Master of Science, The Ohio State University, 1987, Graduate School

Committee: Not Provided (Other) Subjects:

Doctor of Philosophy, The Ohio State University, 2024, Biomedical Sciences

Nearly 50% of the US population is predicted to be obese by 2030. Chronic inflammation is a central feature of obesity, which leads to cardiovascular disease, insulin resistance, cancer, and several other life-threatening complications. A healthy, lean individual is protected from adipose tissue inflammation and associated comorbidities due to well-coordinated interactions between adipocytes and immune cells in adipose tissue that maintain normal systemic metabolism. In obesity, pro-inflammatory changes occur in nearly all adipose immune cells. These proinflammatory changes include increased proinflammatory (T helper1) Th1 cells that produce interferon-gamma, which reduces immunoregulatory T cells (Tregs). T helper type 2 cells (Th2), innate lymphoid type 2 cells (ILC2), and anti-inflammatory M2-type macrophages also decrease, whereas proinflammatory neutrophils, M1-type macrophages, and cytotoxic CD8 T cells increase. The timeline of these events is established in several mouse models of obesity. However, these events in human adipose tissue remain elusive. Over the years, Hsueh laboratory has established a more comprehensive profile of the human AT microenvironment and has identified various mechanisms by which immune cells within the adipose tissue contribute to an obesity-related pro-inflammatory state that raises several metabolic diseases. One of the emerging mechanisms for AT inflammation is the transfer of gut microbes to adipose tissue due to increased gut dysbiosis in obesity. Microbes in AT can cause inflammation. We found that a mere two weeks of high-caloric diet consumption in healthy human subjects has shown signs of leaky gut, which overlapped with the recruitment of neutrophils to the adipose tissue. Interestingly, within the same cohort, we observed a remarkable decline in adipose tissue Tregs, a central player in regulating AT immune homeostasis. Therefore, we explored if a leaky gut resulted in neutrophil recruitment and subsequent proinflammato (open full item for complete abstract)

Committee: Willa Hsueh (Advisor); Daniel Spakowicz (Committee Member); Amy Lovett-Racke (Committee Member); Hazem Ghoneim (Committee Member) Subjects: Biomedical Research

Doctor of Philosophy, The Ohio State University, 2022, Biomedical Sciences

Infections and disease caused by the obligate human pathogen Bordetella pertussis (Bp) are resurging worldwide, despite widespread vaccinations. The current acellular pertussis vaccines remain ineffective against nasopharyngeal colonization, carriage, and transmission. To develop more effective vaccines, it is important to understand how Bp resists killing actions of host immunity. In the current work, we tested the hypothesis that Bordetella polysaccharide (Bps), a member of the poly-􀈕-1,6-N-acetyl-D-glucosamine (PNAG) family, promotes respiratory tract colonization of Bp by resisting killing from two major innate immune factors: (1) antimicrobial peptides (AMPs) and (2) neutrophils. First, we show that Bps enhanced Bp survival in the presence of AMPs by functioning both as a surface shield and a decoy. While surface-bound Bps inhibited AMP binding to the bacterial surface, soluble Bps neutralized AMP activity by direct binding interactions. Simultaneous infection of mice with Bps-proficient and Bps-deficient strains resulted in greater survival of the Bps-deficient strain in the mouse respiratory tract. Furthermore, production of Bps in a nonpathogenic E. coli strain increased AMP resistance in vitro, and increased bacterial survival and augmented pathology in the mouse respiratory tract. Second, we show that Bps enhanced survival in the presence of hydrogen peroxide and hypochlorous acid in vitro. Consistent with these data, Bps enhanced resistance to killing by human neutrophils and suppressed oxidative burst and neutrophil extracellular trap (NET) formation. Together, the results obtained from studies involving AMPs and neutrophils serve as a foundation for studying other PNAG polysaccharides, as well as for the development of an effective Bp vaccine that includes Bps. Traditionally, pertussis is described as an acute disease with coughing fits and other severe symptoms. However, in recent years, many individuals who contract pertuss (open full item for complete abstract)

