Pterygium is an ocular surface disorder characterised by centripetal invasion of the cornea by altered limbal cells, accompanied by fibrosis and neovascularisation. One of the enigmatic features of pterygium is its wing-like shape and the mechanism(s) supporting its centripetal growth remain to be elucidated. As the growth pattern of pterygia mirrors the radial arrangement of corneal nerves, we hypothesised that neuropeptides may facilitate its directional growth. In this thesis, we investigated the roles that the sensory neuropeptide substance P (SP) may play in the pathogenesis of pterygia given its known functions in corneal cell migration, proliferation, wound healing and neurogenic inflammation. Using a modified Boyden chamber method, SP was shown to act as a chemoattractant to pterygium fibroblasts and vascular endothelial cells, and this activity was diminish by blockade of its receptor (NK1). 3H-thymidine incorporation assays confirmed that our cell migration results were unrelated to SP-stimulated proliferation. A bead-based multiplex cytokine array detected secretion of pro-inflammatory cytokines (IL-6, IL-8 and CCL2) from SP stimulated pterygium and limbal epithelial cells. Using real-time RT-PCR and immunoblotting, we show that UVB stimulated transcription of the TAC1 gene followed by secretion of SP in ocular surface epithelial cell cultures. Finally, SP and NK1receptor immunoreactivity was identified in pterygium tissue, where overall, NK1receptors were up-regulated in pterygia. Furthermore, we identified a population of NK1 receptor positive mononuclear cells in pterygia that did not express lineage markers for T or B-Iymphocytes, macrophages or mast cells, but may represent immature haemopoietic cells that may have migrated in from the blood since these cells were also present in autologous conjunctival tissue. In summary, SP may contribute to the shape of pterygia by facilitating migration of fibroblasts and vascular endothelial cells into the normally avascular cornea. Additionally, UVB stimulates SP production in epithelial cells and the presence of SP contributes to inflammation in pterygia by inducing pro-inflammatory cytokine release. Finally, we identified a population of relatively immature, NK1 receptor positive cells in pterygia that may have been attracted by the presence of SP. Collectively, these results imply that SP may contribute to the pathogenesis of pterygia.

No description available.

Thesis advisor: Kenneth C. Williams / HIV infection elicits dysregulation of the immune system and is associated with a number of comorbidities. A common link among HIV-associated comorbidities is monocyte and macrophage activation, accumulation, and high turnover. Anti-retroviral therapy (ART), while useful in preventing HIV infection from progressing to immune dysfunction characterized by AIDS, does not eliminate infection. Even with ART, individuals retain a low level of virus that is able to reseed infection and a higher rate of comorbidities than uninfected people. Prior research has revealed an inflammatory monocyte/macrophage cell population that is uniquely in tissues with infection in an HIV model that uses simian immunodeficiency viral (SIV) infection of rhesus macaques. This cell type is characterized by expression of the inflammatory marker calprotectin. Through measurements of soluble calprotectin present in the plasma of SIV-infected rhesus macaques, I found that calprotectin levels remained low within the first two weeks of infection, sharply increased around three weeks post-infection, typically increased to a maximum during late stage chronic disease, and positively correlated with plasma viral load. Initial calprotectin levels suggests a trend that high pre-infection levels are associated with not progressing to AIDS or SIV encephalitis. Through immunostaining monocytes and flow cytometry, I found that calprotectin expression on classical, intermediate, and non-classical monocytes initially decreases with SIV infection, rebounds for most of infection, and sharply decreases again with late-stage chronic disease. Flow cytometry further showed that the calprotectin-expressing monocyte expresses CD163, CD169, and CCR2, but lacks expression for CX3CR1 and CCR5. Analysis of RNAseq data illustrates trends that suggest an increase in gene expression of genes involved in antiviral/antibacterial and chemotactic functions during conditions when calprotectin gene expression is also increasing. In summary, the data presented in this thesis suggest that the calprotectin-expressing monocyte/macrophage may come from an intermediate monocyte and play a role in inflammation through calprotectin secretion, activation, and increased chemotactic function. / Thesis (MS) — Boston College, 2020. / Submitted to: Boston College. Graduate School of Arts and Sciences. / Discipline: Biology.

