Objective:To discuss the abnormal expression of Wnt inhibitory factor(WIFI) in hepatocellular carcinoma cells and its regulating effect on the hepatocellular carcinoma invasion and metastasis factors of tissue inhibit...Objective:To discuss the abnormal expression of Wnt inhibitory factor(WIFI) in hepatocellular carcinoma cells and its regulating effect on the hepatocellular carcinoma invasion and metastasis factors of tissue inhibitor of matrix metalloproteinases-3(TIMP-3)and caveolin-1.Methods:RT-PCR and Western blot were employed to detect the expression of WIF1 in six hepatocellular carcinoma eell lines of HepG2,Hep3 B,Huh7,PLC/PRF/5.SMMC-7721 and MHCC97 and the immortalized human liver cell line THLE-3.Besides,Lipofectamine 2000 was employed to transfect the eukaryotic expression vector pcDNA3.lWIF1 and blank plasmid pcDNA3.1 into hepatocellular carcinoma cell lines.Transwell assay was used to detect the effect of WIF1 on the invasion ability of hepatocellular carcinoma cells;Western blot was used to detect the effect of WIF1 on the expression of TIMP-3 and caveolin-1in hepatocellular carcinoma cells,it also discussed the effect on the expression of β-catenin.Results:The expression of WIF1 in hepatocellular carcinoma cell lines was lower than that in the normal liver cell lines(P<0.01);while there was basically no expression of WIF1 in the human highly metastatic cell line MHCC-97 and moderate expression in HepG2 and SMMC-7721.Therefore,HepG2 and SMMC-7721 were chosen as the further experimental cell lines.After transfecting the eukaryotic expression vector peDNA3.1-WEFl and blank plasmid pcDNA3.1 into hepatocellular carcinoma eell lines,compared with(he blank plasmid group,the cell viability and invasion ability in the WIF1 group were all reduced(P<0.01),the expression of TIMP-3,caveolin-1 and mRNA were all down-regulated(P<0.01),and the expression of β-catenin was decreased(P<0.01).Conclusions:Because of down-regulation or missing of expression of WIFI in hepatocellular carcinoma cell lines,the up-regulation of WIFI expression can significantly inhibit the invasion and metastasis of HepG2 and SMMC-7721 of hepatocellular carcinoma cell lines,which are related to the up-regulated expression of TIMP-3 and down-regulated ex展开更多
AIM:To investigate the promoter methylation status and mRNA expression of DKK-3 and WIF-1 gene in hepatocellular carcinoma(HCC).METHODS:DKK-3 and WIF-1 acted as Wnt-antagonists and tumor suppressors,but hypermethylati...AIM:To investigate the promoter methylation status and mRNA expression of DKK-3 and WIF-1 gene in hepatocellular carcinoma(HCC).METHODS:DKK-3 and WIF-1 acted as Wnt-antagonists and tumor suppressors,but hypermethylation of the gene promoter and low mRNA expression activated Wnt signaling aberrantly and induced the development of HCC.Methylation status of the DKK-3 and WIF-1 gene promoter was investigated using methylation specific polymerase chain reaction(PCR) in tumor and adjacent non-cancerous tissues from 33 HCC patients and 20 normal liver tissues served as control.The expression of DKK-3 and WIF-1 mRNA was also determined by real-time quantitative reverse transcriptase PCR.The relationship between methylation,mRNA expression,and clinical data,as well as methylation and mRNA expression of the two genes were analyzed.RESULTS:The methylation of DKK-3 and WIF-1 genes in HCC increased significantly compared with adjacent non-cancerous tissues and normal control tissues(χ2 =7.79,P < 0.05;χ2 = 4.89,P < 0.05),and no significant difference in methylation between adjacent non-cancerous tissues and normal control tissues was observed.In HCC tissues,significant differences in the DKK-3 promoter methylation were observed in age and cirrhosis,and significant differences of the WIF-1 promoter methylation were observed in HBsAg and cirrhosis.The average expression of DKK-3 mRNA in HCC and adjacent non-cancerous tissues was increased significantly compared with normal control tissues.The average expression of WIF-1 mRNA showed no significant difference among the three tissues.The mRNA expression of DKK-3 gene in HCC was decreased as the pathological grade increased.CONCLUSION:The aberrant promoter methylation and decreased expression of DKK-3 and WIF-1 may be an important mechanism in HCC,and may be a far-reaching significance in early diagnosis and therapy of HCC.展开更多
