Master of Science, University of Akron, 2017, Physics

Fluorescence correlation spectroscopy (FCS) is a useful experimental technique, that uses statistical analysis of fluorescence intensity fluctuations and a correlation function. The method permits studying the kinetics and interactions of particles not only on the surface of the solution, but also in the bulk. Another benefit of FCS is that it does not require the selection of specific molecules or time intervals for its measurements. By analyzing biological dynamics, we can learn about concentration, viscosity, individual movement and diffusion of particles as well as interactions and the possibility of binding between them. In this work, we present a brief explanation on the analytic formalism of the governing concepts of FCS, as well as detail the experimental setups. We also discuss protein-vesicle interactions and the kinematics of double-stranded DNA and labeled nucleotides. The brightness and size of molecules, before and after adding vesicles to the protein solution, are calculated. Finally the possibility of vesicle-protein binding is validated.

Committee: Adam Smith Dr. (Advisor); Ben Hu Dr. (Committee Chair); Jutta Luettmer-Strathmann Dr. (Committee Member) Subjects: Biochemistry; Biophysics; Molecular Biology; Molecular Chemistry; Physical Chemistry; Scientific Imaging

Master of Science, University of Akron, 2015, Physics

Cellular membranes are complex assemblies and a clear understanding of the physical interactions during their function is of paramount importance. Here, we perform two separate studies for a better understanding of the interactions between membrane compartments and other biomolecules. In the first study, we developed a coupler to integrate a high sensitivity spectrometer with an epi-fluorescence microscope to measure fluorescence spectra of small area samples (400 micrometer squared). We applied our measurements on standard samples, performed three corrections on them and after a linear demixing process, the percentage of FRET efficiency was obtained. The development of this method will be advantageous in future single cell studies for detecting population heterogeneity. In the second study, we investigated the dynamics of membrane lipids in a supported lipid bilayer. Single particle tracking total internal reflection fluorescence microscopy (TIRF) was used to study the lateral mobility of phosphatidylinositol phosphate (PIP) lipids with and without an adsorbed polycationic polymer, quaternized polyvinylpyridine (QPVP). Diffusion coefficients were determined with Brownian and anomalous models. Our results indicate a decrease in diffusion coefficient of the lipids in the presence of QPVP in comparison to its absence, revealing their interaction.

Committee: Adam Smith (Advisor); Jutta Luettmer-Strathmann (Committee Chair); Sergei Lyuksyutov (Committee Member) Subjects: Biophysics; Chemistry; Physical Chemistry; Physics

Master of Science, The Ohio State University, 2011, Vision Science

Introduction: The meibum is a lipid rich secretion that is the primary component of the external layer of the two-layered corneal tear film. The meibomian glands produce the meibum, and meibomian gland dysfunction leads to the degradation of the tear film. This dysfunction is known to lead to ocular irritation, inflammation, and pathology. Understanding this relationship is critical to preventing ocular disease. Methods: An extensive search of peer-reviewed literature focusing on the collection, quantification, and analysis of normal and abnormal meibum and tear lipids was performed by searching PubMed. Results: Numerous collection and quantification techniques are described and their advantages and disadvantages are listed. Studies demonstrate that the meibum consists of a large array of polar (phospholipids, sphingolipids, and ω-hydroxy fatty acids) and non-polar (wax esters, cholesterol esters, diesters, free sterols, monoglycerides, diglycerides, triglycerides, free fatty acids, fatty acid amides, and hydrocarbons) lipids, and changes to these lipids are linked to ocular disease. Individual lipids correlated to pathology are described when possible. Lipid deposition patterns on silicone hydrogel contact lenses are also analyzed. Conclusion: Research shows that the normal meibum and tear lipids are essential for normal ocular health. There is still a dearth of information about each lipid's structure and quantity present due to the inherent difficulties of working with small sample sizes. Additional work needs to be performed to resolve these unanswered questions, so better treatments for blepharitis and dry eye syndrome can be devised and better contact lens materials can be engineered.

