Dietary glutamine supplementation affects macrophage function, hematopoiesis and nutritional status in early weaned mice

Background Et aims: To investigate the effect that early weaning associated with the ingestion of either a glutamine-free or supplemented diet has on the functioning of peritoneal. macrophages, hematopoiesis and nutritional status of mice. Methods: Swiss Webster mice were early weaned on their 14th day of life and distributed to two groups, being fed either a glutamine-free diet (-GLN) or a glutamine-supplemented diet (+GLN). Animals belonging to a control group (CON) were weaned on their 21st day of life. Results: The -GLN and +GLN groups had a lower lean body mass, carcass protein and ash content, plasma glutamine concentration and lymphocyte counts both in the peripheral blood and bone marrow when compared to the CON group (P < 0.05). Dietary supplementation with glutamine reversed both the lower concentrations of protein and DNA in the muscle and liver, as well. as the reduced capacity of spreading and synthesizing nitric oxide, hydrogen peroxide, TNF-alpha, IL-1 beta and IL-6 in cultures of peritoneal. macrophages obtained from the -GLN group (P < 0.05). Conclusion: These data indicate that the ingestion of glutamine modulates the function of peritoneal macrophages in early weaned mice. However, a glutamine-supplemented diet cannot substitute maternal milk in respect to immunological and metabolic parameters. (C) 2008 Elsevier Ltd and European Society for Clinical Nutrition and Metabolism. All rights reserved.

Glutamine in vitro supplementation partly reverses impaired macrophage function resulting from early weaning in mice

Objective: Glutamine is one of the most abundant amino acids found in maternal milk, and its concentration increases throughout lactation. Because glutamine is essential for macrophage functionality, it is hereby suggested that early weaning in conjunction with the absence of glutamine consumption impairs the functioning of macrophages, which could in turn be reversed with an in vitro supplementation with glutamine. Methods: Swiss Webster mice were early weaned at 14 d of age (EW group) or at 21 d of age (control group, n = 8 per group). The EW group was fed a glutamine-free diet from days 14 to 21 of life. Results: Mice in the EW group presented a significant decrease in plasma and muscle concentrations of glutamine and an increase in the activity of glutamine synthetase in the muscle. Peritoneal macrophages obtained from animals in the EW group presented a significant increase in the production of interleukin (IL)-10 and a significant decrease in the synthesis of IL-1 beta, IL-6, tumor necrosis factor-a, nitric oxide, and hydrogen peroxide and in their ability to adhere, spread, phagocytize, and kill fungi. Glutamine in vitro supplementation reversed the decrease in IL-6, nitric oxide, and hydrogen peroxide synthesis and the decrease in the capacity to adhere, spread, and phagocytize in animals of the EW group. Conclusion: These new. data may imply that a lack of glutamine intake in early weaned mice hampers the functioning of peritoneal macrophages. (C) 2008 Elsevier Inc. All rights reserved.

Dietary glutamine supplementation increases the activity of peritoneal macrophages and hemopoiesis in early-weaned mice inoculated with Mycobacterium bovis bacillus Calmette-Guerin

Infants who are breast-fed have been shown to have a lower incidence of certain infectious diseases compared with formula-fed infants. Glutamine is one of the most abundant amino acids found in maternal milk and it is essential for the function of immune system cells such as macrophages. The purpose of this study was to investigate the effect of glutamine supplementation on the function of peritoneal macrophages and on hemopoiesis in early-weaned mice inoculated with Mycobacterium bovis bacillus Calmette-Guerin (BCG). Mice were wearied at 14 d of age and distributed to 2 groups and fed either a glutamine-free diet (n = 16) or a glutamine-supplemented diet (+Gln (n = 16). Both diets were isonitrogenous (with addition of a mixture of nonessential amino acids) and isocaloric. At d 21, 2 subgroups of mice (n = 16) were intraperitoneally injected with BCG and all mice were killed at d 28. Plasma, muscle and liver glutamine concentrations and muscle glutamine synthetase activity were not affected by diet or inoculation with BCG. The +GIn diet led to increased leukocyte and lymphocyte counts in the peripheral blood (P < 0.05) and granulocyte and lymphocyte counts in the bone marrow and spleen (P < 0.05). The +GIn diet increased spreading and adhesion capacities, hydrogen peroxide, nitric oxide, and tumor necrosis factor-alpha (TNF alpha) syntheses and the phagocytic and fungicidal activity of peritoneal macrophages (P < 0.05). The interaction between the +GIn diet and BCG inoculation increased the area under the curve of interleukin (IL)-1 beta and TNF alpha syntheses (P < 0.05). In conclusion, the intake of glutamine increases the function of peritoneal macrophages and hemopoiesis in early-weaned and BCG-inoculated mice. These data have important implications for the design of breast milk substitutes for human infants.

