EPA prevents insulin resistance and glucose intolerance in mice fed a high fat-high sucrose diet

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Supplémentation en AGPI n-3 sur 3 génération, intérêt vis-à-vis du développement de l’obésité et des troubles métaboliques associés

Plusieurs études in vitro ont permis de mettre en évidence la bioactivité des acides gras polyinsaturés oméga 3 (AGPI n-3) sur la cellule adipeuse ou musculaire pour lutter contre les effets des régimes occidentaux et de l’obésité. Les études d’interventions, notamment chez l’Homme sont d’une durée insuffisante pour mettre en évidence des effets métaboliques, les mécanismes impliquant potentiellement l’impression d’empreintes épigénétiques et/ou la régulation très précoce. Cette étude avait pour but d’explorer l’intérêt transgenerationnel d’une supplémentation en EPA (1% du poids du régime) sur 3 générations chez la souris vis-à-vis de la résistance à un challenge nutritionnel. Des mâles (4/6 mois) de chaque génération ont été soumis à un challenge de type high fat+sucrose (sans AGPI) durant 16 semaines. Nous avons suivi le poids et évaluer la tolérance au glucose, la sensibilité à l’insuline des animaux. La réponse des générations 1 et 2 est comparable entre le groupe témoin non supplémenté et le groupe ayant reçu l’EPA. En revanche, les animaux de la 3ème génération prennent moins de poids, pour une prise alimentaire similaire après normalisation par le poids des animaux. La tolérance à l’insuline et au glucose des animaux est aussi partiellement restaurée

L’EPA prévient l’insulino-résistance et l’intolérance au glucose chez la souris sous régime hyperlipidique et hypersucré

L’insulino-résistance (IR) est une conséquence majeure de l’obésité. Elle favorise la progression du syndrome métabolique (SMet) et augmente le risque de diabète de type 2 et de maladies cardiovasculaires. (...

Le DHA restore la sensibilité musculaire à l’insuline en inhibant l’activation de la PKC-theta par les céramides

International audienceIntroduction et but de l’étude Les métabolismes lipidiques et énergétiques sont intrinsèquement liés et jouent un rôle central dans l’apparition de la résistance à l’insuline dans le muscle squelettique. Ce phénomène implique notamment le stress oxydant, l’inflammation et/ou une lipotoxicité qui sont eux-mêmes influencés par le degré de saturation et les taux cellulaires et circulant d’acides gras et de leurs métabolites. Il est bien connu que les acides gras saturés (AGS) favorisent la résistance à l’insuline. En s’opposant à certains effets des AGS, les acides gras polyinsaturés n-3 à longue chaîne (AGPI n-3-LC) seraient protecteurs contre l’insulinorésistance. Les relations entre les mécanismes de transport des acides gras, la signalisation de l’insuline et le métabolisme musculaire sont mal connues. Matériel et méthodes Nous avons analysé les effets d’un AGPI n-3-LC, le DHA (30 μM)sur la cellule musculaire de type C2C12 traitée au palmitate (500 μM).La sensibilité à l’insuline a été évaluée par la quantification de la phosphorylation de la protéine Akt (Western-Blot) et la captation de glucose (Glucose uptake cell-based assay, Cayman). La lipotoxicité a été quantifié par la mesure des taux d’acyl carnitines et de céramides (spectrométrie de masse), la phoshorylation de la PKC-9 (Western-Blot) et l’expression de cytokines pro-inflammatoire (RT-qPCR). Résultats et Analyse statistique Le DHA s’oppose aux effets délétères du palmitate vis à vis de la sensibilité à l’insuline des myotubes C2C12. Il permet de normaliser la signalisation cellulaire et la captation de glucose. L’élévation du taux cellulaire en céramides est associée à une activation de la PKC-9,. À l’inverse, l’inhibition de la synthèse des céramides par la myriocine atténue cet effet et restaure partiellement l’insulinosensibilité. Le profil cellulaire en acyl carnitines est modifié en présence de palmitate suggérant une moindre efficacité de la [3-oxydation. Ce profil est restauré en présence de DHA, suggérant une normalisation du fonctionnement de la (3-oxydation par cet AGPI n-3-LC. Ce résultat est confirmé par une amélioration des taux d’ARNm des Cpt1a et 1b. En outre, le DHA réduit les niveaux d’expression des médiateurs pro-inflammatoires TNFα, IL6 et Ptgs2 qui sont fortement augmentés par le palmitate. Conclusion Nous montrons la première fois un effet bénéfique du DHA sur l’insulinosensibilité musculaire à une dose physiologique. Cet AG régule favorablement la lipotoxicité induite par un AGS en régulant les teneurs cellulaires en céramides et en réprimant les processus inflammatoires et l’activation de la PKC-θ. Ces effets sont associés à des signes de meilleur fonctionnement de la (3-oxydation des acides gras

