EMT在大肠癌细胞奥沙利铂耐药中的作用及分子机制研究

摘要

目的：探讨上皮‐间叶转化（EMT）在大肠癌耐药中的作用及分子机制。方法采用药物浓度递增的方法建立大肠癌LOVO细胞奥沙利铂（L‐OHP）耐药细胞株LOVO／L‐OHP，免疫荧光和蛋白质印迹法（Westernblot）检测LOVO和LOVO／L‐OHP细胞E‐cadherin和Vimentin表达；Westernblot检测核转录因子Snail、Twist表达；噻唑蓝（MTT）检测细胞增殖。结果与LOVO细胞比较，LOVO／L‐OHP皮表型消失，细胞膜E‐cadherin表达减弱（22．63±3．25）％（P＜0．01），获得间叶细胞表型，表达Vimentin（475．42±58．36）％（P＜0．01）。LOVO／L‐OHP细胞株Twist表达轻度增加（116．42±18．36）％（P＞0．05），Snail表达显著增加（382．18±41．33）％（P＜0．01）。siSnail上调E‐cadherin表达（246．82±31．57）％（P＜0．01），下调Vimentin表达（28．75±3．96）％（P＜0．01）；siSnail显著增强LOVO／L‐OHP细胞株L‐OHP化疗敏感性，对照组和siSnail组的IC50分别为23．75μg／mL和12．42μg／mL。结论EMT在大肠癌细胞L‐OHP耐药中可能起重要作用，抑制EMT可恢复耐药细胞化疗敏感性。%Objective To investigate the role and molecular mechanism of epithelial‐mesenchymal transition (EM T ) in che‐moresistance to oxaliplatin in colorectal cancer cells .Methods Oxaliplatin resistant LOVO/L‐OHP cells were established by gradu‐ally increasing the concentration of oxaliplatin and intermittent treatment with high‐dose concentration on parental cells (LOVO) . The expression of E‐cadherin and Vimentin was detected by indirect immunofluorescence and Western blot analysis .The expression of Snail and Twist was detected by Western blot analysis .cell proliferation was detected by MTT .Results Compared with LOVO cells ,the epithelial phenotype of LOVO/L‐OHP cell line was lost ,and the expression of E‐cadherin was decreased (22 .63 ± 3 .25)% (P<0 .01) ,an increase in the mesenchymal marker Vimentin (475 .42 ± 58 .36)% (P< 0 .01) .LOVO/L‐OHP cell line Twist expression was slightly increased (116 .42 ± 18 .36)% (P> 0 .05) ,Snail expression was significantly increased (382 .18 ± 41 .33)% (P<0 .01) .The expression of siSnail increased E‐cadherin (246 .82 ± 31 .57)% (P<0 .01) .The expression of Vimentin (28 .75 ± 3 .96)% (P< 0 .01);siSnail significantly enhanced sensitivity to oxaliplatin based chemotherapy in LOVO/L‐OHP cell line ,IC50 control group and siSnail group were 23 .75 μg/mL and 12 .42 μg/mL .Conclusion EM T may play an important role in chemoresistance to oxaliplatin in colorectal cancer cells ,inhibition of EM T can restore chemosensitivity of resistant colorectal cancer cells.