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Un niveau élevé d'acides gras saturés (AGS) dans le sang a un effet délétère sur les muscles squelettiques en inhibant à la fois la régénération des fibres musculaires et la synthèse des protéines musculaires. Du fait de sa richesse en AGS, l’huile de palme est controversée. Cette étude avait pour objectif d’étudier l’effet d’un régime riche en huile de palme (brute et raffinée) sur l’expression des facteurs myogéniques dans le muscle de rats sédentaires et le comparer avec un régime riche en huile d’olive et en Lard. Quarante rats mâles Wistar ont été répartis en 5 groupes de 8 rats chacun : 1 groupe témoin et 4 groupes nourris par des régimes riches en graisse (HFD) contenant respectivement de l’huile de palme brute, de l’huile de palme raffinée, de l’huile d’olive et du lard. Après 12 semaines de régime, les rats ont été sacrifiés et les tissus prélevés L'expression de Pax7, Myf5, MyoD et MyoG a été évaluée par RT-qPCR dans les muscles gastrocnémiens. Aucune différence significative n’a été observée entre régime témoin et HFD concernant les expressions de Pax7, Myf5 et MyoG. L’expression de MyoD s’est avérée significativement plus élevée dans les animaux HFD (p=0,0004) par rapport au témoin ; en particulier dans le régime riche en huile d’olive par rapport aux autres régimes HFD (p=0,05). Aucune altération significative de l’expression des gènes des facteurs de régulation myogénique n’a été observée avec l’huile de palme sous ses 2 formes, brute et raffinée. En conclusion, l’huile de palme, malgré sa richesse en AGS, n’a pas d’effet délétère sur la régénération du muscle squelettique.Mots clés : Régime riche graisses, muscle squelettique, Pax7, Myf5, MyoD, MyoG.

Context Toll-like receptor 4 (TLR4) activation contributes to obesity-associated insulin resistance in skeletal muscles (SM). TLR4 signaling involves two pathways: the myeloid differentiation primary response gene 88 (MyD88) leading to inflammatory cytokines production and the toll/interleukin-1 receptor domain-containing adapter-inducing interferon (IFN) I (TRIF)-dependent pathways leading to type 1 interferon (IFNI) and interferon stimulated genes (ISG) expression. The E3 ubiquitin ligase RNF41 allows the preferential activation of the TRIF-IFNI pathway; however, its role in insulin response has not been reported. Methods We measured RNF41 level and IFNI pathway activation (ISG expression) in SM biopsies of obese insulin sensitive (OIS) and obese insulin resistant (OIR) women. Then we isolated and differentiated in myotubes, primary human SM cell progenitors from OIS and OIR SM biopsies. We modulated RNF41 and ISG expression in these myotubes and investigated their effects on insulin response. Results RNF41 expression is down-regulated in vivo in OIR SM and myotubes compared to OIS SM and myotubes. TLR4 activation with palmitate induces TRIF-IFNI pathway and ISG in OIS myotubes but not in OIR myotubes. Inhibition of RNF41 expression with siRNF41 in OIS myotubes treated with palmitate attenuates insulin response, IFNI pathway activation and ISG induction, mimicking OIR phenotype. Further, overexpression of RNF41 in OIR myotubes increases insulin response and ISG expression. Exposure to IFNI or to its inducer polyinosinic-polycytidylic acid, restores ISG expression and insulin sensitivity in OIR myotubes and OIS myotubes transfected with siRNF41. Conclusion Our results identify RNF41 as essential to IFNI pathway activation in order to maintain muscle insulin sensitivity during human obesity.

