Full text for dissertations and theses included in this collection dates back to 2011. For older dissertations, check the library’s catalog CatalogUSMAI or Dissertations and Theses.

Recent Submissions

  • Effect of Excipients on the Performance of Spray-dried Amorphous Solid Dispersion (ASD) in Tablets

    Yu, Dongyue; Hoag, Stephen W. (2022)
    Amorphous solid dispersions (ASD) are a proven method of improving the solubility and bioavailability of poorly soluble drugs. Immediate-release tablets are frequently used as the final dosage form for ASDs. The selection of polymers and excipients is critical for the manufacturability and bioavailability of ASD tablets. ASDs were prepared by spray drying; ASD tablets were then generated using a compaction simulator. We first studied the impact of polymer types and drug-polymer ratios on bulk powder properties, morphologies, and compaction behaviors of ASDs. Itraconazole (ITZ) and indomethacin (IND) were used as model drugs, and two polymers were used: hydroxypropyl methylcellulose acetate succinate (HPMCAS) and polyvinylpyrrolidone (PVP). The results indicated that the tabletability increased with decreasing drug loadings, except for ITZ-PVP ASDs. Multivariate analysis revealed that particle surface area was the most significant factor influencing the tensile strength of ASD tablets. Secondly, the contact angle and surface free energy of ITZ ASD tablets containing different HPMCAS grades and drug loadings were evaluated using a Drop Shape Analyzer. A larger contact angle was correlated with a higher dissolution rate, suggesting that contact angle could be a high throughput tool for screening ASDs formulations. Lastly, we investigated the influence of fillers such as microcrystalline cellulose, lactose, mannitol, and starch on drug release and stability of ITZ-HPMCAS ASDs. We discovered that the dissolution performance and physical stability of tablets were influenced by the choice of filler. The results and inferences drawn from this research will provide valuable insights into ASD formulation development downstream tablet production.
  • Novel Bio-interactive Fixed Dental Restoration Cement with Potent Antibacterial and Remineralization Properties

    AlSahafi, Rashed A.; Weir, Michael D.; Xu, Huakun H. (2022)
    Resin-based cement is increasingly used in clinical practice due to its excellent mechanical and physical properties. The cementation and accurate placement of fixed dental restorations with an ideal marginal adaptation to the tooth structure remains a challenging laboratory and clinical task. The marginal gap of fixed dental restoration increases the opportunity for microleakage, plaque accumulation, and tooth demineralization by increasing the potential for food buildup around the margin and exposing the tooth-cement interface to the oral cavity. Thus, there is an increased need to develop a new generation of bio-interactive dental cement with antibacterial, long-term remineralization abilities, and excellent mechanical properties. Therefore, this dissertation aims to invent new bio-interactive resin-based cement containing dimethylaminohexadecyl methacrylate (DMAHDM), nanoparticles of amorphous calcium phosphate (NACP), and nanoparticles of calcium fluoride (nCaF2), which could be a promising approach to increase the chances of success of fixed dental restoration and strengthen tooth structures. All new cement formulations were subjected to a series of mechanical, antibacterial, and ion release assessments. In the first manuscript, we found that the new NACP+DMAHDM cement has excellent potential for fixed restoration cementation, as it efficiently inhibited S. mutans biofilm commonly associated with secondary caries and maintained an excellent mechanical property with high levels of Ca and P ions released. In the second manuscript, we found that incorporating DMAHDM and NACP into resin-based cement provides strong antibacterial action against saliva microcosm biofilm and presents a high level of Ca and P ion recharge abilities. In the third manuscript, we found that the new cement with both NACP and nCaF2 demonstrated the advantages of both types of bio-interactive fillers as it could release a higher level of ions than the resin cement with only nCAF2 and exhibits a better rechargeability compared to the resin cement with only NACP. Lastly, in the fourth manuscript, we found that the novel antibacterial low-shrinkage-stress resin-based cement provided strong antibacterial action and maintained excellent mechanical properties with reduced polymerization shrinkage stress, which could improve the long-term success of the fixed dental restoration.
  • The Clinical Translation of Cardiac Xenotransplantation

