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

Growth factor concentrations and their placental mRNA expression are modulated in gestational diabetes mellitus: possible interactions with macrosomia

<p>Abstract</p> <p>Background</p> <p>Gestational diabetes mellitus (GDM) is a form of diabetes that occurs during pregnancy. GDM is a well known risk factor for foetal overgrowth, termed macrosomia which is influenced by maternal hypergycemia and endocrine status through placental circulation. The study was undertaken to investigate the implication of growth factors and their receptors in GDM and macrosomia, and to discuss the role of the materno-foeto-placental axis in the <it>in-utero </it>regulation of foetal growth.</p> <p>Methods</p> <p>30 women with GDM and their 30 macrosomic babies (4.75 ± 0.15 kg), and 30 healthy age-matched pregnant women and their 30 newborns (3.50 ± 0.10 kg) were recruited in the present study. Serum concentrations of GH and growth factors, <it>i.e</it>., IGF-I, IGF-BP3, FGF-2, EGF and PDGF-B were determined by ELISA. The expression of mRNA encoding for GH, IGF-I, IGF-BP3, FGF-2, PDGF-B and EGF, and their receptors, <it>i.e</it>., GHR, IGF-IR, FGF-2R, EGFR and PDGFR-β were quantified by using RT-qPCR.</p> <p>Results</p> <p>The serum concentrations of IGF-I, IGF-BP3, EGF, FGF-2 and PDGF-B were higher in GDM women and their macrosomic babies as compared to their respective controls. The placental mRNA expression of the growth factors was either upregulated (FGF-2 or PDGF-B) or remained unaltered (IGF-I and EGF) in the placenta of GDM women. The mRNA expression of three growth factor receptors, <it>i.e</it>., IGF-IR, EGFR and PDGFR-β, was upregulated in the placenta of GDM women. Interestingly, serum concentrations of GH were downregulated in the GDM women and their macrosomic offspring. Besides, the expression of mRNAs encoding for GHR was higher, but that encoding for GH was lower, in the placenta of GDM women than control women.</p> <p>Conclusions</p> <p>Our results demonstrate that growth factors might be implicated in GDM and, in part, in the pathology of macrosomia via materno-foeto-placental axis.</p

Taste perception and its effects on oral nutritional supplements in younger life phases

IF 4.534International audiencePurpose of review The current review summarizes the importance of taste perception with regard to acceptance of oral nutritional supplements (ONS) in young children. We also shed light on how basic tastes may influence the orosensory detection of ONS in the light of genetic variations, encoding for different taste modalities, particularly for sweet and bitter (and fat), in children.Recent findings Single nucleotide polymorphism (SNP) of bitter and sweet taste receptor genes, that is, respectively, TAS2R38 and T1R2/T1R3, may influence orosensory perception of ‘bitter-made-sweet’ ONS. The SNP of fat taste receptor gene, that is, CD36, might communicate with bitter taste perception. The emerging new sixth fat taste may interfere with obesity in children.Summary Sweet and bitter taste modalities are innate cues, expressed by children from birth to adolescence, either by a strong preference or by food aversion. Sweet and bitter tastes also communicate with each other as sweeteners can mask bitter phenotype. The fat preference, encoded by specific lingual taste receptors, is also modulated, via its interaction with phenotype and genotype, by bitter taste. Sodium salts might interact with bitter taste. Finally, the taste modalities will impact on the intake of ONS in children as the taste phenotype changes in this population, irrespective to genotype

Perception du goût du gras par le récepteur CD36 dans la papille caliciforme de souris (étude des voies de signalisation)

