胶质瘤TJ905细胞及其干细胞抗凋亡与MRP基因的关系

目的 从人脑胶质瘤细胞TJ905中分离、培养肿瘤干细胞,并探讨其生物学特性及其抗凋亡和耐药基因的表达差异.方法 TJ905细胞经免疫磁珠分离获取CD133+细胞,用无血清培养方法获得肿瘤干细胞球,用含有10%的胎牛血清培养基诱导分化,分化前后分别做兔抗人巢蛋白抗体(nestin)、鼠抗人微小管相关蛋白β(tubulin-β)、兔抗人胶质纤维酸性蛋白质(GFAP)、免疫细胞化学荧光染色,WST-8检测干细胞增殖活性,应用实时荧光定量RT-PCR技术检测Livin、Livinα、Livinβ、生存素和MRP1、MRF3的表达,并观察肿瘤干细胞形态学改变.结果 TJ905细胞中含有少量CD133+细胞,约为0.21%,这些细胞具有典型的干细胞特性,nestin染色为阳性;在无血清培养基中呈悬浮球样生长,能够自我更新和增殖,在有血清培养基中能够分化,tubulin-β、GFAP染色为阳性.TJ905干细胞球Livin、Livinα、Livinβ、生存素和MRP-1、MRP-3 mRNA表达量较TJ905单层培养细胞表达量都有不同程度的降低.结论 TJ905细胞中含有少量肿瘤干细胞,TJ905干细胞球抗凋亡和MRP基因表达量较TJ905单层培养细胞表达量都有不同程度的降低,提示肿瘤干细胞在肿瘤耐药机制中发挥的作用不尽相同,也不总是造成肿瘤耐药的惟一因素.

Expression of anti-apoptotic and multi-drug resistance-associated protein genes in cancer stem cell isolated from TJ905 glioblastoma multiforme cell line

Objective To isolate cancer stem cells glioblastoma cells and detect the expression of anti-apoptotic and multi-drug resistance-associated protein (MRP) genes thereof. Methods CD133 positive ceils were isolated from human glioblastoma muhiforme cells of the line TJ905 by immunomagnetic beads technique and nestin, β-tubulin and GFAP expression were examined by Immunofluoresence staining. RT- PCR was used to detect the expression of livin, livinα, Livinβ, Survivin, MRP1, and MRP3. Results Only 0.21% of the TJ905 cells maintained in serum was CD133+ and showed characteristics of cancer stem ceils, positive in nestin. These cens maintained a sphere-like growth status in serum-free medium in vitro, and could self-renew, proliferate, conditionally differentiate into tubulin-β+ and GFAP+ cells, and produce neurons as well as glial cells. The mRNA expression levels of livin, livinα, survivin, MRP1, and MRP3 of the TJ905 tumor stem cells were significantly lower than those f the of TJ905 cells. Conclusion Cancer stem cells can be isolated from TJ905 glioblastoma muhiforme cells. However, the generating rate of the tumor stem cells is lower than that of the TJ905 cells, and the expression levels of anti-apoptotic and MRP genes are lower than those of the progenitor cells. Showing that cancer stem cells are not the solo factor to maintain tumor growth and resist apoptosis and to pump the anti-tumor drugs out of cells.