Committee: Rajendar Deora Ph.D. (Advisor); Paul Stoodley Ph.D. (Committee Chair); Daniel Wozniak Ph.D. (Committee Member); Santiago Partida-Sanchez Ph.D. (Committee Member); John Gunn Ph.D. (Committee Member) Subjects: Biomedical Research; Immunology; Microbiology

PhD, University of Cincinnati, 2022, Medicine: Molecular and Developmental Biology

Though the basic building blocks of the cytoskeleton were first identified more than fifty years ago, there remains much to learn about how cytoskeletal dynamics impact cellular function, and ultimately contribute to cellular homeostasis and human disease. In this project, we have sought to determine how a microtubule-associated protein, the dynein arm assembly factor Coiled-coiled domain-containing protein 103 (CCDC103) contributes to the normal maturation and effector function of human and zebrafish neutrophils by secondarily impacting α-tubulin lysine 40 (K40) acetylation, a critical post-translational modification affecting microtubule stability. CCDC103 plays a critical role in the proper assembly of the axonemal dynein complex, and, which mutated, causes a disorder known in humans as Primary Ciliary Dyskinesia or PCD. Interestingly, studies have shown that myeloid cells from PCD patients have multiple functional and cytostructural defects, even though myeloid cells themselves do not possess motile cilia. We have previously shown that ccdc103 mutations are sufficient to disrupt neutrophil and macrophage migration and proliferation in zebrafish, mostly likely through its well-characterized role as a microtubule stabilizing factor. Additionally, we identified multiple novel CCDC103 binding partners that may support its function, including the flagellar central-apparatus protein SPAG6 and the NAD-dependent deacetylase SIRT2, and showed that PCD-associated CCDC103 mutations abrogate interactions with both of these proteins. Zebrafish spag6 mutant embryos display multiple phenotypic hallmarks of motile cilia dysfunction, including low fertility and situs inversus, and recapitulate many of the same proliferation, morphology, and directed migration capacity defects also observed in ccdc103 mutants, suggesting that SPAG6 acts together with CCDC103 to stabilize cytoplasmic microtubules. Conversely, SIRT2 is a key deacetylase of α-tubulin, a pos (open full item for complete abstract)

Committee: Joshua Waxman Ph.D. (Committee Member); Lindsey Barske Ph.D. (Committee Member); Ashish Kumar M.D. (Committee Member); Marie-Dominique Filippi Ph.D. (Committee Member); Tony De Falco Ph.D. (Committee Member) Subjects: Cellular Biology

MS, University of Cincinnati, 2021, Medicine: Immunology

NETosis is a regulated form of cell death that neutrophils initiate in order to trap and kill bacteria. During this process neutrophil extracellular traps (NETs) are released. The traps consist of extracellularized DNA coated with antimicrobial proteins. Additionally, the release and rupture of granulocytes helps kill foreign bodies. Little is known about NET formation in the context of fetal infection and inflammation. During in utero infection, fetal lungs and other mucosal tissues come in contact with bacteria and bacterial products present in the amniotic fluid. To study the functionality of fetal neutrophils, two Rhesus macaque models of chorioamnionitis were used. Pregnant animals were injected intra-amniotically with lipopolysaccharide (LPS) or Escherichia coli (E. coli), and fetal lung tissues were harvested after 16 or 48 hours respectively. NETs were identified by immunohistochemistry detection of extracellular citrullinated histone in neutrophils, identified as CD68+HLA-DRneg cells. Compared to controls, the IA LPS fetal lungs did have significantly greater mean NET area. To compare the functionality of fetal neutrophils to adult neutrophils, fetal lung samples were matched with their corresponding chorioamnion. In LPS animals, there was a trend towards higher NET formation in the chorioamnion, suggesting that fetal neutrophils may be capable of NET formation but not as much as adult neutrophils. We also compared the LPS treatment to the E. coli treatment since NETs are classically thought to trap and kill live bacteria. We found no difference between the two treatments demonstrating that fetal neutrophils respond to bacterial products.

