CAJ | 학술논문

目的探讨与脑胶质瘤干细胞相关的CD133表型.方法将无血清培养中的胶质瘤干细胞与含血清贴壁培养的胶质瘤细胞中CD133的表达进行对比.逆转录-聚合酶链反应(RT-PCR)及实时荧光定量PCR检测信使RNA(mRNA)水平的表达,流式细胞术及免疫荧光染色法检测蛋白水平的表达,激光共聚焦扫描成像进行CD133蛋白的亚细胞定位.结果所有胶质瘤细胞中均存在CD133 mRNA及蛋白的表达.AC133抗体所识别的糖基化CD133-AC133分子仅分布于无血清培养中的胶质瘤干细胞,阳性细胞比例为31.8％～99.9％,且定位于此类细胞膜表面.AC133阴性的胶质瘤干细胞及含血清贴壁培养胶质瘤细胞中CD133蛋白的表达位于细胞内,阳性细胞比例约为22.2％～88.6％,其亚细胞定位为内质网和/或高尔基体.结论细胞表面AC133分子,而非CD133 mRNA及细胞内CD133蛋白标记了一类胶质瘤干细胞亚群,但无血清培养环境下也存在AC133呈阴性表达的胶质瘤干细胞亚群.
목적 탐토여뇌효질류간세포상관적CD133표형.방법 장무혈청배양중적효질류간세포여함혈청첩벽배양적효질류세포중CD133적표체진행대비.역전록-취합매련반응(RT-PCR)급실시형광정량PCR검측신사RNA(mRNA)수평적표체,류식세포술급면역형광염색법검측단백수평적표체,격광공취초소묘성상진행CD133단백적아세포정위.결과 소유효질류세포중균존재CD133 mRNA급단백적표체.AC133항체소식별적당기화CD133-AC133분자부분포우무혈청배양중적효질류간세포,양성세포비례위31.8％～99.9％,차정위우차류세포막표면.AC133음성적효질류간세포급함혈청첩벽배양효질류세포중CD133단백적표체위우세포내,양성세포비례약위22.2％～88.6％,기아세포정위위내질망화/혹고이기체.결론 세포표면AC133분자,이비CD133 mRNA급세포내CD133단백표기료일류효질류간세포아군,단무혈청배양배경하야존재AC133정음성표체적효질류간세포아군.
Objective To discuss the CD133 phenotype correlating with glioma stem cells.Methods The expression of CD133 in glioma stem cell lines which were cultured in stem cell permissive serumfree medium was compared with that in conventional adherent glioma cell lines which were established in serum-containing medium.Reverse transcription-polymerase chain reaction (RT-PCR) and real-time quantitative PCR were performed to study the expression level of CD133 mRNA.Flow cytometry and immunofluorescent staining were done to study the protein expression.Confocal microscopy was adopted to study the subcellular localization of CD133 protein.Results CD133 mRNA and protein were distributed both in glioma stem cells and conventional adherent glioma cell lines.The glycosylated CD133-AC133 detected by AC133 antibody was only found in glioma stem cells cultured in serum-free medium and located on the cell membrane,and the proportion of positive cells was 31.8％-99.9％.The CD133 protein in the AC133 negative glioma stem cells and conventional adherent glioma cell lines was found intracellularly and located in the endoplasmic reticulum and/or Golgi apparatus,and the proportion of positive cells was 22.2％-88.6％.Conclusion Neither CD133 mRNA nor intracellular CD133 protein but AC133 phenotype on the cell membrane is closely correlated with glioma stem cells,but a group of AC133 negative glioma stem cells can also exist in serum-free medium.