Severe anemia is a major life-threatening complication of malaria. Inappropriately low reticulocytosis in malaria patients with anemia suggests insufficient erythropoiesis, of which the mechanisms and implications are not clear. The principle growth factor that promotes erythropoiesis is erythropoietin (Epo). Studies determining the serum level of Epo in malaria infected patients have been inconclusive. Furthermore, the role of Epo and the erythropoietic response to Epo stimulation during malaria have never been examined. The purpose of the experiments performed in this thesis was, thus, to investigate the role of Epo and erythropoiesis in relation to anemia during blood-stage malaria using the murine model of Plasmodium chabaudi AS. A murine Epo specific ELISA, which was determined to be less biased by the presence of other cytokines in the samples as compared to the conventional Epo bioassay, was first developed to facilitate the research. The kinetics of Epo production in the kidney and the levels in the serum were characterized. It was demonstrated that Epo production during blood-stage malaria is mainly regulated by the degree of anemia and that renal cytokines may have only a minor effect on this response. Next, the roles of Epo and erythropoiesis during blood-stage malaria were investigated by neutralization of endogenous Epo or by administration of exogenous Epo. Timely onset of Epo-induced reticulocytosis was shown to be important for the alleviation of malarial anemia and survival. However, reticulocytosis in response to Epo stimulation is severely suppressed by infection with malaria. Dissection of the upstream events of erythropoiesis demonstrated that blood-stage malaria compromises the generation of reticulocytes by suppressing the proliferation, differentiation, and maturation of erythroid-lineage cells at various stages of erythroid development. Taken together, our data provide important insights for understanding the patho

Hypermetabolism, observed in Sydenham's chorea (SC); a complication of acute rheumatoid fever (ARF) involving binding of streptococcal M protein antibodies in the brain, may result from an increase in glutamate release. The interaction of mAb 1 OF5, a specific M protein antibody subtype, with brain proteins (e.g. Rabphilin-3A), synaptic vesicles (SVs) and synaptosomal fraction (SF) was examined. Rat brain slices immunostained with mAb l OF5 revealed an interaction with choroid plexus and elements appearing to be neuropils. Dot blotting demonstrated an interaction of mAb I OF5 with both SVs and SFs. Western blotting revealed a smear from mAb 10F5 against the SF fraction. However, both modified SVs and pure liposomes examined by fluorescent and confocal microscopy bound mAbl0F5 suggesting a direct interaction with phospholipids. ELISA demonstrated binding of mAb1OF5 with negatively charged phospholipids involved in antiphospholipid syndrome (APS). Hypermetabolism and binding at the choroid plexus is observed in SC and APS supporting the connection between these disorders. / Department of Physiology and Health Science

No description available.

