下咽癌放疗抵抗细胞放疗抗性与上皮–间质转化的关系

The Relationship Between the Radioresistance of Hypopharyngeal Carcinoma Cells FaduRR and Epithelial Mesenchymal Transformation

该文探讨了下咽鳞癌放疗抵抗细胞FaduRR发生放疗抵抗与上皮间质转化(epithelial-mesenchymal transition,EMT)的关系及可能的机制。课题组前期成功构建了放疗抵抗细胞株FaduRR。平板克隆形成实验、CCK8实验检测放疗抵抗性,流式细胞术检测细胞凋亡、细胞划痕和Transwell侵袭迁移实验检测迁移侵袭能力,Western blot和qRT-PCR实验检测蛋白和mRNA的表达。与亲本细胞Fadu相比,FaduRR细胞的放疗敏感性明显降低,X线照射后第4天的存活率明显增高、细胞凋亡率降低、细胞形态呈现上皮间质化的改变、细胞迁移侵袭能力增强、E-cadherin的蛋白和mRNA表达水平显著下调,而N-cadherin、Vimentin、p-Akt、p-GSK-3β、Snail的蛋白和mRNA表达水平显著上调,但Akt和GSK-3β的表达无明显变化。以上结果提示,下咽鳞癌放疗抵抗细胞株FaduRR发生了EMT改变,其机制可能与Akt/GSK-3β/Snail信号通路的激活有关。

The aim of this study was to explore the relationship between the radioresistance FaduRR cell and the EMT (epithelial-mesenchymal transformation) as well as its potential mechanism in hypopharyngeal squamous carcinoma.The radioresistant hypopharyngeal cancer cell FaduRR was previously established with a gradient cumulative irradiation dose.The radioresistance of FaduRR was measured by colony forming assay and CCK8 cell viability kits.The apoptosis rate was measured by flow cytometry.The migration and invasion abilities were investigated by wound healing assay and transwell assay.Moreover,the mRNA and protein levels were detected by Western blot and qRT-PCR assay.Compared with Fadu cells,FaduRR cells radiation sensitivity was significantly lower.The CCK8 cell viability kits results showed that the survival rate of FaduRR cells in different doses on the fourth day after irradiation were higher than that of parental cells,the apoptosis rate of FaduRR cells was lower.The radioresistant cells FaduRR demonstrated typical ‘cobblestone’ or spindle-like shape,with pseudopodia and enlarged intercellular space.The invasiveness and migration abilities of the FaduRR cells were enhanced.The mRNA and protein expression levels of E-cadherin were significantly decreased,while the mRNA and protein expression levels of N-cadherin and Vimentin were markedly increased after as compared with parental cells.In the Akt/GSK-3β/Snail signaling pathway,the protein levels of p-Akt,p-GSK-3β and Snail were significantly increased,while the expression of Akt and GSK-3β was not significantly changed.According to the results,the radioresistant hypopharyngeal cancer cell FaduRR can induce the epithelial mesenchymal transformation,and the specific mechanism may be related to the activation of Akt/GSK-3β/Snail signaling pathway.