Immune potential and differentiation of equine induced pluripotent stem cells (eiPSC)

Induced pluripotent stem cells (iPSC) have the capacity to self renew and differentiate into a myriad of cell types making them potential candidates for cell therapy and regenerative medicine. The goal of this thesis was to determine the characteristics of equine iPSC (eiPSC) that can be harnessed for potential use in veterinary regenerative medicine. Trauma to a horse’s limb often leads to the development of a chronic non-healing wound that lacks a keratinocyte cover, vital to healing. Thus, the overall hypothesis of this thesis was that eiPSC might offer a solution for providing wound coverage for such problematic wounds. Prior to considering eiPSC for clinical applications, their immunogenicity must be studied to ensure that the transplanted cells will be accepted and integrate into host tissues. The first objective of this thesis was to determine the immune response to eiPSC. To investigate the immunogenicity of eiPSC, the expression of major histocompatibility complex (MHC) molecules by the selected lines was determined, then the cells were used in an intradermal transplantation model developed for this study. While transplantation of allogeneic, undifferentiated eiPSC elicited a moderate cellular response in experimental horses, it did not cause acute rejection. This strategy enabled the selection of weakly immunogenic eiPSC lines for subsequent differentiation into lineages of therapeutic importance. Equine iPSC offer a potential solution to deficient epithelial coverage by providing a keratinocyte graft with the ability to differentiate into other accessory structures of the epidermis. The second objective of this thesis was to develop a protocol for the differentiation of eiPSC into a keratinocyte lineage. The protocol was shown to be highly efficient at inducing the anticipated phenotype within 30 days. Indeed, the eiPSC derived vi keratinocytes (eiPSC-KC) showed both morphologic and functional characteristics of primary equine keratinocytes (PEK). Moreover, the proliferative capacity of eiPSC-KC was superior while the migratory capacity, measured as the ability to epithelialize in vitro wounds, was comparable to that of PEK, suggesting exciting potential for grafting onto in vivo wound models. In conclusion, equine iPSC-derived keratinocytes exhibit features that are promising to the development of a stem cell-based skin construct with the potential to fully regenerate lost or damaged skin in horses. However, since eiPSC do not fully escape immune surveillance despite low MHC expression, strategies to improve engraftment of iPSC derivatives must be pursued.

Étude comparative de l'imagerie dans l'évaluation de l'ostéoarthrose du grasset chez le cheval

Les objectifs de cette étude étaient (1) de décrire la localisation et la sévérité des lésions d’ostéoarthrose (OA) chroniques acquises naturellement au niveau du grasset équin grâce à l’échographie (US), la radiographie (XR), la tomodensitométrie (CT) et l’évaluation macroscopique (ME), (2) de comparer la performance diagnostique de chacune des modalities d’imagerie avec ME et (3) d’évaluer quantitativement la densité osseuse sous-chondrale lors d’OA du grasset chez le cheval à la tomodensitométrie. Des évaluations post mortem radiographique, tomodensitométrique et échographique ont été réalisées sur 23 grassets cadavériques et comparées à l’évaluation macroscopique. Des associations significatives ont été notées entre le «osteophytes global score» de toutes les modalités (US, p=0.04; XR, p=0.005; CT, p˂0.0001) et ME. De plus, la tomodensitométrie a démontré la plus forte association. Les ostéophytes étaient principalement localisés au niveau de l’articulation fémorotibiale médiale et cette articulation présentait également les scores d’ostéophytes les plus sévères. Un patron spécifique d’ostéophytes associé à l’insertion de la capsule articulaire sur le condyle fémoral médial a été mis en évidence. La nouvelle projection radiographique (Ca10Pr5L-CrDiMO) a été utile dans la détection des ostéophytes de la région intercondylaire. Les grades d’ostéophytes (0-3) ne différaient pas significativement selon la modalité dans la majorité des sites. La faible sensibilité/spécificité a indiqué que la sclérose de l’os sous-chondral et l’applatissement des condyles fémoraux ne semblent pas être des indicateurs fiables d’OA du grasset équin. L’OA du grasset équin est associée à une réduction de la densité osseuse sous-chondrale et des sites spécifiques de résorption/kystes sous-chondraux ont été notés chez certains spécimens.

