Effect Of Jianpi Yiqi Decoction On Inflammatory Body Of Hepatocellular Carcinoma Cells NLRP3 By Downregulating Vimentin Expression

Objective: Jianpi Yiqi Prescription is obtained through screening and induction of Clinical Prescriptions under the guidance of the theory of Jianpi Therapy of Traditional Chinese Medicine.By studying the effects of Jianpi Yiqi Decoction on proliferation,invasion,ability and apoptosis of hepatocellular carcinoma cells,and detecting the expression of various components of inflammatory body NLRP3 protein,the effect of Jianpi Yiqi Decoction on NLRP3 inflammatory body by down-regulating the expression of Vimentin(vimentin)in hepatocellular carcinoma cells was explored,so as to further understand the mechanism of Jianpi therapy.Methods:Hep G2 cells were divided into seven groups: blank group,model group,spleen-invigorating L-dose group,spleen-invigorating M-dose group,spleen-invigorating H-dose group,Casp1 inhibitory group and Casp3 inhibitory group.The blank group was cultured with rat serum,and the other groups were induced into inflammatory model cells by LPS + ATP.Then,three doses of Jianpi Yiqi recipe containing low,medium and high(L,M,H)serum,casp1 inhibitor and CASP3 inhibitor were used to intervene each cell.CCK8 test,Transwell invasion test,flow cytometry were used to detect the proliferation,invasion and apoptosis of cells in each group.Caspase-1 activity was detected by Caspase-1 activity kit,IL-1βsecreted by culture supernatant was detected by ELISA,and the expression of Vimentin,ASC,NLRP3 and Caspase-1 protein was detected by Western blot.Result: CCK8 method: LPS + ATP combined stimulation of inflammatory model cells after different concentrations of Jianpi Yiqi recipe containing serum intervention,the proliferation of cells was inhibited(P<0.01 or P<0.05),of which the highest inhibition rate was found in the high dose;the highest inhibition rate was found in the caspase-1 inhibition group(P<0.01),but not in the caspase-3 inhibition group.It has a certain promoting effect.2.Transwell invasion experiment: Compared with the blank serum group,the invasive ability of hepatocellular carcinoma cells in LPS + ATP group was enhanced(P<0.01).After the intervention of different concentration of serum containing Jianpi Yiqi Decoction,the invasive ability decreased(P<0.01),and the effect of the high dose group was the best;the inhibition rate of Caspase-1group was the highest in all experimental groups,while that of Caspase-3group increased its invasive ability.Power.3.Annexin V + PI double staining method: Compared with the blank serum group,the apoptotic rate of LPS +ATP model group decreased significantly;after the intervention model of Jianpi Yiqi recipe containing serum,the apoptotic rate of LPS + ATP inflammation model increased significantly(P<0.01);the apoptotic rate of Caspase-1 inhibitor group was still the highest,but there was no significant difference between the caspase-3 inhibitor group and the LPS + ATP model group(P>0.05).4.ELISA: Compared with the blank serum group,the content of caspase-1 and IL-1beta secreted by LPS+ATP model group increased significantly(P<0.01).After intervention with drug-containing serum,the content of caspase-1 and the content of IL-1beta secreted by caspase-1 in hepatocellular carcinoma cells of model group decreased significantly(P<0.01),especially in the high dose group;the caspase-1 inhibition group decreased in all experimental groups.There was no significant difference between caspase-3 inhibition group and model group(P>0.05).5.Western blot results showed that compared with the blank serum group,the expression of Vimentin,ASC,NLRP3 and caspase-1 protein in LPS+ATP model group increased(P<0.01 or P<0.05).After the intervention of serum containing Jianpi Yiqi Decoction,the expression of all the target proteins decreased(P<0.01 or P<0.05).Caspase-1 was still the best in Jianpi high dose group.The expression of ASC,NLRP3 and caspase-1 in the inflammatory model was significantly decreased in the preparation group(P<0.01),which was the lowest in all experimental groups,but the inhibitory effect on Vimentin was not obvious,even higher than that in the blank serum group.There was no significant difference between caspase-3 inhibition group and model group.Conclusion: 1.After the intervention of the serum containing Jianpi Yiqi Recipe in each group,the effect of the high dose group was the best.It may inhibit the proliferation and invasion of Hep G2 cells by down-regulating the activation of NLRP3 inflammatory body,reducing the inflammatory reaction and improving the inflammatory microenvironment,and induce apoptosis of some hepatocellular carcinoma cells.2.The decrease of NLRP3 inflammatory body may be contained by Jianpi Yiqi Recipe.Drug serum plays an important role in inhibiting ASC,NLRP3,caspase-1 and IL-1beta.3.The serum containing Jianpi Yiqi Decoction may inhibit the NLRP3/caspase-1inflammatory signaling pathway by increasing the expression of Caspase-3protein,and then by degrading Vimentin,regulate the inflammatory microenvironment of hepatocellular carcinoma and inhibit the occurrence and development of hepatocellular carcinoma.