海洋单细胞海藻体外对人脑胶质瘤干细胞生物学活性的影响  

Marine phytoplankton effects on biological activity of human glioma stem cells in vitro

作　　者：段小兵[1,2] 关方霞[3] 邓晓辉[1] 杨波[1,2] 张天祥[1] 田毅[1] 乔晓俊[1] 李远[1] 梁硕[3] 朱万万[3] 

机构地区：[1]郑州大学第一附属医院神经外科,河南省郑州市450052 [2]河南省高等学校临床医学重点学科开放实验室,河南省郑州市450052 [3]郑州大学生物工程系,河南省郑州市450001

出　　处：《中国组织工程研究与临床康复》2011年第23期4215-4219,共5页Journal of Clinical Rehabilitative Tissue Engineering Research

摘　　要：背景:海藻具有广阔的药理活性前景,加强其研究对有目的进行应用开发有重要的指导意义。目的:观察海洋单细胞海藻在体外对人脑胶质瘤干细胞生物学活性的影响。方法:以酶消化法培养人脑胶质瘤干细胞,流式细胞分选出CD133阳性细胞,细胞传代获得第3代细胞。流式细胞仪检测海洋单细胞海藻作用前后细胞CD133表达变化,免疫组织化学检测贴壁细胞巢蛋白及胶质纤维酸性蛋白的表达。实验组分别加入不同质量浓度的海洋单细胞海藻,阴性对照加不含药的PBS,将稀释成4,6,8,10g/L的海洋单细胞海藻加入细胞培养液中并作用24,48,72h,流式细胞仪检测胶质瘤干细胞生长周期,应用酶标仪检测细胞生长抑制情况。结果与结论:随着浓度和时间的增加,与对照组相比倒置显微镜下可见实验组胶质瘤干细胞不易聚团成球,出现贴壁分化,并逐渐明显;加药后胶质瘤干细胞CD133表达量明显减少;出现的贴壁细胞免疫组织化学染色巢蛋白及胶质纤维酸性蛋白表达阳性;流式细胞仪检测显示,停滞在S、G2/M期细胞数增加,而G0/G1期细胞数减少;随着浓度和时间的增加,胶质瘤干细胞增殖明显抑制,与对照组相比差异有显著性意义(P<0.05～0.01)。提示海洋单细胞海藻能够抑制人脑胶质瘤干细胞的增殖,并促进其分化,且作用具有浓度和时间依赖性。BACKGROUND：Marine phytoplankton（MPPT） has important guiding significance for application to strengthen the research. OBJECTIVE：To investigate the effect of MPPT on biologic activity of brain gliomastem cells（BGSCs） in vitro. METHODS：BGSCs cultured in the enzyme digestion way. The CD133 positive cells were sorted out by flow cytometry,and the 3rd passage of BGSCs were gathered through sub-culturing. CD133 expression was identified by flow cytometry before and after the effect of MPPT. Immunohistochemisty was used to detect Nestin and glial fibrillary acidic protein（GFAP） expression of adherent cells. The experimental group was added with different concentrations of MPPT,the negative control group was added with just PBS. 4,6,8,10 g/L MPPT was added into the cell culture fluid. The cell cycle of BGSCs was inspected by flow cytometry,and inhibitory effects were detected using a microplate reader respectively at 24,48,72 hours later. RESULTS AND CONCLUSION：Compared to those in the control group,BGSCs in the experimental group could not form a compact,and began to be adherent and differentiated gradually observed by an inverted microscope,with the increasing of time and concentration. CD133 expression in BGSCs after the effect of MPPT was reduced obviously. The adherent cells expressed GFAP and Nestin. The cell number in S and G2/M phase was increased,and that in G0/G1 phase was decreased. Growth curve indicated that the proliferation of BGSCs was inhibited obviously,with the increasing of time and concentration（P 0.05-0.01） . Results indicate MPTT can inhibit the proliferation of BGSCs,and promote them differentiate,which shows a dose and time dependent effect.

关 键 词：海洋单细胞海藻 脑胶质瘤干细胞 CD133 巢蛋白 细胞周期 

分 类 号：R394.2[医药卫生—医学遗传学]