Study On The Antigenicity Of The HVR1 Synthetic Peptides Of HCV

Graduate student : hao chen Major:Infectious disease TutorrYun ChengHepatitis C virus (HCV) is the major cause of , non-non B viral hepatitis .More than one billion people infected with HCV will become chronic and develop cirrhosis and perhaps hepatocellular carcinoma . But its nosogenesis is still not clear. So HCV infection is now a world-wide health problem. In 1996, Farci used E2 protein inoculate gorilla, and found there are neutralizing epitopes in the hypervariable region 1(HVR1) of HCV. Many research have show that there is cross-reaction between anti-HVRl antibody and the HVR1 of different HCV subspecies , and suppose that the conservative epitopes in this region is the molecular basis for the cross-reaction. Seven HVR1 antigen peptides are designed and synthesized according to the analysis of sequence diversity of HCV HVR1 in Chinese patients infected with genotype Il/lb and III/2a HCV.We designed ELISA method to test the cross-reactivity between mimic epitopes of hypervariable region 1 and serum of HCV patients, in the meantime we predict the antigencity of syntheticpeptide by the computer modeling. Results (l)No.l and No.5 synthetic peptides have abroad cross-reactivity with the serum of HCV patients, 43.3% and 30.0% respectively. (2) There is cross-reactivity between these synthetic peptides. (3 ) Combined No. 1 and No.3, No.l and No.4, No.l and No.5 , No.l and No.7, No.3 and No.4 can react with 53.3%, 56.7%, 60.0%, 53.3%, 26.7% of all serum of HCV infected patients (4) 385 01amino acids of hypervariable region 1 of hepatitis C virus may be a antigen epitope, and there is still no report about this antigen epitope .To explore the optimal immunity approach and improve the antigencity of synthetic peptides, we immune BALB/c mice by the routine foot immunity method and in-spleen immunity method , and use glutaral cross-linking method to improve their antigencity. We found (l)No.l and No.5 peptides can induce relatively strong antibody reaction., and get the best result when they are combined. (2) there are cross-reaction between them.(3) glutaral cross-linking can improve their antigencity.(4) the sera of the subject mice can cross-react with other peptides, especially the No.3 peptide.(5)we can improve their antigencity by glutaral cross-linking, and get the best effect by in-spleen immunity method.We use ELISA method to test the level of cytokinesin the culture medium of PBMC of HCV patients when stimulated with those peptides , and in the sera of mice immunized with them. A interesting result was found that the levels of IFN- y ,IL-4,andIL-10 rised ,but there are no obvious changes as to the level of IL-2 and TNF- a . These results suggest the cytokines induced by those peptides incline to be the TH2 group cytokines. Maybe that is a mechanism of HCV escape from the immune surveillance. The rise of IFN- Y mean those peptides can stimulate cellular immune reaction to HCV, for IFN- Y . is one of the important cytokines in elimination of virus.Strong foundation has been built to the study of the neutralizing epitopes of HVR1 and multi-peptide vaccine to HCV.