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The principal aim of this work was to evaluate the impact of the inhibition of the activation of Ras, using a treatment with S-Farnesylthiosalycilic acid (FTS), on the liver regeneration in rats subjected to a partial hepatectomy (PH).
The firs step was the elaboration of the injection scheme of FTS that blocked the activation of the protein Ras after PH. The scheme that was used finally consisted of three injections of FTS (50mg/kg) 1 hour, 8 hours and 16 hours after PH. The rats were sacrificed 12 or 24 hours after PH. The impact of the FTS treatment, on the liver regeneration, was determined by the expression of PCNA, a cell proliferation marker, as well as the incorporation of BrdU into DNA of hepatocytes during S-phase of the cell cycle and its quantification by flow cytometry analysis. The results confirmed that these injection scheme of FTS provokes a significant reduction of the hepatic regeneration on rats subject to PH.
The next step was to verify, by direct and indirect ways, whether the effect of FTP was due to a functional inhibition of the protein Ras. The direct way was based on the determination, by western-blotting, of the localization of Ras, in the cytosol and/or at the plasma membrane in rats, treated with FTS or DMSO, sacrificed 12 and 24 hours after PH. Unlike DMSO-treated rats, where the Ras protein was only localized to the plasma membrane, the latter was found in both the cytosol and the membranefractions in FTS-treated animals resulting in an inactivation af Ras. The indirect way, determined by western-blotting, was based on the expression, of the phosphorylated forms of three proteins : the protein Rad and the protein Erk1/2 (or P44/p42), by western-blots in rats treated with FTS or DMSO after PH. The protein Raf is a direct target of inactivated Ras (RAS-GTP) which recruits it to the plasma membrane where Raf is phsopharylated and activated by unknown mechanisms. The results showed that the phosphorylated form of Raf was only expressed in the membrane fractions of FTS-treated rats after PH whereas its expression occurred normally in DMSO-treated animals. The proteins Erk1/2 are localized downstream of Raf which controls, in part, their activation. In parallel to what was observed with p-Ras, the expression of the phosphorylated forms of Erk ½ was only reduced in FTS-treated rats but not DMSO-treated animals after PH, confirming the functional inhibition of Ras.
Finally, the last step was to verify that FTS acts primarily on the mature Ras protein and does not affect the expression of its messenger RNA. Amplification of mRNA expression by real time PCR showed a similar increase in Ras mRNA after PH regardless whether the animals were treated with FTS or DMSO, suggesting that FTS does indeed have no impact on the expression of Ras mRNA.
Taken together, these observations confirm that the functional inhibition of Ras, by FTS, results in a significant reduction in liver regeneration which is cumulated with an inhibition of membrane recruitment of Raf and downstream activation of Erk1/2 proteins.
In conclusion, the protein Ras seems to play an important role in liver regeneration possibly implicating, at least in part, the Ras/Raf/MER1.2/Erk1.2 pathway L’objectif principal de ce mémoire est d’évaluer l’impact de l’inhibition de l’activation de Ras sur la génération hépatique chez des rats soumis à une hépatectomie partielle (HP) de 70%.
La première étape a été de mettre au point le schéma d’injection d’un inhibiteur compétitif de Ras, l’acide farnésylthiosalicylique (FTS), permettant d’inhiber l’activation de la protéine Ras. Le schéma utilisé a été de trois injections de 50mg/kg de FTS et réparties comme suit : la première injection une heure après HP, la seconde 8 heure après HP et la troisième 16 heures après HP. Le sacrifice ayant lieu 12 ou 24 heures après HP.
Par mesure de l’expression du PCNA, protéine exprimée durant le cycle cellulaire et donc marqueur de prolifération, et la mesure de la synthèse ADN par immunohistochimie et cytométrie de flux, nous avons confirme que le schéma d’injection utilisé provoque effectivement, une diminution significative de la régénération hépatique en réponse à une hépatectomie partielle.
L’étape suivante a été de vérifier, de façon directe et indirecte, que l’impact du FTS sur la régénération hépatique était bien lié à une inhibition fonctionnelle de Ras. L’évaluation, de manière directe, de l’absence d’activation de Ras, s’est basée sur la visualisation de la localisation, membranaires et cytosoliques, des différents échantillons de foie de rats traités au DMSO (solvant) et au FTS (substance active) après HP, ont été préparées et analysées par Western-blot. Les résultats ont montré que le traitement au FTS, des rats ayant subi une HP, est responsable d’une délocalisation incomplète de la protéine Ras, celle-ci étant exprimée tant au niveau des fractions membranaire que cytosoliques. Cette localisation cytosolique étant d’une inhibition de l’activation de Ras.
L’étape suivante a été d’évaluer, de façon indirecte, l’inhibition fonctionnelle de Ras. L’expression de la forme phosphorylée de la protéine Raf, cible directe de la protéine Ras activée (Ras-GTP), a été évaluée, par western-blot, à partir de fractions membranaires d’hépatocytes de rats traités au DMSO et au FTS, sacrifiés 24 heures après HP. Les autres protéines, dont l’expression des formes phosphorylées ont été évaluées, par western-blot, et quantifiées, après densitométrie, à partir d’homogénats de foies de rats traités au DMSO et au FTS, sacrifiés 12 et 24 heures après HP, sont les protéines Erk1 (p44) et Erk2 (p42). Elles sont situées en aval de la protéine Raf, qui module leur activation par l’intermédiaire des protéines MERK1 et MERK2.
Les résultats obtenus montrent l’absence d’expression de la forme phosphorylée de la protéine Raf, au niveau membranaire, et la diminution de l’’expression des formes phosphorylées des protéines p42/p44, chez les rats traités au FTS après HP. Ces résultats sont donc en accord avec ceux obtenus plus haut et ont permis de confirmer l’existence d’une inhibition fonctionnelle de Ras.
Après avoir montré l’inhibition de l’activation de Ras, il était important de pouvoir confirmer que le FTS agissait bien sur la forme mature de la protéine et non sur l’ARN messager de celle-ci, dont l’expression augmente après HP. L’expression de l’ARN messager de Ras a alors été analysée par PCR quantitative en temps réel. Celle-ci confirme bien l’absence d’atteinte de l’expression de l’ARNm par le FTS.
Les résultats obtenus nous permettent de dire que l’inhibition fonctionnelle de Ras a, pour conséquence première, une diminution significative de la régénération hépatique et que cette réduction est associée à l’absence de recrutement membranaire de la protéine Raf et, plus en aval, à l’inhibition de l’activation de la protéine Erk.
On peut donc conclure que Ras joue un rôle important dans le mécanisme de la régénération hépatique et que ce mécanisme pourrait impliquer la voie Ras/Raf/MERK1,2/Erk1,2

Oncogene Proteins --- Rats --- Hepatectomy --- Liver

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This third edition of the standard reference on the nervous system of the rat is a complete and updated revision of the 1994 second edition. All chapters have been extensively updated, and new chapters added covering early segmentation, growth factors, and glia. The book is now aligned with the data available in the Rat Brain in Stereotaxic Coordinates, making it an excellent companion to this bestselling atlas. Physiological data, functional concepts, and correlates to human anatomy and function round out the new edition.*Designed to be used in conjunction with the bestselling

Animal psychology and neurophysiology --- Mammals --- Rats --- Brain --- Anatomy --- Nervous system --- Rat --- Muridae --- Nervous system. --- Anatomy. --- Rats - Anatomy --- Brain - Anatomy --- Rats - Nervous system

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Fructans are soluble fibres, considered as functional food components, which are highly fermented in the caeco-colon. According to previous studies in the PMT laboratory, oligofructose (OFS), a fructan derived from chicory root insulin, positively modifies disorders in lipid metabolism caused by nutritional factors. It was also shown that it up regulates the synthesis of GLP-1 in the colon and, as a consequence, increases its serum level. GLP-1 is an incretin peptide which plays the role of a hormonal relay in the intestine. It also plays a role in the regulation of food intake and could, therefore, be used to regulate obesity and related metabolic disorders. However, the half-life of GLP-1 in the plasma is short because it is rapidly degraded by a serum enzyme: dipeptidyl peptidase IV (DPP IV). We have tested the following hypothesis: dietary fructans could modulate the activity of DPP IV , which would contribute to the favorable metabolic effects of such dietary fibers.
Male Wistar rats were pre-treated with a standard diet or OFS-enriched (10%) standard diet for 35 days followed by 15 days of high fat diet enriched or not with OFS (10%). On day 50, rats were-food deprived 8 hours and anaesthetized for blood, and intestinal tissue sampling for further analysis.
The addition of OFS in the diet protects against high fat diet modulation of energy intake, body weight gain, fat mass development and serum triglycerides accumulation. OFS fermentation leads to an increase in proglucagon/GLP-1 synthesis in the proximal colon with consequences on the portal concentration of GLP-1 (increase). This effect is accompanied by a decrease of the DPP IV activity in the blood but not in the intestines and the liver. After a 24 h-fasting period, the activity of DPP IV in the blood is increased, a phenomenon which could contribute, in this condition, to a decrease plasma GLP-1.
As a conclusion, in rats exposed to high fate diets, OFS is thus able to modulate endogenous production of gut peptides involved in appetite and body weight regulation. This effect could be dependent on reduction of the DPP IV activity in the blood. If this mechanism is confirmed in humans, the use of fructans as nutritional adjuvants for medical therapy for 2 diabetes and associated metabolic could be interesting for future studies Les fructanes sont des fibres solubles fermentescibles de plus en plus utilises en tant que constituants d’aliments fonctionnels. D’après des études réalisées précédemment au laboratoire PMNT, l’oligofructose (OFS) est capable de modifier positivement les désordres du métabolisme lipidique, engendrés par des manipulations nutritionnelles. Il a également été démontré que celui-ci permet d’augmenter la synthèse du GLP-1 colique et ses taux circulants, ce peptide jouant le rôle de « relais hormonal ». Le GLP-1, outre son action insulino-sécrétagogue, aurait pour effet de réguler l’appétit, d’où son intérêt dans les pathologies associées à l’obésité. Cependant, la demi-vie plasmique du GLP-1 actif est réduite car il est rapidement clivé par l’enzyme dipeptidyl peptidase IV (DPP IV). Notre hypothèse de travail est la suivante : les fructanes alimentaires pourraient être une voie de modulation nutritionnelle de l’activité de la DPP IV, ce qui contribuerait aux effets métaboliques favorables de ces fibres alimentaires.
Des rats mâles Wistar ont été prétraités 35 jours avec une diète standard ou supplémentée en OFS (10%), suivie par 15 jours de traitement avec une diète hyper lipidique standard ou supplémentée en OFS (10%). Les échantillons biologiques intestinaux et sanguins ont été collectés 8 h après les derniers repas en vue des différentes mesures.
Face aux désordres métaboliques induits par la diète hyper lipidique, l’OFS semblerait exercer un effet protecteur en amenuisant la prise alimentaire, le gain de poids corporel, la triglycéridémie ainsi que le développement de la masse grasse. La fermentation de l’OFS dans le côlon proximal entraîne une augmentation de la synthèse du proglucagon/GLP-1, avec pour conséquence, l’augmentation du GLP-1 portal. Cet effet s’accompagne d’une diminution de l’activité de la DPP IV plasmique sans modification de son activité dans les différents segments de l’intestin et du foie. Notons que durant un jeûne de 24 h, l’activité sérique DPP IV est augmentée, ce qui pourrait contribuer, dans cette condition, à la diminution du taux de GLP-1 plasmatique.
En conclusion, chez les rats exposés à une diète enrichie en lipides, l’OFS est capable de moduler la production endogène de GLP-1 impliqué dans la régulation de l’appétit et du métabolisme glucidique, notamment via une diminution de l’activité de la DPP IV circulante. Si ce mécanisme se confirme chez l’homme, l’usage de l’oligofructose, en tant qu’adjuvant nutritionnel à la thérapie médicamenteuse du diabète de type 2 et du syndrome pluri métabolique associé, s’avérerait une perspective intéressante à ce travail

Glucagon-Like Peptide 1 --- Dipeptidyl Peptidase 4 --- Rats

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The provision of foraging opportunities may be a simple way of improving an animal's welfare, but this approach has been neglected for laboratory rats (Rattus norvegicus). Standard housing contains little enrichment and food is often provided ad libitum, which may result in inactivity and obesity, especially in mature males. Foraging enrichments may offer a way to correct these deficiencies. This study compared three potential enrichments - a limited-access hopper, gnawing sticks and a foraging device - to standard housing and feeding conditions, in order to examine their effects on rat body weight food consumption, behaviour and preferences. The subjects were 12 mature male Wistar rats. Effects were assessed from daily weighing and from video records of the rats' behaviour over 24 h periods. The rats' preferences were determined using a four-way test system in which they could choose between a standard cage and cages offering the three potential enrichments. Compared to the standard housing and feeding, the limited-access hopper had a tendency to reduce food consumption, but the time spent feeding increased The gnawing sticks provided the rats with the opportunity to gnaw, but did not affect other behaviours or body weight. The foraging device had the benefits of reducing aggression and allowing the rats to search for and manipulate food, but resulted in significant gains in body weight. Additionally, the foraging device was the preferred feeding source. Of the four possible feeding locations, the rats spent the least amount of time in the standard cage. The foraging device provided the most benefits but requires further modification to address problems of obesity

Ad-libitum. --- Aggression. --- Animal welfare. --- Animal-welfare. --- Animal. --- Behavior. --- Behaviour. --- Body weight. --- Body-weight. --- Cage. --- Cages. --- Device. --- Enrichment. --- Environmental enrichment foraging. --- Feeding. --- Food. --- Foraging. --- Housing. --- Inactivity. --- Laboratory rat. --- Laboratory rats. --- Laboratory. --- Male. --- Males. --- Meal patterns. --- Mongolian gerbil. --- Nesting materials. --- Old. --- Periods. --- Preference. --- Preferences. --- Procurement cost. --- Provision. --- Rat. --- Rats. --- Rattus norvegicus. --- Rattus-norvegicus. --- Search. --- Social contact. --- System. --- Test. --- Time. --- Video. --- Weight. --- Welfare. --- Wistar rats.

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The present Study aimed to measure anticipatory activity in farmed mink (Mustela vison) to study the effects of the presence of environmental enrichments in three housing systems differing in cage structure and in the amount of enrichments. In studies on laboratory rats, anticipatory activity is used as a parameter for potentially stressful conditions of different housing systems: enriched housed rats were less sensitive to sucrose-rewards than standard housed rats as shown by a lower anticipatory reactivity suggesting less stressful conditions. Anticipatory activity in 36 adult female mink was elicited in a Pavlovian paradigm with tasty cat food as reward in January 2002. This Pavlovian paradigm was followed by an observation of stereotypical behaviour in February 2002. The Pavlovian paradigm and the observations of stereotypical behaviour were preceded by observations on juvenile mink behaviour (7-11 weeks of age) in the Summer of 2001. The results of this study show (1) that mink juveniles in the most enriched system had more variable behaviour suggesting better coping potentials and (2) that mink, just like rats, can be trained to anticipate on reward. However, in contrast to the behavioural results of the juveniles, no significant long-term effects of the additional enrichments were found as measured by anticipatory activity and stereotypical behaviour, suggesting no differences between the experimental housing systems in terms of stress. (C) 2004 Elsevier B.V. All rights reserved

Activity. --- Adult. --- Age. --- Behaviour. --- Boxes. --- Brain. --- Cage. --- Cat. --- Coping. --- Enriched. --- Enrichment. --- Environmental enrichment. --- Farmed mink. --- Female. --- Food. --- Housed rats. --- Housing system. --- Housing. --- Laboratory rat. --- Laboratory rats. --- Laboratory. --- Long-term. --- Mink. --- Mustela-vison. --- Observation. --- Potentials. --- Rat. --- Rats. --- Reactivity. --- Reward. --- Stress. --- System. --- Systems. --- Time. --- Welfare.