Integrins are important for T cell interactions with endothelial cells. Because the integrin α E β 7 is expressed on some circulating gut-homing T cells and as T cell numbers are reduced in the intestinal lamina propria of α E-deficient mice, we evaluated whether α E β 7 mediates binding to intestinal endothelial cells. We found that anti-α E β 7 mAbs partially blocked the binding of cultured intraepithelial T cells to human intestinal microvascular endothelial cells (HIMEC). Furthermore, α E β 7-transfected K562 cells bound more efficiently than vector-transfected K562 cells to HIMEC. Finally, HIMEC bound directly to an α E β 7-Fc fusion protein. These interactions were partially blocked by anti-α E β 7 mAbs, and endothelial cell binding to the α E β 7-Fc was dependent upon the metal ion-dependent adhesion site within the α E A domain. Of note, the HIMEC lacked expression of E-cadherin, the only known α E β 7 counterreceptor as assessed by functional studies, flow cytometry, and RT-PCR. Thus, HIMEC/α E β 7 binding was independent of E-cadherin. In addition, this interaction appeared to be tissue selective, as HIMEC bound to the α E β 7-Fc, whereas microvascular endothelial cells from the skin did not. Finally, there was evidence for an α E β 7 ligand on intestinal endothelial cells in vivo, as α E β 7 expression enhanced lymphocyte binding around vessels in the lamina propria in tissue sections. Thus, we have defined a novel interaction for α E β 7 at a nonepithelial location. These studies suggest a role for α E β 7 in interactions with the intestinal endothelium that may have implications for intestinal T cell homing or functional responses.