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More than half of the world population is at risk of dengue virus (DENV) infection because of the global distribution of its mosquito vectors. There is neither effective vaccine nor therapeutics. The only available strategy relies on insecticides, against which mosquitoes are developing resistance. Viruses utilize the host metabolome for replication and dissemination. This is particularly true for envelope viruses like DENV that relies on host lipid membranes to complete their life-cycle. To reach an optimal metabolic environment, viruses subvert the host metabolome. Understanding DENV-mosquito metabolic interactions will reveal novel strategies to stop DENV transmission. Here, we characterized how DENV hijacks the Aedes aegypti mosquito lipidome to identify targets for novel transmission-blocking interventions. To describe metabolic changes throughout the mosquito DENV cycle, we deployed a Liquid chromatography-high resolution mass spectrometry (LC-HRMS) workflow at different stages of vector infection. We revealed a major phospholipid reconfiguration throughout the DENV mosquito cycle, in cells, midguts, and whole mosquitoes. To decipher how DENV reconfigures phospholipids, we phylogenetically characterized acylglycerolphosphate acyltransferase (AGPAT) enzyme isoforms and identified those (i.e., AGPAT1) that catalyze a central rate-limiting step in phospholipid biogenesis. We found that DENV infection decreased AGPAT1 expression, which depletion enhances infection by maintaining high aminophospholipid (aminoPL) concentrations, especially phosphatidylcholine (PC) and phosphatidylethanolamine (PE), during DENV mosquito cycle. By demonstrating that DENV-mediated AGPAT1 downregulation provides a proviral environment, these results reveal the first metabolic host factor in mosquitoes and emphasize the role of aminophospholipids in DENV cellular cycle. We then undertook to precise how DENV influences aminoPL biosynthesis and what stage of DENV cellular cycle requires aminoPL reconfiguration. De novo biosynthesis of PC and PE is known as the Kennedy pathway, where a diacylglycerol (DAG) incorporates either a choline or an ethanolamine group. AminoPL remodeling by deacylation/reacylation then ensures membrane dynamism that participates in membrane rearrangements. Using isotopic labelling through ethanolamine or choline supplementation, we showed that DENV modulates PC and PE biosynthesis by interacting with membrane remodeling.[...]; Plus de la moitié de la population mondiale est exposée au risque d'infection par le virus de la dengue (DENV) en raison de la distribution mondiale de ses moustiques vecteurs. Il n'existe ni vaccin ni traitement efficace. La seule stratégie disponible repose sur les insecticides, contre lesquels les moustiques développent une résistance. Les virus utilisent le métabolome de l'hôte pour la réplication et la dissémination. C'est particulièrement vrai pour les virus enveloppés comme le DENV qui dépend des membranes lipidiques de l'hôte pour compléter son cycle de vie. Pour atteindre un environnement métabolique optimal, les virus perturbent le métabolome de l'hôte. La compréhension de ces altérations chez les moustiques vecteurs pourrait révéler de nouvelles stratégies pour bloquer la transmission du DENV. Ici, nous avons caractérisé comment le DENV détourne le lipidome du moustique Aedes aegypti. Pour décrire les changements métaboliques tout au long du cycle du DENV chez le moustique, nous avons débeloppé une méthode de chromatographie liquide et de spectrométrie de masse à haute résolution (LC-HRMS) à différents stades de l'infection chez le vecteur. Nous avons révélé une reconfiguration majeure des phospholipides tout au long du cycle du DENV chez le moustique, dans les cellules, l'intestin moyen et le moustique entier. Pour déchiffrer la façon dont le virus reconfigure les phospholipides, nous avons caractérisé phylogénétiquement les isoformes de l'enzyme acylglycerol-phosphate acyltransférase (AGPAT) et identifié celles qui catalysent une étape limitante dans la biogenèse des phospholipides. Nous avons constaté que l'infection par le DENV diminuait l'expression de AGPAT1, dont la déplétion renforce l'infection en maintenant des concentrations élevées d'aminophospholipides (aminoPL), en particulier la phosphatidylcholine (PC) et la phosphatidyléthanolamine (PE), pendant le cycle du DENV chez le moustique. En démontrant que la sous-régulation de AGPAT1, causé par le virus, fournit un environnement proviral, nous révèlons le premier facteur métabolique hôte chez les moustiques et soulignent le rôle des aminophospholipides dans le cycle cellulaire viral. Nous avons ensuite cherché à confirmer que le virus influence la biosynthèse des aminoPL et déterminer à quel stade du cycle viral la reconfiguration des aminoPL est nécessaire. La biosynthèse de novo de PC et de PE est connue sous le nom de voie de Kennedy, où un diacylglycérol (DAG) incorpore soit un groupe choline, soit un groupe éthanolamine. [...]

