Adhesion of the blood-borne monocyte to the endothelium is the initial event in the extravasation of this leukocyte into the vessel wall or the interstitial space. The long term objectives of this proposal are (1) to characterize biochemically the cell surface molecules on the endothelium which are involved in monocyte-endothelial cell (EC) attachment, and (2) to determine the mechanism of focal adhesion of monocytes to the vessel wall in response to hypercholesterolemia. These objectives will be pursued principally by a series of in vitro experiments utilizing a model system which involves cultured EC and the human monocytic tumor cell line U937. Experiments will be repeated where appropriate with purified blood monocytes. Our working hypothesis is that EC, in response to certain physiological stimuli, express specific cell surface receptors or binding sites for monocytes. Further, we hypothesize that migrating and/or proliferating EC express an increased number of monocyte binding sites and that this phenomenon is responsible for the focality of monocyte binding in vivo. Finally, we hypothesize that by blocking the initial adhesion of monocytes to the endothelium during the initial stages of hypercholesterolemia that subsequent events, such as fatty streak development, will not occur. Our hypotheses will be tested by pursuit of the following specific aims: 1) To characterize and isolate the binding site(s) for monocytes on the EC surface.
This aim i ncludes studies on a) the protease, glycosidase and lectin sensitivity of monocyte-EC attachment and b) studies on the detergent solubilization and purification of the EC surface molecule(s) involved in the cell-cell adhesion. 2) To prepare serum antibodies and/or monoclonal antibodies to EC surface moieties involved in monocyte-EC attachment. These antibodies will be useful in purification, in the localization and quantification of monocyte binding sites in vivo, and in the blocking of monocyte attachment to the endothelium in vivo (Aim 5). 3) To identify culture conditions or exogenous agents which regulate the expression of monocyte binding sites on the EC surface. We will examine the role of a) time in culture, b) EC migration vs. proliferation, c) EC """"""""injury"""""""" agents and d) hypercholesterolemic vs. normal serum, in the ability of EC to bind monocytes. 4) To compare monocyte attachment to EC from hypercholesterolemic vs. normal animals and to compare monocyte binding to cultured EC from large vessels and small vessels. 5) To inhibit monocyte attachment to the pig or rabbit aorta in vivo by the infusion of antibodies against the monocyte binding site on cultured EC. By gaining mechanistic knowledge on the molecular level of this single event in monocyte-vessel wall interactions, we may be one step closer to preventing pathological phenomena that require monocyte infiltration.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Modified Research Career Development Award (K04)
Project #
1K04HL001561-01
Application #
3073871
Study Section
Pathology A Study Section (PTHA)
Project Start
1985-07-01
Project End
1990-06-30
Budget Start
1985-07-01
Budget End
1986-06-30
Support Year
1
Fiscal Year
1985
Total Cost
Indirect Cost
Name
Cleveland Clinic Lerner
Department
Type
DUNS #
017730458
City
Cleveland
State
OH
Country
United States
Zip Code
44195
Faruqi, R; de la Motte, C; DiCorleto, P E (1994) Alpha-tocopherol inhibits agonist-induced monocytic cell adhesion to cultured human endothelial cells. J Clin Invest 94:592-600
Shankar, R; de la Motte, C A; DiCorleto, P E (1992) Thrombin stimulates PDGF production and monocyte adhesion through distinct intracellular pathways in human endothelial cells. Am J Physiol 262:C199-206
Shankar, R; de la Motte, C A; DiCorleto, P E (1992) 3-Deazaadenosine inhibits thrombin-stimulated platelet-derived growth factor production and endothelial-leukocyte adhesion molecule-1-mediated monocytic cell adhesion in human aortic endothelial cells. J Biol Chem 267:9376-82
Shen, X Y; Hamilton, T A; DiCorleto, P E (1990) Thrombin-induced expression of the KC gene in cultured aortic endothelial cells. Involvement of proteolytic activity and protein kinase C. Biochim Biophys Acta 1049:145-50
DiCorleto, P E; de la Motte, C A (1989) Thrombin causes increased monocytic-cell adhesion to endothelial cells through a protein kinase C-dependent pathway. Biochem J 264:71-7
Tannenbaum, C S; Major, J; Poptic, E et al. (1989) Lipopolysaccharide-inducible macrophage early genes are induced in Balb/c 3T3 cells by platelet-derived growth factor. J Biol Chem 264:4052-7
DiCorleto, P E; de la Motte, C A (1989) Role of cell surface carbohydrate moieties in monocytic cell adhesion to endothelium in vitro. J Immunol 143:3666-72
Shen, X Y; Hamilton, T A; DiCorleto, P E (1989) Lipopolysaccharide-induced expression of the competence gene KC in vascular endothelial cells is mediated through protein kinase C. J Cell Physiol 140:44-51
Fox, P L; DiCorleto, P E (1988) Fish oils inhibit endothelial cell production of platelet-derived growth factor-like protein. Science 241:453-6
Uratsuji, Y; DiCorleto, P E (1988) Growth-dependent subcellular redistribution of protein kinase C in cultured porcine aortic endothelial cells. J Cell Physiol 136:431-8

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