Développement d’une approche quantitative pour l’étude du poumon équin : fixation et échantillonnage pour l’application des principes de la stéréologie

La présente étude visait à développer un protocole de fixation et d'échantillonnage pour le poumon équin suivant les directives publiées sur l’utilisation d’une approche stéréologique de type « design-based ». Les poumons gauches de chevaux contrôles et atteints du souffle ont été fixés avec du formaldéhyde 10% pendant 48h à une pression constante de 25-30 cm d’H2O. Les poumons ont été sectionnés en 20-21 tranches d’une épaisseur d'environ 2,5 cm chacune; de 10-11 tranches ont été sélectionnées de façon aléatoire et systématique pour la mesure du volume de référence avec la méthode de Cavalieri. Un protocole d’échantillonnage systématique, aléatoire et uniforme utilisant le principe du « smooth fractionator » et un poinçon à biopsie de 17 mm ont été utilisés pour échantillonner une fraction représentative de chaque poumon. Les méthodes d’échantillonnage de sections verticales, uniformes et aléatoires (VUR) et d’échantillonnage isotropique, uniforme et aléatoire (IUR) ont toutes deux été effectuées pour comparer le nombre de voies respiratoires en coupe perpendiculaire obtenues à partir de chaque méthode. L'architecture globale et la qualité des tissus fixés ont également été évaluées. Des spécimens pulmonaires équins ont été échantillonnés avec succès selon un protocole visant à produire des données morphométriques valides. Les tissus ont été fixés avec un minimum d'artéfacts et contenaient une quantité suffisante de voies respiratoires en coupe perpendiculaire dans les deux types d’échantillons. En conclusion, un protocole de fixation et d'échantillonnage adapté au poumon équin permettant l'utilisation d'une approche stéréologique de type « design-based » a été élaboré pour l’étude du remodelage des voies respiratoires.

Assessment of meat authenticity using bioinformatics, targeted peptide biomarkers and high-resolution mass spectrometry

In recent years, we observed a significant increase of food fraud ranging from false label claims to the use of additives and fillers to increase profitability. Recently in 2013, horse and pig DNA were detected in beef products sold from several retailers. Mass spectrometry has become the workhorse in protein research and the detection of marker proteins could serve for both animal species and tissue authentication. Meat species authenticity will be performed using a well defined proteogenomic annotation, carefully chosen surrogate tryptic peptides and analysis using a hybrid quadrupole-Orbitrap mass spectrometer. Selected mammalian meat samples were homogenized, proteins were extracted and digested with trypsin. The samples were analyzed using a high-resolution mass spectrometer. The chromatography was achieved using a 30 minutes linear gradient along with a BioBasic C8 100 × 1 mm column at a flow rate of 75 µL/min. The mass spectrometer was operated in full-scan high resolution and accurate mass. MS/MS spectra were collected for selected proteotypic peptides. Muscular proteins were methodically analyzed in silico in order to generate tryptic peptide mass lists and theoretical MS/MS spectra. Following a comprehensive bottom-up proteomic analysis, we were able to detect and identify a proteotypic myoglobin tryptic peptide [120-134] for each species with observed m/z below 1.3 ppm compared to theoretical values. Moreover, proteotypic peptides from myosin-1, myosin-2 and -hemoglobin were also identified. This targeted method allowed a comprehensive meat speciation down to 1% (w/w) of undesired product.

