Thesis (M.A.) / Systemic sclerosis (SSc) is a clinically heterogenous chronic fibrotic disease which affects skin and internal organs. While the pathogenesis of SSc remains unknown, the hallmark of both localized and diffuse SSc in the skin is the replacement of normal dermal architecture with excessive deposition of collagen and other connective tissue macromolecules. Progressive replacement of tissue architecture by collagen-rich extracellular matrix (ECM) results in functional impairment of affected organs. Fibrotic damage to these affected organs accounts for much of the morbidity and mortality concomitant with SSc, particularly in the lungs. Myofibroblasts are the primary ECM-secreting cells during wound healing and fibrosis. Myocardin-related transcription factor A (MRTF-A), is an important regulator of myofibroblast differentiation, depending on serum response factor (SRF) for smooth muscle actin (SMA) and Sp1 in the regulation of collagen gene expression. MRTF-A continually shuttles between the nucleus and cytoplasm in unstimulated cells. Signals of stress, mechanical force, and migration control MRTF-A movement by a mechanism in which Rho-activated cytoskeletal actin polymerization induces its relocation from the cytoplasm to the nucleus. The major hypothesis in this thesis is that MRTF-A is dysregulated (impairment of a physiological regulatory mechanisms) and/or activated in SSc patients in part through transforming growth factor beta (TGF-β). To test this hypothesis, immunohistochemistry using MRTF-A antibodies was performed on SSc patient skin lesions and healthy control skin. Staining was observed in the epidermis, epidermal structures, vasculature and dermis of SSc and healthy control skin. In the epidermal layer of patients with SSc, there was significantly more nuclear localization of MRTF-A then in normal controls. Prominent staining is also present in endothelial, perivascular and some perivascular inflammatory cells of SSc patients. Perivascular staining was not seen in healthy controls. Interestingly, there was some accumulation of nuclear MRTF-A in areas typical of myofibroblasts in SSc skin, but this staining is not as striking as vascular staining. TGF-β activates MRTF-A in a cell-specific manner. As SSc typically begins within the skin, human dermal fibroblasts (HDF) were grown in culture. HDFs synthesize and secrete collagen to a greater extent when compared to human lung fibroblasts (IMR90 cells). Treatment with TGF-β enhances cytoplasmic localization of MRTF-A at 4-8 hours in HDFs and prolongs nuclear localization. Transgenic mouse lung cells were isolated from an MRTF-A loss-of-function mouse carrying the 3.6 kb proximal promoter of the rat COL1A1 gene driving topaz green fluorescent protein (GFP) (pOB3.6COLGFPtpz). Since angiotensin II (ANG II) may enhance TGF-β response or collagen transcription directly, wild type (WT) and MRTF-A knockout (KO) cells were treated with ANG II and TGF-β. Quantification of collagen transcription by GFP fluorescence and protein synthesis by Western and secretion by Sircol analysis revealed collagen gene expression is consistently lower in KO fibroblasts compared to WT. Total percentage of fluorescent KO cells were consistently lower in comparison to WT cells as well. KO cells do not respond to TGF-β or ANG II treatment, whereas TGF-β increased collagen gene expression by WT cells, but not KO cells. Furthermore, treatment with ANG II did not up-regulate transcription in WT mouse lung fibroblasts. However, TGF-β receptor kinase 1 (TβR-1) inhibitor SB431542 attenuated collagen transcription in both WT and KO fibroblasts regardless of treatment suggesting that the receptor is active with or without MRTF-A possibly with an endogenous ligand produced by these cells. The activation of MRTF-A is an important protein regulating collagen synthesis and may potentially serve as a therapeutic target in future treatments of fibrotic disease such as SSc.

Thesis (M.S.)--Boston University

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No description available.

