Letteratura scientifica selezionata sul tema "Modèle intestinal in vitro"
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Articoli di riviste sul tema "Modèle intestinal in vitro":
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Uriot, O., C. Deschamps, M. Brun, M. Pouget, L. Etienne-Mesmin, M. Alric, C. Chaudemanche, Y. Boirie e S. Blanquet-Diot. "Développement et validation d’un modèle colique in vitro de dysbiose du microbiote intestinal humain associé à l’obésité". Nutrition Clinique et Métabolisme 37, n. 2 (maggio 2023): e20. http://dx.doi.org/10.1016/j.nupar.2023.03.032.
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Amamou, A., L. Yaker, C. Bôle-Feysot, G. Savoye e R. Marion-Letellier. "Étude de l’interaction entre des dérivés du tryptophane et le récepteur aryl hydrocarbone dans un modèle in vitro de fibrose intestinale". Nutrition Clinique et Métabolisme 33, n. 1 (marzo 2019): 100. http://dx.doi.org/10.1016/j.nupar.2019.01.412.
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Vergères, Guy, Biljana Bogicevic, Caroline Buri, Sandro Carrara, Magali Chollet, Linda Corbino-Giunta, Lotti Egger et al. "The NutriChip project – translating technology into nutritional knowledge". British Journal of Nutrition 108, n. 5 (11 luglio 2012): 762–68. http://dx.doi.org/10.1017/s0007114512002693.
Abstract (sommario):
Advances in food transformation have dramatically increased the diversity of products on the market and, consequently, exposed consumers to a complex spectrum of bioactive nutrients whose potential risks and benefits have mostly not been confidently demonstrated. Therefore, tools are needed to efficiently screen products for selected physiological properties before they enter the market. NutriChip is an interdisciplinary modular project funded by the Swiss programme Nano-Tera, which groups scientists from several areas of research with the aim of developing analytical strategies that will enable functional screening of foods. The project focuses on postprandial inflammatory stress, which potentially contributes to the development of chronic inflammatory diseases. The first module of the NutriChip project is composed of three in vitro biochemical steps that mimic the digestion process, intestinal absorption, and subsequent modulation of immune cells by the bioavailable nutrients. The second module is a miniaturised form of the first module (gut-on-a-chip) that integrates a microfluidic-based cell co-culture system and super-resolution imaging technologies to provide a physiologically relevant fluid flow environment and allows sensitive real-time analysis of the products screened in vitro. The third module aims at validating the in vitro screening model by assessing the nutritional properties of selected food products in humans. Because of the immunomodulatory properties of milk as well as its amenability to technological transformation, dairy products have been selected as model foods. The NutriChip project reflects the opening of food and nutrition sciences to state-of-the-art technologies, a key step in the translation of transdisciplinary knowledge into nutritional advice.
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Jackson, Tim R., Miniver Oliver, Daniel Appledorn, Tim Dale e Kalpana Barnes. "Abstract 3084: Label-free, real-time live cell assays for 3D organoids embedded in Matrigel®". Cancer Research 82, n. 12_Supplement (15 giugno 2022): 3084. http://dx.doi.org/10.1158/1538-7445.am2022-3084.
Abstract (sommario):
Abstract Organoid technologies are increasingly being used as in-vitro models of human development and disease as they exhibit structural, morphogenetic, and functional properties that recapitulate in vivo pathophysiology. To successfully use these models across a variety of research disciplines and applications, approaches that reduce variability and technology pipelines to image & quantify these complex cell models are required. Here, we demonstrate simple, robust workflows for monitoring and automatically quantifying features, such as morphology, growth and death of organoids using real time live cell analysis. To quantitatively optimize and characterize organoid cultures in-vitro, mouse hepatic, intestinal and pancreatic organoids were embedded in Matrigel® domes (50% or 100%) in 24-well plates and imaged over time in an Incucyte® Live Cell System. Organoid growth, differentiation, and maturation was measured using Incucyte’ s automated Organoid Software Analysis Module, which tracks changes in size and morphology. Integrated metrics enabled objective determination of cell-type specific growth conditions and passaging regimes. To illustrate the utility of the Incucyte® Organoid Analysis Software Module to track organoid growth and death in 96-well plates, intestinal and hepatic organoid fragments were embedded in Matrigel® (50%) for 3 days prior to treatment with protein kinase inhibitor staurosporine (1 µM, STP). Vehicle treated organoids increased in size (10-fold; intestinal or 3-fold; hepatic) over time while marked reduction was observed in the presence of STP. Using label-free size and morphology metrics we could distinguish between cytotoxic and cytostatic mechanisms of action (MoA) of known chemotherapeutic compounds. STP, cisplatin (CIS, DNA synthesis inhibitor) or fluorouracil (5-FU, thymidylate synthetase inhibitor) exhibited concentration dependent inhibition of hepatic organoid growth, yielding IC50 values of 3 nM for STP, 9.7 µM for CIS and 0.78 µM for 5-FU. Whilst attenuation of size was observed across all compounds, increases in eccentricity and darkness indictive of 3D structure disruption and cell death respectively were only observed in CIS and STP-treated organoids (cytotoxic MoA). Differences between the size and morphology readouts illustrated the cytostatic mechanism of 5-FU. Use of this approach was extended to visualize and quantify CFTR function. Following forskolin stimulation, a concentration-dependent increase in intestinal organoid size was observed. In the presence of CFTR inhibitor CFTRinh-172 the maximal response was reduced by >50% (~150% at 10 µM) illustrating that swelling was CFTR-dependent. These data demonstrate the capability to kinetically visualize and quantify distinct organoid morphologies, assess drug-induced cellular changes label-free and illustrates the amenability of this approach across a range of disease areas. Citation Format: Tim R. Jackson, Miniver Oliver, Daniel Appledorn, Tim Dale, Kalpana Barnes. Label-free, real-time live cell assays for 3D organoids embedded in Matrigel® [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3084.
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Losa, Marco, Michael Field, Lauren Collen, Jared Barends, Amit Ringel, Mairead Bresnahan, Jessica Yang et al. "BIOACTIVE INTERLEUKIN-1 DETECTED IN IBD PATIENT INTESTINAL BIOPSIES IS A HALLMARK OF ULCERS AND CORRELATES WITH TRANSCRIPTOMIC ASSESSMENTS, INCLUDING AN ULCER-ASSOCIATED GENE MODULE". Inflammatory Bowel Diseases 30, Supplement_1 (25 gennaio 2024): S00. http://dx.doi.org/10.1093/ibd/izae020.117.
Abstract (sommario):
Abstract BACKGROUND Interleukin (IL)-1 and its post-translationally modified and released forms, IL-1α and IL-1β, have been identified as key molecules to maintain mucosal homeostasis and to drive inflammatory bowel disease (IBD). Deep cellular phenotypic, transcriptional, and in vitro data highlight the importance of IL-1-associated cytokine networks and IL-1 producing cells in severe and anti-TNF non-responsive IBD. Nevertheless, little is known about the presence of bioactive IL-1 proteins within the cell-free gastrointestinal (GI) mucosal environment and associated function in severe IBD and ulceration. METHODS We established a highly sensitive functional assay (~500-fold greater than ELISA) to assess bioactive extracellular IL-1 protein from previously cryopreserved GI biopsies. We performed an ex vivo study in a cohort of Crohn’s disease (CD, n=23), ulcerative colitis (UC, n=18) and non-IBD (n=17) patients, with biopsies from paired inflamed and uninflamed intestinal regions per patient (primarily colon but also ileal). We assessed differential IL-1 bioactivity, including IL-1α vs IL-1β contributions, and performed RNA-seq of the matched cellular compartments of the samples. An ulcer-associated gene signature derived from RNA-seq analysis was evaluated in a single-cell RNA-seq cohort (n=42) of very early onset IBD including monogenic disorders. RESULTS The extent of bioactive intestinal mucosal IL-1α and IL-1β corresponded with disease and ulcer severity in both CD and UC. The most extreme signals were seen in select CD patients with deep ulceration. Bioactive IL-1α was the predominant contributor to total IL-1 signal in most patients although several with ulcers displayed an IL-1β-predominant signal. Matched RNA-seq analysis demonstrated enhanced IL-1 transcripts, correlating with IL-1 bioactivity, and a weighted gene co-expression network analysis (WGCNA) revealed a compelling ulcer-specific module enriched in IL-1 signaling genes and wound repair. We validated our findings in several published (e.g., RISK cohort of ileal CD) as well as unpublished independent adult and pediatric datasets from our group. Deconvolution of the ulcer-specific gene module and projection onto an unpublished single-cell RNA-seq cohort of very early onset IBD patients (including IBD-causative mutations such as IL-10R deficiency characterized by deep ulceration) implicated specific stromal and myeloid populations in ulcer biology. CONCLUSION Mucosal ulceration in IBD is associated with bioactive IL-1α and IL-1β proteins, and transcriptomic evaluation of the same correlates with bioactive signal. An ulcer-associated gene module, validated in other datasets, sheds light on IL-1 biology in intestinal epithelial repair. Results strongly suggest the IL-1 signaling pathway being an attractive and precision-based therapeutic target in subsets of IBD patients.
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Rashid, Md Harun Or, e Feng Lin. "Magnetically Driven Biopsy Capsule Robot with Spring Mechanism". Micromachines 15, n. 2 (18 febbraio 2024): 287. http://dx.doi.org/10.3390/mi15020287.
