Date of Award
Doctor of Philosophy
Molecular Biology, Microbiology and Biochemistry
The mucosal surfaces of the gastrointestinal, respiratory, and reproductive tracts, serve as portals of entry for the majority of pathogens. Systemic (parenteral) vaccines can provide protection against some mucosal pathogens, like poliovirus and influenza virus; however, they do not induce mucosal immune responses, which are critical for protection against mucosal pathogens. The development of mucosal vaccines has had limited success thus far, due to several factors such as poor antigen uptake at the mucosal surfaces, lack of immunogenicity of vaccine formulations, and the need for a better understanding of mucosal immune responses. In order to elicit an immune response, mucosal antigens must traverse the epithelium and reach the sub-epithelial immune compartments. Understanding how antigens are internalized at mucosal surfaces and how the nature of antigen and the route of antigen uptake affect their immunogenicity would greatly aid in the development of successful mucosal vaccines. In addition, even currently licensed mucosally-administered vaccines induce an inferior immune memory, and understanding how prime-boost immunization regimens affect mucosal and systemic humoral immune memory remains an insufficiently explored area of research that is critical for vaccine development. In this work, we examined the routes of mucosal antigen internalization and immune responses to mucosal antigens following various mucosal and systemic prime-boost immunization strategies. We showed that epithelial cells (ECs) overlaying the small intestine (SI) and the female reproductive tract (FRT) can internalize NPs < 40 nm in size. In the SI, soluble antigens (such as dextran and Ovalbumin) are internalized via goblet cell associated passageways (GAPs), as shown previously (1). We found that immunization via any mucosal surface (by mouth, intra-vaginal, or intra-nasal) using both soluble Ovalbumin (Ova) and Ova conjugated to nanoparticles (NP-Ova) induced serum immunoglobulin (Ig) IgG1, IgG2c and intestinal IgA antibodies. The dynamics of these responses varied depending on the nature of the antigen and on the prime-boost routes of immunization. Immunizing by mouth (PO) with soluble Ova, induced oral tolerance (OT), which is characterized by a systemic IgG response that cannot be boosted, and an intestinal IgA response that is abrogated following a subcutaneous (SC) secondary immunization. Intra-nasal (IN) administration of a similar dose of Ova, however, was found to be highly immunogenic, prompting systemic IgG and mucosal IgA antibodies in the intestine and the FRT following a secondary SC immunization. Mucosal immunization (PO, per-vaginal, or IN) using NP-Ova induces substantial serum IgG1 antibodies that were boosted following a secondary immunization. In addition, mucosal priming with NP-Ova followed by either a mucosal or systemic secondary immunization induces serum IgG2c as well as intestinal IgA antibodies. By reversing the prime-boost immunization regime (SC priming followed by PO boosting with NP-Ova), systemic IgG1 antibodies were produced, but no systemic IgG2c or mucosal IgA antibodies. This indicates that mucosal priming is required for a Th1/Th2 systemic IgG response and for the induction of intestinal IgA antibodies. In addition, SC boosting after mucosal priming induced superior immune memory compared to boosting via a mucosal route, with systemic and mucosal antibody titers remaining unaltered for at least 6 months. These findings have relevance for the development of effective mucosal vaccines and vaccination strategies to induce long-lasting mucosal and systemic immune responses.
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