Two of the earliest events in atherogenesis are the accumulation of low density lipoprotein (LDL) and macrophages in the arterial intima. Firm adhesion of monocytes to endothelium is necessary for transmigration of monocytes into the vessel wall. Monocyte adhesion depends upon the level of expression of adhesion proteins by endothelial cells and monocytes as well as the fluid forces acting on monocytes. Localization of early lesions to vessel branches may result from a hemodynamic environment which does not cause cell detachment and alters endothelial cell function to promote monocyte adhesion. The applicants will test the following two hypotheses: 1) arterial fluid dynamics and local oxidation of LDL activate the endothelium to express receptors for monocytes, and 2) localization of monocyte adhesion to vessel branch points represents a balance between hydrodynamic and adhesive interactions. Hydrodynamic interactions include cell transport to the endothelium and drag forces and torque acting on the attached or rolling cell. Objectives of the proposed research are to 1) identify conditions which promote monocyte rolling and arrest on endothelial cells at shear stresses encountered in arteries; 2) characterize the kinetics of these receptor-ligand interactions; and 3) determine the effect of the flow field upon localization of monocyte attachment.
Specific aims are to 1) characterize the biophysics of monocyte interactions with endothelial cells activated with minimally modified LDL (mm LDL); 2) determine the effect of the flow field upon monocyte interactions with endothelial cells; and 3) examine monocyte adhesion to and rolling on activated endothelial cells in a region of flow recirculation with and without red blood cells. Experiments will be performed with U937 cells and human umbilical vein endothelial cells. Endothelial cells will be activated by mm LDL. Flow chamber studies will be used to study rolling and adhesion under conditions which simulate shear stresses and flows in the arterial system. Micropipet measurements will be performed to quantify bond lifetimes and rate constants for binding and dissociation. The applicants will use a sudden expansion flow chamber to create a region of flow recirculation. They will examine the effect of flow recirculation on the expression of adhesion molecules by endothelium and the transport and adhesion of monocytes to endothelium. These studies will provide important quantitative data on monocyte adhesion to endothelium and permit the investigators to evaluate how monocytes can adhere to activated endothelium in the high stress environment of the arterial system.
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