Expression Of The E2 Gene Of Hepatitis C Virus In E.coli And Mammiferous Cell And The CTL Response Induced By E2 Protein

Hepatitis C Virus(HCV) is a single-stranded positive-sense RNA virus that account for most cases of post-transfection non-A non-B hepatitis. In more than 70% of the cases, HCV causes a prolonged, persistent infection, in which about 25% of the patients can eventually lead to cirrhosis and hepatocellular carcinoma. Currently, the lack of efficient antiviral treatment against HCV makes the development of a vaccine highly difficult. The envelope glycoprotein of HCV is one of the important targets in the study of efficient vaccine in the world. It has been proved that the DNA vaccine of E2 could induce humoral and cellular immunity in mice. Whether induce strong specific antibody and T lymphocytes, causing specific CTL response to eliminate virus in vivo or not is the main index for the vaccine evaluation . In this study, we focused on specific CTL inducing by E2 vaccination.The four recombinant plasmids has been constructed, pCE2-1(422-661), pCE2-2(383-661), pQE2(417-661) and pET-HIS-E2(417-750), which were transformed into COS-7, JM109 and BL21.The expression of E2 protein has been identified by Western-blot analysis using the E2 monoclonal antibody. However,the express efficiency of E2 protein in all of the four vectors are low, more efforts should focus on how to improve it.In the study of the induction of specific CTL, the recombinant E2 that its contain 6 His at N terminal end was purified with Ni-NTA-Superflow column after expression in E.coli JM109. To study the immunological properties of E2 protein, the BALB/c mice were vaccinated with purified E2 protein. Five weeks later, the vaccinated mice had been killed and extracted spleens used to prepared the effecter cells. After simulated and augment in vitro, they were used to kill the target cells P815, which were transfected by recombinant expression vector pCE2. By LDH assay, we observed that more than 30% of the target cells were killed at the ratio 200:1, and there are different results with different immunity injection routes. Altogether our data suggest that the E2 protein can provoke E2 specific CTLs in the mice, from which we conclude that the E2 protein is a good candidate for developing a vaccine to prevent HCV infection.