Heparan sulfate influences many cellular processes including adhesion, motility, ligand-receptor interaction, and proliferation. B cells account for a major component of the adaptive immune system, in which they aid in the elimination of, and protection against, invading pathogens. Crucial to the initiation and maintenance of a B cell response is B cell localization, cytokine/chemokine responsiveness, and cell-to-cell interaction. We have recently observed that heparan sulfate is drastically upregulated on B-cells. Indeed, whereas naive B-cells do not express heparan sulfate, B-cells isolated from mice infected with viruses, or mice injected with poly I:C, express heparan sulfate. Importantly this phenotype is abolished when interferon receptor deficient mice are used, indicating a requirement for interferon in this process. Accordingly, B-cells treated ex-vivo with interferon alone express high levels of heparan sulfate. Interestingly, we have shown that interferon treated B-cells are more responsive to the B cell-specific cytokine APRIL, a phenotype associated with increased heparan sulfate-expression. Altogether, these experiments suggest that upon viral infection, interferon induces heparan sulfate expression on B-cells, rendering these cells more responsive to their environment, and potentially affecting the B-cell antibody response. To test this hypothesis, we have generated mice that are deficient for heparan sulfate expression on B-cells (EXT1-/- mice) and have shown that their B- cell development is not altered. Interestingly, preliminary data generated using 2-photon microscopy indicates that B-cell motility was affected in the presence of interferon in a heparan sulfate-dependent manner. Using influenza as an infectious model, we have demonstrated that both the thymus -dependent and - independent antibody responses are increased in EXT1-/- mice. Despite generating this increased response, viral restriction in EXT1-/- mice is unaffected. This proposal aims to further characterize the role of heparan sulfate expression on B cells during a humoral response. Due to the preliminary nature of our in vivo infection data, we aim to confirm and further our initial findings. To this end, we will infect mice with a range of infectious doses of influenza virus and assess the quality of the antibody response by monitoring host health, viral restriction, responding B cell subsets, and the antibodies produced. This will be accomplished by measuring viral titers, antibody titers, responding B cell populations, and host morbidity and mortality. To address the observations that EXT1-/- mice generate an expanded antibody response, yet are unable to more efficiently restrict influenza, we will assess the specificity and efficiency of the humoral response. This will be done by determining the affinity and the virus-neutralizing capabilities of EXT1-/- antibodies. Finally, we will assess the protective effect of serum antibodies isolated from influenza- immune EXT1-/- mice. We hope to reveal additional alterations in the EXT1-/- antibody response that will further support our hypothesis that IFN-mediated induction of HS on B cells serves as a novel form of immune regulation. Based on this, we believe that this work may aid in the development of vaccines in which an effective B cell response is necessary.
The work we prose here may help in understanding of the antibody response against infectious disease. Furthermore, understanding of the topic may enhance vaccine development.
