Metadherin参与酸性微环境诱导的鼻咽癌紫杉醇耐药

Metadherin Involves in Acidic Extracellular pH-induced Paclitaxel Resistance in Nasopharyngeal Carcinoma

[目的]分析异黏蛋白（metadherin,MTDH）在酸性微环境诱导的鼻咽癌紫杉醇耐药中的作用并探讨其相关机制。[方法]不同浓度紫杉醇作用于鼻咽癌细胞CNE-2,48h后CCK-8法检测细胞生长抑制率,确定紫杉醇对CNE-2细胞的IC30。分别用p H 7.4、p H 6.8的细胞培养基培养CNE-2细胞,CCK-8法检测IC30浓度紫杉醇下CNE-2细胞生存率;相差显微镜下观察细胞形态改变;RT-q PCR、Western blot检测MTDH及上皮-间质转化（EMT）标志物表达情况;酸性环境下用si RNA沉默MTDH表达后,检测CNE-2细胞对紫杉醇敏感性及EMT标志物表达变化。[结果 ]紫杉醇对CNE-2细胞的IC30为6.167 nmol/L。IC30浓度下,p H6.8组细胞的生存率为48.46%±4.39%,明显高于p H 7.4组的31.30%±5.21%（P=0.013）。酸性环境下沉默MTDH表达后,CNE-2对紫杉醇的敏感性增强,IC30浓度下对照组和沉默组的细胞生存率分别为48.70%±2.35%和32.87%±2.97%（P=0.020）。沉默MTDH表达可逆转酸性引起的E-cadherin下降、Vimentin表达增强。[结论]沉默MTDH可逆转酸性微环境诱导的鼻咽癌CNE-2细胞紫杉醇耐药,这一现象与EMT进程逆转密切相关。

[Objective] To determine the role of metadherin（MTDH） in acidic extracellular p H（p He）-induced paclitaxel resistance in nasopharyngeal carcinoma（NPC）,and the related mechanism.[Methods] Nasopharyngeal carcinoma CNE-2 cells were stimulated with various concentrations of paclitaxel for 48 h,the proliferation inhibition rate was evaluated by the cell counting kit（CCK-8）,and IC30 value of paclitaxel was determined. Then,CNE-2 cells were incubated in normal（p H 7.4）or acidic（p H 6.8） medium,following IC30 paclitaxel stimulation for 48 h,the cell survival rate was evaluated by CCK-8 assay. Cell morphology was observed under phase contrast microscope. The expression of MTDH and epithelial-mesechymal transition（EMT） makers were detected by quantitative real-time reverse transcription-PCR（RT-q PCR） and Western blot. MTDH expression was blocked by small RNA（si RNA） silencing in NPC cells cultured at a p He of 6.8. Then,the changes of cell sensitivity to paclitaxel and EMT makers were assessed. [Results]The IC30 value of paclitaxel in CNE-2 cells was 6.167 nmol/L. The cell survival rate of CNE-2 cells cultured in acidic（p H 6.8） medium was significantly higher than that in normal（p H 7.4） medium（48.46%±4.39% vs31.30%±5.21%,P=0.013）. The sensitivity of CNE-2 to paclitaxel was enhanced after MTDH expression was silenced in acid environment,under the IC30 concentration,the cell survival rate in the control group and the silencing group was 48.70% ＋2.35% and 32.87% ＋2.97%,respectively（P=0.02）. Silencing of MTDH expression also sensitized acidic p He-induced down-regulation of Ecadherin and up-regulation of Vimentin. [Conclusion] Down-regulation of MTDH expression can reverse the acidic p He-induced paclitaxel resistance in NPC CNE-2 cells,which is closely correlated with MTDH-mediated EMT.