INTEGRIN LIGAND IN GUT LAMINA PROPRIA

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Mucosal T cells subsets have distinct localization patterns, as CD8+ comprise over 90 percent of the intestinal intraepithelial T lymphocytes while CD4+ T cells predominate within the lamina propria. Little is known about the mechanisms which account for this subset specific localization. The integrin alphaEbeta7 expressed on greater than 90 percent of CD8+ lymphocytes and on 40-50 percent of CD4+ cells diffusely distributed with the intestinal mucosa but on less than 3 percent of peripheral blood lymphocytes, and binds to E-cadherin expressed on epithelial cells. Thus, it is a candidate to function in T cell/epithelial cell interactions which may modify T cell localization. To evaluate the in vivo function(s) of alphaEbeta7, integrin alphaE deficient mice were generated. Study of these animals revealed reduced numbers of T cells within intestinal epithelium and also within the lamina propria where E-cadherin is not expressed. Furthermore, freshly isolated intestinal intraepithelial lymphocytes (iIEL), which express alphaEbeta7, adhered more efficiently to lamina propria tissue sections than iIEL isolated from alphaE deficient mice. These observations led us to hypothesize that alphaEbeta7 mediates an adhesive interaction to a ligand expressed within the lamina propria. In this application, studies are proposed to demonstrate that an additional alphaEbeta7 ligand is expressed within the lamina propria and to define this ligand at the molecular level. In aim 1, alphaEbeta7 transfectants will be tested to confirm that alphaEbeta7 expression confers adhesion of transfected cells to the lamina propria and to determine if human alphaEbeta7 mediates a similar adhesive function. In addition, studies will be performed to identify a cultured cells line which expresses the putative additional alphaEbeta7 ligand, using adhesion assays to identify cell lines which support alphaEbeta7 dependent adhesion. In aim 2, anti-ligand monoclonal antibodies or a soluble recombinant form of alphaEbeta7 which binds to this ligand will be generated and used to determine the biochemical structure of the alphaEbeta7 ligand. In aim 3, the gene encoding this lamina propria ligand will be cloned using either expression cloning or ligand purification followed by amino acid sequence determination. These studies will enable the identification and characterization of a second alphaEbeta7 counter-receptor and will provide insight into the mechanisms of mucosal lymphocytes localization. As an anti-alphaE monoclonal antibody dramatically reduced intestinal inflammation in a murine inflammatory bowel disease model, these studies may have important implications in the treatment of inflammatory bowel disease.