顺铂诱导舌鳞癌细胞发生上皮-间充质转化及获得肿瘤干细胞样特性的实验研究

Cisplatin induces epithelial-mesenchymal transition in tongue squamous carcinoma cells and obtains tumor stem cell-like characteristics

目的:建立舌鳞癌顺铂耐药株,观察其是否具有肿瘤干细胞样特性以及是否发生上皮-间充质转化(epithelialmesenchymal transition,EMT)。方法:使用顺铂梯度浓度递增法构建舌鳞癌顺铂耐药细胞株,通过MTS、流式细胞术和球囊培养对比舌癌顺铂耐药株和其亲本细胞株化疗耐药性及肿瘤干细胞特性的变化,通过qRT-PCR和Western免疫印迹检测肿瘤干性因子SOX2、OCT4、BMI1和ABCG2的表达,以及舌癌顺铂耐药株和其亲本细胞株中EMT的指标E-cadherin和Vimentin的表达水平,通过Transwell侵袭迁移实验研究侵袭和迁移能力的变化。采用SPSS 17.0软件包对数据进行统计学处理。结果:成功构建了2株舌鳞癌顺铂耐药株CAL27-res和SCC9-res,2株细胞对顺铂具有更强的耐药性。与亲本的舌癌细胞CAL27和SCC9相比,CAL27-res和SCC9-res细胞呈现间充质表型,Ecadherin显著下调(P<0.001),Vimentin和N-cadherin显著上调(P<0.001),侵袭、迁移能力显著增强(P<0.001)。同时,CAL27-res和SCC9-res细胞球囊形成能力增强,肿瘤干性因子SOX2、OCT4、BMI1和ABCG2上调(P<0.001),CD44^+/CD24^-细胞的比例增高(P<0.001)。结论:顺铂诱导的舌鳞癌耐药细胞发生EMT且获得了肿瘤干细胞样特性。

PURPOSE:To establish a cisplatin-resistant strain of tongue squamous cell carcinoma and observe whether it has tumor stem cell-like characteristics and whether epithelial-mesenchymal transition(EMT)occurs.METHODS:The cisplatin-resistant cell line of tongue squamous cell carcinoma was constructed by cisplatin gradient concentration method.The chemoresistance and tumor stem cell characteristics of tongue cisplatin-resistant strain and its parental cell line were compared by MTS,flow cytometry and balloon culture.The changes of factors related to tumor stem cell-like including SOX2,OCT4,BMI1 and ABCG2 were detected by qRT-PCR and Western blotting.The expression levels of EMT and Ecadherin and vimentin in tongue cancer cisplatin-resistant strains and their parental cell lines were detected by qRT-PCR and Western blot.Transwell modified chamber assay was used to study the invasion and migration ability of the two cell lines.The data were statistically processed using SPSS 17.0 software package.RESULTS:We successfully constructed two squamous cell carcinoma cisplatin-resistant strains CAL27-res and SCC9-res,and the two cell lines were more resistant to cisplatin.Compared with the parental cancer cells CAL27 and SCC9,CAL27-res and SCC9-res cells showed an interstitial phenotype,E-cadherin was down-regulated,vimentin and N-cadherin were up-regulated,and invasion and migration ability was enhanced.At the same time,CAL27-res and SCC9-res cells had enhanced balloon formation ability,tumor stem factors SOX2,OCT4,BMI1 and ABCG2 were up-regulated,and the proportion of CD44^+/CD24^-cells was increased.CONCLUSIONS:Cisplatin-induced tongue squamous cell carcinoma cells developed EMT and obtained tumor stem cell-like properties.