With cardiovascular disease being the leading cause of death worldwide, new and improved disease models are required to facilitate the research and production of new treatments. In this research, we are developing improved methods for modeling hypertension in engineered heart tissues (EHTs) to investigate resulting tissue remodeling at the tissue and cellular levels. We developed a model of hypertension using 3D-printed polylactic acid braces that enable stiffness adjustment of the flexible polydimethylsiloxane (PDMS) EHT platform. The braces were validated by mechanical testing to quantify their stiffening effect. Braces were shown to increase stiffness according to beam bending theory, with bracing half of the post’s length resulting in a 7-fold increase in stiffness. Next, we applied braces to tissues that only contain stromal cells, which are responsible for remodeling the extracellular matrix (ECM). The next steps are to quantify how stiffness affects ECM remodeling, including tissue-wide effects such as changes in tissue length and width, and micro-scale effects such as changes in cell migration, apoptosis, and cytoskeletal structure, which are quantified in 3D using IHC and confocal microscopy. We hypothesize that hypertension results in tissue thinning and lengthening, as well as decreased cell density, increased apoptosis, and increased expression of cytoskeletal markers. Next, the braces are applied to EHTs containing human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). The effect of hypertension on EHT remodeling and hiPSC-CM maturation is then quantified using a custom MATLAB image processing suite. Finally, increasing systolic resistance of novel varieties of EHTs that incorporate hiPSC-derived cardiac fibroblasts (hiPSC-CFs) alongside cardiomyocytes enables evaluation of fibrotic remodeling. We hypothesize that hypertension promotes a hypertrophic response in both hiPSC-CMs and hiPSC-CFs in EHTs, displaying increased sarcomerization and fibrosis respectively. Developing this improved hypertension model will accelerate cardiac regenerative medicine research, and provide new approaches for drug discovery.

Melusin, a chaperone protein expressed in cardiac tissue, induces a protective hypertrophic response in response to chronic mechanical stress. This protective hypertrophic response prevents the progression of cardiomyopathy into heart failure. In previous work done in wild-type (WT) and melusin knockout (melKO) mice, the absence of melusin was correlated with a hypertrophic response indicative of heart failure. I plan to investigate the biomechanical role of melusin in humans using human-engineered heart tissues (EHTs) created from human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) that lack melusin and their isogenic controls. EHTs are more representative of the human heart, making them an ideal model for studying the role of melusin in humans. I hypothesize that WT EHTs subjected to mechanical stress, i.e., high afterload, will outperform the melKO EHTs. To measure this, I will increase the stiffness of the EHT posts and measure contractile force. I have successfully differentiated high-purity WT and melKO cardiomyocytes from iPSCs, essential for creating healthy EHTs. I will cast both WT and melKO tissues on a bed of silicone posts that can be stiffened to varying extents to induce different amounts of mechanical stress on the cells. I will conduct a western blot on EHTs from all treatment groups to determine the level of melusin expression and examine the expression of Heat Shock Proteins (Hsp) 70 and 90, due to their coregulation with melusin. The EHTs that undergo mechanical stress are expected to express melusin and these results will work to establish whether melusin expression in humans is activated by mechanical stress. I will measure and compare the contractile force between the WT and melKO tissues. Improving our understanding of the role of melusin in humans can lead to further research into therapies and treatments for heart failure.

Over the last decade, the electrohydrodynamic (EHD) interaction produced by dielectric barrier discharge (DBD) actuators has seen strong interest for active flow control applications. Plasma synthetic-jet actuators (PSJAs) are DBD jets capable of producing zero-net mass flux momentum injection through ionic interactions in air at atmospheric pressure. These devices are of particular interest due to their rapid response time, lack of moving parts, variety of tunable parameters, and repeatably demonstrated efficacy in boundary layer modulation under a range of flow conditions. Due to the edge-normal momentum injection characteristics of PSJAs, previous studies naturally focused on surface wall jet configurations with spanwise cartesian geometries (see straight-edge, serrated, fingered electrodes) in quiescent, co-, counter-, and cross flow external conditions. We present an alternative 2-dimensional polar axisymmetric geometry; a circular ground electrode that is concentric and equal in diameter to the inner edge of an active high-voltage electrode ring. Steady-state inward injection develops an inward impinging jet and a resulting wall-normal net momentum directed away from the surface. A thorough electro-mechanical characterization led to the development of an empirical model of axisymmetric wall-normal plasma synthetic-jet actuators. The actuators utilize 0.0625” quartz glass as the dielectric material and have critical diameters of Dg = 40 mm, 60 mm, and 80 mm with an active electrode ring thickness of 10mm. The actuators are tested over an AC current operating range of Vpp = 8 – 60kV and f = 0.5 – 8 kHz. Direct thrust measurements are taken at all electrical conditions and intermittently validated through pitot-tube flow-profile tests. This provides rapid insight into the voltage-frequency-diameter-thrust relationships for axisymmetric PSJAs as well as finer intricacies, including entrainment patterns, high-Reynolds eddy formation, and jet dissipation. Combined with electrical current analysis, the axisymmetric PSJA is found to be an effective method of producing wall-normal momentum-injection. This research will directly contribute to the furthering of electric propulsion and flow control systems, opening the doors to more capable, more efficient, and greener flight technology.