Scope Enhanced adiposity and metabolic inflammation are major features of obesity associated with altered gut microbiota and intestinal barrier. How these metabolic outcomes can be impacted by milk polar lipids (MPL), naturally containing 25% of sphingomyelin, is investigated in mice fed a mixed high-fat (HF) diet . Methods and results Male C57Bl/6 mice receive a HF-diet devoid of MPL (21% fat, mainly palm oil, in chow), or supplemented with 1.1% or 1.6% of MPL (HF-MPL1; HF-MPL2) via a total-lipid extract from butterserum concentrate for 8 weeks. HF-MPL2 mice gain less weight versus HF (p < 0.01). Diets do not impact plasma markers of inflammation but in the liver, HF-MPL2 tends to decrease hepatic gene expression of macrophage marker F4/80 versus HF-MPL1 (p = 0.06). Colonic crypt depth is the maximum in HF-MPL2 (p < 0.05). In cecal microbiota, HF-MPL1 increases Bifidobacterium animalis versus HF (p < 0.05). HF-MPL2 decreases Lactobacillus reuteri (p < 0.05), which correlates negatively with the fecal loss of milk sphingomyelin-specific fatty acids (p < 0.05). Conclusion In mice fed a mixed HF diet, MPL can limit HF-induced body weight gain and modulate gut physiology and the abundance in microbiota of bacteria of metabolic interest. This supports further exploration of how residual unabsorbed lipids reaching the colon can impact HF-induced metabolic disorders.

Open access original paper Source : 10th EPR Workshop on Applications of EPR in Biology and Medicine, 2016 in Krakow, POLAND.; International audience; The mammalian retina contains a high level of polyunsaturated fatty acids, including docosahexaenoic acid (22:6) (DHA), which are highly susceptible to oxidation. It has been shown that one of the products of DHA oxidation-carboxyethylpyrrole (CEP), generated in situ, causes modifications of retinal proteins and induces inflammation response in the outer retina. These contributing factors may play a role in the development of age-related macular degeneration (AMD). It is also possible that some of the lipid oxidation products are photoreactive, and upon irradiation with blue light may generate reactive oxygen species. Therefore, in this work we analysed oxidation-induced changes in photoreactivity of lipids extracted from bovine neural retinas. Lipid composition of bovine neural retinas closely resembles that of human retinas making the bovine tissue a convenient model for studying the photoreactivity and potential phototoxicity of oxidized human retinal lipids. Lipid composition of bovine neural retinas Folch' extracts (BRex) was determined by gas chromatography (GC) and liquid chromatography coupled to an electrospray ionization source-mass spectrometer (LC-ESI-MS) analysis. Liposomes prepared from BRex, equilibrated with air, were oxidized in the dark at 37 A degrees C for up to 400 h. The photoreactivity of BRex at different stages of oxidation was studied by EPR-oximetry and EPR-spin trapping. Photogeneration of singlet oxygen (O-1(2), (1)Delta(g)) by BRex was measured using time-resolved detection of the characteristic phosphorescence at 1270 nm. To establish contribution of lipid components to the analysed photoreactivity of Folch' extract of bovine retinas, a mixture of selected synthetic lipids in percent by weight (w/w %) ratio resembling that of the BRex has been also studied. Folch's extraction of bovine neural retinas was very susceptible to oxidation despite the presence of powerful endogenous antioxidants such as alpha-tocopherol and zeaxanthin. Non-oxidized and oxidized BRex photogenerated singlet oxygen with moderate quantum yield. Blue-light induced generation of superoxide anion by Folch' extract of bovine neural retinas strongly depended on the oxidation time. The observed photoreactivity of the studied extract gradually increased during its in vitro oxidation.