Committee: Claire Chougnet Ph.D. (Committee Chair); Ian Lewkowich (Committee Member); Jonathan Katz Ph.D. (Committee Member) Subjects: Immunology

Doctor of Philosophy, The Ohio State University, 2021, Biomedical Sciences

Tuberculosis (TB), caused by the bacterium Mycobacterium tuberculosis (M.tb), is a leading cause of death due to an infectious disease, and it is estimated that 1.66 million people died from TB in 2020. An infected person may progress immediately to active TB disease, characterized by significant clinical symptoms and M.tb transmission, but the majority of those infected with M.tb may contain the disease in a latent infection, with no clinical symptoms or disease transmission. However, latently infected persons may reactivate their infection and progress to active disease later in life. The distinct reasons for the differential TB outcomes in the human population are not clear, but may be linked to the immune process of inflammation. Comorbidities which increase susceptibility to develop active TB and/or succumb to the disease, such as HIV co-infection, COPD, and natural aging, are associated with an increased systemic or pulmonary inflammatory state. Furthermore, as late stages of TB disease are characterized by significant amounts of pulmonary inflammation and tissue destruction, is suggests increased inflammation can drive TB susceptibility. However, animal models and in vitro studies suggest basic and enhanced inflammatory mechanisms can improve M.tb control. Additionally, a subpopulation of humans may be capable of early clearance of M.tb, using enhanced innate immunity to clear the infection. Together, these suggest the relationship between TB and inflammation is complex, and more study is needed to understand the true positive and negative roles of inflammation in TB. In this work we study how acute inflammation may directly impact M.tb control in a mouse model, and assess the impacts of inflammation on myeloid and lymphocyte populations in murine lungs. We found that increasing inflammation using lipopolysaccharide (LPS) treatment prior and during early M.tb infection induces enhanced control of M.tb evident as early as seven days of infection, a protecti (open full item for complete abstract)

Committee: Joanne Turner PhD (Advisor); Ian Davis DVM, PhD (Committee Member); Mireia Guerau-de-Arellano PharmD, PhD (Committee Member); William Lafuse PhD (Committee Member); Traci Wilgus PhD (Committee Chair) Subjects: Biology; Biomedical Research; Immunology; Microbiology

Doctor of Philosophy, The Ohio State University, 2020, Neuroscience Graduate Studies Program

Dermatological complications frequently affect people who suffer a spinal cord injury (SCI). These complications greatly impact patient outcomes; thus, studies investigating how they develop are highly relevant to the SCI community. However, the mechanisms by which SCI affects the skin are poorly understood. Evaluation of these mechanisms is further complicated by heterogeneity in the etiologies and manifestations of SCI. Experimental SCI models can more definitively evaluate the contributions of specific biological mechanisms to SCI-induced skin changes; however, such experimental models are rare. In this thesis, experiments using a controlled model of high-level (T3) SCI were designed to analyze SCI-induced skin changes in the context of two inflammatory skin models and in non-disrupted control skin. First, SCI-induced changes in cutaneous immunity were evaluated in response to a subcutaneous immune stimulus (Complete Freund's Adjuvant) delivered below the level of injury. In vivo imaging of V-sense, a macrophage-specific tracer, was used to monitor skin inflammation in real time. These experiments revealed that within the first 5 days after adjuvant injection, SCI reduced the macrophage response at the injection site by ~50% relative to sham-injured animals. Such experiments definitively showed that SCI alters skin immunity, with relevance to skin infections. Because macrophages are essential for wound repair, the next model investigated how SCI affected the macrophage response to skin wounds. In this model, SCI or sham-injured (control) mice received bilateral 3.5-mm punch biopsies on the back below the injury level. Daily photographs were taken of wounds to create a temporal healing profile, which revealed that SCI significantly impaired wound closure relative to sham injury beginning at 6 days post-wounding (dpw). This corresponded with significantly increased macrophage accumulation within SCI wounds. Protracted macrophage accumulation is also a hallmar (open full item for complete abstract)