    Goerlich, Corbin; Mohiuddin, Muhammad M.; Singh, Nevil (2022)
    Patients with end-stage heart failure requiring heart transplantation die simply because allografts are in short supply. Cardiac xenotransplantation from genetically-modified pig source animals has been proposed to bridge the gap between supply and demand. However, there are two predominant barriers to clinical translation and are poorly understood. The first, is a type of primary graft dysfunction, termed perioperative cardiac xenograft dysfunction (PCXD), which causes failure of the xenograft within 48 hours after transplantation. The second, post-transplantation xenograft growth, which causes a life-limiting diastolic heart failure within the first month after transplantation. We demonstrate that PCXD can be overcome with cardiac preservation techniques that minimize ischemia, either with blood-based cardioplegia induction or non-ischemic continuous preservation (NICP). In a heterotopic PCXD model, we further demonstrate that PCXD is likely a phenomenon rooted in xenograft dysfunction resulting from activation of innate immunity within the xenograft. While further studies need to be done, we demonstrate evidence that PCXD is augmented by the synergistic inflammatory effects of both cardiopulmonary bypass and cross-species transplantation. We also provide evidence that polymorphisms from TLR4 of S. scrofa (swine) compared to H. sapiens, may explain a possible target for the unique inflammatory signaling, that results in xenograft dysfunction after cardiac xenotransplantation. We also demonstrate that post-transplantation xenograft growth is multifactorial, but can be limited by growth hormone receptor knockout donors, along with genetic modifications that reduce immunogenicity of the xenograft. This growth, within 6 months after transplantation, is not a result of physiologic mismatch or cardiomyocyte hypertrophy. Therefore, recipients do not need to be treated for tachycardia and hypertension as previously thought. Lastly, we demonstrate the clinical translation of cardiac xenotransplantation by applying knowledge and expertise obtained from these studies. Expanded access (“compassionate use”) FDA authorization was based on demonstration of principle from our preclinical model. An ECMO-dependent patient without other therapeutic options was transplanted a genetically-modified cardiac xenograft, with 10-gene edits, combined with non-ischemic cardiac preservation and anti-CD40 monoclonal antibody-based immunosuppression. The patient was able to successfully wean from ECMO, participate in active rehabilitation and survived 60 days after transplantation without evidence of rejection.
  • Exploring Child Welfare Placement Pathways to Residential Treatment Settings: A Three Paper Dissertation

    McCarthy, Lauren; Lee, Bethany R. (2022)
    Residential treatment settings (RTS) provide access to services in a safe environment for child welfare involved youth with behavioral health challenges. Despite the potential benefits of RTS, there are concerns that have led to legislation aiming to reduce their use. Policy that aims to reduce the use of RTS should be informed by an understanding of how youth enter RTS. This three-paper dissertation aimed to increase our understanding of how youth placement histories are related RTS entry in the context of youth behavior and development using secondary analysis of state administrative child welfare data. This dissertation also aimed to understand how caregivers experience accessing mental health care for youth through interviews. The first paper explored aggregate patterns of placement transitions and individual factors associated with risk of RTS entry. Findings include that transitions are most likely to occur between similar types of placement settings and that developmental period at first entry to out-of-home care is associated with RTS entry. The second paper explored the presence of unobserved subgroups of child welfare involved youth based on placement histories and whether youth move through these groups over time. The second paper found two subgroups of placement histories, multiple placements in group settings and stability in family care settings. The second paper further found that child behavior and developmental period at first entry to care were associated with group membership and transitions. The third paper identified that caregivers gain empowerment in decision making when accessing mental health services, but that this empowerment declines when accessing acute inpatient services. The third paper further found that accessing RTS was a challenging journey that impacted caregiver well-being, and that the decision to place a child in RTS came following a safety inflection point. Implications include that youth who enter out-of-home care in pre- and early adolescence and their caregivers may require additional support to remain stable in family placement settings. Further implications include the need to improve caregiver access to knowledge about the mental health care system to improve empowerment when making decisions and more resources for acute mental health service settings.
  • Investigating the Role of Microglia and Extracellular Vesicles in Spinal Cord Injury-Induced Brain Dysfunction

    Khan, Niaz; Faden, A. I. (2022)
    Spinal cord injury (SCI) causes brain neurodegeneration leading to cognitive and affective changes, including memory loss and mood alterations. Using SCI models, we and others have demonstrated that progressive neurodegeneration is accompanied by neuroinflammation, including sustained microglial activation. The primary goal of this dissertation was to test the hypothesis that SCI triggers brain microglia-mediated neuroinflammation and secondary neurological dysfunction and to study the underpinning mechanisms including changes in systemic and central extracellular vesicles (EVs). First, we probed the mechanisms responsible for microglia activation and examined the effect of pharmacological depletion of microglia on posttraumatic neuropathology and cognitive/depressive-like behavior in a mouse SCI model. Microglial depletion significantly improved neuronal cell loss in key brain regions and associated cognitive/depressive-like behavioral outcomes after SCI. The transcriptomes of the spinal cord and brain were also substantially altered, supporting our hypothesis that microglia significantly contribute to changes related to inflammation, neurotransmission, and apoptosis after SCI. Second, we studied changes in circulating EVs after SCI. EVs are biological nanoparticles released from cells that contribute to intercellular communication and can become altered with disease. We found a significant increase in plasma tetraspanin CD81+ EVs after SCI at 1d post-injury. Surface CD81 was decreased on astrocytes at the injury site, suggesting that these cells may release CD81+ EVs into circulation. Total plasma EV microRNA content was also significantly modified, similar to the profile previously described in inflammatory astrocyte EVs. Notably, when injected into the cerebroventricular system, plasma EVs from SCI mice increased brain expression of several inflammatory genes, including markers of astrocyte reactivity. Finally, we examined the brain transcriptional profile and EV changes 19 months post-SCI in male and female mice. While we observed strong sex-dependent differences in the overall brain transcriptome after SCI, the homeostatic microglial phenotype was reduced in both sexes. Chronic SCI increased EV count in the brain and modified their microRNA content, which may explain the observed transcriptional changes. Plasma EV markers were also elevated late after injury, especially in males. Collectively, these experiments are the first to characterize EV dynamics after SCI and suggest that EVs may be involved in posttraumatic brain inflammation.
  • The Regulatory Role of the Cytoplasmic Heme Binding Protein PhuS in Pseudomonas aeruginosa