Committee: Jason Nichols OD, MPH, PhD (Advisor); Kelly Nichols OD, MPH, PhD (Committee Member); Heather Chandler PhD (Committee Member) Subjects: Ophthalmology

Doctor of Philosophy (Ph.D.), Bowling Green State University, 2023, Photochemical Sciences

The lipid bilayer's integrity is essential for cell function as it acts as the primary barrier against external molecules like drugs and peptides, which can alter the bilayer's physical properties. This dissertation investigates how amphetamine (AMPH) and methamphetamine (METH), and the charged HIV1-TAT peptide impact the stability of lipid bilayers, using a home-built lipid bilayer apparatus that enables real-time monitoring through electrical and fluorescence measurements. Our findings indicate that AMPH and METH increase the lipid bilayer's ion permeability, with METH having a greater destabilizing effect. High concentrations of these stimulants, akin to levels in blood plasma of individuals with stimulant-related brain injuries, lead to pore formation in the bilayer. The extent of destabilization correlated with the drug concentration. We also studied the translocation dynamics of the charged HIV1-TAT peptide across the lipid bilayer. The analysis of current fluctuations showed that successful translocation of the TAT peptide is concentration-dependent, highlighting the significance of charge in inducing membrane deformation or pore formation. Additionally, molecular dynamic simulations were used to explore AMPH interactions with the lipid bilayer in greater detail. The results revealed AMPH's preferred orientation during interaction and its hydrophobic nature, as evidenced by the larger energy barrier encountered in the hydrophilic head group regions of the lipid bilayer. To complement these findings, we utilized surface-enhanced Raman spectroscopy (SERS) to estimate the concentrations of AMPH within lipid bilayers. The data showed a positive correlation between characteristic peak heights and AMPH concentrations. Moreover, whole-cell patch clamp measurements on neuronal cells were employed to examine AMPH's effects in a more intricate lipid environment. This research contributes to the understanding of how stimulants and charged peptides interact with lipid bi (open full item for complete abstract)

Committee: Hong Lu Ph.D. (Committee Chair); Dryw Dworsky Ph.D. (Other); Joseph Furgal Ph.D. (Committee Member); John Cable Ph.D. (Committee Member) Subjects: Chemistry

Doctor of Philosophy (PhD), Ohio University, 2021, Chemical Engineering (Engineering and Technology)

Molecular dynamics (MD) simulation is a computational methodology to probe molecular-level details of physical systems. In MD, the motion of molecules is simulated and from the molecular trajectories, thermodynamic and kinetic properties are ascertained. Machine learning (ML) techniques are a set of computational tools that allow us to identify complex, non-linear relationships in data. ML methods are particularly useful when a system property of interest depends on a large number of variables, and there are no accurate physics-based models relating the property of interest to the variables. ML methods rely on the availability of large datasets to tease out the relationships between the variables. In this research, we have employed MD simulations to study the structural and thermodynamical properties of the simplified plasma membrane of eukaryotic cells, known as the lipid bilayer. We have applied ML methods to study the phase diagram of lipids. Furthermore, we have used ML methods for various corrosion-related applications. By studying asymmetric lipid bilayers using MD simulations, we have shown that in equimolar, asymmetric lipid bilayers, the two leaflets of the bilayer are in tensile and compressive mechanical stress. In response, cholesterol molecules redistribute between the leaflets to relieve these stresses. As a result, the distribution of cholesterol molecules depends on the relative ordering of lipids in the two leaflets. We show that there is a quantitative relationship between cholesterol distribution and the ordering of lipids. We have also studied the distribution of cholesterol molecules in lipid bilayers as a function of temperature and phase of the lipids. We have applied ML to study phase behavior in three-component lipid mixtures and have shown that trained ML methods are quite effective in reproducing the phase diagram of lipids. Furthermore, we have applied ML tools to model the time-dependent corrosion rate of mild steel in presence (open full item for complete abstract)

Committee: Sumit Sharma Associate Professor (Advisor); Douglas Goetz Professor (Committee Member); Gary Weckman Professor (Committee Member); Alexander Neiman Professor (Committee Member); Marc Singer Associate Professor (Committee Member) Subjects: Biomedical Engineering; Chemical Engineering

Master of Science (MS), Ohio University, 2020, Biomedical Engineering (Engineering and Technology)

