The Study Of The Mechanism Of TNFα And Gemcitabine Inducing Pyroptosis In Pancreatic Cancer Cell

Objective:To investigate the effects and related mechanisms of cytokine TNFαand chemotherapy drug gemcitabine on pancreatic cancer cell.Methods:The pancreatic cancer Panc-1 was selected,treated with different concentration gradients of gemcitabine.The cell activity was detected by the CCK-8 method.The appropriate concentrationwas selected to act on the cells,and three time periods were selected:12h,24h and 48h,the morphology of the cells was observed by microscopic imaging.Cells were treated with TNFα and gemcitabine separately,and the release of IL-1β and IL-18 in the supernatant was detected by Elise method WB and qPCR methods were used to detect the relative expression levels of proteins and mRNAs of GSDMD,GSDME,Caspase-1/3/4/5 in each time period.Results:1.TNFα and gemcitabine in Panc-1 can be observed under the microscope under the characteristics of cell scorching characteristics of Pyroptosis.2.The release of IL-1β and IL-18 in the supernatant of TNFα group and gemcitabine group was significantly higher than that in NC group(P<0.05),and the difference was statistically significant.3.The results of qPCR showed that TNFα stimulated Panc-1,the gene expression of GSDME and Caspase-3 was not significantly different from that of NC group(P>0.05),while the gene expression of GSDMD and Caspase-4 Compared with the NC group was a statistical difference(P<0.05).As time went by,the Caspase-1 gene had a statistical difference compared with the NC group(P<0.05).Gemcitabine stimulated Panc-1,the gene expression of Caspase-3 and GSDME was statistically different from that of NC group(P<0.05).With the progress of time,the gene expressions of Caspase-1/4/5 and GSDMD were statistically different from those of NC group(P<0.05).4.WB results showed that the protein expressions of Caspase-3 and GSDME after TNFα and gemcitabine stimulated Panc-1 cells were statistically different from those of NC group(P<0.05)And in the TNFα group,the expression levels of Caspase-1 and Caspase-5 proteins were statistically different from those in the NC group(P<0.05);In the gemcitabine group,the expression levels of Caspase-1 and Caspase-4 proteins were statistically different from those in the NC group(P<0.05).Conclusion:TNFα and gemcitabine can induce pancreatic cancer pyroptosis through the Cas-3/GSDME pathway rather than the traditional GSDMD pathway,and released a large number of inflammatory factors IL-1β and IL-18.