Characterization of immune responses induced by plasmids encoding bovine rotavirus antigens in the Murine model
Date
1999-01-01
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ORCID
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Degree Level
Doctoral
Abstract
DNA immunization effectively induces humoral and cell-mediated immunity to numerous infectious diseases. To investigate the potential use of DNA-based vaccine for induction of bovine rotavirus (BRV) specific immune responses, I characterized the immune responses induced by plasmids encoding 2 important neutralizing antigens, VP4 and VP7, in a murine model. Immunization with plasmids encoding VP4 protein did not induced detectable BRV-specific antibody responses in mice. However, it induced BRV-specific cell-mediated immune responses characterized by an increased number of BRV-specific cytokine secreting cells in the spleens of immunized mice. In addition, the immunized animals were primed for both antibody and cellular immune responses following BRV boost. However, plasmids encoding VP4 did not boost the BRV-specific immune responses in the BRV-primed mice. The BRV-VP7 is also considered an important antigen for inducing protective immunity against rotaviruses. However, this study indicated that the VP7, either by itself or in the context of viral particle, was poorly immunogenic in mice. This project then explored several approaches to enhance the immunogenicity of the VP7 protein when expressed by the plasmid. These approaches included: (i) enhancing the level of gene expression by modifying the expression cassette of the plasmids; (ii) changing the cellular localization of the plasmid-expressed protein; (iii) co-administration of plasmids encoding VP4 and VP7; (iv) creating plasmids encoding a deletion mutant of VP7; and (v) construction of plasmids encoding chimeric proteins of VP7 and complement C3d or bovine herpes virus-1 glycoprotein D (BHV-1 gD). The results from these studies demonstrated that cellular localization and nature of the VP7 antigen could greatly influence the immunogenicity of the plasmid-expressed antigen and the pattern of immune responses in immunized mice. Plasmids encoding membrane-bound VP7 induced the best BRV-specific immune responses, following BRV boost or in BRV-primed mice, when compared with the authentic and the secretory versions. In addition, the plasmid encoding a deletion mutant VP7, induced predominantly BVR-specific IL-4 production. Strategies that had previously been shown to enhance the immunogenicity of protein antigens were also examined. However, different outcomes were observed when these strategies were applied to plasmid immunization. First, addition of the C3d gene into the expression cassette which encoded antigen-C3d chimeras inhibited the induction of both humoral and cell-mediated, antigen-specific immune responses. Secondly, co-expression of the gD molecule as a gD-VP7 chimera enhanced the level of BRV-specific antibody responses following viral boost or in the BRV-primed mice. These results with the gD protein suggested an alternative approach for improving the immunogenicity of poorly immunogenic antigens. None of the plasmids used in this study induced BRV-specific antibody responses following a primary immunization. However, immunological activity was evident following BRV administration, either prior to or following plasmid immunization. Each plasmid induced a distinct pattern of immune responses in both naive and BRV-primed mice. These investigations demonstrated specific ways that DNA immunization could be modified to modulate the outcome of BRV-specific immune responses. (Abstract shortened by UMI.)
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Degree
Doctor of Philosophy (Ph.D.)
Department
Veterinary Microbiology
Program
Veterinary Microbiology