There is a growing interest in the scientific research community to develop nanoparticles for use in novel commercial and biomedical applications, fueled by recent advances in nanotechnology and nanoparticle synthesis. Potential applications for nanoparticles include use as catalysts during chemical manufacturing processes, use as drug delivery vehicles and imaging agents for biomedical applications, and as surfaces for adsorption during removal of environmental pollutants. The use of nanoparticles in such applications has raised questions concerning their safety and impact on human health. Answers to these questions require a greater understanding of the interactions between nanoparticles and living cells. Models of the cell membrane have been employed to investigate how nanoparticles may adsorb to, fuse with, or penetrate the cell membrane, however careful consideration of the membrane model for such mechanistic studies is necessary. This thesis investigates the role of membrane lipid domains, which are lipid phase segregations comprised of saturated lipids and sterols, in modulating nanoparticle-membrane interactions and further explores how sterol chemistry impacts said interaction. Model membranes were synthesized with an equimolar ratio of sphingomyelin, 1,2-dioleoyl-sn-glycero-3-phosphocholine, and varied sterol composition to yield vesicles with varied lipid domain properties. Fluorescence anisotropy and Forster resonance energy transfer of fluorescent probes was measured to quantify the degree of ordered domain formation in model vesicles. Additionally, confocal microscopy was performed to visualize lipid domains. Following lipid domain characterization, vesicles in which a self-quenching fluorescent dye was encapsulated were exposed to plain silica nanoparticles (diameter 37.5 ± 1.8 nm) and leakage of dye was measured to determine the degree of membrane disruption. By analyzing the results of vesicle leakage assays alongside the results from domain charact (open full item for complete abstract)

Committee: Amir Farnoud (Advisor); Douglas Goetz (Committee Member); Sumit Sharma (Committee Member); Shiyong Wu (Committee Member) Subjects: Biomedical Engineering; Biophysics; Engineering; Nanoscience

Doctor of Philosophy, University of Toledo, 2016, Engineering

A major challenge to the economic viability of outdoor cultivation of microalgae is the high cost of CO2 supply, even when microalgae farms are co-located with point sources of CO2 emissions. In addition, the global capacity for algae biofuel generation is severely restricted when farm locations constrained by proximity to CO2 sources along with the additional limitations of low slope lands and favorable climate. One potential solution to the impediments of CO2 cost and availability is through the cultivation of microalgae in highly alkaline pH solutions (pH>10) that are effective at scavenging CO2 from the atmosphere at high rates. The extreme alkaline pH media would also mitigate culture crashes due to microbial contamination and predators. In this thesis, we report the indoor and outdoor cultivation of a microalgae isolate (Chlorella sp. str. SLA-04) adapted to grow in unusually high pH environments. The isolate was cultivated in a growth medium at pH>10 without any inputs of concentrated CO2. Initial cultivation studies (both indoor and outdoor) resulted in biomass and lipid productivities that were comparable to those reported for other microalgae cultures cultivated in near-neutral media (pH 7-8.5). SLA-04 cultures also showed high lipid productivity and high glucose-to-lipid conversion efficiency when cultivated indoors mixotrophically in the presence of glucose as an organic carbon source. Following this, experiments were performed to determine the effect of pH and bicarbonate (HCO3-) concentrations on biomass productivity of str. SLA-04. Increased HCO3- concentrations in the culture medium resulted in an increase in the overall biomass productivity of str. SLA-04 (22 g-biomass·m-2·d-1). Simultaneously, the high medium pH (pH >10) led to increased mass transfer rates of CO2 from the atmosphere. The improved CO2 uptake rates from the atmosphere resulted in the replenishment of HCO3- utilized during the growth of SLA-04. The quantum yields and photosyn (open full item for complete abstract)

Committee: Sridhar Viamajala (Committee Chair); Sasidhar Varanasi (Committee Member); Patricia Relue (Committee Member); Arunan Nadarajah (Committee Member); Ronald Fournier (Committee Member) Subjects: Alternative Energy; Chemical Engineering

PHD, Kent State University, 2016, College of Arts and Sciences / Department of Biological Sciences

