CHARACTERIZATION OF ADHESION RECEPTORS ON CULTURED MICROVASCULAR ENDOTHELIAL-CELLS DERIVED FROM THE RETROORBITAL CONNECTIVE-TISSUE OF PATIENTS WITH GRAVES OPHTHALMOPATHY

T lymphocytes have been demonstrated recently to play an important rol e in the pathogenesis and propagation of Graves' ophthalmopathy (GO). Recruitment of T cells to the retroorbital tissue in GO involves the a ctivation of certain adhesion molecules both in the vascular endotheli um and in the extravascular connective tissue within the retroorbital space. To characterize the interactions between orbital endothelial ce lls (OECs) and circulating T cells in vitro, we designed a two-step im munopurification procedure with bead-immobilized Ulex europaeus I lect in and anti-human endothelial cell antigen (CD31) monoclonal antibody for rapid and reproducible isolation of highly pure microvascular endo thelial cell populations from small quantities of retroorbital connect ive tissue. Endothelial origin of the resulting cell populations was c onfirmed by positive immunoreactivity for von Willebrand factor, CD31 and thrombomodulin. Under baseline conditions, GO-OECs, but not normal OECs, expressed intercellular adhesion molecule 1 (ICAM-1) and CD44 i mmunoreactivity but no immunoreactivity for endothelial leukocyte adhe sion molecule 1 (ELAM-1) and vascular cell adhesion molecule 1 (VCAM-1 ) was detected. Exposure of GO-OEC and normal OEC monolayers to interf eron gamma, interleukin 1 alpha and tumor necrosis factor alpha result ed in marked up-regulation of immunoreactivity for ICAM-1 and in induc tion of ELAM-1 and VCAM-1. Blocking experiments using monoclonal antib odies directed against various adhesion molecules demonstrated that in teractions between matched activated T lymphocytes and OECs were media ted by integrin-dependent (ICAM-1/leukocyte function-associated antige n 1 (LFA-1); VCAM-1/very late antigen 4 (VLA-4)) and integrin-independ ent (CD44) pathways, and revealed marked differences when comparing GO -OECs and normal OECs, In conclusion. the availability of OECs from af fected retroorbital tissue of patients with GO provides a valuable too l for studying further the mechanisms responsible for orbit-specific l ymphocyte recruitment in GO.