上皮间质转化与卵巢癌细胞顺铂耐药的相关研究

Study on correlation between epithelial-mesenchymal transition and cisplatin resistance in ovarian cancer cells

目的研究人卵巢癌细胞顺铂耐药与上皮间质转化(EMT)的关系。方法以顺铂低剂量诱导结合间歇大剂量冲击的方法建立卵巢癌顺铂耐药细胞株A2780/DDP、OVCAR3/DDP。观察亲本细胞(A2780、OVCAR3)与耐药细胞(A2780/DDP、OVCAR3/DDP)形态;采用Transwell细胞迁移实验及细胞侵袭实验检查各细胞迁移、侵袭能力,Western blot检测EMT标记物E-cadherin、Vimentin的表达,免疫荧光检测E-cadherin的亚细胞定位。以5ng/mL TGF-β体外诱导卵巢癌细胞A2780发生EMT改变,MTS检测其对顺铂化疗敏感性的影响。结果 A2780/DDP、OVCAR3/DDP较A2780、OVCAR3的IC50分别提高了9.58、13.19倍。A2780/DDP、OVCAR3/DDP呈间充质细胞特点,获得侵袭、迁移能力,且上皮分子标志物E-cadherin表达下调,间充质分子标志物Vimentin表达上调。TGF-β体外诱导卵巢癌细胞发生EMT改变后对顺铂敏感性明显降低。结论 EMT与卵巢癌细胞顺铂耐药密切相关。

Objective To study the relationship between cisplatin resistance and epithelial-mesenchymal transition(EMT)in human ovarian cancer cells.Methods The cisplatin resistant cell lines A2780/DDP and OVCAR3/DDP of ovarian cancer were established by low dose induction combined with interval large dose of cisplatin.The morphology of parent cells(A2780,OVCAR3)and drug-resistant cells(A2780/DDP,OVCAR3/DDP)were observed.The migration and invasion abilities of above cells were detected by Transwell cell migration test and cell invasion experiment.The expressions of EMT markers E-cadherin and Vimentin were detected by Western blot.And the subcellular localization of E-cadherin was detected by immunofluorescence.The EMT change in ovarian cancer cells A2780 occurred by 5 ng/mL TGF-βin vitro induction.Its influence on the sensitivity of cisplatin chemotherapy was detected by MTS.Results The IC50 of A2780/DDP and OVCAR3/DDP increased by 9.58 times and 13.19 times of their parent cell lines A2780 and OVCAR3.Both A2780/DDP and OVCAR3/DDP cells were characterized by mesenchymal cell morphology,and acquired the ability of invasion and migration,moreover the expression of epithelial marker protein E-cadherin was downregulated and that of mesenchymal marker protein Vimentin was up-regulated.In addition,the sensitivity of ovarian cancer cells to cisplatin after TGF-βinducing EMT change in vitro was significantly reduced.Conclusion EMT is closely correlated with the cisplatin resistance in ovarian cancer cells.