The major objective of this renewal grant application is to better understand the important immunobiologic effects of the major cell wall (CW) and cell surface components of Streptococcus mutans. Although we and others have shown that induction of salivary IgA antibodies to either whole S. mutans cells or to partially purified products induces an effective caries immunity in rodents, we have almost no understanding of the cellular events which occur in the induction of this salivary immune response. In our studies, we will systematically isolate and purify the major CW components including serotype carbohydrate (CHO), lipoteichoic acid (LTA), peptidoglycan (PG) and its component parts, and cell surface associated proteins. In the first stage of these experiments, we will examine lymphoproliferative responses of murin and rat T and B lymphocytes to these purified components. In this regard, our past studies have shown that serotype CHO is a murine B cell mitogen and polyclonal activator and induces thymic independent (TI) immune responses. We will determine the precise B cell subpopulation which is stimulated by serotype CHO and use this purified substance to investigate induction of IgA responses. Work of others has indicated that LTA and PG from other bacteria exhibit lymphoproliferative properties and we will evaluate both immune responses and adjuvant activity of S. mutans LTA and determine which components of PG exhibit immunopotentiation. In additional studies, S. mutans CW proteins will be isolated by ID and 2D gel electrophoresis and major surface structural antigens identified using monoclonal antibodies. These surface proteins will be examined in the whole cell with monclonal antibodies to individual determinants by both immunofluorescence and electron microscopy. Major surface proteins will be purified by use of monoclonal antibody-immunoadsorbent columns and purified proteins tested for lymphoproliferative and thymic dependent (TD) and TI immune responses, with emphasis on IgA responses. Gnotobiotic rats will be orally immunized with appropriate S. mutans antigen(s) in combination with purified CW adjuvant and levels of salivary sIgA antibodies to antigen determined. Purified antigen and adjuvant which induce significant protective sIgA antibodies will then be determined. These proposed studies will significantly advance our understanding of immunologic properties of S. mutans CW components and their contribution to an effective caries vaccine.
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