Doctor of Philosophy, University of Akron, 2019, Chemical Engineering

Breast Cancer is the most common cancer among women and nearly 1 in every 8 American women suffer from it. Triple negative breast cancer, a type of breast cancer that can be only treated using chemotherapy, affects nearly 20% of the breast cancer diagnosed women. Early detection of breast cancer increases survival rates among patients. This dissertation discusses the development of a new two-functional folate-targeted poly(ethylene glycol)-based fluorescein-labeled conjugate (FA-FL-PEG-FL-FA) for potential applications in selective diagnosis of triple negative breast cancer. Since folate, or vitamin B9, receptors (FRs) are overexpressed in breast cancer cells, the diagnosis of these cells is targeted by using a γ-thiolated folic acid (FA-γ-SH) as the targeting precursor. α-carboxyl group present on folate is necessary for binding with the FRs, therefore, exclusive γ-thiolation was achieved by using n-butyllithium that selectively forms the lithium salt (FA-γ-Li) at the γ-carboxyl position due to its higher pKa. The subsequent thiolation was achieved by reducing FA-γ-S-S-γ-FA, which was prepared by reacting a dibrominated disulphide compound (Br-S-S-Br, synthesized via enzyme-catalyzed transesterification) with FA-γ-Li. To increase the retention time in the body and improve the solubility of the agent in water, functionalized PEGs were used as the hydrophilic linkers. Diamine, dithiol, and dibromide functionalization of PEGs were achieved using novel enzyme (Candida antartica Lipase B)-catalyzed esterifications and transesterifications. Specifically, three strategies to synthesize FA-FL-PEG-FL-FA were investigated for their synthetic convenience and feasibility. Strategy 1 used diamine-functionalized PEG (H2N-PEG-NH2), Strategy 2 used dithiol-functionalized PEG (HS-PEG-SH), and Strategy 3 used dibromide-functionalized PEG (Br-PEG-Br) as the precursors. The first strategy and the third strategy were not successful owing to not quantitatively yielding the interm (open full item for complete abstract)

Committee: Lingyun Liu (Advisor); Jie Zheng (Committee Member); Rebecca Kuntz Willits (Committee Member); Chrys Wesdemiotis (Committee Member); Mark Soucek (Committee Member) Subjects: Chemical Engineering; Chemistry; Molecules; Organic Chemistry; Polymer Chemistry; Polymers

PHD, Kent State University, 2022, College of Arts and Sciences / Department of Physics

Nucleic acids play a significant role in biological processes involving genetic information, such as replication, transcription, and translation. Aside from the well-known canonical double helix structure, DNA can adopt a variety of alternative non-canonical structures such as DNA hairpins, holiday junctions, cruciforms, triplexes, or G-quadruplexes (GQ). GQ structures may form in the guanine-rich (G-rich) regions of the genome, including telomeres, promoters, and other significant regulatory sites. Formation of GQs at telomeres protects chromosome ends against degradation and end-to-end joining. The cytosine-rich (C-rich) sequences that are complementary to GQ forming sequences can adopt an intercalated motif (i-motif) structure at acidic pH. In addition to their physiological functions, both the GQ and the i-motif have potential biotechnological applications. In my dissertation, I will describe several projects on single molecule fluorescence and force spectroscopy studies of GQ and i-motif structures. In the first study, we developed a multiplexed, high throughput single-molecule force sensor that converts fluorescence signal into force information by hybridizing the ends of short looped double-stranded DNA (dsDNA) with a nucleic acid secondary (NAS) structure, such as a hairpin or a GQ. When looped, the dsDNA maintains a constant tension across the ends of the NAS. Initially, we completed proof-of-principle studies on this system using different lengths of dsDNA and hairpin structures under various salt conditions. Later, we applied this method to study of the interaction of an under-tension GQ structure with a destabilizing protein, Replication Protein A (RPA), and a stabilizing small molecule, an oxazole telomestatin derivative. The force applied to the GQ via the looped dsDNA was calibrated, with the help of a collaborator, against high-resolution optical tweezers measurements, which is only a few picoNewtons for the construct and assay conditions we used. (open full item for complete abstract)

Committee: Hamza Balci (Advisor); John Portman (Committee Member); Liang-Chy Chien (Committee Member); Soumitra Basu (Committee Member); Thorsten-Lars Schmidt (Committee Member) Subjects: Biophysics; Physics