Abstract (sommario):
In recent years, capsule endoscopes (CEs) have appeared as an advanced technology for the diagnosis of gastrointestinal diseases. However, only capturing the images limits the advanced diagnostic procedures and so on in CE’s applications. Herein, considering other extended functions like tissue sampling, a novel wireless biopsy CE has been presented employing active locomotion. Two permanent magnets (PMs) have been placed into the robots, which control the actuation of the capsule robot (CR) and biopsy mechanism by employing an external electromagnetic actuation (EMA) system. A spring has been attached to the biopsy mechanism to retract the biopsy tool after tissue collection. A camera module has also been attached to the front side of the CR to detect the target point and observe the biopsy process on the lesion. A prototype of CR was fabricated with a diameter of 12 mm and a length of 32 mm. A spring mechanism with a biopsy needle was placed inside the CR and sprang out around 5 mm. An in vitro experiment was conducted, which demonstrated the precise control translation (2 mm/s and 3 mm/s in the x and y directions, respectively) and desired extrusion of the biopsy mechanism (~5 mm) for sampling the tissue. A needle-based biopsy capsule robot (NBBCR) has been designed to perform the desired controlled locomotion and biopsy function by external force. This proposed active locomoted untethered NBBCR can be wirelessly controlled to perform extended function precisely, advancing the intestinal CE technique for clinical applications.
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Dupont, C., M. E. Bougnoux, J. Matéo, P. Saulnier, D. Payen e M. H. Nicolas-Chanoine. "Diagnostic des candidoses profondes par PCR modèle in vitro et modèle animal". La Revue de Médecine Interne 17 (gennaio 1996): 356s. http://dx.doi.org/10.1016/s0248-8663(97)80878-8.
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Dupont, C., M. E. Bougnoux, J. Mateo, P. Saulnier, D. Payen e M. H. Nicolas-Chanoine. "Diagnostic des candidoses profondes par PCR. Modèle in vitro et modèle animal". Médecine et Maladies Infectieuses 27 (novembre 1997): 1005. http://dx.doi.org/10.1016/s0399-077x(97)80272-7.
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Durix, A., L. Alves de Oliveira, S. Komizarczuck-Bony, M. Carcelen e C. Jean-Blain. "Caractéristiques fermentaires d'un modèle d'acidose in vitro (RUSITEC)". Annales de Zootechnie 43, Suppl. 1 (1994): 26s. http://dx.doi.org/10.1051/animres:19940530.
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Breton, J., P. Tirelle, S. Hasanat, A. Pernot, C. L’Huillier, J. C. do Rego, P. Déchelotte, M. Coëffier, L. B. Bindels e D. Ribet. "Altérations du microbiote intestinal dans un modèle murin d’Anorexie mentale". Nutrition Clinique et Métabolisme 34, n. 1 (aprile 2020): 37–38. http://dx.doi.org/10.1016/j.nupar.2020.02.238.
Tesi sul tema "Modèle intestinal in vitro":
1
Ponce, de Leon Rodriguez Maria del Carmen. "Développement d’un modèle in vitro d’inflammation intestinale par l’utilisation de lignées cellulaires humaines en co-culture pour l’étude des interactionsavec les micro-constituants alimentaires". Thesis, Montpellier, 2019. http://www.theses.fr/2019MONTG009/document.
Abstract (sommario):
L’épithélium intestinal, siège de l’absorption des (micro)-nutriments est aussi le premier système de défense de l’organisme. Un déséquilibre dans l’homéostasie peut être à l’origine d’une réaction inflammatoire associée à des défauts de la barrière intestinale et de la fonction immunitaire, ainsi qu’une malabsorption des nutriments, comme rencontré dans les MICI (Maladies Inflammatoires Chroniques de l’Intestin), dans les stratégies de fortification en micronutriments et les pathologies non transmissibles (obésité). Il est donc important de trouver des moyens d’action, via l’alimentation par exemple, pour prévenir ou au minima réduire, les conséquences nutritionnelles et pathologiques de l’inflammation intestinale, et de comprendre les mécanismes impliqués. Parmi les modèles d’études de l’intestin, les modèles in vitro de culture cellulaire sont de plus en plus utilisés et permettent d'évaluer les mécanismes moléculaires d'une manière simple et reproductible et de réduire l'expérimentation animale.Dans ce contexte et dans le but d’étudier l’interaction de composés bioactifs de l’alimentation avec l’intestin en état d’inflammation, le premier objectif de ce travail de thèse a été la mise au point d’un modèle in vitro d’intestin enflammé associant en co-culture deux lignées intestinales humaines : les Caco-2 TC7 (entérocytes) et HT29-MTX (cellules caliciformes) et une lignée immunitaire de macrophages (THP1). Plusieurs marqueurs d’inflammation ont été évalués et nous avons pu montrer que le modèle de tri-culture répondait à un stimulus inflammatoire (LPS/IFNγ), par une augmentation de la production de cytokines pro-inflammatoires (TNF-α, IL6 et IL8) et d’enzymes (INOS et COX2) ainsi que l’expression de leurs gènes. Par ailleurs, une augmentation de la perméabilité épithéliale via une altération des jonctions serrées (TJs) a également pu être mise en évidence ainsi qu’une surproduction de mucus, lesquels sont des caractéristiques reconnus d’inflammation.Le deuxième objectif était d’étudier l’interaction de la β-cryptoxanthine (BCX), caroténoïde des agrumes, lipophile et anti-oxydant, avec le modèle enflammé. Nous avons utilisé pour solubiliser la BCX deux types de micelles (artificielles et physiologiques) et étudié les marqueurs d’inflammation. Bien qu’il semble d’après les résultats préliminaires que les micelles de BCX montrent une tendance à diminuer la production de certaines cytokines (IL6 et IL8), le rôle des constituants des micelles (Tween 40 ou sels biliaires/phospholipides) dans ce phénomène observé et dans la perméabilité épithéliale reste à clarifier par la suiteThe intestinal epithelium, main place of the absorption of (micro)-nutrients is also the first body's defense system. An imbalance in homeostasis can lead to an inflammatory reaction associated with defects in the intestinal barrier and immune function as well as malabsorption of nutrients, as seen in IBD (Inflammatory Bowel Diseases), in micronutrient fortification strategies and noncommunicable diseases (obesity). It is therefore important to find ways of action, for example through diet, to prevent or at least reduce the nutritional and pathological consequences of intestinal inflammation, and to understand the mechanisms involved. Among intestinal models, in vitro cell culture models are increasingly used and allow to evaluate the molecular mechanisms in a simple and reproducible way and to reduce animal experimentation.In this context and in order to study the interaction of dietary bioactive compounds with the intestine in state of inflammation, the first objective of this work was the development of an in vitro model of inflamed intestine combining in co-culture two human intestinal cell lines: Caco-2 TC7 (enterocytes) and HT29-MTX (goblet cells) and an immune cell line of macrophages (THP1). Several inflammation markers were evaluated and we were able to show that the tri-culture model responded to an inflammatory stimulus (LPS / IFNγ), by increasing the production of pro-inflammatory cytokines (TNF-α, IL6 and IL8) and enzymes (INOS and COX2) as well as the expression of their genes. In addition, an increase of epithelial permeability via tight junctions (TJs) alteration has also been demonstrated, as well as overproduction of mucus, which are recognized inflammation characteristics.The second objective was to study the interaction of β-cryptoxanthin (BCX), a lipophilic and antioxidant carotenoid of citrus, with the inflamed model. To solubilize BCX, we used two types of micelles (artificial and physiological) and studied markers of inflammation. Although it appears from the preliminary results that BCX micelles show a tendency to decrease the production of some cytokines (IL6 and IL8), the role of micelle constituents (Tween 40 or bile salts / phospholipids) in the phenomenon observed and in the epithelial permeability remains to be therefore clarified
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Roy, Isabelle. "Contribution à la mise en place d'un modèle "in vitro" prédictif de l'absorption et du métabolisme intestinal". Paris 5, 1994. http://www.theses.fr/1994PA05P159.
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Fleury, Mickaël. "Impact de traitements antibiotiques sur la flore digestive du porcelet : Etude in vivo et développement d'une approche en système de fermentation in vitro". Thesis, Rennes 1, 2015. http://www.theses.fr/2015REN1B002/document.
Abstract (sommario):
Dans le contexte de l'antibiorésistance, l'objet de ce doctorat vise à évaluer l'impact d'antibiotiques sur le microbiote intestinal de porcelets. La colistine et le ceftiofur, pour lesquels les résistances incluent essentiellement et respectivement mutations chromosomiques et gènes plasmidiques, ont été utilisés. La colistine a significativement réduit la population des entérobactéries, mais aucun E. coli résistant n'a été détecté. L'administration de ceftiofur a eu un impact limité sur les populations bactériennes composant l'écosystème digestif mais a conduit à une forte sélection et à la diffusion d'un gène plasmidique codant pour une bêta-lactamase à spectre étendu. Puis, dans le cadre de la réglementation visant à diminuer l'expérimentation animale, un modèle in vitro colique porcin, nommé PigutIVM, a été mis au point afin de simuler l'environnement digestif du porcelet et a permis de confirmer, in vitro, l'effet observé in vivo de la colistine sur le microbiote. Cet outil a ensuite été utilisé pour évaluer l'impact d'un probiotique, Saccharomyces cerevisiae, comme alternative aux antibiotiques. Le modèle PigutIVM devrait se positionner comme un outil de prédiction pertinent dans les domaines d'investigation aussi bien nutritionnels que pharmacologiquesIn the context of antibiotic resistance, the aim of the current PhD work is to assess the impact of antibiotics on intestinal microbiota of piglets. Two antibiotics i.e. colistin and ceftiofur, for which the main resistances include respectively chromosomal mutations and plasmid genes have been used. Colistin significantly reduced the population of Enterobacteriaceae, but there was no selection of resistant E. coli. The administration of ceftiofur had a limited impact on the bacterial populations that make up the digestive ecosystem but it led to strong selection and dissemination of a plasmid gene encoding an extended-spectrum beta-lactamase. Then, in the framework of regulations to reduce animal testing, an in vitro model of colonic pig named PigutIVM was developed in order to simulate the digestive environment of the piglet and then check the effect of colistin on the microbiota simulated in PigutIVM in vitro. Therefore both the approaches i.e. in vivo and in vitro were compared in order to check the effect of colistin on intestinal microbiota of piglets. This tool was then used to evaluate the impact of a probiotic i.e. Saccharomyces cerevisiae, as alternative to antibiotics. Therefore we assume that this PigutIVM model should be positioned as a relevant predictive tool in the fields of nutritional and pharmacological investigations
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Altay, Gizem. "Towards the development of biomimetic in vitro models of intestinal epithelium derived from intestinal organoids". Doctoral thesis, Universitat de Barcelona, 2018. http://hdl.handle.net/10803/664864.