Committee: Phillip Popovich PhD (Advisor); Dana McTigue PhD (Committee Member); Karl Obrietan PhD (Committee Member); Leah Pyter PhD (Committee Member); Chandan Sen PhD (Committee Member) Subjects: Biology; Biomedical Research; Neurobiology

Doctor of Philosophy (PhD), University of Toledo, 2019, Biomedical Sciences (Medical Microbiology and Immunology)

Borrelia burgdorferi (Bb) is an extracellular spirochetal bacterium and the causative agent of Lyme disease (LD). In vitro, Bb induces robust immune responses which suggests that Bb could be effectively cleared. However, neither innate nor adaptive immune responses are able to eliminate Bb in vivo resulting in persistent infection. Hence, there is a dysregulation of the immune response occurring in vivo, which cannot be recapitulated in an in vitro system due to the intricacy and complexity of the host tissue milieu. In this study, we utilized intravital microscopy (IVM) along with several transgenic mouse lines, to delineate and identify the immune dysregulation occurring within skin tissues in vivo, which permits this extracellular pathogen to maintain a persistent infection. Our IVM studies identified that Bb undergo a massive proliferation between days 2 to 7 post-infection and peak bacterial numbers are achieved in skin tissues adjacent to the infection site around day 8 post-infection. During this period of acute infection, an activated innate response should be identifying and clearing this extracelluar pathogen, since Bb possess potent agonists. However, host innate responses fail to control the spirochete and Bb numbers undergo almost a 50-fold increase during this period. iv Attainment of peak bacterial numbers is followed by a significant decrease in Bb numbers until day 14 post-infection; subsequently, this low level is maintained for over 2 years. Based on this in vivo persistence data, the two major goals of our study were to 1) delineate whether adaptive immune responses were responsible for the significant decrease in Bb numbers after day 8 post-infection, and 2) determine whether Bb-elicited IL-10 is involved in the dysregulation of innate responses necessary to control Bb numbers during early infection. To test the contribution of the host adaptive responses in reducing Bb numbers, we used several transgenic mice with B cell- and T cell-defi (open full item for complete abstract)

Committee: R. Mark Wooten (Committee Chair); Andrea Kalinoski (Committee Member); Kevin Pan (Committee Member); Randall Worth (Committee Member); Robert Blumenthal (Committee Member) Subjects: Biomedical Research; Immunology; Microbiology

Doctor of Philosophy, The Ohio State University, 1985, Graduate School

Committee: Not Provided (Other) Subjects: Chemistry

Doctor of Philosophy, The Ohio State University, 1982, Graduate School

Committee: Not Provided (Other) Subjects: Biology

Doctor of Philosophy, The Ohio State University, 1978, Graduate School

Committee: Not Provided (Other) Subjects: Health Sciences

Doctor of Philosophy (PhD), Wright State University, 2014, Biomedical Sciences PhD

The airway epithelium poses a formidable barrier for the entry of pathogenic viruses due to the formation of tight junctions between adjacent epithelial cells. The coxsackievirus and adenovirus receptor (CAR), a member of the Ig superfamily of cell junction adhesion proteins, is the primary receptor for adenovirus entry and infection. As a result of alternative splicing, two transmembrane isoforms of CAR are generated. While the seven-exon isoform of CAR (CAREX7) is hidden on the basolateral surface of polarized epithelia, the eight-exon isoform of CAR (CAREX8) localizes within the sub-apical region and at the air-exposed apical surface. Apical localization of CAREX8 makes it accessible to invading adenovirus entering the lumen of the airway and able to facilitate viral entry into the epithelium. Previous studies have shown that Interleukin-8 (IL-8), a proinflammatory cytokine and a neutrophil chemoattractant, increases the susceptibility of the airway epithelium to adenoviral infection. I hypothesized that the apical CAREX8 protein expression level and localization are responsible for the susceptibility of a polarized epithelium to viral infection. Moreover, I hypothesized that CAREX8expression is tightly regulated by mediators of IL-8 signaling and the endogenous function CAREX8is to tether neutrophils at the apical surface of the polarized epithelium. Finally, I hypothesized that adenovirus has co-opted CAREX8 and neutrophil transmigration to enhance infection of the polarized epithelium. Consistent with these hypotheses, I demonstrate that IL-8 increases the expression and the apical localization of CAREX8 in polarized airway epithelial cells. In addition, IL-8 differentially activates AKT/S6K and inactivates GSK3ß to augment the protein synthesis of CAREX8. Increased CAREX8 is able to mediate increased neutrophil binding at the apical surface of the epithelium that is completely abolished by competition with CAR-binding adenovirus fiber-knob. Finally, I a (open full item for complete abstract)