International audience; In many liposome applications, the nanomechanical properties of the membrane envelope are essential to ensure, e.g., physical stability, protection, or penetration into tissues. Of all factors, the lipid composition and its phase behavior are susceptible to tune the mechanical properties of membranes. To investigate this, small unilamellar vesicles (SUV; diameter < 200 nm), referred to as liposomes, were produced using either unsaturated 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) or saturated 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) in aqueous buffer at pH 6.7. The respective melting temperatures of these phospholipids were −20 and 41 °C. X-ray diffraction analysis confirmed that at 20 °C DOPC was in the fluid phase and DPPC was in the gel phase. After adsorption of the liposomes onto flat silicon substrates, atomic force microscopy (AFM) was used to image and probe the mechanical properties of the liposome membrane. The resulting force–distance curves were treated using an analytical model based on the shell theory to yield the Young’s modulus (E) and the bending rigidity (kC) of the curved membranes. The mechanical investigation showed that DPPC membranes were much stiffer (E = 116 ± 45 MPa) than those of DOPC (E = 13 ± 9 MPa) at 20 °C. The study demonstrates that the employed methodology allows discrimination of the respective properties of gel- or fluid-phase membranes when in the shape of liposomes. It opens perspectives to map the mechanical properties of liposomes containing both fluid and gel phases or of biological systems.

This chapter focuses on protein-based and lipid-based native nanostructures. The naturally occurring nanostructures coming from milk and eggs constitute a large part of the building blocks already used to design new nanostructures. The chapter also focuses on the description of the structure of several food proteins as they exist naturally in food. Food proteins exist mainly in the monomeric form, but are able to self-assemble into oligomers or aggregates in some specific conditions. These conditions are also addressed. The lipid droplets are spherical particles constituted by a hydrophobic core rich in triacylglycerols (TAG) covered by surfaceactive molecules such as phospholipids and proteins. These TAG-rich droplets are involved in the storage and delivery of energy. Milk fat globules have a core of TAG covered by a biological membrane composed by a trilayer of phospholipids and proteins.

Metabolic diseases are characterized by an altered lipid metabolism and metabolic inflammation. Numerous food products contain polar lipid (PL) emulsifiers that could impact these risk factors. We evaluated the impact of using PL from milk (MPL) (i) acutely on lipid digestion and postprandial lipemia and (ii) in the longer term in addition to a high fat diet on adiposity and adipose tissue inflammation. We compared MPL to soybean PL (SPL) that is currently the main commercial source of PL.In mice, an emulsion stabilized by MPL results in a more rapid postprandial lipemia than an emulsion stabilized by SPL, with an early increase in lipemia and a faster clearance. Differences in lipemia can originate from differential kinetics of lipid hydrolysis in the mouse gut, as an increase intestinal TG hydrolysis is observed in vitro. Moreover, early MPL-derived chylomicrons are smaller than SPL-derived chylomicrons. In the longer term, compared with HF diet, HF-SPL diet increases hepatic lipids, white adipose tissue (WAT) mass, with larger and more numerous adipocytes and increases expression of pro-inflammatory adipokines. This is not observed with HF-MPL diet despite similar dietary intakes. HFP-MPL mice have a lower expression in WAT of marker of macrophage infiltration and more numerous goblet cells in the colon, suggesting an improved gut barrier function with this diet.Postprandial lipemia in mice can be modulated by emulsifying with MPL compared with SPL, partly through differences in chylomicron assembly, and intestinal TG hydrolysis rate. Moreover unlike SPL, MPL in a high fat diet do not induce WAT hypertrophy and inflammation, Les maladies métaboliques d’origine nutritionnelle sont caractérisées par un métabolisme des lipides perturbé et une inflammation métabolique. Les lipides polaires (LP) sont des agents émulsifiants utilisés dans l’industrie agroalimentaire. Le but de notre étude a été d’évaluer l’impact de l’utilisation de nouveaux LP issus du lait sur (i) la digestion et le métabolisme postprandial des lipides, et (ii) à plus long terme sur l’adiposité et l’inflammation dans le tissu adipeux (TA), en comparaison avec les LP de soja, actuellement les plus consommés.Chez la souris, les LP laitiers induisent une cinétique de lipémie postprandiale plus précoce en comparaison avec les LP de soja, associée à une augmentation de la lipolyse intestinale in vitro. De plus, les chylomicrons sécrétés durant le pic de lipémie sont plus petits avec les LP laitiers. D’autre part, la substitution d’une partie des lipides d’un régime hyper-lipidique par des LP laitiers ne modifie pas le stockage des lipides dans le foie et le TA contrairement aux LP de soja qui induisent une augmentation des lipides hépatiques, des adipocytes et des marqueurs inflammatoires dans le TA. Par ailleurs, Les LP laitiers induisent une diminution de l’expression génique de marqueurs de l’infiltration macrophagique dans le TA et du nombre de cellules à gobelet dans le côlon, suggérant une barrière intestinale renforcée.Ces travaux démontrent que les LP laitiers, par rapport aux LP de soja, stimulent la digestion des lipides et induisent une cinétique plus rapide de lipémie postprandiale. A long terme ils n’induisent pas les altérations métaboliques du TA observées en présence de LP de soja dans un régime hyper-lipidique

