Date of Award
Doctor of Philosophy (PhD)
Chemical and Biomolecular Engineering
Scott L. Diamond
Platelets and coagulation proteins work in concert to maintain proper blood flow through the vasculature. When an injury occurs, this hemostatic system must respond efficiently by sealing the wound to prevent excessive bleeding. Deviations from normal hemostasis arise in the clinic frequently; one such condition that is characterized by uncontrolled bleeding is known as trauma-induced coagulopathy (TIC). Severe platelet dysfunction is one key contributing factor to TIC, though its mechanistic causes are still yet to be fully understood. In order to investigate biological explanations for platelet dysfunction during trauma, various cell-based assays were designed and conducted in both healthy and patient populations. Specifically, intracellular calcium mobilization and other fluorescently tracked biomarkers were used as dynamic indicators of platelet activation in response to common agonists. Microtiter well plates prepared with liquid handling systems enabled high-throughput data collection and minimal manual pipetting. Significant platelet dysfunction in response to 31 unique stimulation conditions spanning several signaling pathways was observed in a cohort of trauma patients and tracked at multiple timepoints after initial hospital admission. In experiments designed to interrogate plasma effects on healthy platelet function, patient-derived plasma imparted significant inhibition which implied the presence of a unique soluble plasma species with downregulatory effects on endogenous and transfused platelets. With established knowledge of coinciding coagulant and lytic states during trauma, strategic addition of agonists to healthy platelet suspensions led to generation of soluble fibrin species and desensitization to agonist stimulation through glycoprotein VI (GPVI). Downstream platelet dysfunction was only observed when thrombin was added to the system to polymerize fibrin, whereas stimulation with other agonists or inhibition of various stages of coagulation had no effect on subsequent GPVI function. Maximal inhibition (~95%) was attained when tissue plasminogen activator (tPA) was also incorporated to lyse fibrin polymers into fibrin degradation products (FDP). Concentrations of a small FDP called D-dimer were elevated in trauma patient samples and inversely correlated with a quantitative measure of platelet function. Finally, preliminary results indicate potential binding affinity between platelet receptors and D-dimer. These results shed light on specific biological entities that may be responsible for platelet dysfunction in trauma patients.
Verni, Christopher, "Role Of Soluble Fibrin And Fibrin Degradation Products On Platelet Signaling During Trauma" (2020). Publicly Accessible Penn Dissertations. 4176.