This proposal focuses on the biology of endothelial cells (EC), with the goal of understanding their differentiation in relation to inflammatory stimuli, and their role in supporting the adhesion and extravasation of leukocytes. Since our ability to study EC differentiation is currently limited by a paucity of differentiation markers, particularly in animal models, one important aim will be to use in vivo and in vitro EC isolates to generate monoclonal antibodies (MAbs) defining several classes of endothelial differentiation antigens (inflammation-associated, organ-specific, subset-specific, etc.---identified immunohistologically), whose biochemistry, regulation and (if possible) function will be characterized subsequently in vitro and/or in vivo. We have previously shown that EC in mucosal versus non-mucosal (e.g., lymph nodes) tissues express distinct organ-specific determinants for leukocyte recognition (OSDLR) that control the adhesion and tissue-specific extravasation of circulation lymphocytes, neutrophils, and probably other leukocytes. The following tools are available to study the expression of OSDLR by endothelium: lymphoid cells bearing receptors specific for lymph node or mucosal OSD; enzymatic, polysaccharide, and mono and polyclonal antibody inhibitors of specific leukocyte receptors for OSD; and simple in vitro assays of organ-specific leukocyte-EC interactions. These tools will be applied here to dissect the nature and regulation of OSDLR. The specialized high endothelial cells (HEC) in lymphoid organs express high levels of OSDLR. An in vitro rosetting assay of lymphocyte interaction with viable HEC will be used a) to assess the active role of EC in leukocyte adhesion; and b) to explore the chemistry and regulation of OSDLR by assessing their sensitivity to various enzymatic (particularly proteolytic) and chemical treatments, and by determining whether individual EC can display both mucosal and nodal specificities simultaneously. In addition, we will ask whether the adhesion of leukocytes to cultured human EC treated with IL-1 or tumor necrosis factor (which enhance EC adhesiveness is mediated by OSDLR; and whether these or other monokines, or epithelial-derived cytokines, regulate OSDLR expression. Most importantly, we will combine serologic approaches, functional assays and biochemical isolation techniques to identify, characterize and produce MAbs against OSDLR. Finally, we will explore the specificity of OSDLR in diverse lymphoid tissues in man. These studies are important to understand the biology of endothelium and its role in normal and abnormal inflammatory and immune processes; and the pathology of endothelium in, for example, the early leukocyte-mediated events in atherosclerosis.
Showing the most recent 10 out of 94 publications
