The Mechanism Of Notch-1 In The Epithelial-Mesenchymal Transition And Stem Cell Properties Of Pancreatic Cancer Cells

Part one:The mechanism of Notch-1 in the epithelial-mesenchymal transition of pancreatic cancer cells induced by gemcitabineObjectives:To explore the mechanism of Notch-1 in the epithelial-mesenchymal transition of pancreatic cancer cells induced by gemcitabine.Methods:The EMT phenotype of Panc-1 cells was induced by GEM, as well as the expression of EMT markers, Notch-1 and relevant transcription factors were tested by Western Blot and RT-PCR. The immunofluorescence was used to detect expression of E-cadherin and vimentin. Then transwell migration assay and transwell invasion assay were used to detect the EMT cells.Results:5nM to lOnM GEM and 9 days continuous culture were used for maintaining EMT phenotype of Panc-1 cells. For EMT phenotype, the expression of E-cadherin was reduced, but vimentin, Notch-1 and the transcription factors were increased. The migration and invasion were increased.Conclusion:GEM was a critical drug for EMT phenotype of Panc-1 cells. Notch-1 signaling pathway may be mechanistically linked with EMT phenotype. EMT could make the pancreatic cells more migratory and invasive.Part two:The mechanism of side population cells andABC transporter family in the epithelial-mesenchymal transition of pancreatic cancer cells induced by gemcitabineObjectives:To explore the mechanism of side population cells and ABC transporter family in the epithelial-mesenchymal transition of pancreatic cancer cells induced by gemcitabine.Methods:The Panc-1 cells were exposed in GEM, the expression of ABC transporter family were detected by Western Blot and RT-PCR. Fluorescence-activated cell sorting was used to observed the content of side population cells.Results:The expression of ABC transporter family was up-regulated. The content of side population cells was increased.Conclusion:EMT of Panc-1 cells could make the CSC associated makers up-regulated and lead to the increase of the content of side population cells.Part three:The mechanism of Notch-1 siRNA in the mesenchymal-epithelial transition and side population cells, ABC transporter family of pancreatic cancer cellsObjective:To explore the mechanism of Notch-1 siRNA in the mesenchymal-epithelial transition and side population cells, ABC transporter family of pancreatic cancer cells.Methods:GEM resistance cells were transfected with Notch-1 siRNA. The expression of EMT markers, Notch-1, relevant transcription factors and ABC transporter family were tested by Western Blot and RT-PCR. Fluorescence-activated cell sorting was used to observed the content of side population cells. Then transwell migration assay and transwell invasion assay were used to detect the EMT cells.Results:Notch-1 siRNA could make GEM resistance cells come into mesenchymal-epithelial transition. The expression of E-cadherin was increased, and the expression of vimentin, Notch-1, relevant transcription factors and ABC transporter family were decreased. The migration and invasion were decreased, and the content of side population cells were down-regulated too.Conclusion:Notch-1 siRNA could inhibit the Notch-1 signaling pathway, and Notch-1 is an important signaling pathway for EMT of Panc-1 cells. EMT can promote the invasion, metastasis and the expression of CSC features, when Notch-1 signaling pathway was inhibited, the invasion, metastasis and drug resistance features were suppressed.