L’origine géographique en tant que facteur explicatif de la cyberdélinquance

Ce mémoire propose que l’origine d’un cyberdélinquant soit un facteur explicatif du phénomène de la cybercriminalité. Il comporte deux objectifs : premièrement, décrire les profils des cybercriminels recensés dans les médias internationaux; deuxièmement, vérifier si ces profils varient selon le lieu d’origine du cyberdélinquant. Une base de données, comportant 90 cas de cybercriminels répertoriés à travers le monde, fut créée. Quinze (15) cybercriminels par territoire ont été sélectionnés, les régions ciblées allant comme suit : Amérique du Nord, Amérique latine, Australasie, Europe de l’Ouest, Eurasie et Afrique/péninsule Arabique. En premier lieu, des analyses descriptives ont été exécutées afin de dresser un portrait de ce phénomène. En second lieu, des analyses de tableaux de contingence ont été effectuées entre les variables à l’étude afin de voir si des relations existaient. Enfin, d’autres analyses de tableaux de contingence ont été réalisées afin d’établir les différences des paramètres en fonction de l’origine. Les résultats de ces divers tests démontrent que ce sont généralement de jeunes hommes âgés en moyenne de 25 ans qui seront susceptibles de commettre des délits informatiques. Quelques profils types se sont dégagés des analyses et peuvent s’expliquer par l’accès au matériel informatique, les inégalités économiques entre classes sociales, tout comme la vitesse d’Internet, et ce, en fonction de l’origine du cyberdélinquant.

L’effet de l'interleukine-17 sur la survie et la glucocorticorésistance du neutrophile équin

Les glucocorticoïdes sont les médicaments les plus efficaces pour le contrôle de l'obstruction respiratoire chez les chevaux atteints du souffle, et de l'asthme humain. Toutefois, les neutrophiles persistent dans les voies respiratoires suite à ce traitement. Nous avons précédemment rapporté que les neutrophiles sanguins humains et équins sont sensibles à l'action des glucocorticoïdes. Comme elle contribue à l'insensibilité des cellules épithéliales pulmonaires humaines aux glucocorticoïdes, nous avons émis l'hypothèse que l'IL-17 a un effet similaire sur les neutrophiles et qu’elle contribue à leur persistance dans les voies respiratoires asthmatiques. Objectifs : Évaluer 1. L’expression des deux sous-unités du récepteur de l’IL-17 (l'IL-17RA/IL-17RC) chez les neutrophiles équins hautement purifiés. 2. Si l'IL-17 active directement les neutrophiles et si cette réponse est sensible à l'action des glucocorticoïdes. 3. L'effet de l'IL-17 sur la viabilité et l'apoptose des neutrophiles. Résultats: 1. Les neutrophiles expriment l’IL-17RA/IL-17RC aux niveaux translationnel et protéique. 2. L’IL-17 induit une activation sélective des neutrophiles (surrégulation de l’IL-8), qui n’est pas atténuée par dexaméthasone et 3. l’IL-17 augmente la viabilité des neutrophiles stimulés (LPS) par une diminution de l'apoptose. Nos résultats indiquent que l'IL-17 active directement le neutrophile équin, et que l’augmentation de l’IL-8 (puissant chimioatractant des neutrophiles) qui en résulte n’est pas contrôlée par la dexaméthasone. L'IL-17 pourrait aussi contribuer à la persistance de neutrophiles dans les voies respiratoires chez les chevaux atteints du souffle, en diminuant l'apoptose.

A Diagnostic and Modeling Study on the Onset of Summer Monsoon over South Asia

This study focuses on the onset of southwest monsoon over Kerala. India Meteorological Department (IMD) has been using a semi-objective method to define monsoon onset. The main objectives of the study are to understand the monsoon onset processes, to simulate monsoon onset in a GCM using as input the atmospheric conditions and Sea Surface Temperature, 10 days earlier to the onset, to develop a method for medium range prediction of the date of onset of southwest monsoon over Kerala and to examine the possibility of objectively defining the date of Monsoon Onset over Kerala (MOK). It gives a broad description of regional monsoon systems and monsoon onsets over Asia and Australia. Asian monsoon includes two separate subsystems, Indain monsoon and East Asian monsoon. It is seen from this study that the duration of the different phases of the onset process are dependent on the period of ISO. Based on the study of the monsoon onset process, modeling studies can be done for better understanding of the ocean-atmosphere interaction especially those associated with the warm pool in the Bay of Bengal and the Arabian Sea.

