Inhibiting Monocyte Migration Reduces Arterial Thrombosis and Facilitates Thrombolysis

Abstract

BACKGROUND: Monocytes contribute to the initiation and propagation of venous thrombosis. Little is known about the roles monocytes play in arterial thrombosis, the cause of stroke and myocardial infarction. METHODS: We investigated how CCR2 (CC chemokine receptor 2) knockout ((-/-))-mediated monocyte deficiency affects platelet function, blood coagulation, thrombus volume, and thrombolytic susceptibility in 666 male mice with FeCl3-mediated carotid arterial thrombosis, including 365 C57BL/6 wild type (WT) mice, 295 CCR2(-/-) mice, and 6 CX3CR1-GFP (CX3C chemokine receptor 1-green fluorescent protein) mice. RESULTS: Intravital microscopy and flow cytometry showed that both neutrophils and monocytes were recruited to the acute arterial thrombus, as observed 30 minutes post-thrombosis. Platelet function tests demonstrated platelet aggregation to be lower in the whole blood of CCR2(-/-) mice (versus C57BL/6 WT mice) but not in their leukocyte-free platelet-rich plasma, suggesting this platelet dysfunction is cell-mediated. Flow cytometry experiments revealed lower numbers of monocyte-platelet aggregates in the blood of CCR2(-/-) mice, compared with C57BL/6 WT mice. Blood levels of FXIII (factor XIII) and monocyte levels of FXIII-A were increased after carotid thrombosis in C57BL/6 WT mice but not CCR2(-/-) mice. Further, in vivo micro-computed tomography-based thrombus imaging using fibrin-targeted gold nanoparticles and histology showed that CCR2(-/-) mice had smaller thrombi (0.112 +/- 0.002 mm(3), n=22) than C57BL/6 WT mice (0.125 +/- 0.007 mm(3), n=27; P<0.01), with increased porosity and reduced fibrin cross-linking. Moreover, tPA (tissue-type plasminogen activator) mediated thrombus volume reduction progressed up to approximate to 1 hour faster during the initial 3-hour period in CCR2(-/-) mice and CCR2-siRNA-treated mice, compared with C57BL/6 WT mice. In addition, clopidogrel reduced baseline thrombus volume more, but CCR2(-/-) better facilitated tPA-mediated thrombolysis. CONCLUSIONS: CCR2 antagonism decreases platelet aggregation and reduces FXIII levels in blood and monocytes, thus driving arterial thrombosis towards the generation of a relatively small, porous, more lysable clot. [GRAPHICS]