Suppression of the Epidermal Growth Factor Receptor Inhibits Epithelial-Mesenchymal Transition in Human Pancreatic Cancer PANC-1 Cells
Suppression of the Epidermal Growth Factor Receptor Inhibits Epithelial-Mesenchymal Transition in Human Pancreatic Cancer PANC-1 Cells
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DOI：
10.1007/s10620-012-2036-4
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发表时间：
2012-05-01
影响因子：
3.1
通讯作者：
Yang, Yin-Mo
中科院分区：
文献类型：
--
作者：
Chang, Zhi-Gang;Wei, Jun-Min;Yang, Yin-Mo
表皮生长因子受体(EGFR)在胰腺癌中有异常表达，但其诱导胰腺癌发生的机制尚不清楚。为探讨表皮生长因子受体(EGFR)在胰腺癌细胞上皮-间充质转化(EMT)中的作用，采用慢病毒表达载体将EGFR小干扰RNA导入胰腺癌细胞系PANC-1，建立了EGFR基因敲除的胰腺癌细胞系(SI-PANC-1)。以表达阴性对照序列慢病毒载体的PANC-1细胞作为阴性对照(NC-PANC-1)。采用划痕实验和Transwell实验分析细胞迁移和侵袭情况。应用实时定量聚合酶链式反应和Western blotting检测EMT标志物E-钙粘蛋白、N-钙粘蛋白、波形蛋白和纤维连接蛋白以及转录因子Snail、Slug、Twist1和SIP1在PANC-1、NC-PANC-1和SI-PANC-1细胞中的表达。免疫荧光染色和共聚焦显微镜观察其细胞定位和形态变化。经EGFR的RNA干扰后，si-panc-1细胞的迁移和侵袭能力显著降低。上皮表型标记物E-cadherin表达增加，间充质表型标记物N-cadherin、波形蛋白和纤维连接蛋白表达降低，提示EMT逆转。我们还观察到E-钙粘素在细胞内的移位。转录因子Snail和Slug在si-panc-1细胞中的表达显著降低，抑制EGFR的表达可显著抑制胰腺癌Panc-1细胞的EMT。其作用机制可能与下调转录因子Snail和Slug的表达有关。
Aberrant expression of epidermal growth factor receptor (EGFR) has been detected in pancreatic cancer; however, the mechanisms of EGFR in inducing pancreatic cancer development have not been adequately elucidated. The objective of this study was to determine the role of EGFR in mediating epithelial-mesenchymal transition (EMT) in pancreatic cancer cells.Pancreatic cancer cell line PANC-1 was transfected with small interfering RNA of EGFR by use of a lentiviral expression vector to establish an EGFR-knockdown cell line (si-PANC-1). PANC-1 cells transfected with lentiviral vector expressing negative control sequence were used as negative control (NC-PANC-1). Scratch assay and transwell study were used to analyze cell migration and invasion. Real-time PCR and Western blotting were used to detect the expression of EMT markers E-cadherin, N-cadherin, vimentin, and fibronectin and transcription factors snail, slug, twist1, and sip1 in PANC-1, NC-PANC-1, and si-PANC-1 cells. Immunofluorescent staining with these antibodies and confocal microscopy were used to observe their cellular location and morphologic changes.After RNA interference of EGFR, the migration and invasion ability of si-PANC-1 cells decreased significantly. The expression of epithelial phenotype marker E-cadherin increased and the expression of mesenchymal phenotype markers N-cadherin, vimentin, and fibronectin decreased, indicating reversion of EMT. We also observed intracellular translocation of E-cadherin. Expression of transcription factors snail and slug in si-PANC-1 cells decreased significantly.Suppression of EGFR expression can significantly inhibit EMT of pancreatic cancer PANC-1 cells. The mechanism may be related with the down-regulation of the expression of transcription factors snail and slug.