Fischell Department of Bioengineering Theses and Dissertations
Permanent URI for this collectionhttp://hdl.handle.net/1903/6628
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Item Functionalized Nanoparticles for the Controlled Modulation of Cellular Behavior(2023) Pendragon, Katherine Evelyn; Fisher, John; Delehanty, James; Bioengineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)The ability to control cellular behavior at the single-cell level is of great importance for gaining a nuanced understanding of cellular machinery. This dissertation focuses on the development of novel hard nanoparticle (NP) bioconjugate materials, specifically gold nanoparticles (AuNPs) and quantum dots (QDs), for the controlled modulation of cellular behavior. These hard NPs offer advantages such as small size on the order of 1 – 100 nm, high stability, unique optical properties, and the ability to load cargo on a large surface area to volume ratio, making them ideal tools for understanding and controlling cell behavior. In Aim 1, we demonstrate the use of AuNPs to manipulate cellular biological functions, specifically the modulation of membrane potential. We present the conception of anisotropic-shaped AuNPs, known as gold nanoflowers (AuNFs), which exhibit broad absorption extending into the near-infrared region of the spectrum. We demonstrate the effectiveness of utilizing the plasmonic properties AuNFs for inducing plasma membrane depolarization in rat adrenal medulla pheochromocytoma (PC-12) neuron-like cells. Importantly, this is achieved with temporal control and without negatively impacting cellular viability. Aim 2 explores the use of QDs as an optical, trackable scaffold for the multivalent display of growth factors, specifically erythropoietin (EPO), for the enhanced induction of protein expression of aquaporin-4 (AQPN-4) within human astrocytes. This results in enhanced cellular water transport within human astrocytes, a critical function in the brain's glymphatic system. We show that EPO-QD-induced augmented AQPN-4 expression does not negatively impact astrocyte viability and augments the rate of water efflux from astrocytes by approximately two-fold compared to cells treated with monomeric EPO, demonstrating the potential of EPO-NP conjugates as research tools and prospective therapeutics for modulating glymphatic system function. Overall, the body of work presented in this dissertation develops new NP tools, namely solid anisotropic AuNFs and growth factor-delivering QDs, for the understanding and control of cell function. These new functional nanomaterials pave the way for the continued development of novel NP-based tools for the precise modulation of cellular physiology.Item SPRAYABLE, BIODEGRADABLE POLYMER BLENDS FOR TISSUE ADHESION(2019) Daristotle, John L; Kofinas, Peter; Bioengineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Tissue adhesive materials can revolutionize surgical procedures, but they are often difficult to apply safely because of a required curing step where the viscous components of a glue solidify and become sticky. To simplify their deposition and improve their usability, this dissertation introduces tissue adhesive polymer blends that can be sprayed using a fiber production technique called solution blow spinning. The polymer blends studied here are innovative because they are non-curing: the polymer accumulates as a solid material directly on the tissue substrate of interest during spraying, quickly forming a strong bond. To achieve a rapid increase in tissue adhesion, we developed a surgical sealant composed of poly(lactic-co-glycolic acid) and poly(ethylene glycol) (PLGA/PEG) that becomes adhesive in response to warming to body temperature. We then evaluated PLGA/PEG in small and large animal models of intestinal anastomosis and partial thickness skin wounds. Additional improvements to hemostasis, flexibility, and adhesion were made by incorporating micron-sized silica particles, which produced textured fibers with suppressed crack formation. We also developed the first pressure-sensitive tissue adhesive by formulating elastomeric copolymer blends with two components of different molecular weights. An additional objective of this dissertation was to study sprayable polymers that can be used as a controlled release system for various drugs. Towards this goal, we incorporated antimicrobial silver into solution blow spun PLGA/PEG fibers. At the optimal concentration, silver ions released over 14 days at levels that were effectively antimicrobial with minimal cytotoxicity. Coating strategies for controlling the delivery of polyelectrolyte complexes were also investigated.Item THE APPLICATION OF MICRODEVICES FOR INVESTIGATING BIOLOGICAL SYSTEMS(2018) Shang, Wu; Bentley, William E; Bioengineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)The gastrointestinal (GI) tract is a complex ecosystem with cells from different kingdoms organized within dynamically-changing structures and engaged in complex communication through a network of molecular signaling pathways. One challenge for researchers is that the GI tract is largely inaccessible to experimental investigation. Even animal models have limited capabilities for revealing the rich spatiotemporal variation in the intestine and fail to predict human responses due to genetic variation. Exciting recent advances in in vitro organ model (i.e., organ-on- chips (OOC)) based on microfluidics are offering new hope that these experimental systems may be capable of recapitulating the complexities in structure and context inherent to the intestine. A current limitation to OOC systems is that while they can recapitulate structure and context, they do not yet offer capabilities to observe or engage in the molecular based signaling integral to the functioning of this complex biological system. This