Discriminating between the activities of human cathepsin G and chymase using fluorogenic substrates

Cathepsin G (CG) (EC 3.4.21.20) and chymase (EC 3.4.21.39) are two closely-related chymotrypsin-like proteases that are released from cytoplasmic granules of activated mast cells and/or neutrophils. We investigated the potential for their substrate-binding subsites to discriminate between their substrate specificities, aiming to better understand their respective role during the progression of inflammatory diseases. In addition to their preference for large aromatic residues at P1, both preferentially accommodate small hydrophilic residues at the S1' subsite. Despite significant structural differences in the S2' subsite, both prefer an acidic residue at that position. The Ala226/Glu substitution at the bottom of the CG S1 pocket, which allows CG but not chymase to accommodate a Lys residue at P1, is the main structural difference, allowing discrimination between the activities of these two proteases. However, a Lys at P1 is accommodated much less efficiently than a Phe, and the corresponding substrate is cleaved by ?2-tryptase (EC 3.4.21.59).?We optimized a P1 Lys-containing substrate to enhance sensitivity towards CG and prevent cleavage by chymase and ?2-tryptase. The resulting substrate (ABZ-GIEPKSDPMPEQ-EDDnp) [where ABZ is O-aminobenzoic acid and EDDnp is N-(2,4-dinitrophenyl)-ethylenediamine] was cleaved by CG but not by chymase and tryptase, with a specificity constant of 190?mm(-1) ·s(-1) . This allows the quantification of active CG in cells or tissue extracts where it may be present together with chymase and tryptase, as we have shown using a HMC-1 cell homogenate and a sputum sample from a patient with severe asthma

A substrate-based approach to convert SerpinB1 into a specific inhibitor of proteinase 3, the Wegener's granulomatosis autoantigen.

International audienceThe physiological and pathological functions of proteinase 3 (PR3) are not well understood due to its close similarity to human neutrophil elastase (HNE) and the lack of a specific inhibitor. Based on structural analysis of the active sites of PR3 and HNE, we generated mutants derived from the polyvalent inhibitor SerpinB1 (monocyte/neutrophil elastase inhibitor) that specifically inhibit PR3 and that differ from wt-SerpinB1 by only 3 or 4 residues in the reactive center loop. The rate constant of association between the best SerpinB1 mutant and PR3 is 1.4 × 10⁷ M⁻¹ * s⁻¹, which is ∼100-fold higher than that observed with wt-SerpinB1 and compares with that of α1-protease inhibitor (α1-PI) toward HNE. SerpinB1(S/DAR) is cleaved by HNE, but due to differences in rate, inhibition of PR3 by SerpinB1(S/DAR) is only minimally affected by the presence of HNE even when the latter is in excess. SerpinB1(S/DAR) inhibits soluble PR3 and also membrane-bound PR3 at the surface of activated neutrophils. Moreover, SerpinB1(S/DAR) clears induced PR3 from the surface of activated neutrophils. Overall, these specific inhibitors of PR3 will be valuable for defining biological functions of the protease and may prove useful as therapeutics for PR3-related inflammatory diseases, such as Wegener's granulomatosis

EPA prevents fat mass expansion and metabolic disturbances in mice fed with a Western diet

The authors acknowledge the staff from Auvergne University Experimental Animal Laboratory and Christophe Del’Homme, Philippe Denis, Anne Terisse-Lottier and Alexandre Teynie from the Experimental Animal Facility of the Human Nutrition Unit (INRA of Clermont-Ferrand) for their assistance throughout the animal protocol. We thank Celine Bobby for her help for TaqMan gene expression assays. We gratefully acknowledge financial support and doctoral fellowship (to A.P.) fromLesieur and AvrilImpact of ALA, EPA and DHA on obesity and metabolic complications were studied in mice fed a high-fat high-sucrose diet (HF). HF diets were supplemented with ALA, EPA or DHA (1%w/w) and given to C57BL/6J mice for 16 weeks and to Ob/Ob mice for 6 weeks. In C57BL/6J mice, EPA reduced plasma cholesterol (-20%), limited fat mass accumulation (-23%), adipose cell hypertrophy (-50%), and reduced plasma leptin concentration (-60%), compared to HF fed mice. Furthermore, mice supplemented with EPA exhibited a higher insulin sensitivity (+24%) and glucose tolerance (+20%) compared to HF fed mice. Similar effects were observed in EPA supplemented Ob/Ob mice, although fat mass accumulation was not prevented. By contrast in comparison to HF fed mice, DHA did not prevent fat mass accumulation, increased plasma leptin concentration (+128%) in C57BL/6J mice and did not improve glucose homeostasis in C57BL/6J and Ob/Ob mice. In 3T3-L1 adipocytes, DHA stimulated leptin expression whereas EPA induced adiponectin expression, suggesting that improved leptin / adiponectin balance may contribute to the protective effect of EPA. In conclusion, supplementation with EPA, but not ALA and DHA, could preserve glucose homeostasis in an obesogenic environment and limit fat mass accumulation in the early stage of weight gain