In the past, lipid droplets were considered as inert cell organelles that store cellular energy in the form of neutral lipids. But today, they are known as cell organelles carrying out a myriad of functions required for the sustenance of life. The structure of intracellular lipid droplets closely resembles that of extracellular particles called lipoproteins. Both structures contain a core of neutral lipids surrounded by a monolayer of phospholipids and proteins. However, their functional roles within living organisms are different, ie, lipoprotein particles transport neutral lipids extracellularly in an organism's circulatory system, whereas lipid droplets store neutral lipids intracellularly. These two structures differ in composition as well. However, literature shows that proteins found on the surface of lipoproteins and lipid droplets share a high degree of sequence and structural similarity. Although structural and functional details of lipoproteins and their components have been widely studied in the past, many structural and functional aspects of lipid droplets and their protein components are still unclear. This work describes the lipid binding properties of two structurally homologous proteins carrying amphipathic alpha helix bundles, apoLp-III and perilipin 3 which are found across two different biological systems, namely, lipoproteins in insects and lipid droplets in humans. Both these proteins are exchangeable in their lipid binding behavior. Studying the lipid interactions of the above proteins is of key importance to understand the basics of neutral lipid metabolism and lipid transport of multicellular organisms. Large quantities of recombinant proteins required for our biophysical characterization studies were expressed in E. coli cells and purified using a combination of chromatography techniques. In order to study the lipid interactions of apoLp-III and perilipin 3, we used Langmuir monolayers composed of various types of phospholipids. Our monolay (open full item for complete abstract)

Committee: Edgar Kooijman (Advisor); Srinivasan Vijayaraghavan (Committee Member); Derek Damron (Committee Member); Soumitra Basu (Committee Member); Bansidhar Datta (Other) Subjects: Biophysics

Doctor of Philosophy, The Ohio State University, 2012, Chemical and Biomolecular Engineering

The field of nucleic acid based therapeutics offers treatments for diseases at the most basic level of cellular biochemistry and its potential is boundless. Oligonucleotide therapeutics is also an invaluable tool in studying cellular processes and the precise effect certain genes have on disease models. Since passive cellular uptake of these highly charged molecules is low, an effective delivery method is imperative in order to achieve an observable effect. Lipid based nanoparticles with their numerous, well-documented advantages are often used for delivery and in this work, both pharmaceutical and engineering based approaches were explored to construct lipoplex nanoparticles. From the pharmaceutical side, the effect of lipid chirality on transfection efficiency was investigated. From an engineering perspective, even though liposomal nanoparticles represent an especially promising class of drug delivery vectors, conventional bulk mixing preparation methods have room for improvement with regards to yield and consistent control over structure and composition. Therefore, a device-based approach using arrays of microwells was studied to not only deliver particles directly to cells in a more controlled way but also to introduce control over the typically uncontrolled complexation process of lipids and nucleic acids. DOTAP, as a racemic mixture, is a cationic lipid and a widely used transfection reagent. In this study, racemic and enantiomerically pure DOTAP were used in lipoplex formulations to deliver siRNA to MCF-7 cells, targeting the aromatase enzyme. The R enantiomer of DOTAP was found to be more efficacious than the S enantiomer or the racemate when used in combination with cholesterol. Specifically, the aromatase activity of cells treated with R-DOTAP lipoplexes was 50% lower than those treated with S-DOTAP or racemic lipoplexes at a 10 nM siRNA concentration. In other words, R-DOTAP lipoplexes were twice as effective at the low concentration. Amongst the DOTAP en (open full item for complete abstract)

Committee: L. James Lee (Advisor); Robert J. Lee (Advisor); Jeffrey J. Chalmers (Committee Member); Zhonga Liu (Committee Member) Subjects: Chemical Engineering

Master of Science in Nutrition and Dietetics, University of Akron, 2010, Nutrition and Dietetics

Committee: Lonnie Lowery Dr. (Committee Chair); Sandra Hudak Dr. (Committee Member); Deborah Marino Dr. (Committee Member) Subjects: Nutrition

Doctor of Philosophy in Regulatory Biology, Cleveland State University, 2024, College of Arts and Sciences