    Wilson, Tyree; Wilks, Angela (2022)
    Pseudomonas aeruginosa is an opportunistic pathogen that requires iron for its survival and virulence. P. aeruginosa can acquire iron from heme via the nonredundant heme assimilation system and Pseudomonas heme uptake (Phu) systems. Heme transported by either system is eventually sequestered by the cytoplasmic protein PhuS, which specifically shuttles heme to the iron-regulated heme oxygenase HemO. Furthermore, a conformational rearrangement upon heme binding is necessary for the protein-protein interaction with HemO and a ligand switch between the heme coordinating ligands (His209 and His212) was proposed ot be required for translocation of heme to HemO. As the PhuS homolog ShuS from Shigella dysenteriae was observed to bind DNA as a function of its heme status, we sought to further determine if PhuS, in addition to its role in regulating heme flux through HemO, functions as a DNA-binding protein. Herein, through a combination of chromatin immunoprecipation-PCR, EMSA, and fluorescence anisotropy, we show that apo-PhuS but not holo-PhuS binds upstream of the tandem iron- responsive sRNAs prrF1, F2. Previous studies have shown the PrrF sRNAs are required for sparing iron for essential proteins during iron starvation. Furthermore, under certain conditions, a heme-dependent read through of the prrF1 terminator yields the longer PrrH transcript. Quantitative PCR analysis of P. aeruginosa WT and ΔphuS strains shows that loss of PhuS abrogates the heme-dependent regulation of PrrF and PrrH levels. Taken together, our data show that PhuS, in addition to its role in extracellular heme metabolism, can also modulate PrrF and PrrH levels in response to heme. The dual function of PhuS is central to integrating extracellular heme utilization into the PrrF/PrrH sRNA regulatory network that is critical for P. aeruginosa adaptation and virulence within the host. Additional biophysical, genetic and metabolic approaches have been conducted to determine the role of the PhuS heme coordinating residues regulate the mutual exclusivity of heme and DNA binding and the resulting effects on PrrF and PrrH expression.
  • Characterization of Temperature Dependent Activity in a Model Polyextremophilic Beta-galactosidase Enzyme Through Kinetic and Structural Analysis

    Laye, Victoria; DasSarma, Shiladitya (2022)
    The Antarctic haloarchaeon, Halorubrum lacusprofundi, contains a polyextremophilic family 42 β-galactosidase, which we are using as a model for polyextremophilic enzymes. Divergent amino acid residues in this 78 kDa protein were identified through comparative genomics and hypothesized to be important for cold activity. Six amino acid residues were previously mutated and five were shown by steady-state kinetic analysis to have altered temperature-dependent catalytic activity profiles via effects on Km and/or kcat compared to the wild-type enzyme. Double-mutated enzymes were constructed and tested for temperature effects from 0-50 ºC, including two new tandem residue pairs, two potential loop insertions, and pairwise combination of the single residue mutations. The observed temperature and salinity dependent kinetic effects were compared to root-mean-square fluctuations to determine structural mechanisms of polyextremophilic activity. All the mutated enzymes were found to be more catalytically active at higher and/or less active at colder temperatures compared to the wild-type, with both Km and kcat effects observed for the two tandem mutations. For pairwise combinations, a Km effect was seen when the surface-exposed F387L mutation located in a domain A TIM barrel α helix 19 Å from the active site was combined with two internal residues, N251D or V482L. When another surface-exposed residue, I476V, was paired with N251D or V482L, primarily a kcat effect was observed. The temperature dependent kinetic effects were continued to 25-50 ºC with a dominant Km effect observed in all mutated enzymes. By deleting the identified insertions/loops, we are able to determine the kinetic effects of these insertions. The domain B deletion resulted in a less active enzyme overall converging with the wild-type at higher temperatures while the domain C deletion resulted in reduced cold-activity and improved activity at higher temperatures. Domain B may confer cold-activity without changing the thermal stability while domain C confers cold activity, at least in part, at the expense of activity at higher temperatures. With molecular dynamics simulations, increased flexibility was observed with higher temperature and salinity and in the mutations, indicating that the enzyme’s function is related to its flexibility and there is a balance required for optimal polyextremophilic activity.
  • Visual Deficits in a Model of Gestational Hypothyroidism

