Biological Effects Of Down-regulating DACH1 Expression With ShRNA On The Human Pancreatic Cancer Capan-1 Cells

Objective: To investigate the expression of the cell fate determination factor 1(DACH1) in pancreatic cancer cell lines, and construct and screen out the recombinant plasmid pshRNA-DACH1 which was evaluated the best control effect in inhibition of DACH1 expression, then to explore its effect on proliferation, cell cycle, cell apoptosis, invasion and migration of human Capan-1 cells in vitro, which may lay the foundation for gene diagnosis and molecular targeted therapy of pancreatic cancer.Methods: DACH1 expression in human pancreatic cancer cell lines(PANC-1、BxPC-3、AsPC-1、Capan-1) and immortalized pancreatic ductal epithelial cell(HPDE6c7) was analyzed by semi reverse transcription PCR(RT-PCR) and western blot. The recombinant plasmids target DACH1 were designed, constructed and transfected into Capan-1 cell via Lipofectamine2000, the transfection efficiency were observed by fluorescence microscopy, RT-PCR and western blot were then used to screen out the highest inhibitory efficiency plasmid. The inhibition of cell proliferation was tested by colony formation and cck8 assays. Cell apoptosis and cell cycle were tested by flow cytometry combined with Annexin V-PE/7ADD staining. TranswellTM assay was used to monitor the invasion and migration abilities of Capan-1 cells.Results: DACH1 expression is obviously overexpressed in Capan-1 pancreatic cancer cell compared to the normal epithelial cell. Recombinant plasmid pshRNA-DACH1 was successfully constructed and transfected into Capan-1 cells, and choose the most inhibitory efficiency plasmid. Colony formation and cck8 assays showed the proliferation of Capan-1 cells was significantly inhibited. FCM revealed that cell apoptosis was promoted in interfering plasmid group compared with control groups, whereas cell cycle has no significant difference among the groups. Transwell chamber experiments validated the abilities of migration and invasion reduced in treatment groups.Conclusion: The expression of DACH1 is overexpressed in Capan-1 pancreatic cancer cell, knockdown of DACH1 via shRNA-mediated transfection can significantly enhance the cell apoptosis, restrain the proliferation, migration and invasion of Capan-1 cells, which indicates that DACH1 may has a close relationship with biological function of pancreatic cancer Capan-1 cells.