Guillaume Poupeau

Supplementation With Galactooligosaccharides and Inulin Increases Bacterial Translocation in Artificially Reared Newborn Rats

Supplementation of formulas with prebiotics enhances the growth of lactate producing bacteria, and fecal lactate, and acetate levels in infants. High concentrations of organic acids in intestinal lumen have, however, been shown to impair the intestinal barrier function. To determine whether stimulating the colonic microbiotal metabolism with prebiotics would impair the neonatal intestinal barrier function, artificially reared rats were fed milk formula with or without a mixture of galactooligosaccharides/inulin (GOS/Inulin, 88/12; 5.6 g/L) from the 7th d of life (d7) until weaning (d20). At d18, GOS/inulin supplementation had increased the concentrations of acetate and lactate in colonic lumen. Although neither ileum-associated microbiota nor colonic permeability (assessed in Ussing chambers), nor the expression of tight junction claudin-2 and claudin-3 mRNA were altered, GOS/inulin supplementation was associated with increased bacterial translocation (BT) toward spleen. None of these effects persisted at d40. We conclude that GOS/inulin supplementation may increase BT in an immature gut. The balance between the potential infectious risk of BT vs. its putative beneficial effect on the maturation of neonatal immune system clearly warrants further study.

Simultaneous detection of stable isotope-labeled and unlabeled l-tryptophan and of its main metabolites, l-kynurenine, serotonin and quinolinic acid, by gas chromatography/negative ion chemical ionization mass spectrometry

A method for the detection of unlabeled and (15)N2 -labeled L-tryptophan (L-Trp), L-kynurenine (L-Kyn), serotonin (5-HT) and quinolinic acid (QA) in human and rat plasma by GC/MS is described. Labeled and unlabeled versions of these four products were analyzed as their acyl substitution derivatives using pentafluoropropionic anhydride and 2,2,3,3,3-pentafluoro-1-propanol. Products were then separated by GC and analyzed by selected ion monitoring using negative ion chemical ionization mass spectrometry. L-[(13)C11, (15)N2]-Trp, methyl-serotonin and 3,5-pyridinedicarboxylic acid were used as internal standards for this method. The coefficients of variation for inter-assay repeatability were found to be approximately 5.2% for L-Trp and (15)N2-Trp, 17.1% for L-Kyn, 16.9% for 5-HT and 5.8% for QA (n = 2). We used this method to determine isotope enrichments in plasma L-Trp over the course of a continuous, intravenous infusion of L-[(15) N2 ]Trp in pregnant rat in the fasting state. Plasma (15)N2-Trp enrichment reached a plateau at 120 min. The free Trp appearance rate (Ra) into plasma was 49.5 ± 3.35 µmol/kg/h. The GC/MS method was applied to determine the enrichment of (15)N-labeled L-Trp, L-Kyn, 5-HT and QA concurrently with the concentration of non-labeled L-Trp, L-Kyn, 5-HT and QA in plasma. This method may help improve our understanding on L-Trp metabolism in vivo in animals and humans and potentially reveal the relative contribution of the four pathways of L-Trp metabolism.

Maternal and fetal tryptophan metabolism in gestating rats: effects of intrauterine growth restriction

Abstractl-Tryptophan (l-Trp) is a precursor for serotonin (5-HT) and nicotinamide adenine dinucleotide (NAD) synthesis. Both 5-HT and NAD may impact energy metabolism during gestation given that recent studies have demonstrated that increased 5-HT production is crucial for increasing maternal insulin secretion, and that sirtuin, an NAD+-dependent protein deacetylase, regulates endocrine signaling. Infants born with intrauterine growth restriction (IUGR) are at a higher risk of metabolic disease once they reach adulthood. IUGR is associated with altered maternal–fetal amino acid transfer. Whether IUGR affects l-Trp metabolism in mother and fetus has not been fully elucidated. Recently, we developed an analytical method using stable isotope-labeled l-Trp to explore the metabolism of l-Trp and its main metabolites, l-kynurenine (l-Kyn), 5-HT and quinolinic acid (QA). In this study, dams submitted to dietary protein restriction throughout gestation received intravenous infusions of stable isotope-labeled 15N2-l-Trp to determine whether l-Trp metabolism is affected by IUGR. Samples were obtained from maternal, fetal and umbilical vein plasma, as well as the amniotic fluid (AF), placenta and liver of the mother and the fetus after isotope infusion. We observed evidence for active l-Trp transfer from mother to fetus, as well as de novo synthesis of 5-HT in the fetus. Plasma 5-HT was decreased in undernourished mothers. In IUGR fetuses, maternal–fetal l-Trp transfer remained unaffected, but conversion to QA was impaired, implying that NAD production also decreased. Whether such alterations in tryptophan metabolism during gestation have adverse consequences and contribute to the increased risk of metabolic disease in IUGR remains to be explored.

