T cells bind to cytokine-activated endothelial cells via a novel, inducible sialoglycoprotein and endothelial leukocyte adhesion molecule-1.
The action of human rIL-1 beta on confluent, quiescent monolayers of human umbilical vein endothelial cells (HUVEC) has been studied for the induction of new membrane proteins. Two approaches have been taken. The first is a quantitative two-dimensional gel analysis of [35S]cysteine-labeled membrane proteins of HUVEC with and without cytokine treatment. This analysis indicates that there are a restricted number of new membrane proteins synthesized in the first 6 h of IL-1 treatment, on the order of 19 out of a total of over 600 detectable proteins. Second, we have prepared two mAb (1E7 and 2G7) to different epitopes of a major inducible sialoglycoprotein with molecular mass of 114 kDa and an isoelectric point of 4.6 to 4.8. These antibodies were compared with two additional antibodies, 3B7 and 7A9, which were shown to react with the endothelial leukocyte adhesion molecule-1 (ELAM-1) protein as expressed in COS cells. The 1E7/2G7 protein is distinct from ELAM-1, based upon biochemical comparisons as well as the inability of the 1E7 and 2G7 antibodies to react with ELAM-1-transfected COS cells. The protein defined as 1E7/2G7 is neither expressed constitutively nor in an inducible manner on PBMC, granulocytes, platelets, fibroblasts, or keratinocytes. The 7A9 and 3B7 antibodies are shown to block granulocyte binding to IL-1-activated HUVEC. The 2G7 antibody is effective at inhibiting the binding of T cells but not granulocytes to IL-1-activated endothelium, suggesting this new protein is an adhesion protein that may be active in vivo in T cell-endothelial cell adhesion-related events such as inflammation or lymphocyte recirculation. In addition, T cells were shown to utilize the ELAM-1 protein in binding to cytokine-activated HUVEC. Antibodies directed to both proteins had additive effects on inhibition of T cell adhesion.
Department of Immunology, Otsuka America Pharmaceutical, Inc., Rockville, MD 20850.
N
N
Graber
T V
TV
Gopal
D
D
Wilson
L D
LD
Beall
T
T
Polte
W
W
Newman
D000941:Antigens / Q000096:biosynthesis; D001667:Binding, Competitive; D002448:Cell Adhesion; D015815:Cell Adhesion Molecules / Q000032:analysis / Q000502:physiology*; D019040:E-Selectin; D004730:Endothelium, Vascular / Q000502:physiology*; D006801:Humans; D007249:Inflammation / Q000276:immunology; D007375:Interleukin-1 / Q000494:pharmacology*; D008565:Membrane Proteins / Q000032:analysis; D011233:Precipitin Tests; D012795:Sialoglycoproteins / Q000302:isolation & purification / Q000502:physiology*; D013601:T-Lymphocytes / Q000502:physiology*
D000941:Antigens; D015815:Cell Adhesion Molecules; D019040:E-Selectin; D007375:Interleukin-1; D008565:Membrane Proteins; D012795:Sialoglycoproteins