【作者】 胡峰；

【导师】 雷霆；

【作者基本信息】 华中科技大学 ， 神经外科， 2011， 硕士

【摘要】 目的通过诱导胶质瘤干细胞样细胞的分化,对分化细胞及原肿瘤细胞干细胞特性及其生物学特性的差异进行对比,比较其干细胞表型特性及致瘤性,评价分化治疗恶性胶质瘤的可行性。方法采用干细胞条件下神经球方法培养胶质瘤细胞C6,胶质瘤原代细胞。利用添加全反式维甲酸(ATRA)诱导细胞分化,研究其不同的生长特点,包括细胞形态、自我更新能力等。RT-PCR、Western Blotting和流式细胞技术检测干细胞和分化细胞表面标志物的表达差异;Hoechst33342荧光染色流式细胞术检测其侧群细胞的含量;裸鼠颅内原位成瘤实验分析肿瘤干细胞及分化细胞肿瘤形成能力差异。采用统计学软件SPSS 17.0进行数据统计学分析。结果在干细胞条件培养的C6及原代胶质瘤干细胞通过添加全反式维甲酸引导细胞分化,RT-PCR和Western Blot提示分化后CD133、Nestin表达降低,GFAP表达升高。流式细胞技术检测提示CD133、SP比例降低。裸鼠颅内原位成瘤实验提示胶质瘤干细胞样细胞种植的裸鼠生存时间明显比分化后细胞种植的裸鼠生存时间降低。结论1.全反式维甲酸可以诱导胶质瘤干细胞样细胞的分化。2.分化后的胶质瘤细胞在自我更新能力及表面标记物的表达上较干细胞样细胞降低,其致瘤性也明显降低。3.分化治疗可能成为胶质瘤治疗的可行方法。更多还原

【Abstract】 Objective Use ATRA to induce stem-like glioma cells to differentiate and compare stem-like cells properties and biologic characteristics between stem-like glioma cells, including C6 cell lines, primary glioma cells and their differentiated cells.Methods Glioma cells were cultured with neurosphere-formation assay in serum-free supplemented medium. Stem-like glioma cells (SLGC) were differentiated with all-trans retinoic acid(ATRA)-containing medium. Growth characteristics, including morphological differences, capacity of self-renewal, were detected contrastively through neurosphere formation assay. Expressions of stem cell and differentiated cell markers were determined by reverse transcription polymerase chain reaction (RT-PCR), Western Blotting and Flow Cytometry(FCM). SLGC and differentiated cells were implanted stereotactically and intracranially into the Balb/c nude mice to contrast the capability of tumor initiation and the tumor models were detected by hematoxylin and eosin (HE) staining. All data analyses were performed with the SPSS software, version 17.0.Results Our data suggest that in vitro differentiation of SLGCs with ATRA induces the growth characteristics changes of glioma cells ,down-regulation of CD133 and Nestin expression and up-regulation of GFAP. In vivo mouse model, differentiated cells inoculated mice have better survival outcome. Conclusions1.ATRA can induce the differentiation of SLGCs.2.The tumorigenicity of stem-like glioma cells is reduced after ATRA differentiation.3.Differentiation treatment (ATRA) targeted the stem-like cell population is a candidate for adjuvant therapy against malignant gliomas.更多还原