摘要
目的在体外通过大鼠胶质瘤C6细胞与大鼠骨髓间充质干细胞(bone marrow mesenchymal stem cells,BMSCs)非接触共培养,以探明BMSCs是否受到肿瘤微环境的作用获得肿瘤细胞的相关生物学特性。方法通过6孔板结合Transwell小室建立BMSCs和C6胶质瘤细胞的共培养体系,以共培养组为实验组,设单独培养的BMSCs为对照组;相差显微镜下观察培养后2组细胞形态学的改变;核干细胞因子(nucleostemin,NS)检测细胞增殖力的变化;荧光定量PCR和Western blot检测细胞中mdm2、p53mRNA水平及其蛋白的变化;免疫荧光法检测神经胶质细胞特异的胶质纤维酸蛋白(glial fibrillary acidic protein,GFAP)在细胞中的定位及表达。结果共培养后实验组细胞呈现类似胶质瘤细胞样的形态,表达神经胶质细胞特异的GFAP蛋白;NS蛋白反映的细胞增殖能力未发生改变;共培养后实验组p53mRNA水平明显低于对照组(P<0.01),而p53蛋白的表达水平显著高于对照组[(1.63±0.31)vs(0.85±0.12),P<0.05];实验组mdm2mRNA及其蛋白的表达水平均明显高于对照组(P<0.05)。结论肿瘤微环境可能会使大鼠骨髓间充质干细胞获得肿瘤细胞的相关生物学特性。
Objective To study whether rat bone marrow-derived mesenchymal stem cells（BMSCs）obtain tumor characteristics when co-cultured with rat glioma cells by non-contact culture in vitro.MethodsA co-cultural system was established between rat BMSCs and rat glioma cell line C6 by using 6-well plate and Transwell chamber.Cells were divided into 2 groups,experimental group（co-cultural BMSCs）,and control group（BMSCs cultured alone）.The morphological changes of BMSCs were observed by phase-contrast microscopy.Cell proliferation was determined by the level of nucleostemin（NS）protein.p53 and mdm2 mRNA were examined by real-time quantitative polymerase chain reaction（PCR）.The expression of p53 protein and mdm2 protein were detected by Western blotting.The expression and localization of glial fibrillary acidic protein（GFAP）was studied by immunofluorescence staining.ResultsExperimental cell group demonstrated glioma-like cell morphology and expressed GFAP protein.The ability of prolification had no significant change in experimental group compared with control group.p53 mRNA was decreased significantly（P〈0.01）and the level of p53 protein was increased also（P〈0.05）in experimental group as compared to those of control group.mdm2 mRNA and mdm2 protein were significantly increased as compared to control group（P〈0.05）.ConclusionCertain cytokines of tumor microenvironment can cause rat BMSCs to obtain tumor characteristics,which provides a preliminary experimental basis for the clinical safe application of BMSCs during cancer treatment.
出处
《第三军医大学学报》
CAS
CSCD
北大核心
2010年第7期630-633,共4页
Journal of Third Military Medical University
基金
国家自然科学基金(30670871)~~