FAT/CD36 est impliqué dans la détection et la perception de goût lipidique. Cependant, les mécanismes moléculaires impliqués dans la signalisation déclenchés par ce récepteur dans ces cellules restent inconnus. Dans ce travail, nous avons purifié les cellules qui expriment CD36 à partir des bourgeons du goût de la papille caliciforme chez la souris. Pour élucider les voies de transduction du signal, générées par les AGPI, nous avons utilisé l acide linoléique (AL). Nous avons observé que l AL induit une augmentation de la concentration du calcium libre intracellulaire, [Ca2+]i, via l activation du FAT/CD36. Le SSO, un inhibiteur de FAT/CD36, réduit cette augmentation de [Ca2+]i induite par l AL. Nous avons aussi observé que l AL provoquait la phosphorylation des résidus tyrosines de la protéine Fyn59 et Yes62 de la famille SRC kinases. Les inhibiteurs des SRC-kinases et ceux des canaux SOC baissent l augmentation de [Ca2+]i dans ces cellules. Ces résultats nous permettent d avancer l hypothèse selon laquelle l AL induit l activation des SRC-kinases et favorise l influx calcique dans les cellules CD36-positives. La RT-qPCR a révélé que les cellules gustatives CD36-positives expriment les ARNm des enzymes telles que la TPH-1, l AADC, l TH, la DBH qui sont impliquées dans la synthèse des neurotransmetteurs monoaminergiques. L addition d AL à ces cellules a induit la libération de 5-HT et de noradrénaline à partir des cellules CD36-positives. Cette libération a été bloquée par les inhibiteurs des canaux SOC et ceux des SRC-PTK. Ces résultats ont montré que la liaison d AL avec CD36 augmente la [Ca2+]i, induit la phosphorylation des SRC-PTK et, la libération de 5-HT et NA qui pourraient ainsi être impliqués dans la signalisation vers les fibres nerveuses afférentes et par conséquent, la transmission du signal du goût du gras vers le système nerveux central.We isolated CD36-positive cells from mouse CVP and investigated intracellular signaling events triggered by a polyunsaturated long chain fatty acid, i.e., linoleic acid (LA). LA induced increases in free intracellular calcium concentrations, [Ca2+]i, by recruiting calcium from endoplasmic reticulum (ER) pool via IP3 production, followed by calcium influx via opening of store operated calcium (SOC) channels. LA also induced phosphorylation of SRC-protein tyrosine kinases (PTKs), particularly of Fyn59 and Yes62. LA-evoked phosphorylation of Fyn59 and Yes62 was implicated in the activation of SOC channels. RT-qPCR revealed that the CD36-positive lipid gustatory cells possessed mRNA of enzymes like tryptophan hydroxylase-1 (TPH-1), L-aromatic amino acid decarboxylase (AADC), tyrosine hydroxylase (TH)), dopamine beta-hydroxylase (DBH) which are involved in the synthesis of monoamine neurotransmitters. Interestingly, addition of LA to these cells induced the release of 5-HT and noradrenalin (NA) to the extracellular environment. The LA-induced release of these neurotransmitters was curtailed by a SOC channel blocker and a SRC-PTK inhibitor. These results altogether demonstrate that LA binds to mouse CD36-positive lipid gustatory cells, triggers calcium signaling and SRC-PTK phosphorylation, and induces the release of 5-HT and NA which, in turn, may be implicated in the downstream signaling to the afferent nerve fibers, thus transmitting the output signal from taste buds to the central nervous system.DIJON-BU Sciences Economie (212312102) / SudocSudocFranceF

Un nouvel acteur dans la perception gustative des lipides alimentaires : le canal calcique TRPC3

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A cross-talk between fat and bitter taste modalities

International audienceThe choice of food is governed largely by the sense of taste. To date, five basic taste modalities have been described; however, there is an increasing agreement on the existence of a 6th fat taste. The taste modalities might interact with each other and also with other senses. The advancements in cellular and molecular biology have helped the characterization of taste signaling mechanisms, down to the receptor level and beyond. CD36 and GPR120 have been shown to be involved in the detection of fat taste while bitter taste is perceived by a number of receptors that belong to a family of taste-type 2 receptors (T2R or TAS2R). Hence, the most common role is played by TAS2R16 and TAS2R38 in bitter taste perception in humans. Increasing evidences from behavioural studies suggest that fat and bitter taste modalities might interact with each other, and this interaction might be critical in obesity. In the current review, we will discuss the evidence from genetic and behavioural studies and propose the molecular mechanism of a cross-talk between fat and bitter tastes

