The adult B-cell repertoire is extraordinarily diverse and consists of greater than 10 million unique antibody specificities. The long range objectives of this project are to understand and analyze the variables which regulate the expression of this B-cell repertoire. To achieve these goals, both the role of genetics as well as the influence of the host environment on B-cell repertoire expression must be clarified. Since the contributions of environmental selective forces to the ultimate expression of the B-cell repertoire remain unresolved, the experiments described herein are designed to address this issue. To date, the antigens which have been used to evaluate the influence of the environment on B-cell repertoire expression have been simple, synthetic antigens such as phosphocholine or the relatively complex antigen influenza virus. However, a more direct test for the physiological relevance of B cell-antigen interactions in the regulation of B-cell repertoire expression would be to analyze the influence of the host environment on the expression of B cells specific for a bacterial antigen which is a member of, or cross-reactive with, the normal environmental bacterial flora of the host. To this end, a modification of the limiting dilution splenic fragment culture system has been developed which will permit us to analyze the responsiveness of individual B cells specific for the Gram-negative bacteria, Salmonella typhimurium. Therefore, we plan to utilize this system to study more directly the role of the host environment on shaping the B-cell repertoire by: a) comparing the characteristics of the splenic salmonella-specific B-cell repertoire in germfree (GF) mice, GF mice re-associated with bacteria, and conventional (CV) mice; b) assessing the primary salmonella-specific B-cell repertoire in the Peyer's patches, lymph nodes, and bone marrow in CV and GF mice; and c) analyzing the salmonella-specific pre-receptor B-cell repertoire in the bone marrow of normal CV mice and salmonella-susceptible CBA/N mice. It is anticipated that an increased understanding of the role of the host environment on B-cell repertoire expression may permit the manipulation of this environment to either enhance host immune responses or inactivate host responses. Thus, the results of the experiments described herein will contribute to our understanding of the potential effects of either vaccination or natural exposure to infectious agents. Finally these studies should also further our understanding of the mechanisms of host-parasite interactions.
