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目的探讨PIB5PA(phosphatidylinositol 4,5-bisphosphate5-phosphatase)基因转染人三阴性乳腺癌MDA-MB-231细胞后对紫杉醇敏感性的影响。方法体外培养人乳腺癌MCF-7和MDA-MB-231细胞株,MTT法观察不同浓度(0,0.1,0.2,0.3,0.4,0.5μmol/L)紫杉醇处理后对两株细胞生存率的影响;流式细胞PI单染法检测两株细胞凋亡率的差异;蛋白免疫印迹法检测0.3μmol/L紫杉醇处理两株细胞不同时间点Bcl-2家族成员蛋白表达差异。将p CGNPIB5PA质粒转染对紫杉醇相对不敏感MDA-MB-231细胞,0.3μmol/L紫杉醇处理24h,免疫蛋白印迹法检测PIB5PA,磷酸化Akt以及Bcl-2家族成员蛋白的表达,MTT和PI单染法检测MDA-MB-231细胞对紫杉醇敏感性的改变。结果MDA-MB-231细胞对紫杉醇敏感性较差,过表达PIB5PA后细胞生存率明显降低,细胞中磷酸化Akt水平降低,Bim(Bcl-2 interacting mediator of cell death)表达明显增高,对紫杉醇敏感性明显增强。结论 PIB5PA可能通过上调Bim表达增强三阴性乳腺癌MDA-MB-231细胞对紫杉醇的敏感性。
Objective To investigate the effect of PIB5PA (phosphatidylinositol 4,5-bisphosphate 5-phosphatase) gene on paclitaxel sensitivity in human triple negative breast cancer cell line MDA-MB-231. Methods The human breast cancer MCF-7 and MDA-MB-231 cell lines were cultured in vitro. The effects of different concentrations of paclitaxel (0, 0.1, 0.2, 0.3, 0.4 and 0.5 μmol / L) Flow cytometry was used to detect the difference between the two cell apoptosis rates. Western blotting was used to detect the protein expression of Bcl-2 family members at different time points after treated with 0.3μmol / L paclitaxel. P CGNPIB5PA plasmid was transfected into paclitaxel-sensitive MDA-MB-231 cells and treated with 0.3 μmol / L paclitaxel for 24 h. The expression of PIB5PA, phospho-Akt and Bcl-2 protein was detected by Western blotting. MTT and PI Dye assay was used to detect the change of paclitaxel sensitivity in MDA-MB-231 cells. Results The sensitivity of paclitaxel to MDA-MB-231 cells was poor. The survival rate of MDA-MB-231 cells was significantly decreased after overexpression of PIB5PA and the phosphorylation of Akt was decreased. The expression of Bim-2 interacting mediator of cell death was significantly increased, Sex is obviously enhanced. Conclusion PIB5PA may enhance the sensitivity of paclitaxel to triple negative breast cancer MDA-MB-231 cells by up-regulating Bim expression.