The effects of enzymatic interesterification on the physical-chemical properties of blends of lard and soybean oil

The main goal of the present research effort was to evaluate the physical-chemical properties of blends of lard and soybean oil following enzymatic interesterification catalyzed by an immobilized lipase from Thermomyces lanuginosa (Lipozyme (TM) TL IM). Lipase-catalyzed interesterification produced new tri-acylglycerols that changed the physical-chemical properties of the fat blends under study. Solid fat content (31.3 vs 31.5 g/100 g), consistency (104.7 vs 167.6 kPa), crystallized area (0.6 vs 11.8) and softening point (31.8 vs 32.2 degrees C) of lard increased after interesterification, and this was mostly due to the increase of SSS (saturated) + SSU (disaturated-monounsaturated) triacylglycerols. These contents (SSU + SSS) increased in lard after interesterification from 42.9 to 46.7 g/100 g. The interesterified blends exhibited lower values for the physical properties when compared with their counterparts before enzymatic interesterification. The interesterification of blends of lard with soybean oil increased the amounts of UUU (triunsaturated) and SSS triacylglycerols and reduced the amounts of UUS (diunsaturated-monosaturated) triacylglycerols. The interesterified blends of lard and soybean oil demonstrated physical properties and chemical composition similar to human milk fat and they could be used for the production of a human milk fat substitute. (C) 2009 Elsevier Ltd. All rights reserved.

Metformin therapy and diabetes in pregnancy

No adverse pregnancy outcomes with metformin use have been reported, except in one unmatched study. Otherwise, the studies are small and non-randomised, with the exception of one prospective, randomised controlled trial, currently under way, comparing metformin with insulin in women with gestational diabetes mellitus (the MiG trial). No long-term follow-up data for offspring of mothers receiving metformin have been published. Any woman with diabetes should be as close to euglycaemia as possible before pregnancy. In some circumstances (eg, severe insulin resistance), metformin therapy during pregnancy may be warranted. When metformin treatment is being considered, the individual risks and benefits need to be discussed with the patient so that an appropriate decision can be reached.

Respiratory Infection, Exposure to Mouse Allergen and Breastfeeding: Role in Recurrent Wheezing in Early Life

Background: Environmental factors may influence the development of allergen sensitization and asthma. The aim of this study was to evaluate the role of endotoxin and allergen exposure in early life as a risk factor for recurrent wheezing. Methods: One hundred and four infants from low-income families, at high risk of asthma, were enrolled at birth. Dust samples were collected from the bedding and bedroom floor within 6 months after birth. Recurrent wheezing was defined as 3 or more wheezing episodes in the past year. Endotoxin was determined by Limulus amebocyte lysate assay, and major indoor allergens were quantitated by ELISA in dust extracts. IgE antibodies were measured by ImmunoCAP at 30 months of age. Results: At 30 months, 51 of the 99 infants who completed the study (51.5%) had recurrent wheezing. Respiratory infection was strongly associated with recurrent wheezing (OR 6.67, 95% CI 1.96-22.72), whereas exclusive breastfeeding for at least 1 month was a protective factor (OR 0.09, 95% CI 0.01-0.51). Exposure to high levels of mouse allergen was more frequent among non-recurrent wheezers, approaching significance (OR 0.12, 95% CI 0.01-1.13; p=0.064). None of the children were sensitized to mouse. Sensitization to mite was found in 26/90 (28.8%) children, with no association with recurrent wheezing. Conclusion: Respiratory infection was strongly associated with recurrent wheezing in the first 30 months of life, in children at high risk of asthma, living in a socially deprived community in Brazil. Copyright (C) 2009 S. Karger AG, Basel

Reproductive development and function of female rats exposed to di-eta-butyl-phthalate (DBP) in utero and during lactation