Retinal cell membranes are known to be rich in ether-linked phospholipids also termed as “plasmalogens”. Although their functions remain enigmatic, plasmalogens are considered as reservoirs of polyunsaturated fatty acids (PUFAs). These PUFAs were shown to be released from plasmalogens by specific enzymes under specific conditions in order to play several key functions within cells. Data from our laboratory suggest that the metabolism of plasmalogens and PUFAs may be implicated in the pathophysiology of glaucoma, the leading cause of blindness worldwide. Through several experiments on human patients and rodent models of ocular hypertony, we have shown that the metabolism of plasmalogens and PUFAs is affected in patients with primary open-angle glaucoma (POAG) in a stage-dependent manner, that the modifications in plasmalogen metabolism may start years prior the first clinical symptoms of POAG, and that several combinations of dietary PUFAs could be efficient in preventing retinal glaucomatous damage in rodents.

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Milk phospholipids and lipids have been associated with modulation of membrane properties and intestinal inflammation, but less is known on their effects on the development of gut physiology and immune system in neonates. Two infant formulas differing by the presence of bovine milk phospholipids and lipids (CRTL: a standard-based formula adapted to neonatal pig requirement; PLUS: CRTL plus phospholipids and lipids) were distributed with an automatic milk feeder to 36 piglets during the nursing period (i.e. 28 days). The residual immunoreactivity of β-lactoglobulin and caseins present in the digestive compartments was determined by ELISA. The intestinal permeability was evaluated using Ussing chambers. Mononuclear cells were isolated from mesenteric lymph nodes (MLN) and cultured in presence of a mitogen to evaluate their cytokine pattern. Physiological data were compared to previous data obtained on same aged-sow suckling piglets. Moreover, immunoregulatory capacity of peptide and lipid fractions extracted from jejunal digestas was evaluated in vitro on mitogen-stimulated MLN cells from sow suckling 7 days-old piglets. The immunoreactivity of β-lactoglobulin and caseins was higher in PLUS than CRTL digestas both in jejunum and ileum. Higher intestinal thickness observed in PLUS group was not associated with changes in permeability. The reduced mononuclear cell density in MLN of PLUS-fed piglets indicated a modification of the mobilization or recruitment of MLN cells. PLUS formula also increased IFNγ secretion suggesting an improved intestinal immune system maturation such as observed in sow suckling piglets. Finally, a major immunosuppressor effect of the lipid fractions was evidenced with both formulas. However, the less severe inhibitor activity of lipids extracted from PLUS digesta may participate in the in vivo increased IFNγ secretion in PLUS formula-fed piglets. In conclusion, introduction of milk phospholipids and lipids in infant formula induced modifications of the intestinal immune system maturation that may promote intestinal immune system maturation similar to sow suckling piglets.

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Les données récentes montrent que les bénéfices nutritionnels associés aux acides gras polyinsaturés n-3 dépendent du devenir des structures des lipides et de leur oxydation au cours de la digestion. L’objectif de cette thèse était de préciser l’importance de paramètres liés à la composition des milieux de digestion sur les évolutions des structures des lipides émulsifiés, la bioaccessibilité des acides gras, et leur oxydation. Après la mise au point de conditions de digestion in vitro basées sur l’analyse de la bibliographie et prenant en compte des contraintes liées aux échanges gazeux, des digestions gastro-intestinales ont été effectuées sur des émulsions stabilisées par une protéine ou des tensioactifs. Les résultats montrent des effets contrastés des conditions gastriques, de la nature de l’émulsifiant, de la composition en acides gras et en tocophérols des huiles et de la présence de metmyoglobine sur la floculation et la coalescence des émulsions et sur les cinétiques de lipolyse intestinale et d’oxydation. L’importance de la balance entre pro et antioxydants, de l’insaturation et de la longueur de chaîne des acides gras, ainsi que de la présence des sucs digestifs ont ainsi été démontrés sur l’oxydation des acides gras selon la nature de l’émulsifiant et l’état de dispersion des lipides lors de leur arrivée en phase intestinale. Ainsi, le malonaldéhyde, les 4-HNE et 4-HHE sont formés pendant les phases gastrique et intestinale. Les données sur l’oxydation in vitro ont été comparées avec des résultats obtenus in vivo dans le contenu digestif et dans le plasma chez la souris., Recent data show that the nutritional benefits associated to the consumption of n-3 polyunsaturated fatty acids depend on the fate of lipid structures, including lipid oxidation, during digestion. The aim of this work was to determine the influence of the composition of digestion media on the evolution of the structures of emulsified lipids and the bioaccessibility and the oxidation of fatty acids. In vitro gastrointestinal digestion conditions, based on analysis of the literature and taking into account the constraints related to gas exchanges, were first set up and then used to digest emulsions stabilized by a protein or by surfactants. Interestingly, malonaldehyde, 4-HNE and 4-HHE were formed during both the gastric and intestinal phases. The results show contrasting effects of the gastric conditions, the emulsifier, the presence of metmyoglobin, and the fatty acid and tocopherols compositions of the oils on flocculation and coalescence of the emulsions, kinetics of lipolysis during the intestinal phase, and oxidation. The importance on oxidation of the balance between pro- and anti-oxidants, unsaturation and chain length of the fatty acids has been also confirmed. Depending on the nature of the emulsifier and the dispersion state of lipids at the beginning of intestinal phase the digestive juices exhibit or not protective effect on oxidation. The in vitro oxidation data were compared with results obtained in vivo in the gut contents and plasma of mice.

