Effets d'un épisode de séparation maternelle sur l'intégrité de la barrière intestinale et sur le transcriptome hépatique chez le nouveau-né

Aujourd'hui, de plus en plus d'études épidémiologiques témoignent que des facteurs environnementaux apparaissant tôt dans la vie peuvent influencer le développement de l'enfant, jusqu'à le prédisposer à développer plus tard dans la vie des maladies chroniques, d'ordre métaboliques ou encore auto-immunes. Pour l'intestin de nombreuses études chez l'animal ont mis en lumière l'importance de la présence maternelle au cours des premiers jours de vie dans l'apparition à l'âge adulte de pathologies spécifiques, tels que des troubles fonctionnels digestifs, comme le syndrome de l'intestin irritable (SII). Un modèle expérimental de stress de séparation maternelle (SM) chronique s'est alors développé, permettant de mimer les symptômes associés au SII tels qu'une altération du transit intestinal, une augmentation de la perméabilité intestinale ainsi qu'une hyperalgésie viscérale. . Les défauts de barrière intestinale sont associés à une altération persistante de l'axe corticotrope et à l'action de médiateurs du stress au niveau du côlon. Toutes ces observations ont conclu qu'une SM chronique durant la période postnatale pouvait avoir des effets délétères favorisant le passage incontrôlé de bactéries vers les organes systémiques et aboutissant à un état de micro inflammation. Alors que toutes ces manifestations ont essentiellement été décrites chez l'adulte, aucune étude n'a en retour considéré l'impact immédiat du stress accompagnant la SM chez le nouveau-né, quand celui-ci présente une barrière intestinale et des fonctions hépatiques en cours de maturation, donc sensibles à de nombreux facteurs environnementaux. C'est d'autant plus le cas chez le nouveau-né prématuré qui présente une vulnérabilité plus élevée, car plus sensible à l'action des glucocorticoïdes (GC) qui participent à la maturation des fonctions intestinales, en particulier dans le côlon. De plus, aucune étude ne s'est intéressée aux effets de la SM sur le développement des fonctions hépatiques, alors qu'elles s'établissent en lien étroit avec celui de l'intestin. Au cours de ce travail de thèse, nous avons développé un modèle expérimental de SM chez le rat en utilisant un épisode unique et de courte durée (4 heures) de privation maternelle telle qu'elle est pratiquée en routine lors de la prise en charge d'un nouveau-né dans les services hospitaliers, en particulier chez l'enfant prématuré. Dans l'optique d'en analyser les conséquences selon l'âge de l'individu, nous avons privilégié deux stades de développement chez le rat : 10 jours et 20 jours post-partum. D'après la littérature le rat âgé de 10 jours, constitue un modèle pertinent à plusieurs égards pour l'enfant prématuré, en particulier concernant l'immaturité de son intestin et de ses fonctions de barrière, en plus des fonctions hépatiques. Au 20ième jour post-partum, le jeune rat est à la veille du sevrage et est à un stade de maturité plus avancé, relativement comparable à celui d'un enfant né à terme. A ces deux âges, nous avons déterminé les conséquences immédiates de la SM unique sur la barrière intestinale, en étudiant les paramètres clés tels que la perméabilité intestinale et la translocation bactérienne, et étendu l'étude à l'impact sur le transcriptome hépatique, une approche sans a priori permettant d'apprécier l'impact global de la SM sur le foie en développement. Les résultats de ses travaux montrent qu'une SM de 4h appliquée une seule fois chez le nouveau-né est bien un facteur de stress, augmentant la corticostéronémie tant pour le jeune rat de 10 jours que pour celui de 20 jours, mais avec des conséquences délétères sur la barrière intestinale essentiellement observées chez le raton de 10 jours. Chez ce dernier, l'impact du stress se traduit par une augmentation de la perméabilité colique aux macromolécules, déjà élevée en conditions basales, et par une translocation bactérienne systémique vers le foie et la rate. Nous avons montré que ces deux effets étaient liés à l'activité de l'enzyme MLCK impliquée dans la contraction du cytosquelette des cellules épithéliales. En amont, l'augmentation de la perméabilité intestinale était liée à l'action des GC via le récepteur aux glucocorticoïdes (GR) exprimé dans l'intestin. Une augmentation de la corticostéronémie liée au stress de SM était également observée chez le rat de 20 jours, mais sans pour autant aboutir à des effets délétères sur la muqueuse intestinale. Nous avons constaté que l'intestin du rat de 20 jours était moins sensible à l'action des GC et présentait une faible expression en GR au niveau du côlon en comparaison du rat de 10 jours. Chez ce dernier, les impacts de la SM unique aboutissent également à une modification du transcriptome hépatique, tout particulièrement à une sous-expression de plusieurs gènes impliqués dans la division cellulaire. Ces effets dans le foie sont apparus transitoires et plus subtils que ceux retrouvés au niveau