Thin film photovoltaics (PV) present many advantages which enable their potential replacement of traditional silicon PV, but they are not yet scalable. Current research aims to enable their large-scale production to meet growing energy demands. Alongside scalable manufacturing methods and competitive device lifetimes, the transition from small-area cells to large-area modules poses a scalability barrier. I explore a novel fabrication method accelerating this transition. In fabricating thin film PV modules, large-area films are divided into multiple small-area cells by removing, or “scribing,” thin films between adjacent cells for later electrical connection. Existing scribing methods include laser and mechanical scribing, but both have drawbacks which impede device performance and scalability. My research professor and I invented a third scribing method to address these issues. Lines of solvent are printed onto a thin film device to dissolve target thin films underneath the solvent. As the solution evaporates, the “coffee-stain effect” redistributes the dissolved material to the fluid perimeter, exposing linear areas of underlying thin films. This effect is characteristic of drying liquids containing dispersed solids: liquid from the interior flows to restore liquid evaporating at the edge, carrying nearly all dispersed material to the fluid perimeter. This effect enables thin film scribing without forming performance-inhibiting film defects or toxic residues. The technology also requires low capital expenditure and is compatible with scalable roll-to-roll manufacturing. In this work, I demonstrate the invention’s feasibility and competitiveness by producing scribes comparable with existing technologies using electrohydrodynamic inkjet (EHDIJ) printing of solvent on perovskite solar cells. I authored a report, filed a provisional patent application, and now collaborate with another university to advance the technology. This invention poses a scalable solution to the transition from small-area cells to large-area modules in the thin film PV space, breaking a pivotal barrier to meeting growing energy demands.

Over the past 50 years, breast cancer mortality rates in high-income countries have dropped significantly while low and middle-income countries (LMICs) have made few advancements towards early detection and rapid diagnosis. A lack of infrastructure in LMIC healthcare settings causes patients to receive their diagnosis several months after testing, while some never hear back due to travel and communication barriers. We have partnered with Peace and Love Hospitals in Ghana to develop CoreView Imaging on the Needle (ION), a portable, rapid point-of-care (POC) diagnostic system for use in LMIC. Our team developed ION after discovering breast core needle biopsies (CNBs) were too delicate and sticky for transport in our original millifluidic design. Our research is exploring methods of integrating the ION system into CoreView to improve the testing reliability and image quality of biopsies for diagnosis by designing, testing, and iterating fixtures to image CNBs while still on the needle. We modeled our prototypes in Solidworks and printed them using a Prusa 3D printer. Our goal is to load the CNB under a microscope, stain the tissue nuclei, compress its surface against a clear glass window, and image the biopsy surface within 5 minutes. We are using Microscopy with UV Surface Excitation (MUSE) imaging to take panoramic images along the 20 mm biopsy length. The current prototype takes 1 minute to prepare the biopsy (stain and load into ION fixture), 0.5 minutes for each MUSE image and step to the next image (total of 6 images), and 0.5 minutes to unload the biopsy, resulting in 4.5 minutes between biopsy collection and obtaining images for diagnosis. The improved image quality and expected high reliability verifies the success of our ION design for further validation testing to provide earlier breast cancer detection and rapid treatment to rural patients.