International audience; One of the major insulin resistance instigators is excessive adiposity and visceral fat depots. Individually, exercise training and polyphenol intake are known to exert health benefits as improving insulin sensitivity. However, their combined curative effects on established obesity and insulin resistance need further investigation particularly on white adipose tissue alterations. Therefore, we compared the effects on different white adipose tissue depot alterations of a combination of exercise and grape polyphenol supplementation in obese insulin-resistant rats fed a high-fat diet to the effects of a high-fat diet alone or a nutritional supplementation of grape polyphenols (50 mg/kg/day) or exercise training (1 hr/day to 5 days/wk consisting of treadmill running at 32 m/min for a 10% slope), for a total duration of 8 weeks. Separately, polyphenol supplementation and exercise decreased the quantity of all adipose tissue depots and mesenteric inflammation. Exercise reduced adipocytes' size in all fat stores. Interestingly, combining exercise to polyphenol intake presents no more cumulative benefit on adipose tissue alterations than exercise alone. Insulin sensitivity was improved at systemic, epididymal, and inguinal adipose tissues levels in trained rats thus indicating that despite their effects on adipocyte morphological/metabolic changes, polyphenols at nutritional doses remain less effective than exercise in fighting insulin resistance.

How glucose sensing by the nervous system impacts the regulation of β cell mass and function during postnatal development and throughout adulthood is incompletely understood. Here, we studied mice with inactivation of glucose transporter 2 (Glut2) in the nervous system (NG2KO mice). These mice displayed normal energy homeostasis but developed late-onset glucose intolerance due to reduced insulin secretion, which was precipitated by high-fat diet feeding. The β cell mass of adult NG2KO mice was reduced compared with that of WT mice due to lower β cell proliferation rates in NG2KO mice during the early postnatal period. The difference in proliferation between NG2KO and control islets was abolished by ganglionic blockade or by weaning the mice on a carbohydrate-free diet. In adult NG2KO mice, first-phase insulin secretion was lost, and these glucose-intolerant mice developed impaired glucagon secretion when fed a high-fat diet. Electrophysiological recordings showed reduced parasympathetic nerve activity in the basal state and no stimulation by glucose. Furthermore, sympathetic activity was also insensitive to glucose. Collectively, our data show that GLUT2-dependent control of parasympathetic activity defines a nervous system/endocrine pancreas axis that is critical for β cell mass establishment in the postnatal period and for long-term maintenance of β cell function.

Homeostasis of serotonergic transmission critically depends on the rate of serotonin reuptake via its plasma membrane transporter (SERT). SERT activity is tightly regulated by multiple mechanisms, including physical association with intracellular proteins and post-translational modifications, such as phosphorylation, but these mechanisms remain partially understood. Here, we show that SERT C-terminal domain recruits both the catalytic and regulatory subunits of the Ca2+-activated protein phosphatase calcineurin (CaN) and that the physical association of SERT with CaN is promoted by CaN activity. Coexpression of constitutively active CaN with SERT increases SERT cell surface expression and 5-HT uptake in HEK-293 cells. It also prevents the reduction of 5-HT uptake induced by an acute treatment of cells with the protein kinase C activator β-PMA and concomitantly decreases PMA-elicited SERT phosphorylation. In addition, constitutive activation of CaNin vivofavors 5-HT uptake in the adult mouse brain, whereas CaN inhibition reduces cerebral 5-HT uptake. Constitutive activation of CaN also decreases immobility in the forced swim test, indicative of an antidepressant-like effect of CaN. These results identify CaN as an important regulator of SERT activity in the adult brain and provide a novel molecular substrate of clinical interest for the understanding of increased risk of mood disorders in transplanted patients treated with immunosuppressive CaN inhibitors.

International audience; In vertebrates, skeletal muscle myofibers display different contractile and metabolic properties associated with different mitochondrial content and activity. We have previously identified a mitochondrial triiodothyronine receptor (p43) regulating mitochondrial transcription and mitochondrial biogenesis. When overexpressed in skeletal muscle, it increases mitochondrial DNA content, stimulates mitochondrial respiration, and induces a shift in the metabolic and contractile features of muscle fibers toward a slower and more oxidative phenotype. Here we show that a p43 depletion in mice decreases mitochondrial DNA replication and respiratory chain activity in skeletal muscle in association with the induction of a more glycolytic muscle phenotype and a decrease of capillary density. In addition, p43(-/-) mice displayed a significant increase in muscle mass relative to control animals and had an improved ability to use lipids. Our findings establish that the p43 mitochondrial receptor strongly affects muscle mass and the metabolic and contractile features of myofibers and provides evidence that this receptor mediates, in part, the influence of thyroid hormone in skeletal muscle.

