To Study The Effect Of Gupi Xiaoji Decoction On Pyroptosis Of Human Liver Cancer HepG2.2.15 Cells Based On Caspase-1/GSDMD Pathway

Objective:This study is based on the GuPi Xiao Ji Decoction,to clarify the role of the prescription on pyroptosis of hepatoma cells,to reveal the mechanism and key molecular targets of the prescription based on the Caspase-1/GSDMD pathway,and to provide a new thingking and scientific basis for exploring the effective prescription of regulating and delaying the progress of liver cancer to realize"tumor-bearing survival"Methods:①Stably cultured HepG2.2.15 cells and observed the effect of plasma plasma on cell proliferation through CCK-8 test.②Cells were divided into blank plasma group,VX-765 group(Caspase-1 inhibitor),VX-7 65 with drug-containing plasma group and drug-containing plasma group,Determine drug plasma intervention concentrations according to the results of CCK-8 test,and then conducting drug intervention in each group.③Detected the expression and distribution of cell membrane surface GSDMD-N by immunofluorescence staining.④Collected cell serum and the LDH kit detected LDH release after HepG2.2.15 cell lysis.The ⑤ELISA method detected IL-18,IL-1β release in the cell upper serum.⑥Collected cells and Western blot detected the expression level of characteristic protein Caspase-1,GSDMD-N in cells.Results:①The CCK-8 trial found that at the same activity time the cells decreased dose-dependent with increased plasma concentration,and the time decreased at the same intervention concentration.②After CCK-8 test,the plasma concentration was determinded as 15%.③ImmunoFluorescence staining detection found that the main expression of GSDMD-N(green fluorescence)was mainly distributed on the cell membrane,while the green fluorescence expression in the VX-765 group decreased significantly in the drug plasma group,and the VX-765 with plasma expression increased than that in the VX-765 group.④ LDH kit testing found a significant decrease in LDH release in VX-765 group(P<0.01),and a significant increase in VX-765 with drug plasma and drug plasma group(P<0.01).⑤ ELISA found that compared with blank plasma,the release of IL-18 and IL-1β were reduced in VX-765 group(P<0.01),while the the release of IL-18 and IL-1β in VX-765 with drug-containing plasma and drug-containing plasma were increased(P<0.0 1).⑥Western blot test found that the expression of protein Caspase-1,GSDMD-N in HepG2.2.15 cells were significantly reduced(P<0.01),and Caspase-1,GSDMD-N protein expression in HepG2.2.15 cells after plasma treatment(P<0.01),the expression of protein Caspase-1,GSDMD-N in HepG2.2.15 cells were increase after drug plasma treatment(P<0.01).Conclusion:(1)GuPi XiaoJi Decoction can induce Pyroptosis of human hepatoma HepG2.2.15 cells;(2)GuPi XiaoJi Decoction activates the pyrolytic classical pathway by upregulating the expression of related molecule Caspase-1 in the pyrolytic classical pathway Caspase-1/GSDMD,and then induces the pyrolytic HepG2.2.15 cells of human liver cancer.