La sclérose en plaques est une maladie chronique, auto-immune et neurodégénérative qui se traduit par l’apparition de plaques inflammatoires démyélinisantes dans le système nerveux central. Pour caractériser la réponse immunitaire innée, j’ai développé des outils d’imagerie optique non linéaire multispectrale intravitale, et des solutions semi automatisées d’analyse d’images. J’ai participé à la mise au point d’un protocole d’identification des phénotypes immunitaires en cytométrie. Ces outils appliqués à l’étude de l’encéphalomyélite auto-immune expérimentale, un modèle murin de SEP, ont permis d’analyser la cascade immunitaire par immunomarquages et marquages fluorescents transgéniques, de décrire les distributions des populations cellulaires dans le SNC, et de les associer à la dégradation axonale sur des échelles de temps de la seconde à la semaine. Sur une lignée de souris transgénique, Thy1-CFP/Cd11c-EYFP/LysM-EGFP, j’ai suivi en microscopie biphotonique l’évolution des densités de cellules fluorescentes dans la moelle épinière pendant plusieurs semaines. J’ai conclu que la dégénération axonale et les déficits moteurs sont corrélés avec l’infiltration par les méninges de neutrophiles et monocytes. Les monocytes se différencient in situ en cellules dendritiques d’origine monocytaire (moDCs) parallèlement à l'activation de la microglie. Ces événements cellulaires, dont la maturation des moDCs sont corrélés avec la résorption de l’inflammation. La méthodologie est en place pour poursuivre ces investigations sur d’autres modèles. L’optimisation du microscope m’a aussi permis d’accéder simultanément au contraste endogène CARS pour visualiser la gaine de myéline. / Multiple sclerosis is a chronic auto-immune neurodegenerative disease characterized by the appearance of inflammatory plaques in the central nervous system. To characterize the innate immune response I have developed nonlinear optical tools for intravital multispectral imaging as well as semi-automated image processing solutions. I also contributed to the development of a flow cytometry protocol allowing the identification of immune cell phenotypes. Applied to study of Experimental Autoimmune Encephalomyelitis, a mouse model of MS, these tools have allowed to analyze the immune cascade thanks to immunolabelings and transgenic expression of fluorescence, describe the distributions of cell populations in the CNS with regard to neuron degeneration on time scales ranging from seconds to weeks. On a transgenic mouse line Thy1-CFP/Cd11c-EYFP/LysM-EGFP, I have followed by two-photon microscopy the evolution of fluorescent cells densities in the same area of the spinal cord for several weeks. I conclude that axonal degeneration and motor deficits are correlated with neutrophils and monocytes infiltration from the meninges. The monocytes differentiate in situ in monocyte-derived dendritic cells (moDCs) along with the recruitment of activated microglia. These cellular events correlated with a stabilization/remission phase of the disease. MoDCS maturation thus seems involved in the dampening of inflammation. Methodology and tools are now set for further investigations with other models. The microscope optimization for multicolor excitation allowed me to access simultaneously to the endogenous CARS contrast to visualize the myelin sheath.

Cette thèse propose d'explorer les capacités en mémoire épisodique de patients atteints de sclérose en plaques (sep). A notre connaissance, l'analyse des processus touchés et préservés a fait l'objet de peu de travaux. L'une des raisons historiques semble être la création, dès les années 1980, de batteries d'évaluation rapide destinées aux neurologues. Ainsi, le golden standard de l'évaluation mnésique dans cette affection est le srt (dujardin et al., 2004) qui permet difficilement de contrôler les troubles d'encodage et de récupération. L'objectif de cette thèse sera dans un premier temps de documenter à l'aide d'épreuves permettant une analyse des processus mnésiques, les capacités de mémoire épisodique verbale dans la sep. Les performances de trois groupes de patients (sep, maladies d'Alzheimer et de Parkinson) seront comparées dans l'épreuve rl/ri-16 items. Cette étude sera destinée à vérifier si une majorité de patients sep présente une amélioration du rappel libre après délai contrairement aux deux autres affections testées. Si tel est le cas, l'hypothèse d'un temps de traitement, insuffisamment long pour les patients sep entre les rappels immédiats, mériterait d'être testée. Pour cela, une version parallèle du cvlt sera créée. Ainsi, les performances des patients seront comparées intra-individuellement lors de l'administration de l'épreuve dans sa version originale et lors d'une version où les temps entre les présentations de la liste de courses seront allongés. Un troisième volet sera destiné à tester les capacités de consolidation à long terme (une étude dans ce domaine pour la sep). Une analyse anatomo-fonctionnelle sera conduite en éprouvant l'hypothèse d'un lien entre les performances de rappel à une semaine et la volumétrie hippocampique des patients sep. / This thesis suggests exploring capacities in memory episodic of patients reached by multiple sclerosis. To our knowledge, the analysis of the affected and protected processes was the object of few works. One of the historic reasons seems to be the creation, from the 1980s, of batteries of fast evaluation intended for the neurologists. So, standard Golden of the mnesic evaluation in this affection is the srt (dujardin and al ., on 2004) which allows with difficulty to check) the disorders of encoding and recovery. The objective of this thesis will at first be to document by means of tests allowing an analysis of the mnesic processes, capacities of verbal episodic memory in frog. The performances of three groups of patients (frog, Alzheimer's diseases and of Parkinson) will be compared in the test rl / ri-16 items. This study will be intended to verify if a majority of patients frog presents an improvement of the free reminder after deadline contrary to two other tested affections. If that was the case, the hypothesis of a processing time, insufficiently long for the patients frog between the immediate reminders), would deserve to be tested. For that purpose, a parallel version of the cvlt will be created. So, the performances of the patients will be compared intra - individually during the administration of the test in its original version and during the version where the times between the presentations of the shopping list will be lengthened. The third shutter will be intended to test the capacities of long-term consolidation (a study in this domain for frog). An anatomo-functional analysis will be led by feeling the hypothesis of a link between the performances of reminder in a week and the hippocampal volumetry of the patients frog.

