亚甲蓝对胶质瘤干细胞增殖作用的影响

Effect of methylene blue on proliferation of glioblastomas stem cells in vitro

目的研究亚甲蓝(MB)对胶质瘤干细胞增殖作用的影响。方法从新鲜人脑胶质母细胞瘤标本中分离脑肿瘤细胞,接种于含生长因子的无血清改良Eagle培养基/F-12(DMEM/F-12)培养基中培养至细胞形成干细胞球,用免疫荧光法对其进行检测和鉴定。随机将细胞分为四组:正常对照组,低剂量组,中剂量组,高剂量组。采用生长曲线法,软琼脂克隆形成法检测亚甲蓝对胶质瘤干细胞增殖作用的影响,流式细胞仪检测亚甲蓝作用后的胶质瘤干细胞的凋亡情况。结果生长曲线、集落形成能力结果和凋亡检测结果均显示,与对照组及低剂量组相比,亚甲蓝中、高剂量组细胞的增殖能力明显下降,且亚甲蓝的最低有效浓度为1μM,最佳有效浓度为10μM。结论亚甲蓝可以降低胶质瘤干细胞的增殖能力。

Objective The effect of methylene blue （MB） on proliferation of glioblastomas stem cells （GSCs）in vitro is investigated.Methods GSCs were isolated and inoculated in serum-free suspension culture.And GSCs were identified by immunofluorescent staining using antibody against nestin,CD133.Cultured GSCs were divided into four groups：control group,100 μM group,1 μM group and 10 μM group.Effect of MB on the proliferation of GSCs was detected by survival rates and the clonaity.The apoptosis of cells treated by MB was observed by annexin-Ⅴ and propidium iodide （PI） and flow cytometry.Results In the serum-free suspension medium,neurospheres that consisted of a great number of nestin-positive cells could be obtained.The proliferation of GSCs was observed.Growth curve assay,colony formation,and apoptosis all showed that the proliferation of GSCs in 100 μM and 10μMgroups was decreased significantly after MB treatment compared to control group and 1 μM group with a minimum effective concentration of 1 μM and an optimum effective concentration of 10 μM.Conclusion Methylene blue may inhibit the proliferation of GSCs efficiently.