The Research Of Suppressor Gene Tg737 In Pathogenesis Of Hepatocellular Carcinoma

Hepatocellular carcinoma (HCC) is one of the most common malignanttumors in China and its mortality rate ranks the third place after stomach cancerand esophagus cancer. There are over 11 million people who died from HCCevery year, which accounts for 45% of deaths in the world. Studies have shownthat the prognosis of primary liver cancer is still not optimistic, mainly due tothe lack of specific indicators of early diagnosis and treatment. The research ofpathogenesis in HCC is helpful to promote the rate of diagnosis in the earlyperiod.Tg737 gene is a kind of tumor suppressor gene in HCC. The initial studyfound that the gene was the same as the target gene of genetic polycystic kidneydisease (autosomal recessive polycystic kidney disease, ARPKD) orpk mutantmouse model. But so far, the mechanism of Tg737 gene in HCC is not very clear.Therefore, it is very important to reveal the biological function of Tg737 geneand develope the theory of HCC for diagnosis and treatment of HCC in the earlyperiod. This study is divided into three parts.Partâ… Study of expression of Tg737 and correlation in hepatocellularcarcinomaObjective Expression of Tg737 in HCC was analyzed and relationshipbetween the expression of Tg737 and mechanism of HCC was investigated.Materials and methods RT-PCR and Western blot methods were used toanalyze the expression of Tg737 mRNA and protein.Results The results showed that Tg737 expression of the mRNA andprotein in liver cells were higher than HepG2 and MHCC97 cells. Thirty casesof Tg737 mRNA expression in 65 cases of HCC tissues were lower than thecorresponding adjacent tissues, while 61 cases of Tg737 mRNA expression inthe adjacent tissues were positive. Fifty-nine (90.78ï¼…) cases of Tg737 proteinexpression in 65 cases of the adjacent tissues were positive. There were 34(52.31ï¼…) cases of protein positive expression in HCC tissues. Fifteen caseswere positive in 18 (55.56ï¼…) cases of early (â… -â…¡) HCC tissues. There were 19(27.66ï¼…) cases of positive expression in 47 cases of advanced (â…¢-â…£) HCCtissues. Statistical analysis showed that the Tg737 protein in tumor tissue wassignificantly lower than the adjacent liver tissue. The Tg737 protein positive rateof early (â… -â…¡) HCC tissue was significantly higher than in advanced (â…¢-â…£)HCC tissues.Conclusions It is indicated that Tg737 gene expression in HCC isreduced, which may be important factor in the process of hepatocarcinogenesis.Analysis of Tg737 gene expression may be reference index of HCC stages.Partâ…¡Tg737 affecting activity of HCC cellsObjective Tg737 gene was amplificated from human normal liver tissueand constructed to the eukaryotic expression vector. Then it is studied that Tg737 influences activity of HCC cells.MaterialMaterials and methods Plasmid pEGFP-Tg737 was transfected intoHepG2 cell and increased expression level of Tg737. The experiment wasdivided into three groups: HepG2-Tg737 group (pEGFP-Tg737 transfectiongroup), HepG2-vect group (empty vector transfection group), and HepG2 group(blank control group). Cell cycles were analyzed by flow cytometry andexpression of Tg737 was detected by Real-time PCR and Western blot methods.Cell growth curves were made by MTT test.Results The eukaryotic expression vector pEGFP-Tg737 was constructedsuccessfully and transfected into hepatocellular carcinoma line. Compared withHepG2-vect group and HepG2 group, Tg737 expression of HepG2-Tg737 groupincreased remarkedly after 48 hours of transfection. Cell growth in HepG2-Tg737 group was significantly inhibited and the proportion of G1 phase cells inHepG2-Tg737 group was higher than that of HepG2-vect group and HepG2group and cells of S phase was significantly lower.Conclusions Tg737 gene can inhibit cell proliferation markedly andinduce block of G1 stage.Partâ…¢Study of loss of heterozygosity on Tg737 geneObjective Loss of heterozygosity (LOH) of Tg737 in HCC was detectedand relationship between LOH status and clinical pathology characters wasanalyzed.Materials and methods Polymerase chain reaction-single strandconformation polymorphism (PCR-SSCP) method was used to detect LOH offive microsatellite sites on Tg737.Results The results showed that LOH frequences of five microsatellitesites in Tg737 gene were 61.54%(32/65), 52.31%(34/65), 65.45%(36/65), 58.33%(28/65) and 74.47%(35/65). There was no significant difference (P>0.05)between LOH of five microsatellite sites. However, LOH frequences of SHGC-57879 and G64212 related to tumor metastasis and TNM staging (P<0.05).Conclusions High frequences LOH of Tg737 gene exist in HCC andmicrosatellite sites had high instability, which can affect on expression of thegene.In conclusion, this study confirms that tumor suppressor gene Tg737 playsan important role in the period of hepatocarcinogenesis and inhibits theproliferation of HCC cells. Nevertheless, loss of heterozygosity of the Tg737gene exists in hepatocellular carcinoma and high instability is shown on itsmicrosatellite sites which affect the gene expression. So Tg737 can serve astargets for gene therapy of HCC and provide new ideas for gene therapy of HCC.