基金supported by National Science&Technology Pillar Program during the 12th Five-year Plan Period under the research and development of new trisacryl gelatin microspheres(No.2012BAH5B06)
文摘Objective:To discuss the abnormal expression of Wnt inhibitory factor(WIFI) in hepatocellular carcinoma cells and its regulating effect on the hepatocellular carcinoma invasion and metastasis factors of tissue inhibitor of matrix metalloproteinases-3(TIMP-3)and caveolin-1.Methods:RT-PCR and Western blot were employed to detect the expression of WIF1 in six hepatocellular carcinoma eell lines of HepG2,Hep3 B,Huh7,PLC/PRF/5.SMMC-7721 and MHCC97 and the immortalized human liver cell line THLE-3.Besides,Lipofectamine 2000 was employed to transfect the eukaryotic expression vector pcDNA3.lWIF1 and blank plasmid pcDNA3.1 into hepatocellular carcinoma cell lines.Transwell assay was used to detect the effect of WIF1 on the invasion ability of hepatocellular carcinoma cells;Western blot was used to detect the effect of WIF1 on the expression of TIMP-3 and caveolin-1in hepatocellular carcinoma cells,it also discussed the effect on the expression of β-catenin.Results:The expression of WIF1 in hepatocellular carcinoma cell lines was lower than that in the normal liver cell lines(P<0.01);while there was basically no expression of WIF1 in the human highly metastatic cell line MHCC-97 and moderate expression in HepG2 and SMMC-7721.Therefore,HepG2 and SMMC-7721 were chosen as the further experimental cell lines.After transfecting the eukaryotic expression vector peDNA3.1-WEFl and blank plasmid pcDNA3.1 into hepatocellular carcinoma eell lines,compared with(he blank plasmid group,the cell viability and invasion ability in the WIF1 group were all reduced(P<0.01),the expression of TIMP-3,caveolin-1 and mRNA were all down-regulated(P<0.01),and the expression of β-catenin was decreased(P<0.01).Conclusions:Because of down-regulation or missing of expression of WIFI in hepatocellular carcinoma cell lines,the up-regulation of WIFI expression can significantly inhibit the invasion and metastasis of HepG2 and SMMC-7721 of hepatocellular carcinoma cell lines,which are related to the up-regulated expression of TIMP-3 and down-regulated ex
基金Supported by The Natural Science Fund of Educational Department of Anhui Province,No. 2006KJ094A
文摘AIM:To investigate the promoter methylation status and mRNA expression of DKK-3 and WIF-1 gene in hepatocellular carcinoma(HCC).METHODS:DKK-3 and WIF-1 acted as Wnt-antagonists and tumor suppressors,but hypermethylation of the gene promoter and low mRNA expression activated Wnt signaling aberrantly and induced the development of HCC.Methylation status of the DKK-3 and WIF-1 gene promoter was investigated using methylation specific polymerase chain reaction(PCR) in tumor and adjacent non-cancerous tissues from 33 HCC patients and 20 normal liver tissues served as control.The expression of DKK-3 and WIF-1 mRNA was also determined by real-time quantitative reverse transcriptase PCR.The relationship between methylation,mRNA expression,and clinical data,as well as methylation and mRNA expression of the two genes were analyzed.RESULTS:The methylation of DKK-3 and WIF-1 genes in HCC increased significantly compared with adjacent non-cancerous tissues and normal control tissues(χ2 =7.79,P < 0.05;χ2 = 4.89,P < 0.05),and no significant difference in methylation between adjacent non-cancerous tissues and normal control tissues was observed.In HCC tissues,significant differences in the DKK-3 promoter methylation were observed in age and cirrhosis,and significant differences of the WIF-1 promoter methylation were observed in HBsAg and cirrhosis.The average expression of DKK-3 mRNA in HCC and adjacent non-cancerous tissues was increased significantly compared with normal control tissues.The average expression of WIF-1 mRNA showed no significant difference among the three tissues.The mRNA expression of DKK-3 gene in HCC was decreased as the pathological grade increased.CONCLUSION:The aberrant promoter methylation and decreased expression of DKK-3 and WIF-1 may be an important mechanism in HCC,and may be a far-reaching significance in early diagnosis and therapy of HCC.
