Abstract 1655: Biological Thrombus Age Predicts Fibrinolytic Efficacy In Vivo: Assessment by Multiwavelength Fluorescence Molecular Imaging of Activated Factor XIII Activity and Fibrin
Background: Acute thrombi possess higher levels of activated factor XIII (FXIIIa) and lyse more efficiently than aged thrombi. To assess thrombus age and its impact on fibrinolysis in vivo, we developed a molecular imaging strategy to concomitantly assess FXIIIa activity, fibrin formation, and fibrinolysis.
Methods: Near-infrared fluorescent nanoparticles (NP, CLIO, MGH-CMIR) derivatized with targeting peptides (TP) were targeted to FXIIIa (F13TP-NP, ex/em 670/700nm, F13TP = GNQEQVSPLTLLWC) and fibrin (fibrinTP-NP, ex/em 750/800nm, fibrinTP = GPRP-PGGSKGC). NP agents were evaluated in human plasma clots. In C57Bl6 mice (n = 10), FeCl3-induced femoral thrombi (0.5h/acute or 18h/subacute) were imaged using multiwave-length intravital fluorescence microscopy (IVFM, resolution 13x13x10 μm). Mice were injected with 10 mg Fe/kg F13TP-NP and fibrinTP-NP (n = 8) or control untargeted NP (n = 2). IVFM 3D stacks obtained over 0.5h were analyzed for target-to-background ratios (TBR, ImageJ). Fibrinolysis (n = 6) was initiated for 1h with plasminogen (100mg/kg bolus), heparin (200 U/kg bolus, 50 U/kg/hr infusion) and tPA (1mg/kg bolus, 9 mg/kg/hr infusion). Serial IVFM 3D stacks were acquired. The change in fibrin content (Δfibrin) was measured as: ((fibrin signal at 1 hour/fibrin signal baseline)-1)*100%).
Results: F13TP-NP and fibrinTP-NP strongly targeted human clots (p < 0.0001 vs. control agents). High FXIIIa TBRs were achieved in acute thrombi (TBR 6.8 = 2.2, p < 0.002 vs. subacute thrombi). In acute thrombi undergoing fibrinolysis, the IVFM-derived Δfibrin decreased (−40.7 ± 12.5%) indicating net fibrinolysis. In contrast, the Δfibrin increased in acute thrombi not undergoing fibrinolysis (137 ± 6.3%, p < 0.001), as well as in subacute thrombi undergoing fibrinolysis (29 ± 6.3%, p < 0.001), indicating net increased fibrinTP-NP binding to thrombi over time. The FXIIIa TBR prior to fibrinolysis correlated strongly with Δfibrin (R2 = 0.98, p < 0.01)
Conclusions: Molecular imaging detection of acute thrombi predicts fibrinolytic efficacy and allows molecular assessment of fibrinolysis in vivo. Clinical translation of biological thrombosis imaging methods could guide the optimal use of plasminogen activators in thrombosis syndromes.
This research has received full or partial funding support from the American Heart Association, AHA National Center.