Eukaryotic initiation factor 2A (eIF2A) is a highly conserved 65 kDa eukaryotic protein that functions in minor initiation pathways, which affect the translation of only a subset of messenger ribonucleic acid (mRNAs), such as internal ribosome entry site (IRES)-containing mRNAs and/or mRNAs harboring upstream near cognate/non-AUG start codons. These non-canonical initiation events are important for regulation of protein synthesis during cellular development, differentiation and/or the integrated stress response. Selective eIF2A knockdown in cellular systems was shown to inhibit translation of such mRNAs, which rely on alternative initiation mechanisms for their translation. However, there existed a gap in our understanding of how eIF2A functions in mammalian systems in vivo (on the organismal level) and ex vivo (in cells). To address this question we have developed the eIF2A-total body knockout (KO) mouse model. Using this model, we presented evidence implicating eIF2A in the biology of aging and metabolic syndrome. We discovered that eIF2A-KO mice have reduced life span and that eIF2A plays an important role in maintenance of lipid homeostasis, the control of glucose tolerance and insulin resistance. We also showed the eIF2A KO affects male and female mice differently, suggesting that eIF2A may affect sex-specific pathways. The metabolic syndrome phenotype has three main etiological categories: obesity and disorders of adipose tissue (particularly increased size of adipocytes); glucose intolerance and insulin resistance; and a constellation of independent factors of hepatic, vascular, and immunologic origin and all of these features were observed in the eIF2A-KO mice. Increased adipocyte size is, in particular, well known to be positively correlated with impaired insulin sensitivity and glucose tolerance leading to metabolic syndrome. To specifically check whether the absence of eIF2A in adipose tissue is responsible for metabolic abnormalities observed in the tota (open full item for complete abstract)

Committee: Anton Komar (Committee Chair); Barsanjit Mazumder (Committee Member); Roman Kondratov (Committee Member); William Baldwin III (Committee Member); Merlin Gnanapragasam (Committee Member); William Merrick (Committee Member) Subjects: Biology; Molecular Biology; Physiology

Master of Sciences, Case Western Reserve University, 2025, Pathology

Apnea of prematurity (AOP) is a common cause of short- and long-term morbidity in preterm infants. There is a growing body of evidence supporting the association of apnea in neonates with hyperbilirubinemia. An area of rapid development in preterm infants is the nucleus tractus solitarius (NTS), which is implicated in the control of respiratory drive. The large (BK) and small (SK1,2,3) conductance calcium- and voltage-activated K+ channels are lipid raft associated proteins that are important in regulating the neuronal activity of the NTS. The percent of BK, SK1, SK2, and SK3 found in lipid rafts of P6 jj Gunn rat pup NTS is significantly reduced after treatment with SDMX. These results demonstrate one mechanism through which bilirubin may perturb neuronal activity of the NTS leading to AOP in preterm infants.

Committee: Cynthia Bearer (Committee Chair); Peter MacFarlane (Committee Member); Ganapati Mahabaleshwar (Committee Member); Pamela Wearsch (Committee Member) Subjects: Medicine; Neurobiology

PhD, University of Cincinnati, 2024, Engineering and Applied Science: Chemical Engineering

Biomembranes play a crucial role in the structure and functions of all cells. The presence of amphiphilic co-solvents leads to a disruption of these structures, leading to cell stress and ultimately, cell death. A proposal has been put forth that amphiphilic co-solvents also impact the lateral structure of biomembranes, leading to changes in membrane organization – a new mode of solvent induced cellular stress that has not yet been investigated. In this dissertation, I have undertaken the investigation into this phenomenon through two major tasks, developing tools to study the membrane structure and directly investigating the effect of co-solvents on membrane structure. The primary method for assessing membrane structure is mainly neutron scattering, along with supporting information from molecular dynamics simulation and fluorescence. These results would primarily impact applications in industrial fermentation and biofuels production, with additional potential implication in the mechanisms of alcohol-related gastrointestinal tract and other cancers.

Committee: Jonathan Nickels Ph.D. (Committee Chair); Anastasios Angelopoulos Ph.D. (Committee Member); George Stan Ph.D. (Committee Member); Vadim Guliants Ph.D. (Committee Member); Gregory Beaucage Ph.D. (Committee Member) Subjects: Biophysics

Doctor of Philosophy, Case Western Reserve University, 2024, Biomedical Engineering

