Proteolytic Pathways in Venous Thrombus Resolution

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Background / Rationale: VTE (Venous Thromboembolism) is a common and serious cardiovascular disease with significant mortality and morbidity. Prompt anticoagulation of patients with deep venous thrombosis (DVT) reduces fatal pulmonary embolism (PE) but does not prevent long-term morbidity of VTE. Post-thrombotic syndrome occurs in 25-50% of patients with clots in the deep veins of the arms and legs that consists of pain, swelling, and recurrent skin ulceration. Common risk factors for DVT include cancer, major trauma, surgery, paralysis, prolonged periods of immobility, and older age. DVT and its complications have increased in the Veteran population over the last decade. Deployed military personnel are at an increased risk due to prolonged air and ground transport, dehydration, tobacco use, and extended immobility during hospitalizations for severe injuries. Thrombus resolution is a critical factor in the pathogenesis of post-thrombotic syndrome since incomplete thrombus resolution can result in obstruction of flow and loss of venous valve function. Using novel 3D serial measurements of thrombus volume; we demonstrated that DVT patients with similar initial DVTs have widely varying rates of thrombus resolution over time despite adequate anticoagulation. Longitudinal studies show that patients with more rapid thrombus resolution have a better prognosis than those patients whose thrombus resolves much slower. Despite its clinical importance, the cellular and molecular mechanisms involved in DVT are poorly understood, and there currently is no therapy to accelerate this process. Objectives: Using clinically relevant experimental models of DVT, a comprehensive picture of interconnected cell-mediated molecular processes that orchestrate a precise inflammatory program is starting to emerge and forms the foundation for this proposal. Our objectives are to build on our previous VA Merit findings to: 1) define mechanisms by which plasminogen activator inhibitor-2 (PAI-2) deficiency modulates inflammatory leukocytes to accelerate venous thrombus resolution; 2) test the therapeutic efficacy of blocking the PAI-2 pathway to accelerate venous thrombus resolution; and 3) determine specific gene signatures for the temporal inflammatory vascular remodeling events that occur during venous thrombus resolution and evaluate the prognostic value of candidate biomarkers in patients with evolving and maladaptive thrombus resolution after VTE. Methods: Studies will utilize genetically deficient mice in experimental models of DVT that accurately mimic many of the clinical and pathophysiological features observed in human DVT. We will define mechanisms by which PAI-2 deficiency calibrates immune regulation to accelerate venous thrombus resolution using ex vivo thrombolysis assays, transmigration assays, and neutrophil adoptive transfer experiments. The translational potential of suppressing PAI-2 expression or activity to accelerate venous thrombus resolution will be tested in human cells and in preclinical models. Finally, we will use our well-established clinically relevant mouse models of DVT resolution to identify gene regulatory signatures/potential biomarkers and then evaluate changes in gene networks over time in blood from patients with evolving thrombus resolution after VTE. Findings/Results: Molecular mechanisms that modulate inflammation during venous thrombus resolution in experimental models and in human patients will be identified and therapies based on these mechanisms tested in preclinical models. Status: This is a new project arising from substantial supportive preliminary data from a previous VA Merit Award. Impact: New knowledge from these studies regarding the inflammatory signatures in DVT patients may form the basis for novel therapies for accelerating this process, and in combination with anticoagulants, control excessive fibrosis, and prevent this disease.