A Model of Insulin Resistance in Mice, Born to Diabetic Pregnancy, Is Associated with Alterations of Transcription-Related Genes in Pancreas and Epididymal Adipose Tissue

Objective. This study is conducted on a model of insulin-resistant (IR) mice born to dams which were rendered diabetic by the administration of streptozotocin. Methods. Adult IR and control offspring were selected and we determined the mRNA expression of transcription factors known to modulate pancreatic and adipose tissue activities and inflammation. Results. We observed that serum insulin increased, and the mRNA of insulin gene transcription factors, Pdx-1, Nkx6.1 and Maf-A, were upregulated in IR mice pancreas. Besides, their pancreatic functional capacity seemed to be exhausted as evidenced by low expression of pancreatic Glut2 and glucokinase mRNA. Though IR offspring exhibited reduced epididymal adipose tissue, their adipocytes seemed to be differentiated into macrophage-like cells, as they exhibited upregulated CD14 and CD68 antigens, generally expressed by macrophages. However, there was no peripheral macrophages infiltration into epididymal adipose tissue, as the expression of F4/80, a true macrophage marker, was undetectable. Furthermore, the expression of IL-6, TNF-α and TLR-2, key players of insulin resistance, was upregulated in the adipose tissue of IR offspring. Conclusion. Insulin resistant state in mice, born to diabetic pregnancy, alters the expression of function-related genes in pancreas and epididymal adipose tissue and these offspring are prone to develop metabolic syndrome

Thapsigargin-stimulated MAP kinase phosphorylation via CRAC channels and PLD activation: inhibitory action of docosahexaenoic acid

AbstractThis study was conducted on human Jurkat T-cells to investigate the role of depletion of intracellular Ca2+ stores in the phosphorylation of two mitogen-activated protein kinases (MAPKs), i.e. extracellular signal-regulated kinase (ERK) 1 and ERK2, and their modulation by a polyunsaturated fatty acid, docosahexaenoic acid (DHA). We observed that thapsigargin (TG) stimulated MAPK activation by store-operated calcium (SOC) influx via opening of calcium release-activated calcium (CRAC) channels as tyrphostin-A9, a CRAC channel blocker, and two SOC influx inhibitors, econazole and SKF-96365, diminished the action of the former. TG-stimulated ERK1/ERK2 phosphorylation was also diminished in buffer containing EGTA, a calcium chelator, further suggesting the implication of calcium influx in MAPK activation in these cells. Moreover, TG stimulated the production of diacylglycerol (DAG) by activating phospholipase D (PLD) as propranolol (PROP) (a PLD inhibitor), but not U73122 (a phospholipase C inhibitor), inhibited TG-evoked DAG production in these cells. DAG production and protein kinase C (PKC) activation were involved upstream of MAPK activation as PROP and GF109203X, a PKC inhibitor, abolished the action of TG on ERK1/ERK2 phosphorylation. Furthermore, DHA seems to act by inhibiting PKC activation as this fatty acid diminished TG- and phorbol 12-myristate 13-acetate-induced ERK1/ERK2 phosphorylation in these cells. Together these results suggest that Ca2+ influx via CRAC channels is implicated in PLD/PKC/MAPK activation which may be a target of physiological agents such as DHA

Implication de la signalisation calcique et des MAP kinases dans la perception gustative lipidique

