The Neutrophil Specific CD177 Is a Novel Counter-Receptor for Endothelial PECAM-1.

CD177 is a glycosyl-phosphatidylinositol-linked member of the Ly-6 superfamily that is expressed on approximately 50% of human neutrophils, though 3–5% of Caucasians and African Americans are CD177 null. CD177 carries the clinically-important neutrophil antigen, HNA-2a - alloantibodies to which can result in profound neutropenia. Although the role of CD177 in neutrophil function is not known, it has been found to be aberrantly expressed on the neutrophils of most patients suffering from polycythemia vera, and in approximately half of patients with essential thrombocythemia. To further elucidate the function of CD177, we produced a CD177/IgG Fc fusion protein, and used it identify its binding partner on endothelial cells. Flow cytometric analysis revealed that CD177/Fc bound tightly and specifically to human umbilical vein endothelial cells, and an antigen-capture ELISA screen employing a panel of monoclonal antibodies (mAbs) to endothelial cell surface proteins further revealed the target antigen to be the endothelial cell junctional protein, PECAM-1. Surface plasmon resonance analysis showed tight binding between PECAM-1 and CD177, having a K_d of 8.17 x 10⁻⁷ M. U937 cells stably transfected with CD177 bound immobilized PECAM-1, and this heterophilic interaction could be blocked by mAbs specific for Ig domains 5 or 6 of the PECAM-1 extracellular domain, or by mAbs specific for CD177. In contrast, no inhibition was observed when mAbs against Ig domains 1 and 2 were applied. The same mAbs were also found to be effective inhibitors of neutrophil transendothelial migration. Taken together, these data establish PECAM-1/CD177 as a previously unrecognized heterophilic receptor/counter-receptor pair that functions in the multi-step adhesion and signaling cascade of events that take place during the inflammatory response.