Thromboxane prostanoid receptor stimulation induces shedding of the transmembrane chemokine [CX.sub.3]CL1 yet enhances [CX.sub.3]CL1-dependent leukocyte adhesion

In atherosclerosis, chemokines recruit circulating mononuclear leukocytes to the vascular wall. A key factor is [CX.sub.3]CL1, a chemokine with soluble and transmembrane species that acts as both a chemoattractant and an adhesion molecule. Thromboxane [A.sub.2] and its receptor, TP, are also critical to atherogenesis by promoting vascular inflammation and consequent leukocyte recruitment. We examined the effects of TP stimulation on processing and function of [CX.sub.3]CL1, using [CX.sub.3]CL1-expressing human ECV-304 cells and primary human vascular endothelial cells. TP agonists promoted rapid shedding of cell surface [CX.sub.3]CL1, which was inhibited by pharmacological inhibitors or specific small interfering RNA targeting tumor necrosis factor-[alpha]-converting enzyme (TACE). Because it reduced cell surface [CX.sub.3]CL1, we predicted that TP stimulation would inhibit adhesion of leukocytes expressing the [CX.sub.3]CL1 cognate receptor but, paradoxically, saw enhanced adhesion. We questioned whether the enhanced ability of the remaining membrane-associated [CX.sub.3]CL1 to bind targets was caused by changes in its lateral mobility. Using fluorescence recovery after photobleaching, we found that plasmalemmal [CX.sub.3]CL1 was initially tethered but ultimately mobilized by TP agonists. TP stimulation provoked clustering of transmembrane [CX.sub.3]CL1 at sites of contact with adherent leukocytes. These data demonstrate that TP stimulation induces two distinct effects: a rapid cleavage of surface [CX.sub.3]CL1, thereby releasing the soluble chemoattractant, plus mobilization of the remaining transmembrane [CX.sub.3]CL1 to enhance the avidity of interactions with adherent leukocytes. The dual effect of TP allows [CX.sub.3]CL1 to recruit leukocytes to sites of vascular inflammation while enhancing their adhesion once recruited. leukocytes; inflammation; endothelial cells doi: 10.1152/ajpcell.00380.2009.