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Diplôme : Dr. d'Université

Pour leur apport energetique et les qualites organoleptiques qu'ils conferent, les lipides alimentaires sont consommes dans des produits de compositions et de structures variables. Leur digestion et leur assimilation par l'organisme sont des processus complexes qui ne sont pas totalement elucides. Cet article fait le bilan des parametres physico-chimiques et technologiques qui peuvent affecter la cinetique de vidange gastrique, la digestion et l'absorption des lipides du lait ainsi que les consequences metaboliques. Au-dela de la composition en acides gras, la structure moleculaire des triglycerides doit etre determinee. De plus, la suprastructure des lipides (emulsifies dans des gouttelettes de differentes tailles, non emulsifies) qui affecte les cinetiques d'hydrolyse des triglycerides par les enzymes digestives doit etre caracterisee. Le role de la membrane des globules gras, et de ses composes bioactifs comme les phospholipides, doit etre elucide. Enfin, l'assimilation des lipides peut etre conditionnee par les autres constituants des produits laitiers tels que le calcium et les proteines. Face aux faibles donnees de la litterature, nous proposons des travaux de recherche qui permettraient, en utilisant des matrices de compositions et de structures maitrisees, d'ameliorer la connaissance des mecanismes impliques dans la digestion et l'absorption des lipides du lait. L'enjeu est d'apporter des elements scientifiques concrets et des leviers technologiques permettant aux industriels d'elaborer de nouvelles strategies pour moduler la biodisponibilite des lipides et ainsi contribuer au concept d'aliments nutri-fonctionnels.

The chemical composition and properties of lipids, both triglycerides and phospholipids, play a major role in the functional and nutritional properties of food products. In this study, the suprastructure of fat, solid fat content, and crystallographic properties of triglycerides were investigated in hard-type cheeses from the microscopic scale to the molecular level using the combination of relevant techniques. Two industrial cheeses with different oiling off properties were compared with experimental cheeses manufactured in the laboratory. Microstructural analysis performed using confocal laser scanning microscopy showed that milk processing led to the disruption of fat globules with the formation of nonglobular fat. For a similar fatty acid composition, oiling off was mainly related to the fat in dry matter content and to the suprastructure of fat in cheese. An exogenous fluorescent phospholipid permitted the localization of milk phospholipids in the cheese matrix, which mainly remain around fat inclusions after disruption of the milk fat globule membrane, and to show heterogeneities. We also showed using differential scanning calorimetry that the suprastructure of fat did not affect the solid fat content in cheese at 4 degrees C: 71.6 +/- 4.9%. The organization of triglyceride molecules in fat crystals, elucidated at a molecular level using X-ray diffraction, corresponded to the coexistence of 2 lamellar structures (2L 40.5 angstroms and 3L 54.6 angstroms) with four polymorphic forms: alpha, two beta' and beta. A schematic representation of the multiscale organization of triglycerides and phospholipids in cheese is proposed.

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Ce rapport de DEA décrit les caractéristiques biochimiques (compositions, distributions) des lipoprotéines de la truite arc-en-ciel, modèle en nutrition des poisson.

International audience