Study On The Mechanism Of Action Of Jianpi Xiaoai Recipe In Inducing Pyroptosis Of Human Colon Cancer HCT116 Cell

Objective:To research the promotion of Jianpi Xiaoai Prescription on pyroptosis of colorectal cancer cell HCT116;To discuss its possible mechanism of action on pyroptosis.Methods:1.By conducting CCK-8 experiments,the proliferation of colorectal cancer cell(HCT116)were detected;HCT116 cells are treated with10%,15% and 20% blank rat serum and 10%,15% and 20%drug-containing serum,and cell proliferation and CCK-8 experiments are conducted to observe the inhibitory effect of Jianpi Xiaoai Prescription elimination side drug-containing serum on HCT116cells;2.Cell morphology was observed under TEM;3.The LDH release test detects the LDH content of the cell supernatant;4.ELISA experiments detects the release of IL-1 β and IL-18 in the cell supernatant;5.Expression of GSDMD,GSDME,Caspase-3,Cleaved Caspase-3,GSDME-N are detected by Western blot.Results:1.Jianpi Xiaoai Prescription showed in vitro inhibitory effect on the proliferation of HCT116;2.Compared with the blank mouse serum group and the normal serum group,significant pyroptosis in drug-containing groups under TEM was observed;3.Compared with blank group,the release of of LDH in drug-containing groups are increased(P<0.05);4.Compared with the blank mouse serum group and the normal serum group,IL-1 β and IL-18 increase in the cell supernatant after serum intervention in HCT116 cells(P<0.05);5.The protein expression levels of GSDME in 10%,15% and 20%drug-containing groups are decreased(P<0.05),as well as the expression level of GSDMD in 10% and 20% drug-containing groups(P<0.05).GSDME protein is downregulated in 10%,15% and 20%serum group(P<0.05),with 20% concentration more obvious,GSDMD protein in 10% and 20% serum group downregulated(P<0.05).T he protein expression of full-length Caspase-3 in 10%,15% and 20% serum-containing group was regulated(P<0.05).The expression of Cleaved Caspase-3 is detected in 20% serum group(P<0.05).And the N-terminal protein fragment of the cleaved GSDME is detected in the 15%,20% concentrations group(P<0.05).Conclusion:Jianpi Xiaoai Prescription can cause the pyroptosis of HCT116,whose mechanism is related to the splicing of GSDMD and GSDME proteins,and mainly through Caspase-3 / GSDME signal channel.