PB1056 - In situ Pulmonary Thrombosis is Regulated by Integrin αIIbβ3-dependent Platelet Procoagulant Activity

Background: In situ pulmonary thrombosis (ISPT) is a major complication associated with a high morbidity. Surprisingly, there are no direct in vivo studies that characterize the pathogenesis of ISPT. Recent clinical studies suggest that platelet activation, which increases platelet procoagulant activity, may contribute to the development of ISPT. However, the exact pathophysiology of ISPT is not yet fully understood. Improved discernment of the underlying mechanisms would enable the development of new therapies for ISPT.

Aims: To investigate the role of αIIbβ3-dependent platelet procoagulant activity in the pathophysiology of ISPT.

Methods: Quantitative fluorescence intravital fluorescence lung microscopy (qFILM) was used to visualize ISPT in C57BL/6 (WT) mice following intravascular (IV) thromboplastin administration in the presence or absence of heparin or eptifibatide. Fluorochrome-conjugated anti-mouse CD49b Ab and dextran were administered IV for in vivo staining of circulating platelets and visualization of blood vessels, respectively. ISPT was defined as the occlusion of blood vessels with platelet-rich aggregates, resulting in pulmonary ischemia. Additionally, modified prothrombin time assay (mPTA) was used to investigate the effect of platelet αIIbβ3 activation/inhibition on mouse and human platelet rich plasma (PRP) clotting.

Results: TF triggered dose-dependent ISPT in mice in vivo, which involved development of platelet-rich thrombi in the pulmonary arteriolar bottlenecks (junction of pulmonary arteriole and capillaries), resulting in a transient ischemia in the arteriole and the down-stream capillary tree. The ISPT triggered by IV TF was completely abrogated following IV administration of heparin, and remarkably, also by αIIbβ3 receptor inhibitor (eptifibatide). In addition, mPTA study further confirmed that inhibition of platelet αIIbβ3 significantly extended both human and mouse PRP clotting times.

Conclusion(s): Our current findings strongly suggest that αIIbβ3-dependent platelet procoagulant activity promotes ISPT. Both our models have potential application in investigating the molecular determinants of pulmonary thrombosis in diverse pulmonary disorders as well as evaluating efficacy of new antithrombotic drugs.