Doctor of Philosophy, University of Akron, 2013, Polymer Science

The development of polymeric drug delivery devices began five decades ago, starting with hydrogels in 1960. After that, nano-sized drug carriers were developed to increase the efficiency of the drug uptake. This has been accomplished by encapsulating the drug in polymer carriers that increase the circulation time of the drug in the blood, preventing early adsorption, elimination and targeting the drug where it needs to be delivered. One of the most important strategies to increase the circulation time of nanocarriers is PEGylation, in which poly(ethylene glycol) coats the device to prevent premature elimination from the bloodstream due to protein attachment. The goal of this project is to compare the protein adsorption onto polymeric micelles that have either a cyclic or a linear architecture on the hydrophilic coating. We synthesized PEG-b-PCL amphiphilic diblock copolymers in which the poly(ethylene glycol) block is hydrophilic and the PCL block is hydrophobic. First, PEG macroinitiators were synthesized with either cyclic or linear architectures. The macroinitiators were then used for ring-opening polymerization of e-caprolactone. These diblock copolymers were self-assembled into micelles suspended in water by a co-solvent evaporation method. These two types of polymers (no end groups and end groups) allowed us to study the role of polymer architecture on protein adsorption and circulation time.

Committee: Coleen Pugh Dr. (Advisor); Matthew Becker Dr. (Committee Member); George Newkome Dr. (Committee Member); William Landis Dr. (Committee Member); Chrys Wesdemiotis Dr. (Committee Member) Subjects: Polymer Chemistry; Polymers

Master of Science in Engineering, University of Akron, 2008, Biomedical Engineering

PLGA scaffolds used for wound healing and drug delivery can be surface modified to control cellular attachment to the fibers. An in situ technique has been developed whereupon surface modifying polymers migrate to PLGA microfiber surfaces during electrospinning processes by thermally induced phase separation. Biodegradable scaffolds capable of blocking or encouraging cellular adhesions have been fabricated. An electrospun scaffold is proposed in which a primary scaffold facilitates enhanced cellular attachment, while a secondary scaffold, that blocks cellular interactions, is intended to carry a drug payload to aid in tissue development. Thus, PLGA is surface modified by blending with collagen (I), fibronectin, PEG-grafted-chitosan, and PEG-PLGA block copolymer. Physical characterization of the surface-modified PLGA scaffolds is performed to define the shape, fiber diameter, and surface texture of the electrospun fibers. Additionally, assessment of contact angle and mechanical strength are examined to distinguish wettability and tensile characteristics, respectively. Biological characterization is performed to assess cellular adhesion characteristics, to examine the cellular preference between two different surface-modified blends in a single scaffold, and for the probing of vinculin proteins constituting focal adhesion sites to determine cellular adhesion mechanisms. Results indicate that electrospinning produces textured surface-modified microfibers not significantly different from pure PLGA fiber scaffolds. Contact angle results indicate that surface modification alters surface hydrophilicity. Tensile testing determines that surface modifications do not alter yield or tensile strength, but have an effect on the elastic modulus. Results of cellular adhesion studies indicate that the addition of collagen promotes cellular binding, but is not significantly greater than the control. Cellular adhesion among competitive pairings of the modified blends shows no signific (open full item for complete abstract)

Committee: Yang H. Yun Ph.D. (Advisor) Subjects: Biomedical Research; Engineering; Polymers

Doctor of Philosophy (PhD), Ohio University, 2023, Chemistry and Biochemistry (Arts and Sciences)

In recent years, there took place a notable advancement observed in single-molecule fluorescence microscopy methods and its use in various biomolecular research. This technique allows for direct visualization of dynamics and its detailed complexities of various biological processes at the molecular level which is not possible in bulk measurement. Usually, in experiments related to single-molecule fluorescence measurements, the abundance of key molecules is intentionally minimized, which reduces the noise and improves the quality of imaging. However, such a strategy does not work when experiments involve weak interaction between biomolecules. In such a situation nonspecific interaction between molecules of interest and glass would lead to an unwanted fluorescence background signal, which compromises the imaging quality and reduces the measurement accuracy. In this work, glass surfaces have been functionalized in multiple steps. In the initial step, the glass coverslip surface is modified with (3-aminopropyl) triethoxysilane (APTES), and in the next step, the surface is functionalized using methoxy-terminated polyethylene glycol (mPEG) and biotin-terminated polyethylene glycol (bPEG) molecules. Each surface is characterized using dye-labeled protein molecules called neutravidin. for a variety of single-molecule fluorescence studies as PEG molecules are known to repel any nonspecific molecules binding on the functionalized surfaces. Then the surface is used for two-end immobilization of lambda DNA using biotin and neutravidin interaction. Once the surface is functionalized and characterized, lambda DNA is two ends immobilized on the surface using biotin and neutravidin interaction. Then we use that platform to study the intercalation and de-intercalation kinetics of various intercalating dyes such as single- intercalator (YO-PRO-1) and doubleintercalator (YOYO-1) at various experimental conditions of ionic strength and flow speed of the buffer (open full item for complete abstract)