The purpose of the present study was to examine whether or not luminance thresholds through static, chromatic perimetry could be used to distinguish visual threshold losses in multiple sclerosis from that of normal functioning. It was proposed that threshold losses would be greater at both the fovea and near foveal eccentricities due to the assumption that the cone system, unlike the rods, would be the most effected by MS. Twenty-two MS patients and thirty age matched normals were tested on an extensively modified version of the Fieldmaster F225 Automatic Perimeter. Thresholds were established for an achromatic, red, and blue stimulus along a 195 - 15 degree meridian. Testing was done using a 45 apostilb background, to which the subjects were preadapted prior to testing. Results indicated that there was extensive cone involvement (loss in chromatic thresholds) for the MS subjects. Significant differences existed at the fovea between normal and clinically definite subjects but not between normal and probable. Correlational analyses indicated great functional changes in retinal sensitivity for the MS patients. Similar results were obtained between MS patients with and without optic neuritis. Discriminant analyses indicated that the red filter could correctly classify 86.27% of the normals and MS patients with few false positives or negatives. Log threshold difference values between the fovea and 30 degree nasal eccentricity were used to determine a threshold value which could separate normal profiles from MS profiles. The typical "swiss cheese" defects reported in the clinical literature were found only for the achromatic and blue filters. No irregular profiles were found for the red filter. A possible theoretical model based on the results was discussed. Limitations of the study as well as possible future research were also discussed. / Arts, Faculty of / Psychology, Department of / Graduate

Thesis (M.D.)--Boston University

As current treatments for multiple sclerosis (MS) do not prevent the accumulation of long-term disability, researchers and persons with MS are interested in wellness behaviours and how they may be used to manage MS. This thesis includes a review of the literature on wellness-based interventions in persons with progressive MS. Following this review, a cross-sectional study was conducted to characterize dietary intake by disability status in this population, and to examine the functional and symptomatic correlates of dietary behaviours. Participants with MS and matched controls completed questionnaires and a three-day food intake record. There were significant differences in dietary intake with and without supplements between the MS and control groups. Correlates of dietary behaviours were also examined. Further research examining dietary intake in MS is necessary to understand how disability and other factors impact dietary intake behaviours, and which other correlates may be useful targets for future nutrition interventions.

Indiana University-Purdue University Indianapolis (IUPUI) / The purpose of this investigation was to evaluate the association between the depth of a carious lesion and the sclerotic dentin deposited beneath a calcium hydroxide methyl cellulose base material. The sample chosen consisted of teeth with deep caries and possible pulp exposure, as evidenced by a critical radiographic examination. Clinical procedures consisted of a preoperative serial radiograph, followed by complete caries removal. A barium sulphate radiopaque solution was then applied to the base of the preparation, followed by a second serial radiograph. The barium sulphate was removed and a calcium hydroxide methyl cellulose base was applied and the tooth restored with a silver amalgam alloy. Subsequently one, three, six, and nine month serial radiographs were taken postoperatively. Calcification change of sclerotic dentin overlying the pulp was measured in relation to pulpal floor width by the television instrumentation. The conclusions of this study are as follows: 1. A calcium hydroxide methyl cellulose base material acts as a "trigger mechanism," in deep cavities, stimulating the deposition of sclerotic dentin. 2. The thinner the initial pre-restored pulpal floor, the more rapid and dramatic is the postoperative pulpal floor increase in width, which is apparently a protective pulpal response. 3. During the periods of time when a significant increase in pulpal floor width was noted, the percent change in calcification of this area decreased. Conversely, when little pulpal floor width increase occurred, the percent change in calcification increased.