International audience; In previous studies, we characterized a new hormonal pathway involving a mitochondrial T3 receptor (p43) acting as a mitochondrial transcription factor. In in vitro and in vivo studies, we have shown that p43 increases mitochondrial transcription and mitochondrial biogenesis. In addition, p43 overexpression in skeletal muscle stimulates mitochondrial respiration and induces a shift in metabolic and contractile features of muscle fibers which became more oxidative. Here we have studied the influence of p43 overexpression in skeletal muscle of mice during aging. We report that p43 overexpression initially increased mitochondrial mass. However, after the early rise in mitochondrial DNA occurring at 2 months of age in transgenic mice, we observed a progressive decrease of mitochondrial DNA content which became 2-fold lower at 23 months of age relatively to control animals. Moreover, p43 overexpression induced an oxidative stress characterized by a strong increase of lipid peroxidation and protein oxidation in quadriceps muscle, although antioxidant enzyme activities (catalase and superoxide dismutase) were stimulated. In addition, muscle atrophy became detectable at 6 months of age, probably through a stimulation of the ubiquitin proteasome pathway via two muscle-specific ubiquitin ligases E3, Atrogin-1/MAFbx and MuRF1. Taken together, these results demonstrate that a prolonged stimulation of mitochondrial activity induces muscle atrophy. In addition, these data underline the importance of a tight control of p43 expression and suggest that a deregulation of the direct T3 mitochondrial pathway could be one of the parameters involved in the occurrence of sarcopenia.

International audience; Although physical interactions with other receptors have been reported, heterodimeric complexes of T3 nuclear receptors (TR) with retinoid X receptors (RXRs) are considered as major regulators of T3 target gene expression. However, despite the potent T3 influence in proliferating myoblasts, RXR isoforms are not expressed during proliferation, raising the question of the nature of the complex involved in TR alpha transcriptional activity. We have previously established that c-Jun induces TRalpha1 transcriptional activity in proliferating myoblasts not expressing RXR. This regulation is specific to the muscle lineage, suggesting the involvement of a musclespecific factor. In this study, we found that MyoD expression in HeLa cells stimulates TRalpha1 activity, an influence potentiated by c-Jun coexpression. Similarly, in the absence of RXR, MyoD or c-Jun overexpression in myoblasts induces TRalpha1 transcriptional activity through a direct repeat 4 or an inverted palindrome 6 thyroid hormone response element. The highest rate of activity was recorded when c-Jun and MyoD were coexpressed. Using c-Jun-negative dominants, we established thatMyoD influence on TRalpha1 activity needs c-Jun functionality. Furthermore, we demonstrated that TRalpha1 and MyoD physically interact in the hinge region of the receptor and the transactivation and basic helix loop helix domains of MyoD. RXR expression (spontaneously occurring at the onset of myoblast differentiation) in proliferating myoblasts abrogates these interactions. These data suggest that in the absence of RXR, TRalpha1 transcriptional activity in myoblasts is mediated through a complex including MyoD and c-Jun.

International audience; The regulation of gene expression by thyroid hormone (T-3) involves binding of the hormone to nuclear receptors [ thyroid hormone receptor (TR)] acting as T-3-dependent transcription factors encoded by TR alpha (NR1A1) and TR beta (NR1A2) genes. Several TR alpha variants have already been characterized, but only some of them display T-3 binding activity. In this study, we have identified another transcript, TR alpha-Delta E6, produced by alternative splicing with microexon 6b instead of exon 6. This splicing leads to the synthesis of a protein devoid of a hinge domain. The TR alpha-Delta E6 transcript is detected in all mouse tissues tested. Although TR alpha-Delta E6 did not bind DNA, its expression induced a TR alpha 1 sequestration in the cytoplasm. Functional studies demonstrated that TR alpha-Delta E6 inhibits the transcriptional activity of TR alpha 1 and retinoic X receptor-alpha, but not of retinoic acid receptor-alpha. We also found that TR alpha-Delta E6 efficiently decreased the ability of TR alpha to inhibit MyoD transcriptional activity during myoblast proliferation. Consequently, when overexpressed in myoblasts, it stimulated terminal differentiation. We suggest that this novel TR alpha variant may act as down regulator of overall T-3 receptor activity, including its ability to repress MyoD transcriptional activity during myoblast proliferation.