Cancer therapy has significantly advanced over the past few decades due to development of improved surgical technique, early detection, and novel therapy development. However, cancer remains one of the most lethal diseases due to metastasis, drug resistance, and recurrence. Immunotherapy holds the promise of utilizing the body's immune system to eliminate cancer cells. Immune checkpoint blockade therapy remains the most common utilization of immunotherapy in the clinical but yields varying degrees of success due to the massively immunosuppressed tumor microenvironment. Discovery of novel immune checkpoints and technology for targeting these checkpoints is critical for the advancement of cancer immunotherapy. The novel immune checkpoint protein VISTA (V-domain Immunoglobulin Suppressor of T cell Activation) impairs the toll-like receptor (TLR)-mediated activation of myeloid antigen presenting cells, promoting the expansion of myeloid derived suppressor cells, and suppressing tumor-reactive cytotoxic T cell function. Gene therapy targeting the VISTA checkpoint protein in conjunction with potent TLR agonists represents a safer yet potent alternative to antibody mediated cancer immunotherapy that has shown toxicity in the clinic. The overall objective of the work in this dissertation is to develop nanoparticle platforms for delivery of gene therapy mediate immune checkpoint blockade cancer immunotherapy regiments. First, a dual action lipid nanoparticle (Dual-LNP) that incorporates a VISTA-specific siRNA and a TLR9 agonist (unmethylated CPG) is developed. The Dual-LNP ensures co-delivery of both cargoes to tumor-infiltrating myeloid cells, leading to simultaneous silencing of VISTA and stimulation of TLR9. Next, the efficacy of the Dual-LNP was tested in multiple solid tumor models. The Dual-LNP treatment achieved a high cure rate in colon carcinoma MC38, melanoma B16F10, and YUMM1.7 models (83%, 60%, and 48%, respectively). Finally, investigation into the immune landsc (open full item for complete abstract)

Committee: Efstathios Karathanasis (Advisor); James Basilion (Committee Chair); Li Lily Wang (Committee Member); Andrew Shoffstall (Committee Member); William Schiemann (Committee Member) Subjects: Biomedical Engineering; Nanotechnology

Doctor of Philosophy in Regulatory Biology, Cleveland State University, 2024, College of Arts and Sciences

Fasting (F) offers many metabolic benefits but interestingly induces hepatic steatosis. Many fasting metabolic adaptations have largely been studied under experimental conditions where food is unexpectedly withheld. Such random fasting is in sharp contrast with Calorie restriction (CR), in which mice self-impose a periodic fasting cycle and spend most of their day without the food. Despite long fasting durations, CR mice are protected from hepatic steatosis via unclear mechanisms. Here, we utilized transcriptomic profiling to identify important players in the differential effect of CR and F on liver steatosis. Serum non-esterified fatty acids were elevated to similar concentrations by both CR and F. Both diets induced hepatic β-oxidation but the magnitude and kinetic were stronger in F than in CR. This revealed that liver energy utilization is a discriminating factor between both diets but did not explain differences in liver TAG accumulation. Our unbiased transcriptomic analysis identified some candidate genes that were induced by F but not CR, these genes code for liver fatty acid (FFA) transporters (Slc27a1 and Slc27a2), TAG synthesis (Gpat4), and lipid storage (Plin2 and Cidec). Correspondingly, liver FFAs increased in F (C16:0, C18:0, C18:1, C18:2), while only C18:2 was increased in CR. F increased liver TAG and lipid droplet accumulation while decreased profiles were observed in CR. We found that the tight control of liver lipid homeostasis in CR was achieved through entrained anticipation of the feeding/fasting cycle in CR. Specifically, when CR mice miss their anticipated daily meal or have an impaired circadian clock (Cry1,2 ablation), they began to accumulate liver TAG and induce metabolic profiles similar to F mice. Our study positions entrainment by the circadian clock in the mechanisms of CR.

Committee: Roman Kondratov (Advisor); Srinivasan Dasarathy (Committee Member); Crystal Weyman (Committee Member); Aaron Severson (Committee Member) Subjects: Molecular Biology