Low Level Jet stream of Asian Summer Monsoon and its Variability

The main objective of the of present study are to study the intraseasonal variability of LLJ and its relation with convective heating of the atmosphere, to establish whether LLJ splits into two branches over the Arabian sea as widely believed, the role of horizonatal wind shear of LLJ in the episodes of intense rainfall events observed over the west coast of India, to perform atmospheric modeling work to test whether small (meso) scale vortices form during intense rainfall events along the west coast; and to study the relation between LLJ and monsoon depression genesis. The results of a study on the evolution of Low Level Jetstream (LLJ) prior to the formation of monsoon depressions are presented. A synoptic model of the temporal evolution of monsoon depression has been produced. There is a systematic temporal evolution of the field of deep convection strength and position of the LLJ axis leading to the genesis of monsoon depression. One of the significant outcomes of the present thesis is that the LLJ plays an important role in the intraseasonal and the interannual variability of Indian monsoon activity. Convection and rainfall are dependent mainly on the cyclonic vorticity in the boundary layer associated with LLJ. Monsoon depression genesis and the episodes of very heavy rainfall along the west coast of India are closely related to the cyclonic shear of the LLJ in the boundary layer and the associated deep convection. Case studies by a mesoscale numerical model (MM5) have shown that the heavy rainfall episodes along the west coast of India are associated with generation of mesoscale cyclonic vortices in the boundary layer.

Geochemistry of Rare Erath Elements and Trace Metals along the Western Continental Shelf of India

Rare earth elements have occupied an important role in marine geochemical research, particularly as used in the format of REE abundance patterns to describe the geochemical pathways in marine sedimentation and authigenesis. This study concentrates on the distribution pattern of Rare earth elements in the sediments, behavior of Eu and Ce with respect to their occurrence in multiple oxidation states. It also concentrate the depth wise variation of sediment REEs from near shore areas (30m) to deeper depths 200m) in the Arabian Sea. It includes the downcore variation of REEs and other trace elements in the sediment cores and a comparison between the REE distributions of Arabian Sea sediment with the sediments of Andaman Sea. The study gives a general introduction regarding the importance of RRE studies, its occurrence and abundance, electronic configuration, lanthanide contraction, oxidations states and REE supply to the ocean, seawater and sediments.

Salient Features of the North Indian Ocean Associated with the Indian summer Monsoon

This study deals with the salient features of the north Indian ocean associated with the summer monsoon. The focus is given on the Arabian sea mini warm pool, which is a part of the Indian ocean. It primarily study the certain aspects of the atmosphere and ocean variability in the north Indian ocean. The attempt were made to understand various aspects of time –scale variability of major features occurring in the Indian summer monsoon. The result from the thesis can be utilized as an input for model studies for prediction of monsoon, understanding ocean dynamics, radar tracking and ranging etc.

Nutrient Dynamics in the EEZ of the West Coast of India with Special Reference to the Oxygen Minimum Zone and Denitrification

The present work aims at deciphering the processes that control the nutrient distribution along the EEZ (Exclusive Economic Zone of India) of the west coast of India and to bring out its linkage with primary and secondary productivity. This work assume utmost importance as very few studies have hitherto focused entirely on the EEZ of the west coast of India to address the biochemical responses brought about by monsoons. The present study examines the seasonal variations in physicochemical parameters and associated primary biological responses along the west coast of India. This study targets to measure and understand the shelf ocean exchange in a typical coastal upwelling region of the southeast Arabian Sea, and the influence of convective mixing along the northern part of the west coast of India. The study focuses more directly on coastal upwelling along the southwest coast of India, within the EEZ. The effects of coastal upwelling, eddy formation and the offshore advection are apparent in the present investigation. This has consequences to fisheries and climate, in energy transfer to the food chain and the increased sequestering of carbon in the ocean. The study also focuses on the Oxygen Minimum Zone (OMZ) and dentrification observed along the EEZ of the west coast of India on a seasonal scale. In the study, an attempt is also made to demarcate the geographical boundaries of the denitrification zone in the EEZ of India and on the nature and magnitude of these variations, on a seasonal and inter annual scales

Mixed layer characteristics and hydrography off the west and east coasts of India

