Clinical Significance And Biological Function Of α1,6 Fucosyltransferase In Hepatocellular Carcinoma

Hepatocellular cellularcarcinoma (HCC) is the fifth most common cancer and the third leading cause of cancer-related death in the world. In China, hepatitis B Virus (HBV) infection is a well-recognized risk factor for chronic liver diseases, such as cirrhosis and liver cancer.Most protein in organism exist in the form of glycoprotein,sugar chains of glycoprotein affect its functions directly.It is known that sugar chains have a variety of functions and play a key role in growth,development,differentiation and adhesion in cells.Structural changes in N-glyeans are one of the critical steps in cellular malignant transformation.It is well known that the structures of complex carbohydrates are altered in cancer and that these changes are lligllly associated with invasion and metastasis.The mechanism by which those changes occur are still unknow,but in most cases the activation of glycosyltransferases plays a major role and this leads to so-called'aberrant glycosylation'in cancer tissues.α1,6-fucosyltransferase (FUT8) catalyzes the reaction of core fucosylation by transferring a fucosyl residue from GDP-fucose to the asparagine-linked GlcNAC residue of complex N-glycans viaα-1,6 linkage. This process is regarded as an important manner of post-translational modification and functional regulation of glycoproteins. In normal tissues, the content of core fucosylated N-linked oligosaccharide is rather low. However, with the tumorigenesis of some tissues such as liver, lung and stomach, its content increases dramatically.Part 1: Expression ofα1,6-fucosyltransferase in hepatocellular carcinoma and its clinical significancePrevious studies have demonstrated that terminal fucosylation is associated with the biological aggressiveness of carcinomas, but the significance of core fucosylation (α1,6-fucosylation) throughα1,6-fucosyltransferase (FUT8) has not been studied in depth. Herein, we investigated the expression and clinical significance of FUT8 in hepatocellular carcinoma.Western blot and Real-time PCR were used to detect the expression of FUT8 in 52 cases of hepatocellular carcinoma, their corresponding adjacent liver tissues and 10 cases of portal vein cancer embolus.The relations with its clinical pathological parameters were analyzed too.Results Western blot and Real-time PCR analysis showed that the expression level of FUT8 in hepatocellular carcinoma was greater than that in corresponding adjacent liver tissues (P<0.05) and portal vein cancer embolus (P<0.05). But it had no significant difference between corresponding adjacent liver tissues and portal vein cancer embolus (P>0.05). FUT8 protein expression was correlated with tumor size and intrahepatic sub-foci (P<0.05). Conclusions FUT8 expression is significantly correlated with the biological behavior of primary hepatocellular carcinoma.The high expression of FUT8 may play an important role in the malignancy transformation, invasion progression and metastasis of primary hepatocellular carcinoma.Part2: Biological Function ofα1,6-fucosyltransferase in hepatocellular carcinomaAfter transfecting FUT8 targeted RNAi expression vector and FUT8 overexpression vector to induce gene silencing or overexpression , the changes of core fucosylation in hepatoma carcinoma cells had been assayed by Real-time PCR, and Lectin blot methods.By studying the changes of biological behavior of hepatoma carcinoma cells induced by RNAi,We tried to reveal the biological importance of FUT8 through cell counting kit-8, scratch assay, gelatin zymography, transwell assay. As the result showed, FUT8 overexpression could promote the multiplication of hepatoma carcinoma cells, FUT8 targeted RNAi has the negative effect. FUT8 targeted RNAi could suppress core fucosylation level apparently. Furthermore,FUT8 could elevate MMP activity so as to promote cancer invasion and get involved in the motility of hepatoma carcinoma cells to metastatic potential.Part3: Application evaluation of alpha-fetoprotein variant (AFP-L3) with lectin microcen- trifugalcolumn method in primary liver carcinomaAFP is a well-known tumour marker for hepatocellular carcinomas (HCC), but it is sometimes also increased in benign liver diseases such as chronic hepatitis and liver cirrhosis. In contrast, AFP with core-fucosylation is a very specific marker for HCC, AFP with core-fucosylation was called AFP-L3,because it was detected the L3 fraction on LCA (Lensculinaris agglutinin) lectin-electorophoresis.a1-6 fucosyltransferase (FUT8) catalyses the core-fucosylation reaction.Serum AFP-L3 variants in 622 AFP-positive PLC patients, 24 AFP-negative PLC patients and 17 AFP-positive with non-neoplastic liver disease patients were quantitatively detected by Lectin microcentrifugalcolumn, to compare and investigate the clinical application of lectin microcentrifugalcolumn method for the measurement of alpha-fetoprotein variant (AFP-L3) in primary liver carcinoma (PLC). As the result showed, In 622 AFP-positive PLC patients,there was no association of AFP-L3 with their clinical and biochemical indicators such as alanine aminotransferase (ALT) and albumin (ALB). In AFP-L3 positivc group, the leverls of gamma glutamyl Transferase (GGT), total bilirubin (TBil) were much higher than (those) in AFP-L3 negative group. The positive rate of Lectin microcentrifugalcolumn method and Lectin affinity immunoelectrophoresis autoradiography method, were 46% and 50% respectively.The coincidence of the 2 methods was 92%. Stratified analysis by AFP concentration of AFP-L3 positive rate showed that the positive rate of AFP-L3 increased with the rising of AFP concentration. In PLC patients with AFP concentration arrange from 10 to 20μg/L and patients with benign liver diseases whose concentration of AFP exceed 20μg/L, the AFP-L3 positive rates were 16.67% and 23.53% respectively, which were obviously lower than that of AFP-positive PLC patients (P<0.05). Conclusion In AFP-positive PLC patients, the detectable rate of AFP-L3 is higher, and AFP-L3 has clinical value in the auxiliary diagnosis and differential diagnosis.In consequence,from the present study, we concluded thatα1,6-fucosyltransferase is closely associated with HCC development, it could promote cancer proliferation,invasion and metastasis, correlated with cancer metastasis and prognosis of liver cancer patients closely.