Doctor of Philosophy (Ph.D.), University of Dayton, 2024, Biology

Vertebrate freeze tolerance is an extraordinary phenomenon in which up to 70% of an animal's extracellular fluids are converted to ice while circulation, respiration, and neuronal function are all simultaneously suspended during freezing. Upon thawing, animals must tolerate the resumption of physiological function, restore intracellular fluid volumes, and repair injuries that occurred during freeze-thaw. The mechanisms that enable animal freeze tolerance vary by species and represent a myriad of biochemical, cellular, systems, and organismal strategies. Cope's gray treefrog Dryophytes (Hyla) chrysoscelis is a freeze tolerant anuran that repeatedly freezes and thaws each winter in part by utilizing a complex system of cryoprotectants including glycerol, glucose, and urea. Intracellular transport is facilitated by specialized aquaglyceroporin proteins that enable transmembrane movement of water and the cryoprotectants glycerol and urea during freeze-thaw. The physiological mechanisms that enable freeze tolerance in D. chrysoscelis are not entirely understood and cannot be explained by cryoprotectant accumulation alone. The aim of this dissertation is to explore the unknown physiological mechanisms of freeze tolerance in D. chrysoscelis by using an integrative perspective that incorporates all levels of biological organization. Novel experimental protocols were used to evaluate the ecophysiological effects of repeated freezing and thawing, characterize organismal responses to seasonal and cold acclimation, and determine the effects of cold acclimation and freeze-thaw cycles on membrane lipid composition using 1H-NMR analysis. The results from these studies emphasized the complexity of freeze tolerance in D. chrysoscelis and revealed several novel aspects of freeze tolerance in this species including dynamic blue and green dorsal coloration in frozen and thawing frogs, “freeze resistance” in a freeze tolerant vertebrate, evidence of seasonal (open full item for complete abstract)

Committee: Carissa Krane (Advisor); David Goldstein (Committee Member); Thomas Williams (Committee Member); Yvonne Sun (Committee Member); Amit Singh (Committee Member) Subjects: Biochemistry; Biology; Cellular Biology; Ecology; Molecular Biology; Organismal Biology; Physiology

Master of Science (MS), Ohio University, 2024, Biological Sciences (Arts and Sciences)

Eastern Hellbenders (Crytobranchus a. alleganiensis) have suffered enigmatic, range- wide declines over the past decades. Persisting populations are skewed towards larger, older adults, suggesting that reduced recruitment is responsible for these declines, with degraded water quality, specifically elevated conductivity, implicated as a main contributor. Successful fertilization and the resilience of eggs under high conductivity conditions suggest deleterious effects during larval development. We experimentally assessed the effects of chronic exposure to elevated conductivity (1000 μS/cm) on wild Eastern Hellbender larvae hatched in a lab, as well as the effects of switching from low conductivity (100 μS/cm) to high, and vice versa, on Eastern Hellbenders during early larval development. We assessed mortality over 72 days post-hatching, with half of the larvae switched from their original conductivity treatments to the other over five days beginning at 33 days post-hatching. Chronic exposure to elevated conductivity resulted in significant mortality. Additionally, switching larvae from low conductivity to high resulted in increased mortality, while switching larvae from high conductivity to low increased survival. We also assessed larval morphology and swimming performance and found significant negative effects of chronic exposure to elevated conductivity on both body mass and multiple measures of morphology (length and width). We observed similar effects in animals switched from low conductivity to high, while switching animals from high conductivity to low resulted in only marginally increased mass and morphological measures, demonstrating their inability to compensate for initially depressed growth rates even after being returned to more optimal conditions. Despite altered size and morphology, elevated conductivity did not impact locomotor performance, though switching conductivities, regardless of direction, did result in increased burst distance. We measured who (open full item for complete abstract)

Committee: Shawn Kuchta (Advisor); Viorel Popescu (Committee Member); Kelly Johnson (Committee Member) Subjects: Biology; Conservation; Ecology; Endocrinology; Freshwater Ecology; Molecular Biology; Morphology; Organismal Biology; Wildlife Conservation

Doctor of Philosophy, Case Western Reserve University, 2024, Biomedical Engineering

Much of the human genome contains genes that do not encode proteins, but that are still transcribed into RNA that has essential functions toward regulating gene expression. Among these are long noncoding RNAs (lncRNA), a heterogeneous group of RNAs over 200 nucleotides in length that are often expressed in tissue-, disease-, or developmental stage-specific manners. In addition to its normal roles, lncRNA has been found to be deregulated in multiple human cancers, particularly aggressive ones like non-small cell lung cancer (NSCLC) and triple negative breast cancer (TNBC). NSCLC and TNBC are among the leading causes of cancer-related death globally, in part due to a lack of curative treatment options in a disease known to develop resistance to chemotherapies. Regulating the expression of oncogenic lncRNAs through RNA interference (RNAi) with small interfering RNA (siRNA) could serve as a promising strategy to inhibit tumor growth in aggressive cancers. To achieve the maximum benefit from siRNA therapy, it requires a proper delivery system to reach the site of interest. At the same time, non-invasively monitoring the effects of the therapy can assist in treatment planning and determining its success. This dissertation focuses on delivering siRNA against a long noncoding RNA called DANCR, which is overexpressed in both NSCLC and TNBC, using a pH-sensitive amino lipid carrier, ECO, along with applying magnetic resonance molecular imaging (MRMI) to track the treatment progress. Silencing DANCR with ECO/siDANCR nanoparticles was previously shown to inhibit TNBC cell migration and invasion, but its effects in NSCLC had not been evaluated. Further, ECO/siDANCR nanoparticles were optimized to target a protein abundant in the tumor extracellular matrix but limited in normal tissue, extradomain B fibronectin (EDB-FN). A peptide specific to EDB-FN, ZD2, was previously developed and applied in this work for targeting ECO nanoparticles. At the same time, an EDB-targeted MRI co (open full item for complete abstract)