Committee: Katherine Excoffon Ph.D. (Advisor); David Goldstein Ph.D. (Committee Member); Robert Putnam Ph.D. (Committee Member); Dawn Wooley Ph.D. (Committee Member); Julian Gomez-Cambronero Ph.D. (Committee Member) Subjects: Cellular Biology; Virology

Doctor of Philosophy, The Ohio State University, 2014, Integrated Biomedical Science Graduate Program

Chronic respiratory infections with the bacterium Pseudomonas aeruginosa are the leading cause of mortality in cystic fibrosis (CF) patients. During the course of infection, P. aeruginosa undergoes a phenotypic change from a non-mucoid to a mucoid appearance, which directly correlates with a worsening clinical prognosis. Mucoid conversion results from the acquisition of mutations leading to overproduction of alginate, a capsular polysaccharide, which confers a selective advantage for P. aeruginosa. The most frequent mutation observed in mucoid P. aeruginosa isolated from CF patients occurs in the gene encoding the anti-sigma factor mucA. MucA negatively regulates the sigma factor AlgT and upon mucA disruption AlgT constitutively upregulates alginate production. The overall goal of this study was to identify both the bacterial mechanisms and the CF host factors responsible for promoting mutations leading to P. aeruginosa mucoid conversion. In Chapter 2 we present evidence that CF host inflammatory factors directly contribute to the induction of mutations leading to mucoid conversion, including neutrophils derived from healthy and CF donors, reactive oxygen species, and cationic antimicrobial peptides. In particular, we identified a novel role for the host defense peptide LL-37 as a bacterial mutagen. LL-37 was found to enter bacterial cells at sub-inhibitory concentrations and interact with bacterial DNA. LL-37/DNA interactions promote error-prone replication by the translesion DNA polymerase DinB, which potentiates the mutations responsible for mucoid conversion. We determined that DinB is essential for the induction of mutations within mucA and mucoid conversion in a dinB-deficient strain yields a low frequency of mucoid isolates, which maintain a wild type mucA gene. Approximately 20% of mucoid isolates from CF patients are also found to possess unaltered mucA and in Chapter 3 we interrogate the genetic alterations and mechanism required for mucoid conversion in t (open full item for complete abstract)

Committee: Daniel Wozniak PhD (Advisor); John Gunn PhD (Committee Chair); Larry Schlesinger MD (Committee Member); Sheryl Justice PhD (Committee Member) Subjects: Microbiology

Doctor of Philosophy, The Ohio State University, 2013, Integrated Biomedical Science Graduate Program

