An Experiment Study Of The Effects On The Chemosensitivity And Invasive Of Panc-1after XIAP And Survivin Expression Being Inhibited By RNAi

Part1The expression of XIAPand Survivin in pancreatic cancer cell lines Panc-1and its correlation with chemoresistanceObjective:To explore the expression of XIAP and Survivin in pancreatic cancer cell lines Panc-1, and its correlation with Gem chemoresistance of cells.Methods:Human pancreatic cancer cell lines, Panc-1, Sw1990, and Miapaca-2were cultured in vitro, and the expression of XIAP and Survivin was detected by real-time quantitative RT-PCR and Western Blot, the chemosensitivity of cell lines was assessed by MTT. Panc-1Gem resistant cell line was induced by intermittent cultivation method. The expression of XIAP and Survivin was detected by real-time quantitative RT-PCR and Western Blot, and the chemosensitivity of cell lines was detected by MTT, a correlation analysis was conducted.Results:(1) XIAP and Survivin were simultaneously expressed in3pancreatic carcinoma cell lines (Panc-1, Sw1990, and Miapaca-2), and we found an inverse relationship between expression of XIAP and Survivin (mRNA and protein) and chemosensitivity. PANC-1cells, which had the highest XIAP and Survivin mRNA levels, was most resistant to chemotherapy; Sw1990cells, which had moderate XIAP and Survivin mRNA levels, had a modest sensitivity to chemotherapy; MIAPaCa-2cells, which showed the least XIAP and Survivin mRNA expression, showed the most sensitivity to chemotherapy.(2) The chemosensitivity of induced Panc-1Gem resistant cell line increased significantly (IC50value), while the expression of XIAP and Survivin also increased significantly, demonstrating a positive correlation between them.Conclusion:There is a correlation between the expression level of XIAP and Survivin, they are constitutive and inducible chemotherapy resistance factor of pancreatic cancer cells panc-1at the same time.Part2The effects on cell proliferation, apoptosis, and chemosensitivity afer inhibition of XIAP and Survivin being inhibited by lentivirus-mediated RNAi in Panc-1cellsObjective:To explore the effects on cell proliferation, apoptosis, and chemosensitivity afer inhibition of XIAP and Survivin by lentivirus-mediated RNAi in Panc-1cellsMethods:Separate or simultaneous use of XIAP-shRNA and Survivin-shRNA lentiviral infection of pancreatic cancer cell lines Panc-1, was conducted to establish cell lines in which XIAP and/or Survivin was inhibited stably.The expression of XIAP and Survivin was detected respectively by real-time quantitative RT-PCR and Western Blot; cell proliferation was detected by cell counting and colony formation assay. The chemosensitivity of cell lines was detected by MTT.Results:(1) Three pancreatic cancer cell line, Panc-1-X, Panc-1-S, Panc-1-XS were successfully cultured in which XIAP and/or Survivin expression are stably suppressed respectively was successfully set up.(2) Cell proliferation was significantly suppressed and Gem chemosensitivity of cells was significantly increased after XIAP and Survivin were inhibited simultaneously.(3) Gem chemosensitivity of Panc-XS was significantly higher than that of Panc-1-X and Panc-1-S respectively.Conclusion:Simultaneously inhibiting the expression of XIAP and Survivin in Panc-1cells could suppress cell proliferation, and enhance the Gem chemosensitivity significantly. The effects of simultaneous inhibition strategy on enhancement of Gem chemosensitivity is better than single inhibition strategy by which the expression of XIAP and Survivin were inhibited respectively. Part3The study of the mechanism and effect on EMT、invasion and migration of pancreatic cancer cell lines Panc-2after XIAP expression being simultaneously inhibitedObjective:To explore the the mechanism and effect on EMT、 invisive and migration of pancreatic cancer cell lines Panc-2after XIAP expression being simultaneously inhibitedMethods:The expression of XIAP and Survivin were inhibited simultaneously in cultured Panc-1-XS cells, and the changes in protein expression of E-Cadherin、Slug、PTEN and P-Akt was detected by Western Blot. Cell invasion and migration ability was detected by wound healing assay and Transwell chambers assay.Results:(1) E-cadherin protein expression was significantly elevated and the expression of Slug protein was significantly reduced in Panc-1cells after XIAP and Survivin were inhibited simultaneously.(2) PTEN protein expression was significantly elevated and the expression of P-Akt protein was significantly reduced in Panc-1cells after XIAP and Survivin were inhibited simultaneously.(3) The cell invasion and migration ability was significantly reduced after XIAP and Survivin were inhibited simultaneously. Conclusion:Simultaneously inhibiting the expression of XIAP and Survivin in Panc-1cells possibly through PTEN/PI3K/Akt pathway partially reversed EMT phenotype of Panc-1cells, and reduced its invasiveness and migration ability.