Phthalates are environmental contaminants used in the production of plastics, cosmetics and medical devices. Studies on the effects of phthalates on female reproductive health are particularly sparse and mostly restricted to high-dose exposure in rats. In the present study, pregnant rats were treated with 100 mg/kg-d of di-eta-butyl-phthalate (DBP) or only the vehicle (control group), from GD 12 to GD 20 for evaluation of reproductive outcomes and fetal gonads analysis (F0), and from GD 12 to PND 21 to evaluate reproductive development and function on F1 female offspring. Results showed that all parameters were comparable between groups, although there was a significant increase in the fetal weight after DBP exposure. However, the body weight at birth was normal. Based on these data we can conclude that, in these experimental conditions, DBP did not disturb the reproductive development or function of female rats. (C) 2009 Elsevier Inc. All rights reserved.

Perfil de citocinas no colostro em função da idade gestacional e do crescimento fetal

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Breastfeeding: making the difference in the development, health and nutrition of term and preterm newborns

Breastfeeding is the natural and safe way of feeding small infants, providing nutritional, immunological, psychological and economic recognized and unquestionable advantages. These qualities are especially important in premature infants, because of their vulnerability. Despite highly desirable, there is, in general, little success in breastfeeding preterm infants, especially in special care neonatal units. There are evidences that a high supportive hospital environment, with an interdisciplinary team, makes possible to these infants to be breastfed. In this article, the authors present an up-to-date review about the components of human milk and its unique characteristics, as well as describes aspects that make the breast milk particularly suitable for feeding the premature newborn.

Evolució i estabilitat d’àcids grassos i vitamines antioxidants a la llet materna

Recerca realitzada durant el periode Gener 2006-Abril 2007, a la Facultat de Farmàcia de la Universitat de Barcelona sota la direcció de la Dra. M. Carmen López Sabater. La nutrició durant els primers anys de vida té una enorme importància, amb repercussions en el creixement i desenvolupament del nen i en les possibles malalties futures. Tenint en compte que la llet materna és l’aliment ideal per al nadó durant els primers sis mesos de vida, és important conéixer la composició nutricional de la mateixa i la seva evolució al llarg de la lactància (calostre, llet de transició i madura). La llet conté nutrients majoritaris (proteïnes, carbohidrats i lípids) i nutrients minoritaris, com vitamines i minerals. S'ha desenvolupat i validat un mètode per a la determinació d'àcids grassos (AG) i Àcid Linoleic Conjugat (CLA) en llet materna per Cromatografia de Gasos "fast" (fast-GC), permetent reduir el temps d'anàlisi considerablement en comparació amb els mètodes utilitzats fins el moment. La repetibilitat i reproducibilitat trobades han estat bones, amb coeficients de variació inferiors al 10% en tots els casos. L'aplicació a 6 mostres de llet materna ha demostrat que es tracta d'un mètode senzill, ràpid, eficient i pràctic per a l'anàlisi rutinari d'un elevat nombre de mostres. També s'ha començat a desenvolupar un mètode per a l'anàlisi dels anàlegs de tocoferol de la llet materna, degut a l'interès de la vitamina E com a antioxidant natural. És un mètode per Cromatografia Líquida d'Ultra Resolució (UPLC) amb detecció per Fluorescència (FD) i per Photodiode Array (PDA) que permet treballar amb quantitats de mostra petites mantenint o inclús millorant la sensibilitat i reduint el temps d’anàlisi.

Prediction of infant drug exposure through breastfeeding: population PK modeling and simulation of fluoxetine