dissertation focuses on developing microfluidic tools that provide access to interrogating signaling events amongst populations in the GI tract (e.g., microbes and enterocytes). First, a membrane-based gradient generator is built to establish linear and stable chemical gradients for investigating gradient-mediated behaviors of bacteria. Specifically, this platform enables the study of bacterial chemotaxis and potentially facilitates the development of genetically rewired lesion-targeted probiotics. Second, “electrobiofabrication” is coupled with microelectronics, for the first time, to create molecular-to-electronic (i.e., “molectronic”) sensors to observe and report the dynamic exchange of biochemical information in OOC systems. Last, to address the issue of poor compatibility between OOCs and sensors, we assemble OOCs with molectronic sensors in a modular format. The concept of modularity greatly reduces the system complexity and enables sensors to be built immediately before applications, avoiding functional decay of active biorecognition components after long-term device storage and use. We envision this work will “open” OOC systems for molecular measurement and interrogation, which, in turn, will expand the in vitro toolbox that researchers can use to design, build and test for the investigation of GI disease and drug discovery.Item Design of Self-Assembling Nanostructures to Promote Immune Tolerance(2018) Hess, Krystina; Jewell, Christopher M; Bioengineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)In autoimmune diseases, which affect more than 23 million Americans, the immune system mistakenly attacks healthy tissue. This occurs when the process that normally controls self-reactive inflammatory cells (i.e. tolerance) fails. In multiple sclerosis (MS), the myelin sheath, which insulates nerves, is recognized as a foreign antigen. Demyelination by immune cells results in serious symptoms of neurodegeneration. Current treatments for MS are not curative, but rather manage symptoms by broadly suppressing the immune system, leaving patients unable to fight infection. New therapies that are more specific and effective could greatly improve the quality of life for patients. Biomaterials offer specific advantages for generating antigen-specific tolerance, such as cargo protection, targeted delivery, and controlled release of signals. Additionally, recent reports demonstrate that materials themselves can be intrinsically immunogenic. Two promising biomaterials-based strategies for combating autoimmunity involve: 1) delivery of self-antigen with a regulatory molecule or 2) delivery of self-antigen alone. Aim 1 of this dissertation focuses on the first strategy, creating a novel delivery system for myelin peptide and GpG, an immunomodulatory oligonucleotide. This approach involves electrostatic self-assembly of the two immune signals, eliminating the need for a carrier that could exacerbate inflammation, while still offering attractive features of biomaterials, such as co-delivery. The goal is for immune cells to encounter both signals simultaneously, biasing the response towards tolerance. This work represents the first studies using self-assembled materials to target toll-like receptor signaling, recently shown to be implicated in many autoimmune diseases. Aim 2 of this dissertation is based on the second strategy above, which relies on evidence that changing the trafficking and processing of a self-antigen can impact the development of inflammation or tolerance. Quantum dots, NPs that are intrinsically fluorescent and rapidly drain to lymph nodes, can be decorated with a large and controllable number of myelin peptides. These key features of QDs were exploited to reveal that parameters of self-antigen display (i.e. dose, density) impact biodistribution and immune cell uptake, and are directly correlated to the level of tolerance induced. Together, the described nanotechnologies offer opportunities to probe important questions towards the design of antigen-specific therapies.Item Optical and Thermal Systems for Automation of Point-of-Care Assays(2018) Goertz, John; White, Ian M; Bioengineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Modern medicine has detailed 70,000 different diagnoses; the 21st century challenge is bringing those diagnoses to over 7 billion people. This phenomenal feat requires precision biosensing strategies that minimize necessary training and manual effort while maximizing portability and affordability. Microfluidic strategies, both fabricated chips and paper-based devices, held the promise to facilitate point-of-care diagnostics but have been inadequate for many applications due to the trade-off between bulky pumps or limited control and complexity. This dissertation details novel strategies that control the progression of biochemical reactions with high functionality, portability, and ease-of-use. First, I will describe an amplified signaling reaction that leverages both positive and negative feedback loops to achieve optically-regulated control. This assay, termed “Peroxidyme-Amplified Radical Chain Reaction” enables naked-eye detection of catalytic reporter DNA structures at concentrations across five orders of magnitude down to 100 pM while eliminating the need for manual addition of hydrogen peroxide common to other such detection reactions. Next, I will describe the development of a platform for thermal regulation of generic reactions. To address the need for a broadly capable automation platform that provides equal utility in the lab and field alike, we recently developed “phase-change partitions”. In our system, purified waxes segregate reagents until incremental heating melts the partitions one by one, causing the now-liquid alkane to float and allowing the desired reagents to interact with the sample on demand. This tight control over