Committee: Jixin Chen (Advisor) Subjects: Chemistry; Physical Chemistry

Doctor of Philosophy, The Ohio State University, 2023, Chemical Engineering

Oxygen therapeutics are being developed for a variety of applications in transfusion medicine. In order to reduce the side-effects (vasoconstriction, systemic hypertension, and oxidative tissue injury) associated with previous generations of oxygen therapeutics, new strategies are focused on increasing the molecular diameter of hemoglobin obtained from mammalian sources via polymerization and encapsulation. Another approach towards oxygen therapeutic design has centered on using naturally occurring large molecular diameter hemoglobins (i.e., erythrocruorins) derived from annelid sources. Therefore, the goal of Chapter 3 was to purify erythrocruorin from the terrestrial worm Lumbricus terrestris for oxygen therapeutic applications. Tangential flow filtration (TFF) was used as a scalable protein purification platform to obtain a >99% pure LtEc product, which was confirmed by size exclusion high performance liquid chromatography and SDS-PAGE analysis. In vitro characterization concluded that the ultra-pure LtEc product had oxygen equilibrium properties similar to human red blood cells, and a lower rate of auto-oxidation compared to human hemoglobin, both of which should enable efficient oxygen transport under physiological conditions. In vivo evaluation concluded that the ultra-pure product had positive effects on the microcirculation sustaining functional capillary density compared to a less pure product (~86% purity). In summary, we purified an LtEc product with favorable biophysical properties that performed well in an animal model using a reliable and scalable purification platform to eliminate undesirable proteins. The long-term storage stability and portability of hemoglobin (Hb)-based oxygen carriers are important design criteria in the development of these therapeutics for use in emergency medicine in austere environments. Lyophilization or storing proteins in a freeze-dried form is known to increase storage lifetime and reduce overall weight. In Chapter 4, we (open full item for complete abstract)

Committee: Andre Palmer (Advisor); Jeffrey Chalmers (Committee Member); Dana McTigue (Committee Member); Eduardo Reategui (Committee Member); David Wood (Committee Member) Subjects: Biochemistry; Chemical Engineering

Master of Arts in Speech Pathology and Audiology, Cleveland State University, 2021, College of Sciences and Health Professions

Speech language pathologists (SLPs) working in medical settings often evaluate and treat individuals with dysphagia. When a patient with dysphagia is not safely receiving sufficient nutrients per oral, an alternative feeding method may be recommended especially for those with a neurological disorder. Percutaneous endoscopic gastrostomy is the most common enteral feeding method and is often recommended by the speech language pathologist. However, the role of the speech language pathologist treating these patients is not clearly defined in the literature. This qualitative study aims to better understand the role that the SLP plays in treating pediatrics and adults with alternative feeding methods, specifically percutaneous endoscopic gastrostomy tubes. To achieve this, a survey was sent to 67 medical SLPs across the country and asked them questions pertaining to the domain areas of demographics, PEG tube duration, PEG tube indicators, education, and counseling, as well as intervention. Results of the study presented as descriptive statistics suggest that there may be discrepancies between treating pediatric and adult patients with PEG tubes. Results also suggest that years of experience impacts some domain areas.

Committee: Violet Cox Ph.D., MLS (Committee Chair); Emily St. Julian M.S. (Committee Member); Anne Su Ph.D. (Committee Member) Subjects: Speech Therapy

Master of Sciences, Case Western Reserve University, 2020, Biomedical Engineering

Although the average cancer mortality rate has steadily declined over the past 25 years, the majority of advanced disease remains resistant to current treatment methods and yields dismal prognoses.1 Novel immunotherapies such as CAR T-cell transfer therapy and ICB provide promising avenues to overcome tumor-mediated immune evasion and initiate an adaptive immune response towards cancer cells. The strongly immunosuppressive microenvironment in advanced tumors, however, can limit the downstream potential of these therapies, and to date these therapies have provided mixed clinical results.2,3 In an attempt to reverse the immunosuppressive tumor microenvironment, we have designed a MSN that can traffic a STING agonist—cdGMP—into innate immune cells and produce a nearly 9-fold increase in pro-inflammatory IFN-훽 secretion compared to the free agonist. In a metastatic 4T1 model, the surface density of PEG molecules dictates what innate immune cells have access to systemically injected MSNs. Following treatment with cdGMP-loaded MSNs, this differential distribution and uptake of particles leads to differences in innate immune cell recruitment at metastatic sites and in the blood. It is displayed that an immunostimulatory nanoparticle can be engineered to enhance innate immune cell activation in advanced, metastatic disease, providing a tool that may be able to work in combination with other current immunotherapies to yield more robust and consistent therapeutic responses.