Stress is commonly considered to be immunosuppressive, but in some diseases states, such as asthma or infection, stress can be immunoenhancing. This immunoenhancement has been associated with immune cell glucocorticoid resistance that renders the cells insensitive to the anti-inflammatory effects of glucocorticoids. A unique murine social disruption stress paradigm, SDR, can model the stress-induced glucocorticoid resistance and exacerbation of inflammation, which can be relevant to inflammatory diseases in humans. In the context of SDR, stress enhances inflammation and delays resolution in an Aspergillus fumigatus (Af) allergic airway inflammation model. In stressed and Af challenged mice, gene expression data suggested increased inflammation (IL-1ß, TNF-a, GM-CSF) with histological data supporting that the increase was due to infiltrating inflammatory cells. Furthermore, stress and Af challenge most prominently increased granulocytes in the lung compared to controls. Bone marrow chimeras demonstrated that the increase in immune cells was bone marrow-derived, and that stress induced myeloid progenitor cell egress and trafficking to lung. Closer examination of the granulocytic population identified many as neutrophilic populations. Using the antibodies to CD16 and CD49d, several distinct neutrophil populations were visualized including apoptotic, mature, activated, or immature neutrophils. Stress and Af challenge significantly increased the immature neutrophil population in both the lung and blood. In the clinic, it has been shown that a rapid release of immature neutrophils from the bone marrow can occur during times of stress and immune challenge. The consequences of this state of neutrophilia on disease are still being determined, but it is known these neutrophils have a higher capacity to induce inflammation and exacerbate patient symptoms. In a second study, we examined the consequences of Y1 receptor (Y1R), ß-adrenergic receptor (ßAR), and IL-1 receptor t (open full item for complete abstract)

Committee: John Sheridan PhD (Advisor); Michael Bailey PhD (Committee Member); Jonathan Godbout PhD (Committee Member); John Walters DDS, MSc (Committee Member) Subjects: Dentistry; Immunology; Medicine; Microbiology; Neurosciences; Psychobiology; Psychology

Doctor of Philosophy (PhD), University of Toledo, 2013, College of Medicine

Infection of susceptible hosts by the encapsulated Gram-negative bacterium Burkholderia pseudomallei (Bp) causes melioidosis, with septic patients attaining mortality rates greater than or equal to 40%. Due to its high infectivity through inhalation and lack of effective therapies, Bp is considered as a potential bioweapon. Thus, there is great interest in identifying immune effectors important in Bp clearance. Our goal was to compare the relative abilities of murine macrophages and neutrophils to clear Bp in vitro, as well as determine the importance of serum opsonins. Our studies showed that murine macrophages and neutrophils were inherently unable to clear unopsonized Bp or the related relatively-avirulent acapsular bacterium B. thailandensis (Bt). Opsonization of Bp with complement or specific antibodies increased uptake by macrophages, but did not promote clearance, although antibody-binding enhanced complement deposition. However, complement opsonization of Bp resulted in enhanced uptake and rapid killing by neutrophils. Bp killing was linked with rapid induction of reactive oxygen species by neutrophils infected with Bp opsonized with a threshold level of complement deposition (greater than or equal to 5% serum in our system). Addition of Bp-specific antibodies enhanced complement deposition, but antibodies alone were unable to elicit clearance by neutrophils. Macrophages are only observed to efficiently clear Bp after pre-activation with IFN-gamma, which is independent of serum opsonization. These studies indicated that murine macrophages were inefficient in clearing even opsonized Bp unless pre-activated with IFN-gamma, and suggested the requirement of a Th1 response for macrophages to effectively clear Bp. Conversely; neutrophils appeared to efficiently clear serum-opsonized Bp. These findings provide important insight regarding the cell types and immune mediators that should be targeted for melioidosis-related therapies.

Committee: R. Mark Wooten (Advisor); Robert Blumenthal (Committee Chair); Viviana Ferreira (Committee Member); Jason Huntley (Committee Member); Marcia McInerney (Committee Member) Subjects: Immunology; Microbiology