reaction progression enabled us to construct hands-free detection systems for isothermal DNA amplification, heavy metal contamination, and antibiotic resistance profiling. My work has demonstrated a broadly capable suite of assay control systems with the potential to enable simple, inexpensive automation of a broad array of chemical and biological analysis across human medicine, environmental surveillance, and industrial chemical synthesis.Item DEVELOPING AN IN SITU SPRAYABLE BIOSCAFFOLD WITH ANTIMICROBIAL PROPERTIES AS AN EMERGENCY BURN WOUND DRESSING(2018) Hunter, Joseph William; Kofinas, Peter; Bioengineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Commercially available silver dressings and their associated application/removal protocols suffer from several serious drawbacks, including: inability to monitor the burn wound beneath the opaque dressing, high costs, traumatic debridement due to a non-degradable mesh, delays involved with transporting a patient to a location where sterile conditions can be maintained, and restrictions upon when a silver dressing can be used. The results of this work present a promising proof of concept for an in situ sprayable synthetic polymer containing a silver salt that was found to allow for wound observation (transitions to clear at body temperature), degrade in a biocompatible manner, release broad-spectrum antimicrobial silver in a controlled manner, and can be safely applied in the acute period following a burn in the absence of sterile conditions with little detriment to wound healing in vivo and less than 20% reduced viability in vitro.Item Engineering biomaterials to promote systemic, antigen-specific tolerance(2017) Tostanoski, Lisa Hoban; Jewell, Christopher M; Bioengineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)In autoimmune diseases, such as multiple sclerosis (MS) and type 1 diabetes, the immune system incorrectly identifies and attacks “self” molecules. Existing therapies have provided important benefits, but are limited by off-target effects, reduced efficacy as disease progresses, and lack of cure potential, necessitating frequent, life-long dosing. An exciting strategy being explored is the design of vaccine-like therapies that selectively reprogram immune responses to self-molecules. This approach could, for example, control the attack of myelin – the protective coating around neurons – that occurs during MS, without leaving patients immunocompromised. However, the realization of this idea has proven difficult; once injected, conventional approaches do not provide control over the combinations, concentrations, and kinetics of signals that reach key tissues that orchestrate immune responses, such as lymph nodes (LNs). Biomaterials have emerged as a promising strategy to confront this challenge, offering features including co-delivery of cargos and controlled release kinetics. The research in this dissertation harnesses biomaterials to develop novel strategies to promote effective, yet selective control of autoimmunity, termed antigen-specific tolerance. In the first aim, direct injection was used to deposit degradable microparticles in LNs, enabling local controlled release of combinations of myelin peptide and Rapamycin, a drug shown to promote regulatory immune function. This work demonstrates the potency of intra-LN delivery in mouse models of MS, as a single dose of co-loaded microparticles permanently reversed disease-induced paralysis in a myelin-specific manner. The results also support this approach as a platform to study the link between local LN signaling and resultant responses in non-treated tissues and sites of disease during autoimmunity. In the second aim, myelin peptide and GpG, a regulatory ligand of an inflammatory pathway overactive in mouse models and patients with autoimmunity, were self-assembled. This approach generated microcapsules that mimic attractive features of conventional biomaterials, but eliminate synthetic carrier components that can complicate rational design and, due to intrinsic inflammatory properties, might exacerbate autoimmunity. These materials promoted tolerance in mouse cells, mouse models of MS, and samples from human MS patients. Together, these strategies could offer novel, modular approaches to combat autoimmune diseases and inform design criteria for future therapies.Item TOWARDS AN UNDERSTANDING OF THE DEGRADATION MECHANISMS OF UHMWPE-BASED SOFT BALLISTIC INSERTS(2016) TSINAS, ZOIS; Al-Sheikhly, Mohamad; Bioengineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)The objective of this work is to advance the field of lightweight and soft ultra-high molecular weight polyethylene (UHMWPE) inserts used in ballistic resistant-body armor, through the evaluation of chemical and physical degradation, and provide critical insight into the mechanisms involved. These inserts are comprised of non-woven UHMWPE fibers, foil-matrix low density polyethylene (LDPE), and a binder resin. Degradation of these components can be initiated by mechanical stress induced by routine use of the armor, thermal exposure due to storage and wear, and exposure to humidity and oxygen. Degradation of this system may include C-C and C-H bond ruptures resulting in C-centered radicals, thermo-oxidative reactions, as well as changes in the degree of crystallinity and the crystalline morphology of the UHMWPE fibers. This is the first comprehensive study on degraded UHMWPE-fibers extracted from body armor that have been subjected to accelerated aging. Previous studies have only focused on oxygen uptake and changes in the tensile strength of virgin UHMWPE fibers as markers of degradation. This work extends beyond oxygen uptake, to examine changes in the topography, the degree of crystallinity, and the crystal phases of UHMWPE fibers. Mechanical stress was found to be the main cause of kink band formation