BACKGROUND: Risks of significant infant drug exposurethrough breastmilk are poorly defined for many drugs, and largescalepopulation data are lacking. We used population pharmacokinetics(PK) modeling to predict fluoxetine exposure levels ofinfants via mother's milk in a simulated population of 1000 motherinfantpairs.METHODS: Using our original data on fluoxetine PK of 25breastfeeding women, a population PK model was developed withNONMEM and parameters, including milk concentrations, wereestimated. An exponential distribution model was used to account forindividual variation. Simulation random and distribution-constrainedassignment of doses, dosing time, feeding intervals and milk volumewas conducted to generate 1000 mother-infant pairs with characteristicssuch as the steady-state serum concentrations (Css) and infantdose relative to the maternal weight-adjusted dose (relative infantdose: RID). Full bioavailability and a conservative point estimate of1-month-old infant CYP2D6 activity to be 20% of the adult value(adjusted by weigth) according to a recent study, were assumed forinfant Css calculations.RESULTS: A linear 2-compartment model was selected as thebest model. Derived parameters, including milk-to-plasma ratios(mean: 0.66; SD: 0.34; range, 0 - 1.1) were consistent with the valuesreported in the literature. The estimated RID was below 10% in >95%of infants. The model predicted median infant-mother Css ratio was0.096 (range 0.035 - 0.25); literature reported mean was 0.07 (range0-0.59). Moreover, the predicted incidence of infant-mother Css ratioof >0.2 was less than 1%.CONCLUSION: Our in silico model prediction is consistent withclinical observations, suggesting that substantial systemic fluoxetineexposure in infants through human milk is rare, but further analysisshould include active metabolites. Our approach may be valid forother drugs. [supported by CIHR and Swiss National Science Foundation(SNSF)]

Identification and quantification techniques in mass spectrometry-based proteomics

Résumé La protéomique basée sur la spectrométrie de masse est l'étude du proteome l'ensemble des protéines exprimées au sein d'une cellule, d'un tissu ou d'un organisme - par cette technique. Les protéines sont coupées à l'aide d'enzymes en plus petits morceaux -les peptides -, et, séparées par différentes techniques. Les différentes fractions contenant quelques centaines de peptides sont ensuite analysées dans un spectromètre de masse. La masse des peptides est enregistrée et chaque peptide est séquentiellement fragmenté pour en obtenir sa séquence. L'information de masse et séquence est ensuite comparée à une base de données de protéines afin d'identifier la protéine d'origine. Dans une première partie, la thèse décrit le développement de méthodes d'identification. Elle montre l'importance de l'enrichissement de protéines comme moyen d'accès à des protéines de moyenne à faible abondance dans le lait humain. Elle utilise des injections répétées pour augmenter la couverture en protéines et la confiance dans l'identification. L'impacte de nouvelle version de base de données sur la liste des protéines identifiées est aussi démontré. De plus, elle utilise avec succès la spectrométrie de masse comme alternative aux anticorps, pour valider la présence de 34 constructions de protéines pathogéniques du staphylocoque doré exprimées dans une souche de lactocoque. Dans une deuxième partie, la thèse décrit le développement de méthodes de quantification. Elle expose de nouvelles approches de marquage des terminus des protéines aux isotopes stables et décrit la première méthode de marquage des groupements carboxyliques au niveau protéine à l'aide de réactifs composé de carbone 13. De plus, une nouvelle méthode, appelée ANIBAL, marquant tous les groupements amines et carboxyliques au niveau de la protéine, est exposée. Summary Mass spectrometry-based proteomics is the study of the proteome -the set of all expressed proteins in a cell, tissue or organism -using mass spectrometry. Proteins are cut into smaller pieces - peptides - using proteolytic enzymes and separated using different separation techniques. The different fractions containing several hundreds of peptides are than analyzed by mass spectrometry. The mass of the peptides entering the instrument are recorded and each peptide is sequentially fragmented to obtain its amino acid sequence. Each peptide sequence with its corresponding mass is then searched against a protein database to identify the protein to which it belongs. This thesis presents new method developments in this field. In a first part, the thesis describes development of identification methods. It shows the importance of protein enrichment methods to gain access to medium-to-low abundant proteins in a human milk sample. It uses repeated injection to increase protein coverage and confidence in identification and demonstrates the impact of new database releases on protein identification lists. In addition, it successfully uses mass spectrometry as an alternative to antibody-based assays to validate the presence of 34 different recombinant constructs of Staphylococcus aureus pathogenic proteins expressed in a Lactococcus lactis strain. In a second part, development of quantification methods is described. It shows new stable isotope labeling approaches based on N- and C-terminus labeling of proteins and describes the first method of labeling of carboxylic groups at the protein level using 13C stable isotopes. In addition, a new quantitative approach called ANIBAL is explained that labels all amino and carboxylic groups at the protein level.

Influence of nutrition on urinary oxalate and calcium in preterm and term infants.

