OC 37.4 - Plasminogen retention is attenuated in thrombi from Plg-RKT deficient mice

Background: Human and murine platelets express the transmembrane lysine-dependent plasminogen receptor, Plg-RKT. This receptor functions to retain platelet-derived plasminogen on the activated platelet surface promoting plasmin generation.

Aims: To define the importance of Plg-RKT in retention of plasminogen within the thrombus microenvironment.

Methods: Blood was collected from wild type (WT) or Plg-RKT-/- deficient mice. Thrombi were formed ex vivo by flowing blood over collagen and tissue factor coated microfluidic chambers at 250 – 1000 s-1. Dylight633 (DL633)-labelled plasminogen and AlexaFluor488-fibrinogen were incorporated ± tissue plasminogen activator (tPA). Thrombus formation was imaged using a fluorescence microscope. Thrombosis was induced in the carotid artery using 30% FeCl3 with prior infusion of AlexaFluor647-labelled fibrinogen and DyLight488-GP1bβ antibody to label platelets. After 5 min tPA was infused. Thrombus formation was imaged for 30 min in real-time using a Zeiss Examiner upright intravital microscope equipped with spinning disk confocal.

Results: Shear stress has an impact on plasminogen accumulation in thrombi formed ex vivo, being more abundant at high shear (1000 s-1). Plasminogen accumulation in thrombi formed ex vivo was attenuated in Plg-RKT-/- deficient mice compared to WT mice. In a model of FeCl3 induced thrombi initial fibrinogen accumulation was similar in Plg-RKT-/- and WT mice. However, post tPA infusion, fibrin(ogen) accumulation in Plg-RKT-/- mice was increased compared to WT. This indicates that in Plg-RKT-/- mice the fibrinolytic potential was reduced tipping the balance in favour of fibrin formation. Interestingly, initial platelet accumulation was higher in Plg-RKT-/- mice compared to WT.

Conclusion(s): These data indicate that under physiological flow conditions Plg-RKT functions to incorporate and retain plasminogen into the growing thrombus. A novel in vivo thrombolysis model revealed that deficiency in Plg-RKT impacts on initial platelet accumulation and fibrin deposition. These data demonstrate the functional importance of Plg-RKT in localising plasminogen within thrombi which in turn impacts thrombus persistence in vivo.