Research In Meachanism Of Effects Of Corilagin On Migratory And Invasive Abilities Of Human Liver Cancer Cells

Objective:To explore the effect of corilagin on the migration and invasion of human liver cancer cells lineSMMC-7721,MHCC97H cell.in vitro and analyze possible mechanism.(1)The conventional culture of SMMC-7721 and MHCC97H cells in general.Inhibition of different concentrations of corilagin.were treated with 24h.the proliferation of cells was determined by MTT method.(2)The wound healing method were used to evaluate the effect of different concentrations of corilagin on the migrations of SMMC-7721 and MHCC97H cells by detecting the width of wounds.(3)The transwell chamber assay were used to evaluate the effect of corilagin on the migrations of SMMC-7721 and MHCC97H cells by detected amounts of cell migrating.(4)The transwell chamber assay were used to evaluate the effect of corilagin on the invasions of SMMC-7721 and MHCC97H cells by detected amounts of cell invasioning.5)The immunofluorescence assay was applied to observe the effects of corilagin on the actin cytoskeleton rearrangement and the formation of filopodia and lamellipodia in SMMC-7721 and MHCC97H cells.(6)The western blot assays were performed to explore the effects of EMT related proteins(E-cadherin、β-catenin、N-cadherin、Vimentin)and the MAPK pathway related proteins(ERK、p-ERK、JNK、p-JNK、P38、p-P38).Results:(1)MTT assay demonstrated that the proliferation of SMMC-7721 and MHCC97H cells were inhibited by different concentrations of corilagin.The sensitivity of two cells were different,and the MHCC97H cells were more sensitive to the concentration of corilagin.TheSMMC-7721cellsweresignificantlyinhibitedat300μmol/L,andtheMHCC97H cells was signif icantly inhibited at 200 μmol/L.(2)Results of cell wound-healing assays showed that,corilagin(50、100、200 μmol/L)inhibited the migration of SMMC-7721 cells with 0、24h、48h,corilagin(30、60、120 μmol/L)inhibited the migration of MHCC97H cells with 0、24h、48h,and corilagin inhibited SMMC-7721 and MHCC97H cells migration with a concentration and time dependence.(3)Transwell cell migration showed that corilagin inhibited the migration of SMMC-7721 and MHCC97H cells with a concentration dependence.The more higher concentration,the more significant inhibition of migration.Transwell cell migration assays showed that,corilagin inhibited the migration of SMMC-7721 at the concentration 50、100、200 μmol/L,and inhibition ratios of transwell cell migration assays showed were 45.20%、52.03%、62.09%,corilagin inhibited the the abilities of migration of MHCC97H at the concentration of 30、60、120μmol/L.(p<0.05);corilagin inhibited the migration of MHCC97H cells at the concentration 30、60、120 μmol/L and inhibition ratios of transwell cell migration assays showed were 13.62%、51.38%、64.77%;corilagin inhibited the abilities of migration of MHCC97H cells at the concentration of 60、120 μmol/L(p<0.05),but there were no significant effect of corilagin on cell migration at the concentration of 30 μmol/L(p>0.05)(4)Transwell cell invasion assays showed that,corilagin inhibited the invasion of SMMC-7721 at the concentration 50、100、200 μmol/L,and inhibition ratios of transwell cell invasion assays showed were 7.82%、25.23%、60.09%,corilagin inhibited the abilities of migration at the concentration of 200 μmol/L(p<0.05),but there were no significant effect of corilagin on cell migration at the concentration of 50、100μmol/L(p>0.05).transwell cell invasion assays showed that,corilagin inhibited the invasion of MHCC97H at the concentration 30、60、120μmol/L,and inhibition ratios of transwell cell invasion assays showed were 25.20%、38.96%、71.19%,corilagin inhibited the abilities of migration at the concentration of 60、120μmol/L(p<0.05),but there were no significant effect of corilagin on cell migration at the concentration of 30μmol/L(p>0.05)(5)Using immunofluorescence staining,corilagin can make SMCC-7771 and MHCC97H stayed in F-actin skeleton remodeling,the higher the concentration,the cell edge filopodia and lamellipodia formation gradually decreased,the fiber tension number increases gradually,this effect in SMMC-7721 200 μmol/L significantly,while the MHCC97H 120μmol/L is obvious.(6)The Western blot showed that corilagin can significantly up-regulate the expression of N-cadherin and Vimentin in EMT,but there were no significant effect to E-cadherin and β-catenin.the MAPK pathway by up-regulate the expression of p-ERK expression and down-regulate the expression of p-p38,the expression of the change of p-P38,p-JNK,JNK,ERK,P38 expression was not statistically significant.Conclusion:(1)Corilagin can inhibit the abilities of migration and invasion SMMC-7721 and MHCC97H cells.(2)Corilagin inhibits the migration and invasion of SMMC-7721 and MHCC97H,and its mechanism may be through inhibition of EMT in N-cadherin,Vimentin protein expression affect cytoskeleton remodeling.(3)Corilagin inhibited the migration and invasion of SMMC-7721 and MHCC97H,and its mechanism might be through the regulation of ERK pathway-mediated remodeling of cytoskeleton.