Committee: Efstathios Karathanasis PhD (Committee Chair); David Wald MD-PhD (Committee Member); Horst von Recum PhD (Committee Member) Subjects: Biomedical Engineering

Doctor of Philosophy, University of Akron, 2019, Polymer Science

PEG and PEG-based hydrogels offer many advantages for tissue engineering scaffolds as a consequence of their hydrophilicity, wide range of cross-linking chemistries, low immunogenicity, and tunable mechanical properties. Human mesenchymal stem cells (hMSCs) have demonstrated varying lineage commitment in response to the mechanical and degradation properties of PEG hydrogels within which they were encapsulated. Furthermore, the proliferation and differentiation of pre-osteoblasts have been shown to be influenced by cell scaffold features such as porosity and surface area. Therefore, there is a need to develop materials that provide precise control over mechanical properties and final scaffold morphology to allow for the accurate assessment of material properties on cell behavior. Few hydrogel systems can alter mechanical moduli without changing design elements such as composition, concentration, and stoichiometry. By employing the pH-sensitive kinetics of oxime bond formation, PEG-oxime hydrogels with a shear modulus range of G' = 5-25 kPa were created from solutions of identical precursor composition, concentration, and stoichiometry simply by varying the pH and buffer concentration of the gelation solution. Equilibrium gel swelling and small angle neutron scattering (SANS) experiments indicated fundamental differences in bond connectivity (i.e. increased loops and dangling ends) were responsible for the observed range in G'. Finally, a 24 h cell viability assay was performed to demonstrate the cell viability of these materials. This study provided important insight in isolating material properties from the chemistry of cross-linking and will inspire further investigations into the effects of material properties on cell behavior. Hydrogel processing is often limited to molding techniques, preventing the formation of complex scaffold morphologies. By utilizing linear PEG as a macro-initiator for the ring opening polymerization of poly(propylene fumarate (open full item for complete abstract)

Committee: Matthew Becker PhD (Advisor); Andrey Dobrynin PhD (Committee Chair); Yu Zhu PhD (Committee Member); Coleen Pugh PhD (Committee Member); Bryan Vogt PhD (Committee Member) Subjects: Biomedical Research; Organic Chemistry; Polymer Chemistry; Polymers

Doctor of Philosophy, University of Akron, 2019, Polymer Science

This dissertation describes the development of novel mass spectrometry (MS) methodologies and techniques to improve, simplify and allow for specific characterizations of synthetic materials. Successful MS analyses are governed by the mass spectrometry principles surrounding specific ionization sources, mass analyzers and ion detecters. The methods described within this dissertation provide information about polymer materials that is historically difficult to determine using other available analytical techniques. Self-assembled terpyridine (tpy)-based metallomacrocycles that are coordinatively-bound using various transition metals were investigated through the analysis of their collision cross-section (CCS) values obtained by travelling wave ion mobility (TWIM)-MS separation, and through comparison to modeled/simulated structures and theoretical CCS values. This work demonstrated the uniquely powerful ability of TWIM-MS separation for architectural verification in samples containing stoichiometrically-defined isobaric ion species ranging from dimeric macrocycles to hexameric macrocycles. Furthermore, TWIM-MS was utilized in a separate project which allowed for the development of a method to monitor concentration dependent supramolecular structural equilibria and conformational trends. Using the methodology in this work, equilibrium constants were derived from these data; this was illustrated for two equilibria concerning the self-assembly of a tpy-based ligand containing a flexible crown ether moiety with Zn2+ ions, and a tetrakisterpyridinyl ligand with Cd2+ ions, which have previously been shown to reversibly interconvert. Additionally, experimental CCS values were also derived from drift time measurements and compared with theoretical predictions for these complex self-assembled terpyridine-based supramolecules to provide insight into their size, stoichiometry and architecture. Parallel studies have focused on the development of methods that can obtain conne (open full item for complete abstract)

Committee: Chrys Wesdemiotis (Advisor); Mark Foster (Committee Chair); George Newkome (Committee Member); Matthew Becker (Committee Member); Kevin Cavicchi (Committee Member) Subjects: Analytical Chemistry; Chemistry; Materials Science; Polymer Chemistry; Polymers

Doctor of Philosophy, Case Western Reserve University, 2018, Chemical Engineering