Antiretroviral therapy (ART) prevents the progression of human immunodeficiency virus (HIV) by suppressing viral load, limiting transmission. Of the ~20 million people receiving ART, 30-40% do not maintain adequate medication adherence, resulting in treatment failure and drug resistance. Monitoring HIV medication adherence improves the efficacy of ART but requires bulky and expensive instruments that are not widely accessible at the point-of-need (e.g. doctor’s office or patient’s home), so a rapid and accessible diagnostic alternative is necessary. Our group developed the REverSe TRanscrIptase Chain Termination (RESTRICT) enzymatic assay to provide rapid and inexpensive measurement of HIV drug adherence by measuring antiretroviral drug activity indicated by fluorescence. However, one current limitation of RESTRICT is the need for trained operators to complete multiple precisely timed steps required in the enzymatic activity assay. We aim to create a 3D-printed microfluidic device that will automate the liquid handling steps required for RESTRICT via precisely tuned capillary action for rapid and user-friendly measurement of antiretroviral drugs. To that end, we first demonstrated a proof of concept by creating a microchip with a controlled 15 minute liquid delivery time and consistent liquid extraction. Through optimization of 3D-printing methods, channel geometry, and surface treatment, we created a microchip designed to deliver liquid in 14.64 minutes that ran experimentally in 19.12 ± 2.33 minutes. In the future, we will demonstrate the feasibility of RESTRICT run on-chip and fluorescence measured off-chip by testing clinically-relevant drug concentrations using the controlled liquid delivery time and liquid extraction methods developed. By creating an automated and rapidly fabricated microfluidic chip for therapeutic drug monitoring, we hope to achieve a hands-off device that removes external manipulation to increase the accessibility of RESTRICT-on-a-chip for point-of-need settings without specialized equipment or highly trained operators.

Treatment of individuals with HIV using antiretroviral therapy (ART) is highly effective, but effective clinical management depends on maintaining therapeutic drug concentrations. Antiretroviral (ARV) drug concentrations in patients with HIV can vary due to differences in drug metabolism, medication adherence, or interactions between multiple drugs. These individuals may have subtherapeutic or supratherapeutic drug concentrations, putting them at risk of treatment failure, acquisition of drug resistance, and risk of hospitalization or death. Current measurement of ARV concentration is done through liquid chromatography tandem mass spectrometry, which requires expensive equipment and requires a labor-intensive protocol. This restricts accessibility to specialized laboratories, making it difficult for persons with HIV to have routine measurements of ARV drug concentrations. The goal of the project is to develop an assay that is simple to perform and uses standard equipment to increase access to routine clinic-based drug level monitoring to improve HIV care. We designed an assay using a 2-step process of DNA strand transfer and quantitative polymerase chain reaction (qPCR) to quantify integrase strand transfer inhibitors (INSTIs). We tested for dolutegravir (DTG) and cabotegravir (CAB) in both buffer and plasma -- the latter to simulate patient blood samples. We were able to demonstrate that the assay could quantify clinically relevant drug concentrations of DTG and CAB. By developing an assay that can be readily integrated into most clinical laboratories, we will contribute to increasing access to routine HIV drug level monitoring to improve clinical HIV care and maintaining viral suppression in persons with HIV.

Capillary microfluidics devices automate point-of-care diagnostic assays because of their instrument-free operation, small size, and low material cost. However, capillary microfluidics currently require expensive fabrication instruments limiting rapid prototyping and deployment in low-resource settings. State-of-the-art capillary microfluidics are fabricated by using digital light project stereolithography (DLP-SLA) 3D-printers that offer high resolution (40 µm) but are expensive ($10,000 – $20,000). Liquid crystal display (LCD) SLA printers have recently emerged with comparable resolution and much lower cost ($300 – $1000). However, our initial experiments with LCD-SLA printers exhibited defects and post-processing issues. Our goal is to optimize the fabrication process of capillary microfluidics using an inexpensive LCD-SLA printer (Anycubic Photon Mono 6K, ~$400) and calibrate performance relative to a DLP-SLA printer (CADWorks Pr 4K, ~$15,000). By varying printing parameters including UV power, exposure time, layer height, retraction speed, and resin choice, we found optimal conditions that produced microfluidic channels with comparable dimensions on the Anycubic and CADWorks printers (i.e. coefficient of variation <20%). With printer settings of 40% UV Power, 1.8s exposure time, 20 µm layer height, and 0.1 mm/s retraction speed with CADWorks clear resin, the Anycubic 3D printer produced microchannels down to 100 μm, the smallest feature size we achieved with the CADWorks printer. Optimizing 3D-printing of capillary microfluidics using inexpensive LCD-SLA printers like the Anycubic has the potential to enable rapid prototyping of point-of-care diagnostics in low-resource settings. Next steps include printing more complex microfluidic components including domino valves and trigger valves.

