乳腺癌细胞蛋白质O-GlcNAc糖基化水平与紫杉醇抗肿瘤作用的关系研究

Relationship of protein O-GlcNAcylation levels with antitumor effect of Taxol in breast cancer

目的研究紫杉醇对乳腺瘤细胞蛋白质O-Glc NAc糖基化水平的影响及其机制,探讨蛋白质O-Glc NAc糖基化水平变化对紫杉醇抗肿瘤活性的影响。方法使用Western blot检测紫杉醇对乳腺癌细胞蛋白质O-Glc NAc糖基化水平及多种相关酶蛋白表达水平的影响;RT-q PCR检测紫杉醇对N-乙酰氨基葡萄糖苷酶(β-N-acetylglucosaminidase,OGA)和N-乙酰氨基葡萄糖转移酶(β-N-acetylglucosaminyltrans-fevase,OGT)mRNA水平的影响;采用OGA/OGT抑制剂PUGNAc/alloxan改变蛋白质O-Glc NAc糖基化水平,磺基罗丹明B法测定紫杉醇对肿瘤细胞增殖的影响。结果紫杉醇可诱导乳腺癌细胞MDA-MB-231蛋白质O-Glc NAc糖基化水平升高(P<0.05),且具有时间和浓度依赖性;紫杉醇还可同时诱导OGA和OGT mRNA和蛋白水平的升高(P<0.05),且具有浓度依赖性;采用OGA的抑制剂PUGNAc提高蛋白质O-Glc NAc糖基化水平后,细胞对紫杉醇的敏感性升高;采用OGT的抑制剂alloxan降低蛋白质O-Glc NAc糖基化水平后,细胞对紫杉醇的敏感性降低。结论紫杉醇可诱导蛋白质O-Glc NAc糖基化水平升高,蛋白质O-Glc NAc糖基化水平的改变也会影响乳腺癌细胞对紫杉醇的药物敏感性。

Objective To study effect of Taxol on protein O-GlcNAcylation levels and investigate whether protein O-GlcNAcylation levels can affect the sensitivity of breast cancer cells to Taxol.Methods Western blot analysis was performed to examine protein O-GlcNAcylation levels and the expression of enzymes related to O-GlcNAcylation biosynthesis in Taxol treated breast cancer cells.RT-qPCR was used to analyze the effects of Taxol on OGA and OGT mRNA expression in cancer cells.The sulforhodamine B colorimetric assay was used to determine the effect of alteration of protein O-GlcNAcylation on the anti-proliferation of Taxol in breast cancer cells by adding OGA inhibitor and OGT inhibitor, respectively.Results Taxol treatment enhanced protein O-GlcNAcylation levels in dose-and time-dependent manners in breast cancer cell MDA-MB-231 （ P 〈0.05 ） .Taxol increased the mRNA levels of OGT and OGA after MDA-MB-231 cells were treated for 24 h（P〈0.05）.As OGA inhibitors increased protein O-GlcNAcylation levels, the sensitivity of MDA-MB-231 to Taxol was increased.As OGT inhibitor decreased protein O-GlcNAcylation levels, the sensitivity of MDA-MB-231 to Taxol was reduced.Conclusion Taxol treatment can enhance protein O-GlcNAcylation levels and the changes of O-GlcNAcylation levels alter the sensitivity of breast cancer cell MDA-MB-231 to Taxol.