Commercial plating systems typically utilize an electrolyte containing the plated ion in combination with additive mixtures consisting of multiple organic surface active species. These additives adsorb on the plated electrode, modifying the deposit properties, texture, and distribution. While some of the mechanistic details of the additives adsorption and interactions have been characterized, the effects of convective flow and particularly of complex current waveforms remain uncharted. A specific motivation for the research reported herein, is the preferential fill, by electroplated copper, of blind and open vias in printed circuit boards that is achieved utilizing special additives in combination with the application of periodic reverse current waveform in the presence of ferric ions and complex flow. The process, which is widely utilized, has been developed empirically. Its optimization requires understanding the effects of each process parameter, its quantification, and the development of a comprehensive quantitative model. The additives utilized in this study are polyethylene glycol (PEG) which is a copper deposition inhibitor, and bis-sodiumsulfopropyl-disulfide (SPS), which is a weak accelerator. Those very same additives enable the bottom-up metallization of semiconductor interconnects; however, due to the much larger metallized features (hundreds of microns vs. few nanometers) and much longer deposition time (order of hour instead of a few seconds), the challenges facing the plating process herein, are different and, in many cases, more complex. A quantitative model describing competitive adsorption of additives and polarization effects was developed to address deficiencies in current theories. This model, invoking heterogeneous adsorption energy sites, accounts for the steady-state additives (SPS and polyethylene glycol, `PEG') coverages, subject to competitive adsorption, as a function of additives concentration in solution, and accurately predic (open full item for complete abstract)

Committee: Uziel Landau (Advisor); Rohan Akolkar (Committee Member); Heidi Martin (Committee Member); Daniel Scherson (Committee Member) Subjects: Chemical Engineering

Doctor of Philosophy (PhD), Ohio University, 2018, Chemistry and Biochemistry (Arts and Sciences)

Advancing age is the primary risk factor for cardiovascular, cerebrovascular and renal diseases. The imbalance between prooxidative and antioxidative processes increase with senescence. The decrease in cytoprotective nitric oxide (NO) and increase in reactive oxygen species (ROS) such as superoxide (O2¯) and peroxynitrite (ONOO¯) are suggested to be the main factors of endothelial dysfunction and aging. Endothelial dysfunction is associated with the impaired generation of NO and overproduction of O2¯ resulting in the formation of cytotoxic ONOO¯. Replicative senescence of human endothelial cells (ECs) was analyzed here using human umbilical vein endothelial cells (HUVECs). In this study, we employed a nanomedical system to measure the concentration of NO, O2¯, and ONOO¯ simultaneously. Peroxynitrite is a powerful cytotoxic oxidant formed from the reaction between NO and O2¯. Nanosensors were placed near the endothelium and calcium ionophore-stimulated NO, O2¯, and ONOO¯ were measured. The present work was performed to investigate endothelial nitric oxide synthase (eNOS) dysfunction during aging of endothelial cells. The age-related cardiovascular diseases such as heart failure, atherosclerosis, ischemic heart disease and myocardial infarction can be associated with endothelial dysfunction. The nanomedical approach enabled us to characterize changes in eNOS coupling/uncoupling as a function of biological aging. The release of NO and ONOO¯ was studied in different passages in ethnic groups (Caucasian Americans (CAs), African Americans (AAs) and Asian Americans (ASAs)). Our results suggest that HUVECs of AAs are more susceptible to endothelial dysfunction during aging. The shortening of relative telomere length in aging coincided with a decrease of NO and increase of ONOO¯ concentrations. The imbalance between [NO] and [ONOO¯] was due to endothelial dysfunction. The treatment of aging endothelium with factors affecting the eNOS pathway (e.g., VAS2870, PEG-SOD, and L-ar (open full item for complete abstract)

Committee: Tadeusz Malinski PhD (Advisor) Subjects: Aging; Alternative Medicine; Cellular Biology; Chemistry; Developmental Biology; Endocrinology; Environmental Science; Environmental Studies; Epidemiology; Health Care; Health Sciences; Immunology; Medicine; Microbiology; Molecular Biology; Molecular Chemistry; Molecules; Nanoscience; Nanotechnology; Neurobiology; Neurology; Neurosciences; Oncology; Organic Chemistry; Organismal Biology; Pharmaceuticals; Pharmacology; Pharmacy Sciences; Physiology; Polymer Chemistry; Sports Medicine; Therapy; Toxicology; Zoology

