PCR analysis has characterized tissue-specific expression of CD44 isoforms resulting from alternative splicing of up to 10 exons. To examine isoform expression at the level of cell surface protein and to probe the possible biological functions of CD44 isoform products in vitro or in vivo, we have generated monoclonal antibodies specific for variable exon products. Immunization with fusion proteins containing variable CD44 exon products produced hybridomas with multiple CD44 variable exon specificities. Flow and immunohistochemical techniques detected variable exon determinants on epithelial cells and on activated lymphocytes. mAbs and fusion proteins are being used to assess the function of CD44 variable isoforms. To identify functional cell surface molecules expressed on activated ymphyocytes, mAb were generated by immunizing rates with activated mouse B cells. One of these mAb (GL7) reacts with subpopulations of activated B and T cells, as well as on a functionally distinct supopulation of CD4+8-thymocytes with unique cytokine secretory capacity, and on bone marrow pre-B cells. GL7 administered in vivo influences early B cell development. Expression cloning identified the determinant recognized by GL7 as an alpha 2, 6-sailyl transferase-dependent epitope(s) present on human as well as mouse lymphoid cells. Characterization of the activation antigen-specific mAb GL1 led to identification of the mouse B7-2 costimulatory molecule, which has predominant functional costimulatory role both in vivo and in vitro. Anti-B7-2 mAb inhibited accessory cell-dependent responses of T cells in vitro and in vivo, indicating that B7-2 is a functional costimulatory molecule for T cell-dependent (TD) responses. Expression of B7-2 costimulatory molecules during in vivo TB antibody responses correlated with somatic hypermutation and affinity maturation, and in in vivo treatment with anti B7-2 inhibited TD responses as well as memory formation and somatic mutation. T cell dependent (TD) B cell activation was analyzed in the antibody responses of mice defective in the ataxia telangiectasia (AT)mutated (Atm) gene. These mice were found to have a profound defect in vivo in Ig class switching during TD antibody responses, with most extensive defects observed in IgE and IgA response, paralleling the clinical AT syndrome. In vitro studies have identified a defect intrinsic to B lymphocytes in the ability to Ig switch in response to activation with cytokines plus LPS or anti-CD40. Parallel studies are in progress in Atm mice and AT patient lymphocytes to further characterize T and B cell functional status.
| Lumsden, Joanne M; Williams, Joy A; Hodes, Richard J (2003) Differential requirements for expression of CD80/86 and CD40 on B cells for T-dependent antibody responses in vivo. J Immunol 170:781-7 |
