Improving arterial bleeding cessation by modulating thrombus formation
How to quickly stop arterial bleeding is one of the most important survival issues. Surprisingly, our current understanding of the platelet-mediated thrombus formation is primarily based on models in which the blood vessel wall remains intact and therefore little bleeding occurs. Hence, the existing models, developed to understand inward thrombus growth and arterial plaque formation (thrombosis), are of limited predictive value for the management of bleeding cessation (hemostasis). We propose that fresh insights need to be gained from a new animal model of arterial puncture in order to enhance hemostasis without potential systemic thrombotic events. In this regard, we have recently applied the wide-area electron microscopy to visualize the subcellular structure of entire thrombi after mouse jugular venous puncture wound. Interestingly, the plate-to-plate adhesion that forms an extravascular ‘cap’ appears to be a critical step in venous bleeding cessation. This finding, in contrast to the conventional core-and-shell model of thrombus formation in non-puncture models, opens up the possibility of an external, topical application of agents to modulate thrombus formation. However, venous bleeding does not closely model clinically relevant trauma because of the low pressures involved. Without the ultrastructural understanding of thrombus formation, current clinical practices include intravenous administration of tranexamic acid, a fibrinolysis inhibitor/clot stabilizer presently administered intravascularly in trauma centers. In this project, we will establish a true arterial puncture model in mice and evaluate efficacy of external pro-thrombotic agents in enhancing bleeding cessation.
Brief experimental plans are:
(1) Optimal surgical conditions for femoral arterial puncture will be established while monitoring bleeding time. An important variable to be determined is the size of the puncture needle.
(2) Whole thrombus before and after bleeding cessation will be imaged by correlative fluorescent microscopy and wide-area electron microscopy. Ultrastructure of the thrombus including platelet activation states and interaction will be analyzed at sub-platelet level.
(3) Using the puncture model established in (1), effects of external application of tranexamic acid on bleeding time will be compared to the intravenous application and vehicle control. Once efficacy of external application is established, embedding therapeutic agents into biodegradable material (e.g., collagen patches) will be explored.
This work represents a new collaboration of three faculty members with complementary expertise: Dr. Sung Rhee (vascular pharmacology), Dr. Jerry Ware (platelet physiology), and Dr. Avi Bhavaraju (trauma surgery). Additionally, Mr. Hunter Rose (medical student) will be trained in rodent surgery, imaging, and analysis.