Real-Time, Non-Invasive Thrombus Characterization using Ultrasound Responsive Nanodroplets - ABSTRACT Thrombosis is a leading cause of morbidity and mortality worldwide, associated with several serious health complications including pulmonary hypertension, cardiovascular disease, and death. As thrombi mature, their structural and mechanical properties change, becoming mechanically less compliant due to fibrin crosslinking and gradual collagen deposition. The evolving mechanical properties of thrombi over time necessitates a dynamic and adaptable approach for timely diagnosis and treatment management. However, current diagnostic methods are primarily for detection of thrombi and fail to provide critical information about thrombus age and mechanical characteristics. This limitation highlights the importance of mechanistic understanding of histological (e.g., composition) and mechanical (e.g., elastic modulus) changes that occur as thrombi age, which is critical for refining treatment strategies tailored to thrombus development stage and optimizing patient outcomes. Due to the dynamic nature of thrombi and limitations of existing diagnostic methods, there is a critical need to develop a methodology for real-time quantification of evolving thrombus characteristics, such as age and elastic modulus. This will help understand the clot aging mechanism better and guide appropriate treatment selection. Our long- term goal is to develop an in situ micro-rheology technique that can non-invasively and spatiotemporally characterize aging thrombi in real-time using ultrasound responsive nanodroplets. These nanodroplets are designed to phase-transition into microbubbles when exposed to focused ultrasound above a threshold amplitude, a process known as acoustic droplet vaporization (ADV). While ADV has been utilized in several biomedical applications, such as drug delivery and super-resolution contrast imaging, its potential in micro- rheology remains unexplored. Ultrasound responsive nanodroplets can be administered intravenously and activated when they reach the thrombus site. These nanodroplets have several advantages, including their ability to form bubbles during ultrasound exposure and then rapidly recondense once the ultrasound is turned off. Additionally, their pharmacokinetic properties, such as longer circulation times due to their small size and liquid core, further enhance their effectiveness. ADV bubbles are highly sensitive to medium properties, enabling quantification of mechanical properties based on their acoustic emissions. The objective is to correlate the acoustic emissions of ADV bubbles with the mechanical properties of aging thrombi. The central hypothesis is that acoustic emissions of the ADV bubbles can be used to quantify thrombus age and elastic modulus. To test this hypothesis, we will pursue three specific aims. 1) Quantify the impact of medium properties on the acoustic emissions of ADV bubbles in thrombus-mimicking hydrogels. 2) Determine thrombus characteristics in an in vitro venous flow model. 3) Demonstrate in situ characterization of thrombus in an inferior vena cava model. The successful completion of the proposed research will be significant because it will establish a novel platform for non-invasive, real-time, and super-resolution characterization of aging thrombi.