Doctor of Philosophy, University of Akron, 0, Chemistry

Presently in the United States, cancer is the second leading cause of death for both men and women, with lung cancer attributing to approximately 28% of all cancer related deaths. Lung cancer can further be separated into two categories: small-cell lung carcinoma (SCLC) and non-small-cell lung carcinoma (NSCLC). Treatment of NSCLC will depend on the stage of the cancer and can include surgery, chemotherapy, and/or radiation therapy. Chemotherapy often involves a mixture of organic-based and heavy metal-based drugs. The downside of these drugs are that they usually have a narrow therapeutic index and can have detrimental side effects. Due to these downsides, there is a continuous need to develop new anti-tumor agents that have a wider therapeutic index to reduce devastating side effects on the patient while having the desired effect on the tumor cell. Recently, the Youngs group had developed silver N-heterocyclic carbene complexes that show in vitro anticancer properties against NSCLC cell lines, however, solubility of the drugs is an issue. This dissertation describes encapsulating these complexes into biodegradable nanoparticles, in vitro and in vivo testing of the nanoparticles as well as various compounds against a panel of cancerous cell lines. Chapter I goes into the background of medicinal N-heterocyclic carbene complexes and the background of various nanoparticles used for drug delivery as well as the fabrication of the nanoparticle. Chapter II discusses the synthesis of the biodegradable nanoparticles made from poly(D,L-lactic-co-glycolic acid) (PLGA) and poly(ethylene glycol) (PEG) nanoparticles, encapsulation of the complexes, and use of various targeting moieties decorating the nanoparticle surface. In vitro testing for antimicrobial and anticancer properties are also reported. Chapter III aims to optimize silver N-heterocyclic carbene complexes into PLGA-PEG nanoparticles and studies the antimicrobial activities of these systems. Chapter IV shows t (open full item for complete abstract)

Committee: Wiley J. Youngs Dr. (Advisor); Claire A. Tessier Dr. (Committee Member); Peter L. Rinaldi Dr. (Committee Member); Sailaja Paruchuri Dr. (Committee Member); Yang H. Yun Dr. (Committee Member) Subjects: Biomedical Research; Chemistry; Medicine; Nanotechnology; Pharmaceuticals; Polymers

PhD, University of Cincinnati, 2016, Engineering and Applied Science: Materials Science

In vivo, different cell types assemble in specific patterns to form functional tissues. Reproducing this process in vitro by designing scaffold materials to direct cells precisely to the right locations at the right time is important for the next generation of biomaterials. Here, using Microarray Amplification of Natural Directional Persistence (MANDIP), we demonstrated simultaneous assembly of fibroblasts and endothelial cells by directing their long range migration. Quantitative analysis of cell migration on different MANDIP designs yielded insights on the relative importance of asymmetric island shape and arrangement. The approach enables spatial patterning of different cell types with micrometer-scale precision over large areas for investigation of cell-cell interactions within complex tissue architecture. To reduce the mortality due to the cancer, it is necessary to study the migrations and proliferation of human epidermal keratinocytes (HEK). Here, we focused on how an individual HEK's mobility that directional property and speed of migration on long-range patterns in micron scale. HEK migrated to one direction of the pattern then turned the moving direction to the previous position and expanded their activity regions during migrating repeatedly as back and forth. HEK secrete a biopolymer, fibronectin, during migration. Our hypothesis is that it causes HEK to remember their previous position and is guided to their moving direction. To demonstrate how the long-range micropatterns were modified to closed patterns as a rotation and anti-fibronectin was applied on the substrate after cell migration. In the results, HEK migrated continuously one direction as either clockwise or counter-clockwise after rotating once on the pattern. The studies of behaviors of the individual keratinocytes provide that an improved design for wound healing and other tissue engineering applications. Disparities in cellular behavior between cultures of a single cell type and (open full item for complete abstract)

Committee: Chia Chi Ho Ph.D. (Committee Chair); Gregory Beaucage Ph.D. (Committee Member); Carlos Co Ph.D. (Committee Member); Jude Iroh Ph.D. (Committee Member); Vesselin Shanov Ph.D. (Committee Member) Subjects: Biochemistry

Doctor of Philosophy (PhD), Ohio University, 2016, Chemistry and Biochemistry (Arts and Sciences)

The T box riboswitch is a regulation mechanism at the level of transcription or translation which controls expression of amino acids related genes, including a lot of essential genes, in many bacteria. The T box riboswitch interacts with cognate tRNAs and senses their aminoacylation status. A charged cognate tRNA allows formation of the thermodynamically more stable terminator structure which induces transcription termination. An uncharged tRNA stabilizes the alternative antiterminator structure and prevents formation of the terminator. Transcription proceeds and leads to expression of the downstream gene(s). The T box riboswitch is a novel and promising drug target since multiple genes essential to bacterial survival are regulated by this mechanism in many pathogenic bacteria. In order to further study the T box riboswitch mechanism and screen a synthetic ligands library, a fluorescently monitored multi-round in vitro antitermination assay with an enhanced throughput was successfully developed and comprehensively evaluated. Using this assay, the effects of molecular crowding, spermidine and DMSO on the T box riboswitch function were studied and 304 ligands were screened. A total of nine ligands showed specific inhibition to the tRNA-induce antitermination. Combining melting temperature analysis and structural probing, the binding of spermidine to the antiterminator was also characterized.

