Effects Of Extracellular Matrix On The Stemness And Epithelial Mesenchymal Transition Of Pancreatic Cancer Cells

ObjectiveThis study aims to investigate the effects of extracellular matrix(ECM)on the stemness,proliferation,migration,invasion and other biological behaviors of human pancreatic cancer cells,and to provide a theoretical basis for the development of targeted pancreatic cancer therapy based on various proteins in the Extracellular matrix.Methods1.Purchase Collagen I(COL I),Fibronectin(FN)and Laminin(LM)in ECM,and the three ECM-related proteins were used to construct a 3D culture microenvironment composed of ECM.2.Pa Tu8988 and PANC-1 cells in pancreatic cancer cell lines were continuously cultured in a 3D culture microenvironment,and the effects of different ECMs on the stemness of pancreatic cancer cells were detected by flow cytometry and stem cell spheroidization assay.3.Clone formation assay,scratch assay,and Transwell migration and invasion assay were used to detect the effects of different ECMs on the proliferation,migration and invasion of pancreatic cancer cells.4.Western blot was used to verify the changes of different ECMs on cell stemness and epithelial mesenchymal transition(EMT)related proteins in pancreatic cancer cells at the protein level.Results1.A 3D culture microenvironment composed of different ECM-related proteins was successfully constructed,and Pa Tu8988 and PANC-1 cells in the pancreatic cancer cell lines can survive and grow continuously in this 3D culture microenvironment.2.Compared with the ordinary 2D culture microenvironment,the Pa Tu8988 and PANC-1cells continuously cultured in the 3D culture microenvironment,flow cytometry proved that the expression of cell stemness-related molecular markers CD133 and CD44 increased,and the stem cell spheroidization assay was the same It proved that the stemness of cells in the 3D culture microenvironment group was enhanced;and among the three different ECM-related proteins,COL I had the most obvious effect on the stemness enhancement of pancreatic cancer cells,followed by LM,and FN had the weakest effect.3.The Pa Tu8988 and PANC-1 cells in the 3D culture microenvironment group showed stronger proliferation,clone formation,migration and invasion capabilities than the cells in the 2D culture microenvironment group;and also showed the most enhanced effect of COL I.LM is the next,and FN has the weakest effect.4.After continuous culturing in a 3D culture microenvironment,Pa Tu8988 and PANC-1cell EMT-related indicators E-cadherin protein expression levels decreased,N-cadherin,Vimentin,Slug,Snail protein expression levels increased,and the expression levels of cell stemness-related indicators CD133,CD44,Nanog,SOX2,OCT4 were increased;for different ECM-related proteins,the protein expression of the COL I group also changed most significantly,followed by the LM group,and the FN group the weakest.ConclusionsThe extracellular matrix enhances the cell stemness and epithelial-mesenchymal transition of pancreatic cancer cells,and has the effect of promoting the proliferation,migration and invasion of pancreatic cancer cells.At the same time,different extracellular matrix-related proteins have different biological effects on pancreatic cancer cells.