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目的探讨热休克蛋白B1(heat shock protein B1,HSPB1)在U251细胞株及其干细胞中差异性表达对肿瘤细胞凋亡的影响。方法采用无血清培养基培养U251细胞,获得U251干细胞;应用细胞免疫荧光染色法检测CD133和nestin鉴定胶质瘤干细胞;应用iTRAQ技术对U251细胞株及其干细胞进行HSPs差异性分析;应用MTT法、Western blot和RT-PCR观察经卡莫司汀干预后,肿瘤细胞凋亡与HSPB1蛋白、基因表达变化的关系。结果在未经卡莫司汀干预的胶质瘤与胶质瘤干细胞的HSPB1的蛋白比值为0.523;经卡莫司汀干预U251及其干细胞后,HSPB1蛋白含量分别升高了1.4、3.3倍,HSPB1 mRNA分别升高1.5、3.6倍,经相同浓度的卡莫司汀干预后的U251细胞生长受到抑制,而U251干细胞无明显抑制。结论 HSPB1参与U251胶质瘤干细胞化疗抵抗过程,对肿瘤细胞增殖、凋亡起着重要的调控作用。
Objective To investigate the effect of heat shock protein B1 (HSPB1) on the apoptosis of tumor cells in U251 cell line and its stem cells. Methods U251 cells were cultured in serum-free medium to obtain U251 stem cells; CD133 and nestin were detected by immunofluorescence staining to identify glioma stem cells; iTRAQ technique was used to analyze the difference of HSPs between U251 cells and stem cells; Western blot and RT-PCR were used to observe the relationship between tumor cell apoptosis and HSPB1 protein and gene expression after intervention with carmustine. Results The protein ratio of HSPB1 in glioma and glioma stem cells without intervention of Carmustine was 0.523. The HSPB1 protein content in U251 and its stem cells was increased by 1.4 and 3.3 times by Carmustine, HSPB1 mRNA increased 1.5 and 3.6 times respectively. The growth of U251 cells was inhibited by the same concentration of carmustine intervention, while the U251 stem cells had no obvious inhibition. Conclusion HSPB1 is involved in the chemoresistance of U251 glioma stem cells and plays an important regulatory role in the proliferation and apoptosis of tumor cells.