in UHMWPE fibers. Additionally, oxidation products and molecular oxygen were found to be at higher concentrations in the kink bands compared to other parts of the fiber. This suggests a synergistic effect between mechanical stress induced kink bands and oxidative degradation. The degree of crystallinity of the fibers did not change significantly, however morphological changes of the crystalline phases and changes in the orientation of the crystals were observed. Finally, this study investigates, for the first time, the degradation of the binder material that retains the fibers together in the laminates. The binder resin used in the laminates was identified to be a copolymer of polystyrene and polyisoprene, which undergoes oxidative degradation accompanied by a decrease in the weight-average molecular weight.Item Engineering Biodegradable Vascular Scaffolds for Congenital Heart Disease(2015) Melchiorri, Anthony John; Fisher, John P; Bioengineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)The most common birth defects worldwide are congenital heart defects. To treat these malformations in a child’s cardiovascular system, synthetic grafts have been used as a primary intervention. However, current grafts suffer from deficiencies such as minimal biological compatibility, inability to grow and adapt, and high failure rates. Additionally, the grafts are not customized to the patient, which may lead to graft failure given that defects may vary significantly from patient to patient. The work presented here aims to adapt tissue engineering paradigms to develop customizable vascular grafts for congenital heart defects using to reduce the long-term risk and the number of surgeries experienced by patients. The first component of this research focuses on solvent-cast vascular grafts. This system of fabrication was used to explore how various strategies and graft modifications affect the graft’s performance in vivo. Grafts were fabricated with the mechanical properties necessary to withstand the stresses of a physiological environment and support neotissue formation. To improve tissue formation, the grafts were modified with bioactive molecules to improve vascular tissue growth. In addition, the grafts were combined with a tissue perfusion bioreactor. The bioreactor applied fluid flow to support cell seeding, differentiation, and growth of endothelial progenitor cells on the grafts, demonstrating a robust strategy for tissue formation prior to implantation. The second component of this research centers on the development of a biomaterial for 3D printing. 3D printing offers unparalleled customizability, as a graft can be designed based on medical images of a patient, tested via computer modeling, and then printed for implantation. A resin was developed consisting to produce grafts that were mechanically compatible with native blood vessels and maintained patency and tissue formation six months after implantation. The library of 3D printed vascular graft materials was also expanded by creating a novel copolymer resins, which varied in mechanical properties and degradation profiles. In addition, the concepts and strategies of biofunctionalization developed in the solvent-cast vascular grafts can be combined with the 3D printed graft strategies. Grafts designed, printed, and modified using these combinatorial approaches can greatly improve the long-term outcomes of treating congenital heart disease.Item Polymeric Materials for Hemostatic and Surgical Sealant Applications(2015) Behrens, Adam Michael; Kofinas, Peter; Bioengineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Commercial hemostatic agents and surgical sealants do not meet the current clinical need. The available options suffer from a variety of shortcomings including high costs, short shelf lives, difficult preparations, and concerns over safety. This work aims to utilize synthetic polymers to develop alternative approaches that have the potential to improve outcomes from traumatic injuries and surgeries while minimizing risk and cost. The first aspect of this research focuses on the development of hemostatic hydrogel particles. These spherical hydrogels with a narrow size distribution were synthesized via inverse suspension polymerization. A cationic monomer was utilized in the hydrogel formulation to facilitate rapid swelling, leading to the formation a physical barrier to blood loss. Coagulation studies demonstrated the ability to cause localized aggregation through charge interactions with erythrocytes while reducing clotting activity in the bulk. This mechanism allows the hydrogel to quickly block blood flow and may mitigate thrombotic complications at distal sites. Hemostatic efficacy was exhibited by decreases in both the time to hemostasis and mass of blood loss in rat liver puncture and tail amputation injury models when compared to compression with gauze alone. The second aspect of this research focuses on the development of a synthetic surgical sealant. This work is centered on the investigation of a polymer fiber mat deposition method called solution blow spinning. This fabrication technique allows for the rapid in situ generation of polymer fibers, offering the ability to conformally deposit polymeric materials directly on the surgical site of interest. Solution blow spinning was utilized to deposit a body temperature responsive, biodegradable polymer blend. Above a critical temperature, the two phase fibrous polymer mat transitioned into a one phase polymer film. This transition resulted in plasticization and promoted polymer-substrate interaction, leading to increased adhesion. Sealant efficacy was demonstrated in a cecal intestinal anastomosis mouse model, where the polymer blend was used to supplement sutures. Both burst pressure and survival rate were significantly improved over the suture-only control.