摘要
目的探讨胰岛素样生长因子Ⅰ型受体(IGF-1R)信号通路及上皮-间质转化(EMT)对EGFR野生型、K-ras突变型非小细胞肺癌A549细胞表皮生长因子酪氨酸激酶抑制剂(EGFR-TKIs)获得性耐药的作用。方法选用EGFR野生型、K-ras突变型的非小细胞肺癌A549细胞,用吉非替尼将其诱导成耐药细胞A549/GR。MTT法检测细胞增殖,划痕实验及Transwell侵袭实验检测细胞侵袭转移能力,qRT-PCR和Western blot法分别检测EMT标志分子及IGF-1R信号通路相关分子的mRNA和蛋白表达。结果 A549/GR对吉非替尼的敏感性显著下降(P<0.05)。与A549细胞比较,A549/GR细胞形态呈现间质化改变,侵袭和转移能力均显著增加(P<0.05)。A549/GR细胞中间质标志Vimentin的mRNA和蛋白表达量均显著增多(P<0.05),上皮标志E-Cadherin表达量显著减少(P<0.05)。与A549细胞相比,A549/GR细胞IGF-1R的mRNA和蛋白及其磷酸化表达量均未见明显改变(P>0.05),而AKT磷酸化水平及其mRNA水平明显上调(P<0.05),ERK蛋白及其磷酸化和mRNA表达量均明显增加(P<0.05)。A549/GR细胞的EGFR及其磷酸化蛋白表达量较A549细胞明显下降(P<0.05)。结论 EMT及AKT和ERK信号通路的激活均可能在EGFR野生型、K-ras突变型A549细胞的EGFRTKIs获得性耐药中起重要作用,但获得性耐药与IGF-1R表达无关。
Objective To investigate the roles of insulin -like growth factor Ⅰreceptor （ IGF-1R） signal path-way and epithelial-mesenchymal transition （ EMT） in the acquired resistance of epidermal growth factor receptor -tyro-sine kinase inhibitors （EGFR-TKIs） in EGFR wild-type and K-ras mutant non -small cell lung cancer （NSCLC） A549 cells.Methods The EGFR wild-type and K-ras mutant human NSCLC A549 cells were used in this study.The gefitinib-resistant A549 cells （named as A549/GR cells） were created by repeated exposure to gefitinib .MTT assay was used to measure the cell proliferation .Wound-healing array and transwell array were used to determine the invasive and migratory capabilities of cells .The gene and protein expressions of E -cadherin, vimentin, IGF-1R, AKT, ERK and EGFR were determined by qRT -PCR and Western blot, respectively.Results The A549/GR cells acquired resistance to EGFR-TKIs, with significantly reduction in sensitivity to gefitinib （P〈0.05）.Compared with A549 cells, A549/GR cells presented with mesenchymal phenotype , accompanied by significant enhancement of invasion and migration （ P〈0.05）.The expression of mesenchymal cell marker vimentin was also significantly increased in A 549/GR cells （ P〈0.05）, with significant reduction in expression of epithelial cell marker E -Cadherin （P〈0.05）.No significant differ-ence was found in the expression of IGF -1R mRNA, protein or its phosphorylated form between A 549 and A549/GR cells （P〉0.05）;while the expression of ERK in A549/GR cells were significantly increased （P〈0.05）, accompanied with significant up-regulation of mRNA level and phosphorylation level of AKT （P〈0.05）.The expression of EGFR and its phosphorylated form were significantly reduced in A 549/GR cells as compared with those in A549 cells （P〈0.05）.Con-clusion EMT and the activation of both AKT and ERK signal pathways may play important roles in acquired EGFR -TKIs-resistance in EGFR wild-type and K-ras mutant NSCLC A549 cells
出处
《广东医学》
CAS
CSCD
北大核心
2014年第10期1473-1477,共5页
Guangdong Medical Journal
基金
国家自然科学基金面上项目(编号:81172225)