Dans ce travail, nous démontrons que STIM1, un senseur calcique activé par la déplétion du Ca2+ intracellulaire du réticulum endoplasmique, est indispensable pour la signalisation calcique et la préférence oro-sensorielle du gras. Nous observons que l'acide linoléique (LA), en activant les phospholipases A2 via CD36, produit de l acide arachidonique (AA) et de la lyso-phosphatidylcholine (lyso-PC). Cette activation déclenche un influx calcique dans les cellules CD36-positives, et induit la production du facteur CIF (Ca2+ Influx Factor). CIF, AA et lyso-PC exercent différentes actions sur l'ouverture des canaux SOC (Stored Operated Calcium Channel) constitués de protéines Orai et contrôlés par STIM1. Par ailleurs, les souris au phénotype Stim1-/- perdent la préférence spontanée pour les lipides et la libération de la sérotonine à partir des cellules gustatives dans le milieu extracellulaire chez les animaux sauvages. Nous demontrons aussi que la signalisation calcique médiée via CD36 est doublement modulée lors de l obésité. L augmentation de la [Ca2+]i dans les cellules gustatives observée chez le Psammomys obesus, un modèle d obésité nutritionelle, est fortement diminuée chez les souris rendues obèses par un regime hyperlipidique. Nous avons constaté également que l interaction de LA avec le CD36 induit l activation des MAP Kinases de la voie MEK1/2/ERK1/2/Elk-1 qui est non seulement à l origine de l activation des aires cérébrales telles que le NTS, le noyau arqué, l hippocampe mais aussi indispensable pour la préférence spontanée pour les lipides alimentaires. Nos résultats suggèrent pour la prémière fois, que la voie ERK1/2 des MAPK et la signalisation calcique lipidique controlée par STIM1 sont impliquées dans la perception oro-gustative des lipidesIn this work, we demonstrate that stromal interaction molecule 1 (STIM1), a sensor of Ca2+ depletion in the endoplasmic reticulum, mediates fatty acid induced Ca2+ signaling in the mouse tongue and fat preference. We showed that linoleic acid (LA) induced the production of arachidonic acid (AA) and lysophosphatidylcholine (Lyso-PC) by activating multiple phospholipase A2 isoforms via CD36. This activation triggered Ca2+ influx in lingual CD36-positive taste bud cells (TBCs) purified from mouse CVP. LA also induced the production of Ca2+ influx factor (CIF). STIM1 was found to regulate LA-induced CIF production and the opening of store-operated Ca2+ (SOC) channels. Furthermore, CD36-positive TBCs from Stim1 / mice failed to release serotonin, and Stim1 / mice lost the spontaneous preference for fat that was observed in wild-type animals. We also demonstrate that the calcium-mediated signaling via CD36 is doubly modulated in obesity. The increase in [Ca2+]i in taste bud cells observed in Psammomys obesus, a model of nutritional obesity is strongly reduced in diet-induced obese (DIO) mice. We also found that the interaction of LA with CD36 induces activation of MAP Kinases MEK1/2/ERK1/2/Elk-1 pathway that is not only responsible for the activation of NTS, arcuate nucleus, and the hippocampus in the brain but also essential for the spontaneous preference for fat food. Our results suggest for the first time, that ERK1/2 MAPK pathway and lipid-induced calcium signaling controlled by STIM1 are involved in oro-gustatory perception of dietary lipidsDIJON-BU Doc.électronique (212319901) / SudocSudocFranceF

Rôle du récepteur nucléaire d'activation et de prolifération des péroxysomes (PPAR-alpha) dans la modulation de l'inflammation et l'activation des cellules T

