Correlation Between Core,Nonstructural Protein 3 (NS3) Of Chronic Genotype 1b Hepatitis C Virus And Development Of Hepatocellular Carcinoma

Background and aim:Goldfield et al first reported the non-A,non-B hepatitis virus in 1974, Chiron' Corporation(USA)finally succeeded in cloning part of the genome of the virus from the plasma of a chimpanzee chronically infected with the non-A,non-B hepatitis virus that came to be known as the hepatitis C virus(HCV).The data from WHO shown that HCV is a major health problem,it have infected approximately 3%of the world's population and estimated 170 million people infected worldwide,and the infected population was arise about 35 thousand a year.In China,the statistical results showed that the infection rate of HCV was 3.2%,and about 38 million people infected.70%infected individuals would give rise to chronic hepatitis,and part of them would develop to liver cirrhosis or/and hepatocellular carcinoma in 20-30 years.HCV was a global disease,which posed major risk for development of liver cirrhosis and hepatocellular carcinoma,and the HCV infection rate was arise every year in our country.HCV was classified within the Flavivirideae family by International Committee on Nomenclature of Viruses(ICNV).The HCV genome, single-stranded,positive sense RNA of approximately 9.6 kb,encodes a polyprotein of about 3,000 amino acids(aa),HCV precursor protein yields at least 10 distinct cleavage products,in the following order:NH₂-Core-E1-E2-p7-NS2- NS3-NS4A-NS4B-NS5A-NS5B-COOH.The Core,E1,E2 was the structural protein of HCV,and the NS2-NS5 was the non-structural protein of HCV,which were the structure correlation with the virus replication and the function of p7 was still unknown.Those protein composed the HCV together,they not only been involved with the viral repeating,but also been involve in the cellular modulating process.We think that if we recognize the structure and the function of the HCV protein completely,we would clear this disease.The viral genome exhibits a considerable degree of sequence variation,based on which HCV is currently classified into at least 6 genotypes:1a,1b,2a,2b,3a,3b and many subtypes.HCV subtype 1b(HCV-1b)is the most prevalent subtype in most parts of Asia including Japan and China,and is likely more associated with poor responsiveness to interferon treatment,rapid disease progress and development of HCC than the other HCV subtypes.Lots of researches about HCV had been done,but we didin't recognize the HCV very well,many researchers studied HCV from different aspect,and achieved some results.In those achievements that the Core and NS3, NS5A were confirmed that be correlation with the development of HCC.Method:1.Sera were collected and classed into three groups which were HCVcarriers and HCV-infected patients with and without HCC based on the patients clinical data.2.Serum samples were amplified by nested polymerase chain reaction (nested-PCR)to obtain the genotype 1b using the special genotyping primers.3.Obtained the PCR products of Core region by nested-PCR method and directly sequenced.4.The phylogenetic tree was drawn up by using the neighbor-joining method with the N-J plot program in the CLUSTAL W package to constructed to elucidate the relationship between the three groups.5.Obtained the PCR products of NS3_1027-1206region by nested-PCR method and directly sequenced,then the sequences result translated to amino acid sequences by the Chromas software.6.The secondary structure of NS3_1027-1206was predicted using GENETYXMACver. 10.1.7.One-way analysis of variance and x2 test was adopted to compare every index value by SSPS 12.0 statistic analysis software.Results:1.The percentage of genotype 1b was 71%in the carrier group;77.5%in the HCC group;73%in the non-HCC group,and the percentage had no statistically significant difference(P＞0.05)in Changchun and Jilin areas.2.The homology of nucleotide sequences of Core region which were amplified using the 1b special genotyping primers was 90%,and the homologies were 90%-97%compared with the data which published on the GENEBANK,and there was no difference in the nucleotide sequence of the three groups by analysis with the phylogenetic tree,and this part of Core region had no correlation with the development of HCC.3.The homology of nucleotide and amino acid sequence was 95%,there was no particular mutation in the NS3_1027-1206region amino acid sequence.4.On the basis of secondary structure of NS3_1027-1206,the HCV isolates were classified into two groups,group A and group B.5.Group A was predominant in the carriers and non-HCC patients,group B was predominant in the HCC patients.The difference in the distribution pattern of those group between carriers and non-HCC patients was no statistically significant(P＞0.05),but was statistically significant(P＜0.01)between carriers, non-HCC patients and HCC patients.HCV-NS3 had correlation with the development of HCC.Conclusions:1.The genotype 1b became predominant in all the patients,the percentage was 70%and had no statistically significant difference(P＞0.05)in Changchun and Jilin areas.2.Nucleotide sequences of Core region which were amplified using the 1b special genotyping primers were conserved,this part of Core region had no correlation with the development of HCC.3.On the basis of secondary structure of NS3_1027-1206,the HCV isolates were classified into two groups,group A and group B.4.HCV-NS3 had correlation with the development of HCC.