Circulating T and B lymphocytes bind to high endothelial cell venules (HEV) as the first step in their emigration from blood into lymph nodes (LN) and Peyer's patches (PP). We have used an in vitro lymphocyte-HEV adherence assay to probe the molecular basis of this binding and have identified rat lymphocyte surface adhesion molecules or 'homing receptors' specific for HEV of LN. This proposal is divided between experiments specifically directed towards the analysis of the genes encoding receptors specific for HEV of LN or HEV of PP and to analyze the structure and regulation endothelial ligands on HEV. Initially we will analyze the DNA sequences encoding the surface receptor for peripheral lymph node HEV and will use the available cDNA fragments to screen for full-length cDNA clones. In parallel studies, the antibody and nucleic acid probes specific for receptors for PP high endothelium will be employed to identify and characterize genes sequences encoding Peyer's patch HEV receptors. The cDNA clones for LN and PP HEV receptors will then allow us to clone and characterize the genomic sequences and their organization. The expression of receptor gene sequences in immature and mature lymphocytes as well as leukocytes and hematopoietic cells will also be analyzed. Full length cDNA will also be transfected into mammalian cells expression systems to study the molecular basis of lymphocyte migration. Successful expression of cloned cDNAs or of corresponding genomic sequences will not only confirm a specific role of the A.11 and 1B.2 antigen in HEV adherence and in vivo migration, but will also be the most direct and powerful method for determining the specificity of homing receptor encoding cDNAs for lymph node and mucosal lymphoid organs. In parallel experiments, we will test the hypothesis that the panel of monoclonal antibodies, KJ-4, recognize ligands on LN HEV and to investigate whether incubation of isolated HEV cells with specific cytokines can up-regulate their capacity to adhere lymphocytes. The complementary approaches have the potential to elucidate the molecular mechanisms regulating lymphocyte-endothelial cell interactions during lymphocyte migration.
