The broad objective of this research is to elucidate the molecular events involved in regulation of lgA responses at mucosal sites. Our studies have shown that higher mammals possess a common mucosal immune system in which ingested antigens sensitize B and T cells in gut-associated lymphoreticular tissue (GALT), e.g., the Peyer's patches (PP). T helper (Th) cells regulate the inductive phase in PP, and it is thought that Th and lgA-committed B cells home to diverse mucosal effector regions such as the lamina propria of the gastrointestinal (Gi) tract for lgA synthesis. However, the precise mechanisms involved in regulation of this response remain unknown. Our past work has shown that PP dendritic cell (DC)-T cell mixtures significantly enhance lgA synthesis. Further, antigenspecific PP Th cell clones were shown to selectively support lgA responses, clearly indicating that GALT is an lgA inductive site. Recent studies have shown that recombinant interleukins, especially lL-5 and lL- 6, support enhance lgA synthesis in PP B cell cultures. The lL-5- and lL- 6-induced increase in lgA synthesis was confined to nongerminal center, surface lgA-positive (sigA+) B cells. This subset also repopulates lamina propria regions upon adoptive transfer to SCID mice which suggests that mature sigA+ B cells are sensitive to cytokines when they exit GALT. In this new renewal application we will define the T cell and cytokine requirements for sigM+ B cell switches to sigA+ in PP germinal centers by using highly purified B cell subsets and virus-transformed B cell lines from GALT. We will continue our studies to define the cytokine events involved in B cell activation, division and terminal differentiation to lgA synthesis, with special emphasis on cloned PP Th2 cells and lL-5 and lL-6. We will extensively use the single-cell ELISPOT assay to determine the relative frequencies of Th1 (lL-2 and lFNgamma) and Th2 (lL-5 and lL- 6) cells in GALT and lamina propria regions, for their roles in production of switch cytokines (TGFbeta) and cytokines inducing terminal differentiation to lgA synthesis (lL-5 and lL-6). We will extensively use two animals models to insure that the in vitro effects of cytokines represent the actual events that occur in vivo. Mice chronically depleted of CD4+ Th cells by immunotoxin- conjugated anti-CD4 mAbs which inactivate ribosomes and protein synthesis in CD4+ T cells will be used to show that the development of lgA plasma cells in effector mucosal sites is T cell dependent, while SCID mice will be used as adoptive hosts for cloned PP Th cells and slgA+ B cell subsets that respond to interleukins and that repopulate mucosal sites. We will study the antigen presenting cell functions of PP slgA+ and slgA- B cells and DC for effective induction of lgA responses. Finally, we will determine the mechanism(s) involved in PP DC-Th cell induction of pre-B B cell maturation in GALT, including a possible role for lL-7.
Showing the most recent 10 out of 185 publications