de l'intestin, et dont les éventuelles conséquences fonctionnelles restent à être évalués. Les travaux de cette thèse reposant sur le modèle rat ont révélé des effets délétères de la séparation maternelle sur la barrière intestinale en cours de développement et des effets indépendants et plus subtils sur le transcriptome hépatique. Ces observations révèlent la nécessité de prise en compte du stress de la séparation maternelle, pratiquée en routine chez l'homme et notamment chez l'enfant prématuré sur la santé de l'enfant et plus tard chez l'adulte.Currently, an increasing number of epidemiological evidences suggest that environmental factors occurring early in life can influence the development of the child and predispose him later in life to develop chronic, metabolic or autoimmune diseases. Concerning the gut, many animals' studies have highlighted the importance of maternal presence during the first days of life on the development of intestinal diseases occurring later in adulthood, such as functional gastrointestinal disorders as irritable bowel syndrome (IBS). An experimental model of chronic maternal separation (MS) stress was developed to mimic symptoms associated with irritable bowel syndrome such as alteration of motility, increased intestinal permeability and visceral hyperalgesia. Alterations of the intestinal barrier were associated with a persistent impairment of the HPA axis and with the action of stress mediators in the colon. All of these observations evidenced that chronic exposure to MS during the early postnatal period could have deleterious effects on the intestinal barrier favoring bacteremia and leading to a micro inflammation state. All these events were mostly described at adulthood but no study had investigated the immediate impact of the maternal separation stress directly in the newborn while his intestinal barrier and liver functions are still maturing and potentially sensitive to environmental stress. This is especially the case of preterm infant who are more vulnerable to stress because they are more sensitive to the action of glucocorticoids involved in intestinal functions maturation, especially in the colon. In addition, no study had investigated the effects of maternal separation on the development of liver function that are still maturing after birth and are in close crosstalk with the intestine. In this thesis, we have developed an experimental model of maternal separation in rats using a single short-term exposure of 4 hours as it could be observed in practice in maternity, especially in prematurity birth cases. In order to analyze the consequences of MS according to the age, we focused on two stages of development in the rat: on the 10th and 20th day post delivery. The 10-day-old rat is often considered as a good model for the premature infant, notably due to an immature gut barrier. At 20 days of age, the rat is at a higher stage of maturity, comparable to the state of a full-term infant. At both ages, we determined the immediate consequences of a single MS on the intestinal barrier by studying key parameters such as intestinal permeability and bacterial translocation. We also extended the study to the impact of a single MS on the liver transcriptome using a genome-wide approach without a priori to assess the overall impact of SM on the developing liver. Our results show that a 4 hours-MS is a stressful event increasing corticosterone level in 10- and 20-day-old rats but associated with deleterious effects on the intestinal barrier in 10-day-old rats only. At this age, the MS stress induced an increase of colonic permeability to macromolecules, already high at baseline, and systemic bacterial translocation to the liver and spleen. Both effects were linked to the activity of the MLCK enzyme involved in the contraction of the cytoskeleton of epithelial cell. The increase of intestinal permeability was related to glucocorticoids (GC) action through intestinal glucocorticoid receptors (GR). The increase of glucocorticoid level was also observed in 20 days rats but without deleterious effects on the intestinal mucosa. We have found that the intestine of 20-day-old rats was less sensitive to GC action and expressed GR at a lower level in the colon compared to 10-day-old rats. At the younger age, the maternal separation also led to a modulation of the liver transcriptome, most notably a down-regulation of genes involved in cell division. These effects were transient and more subtle than those observed on the intestinal barrier. Their potential functional consequences remain to be evaluated. The work of this PhD based on the 10-day-old rat model highlighted the deleterious effects of maternal separation on the intestinal barrier during development as well as independent and more subtle effects on the liver transcriptome. These findings emphasize the need to take into account the stress induced by maternal separation, practiced routinely in humans and especially in premature infants, on the health of the child and later at adulthood

Perceptions and Practices of Stimulating Children's Cognitive Development Among Moroccan Immigrant Mothers

We explored the perceptions of children's cognitive development among Moroccan Arabic and Berber immigrant mothers who cannot read, who are less educated, middle educated or highly educated in the Netherlands. A series of in-depth interviews was conducted with 22 mothers with young children (mean age = 5 years and 6 months). Qualitative data analyses revealed five major themes that are of significant importance to these mothers: moral attitudes, social values and religiousness; conversation, reading and playing as stimulating activities; importance attached to education; parental expectations; attributions of school success. The parental perceptions about the cognitive develop

Chronic Early-life Stress in Rat Pups Alters Basal Corticosterone, Intestinal Permeability, and Fecal Microbiota at Weaning: Influence of Sex.

Background/aimsWistar rat dams exposed to limited nesting stress (LNS) from post-natal days (PND) 2 to 10 display erratic maternal behavior, and their pups show delayed maturation of the hypothalamic-pituitary-adrenal axis and impaired epithelial barrier at PND10 and a visceral hypersensitivity at adulthood. Little is known about the impact of early life stress on the offspring before adulthood and the influence of sex. We investigated whether male and female rats previously exposed to LNS displays at weaning altered corticosterone, intestinal permeability, and microbiota.MethodsWistar rat dams and litters were maintained from PND2 to 10 with limited nesting/bedding materials and thereafter reverted to normal housing up to weaning (PND21). Control litters had normal housing. At weaning, we monitored body weight, corticosterone plasma levels (enzyme immunoassay), in vivo intestinal to colon permeability (fluorescein isothiocyanate-dextran 4 kDa) and fecal microbiota (DNA extraction and amplification of the V4 region of the 16S ribosomal RNA gene).ResultsAt weaning, LNS pups had hypercorticosteronemia and enhanced intestinal permeability with females > males while body weights were similar. LNS decreased fecal microbial diversity and induced a distinct composition characterized by increased abundance of Gram positive cocci and reduction of fiber-degrading, butyrate-producing, and mucus-resident microbes.ConclusionsThese data indicate that chronic exposure to LNS during the first week post-natally has sustained effects monitored at weaning including hypercorticosteronemia, a leaky gut, and dysbiosis. These alterations may impact on the susceptibility to develop visceral hypersensitivity in adult rats and have relevance to the development of irritable bowel syndrome in childhood

Perceptions and Practices of Stimulating Children’s Cognitive Development Among Moroccan Immigrant Mothers

We explored the perceptions of children’s cognitive development among Moroccan Arabic and Berber immigrant mothers who cannot read, who are less educated, middle educated or highly educated in the Netherlands. A series of in-depth interviews was conducted with 22 mothers with young children (mean age = 5 years and 6 months). Qualitative data analyses revealed five major themes that are of significant importance to these mothers: moral attitudes, social values and religiousness; conversation, reading and playing as stimulating activities; importance attached to education; parental expectations; attributions of school success. The parental perceptions about the cognitive development of young children differed according to their own educational level. Mothers who cannot read and mothers with less education emphasized the development of moral, social and religious values for strengthening the cultural identity of their children. This sense of identity would enable them to function within their own cultural group and help them to perform well at school. School success was attributed in large part to a combination of the efforts of the child and the school. Middle and highly educated mothers, on the other hand, valued scholastic development and attributed school success to their own efforts and to the kind of support the child received. The ethnic background of the parents, whether Arabic or Berber, did not make a difference in the perceptions

Algorithmes Neuro-Flous de Segmentation d'Images IRM

Résumé- Les méthodes de segmentation existantes sont généralement fondées sur des principes très différents comme par exemple les processus de classification, l’utilisation de modèles de contours déformables ou encore de modèles de connaissance. Mais ces dernières n'utilisent en générale qu'un seul volume d'image ce qui cause dans la plupart des cas une perte considérable d'informations. De plus ces méthodes ont tendance à être dépendantes de certains paramètres à savoir le nombre de classes, les fonctions de similarités qui sont souvent imposés ce qui rend l'application de ces techniques aux images médicales multimodales non appropriées. Le but de notre travail est de montrer qu’il est possible de définir un cadre commun de travail pour permettre la mise en œuvre de coopération entre des approches hétérogènes. L’intérêt d’une telle approche est de pouvoir exploiter la complémentarité d’informations qui résultent de l’application de plusieurs méthodes afin de proposer un système complet de segmentation. Mots clefs- Diagnostic flou; Images médicales Multimodales