Abstract (sommario):
Intestinal epithelium is highly specialized tissue organized into crypt-villus units relevant for their effective barrier function and nutrient absorption. In the crypt units reside the proliferative intestinal stem cells (ISCs) that divide and differentiate while migrating along the villi to generate the epithelium. The proliferation, migration and differentiation of ISCs is governed by the tightly controlled spatio-chemical gradients of ISC niche factors; bone morphogenic protein (BMP), wingless/Int (Wnt) and epidermal growth factor (EGF) pathway modulators. In vitro models of the intestinal epithelium, for the most part, based on culturing of intestinal stem cells/crypts in 3D cultures forming structures called organoids. These structures faithfully recapture diverse cell populations and their multicellular organization of native intestinal epithelium. However, 3D closed geometry of intestinal organoids prevents access to the apical region of the epithelium, making them unsuitable for conventional functionality assays. Experimental modeling of intestinal epithelial biology and physiology are limited due to the lack in vitro platforms that recapitulate these key aspects of the small intestinal epithelium: its distinct cell populations, 3D architecture and the gradients of ISC niche biochemical factors along the crypt-villus axis.
Here, we describe development of in vitro models of intestinal epithelium obtained from intestinal organoid-derived crypts. First, we present a method that takes the advantage of substrate stiffness to dictate the formation of monolayers with accessible lumen rather than 3D organoids with a closed geometry. The 2D intestinal epithelium model has in vivo-like crypt-villus cellular organization with all major epithelial cell types and show physiologically relevant tissue barrier function. Then, we describe the development of a more complex model of intestinal epithelium by incorporating a 3D villus-like basement membrane substitute fabricated on hydrogels. For that, poly(ethylene glycol) diacrylate (PEGDA) hydrogels are chosen due to their highly tunable chemical, and mechanical properties, porosity and photocrosslinkable nature allowing easy microstructuring. The formation of 3D bullet-like complex shapes was achieved by photolithography-based crosslinking of PEGDA, a simple, cost-effective approach. The bioactive functionalization of otherwise inert PEGDA for cell adhesion, was achieved by copolymerizing it with acrylic acid and a variety of cell adhesion proteins can be covalently anchored to the 3D villus-like hydrogels. We establish the optimal conditions for the growth of intestinal organoid-derived epithelial monolayers and demonstrated that organoid-derived intestinal epithelial cells successfully formed epithelial monolayers on collagen type I functionalized 3D villus-like PEGDA-acrylic acid hydrogels. Finally, we describe methods to create spatiotemporal gradients of biochemical ISC niche factors on 3D villus-like hydrogels and demonstrate that these gradients can be used to compartmentalize the differentiated epithelial cells. The spatio-chemical gradients of ISC niche biochemical factors on PEGDA hydrogels with proper porosity were successfully generated based on the free diffusion of the factors from a source to a sink chamber in a custom-made microfluidic device allocating the hydrogel and visualized with light-sheet fluorescence microscopy. In silico models were developed to simulate the spatio-chemical gradients formed within the hydrogels. The 3D villus-like PEGDA hydrogels were fabricated on porous membranes and successfully adapted to Transwell® inserts that permitted access to both sides of the hydrogel and the generation of spatio-chemical gradients. The gradients generated in this fashion can be used to compartmentalize the differentiated epithelial cells more towards the tips of the villus-like microstructures.
The 3D villus-like platform improves the current models in providing cells with physiologically representative topographical and mechanical cues and biochemical gradients. Due to its utility, this platform might find uncountable applications. It can be used for the understanding of the basic biology of the intestinal epithelium. In addition, it can be used to culture human intestinal stem cells allowing for the screening of novel therapies and disease modeling.El epitelio intestinal es un tejido altamente especializado, organizado en unidades de criptas y vellosidades que son relevantes para sus eficaces funciones de barrera y absorción de nutrientes. En las unidades de criptas residen las células madre intestinales (ISC) proliferativas que se dividen y diferencian mientras migran a lo largo de las vellosidades, las cuales generan el epitelio maduro. En el epitelio maduro, las ISC y las células proliferativas se localizan en las criptas y las células absorbentes y secretoras diferenciadas en las vellosidades. La proliferación, migración y diferenciación de las ISC se rigen por los gradientes químicos espaciales altamente controlados de los factores de nicho de la ISC; Moduladores de la vía de bone morphogenic protein (BMP), wingless/Int (Wnt) y epidermal growth factor (EGF). El modelado experimental de la biología y la fisiología del epitelio intestinal está limitado debido a la falta de plataformas in vitro que recapitulan estos aspectos clave del epitelio del intestino delgado: sus distintas poblaciones celulares, la arquitectura 3D y los gradientes de factores bioquímicos de nicho ISC a lo largo del eje cripta-vellosidad. Aquí, describimos el desarrollo de modelos in vitro de epitelio intestinal obtenidos de criptas derivadas de organoides intestinales. En primer lugar, presentamos un método para obtener monocapas epiteliales intestinales 2D con lumen accesible y función de barrera fisiológica. A continuación, describimos el desarrollo de andamios biomiméticos 3D similares a vellosidades en hidrogeles de diacrilato de polietilenglicol (PEGDA) utilizando un enfoque fotolitográfico simple y rentable. Demostramos que nuestra plataforma de vellosidades sintéticas apoya la formación de monocapas epiteliales de células epiteliales intestinales derivadas de organoides. Finalmente, describimos métodos para crear gradientes espaciotemporales de factores nicho bioquímicos ISC en hidrogeles 3D similares a vellosidades y demostramos que estos gradientes se pueden usar para compartimentar las células epiteliales diferenciadas. La plataforma 3D que recrea las vellosidades intestinalesmejora los modelos actuales al proporcionar a las células las señales topográficas y mecánicas y los gradientes bioquímicos fisiológicamente representativos. Debido a su utilidad, esta plataforma puede encontrar innumerables aplicaciones. Puede ser utilizada para la comprensión de la biología básica del epitelio intestinal. Además, se puede utilizar para cultivar células madre intestinales humanas que permitan la detección de nuevas terapias y el modelado de enfermedades.
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Gérémie, Lauriane. "Development of an in-vitro intestinal model featuring peristaltic motion". Thesis, Sorbonne université, 2019. http://accesdistant.sorbonne-universite.fr/login?url=http://theses-intra.upmc.fr/modules/resources/download/theses/2019SORUS118.pdf.
Abstract (sommario):
Le Gut-on-chip fait partie d'un thème de recherche plus générale, appelez Organ-on-chip qui a pour objectif de développer des modèles in-vitro qui récapitulent des caractéristiques essentielles de l'organe d'intérêt. Dans le cas de l'intestin, les Gut-on-chip plateformes ont été principalement développés pour reconstituer soit l'architecture 3D de l'intestin, soit sa dynamique et plus particulièrement le péristaltisme. Durant ma thèse j'ai développé une nouvelle et polyvalente Gut-on-chip, présentant ces deux aspects du micro-environnement intestinale. Cette Peristalsis-on-chip nous a permis d'étudier l'influence du mouvement péristaltique sur le comportement cellulaire en fonction de la géométrie de la structure. Pour cette étude nous avons ensemencé des cellules Caco2 sur des substrats 2D ou 3D recouvert de laminine et les avons soumis à un étirement cyclique (à 0.2 Hz et 10\%) pendant 2, 5, 8, 16, 24 et 48 heures. Lors de ces expériences nous avons pu observer une réorientation cellulaire perpendiculaire à l'axe d'étirement que nous avons caractérisé en fonction des conditions de recouvrement, de la confluence initiale, du temps d'étirement et de la géométrie de la structure. Il est intéressant de noter que la réponse cellulaire la plus importante a été obtenue par la combinaison de la géométrie 3D et de l'étirement, ce qui illustre bien le besoin de ces deux éléments pour mieux mimer les conditions intestinales in vivoMy PhD work is part of the organ-on-chip field, and more precisely part of the gut-on-chip field. It is in line with the main objective of this field, which is the development of in-vitro models recapitulating as faithfully as possible the intestinal micro-environment. Through my PhD work I first developed a versatile gut-on-chip platform recapitulating the intestinal 3D architecture as well as its dynamic micro-environment. Therefore, this platform allows us to study the influence of the intestinal dynamic, especially the peristalsis, on cellular behavior in function of the 3D architecture of the scaffold. For this study Caco2 cells have been seeded either on a 2D or a 3D scaffold coated with laminin and submitted to a cyclic stretching (at 0.2 Hz and 10%) for 2, 5, 8, 16, 24 and 48 hours. Our main observation was the cellular reorientation induced by the stretching, therefore we characterized the cell behavior in function of the coating condition, the initial confluency, the stretching time and the scaffold geometry. Interestingly, the strongest cellular response was obtained when the 3D geometry and the stretching was combined illustrating the need of these two stimuli to better mimic the intestinal in vivo conditions
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Garcia, Rodriguez Alba. "The applicability of in vitro models of the intestinal barrier for the risk assessment of engineered nanomaterials used as food additives". Doctoral thesis, Universitat Autònoma de Barcelona, 2018. http://hdl.handle.net/10803/669883.
Abstract (sommario):
Los avances en el campo de la nanotecnología han permitido desarrollar una gran diversidad de nanomateriales sintetizados artificialmente (NMs), los cuales presentan nuevas y prometedoras aplicaciones en diversas industrias. Debido a sus exclusivas propiedades, los NMs son utilizados en la comida o envoltorios de comida como mejora de textura, color, sabor, estabilizador, etc.
A pesar de sus propiedades innovadoras, existe un aumento en la preocupación sobre si las nanopartículas (NPs) de dióxido de titanio (TiO2) puedan llegar a producir efectos adversos en la salud humana. La Agencia Internacional para la Investigación del Cáncer (IARC) clasificó el TiO2 como posible carcinógeno humano (grupo 2B) debido a suficientes evidencias científicas indicando que las NPs de TiO2 pueden causar cáncer de pulmón a través de su inhalación. Sin embargo, en el mismo informe no se obtuvo resultados concluyentes respecto a la exposición de dichas NPs por vía oral debido a la falta de ensayos toxicológicos e información. Es por eso que el objetivo de la presente Tesis es estudiar de manera in vitro los efectos, factores y mecanismos biológicos que la exposición a NPs metálicas puedan causar en el epitelio del intestino delgado humano.
Para este propósito, se desarrolló por primera vez en nuestro laboratorio un modelo epitelial in vitro que mimetiza el intestino delgado humano. En nuestro primer estudio, se definieron y caracterizaron condiciones de cultivo del ya descrito modelo, Caco- 2/HT29/Raji-B. Según nuestros resultados en los estudios de integridad y permeabilidad, confirmamos que la mejor ratio de células Caco-2 (enterocitos) y HT29 (células calciformes) era 90:10, respectivamente. Paralelamente se detectó la inducción de células presentadoras de antígenos o también conocidas cómo células M, y se propuso un listado de genes cómo marcadores para bio-monitorizar la correcta diferenciación celular y formación de la barrera intestinal in vitro. Finalmente se testó la funcionalidad de nuestro modelo epitelial in vitro exponiéndolo durante 24 h tanto a NPs de TiO2 como de SiO2. Utilizando microscopía laser confocal se demostró que las NPs de TiO2 podrían conllevar efectos adversos en el epitelio intestinal ya que tienen la capacidad de internalizar en las células, llegando incluso, a entrar en contacto con el núcleo celular.
Debida a la gran diversidad de NMs que actualmente se pueden sintetizar artificialmente, y a que cada uno de ellos puede presentar propiedades distintas y por ende afectar de forma diferente sobre la salud humana, el segundo objetivo de la presente Tesis fue valorar los efectos de tres formas distintas de TiO2 (nano-esferas, nano-óvalos i nano-filamentos) utilizando el modelo intestinal Caco-2/HT29. Nuestros resultados demostraron que las tres formas de TiO2 son capaces de desestabilizar el epitelio intestinal, cruzar la cubierta de mucosa, e internalizar en las células hasta alcanzar al núcleo celular. Teniendo en cuenta las imágenes obtenidas con microscopía láser confocal, se demostró que tanto las nano-esferas cómo los nano-óvalos traspasan la barrera intestinal intracelularmente mientras que los nano-filamentos lo hacen por vía paracelular. Finalmente, utilizando el ensayo del cometa, detectamos que las tres NPs produjeron un leve pero estadísticamente significativo daño genotóxico general pero no daño genotóxico oxidativo.
Por último, el tercer estudio se llevó a cabo en el departamento de Ingeniería Biomédica de la Universidad de Binghamton (Binghamton, NY, USA) con el propósito de una mención internacional de la presente Tesis. Puesto que la absorción de nutrientes es una de las principales funciones del intestino delgado, en este estudio se evaluó la actividad de tres enzimas digestivas (Fosfatasa Alcalina Intestinal, Aminopeptidasa-N y la bomba de sodio/potasio) tras exponer el modelo Caco-2/HT29-MTX a NPs de TiO2 y SiO2. Con el fin de simular estrictamente las condiciones reales del tracto gastrointestinal humano, las NPs fueron digeridas de manera artificial simulando el proceso de digestión humano (boca, estómago, intestino), y co-cultivadas con bacterias comúnmente encontradas en el primer segmento del intestino delgado humano, el duodeno. Concretamente se utilizaron el comensal grampositivo Lactobacillus rhamnosus GG, conocido por su actividad probiótica, y el oportunista gramnegativo Escherichia coli NCTC 9001. En este estudio se observó que la presencia de ambas bacterias en el modelo in vitro Caco-2/HT29-MTX, disminuía los efectos adversos de las NPs sobre la actividad enzimática del epitelio.Nano-technological approaches are allowing the development of deliberately engineered nanomaterials (ENMs), presenting promising new applications for many industrial fields. Especially, ENMs possess unique properties and novel uses in food or food packaging materials such as the enhancement of texture, colour, flavour, nutrient stability and food packaging safety. Despite their innovative properties, there is an increasing concern about the possibility that human exposure to TiO2NPs may lead to significant adverse health effects. The International Agency for Research on Cancer (IARC) classified TiO2 as a human carcinogen group 2B because there was enough evidence that nano-TiO2 may cause lung cancer by inhalation. Although oral exposure was also debated by IARC, the final report was inconclusive due to non-existing standardized procedures for nano- TiO2 risk assessment. Due to the potential adverse effects of this ENMs and the lack of information regarding toxicological aspects over the oral exposure, in this Thesis we have carried out in vitro studies on the biological effects of TiO2NPs. For the aforementioned purpose, we set up and characterized, for the first time in our laboratory, an epithelial in vitro model that closely mimics the human small intestine. Thus, in our first study, we defined the best culture conditions for the alreadydescribed model, Caco-2/HT29/Raji-B. From our integrity and permeability findings, we confirmed that the best Caco-2/HT29 cell ratio is 90:10, respectively, as TEER values, paracellular LY permeability and the mucus shed formed correlated well with other studies. We also were able to detect the induction of M-like cells by TEM. Moreover, in order to monitor the proper barrier formation, we proposed a set of genes related to the cell junctional complexes, brush border enzymes, mucus shed components and M-cell markers. Finally, we tested the goodness of our epithelial in vitro model by exposing it to both TiO2NPs and SiO2NPs for 24 h. Our confocal results evidenced the potential adverse effects of TiO2NPs and SiO2NPs on the intestinal epithelium, as NPs internalization and NPs-cell nucleus interaction were observed. Because of the heightened interest in the identification, validation and standardization of the effects associated to exposures to new ENMs, our second study aimed to assess the effects of three different shapes of TiO2NPs (spheres, rods and wires) on the Caco-2/HT29 barrier. Our results demonstrated that the three types of TiO2NPs have the ability to impair the membrane’s integrity, translocate through the mucus shed and internalize in the cells, reaching the nucleus. Taking into account our confocal images results, we hypothesize that due to their shapes, nano-wires are more likely to cross paracellularly, while nano-spheres and nano-rods used intracellular passage to cross the intestinal epithelium. Despite previous evidence that relate the capability of TiO2NPs to produce ROS, we have not detected oxidatively DNA damage. However, and in accordance with the confocal images showing a great amount of NPscell nucleus events, we detected a slight but significant general DNA damage in the barrier’s cells. Finally, the third study was performed under the framework of an international mention carried out in the Biomedical Engineering Department at the Binghamton University (Binghamton, NY, USA). Nutrient absorption is one of the main and most important functions of the small intestine. Thus, to understand and evaluate whether ENPs can trigger physiological potential pathologies, the activity of the intestinal alkaline phosphatase (IAP), aminopeptidase-N (APN) and Na+/K+ ATPase enzymes were measured after exposing the Caco-2/HT29-MTX barrier to TiO2NPs and SiO2NPs for 4 h. Moreover, and in order to further mimic the physiological conditions of a real digestion, the Caco-2/HT29-MTX barrier was exposed to both NPs previously digested and co-cultured with both Escherichia coli and Lactobacillus rhamnosus, as examples of commensal microbiota.
7
SUNDARAM, TAMIL SELVI. "ESTABLISHING IN VITRO INTESTINAL EPITHELIAL CELL MODELS IN TRANSLATIONAL ANIMAL NUTRITION". Doctoral thesis, Università degli Studi di Milano, 2022. https://hdl.handle.net/2434/944348.
Abstract (sommario):
Immunomodulatory nutrients as the omega-3 polyunsaturated fatty acids (n-3 PUFA)
and citrus pectins (CPn) are reported to beneficially affect the host intestinal immunity.
However so far, the biological pathways modulated by these nutrients in intestinal
inflammation at the level of intestinal epithelial layer (IEL) remains elusive. To bridge this knowledge gap, the aim of our present study was set in the direction to delineate the effects of n-3 PUFA in porcine IPEC-J2 cell line under (LPS) stress conditions underpinning pig nutrition, combining the state-of-the-art cell-based assays and bioinformatic analysis. The second part of our study was directed towards establishing a primary intestinal epithelial cell (IEC) culture from chicken embryos for the assessment of CPn against LPS stress, underpinning poultry nutrition. Utilizing different cell-based assays, we have successfully demonstrated the proliferative effects of n-3 PUFAs as eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) in the IPEC-J2 cells. Besides, n-3 PUFA pre-treatment (DHA:EPA, 1:2, 10 µM, 24 h) was shown to counteract the cellular damage elicited by different stress factors as LPS, hydrogen-peroxide (H2O2) and dextran sulphate sodium (DSS). In addition, through system biology and multi-omics integration (transcriptomics and proteomics), we have demonstrated the cytoprotective properties of n-3 PUFA in IPEC-J2 cells against LPS-induced inflammatory
and metabolic damage. Specifically, in these cells, we have identified that n-3 PUFA regulate the biological process as, (i) Axon guidance for developmental process; (ii) Defensin and interferon-mediated antimicrobial defense response for homeostasis; (iii) Cell junction assembly under stress-related cell proliferation; (iv) Amelioration of TLR/MyD88 and cytokine signaling in innate immune response; (v) Fatty acid storage in lipid droplets for lipid homeostasis; (vi) Recovery of central carbon metabolic process from dysregulation under infection; (vii) Lipolysis of fatty acids stored in lipid droplets for prevention of cell starvation during infection. To the best of the knowledge, this is the first study to comprehensively map the bioactivity of n-3 PUFA in enterocytes using multi-omics approach. The outcome of the present study, will enable us to better understand the role of n-3 PUFA in intestinal barrier for planning nutritional or
therapeutic strategies. Further in the chicken in vitro study, we have preliminarily demonstrated a simplified method of cell isolation and establishment of primary IEC culture from chicken embryos using mechanical tissue disruptions method. We have also shown that the response of chicken cells is in accordance with the reference IPEC-J2 line using a dose-response study with CPn and LPS. This primary IEC culture model can further be utilized as a starting point for setting up poultry in vitro studies on intestinal barrier.
8
Kratz, Jadel Müller. "Implementação e aplicação do modelo in vitro com células Caco-2 para estudo da permeabilidade intestinal de fármacos". Florianópolis, SC, 2011. http://repositorio.ufsc.br/xmlui/handle/123456789/95611.
Abstract (sommario):
Tese (doutorado) - Universidade Federal de Santa Catarina, Centro de Ciências da Saúde, Programa de Pós-Graduação em Farmácia, Florianópolis, 2011Made available in DSpace on 2012-10-26T03:54:33Z (GMT). No. of bitstreams: 1 294811.pdf: 4760452 bytes, checksum: fa236ca094f2d6addc744e0ad2b1cb85 (MD5)
A absorção oral de um fármaco é controlada fundamentalmente por dois fatores: a solubilidade aquosa/dissolução e a permeabilidade intestinal. Portanto, a determinação dessas características ainda nas fases de descoberta e desenvolvimento de fármacos pode prover moléculas com ótimo perfil biofarmacêutico. Nesse sentido, esta tese teve dois objetivos: implementar e validar o modelo de avaliação da permeabilidade in vitro com células Caco-2 no Laboratório de Virologia Aplicada da UFSC, e aplicar esse modelo na determinação da permeabilidade de fármacos e compostos em estudo. Na implementação do modelo Caco-2 foi demonstrada a adequação morfológica das células, através do monitoramento da resistência elétrica transepitelial e da permeabilidade do Lucifer yellow. Através da avaliação da permeabilidade de fármacos marcadores (aciclovir, carbamazepina, hidroclorotiazida, propranolol, vimblastina e verapamil) em experimentos de transporte bi-direcional foi demonstrado que o modelo foi devidamente validado, já que foi estabelecida correlação entre a permeabilidade in vitro e a absorção em humanos. Adicionalmente, uma metodologia por CLAE-UV foi desenvolvida e validada para a determinação concomitante de todos esses fármacos marcadores. Na segunda etapa, avaliou-se o complexo talidomida:hidroxipropil-?-ciclodextrina, desenvolvido e caracterizado no estado sólido por diferentes técnicas, que comprovaram a complexação do fármaco e a redução da sua cristalinidade. Essas alterações culminaram em um leve aumento da solubilidade aparente do fármaco, bem como propiciaram um perfil de dissolução superior, quando comparado ao da talidomida isolada. No entanto, nenhuma alteração na permeabilidade foi detectada. Esses resultados sugerem que esta complexação poderia aumentar a biodisponibilidade oral da talidomida, através do aumento da sua solubilidade nos fluídos gastrointestinais, bem como facilitando a sua dissolução a partir de uma forma farmacêutica sólida. Também foi avaliado o composto anti-herpético galato de pentila, e foi demonstrado que ele apresenta perfis de permeabilidade intestinal e cutânea favoráveis para sua absorção oral e permeação tópica, respectivamente. Assim, após administração oral, sua biodisponibilidade não seria limitada pela permeabilidade intestinal, e no que se refere à administração tópica, ele ficaria restrito às camadas da pele no local de aplicação. Esses dados fornecem informações valiosas para o desenvolvimento de formas farmacêuticas e para a avaliação da atividade antiviral in vivo. Em suma, o modelo com células Caco-2 foi implementado, validado e aplicado satisfatoriamente, o que permitirá a execução de estudos colaborativos, sobretudo com o enfoque do Sistema de Classificação Biofarmacêutica.
9
Langan, Laura. "Fish intestinal cultures for ecotoxicological studies : in vitro and primary culture models". Thesis, University of Plymouth, 2017. http://hdl.handle.net/10026.1/9486.
Abstract (sommario):
Ecotoxicity testing of chemicals for environmental risk assessment is an area where a high number of vertebrates are used across a variety of industrial sectors. The application of the 3Rs in toxicity testing using fish address both the ethical and societal concerns around this issue in addition to the increasing legislative requests for the incorporation of animal alternatives. This thesis aims to highlight the potential of 3D cell culture models to "bridge the gap" between in vitro and in vivo screening procedures for testing of chemicals with the potential to persist or bioaccumulate thereby improving the predictive power of screening procedures. This thesis examines two alternative methods for their potential use as an intestinal based toxicokinetic tool for environmental risk assessment, utilising an in vitro fish cell line replacement tool (RTgutGC). In addition, for the first time a new intestinal primary cell culture based model was developed to address both intestine region specific response (pyloric, anterior, mid and posterior) and size related adaptability to toxins. Paramagnetic oximetry was used to measure oxygen content within 3D structures (spheroids) in order to better understand the microenvironment of these culture models. Using histology, immunohistochemistry, transepithelial electrical resistance (TEER), transmission electron microscopy (TEM), metabolic, fluorescence and gene expression assays, the comparability of this system to native intestinal response was established. Following exposure to carefully chosen environmental contaminants (Benzo[a]pyrene and Copper), the RTgutGC cell line demonstrated comparable responses to existing literature in terms of uptake, metabolism, DNA damage and the presence an equivalent saturable level. Primary enterocytes cultured on transwell inserts remained viable for upto six weeks, with permeability and metabolic activity comparable to native tissue (both in vitro and ex vivo). Taken in combination, these features of enterocytes represent a profile more closely representative of the intestine then the widely used "gut sac" method. With the potential advantages of incorporating complexity at differing levels (connective tissue layer, intestinal bacteria biome), the intestinal models described offer the potential to screen highly persistent toxins which may require prolonged incubation, in addition to the exploration of complex experimental designs which minimise animal usage (uptake, depuration, uptake). As a consequence, the models developed within this thesis significantly enrich the emerging fish based in vitro testing strategies.
10
Deschamps, Charlotte. "Impact du poids corporel et d'une perturbation antibiotique sur le microbiote intestinal du chien : simulation in vitro et stratégies de restauration". Electronic Thesis or Diss., Université Clermont Auvergne (2021-...), 2023. http://www.theses.fr/2023UCFA0055.
Abstract (sommario):
Différentes tailles de chiens sont associées à des variations de la physiologie digestive, principalement liées au colon et à ses microorganismes résidents. Ce microbiote intestinal joue un rôle clé en santé, soutenant les processus nutritionnels, immunologiques et physiologiques. Néanmoins, les maladies ou l'antibiothérapie peuvent altérer l'équilibre microbien et induire un état perturbé appelé dysbiose. Pour restaurer l'eubiose du microbiote, de nouvelles stratégies de restauration ont été développées telles que les pré, pro ou postbiotiques. Cependant, peu d'études ont évalué leurs effets sur le microbiote dans le cadre de l'antibiothérapie. Cette thèse entre l'unité Microbiologie, Environnement digestif et Santé de l'Université Clermont Auvergne et les deux sociétés Lallemand Animal Nutrition et Dômes Pharma, visait à étudier l'impact du poids corporel et des antibiotiques sur le microbiote colique canin, ainsi que le potentiel de stratégies de restauration microbienne, à l'aide de modèles intestinaux in vitro.Cette thèse a commencé par évaluer l'impact de différentes méthodes de stockage des échantillons fécaux (congélation 48 h à -80°C, 48 h à -80°C avec du glycérol ou lyophilisation avec maltodextrine/tréhalose) sur la cinétique de colonisation du microbiote et ses activités métaboliques dans Mucosal Artificial Colon (M-ARCOL). Par rapport aux selles fraîches, l'inoculation avec des selles congelées brutes est apparue comme la meilleure option. Grâce à une revue de la littérature, le modèle a été adapté pour reproduire les paramètres nutritionnels, physicochimiques et microbiens spécifiques des conditions du petit, moyen et grand chien dans un nouveau modèle appelé Canine M-ARCOL (CANIM-ARCOL), validé avec des comparaisons in vitro-in vivo. Ceci a permis de reproduire in vitro l'augmentation de la production des principaux acides gras à chaîne courte (AGCC), et la prolifération des Bacteroidota et Firmicutes observées in vivo. Puis, le modèle a permis de réaliser une étude mécanistique, révélant que les paramètres nutritionnels et physicochimiques façonnent l'activité du microbiote associé à la taille du chien, mais que l'inoculum fécal est nécessaire pour reproduire sa composition. Enfin, notre modèle a été adapté pour reproduire la dysbiose induite par les antibiotiques. Conformément aux données in vivo, l'antibiothérapie a induit la prolifération des Enterobacteriaceae, Streptococcaceae et Lactobacillaceae, tandis que la diversité et la production d'AGCC diminuaient. Des effets similaires mais moindres ont été observés dans le microbiote mucosal. Enfin, nous avons évalué l'effet de Saccharomyces boulardii CNCM I-1079 et de Lactobacillus helveticus HA-122 tyndallisée sur la résistance du microbiote pendant le traitement antibiotique et la résilience après celui-ci. Les deux stratégies ont réduit la prolifération des Enterobacteriaceae pendant l'antibiothérapie et permis au cours des deux premiers jours, une résilience plus rapide de la composition et l'activité du microbiote, dans le lumen et le mucus.Ce travail a fourni des informations pionnières et significatives sur l'impact de la taille du chien et de l'antibiothérapie sur la composition et l'activité du microbiote luminal et mucosal du colon canin, comblant les lacunes dans ces domaines. Ces travaux améliorent la compréhension de la résilience du microbiote en réponse aux perturbations antibiotiques. Dans un futur proche, en accord avec les règles européennes 3R visant à réduire les expérimentations animales, nos approches in vitro pourraient être utilisées pour des études mécanistiques des interactions entre nutriments, additifs alimentaires ou produits vétérinaires et microbiote. De telles expériences pourraient être réalisées en situations saines ou perturbées (obésité, maladies inflammatoires de l'intestin ou entéropathies chroniques), en tenant compte des variabilités interindividuelles pour progresser vers une nutrition et une médecine personnaliséesDifferent dog sizes are associated with variations in digestive physiology, mainly related to the large intestine and its resident microorganisms. This gut microbiota plays a key role in animal health, supporting nutritional, immunological and physiological processes. Nevertheless, diseases or antibiotherapy can disturb microbial equilibrium and induce a perturbated state called dysbiosis. To restore microbiota eubiosis, new restorations strategies have been developed such as pre-, pro- or postbiotics. However, very few studies have evaluated their effects on gut microbiota in the context of antibiotherapy. This joint PhD between the Microbiology, Digestive Environment and Health unit from Université Clermont Auvergne and the two compagnies Lallemand Animal Nutrition and Dômes Pharma, aimed to investigate the impact of body weight and antibiotic disturbance on canine colonic microbiota, as well as the potential of microbial restoration strategies, using in vitro gut models.This thesis started by evaluating the impact of different methods for faecal sample storage (48-h freezing -80°C, 48-h -80°C with glycerol or lyophilization with maltodextrin/trehalose) on the kinetics of microbiota colonization and metabolic activities in the Mucosal Artificial Colon (M-ARCOL). Compared to fresh stools, inoculating with raw frozen stool without cryoprotectant was the best option among those tested. Second, thanks to a large literature review, the M-ARCOL model was adapted to reproduce the main nutritional, physicochemical and microbial parameters specific from small, medium and large size conditions in a new model called Canine M-ARCOL (CANIM-ARCOL), further validated through in vitro-in vivo comparisons. This adaptation allowed to reproduce in vitro the increase in Bacteroidota and Firmicutes abundances and higher main short-chain fatty acid (SCFA) concentrations observed in vivo. Then, we used the CANIM-ARCOL to perform a mechanistic study, which revealed that nutritional and physicochemical parameters are enough to shape microbiota activity according to dog size, but faecal inoculum was necessary to reproduce size-related microbiota composition. The next step was to adapt the CANIM-ARCOL to diseased situation, focusing on antibiotic-induced dysbiosis. In accordance with in vivo data, antibiotherapy induced an increase in Enterobacteriaceae, Streptococcaceae and Lactobacillaceae relative abundances while alpha-diversity and SCFA production decreased. Similar but lower effects were observed in mucus-associated microbiota. Lastly, we evaluated the effect of the live probiotic yeast Saccharomyces boulardii CNCM I-1079 and the heat-inactivated bacteria Lactobacillus helveticus HA-122 on microbiota resistance during antibiotic treatment and resilience afterwards. Of interest, both microbial strategies decreased the Enterobacteriaceae bloom during antibiotherapy and allowed, in the first two days, a quicker recovery of microbiota composition and activity, in both the luminal and mucosal compartments.This PhD work provided pioneering and significant insights into the impact of dog size and antibiotherapy on canine colonic luminal and mucus-associated microbiota composition and activity, filling gaps in knowledge in these fields. This work also contributed to a better understanding of microbiota resilience in response to antibiotic disturbance. In a near future, in accordance with the European 3R's rules aiming to reduce at a maximum animal experiments, our in vitro approaches could be used for mechanistic studies on the interactions between nutrients, feed additives or veterinary products and canine colonic microbiota. Such experiments could be performed under healthy but also disturbed gut microbial situations (including obesity, inflammatory bowel diseases or chronic enteropathies), always considering interindividual variabilities to move towards personalized nutrition and medicine
Libri sul tema "Modèle intestinal in vitro":
1
Mahmood, Belal A. I. Studies on the effects of cholera toxin and fish oil on intestinal secretion in vitro. Manchester: University of Manchester, 1994.
2
Nicklin, Paul Leslie. Amino acid, peptide and drug transport across monolayers of human intestinal (CAC0-2) cells in vitro. Birmingham: Aston University. Department of Pharmaceutical Sciences., 1993.
3
Essai n° 431 : Corrosion Cutanée In Vitro : Essai sur Modèle de Peau Humaine. OECD, 2014. http://dx.doi.org/10.1787/9789264224216-fr.
4
Essai n° 431 : Corrosion cutanée in vitro : Essai sur modèle de peau humaine. OECD, 2019. http://dx.doi.org/10.1787/9789264264625-fr.
5
Essai n° 431: Corrosion cutanée in vitro :Essai sur modèle de peau humaine. OECD, 2004. http://dx.doi.org/10.1787/9789264071155-fr.
6
Essai n° 431 : Corrosion cutanée in vitro : Essai sur modèle de peau humaine. OECD, 2015. http://dx.doi.org/10.1787/9789264242784-fr.
7
Essai n° 431 : Corrosion cutanée in vitro : Essai sur modèle de peau humaine. OECD, 2013. http://dx.doi.org/10.1787/9789264203839-fr.
8
H.R. Hemati Matin, F. Shariatmadari*, M.A. Karimi Torshizi e L.I. Chiba. Effect of dietary fibre sources on in vitro mineral binding capacity and growth performance, mineral digestibility, tibia and intestinal characteristics in broiler chickens. Verlag Eugen Ulmer, 2018. http://dx.doi.org/10.1399/eps.2018.250.
9
Sohn, Woon-Mok, e Jong-Yil Chai. Anisakiosis (Anisakidosis). Oxford University Press, 2011. http://dx.doi.org/10.1093/med/9780198570028.003.0070.
Abstract (sommario):
The term ‘anisakiosis (anisakidosis)’ or ‘anisakiasis’ collectively defines human infections caused by larval anisakids belonging to the nematode family Anisakidae or Raphidascarididae. Anisakis simplex, Anisakis physeteris, and Pseudoteranova decipiens are the three major species causing human anisakiosis. Various kinds of marine fish and cephalopods serve as the second intermediate hosts and the infection source. Ingestion of viable anisakid larvae in the fillet or viscera of these hosts is the primary cause of infection. The parasite does not develop further in humans as they are an accidental host. Clinical anisakiosis develops after the penetration of anisakid larvae into the mucosal wall of the alimentary tract, most frequently the stomach and the small intestine. The affected sites undergo erosion, ulceration, swelling, inflammation, and granuloma formation around the worm. The patients may suffer from acute abdominal pain, indigestion, nausea, vomiting, and in some instances, allergic hypersensitive reactions. Symptoms in gastric anisakiosis often resemble those seen in peptic ulcer or gastric cancer, and symptoms in intestinal anisakiosis resemble those of appendicitis or peritonitis. Treatments include removal of larval worms using a gastroendoscopic clipper or surgical resection of the mucosal tissue surrounding the worm. No confirmed effective anthelmintic drug has been introduced, though albendazole and ivermectin have been tried in vivo and in vitro. Prevention of human anisakiosis can be achieved by careful examination of fish fillet followed by removal of the worms in the restaurant or household kitchen. Immediate freezing of fish and cephalopods just after catching them on fishing boats was reported helpful for prevention of anisakiosis. It is noteworthy that anisakiosis is often associated with strong allergic and hypersensitivity reactions, with symptoms ranging from isolated angioedema to urticaria and life threatening anaphylactic shock.
Capitoli di libri sul tema "Modèle intestinal in vitro":
1
Sambruy, Yula, S. Ferruzza, G. Ranaldi e I. De Angelis. "Intestinal Cell Culture Models". In Cell Culture Methods for In Vitro Toxicology, 97–113. Dordrecht: Springer Netherlands, 2001. http://dx.doi.org/10.1007/978-94-017-0996-5_7.
2
Vieira, Adriana, Ana Gramacho, Dora Rolo, Nádia Vital, Maria João Silva e Henriqueta Louro. "Cellular and Molecular Mechanisms of Toxicity of Ingested Titanium Dioxide Nanomaterials". In Advances in Experimental Medicine and Biology, 225–57. Cham: Springer International Publishing, 2022. http://dx.doi.org/10.1007/978-3-030-88071-2_10.
Abstract (sommario):
AbstractAn exponential increase in products containing titanium dioxide nanomaterials (TiO2), in agriculture, food and feed industry, lead to increased oral exposure to these nanomaterials (NMs). Thus, the gastrointestinal tract (GIT) emerges as a possible route of exposure that may drive systemic exposure, if the intestinal barrier is surpassed. NMs have been suggested to produce adverse outcomes, such as genotoxic effects, that are associated with increased risk of cancer, leading to a concern for public health. However, to date, the differences in the physicochemical characteristics of the NMs studied and other variables in the test systems have generated contradictory results in the literature. Processes like human digestion may change the NMs characteristics, inducing unexpected toxic effects in the intestine. Using TiO2 as case-study, this chapter provides a review of the works addressing the interactions of NMs with biological systems in the context of intestinal tract and digestion processes, at cellular and molecular level. The knowledge gaps identified suggest that the incorporation of a simulated digestion process for in vitro studies has the potential to improve the model for elucidating key events elicited by these NMs, advancing the nanosafety studies towards the development of an adverse outcome pathway for intestinal effects.
3
Almeida, Hugo, Amélia C. F. Vieira, João Teixeira, Maria João Gomes, Pedro Barrocas, Teófilo Vasconcelos e Bruno Sarmento. "Cell-Based Intestinal In Vitro Models for Drug Absorption Screening". In Drug Discovery and Evaluation: Safety and Pharmacokinetic Assays, 1–22. Cham: Springer International Publishing, 2022. http://dx.doi.org/10.1007/978-3-030-73317-9_94-1.
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Morita, Atsuya, Mizuho Nakayama, Hiroko Oshima e Masanobu Oshima. "In Vitro and In Vivo Models for Metastatic Intestinal Tumors Using Genotype-Defined Organoids". In Methods in Molecular Biology, 19–30. New York, NY: Springer US, 2023. http://dx.doi.org/10.1007/978-1-0716-3331-1_2.
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Wang, Tingting, Xianglong Li e Qinghua Liao. "Stool and Intestinal Microbiota Examination". In In Vitro Diagnostic Industry in China, 341–59. Singapore: Springer Nature Singapore, 2023. http://dx.doi.org/10.1007/978-981-99-3110-1_20.
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Lallés, J. P., e I. P. Oswald. "Chapter 8: Techniques for investigating gut function in vivo, ex vivo and in vitro in monogastric farm animals". In Intestinal health, 191–218. The Netherlands: Wageningen Academic Publishers, 2015. http://dx.doi.org/10.3920/978-90-8686-792-9_8.
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Jardé, Thierry, Genevieve Kerr, Reyhan Akhtar e Helen E. Abud. "Modelling Intestinal Carcinogenesis Using In Vitro Organoid Cultures". In Methods in Molecular Biology, 41–52. New York, NY: Springer New York, 2018. http://dx.doi.org/10.1007/978-1-4939-7568-6_4.
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Chan, Wing Hei, Diana Micati, Rebekah M. Engel, Genevieve Kerr, Reyhan Akhtar, Thierry Jardé e Helen E. Abud. "Modeling Intestinal Carcinogenesis Using In Vitro Organoid Cultures". In Methods in Molecular Biology, 55–69. New York, NY: Springer US, 2023. http://dx.doi.org/10.1007/978-1-0716-3331-1_5.
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Chen, Yun, Chuan Li, Ya-Hui Tsai e Sheng-Hong Tseng. "Intestinal Crypt Organoid: Isolation of Intestinal Stem Cells, In Vitro Culture, and Optical Observation". In Methods in Molecular Biology, 215–28. New York, NY: Springer New York, 2017. http://dx.doi.org/10.1007/7651_2017_21.
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Smith, Philip L. "Methods for Evaluating Intestinal Permeability and Metabolism in Vitro". In Pharmaceutical Biotechnology, 13–34. Boston, MA: Springer US, 1996. http://dx.doi.org/10.1007/978-1-4899-1863-5_2.
Atti di convegni sul tema "Modèle intestinal in vitro":
1
Stan, Miruna, e Alina Strugari. "Nanoparticle Intestinal Transport Characterization Using In Vitro Co-Culture Models". In 1st International Online Conference on Nanomaterials. Basel, Switzerland: MDPI, 2018. http://dx.doi.org/10.3390/iocn_2018-1-05480.
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Elisa Ramos Magalhães, Ana, Juliana Alves Macedo e Mônica Cristina Lopes do Carmo. "AVALIAÇÃO DO EXTRATO DE FOLHAS DE PASSIFLORA EDULIS EM MODELO IN VITRO DE INFLAMAÇÃO INTESTINAL". In XXV Congresso de Iniciação Cientifica da Unicamp. Campinas - SP, Brazil: Galoa, 2017. http://dx.doi.org/10.19146/pibic-2017-77869.
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Lukyanov, Alexandr D., Danila Yu Donskoy, Miroslav A. Vernezi, Maria S. Mazanko e Svetlana G. Studennikova. "EXPERIENCE IN DEVELOPING MODELS OF ARTIFICIAL GASTROINTESTINAL TRACTS OF ANIMALS". In XXVII savetovanje o biotehnologiji. University of Kragujevac, Faculty of Agronomy, 2022. http://dx.doi.org/10.46793/sbt27.193l.
Abstract (sommario):
For „in-vitro“ experimental study of the effect of probiotics on the development of microbiota of poultry and domestic animals, a line of „SHIME“ – like models of artificial gastrointestinal tracts in the form of complexes of miniature bioreactors with microcontroller control and monitoring systems was created. Experience in operating this equipment in an experimental study of the effect of prebiotics on the development of intestinal microbiota has allowed to establish a number of features and limitations, which are brought in this paper. The peculiarities of setting up peristaltic pumps, magnetic mixers and the choice of connecting tube configuration are considered.
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Williams, Courtney M., Jessica L. Harper, Hui Huang, Pat Boland, Qing Zhang, George D. Yancopoulos, Zhe Li e O. Gavin Thurston. "Abstract 2644: In vivo and in vitro stem cell models suggest a role for LGR5 in regulating intestinal epithelial lineage specification." In Proceedings: AACR 104th Annual Meeting 2013; Apr 6-10, 2013; Washington, DC. American Association for Cancer Research, 2013. http://dx.doi.org/10.1158/1538-7445.am2013-2644.
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Hoppensteadt, D., A. Kumar, J. Fareed e J. Mardigian. "STUDIES ON THE ANTICOAGULANT AND ANTITHROMBOTIC ACTIONS OF DERMATANS AND THEIR SULFATED DERIVATIVES". In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1643241.
Abstract (sommario):
Non-antithrombin III mediated effects such as interaction with heparin cofactor II, modulation of endothelium and polymorphonuclear leukocytes contribute to the overall antithrombotic effects of glycosaminoglycans. In order to study the role of these dermatans, we investigated their in vitro anticoagulant effects using the clot based (PT, APTT, TT, and Heptest), antiprotease (anti IIa and anti Xa) and Thromboplastin C activated fibrinopeptide A generation test. The in vivo antithrombotic actions were investigated, against activated and non activated prothrombin complex concentrates, and in combination with Russells viper venom in jugular and femoral vein stasis thrombosis models (rabbit). The dermatans studied consisted of a standard dermatan of porcine intestinal origin and four sulfated dermatans with varying degrees of sulfation. All of the dermatans studied showed weak anticoagulant effects on the routinely performed clot based assays. Marked variability was seen on the protease inhibition (anti Xa and anti IIa) assays. In the in vivo studies all dermatans studied showed varying degrees of antithrombotic actions against various thrombogenic agents in a modified stasis thrombosis model. Sulfation appeared to produce stronger anticoagulant effects as determined by in vitro assays, whereas the intravenous antithrombotic actions of native dermatan were stronger than sulfated derivatives. This data suggests that dermatans produce their antithrombotic actions via non-antithrombin III mediated pathways. Furthermore, in vitro testing methods are of limited value in the evaluation of the biologic actions of dermatans and their derivatives.
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Boyd, Abigail, Joey Talbert e Nuria Acevedo. "Addition of cholesterol esterase substantially enhances phytosterol ester bioaccessibility in emulsions with different droplet sizes using a standardized in vitro digestion model". In 2022 AOCS Annual Meeting & Expo. American Oil Chemists' Society (AOCS), 2022. http://dx.doi.org/10.21748/oplq4898.
Abstract (sommario):
In vitro digestion models are an important tool for the evaluation of foods and supplements, especially foods containing nutraceuticals and bioactive compounds. The INFOGEST network of researchers has published multiple iterations of a static in vitro digestion protocol for the study of food digestion. However, the protocol currently lacks cholesterol esterase (CE) activity during the intestinal phase, and few studies have attempted to evaluate its incorporation into the model. In the present study, the effect of additional CE on phytosterol ester (PSE) bioaccessibility in oil-in-water emulsions with different droplet sizes was evaluated. A range of droplet sizes was obtained by utilizing different homogenization techniques to produce large (9.9 um), medium (3.4 um), and fine (1.3 um) emulsions. Complete lipid digestion was observed for all emulsions. In agreement with previous studies on lipophilic bioactive compounds, emulsion droplet size and PSE bioaccessibility were inversely related to droplet size as smaller droplets led to increased bioaccessibility (12.7, 15.7, and 19.2% for large, medium, and fine emulsions, respectively). This relationship was significantly more pronounced following the addition of CE, with bioaccessibility of large (19.6%), medium (34.2%), and fine (40.8%) emulsions increasing 1.5, 2.2, and 2.1-fold, respectively. Additionally, Beta-sitosterol was significantly greater in bioaccessibility than stigmasterol in medium and fine emulsions with added cholesterol esterase. These findings provide compelling evidence for the inclusion of CE into the standardized INFOGEST protocol and have implications for the in vitro study of esterified micronutrients beyond PSEs. With an improved model, researchers can collect more accurate information about lipid bioaccessibility within different food matrices.
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Akpinar, Ayse Nur, Selvi Secil Sahin, Büşra Moran Bozer, Aziz Tekin e Cansu Ekin Gumus-Bonacına. "Effect of Food Emulsions on the Cytotoxicity of 3-chloropropane-1,2-diol Esters". In 2022 AOCS Annual Meeting & Expo. American Oil Chemists' Society (AOCS), 2022. http://dx.doi.org/10.21748/emiq3563.
Abstract (sommario):
3-Chloropropane-1,2-diol (3-MCPD) esters are food processing contaminants, and are known to be potentially carcinogenic. The main purpose of this study was to determine whether the emulsion technique increases the absorption of this contaminant, and to develop strategies to reduce their absorption. 3-MCPD ester contents in edible oils (hazelnut, walnut, sunflower, soybean, corn, and olive oil) were determined. Refined olive oil contained the highest 3-MCPD ester level (1.7749 ± 0.1262 mg/kg). Then, emulsions (fine, medium, coarse) were prepared with 0.15-2% emulsifier (whey protein isolate) and 10% oil using microfludizier at 3-10 kpsi pressure. The free fatty acid release was investigated using an in vitro digestion model, and the bioaccessibility was calculated. Methylthiazole tetrazolium (MTT) cell viability method was used to perform toxicity tests. The zeta potential and droplet size of the Samples were measured after each digestion phase. The emulsion with the smallest particle size (389 nm) resulted in the highest release of free fatty acid (85.26%) during small intestinal phase. Fine emulsions also resulted in the highest 3-MCPD ester bioaccesibility (95%). During in vitro digestion, droplet size increased at stomach phase for all emulsion types (Figure 1). When an indigestible oil, such as lemon oil, was added to the oil phase of the fine emulsion (up to 30%), the release of free fatty acid decreased by up to 30% as expected and bioaccessibility decreased by up to 45%. The micelle phase of the fine and medium emulsion had a toxic effect on the fibroblast cell line (Figure 2). When the particle size increased and lemon oil was added to the oil phase of the emulsion, the percent viability increased. Briefly, there are two ways to reduce the absorption of 3-MCPD esters in food emulsions: producing emulsions with larger particle size, or adding indigestible oil to the oil phase.
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Hippenstiel, S., B. Brell, C. Zu Dohna, M. Dorenberg, AC Hocke, H. Martens, N. Suttorp e B. Temmesfeld-Wollbruck. "Adrenomedullin Reduces Sepsis-Related Intestinal Epithelial PermeabilityIn VivoandIn Vitro." In American Thoracic Society 2009 International Conference, May 15-20, 2009 • San Diego, California. American Thoracic Society, 2009. http://dx.doi.org/10.1164/ajrccm-conference.2009.179.1_meetingabstracts.a1162.
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Alno, N., F. Clipet, P. Pellen-Mussi, G. Cathelineau e G. De Mello. "Mise au point d’un modèle de culture tridimensionnelle pour l’évaluation de substituts osseux in vitro". In 56ème Congrès de la SFMBCB. Les Ulis, France: EDP Sciences, 2011. http://dx.doi.org/10.1051/sfmbcb/20115602007.
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Pferschy-Wenzig, EM, A. Roßmann, K. Koskinen, K. Ardjomand-Woelkart, G. Meng, E. Koch, C. Moissl-Eichinger e R. Bauer. "Interactions between hawthorn extract WS® 1442 and human intestinal microbiota in vitro". In GA 2017 – Book of Abstracts. Georg Thieme Verlag KG, 2017. http://dx.doi.org/10.1055/s-0037-1608400.
Rapporti di organizzazioni sul tema "Modèle intestinal in vitro":
1
Shpigel, Nahum, Raul Barletta, Ilan Rosenshine e Marcelo Chaffer. Identification and characterization of Mycobacterium paratuberculosis virulence genes expressed in vivo by negative selection. United States Department of Agriculture, gennaio 2004. http://dx.doi.org/10.32747/2004.7696510.bard.
Abstract (sommario):
Mycobacterium avium subsp. paratuberculosis (MAP) is the etiological agent of a severe inflammatory bowel disease (IBD) in ruminants, known as Johne’s disease or paratuberculosis. Johne’s disease is considered to be one of the most serious diseases affecting dairy cattle both in Israel and worldwide. Heavy economic losses are incurred by dairy farmers due to the severe effect of subclinical infection on milk production, fertility, lower disease resistance and early culling. Its influence in the United States alone is staggering, causing an estimated loss of $1.5 billion to the agriculture industry every year. Isolation of MAP from intestinal tissue and blood of Crohn's patients has lead to concern that it plays a potential pathogenic role in promoting human IDB including Crohn’s disease. There is great concern following the identification of the organism in animal products and shedding of the organism to the environment by subclinically infected animals. Little is known about the molecular basis for MAP virulence. The goal of the original proposed research was to identify MAP genes that are required for the critical stage of initial infection and colonization of ruminants’ intestine by MAP. We proposed to develop and use signature tag mutagenesis (STM) screen to find MAP genes that are specifically required for survival in ruminants upon experimental infection. This research projected was approved as one-year feasibility study to prove the ability of the research team to establish the animal model for mutant screening and alternative in-vitro cell systems. In Israel, neonatal goat kids were repeatedly inoculated with either one of the following organisms; MAP K-10 strain and three transposon mutants of K-10 which were produced and screened by the US PI. Six months after the commencement of inoculation we have necropsied the goats and taken multiple tissue samples from the jejunum, ileum and mesenteric lymph nodes. Both PCR and histopathology analysis indicated on efficient MAP colonization of all the inoculated animals. We have established several systems in the Israeli PI’s laboratory; these include using IS900 PCR for the identification of MAP and using HSP65-based PCR for the differentiation between MAV and MAP. We used Southern blot analysis for the differentiation among transposon mutants of K-10. In addition the Israeli PI has set up a panel of in-vitro screening systems for MAP mutants. These include assays to test adhesion, phagocytosis and survival of MAP to/within macrophages, assays that determine the rate of MAPinduced apoptosis of macrophages and MAP-induced NO production by macrophages, and assays testing the interference with T cell ã Interferon production and T cell proliferation by MAP infected macrophages (macrophage studies were done in BoMac and RAW cell lines, mouse peritoneal macrophages and bovine peripheral blood monocytes derived macrophages, respectively). All partners involved in this project feel that we are currently on track with this novel, highly challenging and ambitious research project. We have managed to establish the above described research systems that will clearly enable us to achieve the original proposed scientific objectives. We have proven ourselves as excellent collaborative groups with very high levels of complementary expertise. The Israeli groups were very fortunate to work with the US group and in a very short time period to master numerous techniques in the field of Mycobacterium research. The Israeli group has proven its ability to run this complicated animal model. This research, if continued, may elucidate new and basic aspects related to the pathogenesis MAP. In addition the work may identify new targets for vaccine and drug development. Considering the possibility that MAP might be a cause of human Crohn’s disease, better understanding of virulence mechanisms of this organism might also be of public health interest as well.
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Harpaz, Sheenan, Steven G. Hughes e Pinhas Lindner. Optimization of Diet for Post Larvel/Juvenile Sea Bass and Hybrid Stripped Bass Based on Enzymatic Profiles of their Digestive Tracts. United States Department of Agriculture, dicembre 1995. http://dx.doi.org/10.32747/1995.7604924.bard.
Abstract (sommario):
The overall goal of this research work was to identify the main proteolytic activities which take place in the digestive tracts of young bass fish, and use the knowledge acquired in order to improve feed protein utilization in juvenile fish based on their digestive capacity. The results of the work clearly showed that the young fish possess the entire profile of proteolytic enzymes which is found in adult fish. Yet, in the young fish the level of activity is substantially lower per gram tissue (or gram protein) as compared with the activity found in the digestive tracts of the same fish at an older (larger) age. In addition it was found that the main proteolytic enzyme in these fish is chymotrypsin which accounts for almost 80% of the proteolytic activity. An effort aimed at enhancing this activity has lead to the interesting finding that alcohol substantially enhances the proteolytic activity of fish intestines. Fish intestinal homogenates were used in order to evaluate the suitability of various feeds for the fish. Potential feed proteins were subjected to the proteolytic activity of the fish enzymes in vitro, in a manner simulating the natural process. The proteolytic activity was monitored by the valuation of the products, i.e. amino acid released. This method has proven to be a powerful tool which enables us to predict with a very high degree of accuracy the potential of a feed to promote growth. Selection of feed based on the proteolytic capacity of the fish degestive tracts can now be implemented in feed formulation, as anticipated in the original research proposal.