Mechanism And Significance Of Down-regulation Of Vimentin Expression In Hepatocellular Carcinoma Cells By Jianpi Yiqi Decoction

Objective:This project was focused on the mechanism of down-regulation of Vimentin expression in hepatocellular carcinoma cells by Jianpi Yiqi decoction,and the effects on proliferation,invasion,Caspase-3 apoptotic pathway and NLRP3/Caspase-1 pathway of hepatocellular carcinoma cells.Methods: Medicated serum of Jianpi Yiqi decoction was prepared and used to interfer TGF-β1-induced SMMC-7721 cells with different concentrations(0%,5%,10%,15%,20% and 30%).The proliferation and invasion of human hepatocellular carcinoma SMMC7721 cells were detected by MTT and Transwell invasion assay,respectively.Interfering time and concentration of Jianpi Yiqi decoction were optimized.The effects on biological behavior of TGF-β1-induced SMMC-7721 were examined after treatment by 0%(blank serum group),7.5%,15%,30% concentration medicated serum containing Jianpi Yiqi decoction,and compared with the results by Vimentin breaker(10μmol/L WFA)or by Caspase inhibitor(10μmol/L Z-VAD-FMK).Furthermore,the expression of Vimentin,Caspase-3,PARP-1,NLRP3 and Caspase-1 were detected by Western blot.Results:The TGF-β1-induced SMMC-7721 cells after treatment by 15%concentration medicated serum containing Jianpi Yiqi decoction for 48 hours,showed the greatest inhibition effects on the proliferation and invasion of the cells.Protein expression of Vimentin,PARP-1,NLRP3 and Caspase-1 in the treated cells were decreased as compared with the blankserum group(P<0.01),and protein expression of Caspase-3 increased.The decreased degree of Vimentin and PARP-1 were consistent with the proliferation and invasion inhibition rate and apoptosis rate of SMMC-7721 cells.The proliferation and invasion inhibition rate and apoptosis rate of the Vimentin-breaker group cells were the highest in all experimental groups,but only up-regulation or down-regulation of the proteins expression were more obvious.However,the proliferation and invasion inhibition rate of the Caspase-inhibitor group cells were the lowest in all experimental groups,and the apoptosis rate was only higher than that of the EMT model group,even lower than that of the blank serum group(P<0.05).Compared with the blank serum group cells,down-regulation of Caspase-3,NLRP3 and Caspase-1and up-regulation of PARP-1 protein expression were observed in the Caspase-inhibitor group cells.For the the Caspase-inhibitor group,Vimentin expression difference was not statistically significant(P>0.05),protein cleavages of Vimentin and PARP-1 were not showed.Conclusion:The expression of Vimentin in SMMC-7721 cells is down-regulated by serum containing Jianpi Yiqi decoction,which may be due to the cleavage of Vimentin by Caspase-3.In this process,a positive feedback loop forms,whereby activated caspase-3 degrades Vimentin,and these degraded fragments consequently activate caspases to amplify apoptosis.Furthermore,down-regulation of Vimentin may regulate the development and progression of hepatocellular carcinoma by decreasing the NLRP3/Caspase-1 inflammatory signaling pathway.