Master of Science, The Ohio State University, 2010, Dentistry

Background: Previous studies have shown that clarithromycin inhibits several periodontal pathogens and is concentrated inside gingival fibroblasts and epithelial cells by an active transporter. We hypothesized that polymorphonucelar cells (PMNs) and other cells of myeloid lineage possess a similar transport system for taking up clarithromycin. In addition, we predicted that clarithromycin accumulation inside PMNs would enhance killing of Aggregatibacter actinomycetemcomitans. Methods: To test the first hypothesis, purified PMNs and cultured HL-60 cells were incubated with [3H]-clarithromycin. Clarithromycin transport was assayed by measuring changes in cell-associated radioactivity over time. The second hypothesis was evaluated with PMNs loaded by brief incubation with clarithromycin (5 μg/ml). Opsonized bacteria were incubated at 37° C with control and clarithromycin-loaded PMNs and in the presence and absence of clarithromycin (5 μg/ml). Results: Undifferentiated HL-60 cells, HL-60 cells that had been differentiated into PMN-like granulocytes, and mature human PMNs all took up clarithromycin in a saturable manner. The kinetics of uptake by all three types of cells yielded linear Lineweaver-Burk plots. PMNs and HL-60 granulocytes transported clarithromycin with a Km of approximately 250μg/ml and a Vmax that was not significantly different. When extracellular concentrations were diluted, HL-60 granulocytes loaded with clarithromycin lost approximately 20% of their antibiotic content within 3 minutes and approximately 45% of their original content within 60 minutes. HL-60 granulocytes and PMNs accumulated intracellular levels of clarithromycin that were 28- to 71-fold higher than their extracellular concentration. When assayed at a bacteria-to-PMN ratio of 100:1, clarithromycin-loaded PMNs killed significantly more bacteria and achieved shorter half times for killing than control PMNs (P < 0.04). At a ratio of 30:1, these differences were not consistently significant (open full item for complete abstract)

Committee: John Walters DDS, MMedSci (Advisor); John M. Nusstein DDS, MS (Committee Member); Hua-Hong Chien DDS, PhD (Committee Member) Subjects: Pharmaceuticals

Doctor of Philosophy, Case Western Reserve University, 2012, Pathology

Filamentous fungal infections of the cornea caused by Aspergillus and Fusarium species are an important cause of blindness and visual impairment worldwide accounting for > 65% of corneal ulcers in developing countries and an estimated 80,000 total cases in India alone. Tissue pathology is often severe leading to either extensive scar tissue formation, cornea transplantation (>10% ), or enucleation of the affected eye. The series of studies described herein delineate our multifaceted approach to investigate: 1. the mechanism by which neutrophils are recruited into fungal-infected corneas, 2. the effectors required by neutrophils to kill fungal hyphae, and 3. the microbial factors utilized by hyphae to resist attack. In collaboration with Aravind Eye Hospital in Tamil Nadu, India, we first characterized the human immune response during fungal keratitis using ulcerative material from infected corneas as well as infected whole corneas from patients undergoing corneal transplantation. Analysis of these tissues identified neutrophils as the major cellular infiltrate with smaller populations of macrophages and T cells. In addition, utilizing qPCR and RNA extracted from fungal-infected human corneas we identified elevated expression of: 1. fungal-sensing cell-surface receptors (Dectin-1, TLR2, and TLR4), 2. neutrophil-recruiting cytokines (IL-8, IL-1¿¿¿¿, IL-17), 3. proteins involved in IL-1 ¿¿¿¿ maturation (NALP3, ASC), and 4. cytokines associated with adaptive immune responses (IFN-gamma ,IL-17, but not IL-4). Having characterized the human immune response during fungal keratitis, we next developed and utilized a novel mouse model of trauma-induced fungal keratitis and knockout mice to identify that ¿¿¿¿-glucan on A.fumigatus germinating conidia activates Dectin-1 on resident macrophages to produce IL-1¿¿¿¿, and CXCL1/KC, which together with IL-1R1/ MyD88-dependent activation, results in recruitment of neutrophils to the corneal stroma and TLR4 but not MD-2 dependent fung (open full item for complete abstract)

Committee: George Dubyak (Committee Chair); Brian Cobb (Committee Member); Mahmoud Ghannoum (Committee Member); Clive Hamlin (Committee Member); Amy Hise (Committee Member); Eric Pearlman (Advisor) Subjects: