the proliferation of glioma stem cells. Methods C6 glioma stem cells were cultured by inoculating ceils with an extremely low density. The differentiation of glioma stem cells was identified by serum reduction and immunofluorescence staining. The glioma stem cells were divided into control group, nonsense group and SUMO1 group. The diameter of cloned glioma stem cells was measured. Expressions of Ki67, CD133, nestin, GFAP, and those of Oct4 and SUMO1 were detected by immunofluorescence staining and Western blot, respectively. Results The GFAP was highly expressed in the differentiated glioma stem cells. The diameter of glioma stem cells was significantly longer and the positive rate of Ki67, CD133,nestin was significantly lower in SUMO1 group than in control group and nonsense group （P〈0. 05）. The positive rate of GFAP was significantly higher in SUMO1 group than in control group and nonsense group （57.35%±7.87% vs 1.09±0.27%,57.35%±7.87% vs 0. 87%±0.21%,P〈0.05）. The free and covalent SUMO1 and Oct4 binding rate was significantly lower in SUMO1 group than in control group （P〈0.05）. Conclusion Oct4 can maintain the proliferation potential of glioma stem cells by covalently binding to SUMO1.