The periodontal ligament is a connective tissue that anchors teeth to the bony socket and is crucial in providing nutrition for the survival and functioning of the human body through the mastication of food by teeth. Because the periodontal ligament is anatomically sealed off from the oral cavity, no non-invasive techniques currently exist to investigate it in-vivo, making the development of sound in vitro models critical for periodontal research. With periodontal disease affecting over 743 million people worldwide, in-vitro research to develop regenerative therapies to replace diseased periodontal tissue is urgently needed. To achieve this, we have developed a novel 3D in-vitro model, which closely mimics the in-vivo periodontal ligament. The 3D tissues are fabricated by inverting an array of silicone posts into silicone molds containing a cell-collagen gel mixture in a 24-well plate. Once the tissues have been incubated and the collagen polymerizes, magnetic mechanical force stretches tissues on posts. The post deflection is used to calculate tissue stiffness and contractility. Preliminary data shows a reduction of contractile force in the tissue constructs after 24 hours of mechanical stretching. I expect to find similar outcomes through additional experimentation. Understanding the periodontal ligament’s response to mechanical force is crucial for its effective restoration and ensuring that it is mechanically sound. The novel in-vitro 3D model that we have developed provides a valuable opportunity to better comprehend the ligament's response to these forces. By performing further experiments with this model, we can gain a deeper understanding of the periodontal ligament, allowing for informed decisions when it comes to replacement and repair in patients. This model offers controlled and repeatable experiments, providing more accurate insights into the biology of the periodontal ligament and contributing to the advancement of periodontal disease treatment.

As of 2021, there were approximately 38.4 million people living with HIV who require routine viral load testing. Viral load testing returns a quantitative measure of viral concentrations and is indicative of antiretroviral therapy efficacy and adherence compliance, with lower viral loads correlated to better health outcomes. Quantitative polymerase chain reaction (qPCR) is the gold standard for measuring viral load, but is not accessible to many clinics and patients around the world due to its long assay times and requirements of specialized equipment and highly trained personnel. As a result, qPCR is limited to centralized testing facilities far from the point-of-care (POC), leading to delayed results or loss of follow-up. Our group has addressed this by developing an HIV viral load test using recombinase polymerase amplification (RPA), which has a 20 minute sample-to-answer time and is more appropriate for POC settings. Our test involves the formation of discrete fluorescent nucleation sites which can be counted to estimate the viral load. However, our test fails to accurately quantify higher HIV viral loads (>3,000cps/rxn). We have difficulties distinguishing between individual sites at these higher copy numbers due to sites merging. In this project, I address the limited dynamic range of this test by performing RPA between two glass slides and investigating the effects of different slide thicknesses and concentrations of polyethylene glycol (PEG), a crowding agent used in RPA reactions. I perform nucleation site analysis using computer vision techniques to measure nucleation site radius and intensity and study how these factors affect site diffusion and amplification. By analyzing nucleation site behavior, we demonstrate potential for an HIV viral load test with a higher dynamic range and gain a better understanding for RPA nucleation site formation, ultimately helping to improve access to testing and treatment for people living with HIV.

Particle-laden turbulent flows are important in both natural and industrial contexts. The particles in many of these processes, such as the formation of ice crystals in clouds or the paper-making process, are anisotropic, with directionally-dependent drag coefficients. Generally, anisotropic particles are free to rotate as they are advected by the carrier fluid. However, external forcing from gravitational and magnetic fields, the larger scale flow, and active behavior can restrict the particles’ orientation, fixing their anisotropic resistance with respect to the reference frame. The dynamics and statistics of symmetric particles in isotropic turbulence are well-studied, but the effect of anisotropic forcing on the transport and behavior of asymmetric particles is less well-understood. We studied these dynamics by conducting Lagrangian particle tracking in simulated isotropic turbulence from the Johns Hopkins Turbulence Database. Computer simulations of 54,000 randomly-placed particle tracers were run in Python, with anisotropy introduced by directly scaling the velocity of the tracer-particle at each simulation time step. We examine how increasing a particle’s resistance to motion in one direction in isotropic turbulence impacts the transport and dispersion statistics in all three directions. We find that increasing tracer anisotropy decreases diffusivity in the direction of velocity scaling as expected, but the diffusivity in the unscaled directions increases such that the total diffusivity remains roughly constant. Studying the dynamics of these simulated anisotropic particles in turbulence will provide a better understanding of how turbulence mixes both particles and the fluid itself, which can then be applied to particle-turbulence interactions in both environmental and industrial contexts.