Anti-ovine VCAM-1 monoclonal antibodies inhibit adhesion and proliferation between sheep endothelial and mononuclear cells in vitro

This paper reports the production and characterization of three monoclonal antibodies (mAb) recognizing ovine vascular cell adhesion molecule-1 (VCAM-1). The mAb were raised against sheep umbilical vein endothelial cells (ShUVEC) and flow cytometric analysis demonstrated that one mAb, QE4G9, was crossreactive with human VCAM-1 expressed on Chinese hamster ovary cell transfectants. Protein modulation studies on ShUVEC and immunoperoxidase staining of inflamed renal tissue further indicated the reactivity of the other two ovine mAb, QE1F3 and QE2G4, with ovine VCAM-1. The flow cytometric profile of the three mAb on stimulated ShUVEC was identical to that observed with the cross-reactive anti-human VCAM-1 mAb, HAE2-1. Peak expression occurred between 6-12 h after stimulation, followed by a slight decrease to a plateau extending beyond 48 h. In functional assays, all mAb inhibited adhesion of PMA-activated sheep PBMC to stimulated ShUVEC. In addition, QE4G9 inhibited proliferation of sheep PBMC in the mixed lymphocyte-endothelial cell reaction (MLER) by 56%. The results demonstrate that the three anti-ovine VCAM-1 mAb recognize functional epitopes on sheep vascular endothelial cells. These mAb will be valuable tools in the investigation of VCAM-1 expression in various pathophysiological conditions using sheep models, and in the study of VCAM-1-mediated leucocyte-endothelial cell interactions, both in vitro and in vivo.