    Tarasiewicz, Agnieszka; Medina, Alexandre E. (2022)
    Hypothyroidism prevalence among pregnant women is between 0.5 to 4% (Carney et al., 2014). Most studies look at the prolonged or severe reduction in thyroid hormone (TH) levels. We looked at how the reduced levels of TH during the third trimester of human gestation and the first weeks after birth in rodents impact the visual system. Additionally, we try to answer whether PTU treatment would affect neuronal plasticity in the visual cortex. We used the Visual Evoked Potential recordings to assess contrast sensitivity, spatial frequency acuity, and ocular dominance plasticity. In addition, we look into the expression of the photoreceptors in the retina. PTU exposure impacts the contrast sensitivity but not the spatial frequency acuity or ocular dominance plasticity. The expression level of the photoreceptor Opsin-M was also impacted. The reduced levels of the thyroid hormones during this crucial time have long-lasting consequences for the proper visual system processing.
  • The Role of Colonization Factors CFA/I and CS21 in Enterotoxigenic E.coli (ETEC) Pathogenesis in the Human Enteroid Model

    Smith, Emily; Barry, Eileen M. (2022)
    Enterotoxigenic Escherichia coli (ETEC) is a primary causative agent of diarrhea in travelers and in young children in low-to-middle income countries (LMICs). ETEC adhere to intestinal epithelia via colonization factors (CFs) and secrete heat-stable toxin (ST) and/or heat-labile toxin (LT), causing dysregulated cellular ion transport and water secretion. ETEC isolates often harbor genes encoding more than one CF and are prime targets as vaccine antigens. Many clinical isolates express CFA/I and CS21; however, a role for CS21 alone or with CFA/I has not been defined. We hypothesize that expression of both CFs confers increased adherence and toxin delivery to the human enteroid. Clinical strains expressing CFA/I and/or CS21 were evaluated, and CF-deficient mutants were engineered. After confirming CF expression using western blot and electron microscopy, assays demonstrated CFA/I was important for CFA/I-CS21 ETEC adherence, as CFA/I-deficient mutants and strains pre-incubated with anti-CFA/I antibody had significantly reduced adherence to enteroid monolayers compared to wildtype. In contrast, CS21 was not required as CS21-deficient mutants and strains pre-incubated with anti-CS21 antibody adhered at similar levels as wildtype. These data demonstrate that targeting CFA/I in CFA/I-CS21 ETEC is sufficient for significant adherence reduction. Delivery of ST by CFA/I-CS21 ETEC was evaluated. Strain-specific levels of toxin delivery were detected but CF-dependent ST delivery was not observed, which may reflect the lack of flow and stretch in the current enteroid model. Upon investigation of host responses to ETEC, the enteroid monolayer integrity was not disrupted, as shown by the increase in transepithelial electrical resistance and the lack of inflammatory cytokines produced. Infection with ETEC strains resulted in decreased mucus (MUC2) production, but this was not CF-dependent. Further studies of strain-specific CFA/I expression revealed that it may be transcriptionally or post-transcriptionally regulated, following observation of nearly identical CFA/I operon sequences and many shared CF-specific regulators at the genomic level. Overall, these data support the role of CFA/I in CFA/I-CS21 ETEC adherence and reinforces CFA/I as a main target for vaccines. These data also highlight the human enteroid model to study ETEC pathogenesis and for evaluation of preclinical therapeutics.
  • Novel Modules in the T Cell Signaling Circuit Which Enable Synergy Between Responses to Self and Foreign Peptides