New Fuzzy Clustering Algorithm Applied to RMN Image Segmentation

Abstract—An entirely automatic procedure for the classification of cerebral tissues from magnetic resonance nuclear imaging (MRN) 3D of the head are described in this paper. This procedure doesn't make any assumption nor on the number of classes nor on the shape of the density. Indeed, this last is estimated by a non parametric method, it is about the method of the Parzen's Kernel. A new objective function is proposed to improve the FCM algorithm by the addition of one term of entropy aiming to maximize the number of "good " ordering. A supplementary correction is operated by a probabilistic procedure said of fuzzy relaxation including the probabilities of the neighboring points. The validation of the algorithm is made on simulated data and on real cerebral imaging RMN

Limited Nesting Stress Alters Maternal Behavior and In Vivo Intestinal Permeability in Male Wistar Pup Rats.

A few studies indicate that limited nesting stress (LNS) alters maternal behavior and the hypothalamic pituitary adrenal (HPA) axis of dams and offspring in male Sprague Dawley rats. In the present study, we evaluated the impact of LNS on maternal behavior in Wistar rats, and on the HPA axis, glycemia and in vivo intestinal permeability of male and female offspring. Intestinal permeability is known to be elevated during the first week postnatally and influenced by glucocorticoids. Dams and neonatal litters were subjected to LNS or normal nesting conditions (control) from days 2 to 10 postnatally. At day 10, blood was collected from pups for determination of glucose and plasma corticosterone by enzyme immunoassay and in vivo intestinal permeability by oral gavage of fluorescein isothiocyanate-dextran 4kDa. Dams exposed to LNS compared to control showed an increase in the percentage of time spent building a nest (118%), self-grooming (69%), and putting the pups back to the nest (167%). LNS male and female pups exhibited a reduction of body weight by 5% and 4%, adrenal weights/100g body weight by 17% and 18%, corticosterone plasma levels by 64% and 62% and blood glucose by 11% and 12% respectively compared to same sex control pups. In male LNS pups, intestinal permeability was increased by 2.7-fold while no change was observed in females compared to same sex control. There was no sex difference in any of the parameters in control pups except the body weight. These data indicate that Wistar dams subjected to LNS during the first postnatal week have an altered repertoire of maternal behaviors which affects the development of the HPA axis in both sexes and intestinal barrier function in male offspring

Brain corticotropin‐releasing factor signaling: Involvement in acute stress‐induced visceral analgesia in male rats

BackgroundWater avoidance stress (WAS) induces a naloxone-independent visceral analgesia in male rats under non-invasive conditions of monitoring. The objective of the study was to examine the role of brain CRF signaling in acute stress-induced visceral analgesia (SIVA).MethodsAdult male Sprague-Dawley rats were chronically implanted with an intracerebroventricular (ICV) cannula. The visceromotor response (VMR) to graded phasic colorectal distension (CRD: 10, 20, 40, 60 mm Hg, 20 seconds, 4 minutes intervals) was monitored using manometry. The VMR to a first CRD (baseline) was recorded 5 minutes after an ICV saline injection, followed 1 hour later by ICV injection of either CRF (30, 100, or 300 ng and 1, 3, or 5 μg/rat) or saline and a second CRD, 5 minutes later. Receptor antagonists against CRF1 /CRF2 (astressin-B, 30 μg/rat), CRF2 (astressin2 -B, 10 μg/rat), oxytocin (tocinoic acid, 20 μg/rat), or vehicle were injected ICV 5 minutes before CRF (300 ng/rat, ICV) or 15 minutes before WAS (1 hour).Key resultsICV CRF (100 and 300 ng) reduced the VMR to CRD at 60 mm Hg by -36.6% ± 6.8% and -48.7% ± 11.7%, respectively, vs baseline (P < 0.001), while other doses had no effect and IP CRF (10 µg/kg) induced visceral hyperalgesia. Astressin-B and tocinoic acid injected ICV induced hyperalgesia and prevented the analgesic effect of ICV CRF (300 ng/rat) and WAS, while astressin2 -B only blocked WAS-induced SIVA.Conclusions & inferencesThese data support a role for brain CRF signaling via CRF2 in SIVA in a model of WAS and CRD likely mediated by the activation of brain oxytocin pathway