Effect of Canola oil enrichment with microconsituants against metabolic disorders

Aim/hypothesis: Insulin resistance (IR) favors the progression of metabolicsyndrome (MetS) and increases the risk of type2 diabetes. IR results from metabolic dysfunctions,oxidative stress and inflammation caused by ectopic fat depots. We studied the effect of canola oil enriched with micronutrients naturally present in canola seed on IR and MetS during a high fat (HF)-challenge]. Research design and Methods: Rats were fed with a HF diet containing 30% of lipids, mainly derived from palm oil (diet P) or from a mixture of palm and canola oils (60:40, diet C). In 2 groups of animals, diet C was further supplemented with alpha tocopherol, CoQ with or without Canolol (diets CMC & CM respectively). Insulin sensitivity and glucose tolerance were assessed after 10 weeks. Tissue sampling was carried at sacrifice for gene expression analysis (RT-qPCR), protein expression level (Western-Blot), fatty acid composition (Gas Chromatography). HF animals were compared to control animals receiving a standard diet. ANOVA was performed for statistical analysis (followed by adjusted Fisher post-hoc test when p<0.05).Results: All groups receiving a HF diet gained significantly more weight and accumulated fat mass compared to control (No difference between all HF groups). Glucose tolerance was altered in P and C groups and resored in CM and CMC. Insulin sensitivity and muscle Akt activition were not different between control and P groups, but were increased in CMC group compared to controls. No marked changes in plasma antioxidant defense were detected. A reduction in PDK4 and Srebp1c mRNA levels was observed in skeletal muscle in CM group.Conclusions: The proportion of AGPI could not prevent metabolic disturbance induced by a HF meal. Enrichment with natural micronutrients could have a small but beneficial effect against insulin resistance. These effects might be linked to a better glucose oxidation in skeletal muscl

Association citrulline-statine : un traitement innovant pour l’obésité et le diabète de type 2

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Early sepsis does not stimulate reactive oxygen species production and does not reduce cardiac function despite an increased inflammation status

This work was supported by 2 grants attributed by the Fédération Française de Cardiologie and society ADETEC of patients with a previous cardiac surgeryIf it is sustained for several days, sepsis can trigger severe abnormalities of cardiac function which leads to death in 50% of cases. This probably occurs through activation of toll‐like receptor‐9 by bacterial lipopolysaccharides and overproduction of proinflammatory cytokines such as TNF‐α and IL‐1β. In contrast, early sepsis is characterized by the development of tachycardia. This study aimed at determining the early changes in the cardiac function during sepsis and at finding the mechanism responsible for the observed changes. Sixty male Wistar rats were randomly assigned to two groups, the first one being made septic by cecal ligation and puncture (sepsis group) and the second one being subjected to the same surgery without cecal ligation and puncture (sham‐operated group). The cardiac function was assessed in vivo and ex vivo in standard conditions. Several parameters involved in the oxidative stress and inflammation were determined in the plasma and heart. As evidenced by the plasma level of TNF‐α and gene expression of IL‐1β and TNF‐α in the heart, inflammation was developed in the sepsis group. The cardiac function was also slightly stimulated by sepsis in the in vivo and ex vivo situations. This was associated with unchanged levels of oxidative stress, but several parameters indicated a lower cardiac production of reactive oxygen species in the septic group. In conclusion, despite the development of inflammation, early sepsis did not increase reactive oxygen species production and did not reduce myocardial function. The depressant effect of TNF‐α and IL‐1β on the cardiac function is known to occur at very high concentrations. The influence of low‐ to moderate‐grade inflammation on the myocardial mechanical behavior must thus be revisited