    Wolf, Gideon; Singh, Nevil (2022)
    T cell activation occurs when a T cell receptor (TCR) engages with cognate agonistic peptides in the context of Major Histocompatibility Complexes (pMHCs) on antigen presenting cells (APCs). This initiates a series of intracellular signaling events proximal to the TCR and associated CD3 complexes, mediated by unique kinases together with scaffolding and lattice molecules. The downstream cascades ultimately lead to transcriptional changes that promote the cellular program of conventional T cell activation—for example, cytoskeletal rearrangement, cytokine production, cellular proliferation, and differentiation. Understanding signaling mechanisms not only allows us to decipher the regulation of T cell activation but also to manipulate immune responses pharmacologically. TCR signaling by agonistic pMHCs is well studied, but much less is known about signaling by a parallel universe of self-peptides that also engage TCRs in vivo. The focus of this thesis is to understand how T cells perceive these signals without fully acquiring effector responses as a result. The central hypothesis of my thesis is that self-peptide ligands signal uniquely through the TCR to alter the fate and function of a T cell both with and without presence of their cognate agonist. We evaluated this hypothesis using approaches that globally increased self-peptide presentation in vivo (using FLT3L to generate more DCs), deprived T cells of self-peptide (by culturing away from APCs) or stimulated a TCR with a known self-peptide in the presence or absence of the strong agonist. The significant findings from these studies are that (i) boosting self-peptide presentation transiently increases a narrow T cell effector subset; (ii) depriving T cells of self-engagement lowers basal phosphorylation in the key signaling adapter LAT; (iii) self-peptides do not trigger cellular activation on their own, but synergize to enhance activation as measured by CD69, pERK, and several other parameters (iv) self-peptides initiate TCR signaling up to the level of pMEK and (v) self-peptides elicit a unique transcriptional profile. Together, these results not only define the unique contributions of self-peptides to T cell activation but also demonstrate a distinct wiring profile in the TCR-signaling network that limits self-peptide sensing at the ERK step.
  • The Role of Granzyme B in Tumor Migration and Metastasis

    Ellis, Tibbs; Cao, Xuefang (2022)
    The granule exocytosis pathway has classically been described as our most potent response to pathogens, cancers, as well as autoreactive disorders. This is executed by the action of perforin and a family of granule-associated serine proteases known as granzymes (Gzm). Many studies have shown that these proteases exert their effects by killing affected cells, creating an obvious correlation with cytotoxic T cells (CTLs) and natural killer cells (NK) greatly expressing GzmB. Studies have found that the tumor cells ability to create a suppressive environment is dependent on Gzm expression by regulatory T lymphocytes (Treg). Research has shown GzmB can target and cleave extracellular molecules such as fibronectin, vitronectin and laminin. Bird et al published data showing that GzmB aids in the transmigration of CTLs, however the ability of GzmB to aid in the migration of non-T cells has not been investigated. We hypothesize that GzmB secretion may cause extracellular matrix (ECM) remodeling in the tumor microenvironment, aiding in the outgrowth of the tumor cells via promoting higher rates of invasion and/or metastasis. First, we designed experiments to test in-vivo and in-vitro conditions that generate GzmB production in multiple cell types. We were able to induce GzmB production by Tregs both in-vitro and in-vivo. We also identified multiple mechanisms that allow for CD8 T cells to produce GzmB. We next investigated mechanisms that would allow for GzmB to be found in the extracellular environment. We found that CD8 T cells have the ability to secrete GzmB without the need of an immunological synapse. In fact, GzmB secretion is not dependent on exogenous IL-2. GzmB improves the migration and invasive potential of tumor cells. In Treg-specific GzmBko (Foxp3creGzmBfl/fl) mice, GzmB+ Tregs increase the rate of tumor metastasis in the lungs of the mice. In addition, GzmB+ CD8 T cells can potentiate tumor metastasis, in contrast to GzmB-deficient CD8 T cells. These findings shed light on the protumorigenic potential of secretory GzmB, which could serve as a unique biomarker for predicting metastasis.
  • Development of the Lennard-Jones Parameters for the Polarizable Classical Drude Oscillator Force Field

    Chatterjee, Payal; MacKerell, Alexander D., Jr. (2022)
    The quality of Force Fields (FF) determines accuracy of observations made through molecular simulations. Accuracy of such simulations may be achieved by explicit inclusion of electronic polarization, such as via the implementation of the Drude harmonic oscillator, as in Drude Polarizable FF. Although the Drude Polarizable FF spans a large range of biomolecules including proteins, nucleic acids, lipids and carbohydrates, an expansion of its small molecule FF is essential, given the vastness of chemical space. Such an expansion must be accompanied by the optimization of van der Waals (vdW) interactions, in the context of the Lennard-Jones (LJ) formalism. Optimization of the LJ parameters is a multivariate and multi-objective problem and is one of the most challenging aspects of FF optimization. Through this thesis, we have developed a method that utilizes the sampling power of Latin Hypercube Design (LHD) and learning abilities of Deep Neural Network (DNN) to overcome some of these challenges. The model is trained on empirical pure solvent/crystal properties of a selected set of “training set” compounds, where the final selection is based on the quality of both gas phase and condensed phase properties. The optimized LJ parameters are validated for transferability on “validation set” compounds, while their ability to reproduce other experimental thermodynamic properties such as hydration free energy and dielectric constants, are also verified. Chapter 1 of this thesis presents an introduction to underlying concepts of FFs, with a major focus on polarizable FFs. Chapter 2 details development of the method, using four different chemical classes, i.e., alkenes, 3 and 4 membered rings and nitriles. Chapter 3 updates the method developed in chapter 2, addressing the challenge of parameter correlation. Chapter 4 applies the updated method to another chemical class (alkynes), while Chapter 5 concludes the thesis and is a discussion of the challenges associated with empirical FF development with a focus on LJ parameters. Overall, the method developed through this thesis addresses the most challenging aspect of FF development, i.e., LJ parameter development, implemented in a manner that could be utilized in context of development of both additive and polarizable FF.
  • Using PacBio SMRT platform to assess the genetic variation of Plasmodium falciparum Circumsporozoite Protein (PfCSP) in Kalifabougou, Mali

    Ouedraogo, Nadiatou; Laurens, Matthew B. (2022)
    Objectives: In this study, we assessed the genetic variation in CSP sequences from Kalifabougou, Mali, by evaluating the diversity of haplotypes in the N- and C- Terminal region, the repeat regions, and the T- and B-cell epitopes. Methods: Sequences used for the study were generated from 59 sequences collected during a malaria transmission season conducted in Kalifabougou, Mali. Generated using the PacBio SMRT platform. Results: Results showed that the central repeat and C-terminal Th2R/Th3R epitope regions were highly polymorphic in this study, whereas the N-terminal non-repeat region was less variable. Furthermore findings highlighted CSP polymorphism and suggested that the repeat and B-cell epitope of our samples is similar to those of 7G8 PfCSP, whereas the C-terminal is similar to that of the 3D7 PfCSP strain. Finally, findings the CSP sequences from the Kalifabougou site are more similar to those from Navrongo than to those from Cape Coast.
  • Membrane-Anchored Serine Proteases in Ovarian Cancer Dissemination and Metastasis

    Pawar, Nisha; Antalis, Toni M. (2022)
    Ovarian cancer (OvCa) is the most fatal gynecological malignancy due to delayed clinical presentation, passive metastasis, frequent tumor recurrence, and lack of effective targeted therapies. OvCa tumors predominantly shed as single cells or spheroids and disseminate throughout the peritoneal cavity. Spheroids are considered critical metastatic units that contribute to disease recurrence and chemoresistance. Despite recent advances in treatment strategies, there has been little improvement in patient survival. There is an urgent need to develop a better understanding of molecular mechanisms that are fundamental for OvCa dissemination, in order to provide alternative therapeutic options for long-term remission. Matriptase, a membrane-anchored serine protease (MASP), and its substrate protease-activated receptor-2 (PAR-2) both exhibit elevated expression in OvCa compared to normal ovary tissues which worsens patient survival, suggesting a critical role for activation of this pathway in OvCa progression. Overactive matriptase, induced by an imbalanced ratio with its cognate inhibitor hepatocyte growth factor activator inhibitor-1 (HAI-1) has been implicated in disruption of barrier integrity and epithelial carcinogenesis, although the mechanism is not known. This study, demonstrates a functional role for an imbalanced matriptase:HAI-1 ratio in OvCa progression, and identifies a matriptase/PAR-2/PI3K/Akt/MMP-9/E-cadherin signaling axis that disrupts cell-cell interactions and promotes formation of pro-metastatic loose spheroids in vitro, in in vivo orthotopic xenograft models, and in patient-derived tumor cells. This matriptase/PAR-2 pro-metastatic signaling is in direct contrast to another MASP family member, testisin, suggesting a novel mechanism of biased agonism that orchestrates OvCa dissemination. Since over-activity of MASPs is associated with advanced OvCa, we have developed a MASP-activated pro-drug based on a re-engineered anthrax toxin (PAS:LF) that is selectively activated by zymogen-activating proteases on the tumor cell surface to inhibit tumor cell-survival pathways. This therapeutic strategy is radically different from anti-proliferative mechanisms of standard chemotherapeutic agents. We have demonstrated anti-tumor efficacy in a range of OvCa cells and spheroids in vitro, in various in vivo orthotopic xenograft models of OvCa dissemination, in patient ascites-derived tumor cells and in patient-derived xenograft models in vivo, with no off-target adverse effects. Clinical translation of these preclinical findings could establish PAS:LF as a promising treatment strategy to improve patient outcomes.
  • Caries Experience Based on A Simulated Epidemiological Screening and Routine Examination with Radiographs

    Kuchari, Abrar; Dhar, Vineet (2022)
    Purpose: To assess and compare caries experience based on an epidemiological “screening” examination and a routine examination with radiographs. Methods: 128 patients ages 5 to 10 years were included. An initial examination was done by a pediatric dentist in conditions simulating an epidemiological screening assessment. A second evaluation was done by eight postgraduate pediatric dental resident who conducted a routine clinical exam of the patient in the dental chair including radiographs. Results: By the epidemiological screening, the prevalence of untreated dental caries (Y/N) in primary and/or permanent teeth was 50% and the caries experience was 77%. After the routine examination with radiographs, the prevalence of untreated dental caries was 74.22% and caries experience was 90%. The severity of untreated dental caries was 2.0 and 3.9, and the severity of caries experience was 3.2 and 5.6 by the simulated epidemiological screening and routine examination with radiographs, respectively.
  • The Effects of HIV Exposure and Maternal Antibodies to Hepatitis B Virus on the Immune Response of Nigerian Infants to Hepatitis B Vaccine.