Notre étude a montré l implication de la déficience de PPARa dans la modulation de latranscription des gènes de l insuline et de l inflammation des adipocytes chez les sourisadultes C57BL/6J (WT) et PPARa-null. A jeun, les souris PPARa-null sont hypoglycémiquespar rapport aux animaux témoins WT. La concentration en insuline et l expression de sesARNm pancréatiques, par rapport aux animaux témoins, sont diminuées chez les sourisPPARa-null, suggérant que la suppression du gène de PPARa contribuait à la faibletranscription de ces gènes. De plus, la suppression du gène de PPARa aboutit à la diminutiondes facteurs de transcription des gènes de l insuline comme Pdx-1, Nkx6.1 et MafA. En outre,la capacité pancréatique fonctionnelle est aussi détériorée par la suppression du gène dePPARa puisque le pancréas des souris PPARa-null exprime de faibles taux de Glut2 et deglucokinase. Les souris PPARa-null expriment des taux élevés d adiponectine et de leptinecomparées aux souris témoins. Dans les tissus adipeux, les souris PPARa-null présentent uneaugmentation de l expression de CD14 et CD68 généralement exprimés par les macrophages.La suppression du gène de PPARa diminue, au niveau des adipocytes, l expression de MCP-1, TNFa, IL-1b, IL-6 et RANTES, alors que l expression de TLR-2 et de TLR-4 (récepteurspro-inflammatoires) était élevée dans les tissus adipeux. Ces résultats suggèrent qu encondition normale, la déficience en PPARa, chez les souris est impliquée dans la modulationde la transcription des gènes de l insuline et le statut inflammatoire du tissu adipeux.En outre, l'invalidation du gène de PPARa dans les cellules T a abouti àl'augmentation de T-bet et la diminution de GATA-3 tant aux niveaux de la protéine que del ARNm. Comme prévu, l acide Docosahexaénoïque (DHA) a exercé non seulement un effetinhibiteur sur la prolifération des cellules T, mais aussi a diminué la sécrétion d IFN-g etstimulé la sécrétion de l IL-4 dans les deux types cellulaires. Le DHA a aussi diminué T-bet etaugmenté GATA-3 tant au niveau de la transcription qu au niveau de la protéine. Quoique lescellules T des souris PPARa-null ont exprimé un plus fort niveau de phosphorylation de p38MAP kinase que les cellules T de WT, le DHA a diminué la phosphorylation des MAPkinases (p38 et ERK1/2) dans tous les deux les types cellulaires. Les inhibiteurspharmacologiques des MAP kinases ont aussi diminué T-bet et augmenté GATA-3 dans lescellules T. Ces résultats démontrent que le DHA, via son action sur les MAP kinases, modulel'expression des facteurs de transcription. Ces résultats expliquent aussi le mécanisme d'actionde cet acide gras sur la différenciation des cellules T dans la maladie et la santéWe assessed, in this study, the effects of PPARa deficiency on the expression of mRNAencoding for insulin gene transcription factors in pancreatic b-cells along with thoseimplicated in inflammation in adipose tissues. On fasting, the adult PPARa-null mice werehypoglycemic. Serum insulin concentrations and its pancreatic mRNA transcripts weredownregulated in PPARa-null mice, suggesting that PPARa gene deletion contributes to lowinsulin gene transcription. The PPARa gene deletion downregulates the mRNA expression ofinsulin gene transcription factors, i.e., Pdx-1, Nkx6.1 and MafA. Besides, the pancreaticfunction was diminished by PPARa deficiency as PPARa-null mice expressed low pancreaticGlut2 and glucokinase mRNA. PPARa-null mice also expressed high adiponectin and leptinmRNA levels compared to wild type animals. Adipose tissues of PPARa-null mice exhibitedupregulation of CD14 and CD68 mRNA, generally expressed by macrophages. PPAR-a genedeletion downregulates the adipocyte mRNA of certain pro inflammatory agents, like MCP-1,TNF-a, IL-1b, IL-6, and RANTES, though pro-inflammatory TLR-2 and TLR-4 mRNAswere upregulated in the adipose tissues. Our results suggest that PPAR-a deficiency, in mice,is implicated in the modulation of insulin gene transcription and inflammatory status inadipose tissues.The another part of the study was conducted on CD4+ T-cells, isolated from wild type(WT) and PPARa-null mice, in order to assess the mechanismof action of docosahexaenoicacid (DHA), an n-3 fatty acid, in the modulation of two transcription factors, i.e., T-bet andGATA-3, implicated in T-cell differentiation towards, respectively, TH1 and TH2 phenotype.The T-cells from PPARa-null mice secreted higher IFN-g and lower IL-4 concentrations thanWT T-cells. Furthermore, the deletion of PPAR-a gene in T-cells resulted in the upregulationof T-bet and downregulation of GATA-3 both at mRNA and protein levels. DHA exerted notonly an inhibitory effect on T-cell proliferation, but also diminished IFN-g and stimulated IL-4 secretions in both cell types. DHA also downregulated T-bet and upregulated GATA-3 bothat transcription and protein levels. Though the T-cells from PPARa-null mice expressedhigher p38 phosphorylation than WT T-cells, DHA diminished the MAP kinasephosphorylation (p38 and ERK1/2) in both the cell types. The pharmacological inhibitors ofMAP kinases also downregulated T-bet and upregulated GATA-3 in T-cells. Altogether, theseresults demonstrate that DHA, via its action on MAP kinases, modulates the expression oftranscription factors. These results also explain the mechanism of action of this fatty acid onT-cell differentiation in disease and healthDIJON-BU Doc.électronique (212319901) / SudocSudocFranceF

