Study On The Effect Of MMP16 On Invasion Mechanism And Prognosis Of Hepatocellularcarcinoma

Part ? Study on the prognostic value of MMP16 in TCGA database and validation cohortObjective:To investigate the effect of MMP16 on the survival of patients with liver cancer in the TCGA database and validation cohort,and to determine whether the expression of MMP16 in liver cancer patients has a prognostic effect.Methods:(1)Select 330 patients with liver cancer in the TCGA database,and use univariate Cox regression analysis to determine the relationship between MMP16 expression and survival of liver cancer patients;(2)In terms of overall survival,use X-tile program determine the optimal cut-off value of MMP16 expression in 330 liver cancer patients to distinguish between high-expression and low-expression patients,and to verify whether the 5-year overall survival rate of the two groups was statistically different;(3)Follow-up of 330 patients with HCC,chi-square test to investigate whether the percentage of MMP16 high expression in all death patients is statistically different from the percentage of patients with high MMP16 expression in surviving patients;(4)Kaplan-Meier survival curve To investigate the relationship between the expression of MMP16 and the 5-year overall survival and disease-free survival in another 69 patients invalidation cohort.Results:(1)In patients with liver cancer,MMP16 is a predictor of overall survival(HR:1.169,95%CI:1.034-1.321,P=0.013);(2)X-tile program distinguishes MMP16 from high expression and The best threshold for low expression is-2.50.Using this optimal cut-off value,all patients were divided into two groups of patients with high and low MMP16 expression.The 5-year overall survival rate of the former was significantly higher than the latter(x2=9.259,P=0.002);(3)In the 330 liver cancer patients,107 patients eventually died of the disease.The percentage of MMP16 high expression in all death patients was significantly higher than that in the surviving ones,the difference was statistically significant(x2=9.188,P=0.002);69 patients with HCC were divided into MMP16 high expression group and low expression group.Kaplan-Meier survival curve analysis showed that the 5-year overall survival rate of MMP16 high expression group was significantly lower than that of MMP16 low expression group(30.7%vs.59.2%,x2).=5.172,P=0.023),5-year disease-free survival was also significantly lower than in the low-expression group(43.2%vs.22.1%,x2=4.119,P=0.042).Conclusions:In the TCGA database,MMP16 isa predictor of overall survival in patients with liver cancer.In the validation cohort,MMP16 is an independent predictor of survival in univariate and multivariate studies.Part ? The Effect of MMP16 on the invasion ability of hepatoma cell linesObjective:To study the expression of MMP16 in HCC tissues and normal liver tissues,to predict the function of MMP16 in HCC,and to further explore the role of MMP16 in the development of HCC.To investigate whether it has an effect on the proliferation and invasion of HCC tumor cells.Methods:(1)We used RT-PCR and IHC to study the expression of MMP16 in 25 pairs of cancer tissues and adjacent normal tissues on the mRNA level and the protein level;(2)Transfecting MMP16 with lentivirus to HepG2 and HCCLM3 cells were assayed for the effect of MMP16 transfection by RT-PCR and Western-blot;(3)The effect of silencing MMP16 expression on the growth rate of HCC cells was determined by CCK-8;(4)Transwell analysis was used to study the effect of MMP16 on HCC cell invasion ability.Results:The results of RT-PCR and IHC showed that the expression level of MMP16 mRNA in HCC tissues was significantly higher than that in the adjacent normal tissues(P<0.001).IHC studies showed that only a small amount of MMP16 was stained in normal liver tissues,but the mount was more in tumors.These data suggested that MMP16 may be an oncogene in HCC.After transfecting of HCC cells,RT-PCR and Western-blot assays confirmed that the expression of MMP16 in HCC cells after transfection was significantly lower than that in untransfected cells.The CCK-8 assay indicated that silencing MMP16 expression did not affect the growth rate of HCC cells.Transwell analysis showed that the number of transmembrane invasion cells in the MMP16-interfering group was significantly reduced compared with the control group(P<0.05).Overall,the decreased expression of MMP16 attenuated the invasion ability of HCC cells.Conclusion:Interfering with the expression of MMP16 in HCC cells has no significant effect on the proliferation of tumor cells,but significantly attenuates the invasive ability of tumor cells.Part ? Study on the effect of MMP16 on invasion mechanism in hepatoma cell linesObjective:Since epithelial-mesenchymal transition(EMT)is closely related to the ability of cancer cells to metastasize,we examined the EMT markers of MMP16 in interfering cells and their control cell lines to investigate whether MMP16 affects the invasion of HCC cells by interfering with EMT.Method:(1)At the mRNA level,we used RT-PCR to detect the expression levels of E-cadherin,vimentin and N-cadherin in HepG2 and HCCLM3 HCC cells before and after interference with MMP16;(2)At the protein expression level,we used Western-blot to detect the expression levels of E-cadherin,vimentin and N-cadherin in HepG2 and HCCLM3 HCC cells before and after interference with MMP16.Results:Simultaneously at the mRNA and protein levels,silencing of MMP16 expression resulted in increased expression of the epithelial-specific protein,E-cadherin,and decreased expression of the mesenchymal markers,vimentin and N-cadherin.Conclusion:Ectopic MMP16 expression promotes the invasion of HCC cells by inducing EMT.