    Bada, Florence; Stafford, Kristen Alyce; Campbell, James Daniel (2022)
    Introduction: Chronic hepatitis B virus (HBV) infection leads to considerable morbidity and early mortality. Development of chronic HBV infection can be prevented by a three to four dose schedule of Hepatitis B vaccines in immunocompetent infants. However, HIV exposed uninfected infants (HEU) are thought to exhibit an attenuated immune response to some vaccines. In addition, a significant proportion of pregnant women in Nigeria have antibodies to HBV, specifically Hepatitis B surface antibodies (HBsAb). Maternal antibodies inhibit infant immune responses in some instances. Objective: To estimate the effect of antenatal and perinatal exposure to maternal HIV, and the effects of maternal HBsAb on the immune response of Nigerian infants to hepatitis B vaccine. Methods: Using a retrospective cohort design, we determined the relationship between infant HIV-exposure status, and infant exposure to detectable concentrations of maternal HBsAb, and infant immune response to Hepatitis B vaccine separately using general linear models adjusted for potential confounders. Subsequently, we used Fisher’s Exact tests to compare the proportion of infants with HBsAb concentrations above 10mIU/mL: for HEU as compared to HIV unexposed uninfected infants (HUU), and for infants exposed to detectable concentrations of maternal HBsAb as compared to infants unexposed to detectable concentrations of maternal HBsAb at 24 and 52 weeks of age separately. Results: We found HIV exposure to be associated with infant immune response to hepatitis B vaccine at birth, Weeks 4 and 52 (P<0.0001, P=0.05 and P<0.0001) and also to be associated with the proportion of infants with HBsAb concentrations ≥ 10mIU/mL at Week 52 (P=0.04). Exposure to detectable concentrations of maternal HBsAb was also associated with infant immune response at Weeks 24 and 52 respectively. Conclusion: Though antenatal and perinatal exposure to HIV, and to detectable concentrations of maternal HBsAb were found to be associated with infant immune response to hepatitis B vaccine, these exposures did not appear to attenuate immune response. In addition, differences observed in the proportion of infants with HBsAb concentrations ≥ 10mIU/mL were small and do not appear to be clinically relevant.
  • Surviving High School Transfers: A Multilevel Study of Student and School Characteristics Related to School Transfer, Graduation, and College Entry

    de Tablan, Dante; DeForge, Bruce R. (2022)
    Background: Transferring schools frequently happens across the United States from kindergarten to college. While student mobility studies have focused more on elementary and middle school grades, research at the high school level is limited. In addition, studies on transfer during high school related to postsecondary education are even more scarce. This dissertation investigates transfer risk associated with student and school characteristics and its association with high school graduation and college enrollment. Method: The dissertation used analytic samples from a cohort of 6,810 first-time ninth graders enrolled in Baltimore City Public Schools from 2012-2013 to 2017-2018. To examine the factors related to college entry, a second analytic sample included only students who obtained a high school diploma, a certificate of completion, or a GED (N = 4,297). The study employed mixed-effects parametric proportional hazards modeling to investigate student- and school-level characteristics associated with time to the transfer event, and multilevel binary logistic regressions to analyze student and school factors related to odds of high school graduation and college enrollment. Results: Thirty-four percent of students transferred schools during the study period. White students, those with missing 8th-grade math test scores, students who received a suspension, and employed students were at a reduced risk of transfer. Chronic absentees and those with standardized 8th-grade math scores had increased transfer risk. While the school percentage of students eligible for free and reduced meals related to a reduced transfer risk, the school special education rate was associated with increased transfer risk. Transferring schools was associated with lower odds of graduation and college entry, and percentage of school transfer was associated with lower odds of college enrollment. Transfer, as it related to graduation and college entry, varied across schools. Conclusion: School transfer is a multidimensional event related to adverse educational outcomes for many students. This dissertation identified student and school characteristics associated with time to transfer risk. Moreover, the study highlighted the adverse effects of transfer on graduation and college entry. Finally, a discussion of the limitations, strengths, and implications for research, policy, and practice are presented.
  • Hippocampal Contributions to Stress: What the Hippocampus Tells the HPA Axis and What Could Go Wrong?

