Monocyte binding to the endothelium is a requisite first step in the migration of this leukocyte from the circulation to tissue during atherogenesis, inflammation and wound healing. The overall objective of this research project is to better understand the monocyte/endothelium interaction as it relates to atherosclerotic plaque development and to define the intracellular signalling pathways and agonist-induced genes that are involved in the regulation of monocyte binding to endothelial cells (EC). Our working hypothesis is that monocyte adhesion to EC is stimulated either by the migration and proliferation (turnover) of EC or by cytokines and injury-associated modulators, such as thrombin, through a cascade of specific intracellular events. This process is presumed to involved the activation of second messenger systems and the induction of multiple """"""""early response"""""""" genes. Our approach to understanding the regulation of monocyte binding to the endothelium involves several independent tacts as reflected in the following five specific aims: 1 To clone from a human aortic EC library lipopolysaccharide (LPS)- inducible genes that are potentially involved in the activation of EC and in the expression of cell surface binding sites for monocytes. Sequence data will be obtained to establish their identity or uniqueness; 2) To employ two distinct approaches, anti-sense DNA and antibody microinjection, to test the involvement of the above cloned genes in the induction of monocyte adhesion to EC; 3) To characterize further the intracellular signalling pathways (e.g., the Na+/H+ antiporter, calcium fluxes, and guanine nucleotide binding proteins) that may be involved in thrombin- induced monocyte adhesion to EC; 4) To determine whether other agonists of EC function such as bradykinin, serotonin, and histamine are capable of stimulating monocyte adhesion to EC and whether the same second messenger systems invoked by thrombin are employed in the expression of the response; 5) To determine whether EC derived from human vessels with very different propensity for development of atherosclerosis (aorta, internal mammary artery, and saphenous vein) differ in their basal or stimulated level of monocyte adhesion. We plan to relate our findings to the pathophysiology of atherosclerotic plaque development and would hope to identify means of arresting the atherogenic process at the point of monocyte involvement.
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