Profiles Of Immune Responses Elicited By Different Forms Of The Hepatitis B Virus Surface Antigen As Studied By The DNA Immunization Approach

Objective: To investigate the immune profiles of three forms of the Hepatitis B virus surface antigen (HBsAg), large, middle and small surface antigens,and further to provide the therapeutic basis for developing new vaccine against HBV.Methods: Three forms of HBsAg DNA vaccines LHBs,MHBs and SHBs were constructed to express the large, middle and small forms of HBsAg, respectively. A consensus S gene for HBV adr subtype (adr-S-Cons) was used as the template for these DNA vaccines. Balb/c mice received DNA immunization at weeks 0, 2, and 4 by the electroporation method. Serum samples were taken prior to each immunization and two weeks after the last immunization. S antigen-specific antibody responses were analyzed by ELISA against either commercial HBsAg or different forms of HBsAg from the above three DNA vaccines (LHBs, MHBs and SHBs). In addition, Western blot was conducted to verify the specific antigens recognized by mouse immune sera. Enzyme-linked immunospot (ELISPOT) assays for antigen- specific antibody secreting cells (ASCs) were performed to detect the profiles of B cell immune responses against different forms of S antigens. Finally, S antigen-specific T cell immune responses were analyzed by ELISPOT to detect IFN-γ? and IL-2 responses in immunized mice. Results:1. Three forms of S DNA vaccines (LHBs, MHBs and SHBs) were successfully constructed.2. Expression of LHBs, MHB and SHBs DNA vaccines were confirmed by transfection of 293T cells and Western Blot analysis.3. Positive antibody responses against either HBsAg,LHBs,MHBs or SHBs antigens were detected in DNA immunized mice sera. However, the levels of antibody responses against different forms of antigens are different among different immunized animal groups suggesting that each form of S antigens may have its unique conformation.4. Study of antigen-specific ASCs showed that different forms of S antigens have different levels of immunogenicity in eliciting S specific B cell responses against different forms of S antigens.5. No difference of T cell immune responses was observed among different forms of S antigens when measured by IFN-γor IL-2 ELISPOT analysis.Conclusions:1. All three forms of S DNA vaccines were immunogenic by eliciting S-specific immune response in Balb/c mice.2. Different forms of S antigens have different levels of immunogenicity in eliciting S specific B cell immune response. Overall, MHBs DNA vaccine containing preS2 antigen was the most immunogenic among three forms of S antigens in eliciting S-specific antibody and B cell immune responses.3. No significant difference was observed among different forms of S antigens in eliciting S specific T immune responses.