The Antiviral Effect Of Alpha Interferon For Anti Hepatitis B Virus Grnotype ?

Hepatitis B virus(HBV)belongs to the family Hepadnaviridae,it is an enveloped virus with a circular,partially double-stranded DNA genome of 3.2kb.Hepatitis B virus infection can induce diseases,such as acute hepatitis,chronic hepatitis,hepatocirrhosis,and hepatocellular carcinoma,and thus severely threats global human health.HBV is one of the most successful human pathogens,with an estimated 2 billion people have serological evidence of past or present infected with HBV worldwide,of whom 400 million have chronic hepatitis B(CHB)infection.More than 75%patients with hepatitis B virus live in the western Pacific and Southeast Asia.China is a country that has a high incidence of HBV,with more than 120 million hepatitis B patients.Approximately 15%?40%of the hepatitis B virus carriers eventually developed HBV-related cirrhosis or HCC.Each year about 600 thousand people die from liver disease caused by HBV infection.Nucleoside analogues(NA)and alpha interferon(a-IFN)was the two effective antiviral drugs for hepatitis B therapy.A large number of studies have shown that NA prone to drug resistance mutation after long-term treatment.The HBV viral load will be reincreased and might be caused abnormal liver function when stop taking drugs Alpha interferon could inhibit viral replication and might improve the ability of the host immune response,and prevent virus invade,and removal the infected cells.However,the antiviral effect of a-IFN was dependent on HBV genotype.For a-IFN treatment,the response efficiency of HBV genotype B is better than that of C genotype.There was not the report about the response efficiency of a-IFN for HBV genotype I.The efficient eukaryotic expression vector of pcDNA3.1(+)and whole genome sequence with different HBV genotype were used to construct recombinant plasmids.The constructed plasmids were introdueced into HepG2 cell line with the lipidosome,the expressed hepatitis B surface antigen was qualitative and quantitative analysis determined with ELISA,MTS method was used to detect the toxic effects of a-IFN to the HepG2 cell with transient transfecting plasmid of HBV genotype B,C and I.,The anti-viral effect of ?-IFN was valuated through the expressed HBsAg which detect by ELISA.This study result showed that we successfully constructed pcDNA3.1(+)/HB V recombinant plasmids with different genotypes.The recombinant plasmids were successfully introduced into HepG2 cell line with the lipidosome,and then we detected HBsAg expressed in the supernatant.MTS method result show that one-fold-overlength genome of HBV genotype B and C have had little toxicity,one-fold-overlength genome of HBV genotype I had toxic effective for the HepG2 cell.?-1b had no inhibition effect for the HepG2 cell of transient transfection with one-fold-overlength genome of HBV genotype B,C and I.The antiviral effect of a-lb with 8000 IU/ml was better than other concentration groups for the HepG2 cell with transient transfecting plasmids of HBV genotype B,C and I.Under the circumstances of ?-1b,the HepG2 cell with transient transfected HBV genotype B showed sustained response effect,HBV genotype I had a short response effect in the early period,and HBV genotype C had an earlier response effect,however,the response effect was unstable.A total of 1148 HBV genome sequences from patients throughout China were collected via the NCBI(National Center For Biotechnology Information)database(information including:genotype,territory and clinical status).HBV genotypes were classified by a direct reference from the Genbank sequence annotation,phylogenetic tree and online software analysis(http//www.ncbi.nlm.nih.gov/projects/genotyping/formpage.cgi).The phylogenetic tree was constructed based on the neighbor-joining method by MEGA5.0 software.HBV sequences were grouped based on phylogenetic tree and the distance between the groups was calculated by using the computer between group mean distance methods.712 HBV sequences with clear annotation of clinical symptoms were selected to,analyses the correlation of mutation and clinical symptoms.Characteristics of sequences were analyzed by using DNAStar and BioEdit software packages.The codon usage bias and RNA secondary structures analysis were performed by RNAdraw software.Recombination analysis was performed by using Simplot software.The results suggested that,in China,HBV genotype C was the predominant in Northeastern,genotype B was predominant in Central Southern areas,genotype B and C were both dominant in Southwestern areas,and the recombinant genotype C/D was predominant in Northwestern areas.C2 and B2 were identified as the two major sub-genotypes,FJ386674 might be a new sub-genotype as B10.The BCP(basal core promoter)double mutation and pre-C mutation showed various significant differences between hepatitis symptoms.In addition to ATG,many other HBV initiation codon are also existing.HBV have the codon usage bias,the termination codon of X,C and P open reading frame(ORP)were TAA,TAG,and TGA,respectively.The major stop codon of S-ORF was TAA(96.45%)and TGA(83.60%)in B2 and C2 subtype,respectively.The study showed that 8000 IU/ml of ?-1b had better antiviral effect on the HepG2 cell with HBV genotype B,C and I.The recapitulated results of Chinese epidemiology of HBV might be meaningful critical for the future prevention and therapy of HBV infections.