Signalling mechanisms in B cell differentiation: Studies on specific human immune response in vitro

Sammanfattning: We need to understand the major signalling events in human B cell differentiation in order to be able to regulate the generation of human antigen specific antibodies. This study has focused on the signalling mechanisms involved in generation of germinal centre reaction and secretion of specific antibodies by human B cells, using different in vitro immunisation technologies. Naive human B cells that were stimulated by crosslinking their sIg, CD40 and CD44 acquired a germinal centre phenotype. Signalling via these surface molecules caused increased proliferation and made the B cells enter an apoptotic cycle, which is characteristic for germinal centre B cells. Three different in vitro immunisation protocols were utilised to study the mechanisms behind secretion of human antigen specific antibodies. First, soluble antigen in the cultures was shown to abrogate specific Ig response by reducing the number of memory B cells secreting specific antibodies to a recall antigen. By crosslinking the antigen bound to the sIg of those B cells the antigen specific suppression could be lifted. Secondly, purified B cells together with isolated CD4+ T cells and a superantigen could be induced to secrete antibodies to primary antigens, by crosslinking the antigen bound to their sIg. The specific antibody secretion was totally dependent on CD40 and CD86 ligation of the B cells. The third protocol demonstrates specific IgE production using a system that induces isotype switching in vitro. Specific IgE production appears to be dependent on NFkB activation by IL-4, produced and secreted by Th2 cells generated during the primary response. This thesis provides possible signalling mechanisms for the initiation of germinal centre reactions. It also presents data on antibody production in vitro in terms of antigen presentation and cell-cell interaction.