The oral lipid sensor GPR120 is not indispensable for the orosensory detectionof dietary lipids in the mouse

International audienceImplication of the long-chain fatty acid (LCFA) receptor GPR120, also termed free fatty acid receptor 4 (FFAR4), in the taste-guided preference for lipids is a matter of debate. To further unravel the role of GPR120 in the "taste of fat", the present study was conducted on GPR120-null mice and their wild-type littermates. Using a combination of morphological (i.e. immunohistochemical staining of circumvallate papillae - CVP), behavioral (i.e. two-bottle preference tests, licking tests and conditioned taste aversion) and functional studies (i.e. calcium imaging in freshly isolated taste bud cells - TBC), we show that absence of GPR120 in oral cavity was not associated with changes in i) the gross anatomy of CVP, ii) the LCFA-mediated increases in [Ca2+]i, iii) the preference for oily and LCFA solutions and iv) the conditioned avoidance of LCFA solutions. In contrast, the rise in [Ca2+]i triggered by grifolic acid (GA), a specific GPR120 agonist, was dramatically curtailed when GPR120 gene was lacking. Taken together these data demonstrate that activation of lingual GPR120 and preference for fat are disconnected, suggesting that GPR120 expressed in TBC is not absolutely required for the oral fat detection in the mouse

Taste for fat: the 6th taste modality

IF 4.066International audienc

Inflammation et immunité : implications dans l’obésité et le diabète de type 2

The evidences have been increasingly accumulated on the implication of inflammatory mediators like tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) in the pathological states related to insulin resistance like obesity, type 2 diabetes and atherosclerosis. There seems a link between insulin resistance and these pro-inflammatory agents, secreted by macrophages and adipocytes. Th (helper) cells are differentiated into either Th1 or Th2 phenotypes. It is generally considered that Th1 phenotype is pro-inflammatory whereas Th2 phenotype exerts anti-inflammatory (protective) effects. The upregulation of Th1 phenotype may aggravate these pathologies. One of the adipokines, i.e., adiponectin, and insulin act as anti-inflammatory agents. Insulin also favours the differentiation of Th cells into Th2 phenotype. TNF-α and IL-6 might counter balance the action of insulin by interfering with insulin receptor signalling in these pathological situations. The agonists of PPARα and PPARγ, and n-3 polyunsaturated fatty acids may exert an anti-inflammatory effect by shifting Th1/Th2 balance to Th2 phenotype. The factors, implicated in the secretion of inflammatory mediators are not well known, though the role of glucose-induced oxidative stress has been underlined. In this article, we shed light on the cross-talk between pro- and anti-inflammatory agents in these patho-physiological states related to insulin resistance

Etude des mecanismes de transport transmembranaire des polyamines: leur importance au cours de la proliferation cellulaire cancereuse

SIGLEINIST T 74092 / INIST-CNRS - Institut de l'Information Scientifique et TechniqueFRFranc

Role of T-cells in Diabetic Pregnancy and Macrosomia

344-349A number of studies have recently addressed the correlationship between diabetic pregnancy/macrosomia and differentiation of T-cells into Th1 and Th2 subsets. Diabetic pregnancy has been found to be associated with a decreased Th1 phenotype and IL-4 mRNA expression. In macrosomic offspring, high expression of IL-2 and IFN-ϒ mRNA, but not of Th2 cytokines is observed, indicating that the Th1 phenotype is upregulated during macrosomia. T-cells of gestational diabetic rats and their macrosomic offspring seem to present a defect in signal transduction. Indeed, the recruitment of free intracellular calcium concentrations from intracellular pool in T-cells of these animals is altered. The phenotype of regulatory T-cells (T-Reg) is upregulated in diabetic pregnancy and their infants. T-cells in diabetic pregnancy and macrosomic obese offspring are in vivo activated. Adipokines and peroxisome proliferator-activated receptor-⍺ (PPARα) also seem to modulate the pro-inflammatory cytokines in these pathologies. Hence, activation of the immune system might be considered as one of the regulatory pathways including metabolic abnormalities in these two pathologies