Perinatal Hypercholesterolemia Exacerbates Atherosclerosis Lesions in Offspring by Altering Metabolism of Trimethylamine-N-Oxide and Bile Acids

Objective— Experimental studies suggest that maternal hypercholesterolemia may be relevant for the early onset of cardiovascular disease in offspring. We investigated the effect of perinatal hypercholesterolemia on the atherosclerosis development in the offspring of apolipoprotein E–deficient mice and the underlying mechanism. Approach and Results— Atherosclerosis and related parameters were studied in adult male or female apolipoprotein E–deficient mice offspring from either normocholesterolemic or hypercholesterolemic mothers and normocholesterolemic fathers. Female born to hypercholesterolemic mothers had more aortic root lesions than female born to normocholesterolemic mothers. Lesions in whole aorta did not differ between groups. Higher trimethylamine-N-oxide levels and Fmo3 hepatic gene expression were higher in female born to hypercholesterolemic mothers offspring compared with female born to normocholesterolemic mothers and male. Trimethylamine-N-oxide levels were correlated with the size of atherosclerotic root lesions. Levels of hepatic cholesterol and gallbladder bile acid were greater in male born to hypercholesterolemic mothers compared with male born to normocholesterolemic mothers. At 18 weeks of age, female born to hypercholesterolemic mothers showed lower hepatic Scarb1 and Cyp7a1 but higher Nr1h4 gene expression compared with female born to normocholesterolemic mothers. Male born to hypercholesterolemic mothers showed an increase in Scarb1 and Ldlr gene expression compared with male born to normocholesterolemic mothers. At 25 weeks of age, female born to hypercholesterolemic mothers had lower Cyp7a1 gene expression compared with female born to normocholesterolemic mothers. DNA methylation of Fmo3, Scarb1, and Ldlr promoter regions was slightly modified and may explain the mRNA expression modulation. Conclusions— Our findings suggest that maternal hypercholesterolemia may exacerbate the development of atherosclerosis in female offspring by affecting metabolism of trimethylamine-N-oxide and bile acids. These data could be explained by epigenetic alterations.

SFRP-02 – Recherche expérimentale – Effet mémoire d’une alimentation hyperprotéique chez le raton

Les nouveau-nes de petit poids de naissance ayant souffert d’un retard de croissance intra-uterin (RCIU) sont une population a haut risque pour developper un « syndrome metabolique » (obesite, hypertension, insulino-resistance) a l’âge adulte. Ces bebes sont generalement nourris avec des laits « enrichis » en proteines afin de leur assurer une croissance de rattrapage. Or le risque de pathologies cardiovasculaires ou d’obesite a l’âge adulte est correle a la vitesse de croissance dans les toutes premieres semaines de vie. Notre hypothese est qu’une alimentation hyperproteique prescrite en periode neonatale chez des nouveau-nes RCIU pourrait avoir des consequences nefastes sur la sante a l’âge adulte. Pour etudier les consequences a distance d’une alimentation neonatale hyperproteique, des ratons RCIU, nes de meres carencees en proteines durant la gestation (60 % de carence) ont ete equipes au 5eme jour de vie (J5) d’une canule de gastrostomie posee de facon percutanee (modele de ratons « pup in the cup »). Ils ont recu, de J5 (pose de la canule) a J21 (jour du sevrage) et de facon randomisee, soit un lait « normoproteique » (NP : 87,0 g/l, n = 9), soit un lait « hyperproteique » (HP : 130,5 g/l, n = 7). La croissance des rats et les quantites ingerees ont ete suivies regulierement (de J5 a J160, rat adulte). Le lait HP ne modifie pas la croissance des ratons avant le sevrage. Le lait HP presente des effets a distance : (1) un surpoids des rats nourris au lait hyperproteique vs. normoproteique entre J50 et J65, (HP : 329 ± 14 g vs . NP : 307 ± 28 g pour des rats de J60, p = 0,079), lie a une augmentation des quantites ingerees d’aliments (de J50 a J60, HP : 28 ± 3 g/j vs . NP : 25 ±3 g/j, p = 0.079) ; (2) une augmentation, de la masse du pancreas (0,29 ± 0,15 g vs . 0,17 ± 0,06, p = 0.046) et de la masse de la graisse mesenterique (6,3 ±1,4g vs . 4,5 ± 2, 1, p = 0,088) respectivement chez des rats HP vs. NP. Ces donnees preliminaires suggerent qu’un lait hyperproteique, administre a des ratons RCIU durant la periode neonatale, a des effets memoire sur le poids et la masse grasse mesenterique, traduisant des desordres metaboliques. Des mesures de composition corporel le, glucose et insuline plasmatiques, nous permettront de preciser ces perturbations metaboliques.