In this thesis, a detailed attempt has been made to understand the general hydrography of the upper 300m of the water column, in the eastern Arabian Sea and the western Bay of Bengal, the two contrasting basins in the northern Indian Ocean, using recently collected data sets of Marine Research-Living Resources (MR-LR) assessment programme, funded by Department of Ocean Development, from various cruises, pertaining to different seasons. Initially it discuss the general hydrography of the west and east coasts of India are covered, in the context of mixed layer processes. The study describes the materials and methods . To compare the hydrography of the AS and BOB, a unique MLD(Mixed Layer Depth) definition for AS and BOB is essential, for which the 275 CTD profiles were used. A comparison has been made among the various MLD criteria with the actual MLD. The monthly evolution of MLD, barrier layer thickness and the role of atmospheric forcing on the dynamics of the mixed layer in the AS and BOB were studied. The general hydrography along the west coast of India is described. The upwelling/downwelling, winter cooling processes, in the context of chemical and biological parameters, are also addressed. Finally the general hydrography of the Bay of Bengal is covered. The most striking feature in the hydrography are the signature of an anticyclonic subtropical gyre during spring intermonsoon and a cold core eddy during winter monsoon. The TTS(Typical Tropical Structure) of the euphotic layer was also investigated.

Salient Features of the North Indian Ocean Associated with the Indian summer Monsoon

This study deals with the salient features of the north Indian ocean associated with the summer monsoon. The focus is given on the Arabian sea mini warm pool, which is a part of the Indian ocean. It primarily study the certain aspects of the atmosphere and ocean variability in the north Indian ocean. The attempt were made to understand various aspects of time –scale variability of major features occurring in the Indian summer monsoon. The result from the thesis can be utilized as an input for model studies for prediction of monsoon, understanding ocean dynamics, radar tracking and ranging etc.

Seasonal Studies on Primary Production and Associated Environmental Parameters in the Indian EEZ

The thesis describes the importance of Indian EEZ, definition and the various factors affecting primary production, general account of phytoplankton and its importance in marine ecosystem etc. In review of literature, general oceanography of Arabian Sea and Bay of Bengal and hydrography of eastern Arabian Sea and western Bay of Bengal. It deals with the distribution patterns of primary production, chlorophyll a, phytoplankton composition and particulate organic carbon in the eastern Arabian Sea and western Bay of Bengal during different seasons. Factors that affect primary productivity are irradiance, temperature, stability of the surface waters, nutrients and zooplankton grazing. The differential biological response of eastern Arabian Sea and western Bay of Bengal to monsoonal regimes. A precise estimation on the primary production of the entire EEZ of India on a regional basis and on a seasonal scale would be the only way to achieve any kind of predictive assessment on the fish stock and their sustainable yield. This study mainly envisages the qualitative and quantitative aspects on the magnitude of phytoplankton standing crop and production of organic carbon and their relationship to environmental characteristics during summer monsoon, Inter monsoon and winter monsoon periods in the east and west coasts of the Indian EEZ.This study revealed that the seasonality exerts a great impact on the biological production in the eastern Arabian Sea and western Bay of Bengal. High biological production may be the reason why most of the fish landings are Concentrated in the west coast of India than east coast. The present data on Phytoplankton production rate and the species composition will provide a meaningful ground for evaluations of exploitable renewable resources of the IndianEEZ

A new species of Pseudostomella (Gastrotricha: Macrodasyida: Thaumastodermatidae) from a sandy beach of Kerala, India

During an interstitial faunal survey along the south-west coast of Kerala, India, gastrotrich fauna were found in abundance. Together with species of the genera Xenotrichula, Halichaetonotus and Tetranchyroderma, were present several undescribed thaumastodermatid gastrotrichs belonging to the buccal palp bearing genus Pseudostomella. Adults of the new species are characterized by the following traits: total body length of about 300 μm; cuticular armature made up of medium sized pentancres covering the entire dorsolateral surface; pre-buccal, grasping palps bearing five, large papillae dorsally and 4-6 smaller papillae ventrally; adhesive apparatus made up of six anterior, 22-24 ventrolateral, two dorsolateral and six posterior adhesive tubes; caudal organ pear-shaped; frontal organ spherical. Pseudostomella cheraensis sp. nov. is the fourth taxon of the genus known from India; however, all the previous species reported hitherto from India have tetrancres instead of pentancres.