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目的:观察顺铂(CDDP)诱导的人喉癌耐药细胞(hep-2/CDDP)中的上皮间质转化(EMT)情况,探讨EMT在喉癌顺铂耐药中的机制。方法:利用hep-2细胞和前期培养的hep-2/CDDP细胞株;Transwell、划痕实验检测细胞的侵袭迁移能力;分别提取人喉癌顺铂耐药细胞hep-2/CDDP及其亲代细胞hep-2mRNA、总蛋白,应用实时荧光定量PCR(RT-qPCR)、Western blot方法检测上皮表型E-cadherin、Zo-1,上皮间质转化转录因子Snail、Slug、Twist1,间质表型分子Vimentin mRNA及蛋白表达水平。结果:Transwell结果显示耐药细胞穿过基质胶的数量高于亲本细胞(P<0.05),划痕实验示hep-2/CDDP细胞划痕愈合面积百分比高于hep-2细胞(P<0.05),hep-2/CDDP细胞上皮表型E-cad、Zo-1mRNA及蛋白表达水平较hep-2细胞降低(均P<0.01),EMT转录因子Snail、Slug表达水平较hep-2细胞升高(均P<0.01),Twist1蛋白水平表达无明显变化(P>0.05),间质表型Vimentin mRNA及蛋白表达水平较hep-2细胞升高(均P<0.01)。结论:hep-2细胞的耐药性和侵袭迁移能力改变与顺铂诱导的上皮间质转变可能有关,抑制EMT相关通路可能为逆转喉癌顺铂耐药提供新思路。
OBJECTIVE: To observe the epithelial-mesenchymal transition (EMT) induced by cisplatin (CDDP) in human laryngeal carcinoma cell line hep-2 / CDDP and to explore the mechanism of EMT in cisplatin-resistant laryngeal carcinoma. Methods: Hep-2 / CDDP cells and hep-2 / CDDP cell lines were cultured with hep-2 cells and Transwell, scratch assay to detect the invasion and migration ability of hep-2 / CDDP cells and their parental cells hep-2 mRNA and total protein. The expressions of E-cadherin, Zo-1, Snail, Slug and Twist1 were detected by real-time quantitative reverse transcription polymerase chain reaction (qPCR) Vimentin mRNA and protein expression levels. Results: Transwell results showed that the number of drug-resistant cells crossing the matrigel was higher than that of the parental cells (P <0.05). Scratch experiments showed that the percentage of wound healing area of hep-2 / CDDP cells was higher than that of hep-2 cells (P <0.05) , the expression of E-cad and Zo-1 mRNA and protein in hep-2 / CDDP epithelial cells were lower than those in hep-2 cells (all P <0.01), and the expressions of Snail and Slug in hepatic stellate cells were higher than those in hep-2 cells (All P <0.01). There was no significant change in Twist1 protein expression (P> 0.05) and the expression of interstitial vimentin mRNA and protein in hepatic stellate cells was higher than that in hep-2 cells (all P <0.01). Conclusion: The change of drug resistance and invasion and migration of hep-2 cells may be related to cisplatin-induced epithelial-mesenchymal transition. Suppression of EMT-related pathways may provide new ideas for reversing cisplatin-resistant laryngeal cancer.