    Cole, Anthony; Thompson, Scott M. (2022)
    The links between stress and psychiatric illnesses are numerous and bidirectional. Stress is highly correlated to psychiatric illness, increasing the risk of illness and often precipitating its onset. The primary class of neuroendocrine stress hormones, glucocorticoids, is often dysregulated in patients with major depression. Understanding the neurobiological changes underlying these findings is crucial for the development of improved therapeutic strategies and better patient outcomes. Prior work suggests that the hippocampus is stress-sensitive, substantially altered in major depression, and plays an important regulatory role in HPA axis function. Our understanding of the processes regulating the stress response is incomplete, particularly in regards to the various brain regions responsible for its central regulation. In this document I strive to clarify our understanding of a specific component of stress regulation, namely the hippocampal contribution, which could prove important to understanding the various links between stress and psychiatric illness. To do this, I targeted the ventral hippocampus with injections containing virus that expressed the light-sensitive ion channel channelrhodopsin (ChR-EYFP) in glutamatergic neurons. I determined the extent of glutamatergic hippocampal terminal expression in brain regions thought to regulate the paraventricular nucleus of the hypothalamus (PVN), and then stimulated those terminals while recording from corticotropin-releasing factor-positive (CRF+) neurons in the PVN in a whole-cell voltage clamp recording configuration. Using parasaggital brain sections, I was able to induce optically-evoked inhibitory post-synaptic currents in CRF+ neurons of the PVN, demonstrating the first direct functional evidence of hippocampal inhibition of the PVN. I used a similar injection scheme to record GAD+ neurons in the BNST. Finally, I utilized in vivo optical stimulation of hippocampal terminals within the BNST during acute restraint stress and observed a significant decrease in circulating levels of corticosterone. My data indicates that the hippocampus sends dense projections to the BNST and optical excitation of these hippocampal terminals produce inhibitory currents in CRF+ neurons of the PVN. Our data suggests a disynaptic hippocampus-BNST-PVN circuit of regulation by which the hippocampus inhibits the CRF+ PVN cells. The in vivo results indicate that the hippocampal inhibition of these neurons is capable of functionally inhibiting the HPA axis.
  • Perceived Need for Care and Mental Health Service Use: The Moderating Effect of Race/Ethnicity and Military Veteran Status among a Population-based Sample of U.S Adults

    McNish, Nicole Latoya; Bright, Charlotte Lyn (2022)
    Background: Black, Indigenous, and People of Color (BIPOC) adults and military veterans are susceptible to chronic and severe mental illness given their higher likelihood of exposure to risk factors compared to the general population. Despite the consequences of untreated mental illness and federal initiatives aimed at expanding healthcare access, treatment utilization among both groups remains low relative to treatment need. Research has found significant racial and ethnic differences in perceived need for care (PNC) and severity of psychological distress, important antecedents of treatment seeking. However, the extent to which these factors apply to veterans has not been widely examined. Using the Behavioral Model of Health Services Use (Andersen, 1995) and Behavioral Model for Vulnerable Populations (BMVP, Gelberg et al., 2000), this dissertation aimed to address these gaps in the existing literature given their significance to ongoing research, policy, and intervention efforts. Methods: Data were drawn from the 2018 National Survey on Drug Use and Health (N = 43,026). Binary logistic regression was used to examine the associations of PNC and symptom severity with mental health service use among veterans and nonveterans of diverse racial and ethnic backgrounds with interaction terms for race/ethnicity and veteran status. Results: PNC and symptom severity are associated with higher probabilities of mental health service use. However, there are differences in the adjusted probabilities in mental health service use favoring White nonveterans. Veteran status moderates these associations such that the probabilities of mental health service use were stronger for veterans than nonveterans who reported PNC, except for Asian veterans. Conclusions: Veteran status moderates some longstanding racial and ethnic disparities in mental health service use. Further research is needed, particularly among Asian veterans, to identify factors that contribute to lower probability of mental health service use in comparison to White adults, and to identify ways to leverage the effectiveness of interventions employed among veterans in support of other vulnerable groups.
  • Pharmacometabolomics of clopidogrel: Determining the genetic and metabolic contributors to clopidogrel response

    Bromberek, Sarah Katherine; Beitelshees, Amber L. (2021)
    Clopidogrel is a commonly prescribed antiplatelet drug and there is considerable variability in response. The PAPI study was conducted in 687 individuals to determine the genetic predictors of clopidogrel response. Targeted metabolomic profiling of 42 amines was performed in a subset of 198 PAPI subjects with genetics data available. We identified the metabolic signature of clopidogrel and determined metabolites associated with clopidogrel-induced changes in platelet reactivity. We found tyrosine was most significantly changed after exposure to clopidogrel, and BCAAs baseline levels were associated with clopidogrel-induced changes in platelet aggregation. Gaining insights into factors that influence variability in antiplatelet response is important for identifying novel antiplatelet mechanisms or biomarkers for predicting response. These findings can have long-term implications toward advancing precision medicine in antiplatelet therapy.

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