Background: Crohn's disease is a chronic inflammatory disease of the intestine in humans, with no known cause. Johne's disease is a chronic intestinal disease of ruminants caused by Mycobacterium avium suhspecies paratuberculosis (MAP), and has some features similar to Crohn's disease. Although MAP has been purported to play an etiologic role in Crohn's disease, this causal link is still under debate. Objective: The overall aim of this project is to analyze MAP strains from different hosts (cattle, sheep and humans) and regions in North Dakota by biochemical and epidemiological methods, in order to better understand the pathogenesis and epidemiology of MAP strains and the relationship between MAP and Crohn's disease. The specific aims of this research are the following: Aim 1. Investigate the epidemiological evidence for MAP as a cause of Crohn's disease. Aim 2. Conduct a comparative causality study to investigate whether MAP or other enteric pathogens cause Crohn's disease. Aim 3. Evaluate the occurrence of MAP infections in cattle in North Dakota, 1995-2005. Aim 4. Analyze MAP strains from symptomatic and asymptomatic cattle. Aim 5. Investigate the biochemical variations of rapid and slow growing MAP strains. Aim 6. Evaluate MAP strains from low shedders and high shedders. Methods: MAP isolates were analyzed by biochemical methods of gas chromatography, high performance liquid chromatography and mass spectrometry. In addition, extensive literature review was performed to (1) determine the epidemiologic causal link between MAP and Crohn' s disease and (2) determine whether MAP or other enteric pathogens cause Crohn's disease. Results: Results from our study indicated the availablity of epidemiologic evidence supporting the causal role of MAP in Crohn's disease. It was also demonstrated that MAP is the most implicated organism in the etiology of Crohn's disease when compared to other infectious agents. Investigation of the occurrence of MAP infection in North Dakota showed an increase in number of MAP cases reported, with seasonal trends. Biochemical typing of MAP strains from symptomatic and asymptomatic cattle indicated that the symptoms status of isolates was significantly associated with mass spectra patterns and shedder status (p < 0.05). However, the association between symptoms status and HPLC and GC patterns was not significant (p > 0.05). Investigation of biochemical variations of rapid and slow growing MAP strains showed associations between the biochemical variability of MAP strains and their growth rate and presence of symptoms in the source cattle. Evaluation of MAP strains of different shedding characteristics by univariate logistic regression revealed that the shedder status of isolates was significantly associated with growth rate of isolates, symptom status, and source regions, but not with mass spectra patterns of isolates. Conclusion: Overall, this study strengthens the theories of strain sharing, intraspecies and interspecies transmission, and supports an association between MAP and Crohn's disease. In addition, the understanding of the biochemical variation among MAP isolates will help in the future design of diagnostics, therapeutics and vaccines for Johne's and Crohn's diseases. / North Dakota State University. Department of Chemistry and Biochemistry

No description available.

Parkinson's disease (PD) is a neurodegenerative disorder with no known cure. This thesis explores the degenerative process in two neurotoxin-based models, the 6-hydroxydopamine and the chronic 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine(MPTP)/probenecid mouse models, to yield important information about the pathogenesis of PD. Neuronal survival patterns in Parkinsonian patients and animals are heterogeneous. More dopaminergic neurons are lost from the ventral tier of the substantia nigra (SN) than from the dorsal tier or the adjacent ventral tegmental area, possibly due to differential expression of the calcium-binding protein, calbindin D28K. Brain sections were processed for tyrosine hydroxylase (TH) and calbindin (CB) immunocytochemistry to distinguish the dopaminergic subpopulations. I show that more TH+/CB- and TH-/CB+ than TH+/CB+ neurons are lost in both models, suggesting that CB confers some degree of protection for dopaminergic neurons. With respect to connectivity, I show that both TH+ and CB+ neurons receive striatal and dorsal raphe inputs. I investigated the possibility of a progressive loss in midbrain neurons by prolonging the post-lesion survival period. In both models, there is an irreversible neuronal cell loss of TH+, CB+ and TH+/CB+ neurons but the effects of survival time and lesion treatments differ for the three neuronal types. The lesions also appear to be toxic to GABAergic neurons. I explore whether, once neurodegeneration has started, neurons can be rescued by pharmacological intervention. Salicylic acid appears both to reduce microglial activation and significantly improve TH+, but not CB+ or TH+/CB+ neuronal survival. PD appears multifactorial in origin and may involve complex interactions between genetic and environmental influences. I show that a xenobiotic-metabolising enzyme, arylamine N-acetyltransferase may fulfil a neuroprotective role in the SN by limiting the environmental risks. Taken together, this study provides a body of information on two different mouse PD models and highlights possible genetic predispositions to PD neuropathology.

published_or_final_version / abstract / toc / Medicine / Master / Doctor of Medicine