Committee: Jennifer Hines PhD (Advisor) Subjects: Biochemistry; Chemistry; Molecular Biology

Master of Science, University of Akron, 2015, Polymer Science

Poly(lactic acid) (PLA) plays an important role in biodegradable polymers due to its unique properties. However, PLA only has reactive end groups; therefore it is difficult to make bulk modifications and an efficient way to functionalize the main-chain of this polyester needs to be found. 2-Bromo-3-hydroxypropanoic acid (BHPA) is a halogenated constitutional isomer of lactic acid. In this thesis, BHPA is used to prepared to make statistical copolymers with lactic acid by direct polycondensation to provide reactive sites along the polymer backbone. The halogen is distributed along the backbone of the copolymer, such as a grafting-to structure can be made. Poly(ethylene glycol) (PEG) is most often used as a hydrophilic segment because of its biodegradability and biocompatibility. The PEGylated poly(lactic acid) can serve as a polymer to which various medical and drug delivery systems can be attached. Here, mPEG side chains were formed by grafting mPEG homopolymers with 2,2,6,6-tetramethylpiperidine-1-oxyl (TEMPO) end groups onto the polymer by a radical coupling reaction. TEMPO is one of the most widely used nitroxide radicals due to its remarkable persistency. PEGylated poly(lactic acid) can be prepared via a radical coupling reaction of PEG-supported TEMPO with the brominated poly(lactic acid).

Committee: Coleen Pugh Dr. (Advisor); Yang Hyun Yun Dr. (Committee Member) Subjects: Polymer Chemistry

Master of Sciences, Case Western Reserve University, 0, Materials Science and Engineering

Proteins and viral nanoparticles are important building blocks for the construction of micro- and nano-scale materials. The behavior of these biomolecular systems in crystallization, self-association and molecular recognition, is determined by the long-range interactions between them. Understanding the variation of long-range interactions under different solution conditions facilitates the manipulation and design of novel hierarchical structures. The second virial coefficient is the most significant factor that depends on the strength of long-range interaction in solutions. Composition-gradient multi-angle static light scattering (CG-MALS) was used to characterize the second virial coefficient of a typical globular protein, bovine serum albumin (BSA), and a larger macromolecule, cowpea mosaic virus (CPMV). Since BSA molecules are prone to aggregate, a composition identification of the BSA solution is carried out with fast protein liquid chromatography (FPLC) and dynamic light scattering (DLS) measurements. The FPLC and DLS study show that monomer is the dominating species in the BSA solutions studied. Then the second virial coefficient of the BSA solution is measured by CG-MALS as a function of pH and univalent salt (NaCl) strengths. The value of second virial coefficient remains greater than zero (corresponding to predominant repulsive interaction between BSA molecules) throughout the experimental region , and the magnitude has a systematic variation as a function of pH and NaCl strength, which reveals the change of the net charge on the surface of BSA molecule and the resulting variation of the inter-molecular force. The minimum of the second virial coefficient is determined to be 1.13*10-5 ml*mol/g² at pH ∼4.6, identified as the isoelectric point of BSA, where the electrostatic interaction reaches the minimum value and van der Waals-London dispersion interaction governs the long-range ordering of the proteins. The second virial coefficient of native CPMV and PEGy (open full item for complete abstract)

Committee: Roger French (Committee Chair); Frank Ernst (Committee Member); Nicole Steinmetz (Committee Member); Rudolf Podgornik (Committee Member) Subjects: Materials Science

Doctor of Philosophy, Case Western Reserve University, 2014, Biomedical Engineering

The key role of smooth muscle cell (SMC) migration and proliferation in vascular physiological and pathological remodeling necessitates the exploration of mechanisms underlying these functions. This work focuses on engineering a poly (ethylene glycol) (PEG) hydrogel as a model system to evaluate SMC migration and proliferation in three dimensions (3D). We hypothesized that 3D SMC migration and proliferation can be regulated by the properties of a cell-instructive scaffold, including cell-matrix adhesion, degradability, and cross-linking density. To accomplish this, bio-inert PEG-based hydrogels were designed as the scaffold substrate. To mimic the properties of the extracellular matrix (ECM), cell-adhesive peptide (GRGDSP) and enzyme-sensitive peptide (VPMSMRGG or GPQGIAGQ) were incorporated into the PEG macromer chain. Copolymerization of the biomimetic macromers by photopolymerization resulted in the formation of bioactive hydrogels with the dual properties of cell adhesion and proteolytic degradation. Studies of mass swelling ratio as a function of gel compositions indicated that this hydrogel can be engineered quantitatively to allow for uncoupled investigation of scaffold properties on cell functions. By utilizing these biomimetic scaffolds, we studied the effect of adhesive ligand concentration, proteolysis, and network cross-linking density on 3D SMC migration and proliferation. Our results indicated that 3D SMC migration and proliferation were critically dependent on cell-matrix adhesiveness, proteolysis, and cross-linking density. The incorporation of cell-adhesive ligand significantly enhanced SMC spreading, migration and proliferation, with cell-adhesive ligand concentration mediating 3D SMC migration and proliferation in a biphasic manner. The faster degrading hydrogels promoted SMC migration and proliferation. In particular, higher cross-linking density could impede 3D SMC migration and proliferation despite the presence of cell-adhesive ligands and pro (open full item for complete abstract)

Committee: Kandice Kottke-Marchant (Committee Chair); Anirban Sen Gupta (Committee Member); Horst von Recum (Committee Member); Stuart Rowan (Committee Member) Subjects: Biomedical Engineering; Polymers

Doctor of Philosophy, University of Akron, 2014, Polymer Science

The synthesis of functional polymers with well-defined structure, end-group fidelity and physico-chemical properties useful for biomedical applications has proven challenging. Chemo-enzymatic methods are an alternative strategy to increase the diversity of functional groups in polymeric materials. Specifically, enzyme-catalyzed polymer functionalization carried out under solventless conditions is a great advancement in the design of green processes for biomedical applications, where the toxicity of solvents and catalyst residues need to be considered. Enzymes offer several distinct advantages, including high efficiency, catalyst recyclability, and mild reaction conditions. This reseach aimed to precisely functionalized polymers using two methods: enzyme-catalyzed functionalization via polymerization and chemo-enzymatic functionalization of pre-made polymers for drug delivery. In the first method, well-defined poly(caprolactone)s were generated using alkyne-based initiating systems catalyzed by CALB. Propargyl alcohol and 4-dibenzocyclooctynol (DIBO) were shown to efficiently initiate the ring opening polymerization of e-caprolactone under metal free conditions and yielded polymers with Mn ~4 to 24 KDa and relatively narrow molecular mass distribution. In the second methodology, we present quantitative enzyme-catalyzed transesterification of vinyl esters and ethyl esters with poly(ethylene glycol)s (PEG)s that will serve as building blocks for dendrimer synthesis, followed by introducing a new process for the exclusive gamma-conjugation of folic acid. Specifically, fluorescein-acrylate was enzymatically conjugated with PEG. Additionally, halo-ester functionalized PEGs were successfully prepared by the transesterification of alkyl halo-esters with PEGs. 1H and 13C NMR spectroscopy, SEC and MALDI-ToF mass spectrometry confirmed the structure and purity of the products.

Committee: Judit E. Puskas Dr. (Advisor); Matthew L. Becker Dr. (Advisor); Chrys Wesdemiotis Dr. (Committee Chair); Toshikazu Miyoshi Dr. (Committee Member); Abraham Joy Dr. (Committee Member); Nic D. Leipzig Dr. (Committee Member) Subjects: Biomedical Research; Materials Science; Polymer Chemistry

Master of Science in Engineering, University of Akron, 2013, Biomedical Engineering

Mechanical properties are a critical factor in cell behavior and can signal cells to differentiate, migrate, and proliferate. In vivo, the mechanical properties of tissues are complex and heterogeneous. Mechanical gradients are prevalent in the human body as every inter-connective tissue in every organ has gradients across the spatial volume. In addition, pathologies, such as, cancer and myocardial infarction trigger complex physical changes within the tissue. After myocardial infarction, the cardiac tissue hardens creating a mechanical gradient interface between the diseased cells and healthy tissue. Two weeks after myocardial infarction, the tissue hardens creating a gradient from the healthy tissue (16-20 kPa) to infarcted tissue (~50 kPa). In order to mimic the native environment, it is necessary to have a scaffold in which the mechanical properties can be tuned spatially. Photomasking is a method by which one can restrict the amount of UV irradiation intensity depending on the opacity of the photomask. Reducing the irradiation intensity over a fixed duration can selectively crosslink scaffolds which undergo photopolymerization. Polyethylene glycol (PEG) can be functionalized with a diacrylate group to form a hydrogel which can undergo radical photopolymerization. The methods described in this paper analyze how different opacity photomasks (0, 10, 20, 30, and 40%) affect irradiation intensity, and then how the irradiation intensity affects the elastic modulus (E') and swelling ratio of 9% wt/wt PEG hydrogel. Using the relationship between photomasking and E' a gradient scaffold was designed resulting in a PEG gradient scaffold between 17-57 kPa. Type I collagen was incorporated within the PEG scaffold at a concentration of 1 mg/mL. The incorporation of the collagen has no significant effect on the E'. A live / dead assay was performed on the PEG collagen scaffold showing high viability (95.91 %) of human mesenchymal stem cells (hMSCs). hMSCs wer (open full item for complete abstract)

Committee: Ge Zhang Dr. (Advisor); Rebecca Willits Dr. (Committee Member); Nic Leipzig Dr. (Committee Member) Subjects: Biomechanics; Biomedical Engineering