Committee: Zheng-Rong Lu (Advisor); William Schiemann (Committee Member); Horst von Recum (Committee Member); Samuel Senyo (Committee Chair) Subjects: Biomedical Engineering

PHD, Kent State University, 2023, College of Arts and Sciences / Department of Biological Sciences

Anionic signaling lipids (ASL) play multiple crucial roles in the cell. They perform their signaling functions by interacting with (macro)molecules in their vicinity. The headgroups of ASL possess unique physicochemical properties (ionization, hydrogen bond formation, hydration, etc.) that mediate specific interactions in the cell. The physicochemical properties of ASL are modulated based on the characteristics of lipids that surround the ASL as well as changes to conditions in the cytosol. To understand the functions of ASL in the cell, it is first vital to determine their inherent behavior under normal physiological conditions as well as changes to cytoplasmic and membrane properties that occur during signaling and in diseases. In this study, 31P Nuclear Magnetic Resonance was primarily used to investigate the biophysical properties of some ASLs and their interactions with important molecules in the cell that have been overlooked so far. The interactions of phosphatidic acid (PA) with methylated phosphatidylethanolamine (PE) derivatives, increasing cholesterol concentrations, and with the secondary signaling lipid diacyl glycerol pyrophosphate (DGPP) were investigated. We also looked into the interactions of DGPP with cationic molecules. Additionally, we investigated the ionization properties of the 3 monophosphoinositides (PIxP) namely phosphatidylinositol 3-phosphate, phosphatidylinositol 4-phosphate and phosphatidylinositol 5-phosphate in phosphatidylcholine (PC) and in equal concentrations of PC and PE model membranes. For PA, we find that the presence and the number of methyl groups on the methylated PE derivatives differentially affect its ionization properties. Higher levels of cholesterol impact the properties of the membrane to affect the chemical shift of PA, but similar higher cholesterol levels, surprisingly, do not induce significant changes in the pKa2 of PA. The ionization properties of PA and DGPP are both increased when the two lipids are present (open full item for complete abstract)

Committee: Edgar Eduard Kooijman PhD (Advisor); Thorsten-Lars Schmidt PhD (Committee Chair); Manabu Kurokawa PhD (Committee Member); Gary Koski PhD (Committee Member); Elizabeth Mann PhD (Committee Member) Subjects: Biochemistry; Biophysics

Master of Science, University of Akron, 2023, Biology

The zebrafish (Danio rerio) is an important model organism in human related research. Although commonly used in lab settings, there is a lack of consistency in diets fed to cohorts. This inconsistency is amplified by an incomplete understanding of the impact on offspring because of parental diets. Due to the importance of this animal model in human related studies, we aimed to explore the potential impacts from diet on adult behavior and offspring development through evaluation of feeding preference and the amount of yolk provided to developing embryos. Utilizing a 3D printed arena and machine learning, a spatial preference was seen and was further linked to specific food items. This provided encouragement for the use of machine learning and updated technology to further understand zebrafish behavior. During extended feeding, diets fed to adult female zebrafish resulted in weight gain, variation in standard length, and differences in the yolk to chorion ratios for each of the treatments. High carbohydrate diets impacted the ability for females to gain weight at the same rate of the control, high protein, and high lipid diets. However, the ratio of yolk to the chorion of the eggs for the high carbohydrate diet, high protein diet, and the control diet were significantly higher than the high lipid diet, regardless of the production of eggs following spawning events.

Committee: Brian Bagatto (Advisor); Todd Blackledge (Committee Member); Richard Londraville (Committee Member) Subjects: Animals; Aquatic Sciences; Biology; Physiology