Macrophage accumulation is one of the earliest events in atherosclerosis. Macrophages are derived from blood-borne monocytes that adhere to endothelium and transmigrate across the endothelium. In flowing blood monocytes are transported along streamlines, Contact is more likely at regions where curvature brings streamlines close to the vessel wall and flow reversal first occurs. In order for cells to adhere and activate signaling pathways that promote transmigration, the adhesive force arising from bond formation must resist the hydrodynamic force causing cells to detach. Firm adhesion in regions of flow reversal must still occur rapidly in order to resist higher shears tresses arising during systole. Recently we observed that adhesion of monocytic cells is enhanced by forces acting normal to the surface between the monocyte and endothelium. Bond formation increased leading to more stable adhesion. Normal forces arising from secondary flows may thus play an important role in creating adhesive forces able to resist detachment of monocytes from endothelium in the relatively high shear stress environment found in the arteries. These normal forces may be further enhanced by red cell interactions with monocytes about to contact or adhering to endothelium. Cell rolling and arrest involves bond and tether formation. The objective of the proposed research is to test the hypothesis that normal forces arising in regions of flow recirculation enhance monocyte adhesion by compression of the microvitli and increased bond formation between monocytes and endothelium. A combination of micropipette aspiration experiments, flow chamber adhesion measurements and total internal reflection fluorescence microscopy (TIRF) will be used to test this hypothesis in vitro through the following specific aims: (1) characterize the viscoelastic behavior of monocytic cells, endothelium and tether formation (2) Investigate the effect of normal forces on the biophysics of bond and tether formation; (3) characterize the receptors involved in adhesion mediated by normal forces;(4) Determine the effect of normal; and shear forces on the contact area of rnonocytes; and (5) Examine the role of normal forces in enhanced monocyte adhesion near flow reattachment. Experimental studies will be aided by application of theoretical studies to model the adhesion of monocytes to endothelium. Results from this research will provide new information about the mechanism by which monocytes adhere to arterial endothelium in atherosclerosis-prone regions.
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