Few data for normal urinary oxalate (Ox) and calcium (Ca) excretion related both to gestational age and nutritional factors have been reported in preterm or term infants. We therefore determined the molar Ox and Ca to creatinine (Cr) ratios in spot urines from 64 preterm and 37 term infants aged 1-60 days, either fed formula or human milk (HM). Only vitamin D was supplemented; renal or metabolic diseases were excluded. Urinary Ox/Cr ratio was higher in preterm than in term infants, both when formula fed (1st month 253 vs. 180 mmol/mol and 2nd month 306 vs. 212 mmol/mol; P&lt;0.05) or HM fed (206 vs. 169 mmol/ mol and 283* vs. 232 mmol/mol; *P&lt;0.05). Ox/Cr was also higher in formula- than HM-fed preterm infants. The ratio increased during the first 2 months of life irrespective of nutrition. Urinary Ca/Cr ratio was comparable in all groups during the 1st month of life, except for a lower (P &lt; 0.05) value in term infants fed HM (0.10 mol/mol). It increased in all groups during the 2nd month of life, being highest in HM-fed preterm infants (1.86 mol/mol). In conclusion, urinary Ox and Ca excretion is influenced by both gestational age and nutrient intake in preterm and term infants.

Gene expression profiles in rat mesenteric lymph nodes upon supplementation with Conjugated Linoleic Acid during gestation and suckling

Background Diet plays a role on the development of the immune system, and polyunsaturated fatty acids can modulate the expression of a variety of genes. Human milk contains conjugated linoleic acid (CLA), a fatty acid that seems to contribute to immune development. Indeed, recent studies carried out in our group in suckling animals have shown that the immune function is enhanced after feeding them with an 80:20 isomer mix composed of c9,t11 and t10,c12 CLA. However, little work has been done on the effects of CLA on gene expression, and even less regarding immune system development in early life. Results The expression profile of mesenteric lymph nodes from animals supplemented with CLA during gestation and suckling through dam's milk (Group A) or by oral gavage (Group B), supplemented just during suckling (Group C) and control animals (Group D) was determined with the aid of the specific GeneChip® Rat Genome 230 2.0 (Affymettrix). Bioinformatics analyses were performed using the GeneSpring GX software package v10.0.2 and lead to the identification of 89 genes differentially expressed in all three dietary approaches. Generation of a biological association network evidenced several genes, such as connective tissue growth factor (Ctgf), tissue inhibitor of metalloproteinase 1 (Timp1), galanin (Gal), synaptotagmin 1 (Syt1), growth factor receptor bound protein 2 (Grb2), actin gamma 2 (Actg2) and smooth muscle alpha actin (Acta2), as highly interconnected nodes of the resulting network. Gene underexpression was confirmed by Real-Time RT-PCR. Conclusions Ctgf, Timp1, Gal and Syt1, among others, are genes modulated by CLA supplementation that may have a role on mucosal immune responses in early life.

Gene expression profiles in rat mesenteric lymph nodes upon supplementation with Conjugated Linoleic Acid during gestation and suckling

Background Diet plays a role on the development of the immune system, and polyunsaturated fatty acids can modulate the expression of a variety of genes. Human milk contains conjugated linoleic acid (CLA), a fatty acid that seems to contribute to immune development. Indeed, recent studies carried out in our group in suckling animals have shown that the immune function is enhanced after feeding them with an 80:20 isomer mix composed of c9,t11 and t10,c12 CLA. However, little work has been done on the effects of CLA on gene expression, and even less regarding immune system development in early life. Results The expression profile of mesenteric lymph nodes from animals supplemented with CLA during gestation and suckling through dam's milk (Group A) or by oral gavage (Group B), supplemented just during suckling (Group C) and control animals (Group D) was determined with the aid of the specific GeneChip® Rat Genome 230 2.0 (Affymettrix). Bioinformatics analyses were performed using the GeneSpring GX software package v10.0.2 and lead to the identification of 89 genes differentially expressed in all three dietary approaches. Generation of a biological association network evidenced several genes, such as connective tissue growth factor (Ctgf), tissue inhibitor of metalloproteinase 1 (Timp1), galanin (Gal), synaptotagmin 1 (Syt1), growth factor receptor bound protein 2 (Grb2), actin gamma 2 (Actg2) and smooth muscle alpha actin (Acta2), as highly interconnected nodes of the resulting network. Gene underexpression was confirmed by Real-Time RT-PCR. Conclusions Ctgf, Timp1, Gal and Syt1, among others, are genes modulated by CLA supplementation that may have a role on mucosal immune responses in early life.