Effet de l'acide docosahexaenoïque (DHA), un acide gras polyinsaturé de la famille n-3, dans l'activation des cellules T

Nous avons étudié le rôle de l acide docosahexaenoïque (DHA) dans la régulation de la signalisation calcique, la voie des Src Tyrosines kinases et l activation des canaux TRPC3/6, des étapes clés contrôlant la prolifération cellulaire. De plus, dans les lymphocytes T régulateur murin, nous avons remarqué que le DHA, via son action inhibitrice sur IL-10 induit in vivo et in vitro une inhibition de la prolifération cellulaire. L analyse des chemokines montre que le DHA exerce un effet sur la migration et l adhésion des cellules T régulatrices en régulant l expression des protéines ERK1/2 et Akt. Le DHA, module les fonctions cellulaires en agissant directement sous forme d acide gras libre, ou bien, indirectement sous forme estérifiée en position sn-2, dans les diacylglycérols (DAG). Classiquement, l influx calcique est déclenché suite à la déplétion des réserves calciques intracellulaires, selon un mécanisme dit store-operated Ca2+ (SOC) entry (SOCE), via l ouverture de canaux dits SOC, qui peuvent être les canaux CRAC (Ca2+ release-activated Ca2+ channels) ou les canaux TRPC (Canonical Transient Receptor Potential). Parmi les canaux TRPC, ce sont les canaux TRPC3/6/7 qui sont régulés par les DAG. Dans les cellules Jurkat, seul TRPC3 et TRPC6 sont présents et exprimées différemment en fonction du stade de prolifération et cycle cellulaire. Dans les cellules monocytaires U937, nous avons montré que le DHA, augmente le [Ca2+]i via l ouverture des canaux CRAC et l activation des PKC... D un point de vue physiologique, nous avons démontré que la signalisation calcique déclenchée en amont par le DHA déclenche l apoptose, évaluée par la production de radicaux libres oxygénés (RLO) et l activation de la caspase-3. L ensemble de ces résultats permet de soutenir l hypothèse selon laquelle, le DHA exerce des effets immunomodulateurs, en partie, via l augmentation de la [Ca2+]i, et l inhibition de la voie de signalisation PKC/MAPK.In this study, we investigated the role of docosahexaenoic acid, an n-3 PUFA, in the regulation of calcium, Src tyrosine kinase signaling and activation of TRPC3/6 channels. Furthermore, in mouse T regulatory cells we have shown that DHA via an inhibition of IL-10 induces in vivo and in vitro an inhibition of cell proliferation. The analysis of chemokines shows that DHA exert an effect on T reg migration and adhesion by regulating the expression of ERK1/2 and Akt protein. DHA modulates cell functions directly in the form of free fatty acid or, indirectly in etherified form at the sn-2 position in diacylglycerol (DAG). Classically, calcium influx is achieved following intracellular calcium stores depletion, through a mechanism termed as store-operated Ca2+ (SOC) entry (SOCE), via the opening of SOC channels that can be CRAC (Ca2+ release-activated Ca2+ channels) channels or TRPC (Canonical Transient Receptor Potential) channels. Among TRPC channels, TRPC3/6/7 channels are those which are regulated by DAG. In Jurkat T cells, only TRPC3 and TRPC6 were present and express differentially in relation to cell cycle and proliferation. In U937 cells, DHA induced increases in [Ca2+]i via the opening of CRAC channels and activation of PKC... By a physiological point of view, we have demonstrated that DHA-induced Ca2+ signaling can initiate apoptosis pathway, as assessed by the measurement of reactive oxygen species (ROS) production and caspase-3 activation. Together, these observations suggest that DHA exert immunomodulatory effects in part, by decreasing pHi, increasing [Ca2+]i and inhibiting PKC/MAP kinase pathway.ngl.DIJON-BU Sciences Economie (212312102) / SudocSudocFranceF

Nutrition: From Bench to Bedside

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