A Microfluidic Approach For Investigating Thrombosis And Hemostasis: From Disease Models To Drug Testing
Medicine and Health Sciences
In vitro studies of cardiovascular biology have often relied on static assays using plasma or isolated blood cells in the past. Microfluidic technologies offer a unique opportunity to study the mechanisms of hemostasis and thrombosis under physiologically relevant conditions. This thesis describes a microfluidic approach to model cardiovascular diseases and evaluate therapeutic candidates. In our studies, we recreated sterile occlusive thrombosis under a wide range of pressure drops and investigated the effect of hemodynamic forces on neutrophil activities during clot formation. We discovered that high interstitial hemodynamic forces (> 70 mmHg/mm-clot) can drive physically entrapped neutrophils to rapidly form neutrophil extracellular trap (NET) during sterile occlusive thrombosis, offering an explanation for the rapid neutrophil DNA release in mouse ferric chloride carotid artery injury models and the extracellular nucleosomes detected in thrombi/plasma obtained from patients with stroke, myocardial infarction, and disseminated intravascular coagulation. In addition to modeling occlusive thrombus formation, we also characterized clotting profiles of patient populations and recreated disease phenotypes. Through modifying our microfluidic clotting assay, we reduced the risk of contact activation during sample preparation and increased the dynamic range for drug testing. Hemophilia assays phenocopying the bleeding disorder was developed using healthy adult blood treated with a low-activity factor variant or neutralizing antibodies. The assays could enable rapid screening of novel hemophilic agents in a high-throughput fashion. We also used our microfluidic clotting assays to measure clotting rates of neonatal patients with congenital heart disease, and we found reduced platelet deposition and fibrin generation ex vivo for neonates compared to healthy adults. Neonatal patients also displayed heightened sensitivity to antithrombotic drugs. The majority of the patients who underwent surgical operations had increased platelet deposition 24 hours after surgery whereas the trend for fibrin polymerization was less clear. The significant increase in platelet response demonstrated the need for postoperative pharmacological protections. Our results in a phase I clinical trial showed that cangrelor is an excellent candidate for preventing shunt thrombosis in neonatal patients after surgical operations. Using microfluidic devices, we demonstrated that thrombosis-and-hemostasis-on-a-chip could help us understand the mechanisms of cardiovascular biology and evaluate therapeutic candidates.