Parallèlement à son rôle dans le métabolisme énergétique, l’activité mitochondriale intervient également dans l’induction de l’apoptose, ainsi que dans la régulation de la prolifération et de la différenciation cellulaires. Il existe en particulier une véritable régulation de la différenciation des myoblastes par l’activité mitochondriale, indépendante de la production d’ATP. Elle implique notamment le contrôle de l’expression de myogénine et de l’activité des facteurs myogéniques. Dans cette étude, nous démontrons que l’expression du proto-oncogène c-Myc est respectivement stimulée ou diminuée par une inhibition ou une stimulation de l’activité mitochondriale. Cette régulation s’effectue en grande partie au niveau de la stabilité du messager, et au niveau de la localisation cellulaire de la protéine dans les myoblastes aviaires. De plus, la surexpression de c-Myc reproduit très exactement les effets d’une inhibition de l’activité mitochondriale : i) abrogation de la différenciation terminale ; ii) inhibition de l’expression de Myogénine, sans altération de celle de MyoD ; iii) blocage de l’aptitude des facteurs myogéniques à induire la différenciation ; iv) inhibition de la sortie des myoblastes du cycle cellulaire. Ces résultats démontrent que c-Myc est une cible importante de l’activité mitochondriale, impliquée dans l’influence de l’organite sur la différenciation des myoblastes. Nous avons également mis en évidence l’existence d’un autre gène cible de l’organite qui code la phosphatase calcium dépendante Calcineurine. Son expression est respectivement inhibée ou stimulée par l’inhibition ou la stimulation de l’activité mitochondriale. De plus, l’expression d’une forme constitutivement active de Calcineurine stimule la différenciation des myoblastes et l’expression de Myogénine, alors que ces deux événements sont bloqués par l’expression d’un ARN antisens Calcineurine. Enfin, la stimulation de l’activité mitochondriale, comme l’expression d’une forme constitutivement active de Calcineurine stimule spécifiquement l’expression de l’isoforme lente des chaînes lourdes de myosine. Ces données démontrent donc que, notamment via l’expression de Calcineurine, l’activité mitochondriale régule non seulement la différenciation des myoblastes, mais détermine également le type contractile des fibres musculaires, Mitochondrial activity is a major regulator of myoblast differentiation and of the expression of myosin isoforms. Besides their influence on fuel metabolism, mitochondrial activity is also involved in the induction of apoptosis and in the regulation of cell proliferation and differentiation. In particular, an actual regulation of myoblast differentiation by mitochondrial activity is well established, independently of ATP synthesis, through the control of myogenin expression and myogenic factor activity. In this study, we established that c-Myc expression is respectively up- or down-regulated by inhibition or stimulation of mitochondrial activity. This regulation essentially takes place at a post-transcriptional level and also concerns, in avian myoblast, the cellular localisation of the protein. In addition, c-Myc overexpression exactly mimics the influence of an inhibition of mitochondrial activity: i) inhibition of myoblast differentiation; ii) inhibition of myogenin expression; iii) inhibition of myogenic factor ability to induce differentiation; iv) inhibition of myoblast withdrawal from the cell cycle. These results demonstrate that c-Myc is an important target of mitochondrial activity involved in the myogenic influence of the organelle. We also identified another target gene of mitochondrial activity encoding the calcium-dependent phosphatase calcineurin. Its expression is respectively inhibited or stimulated by inhibition or stimulation of mitochondrial activity. In addition, expression of a "constitutively active form of calcineurin stimulates of myoblast differentiation and myosin isoform expression, whereas these two events are suppressed by expression of antisense calcineurin RNA. Lastly, stimulation of mitochondrial activity or overexpression of the constitutively active form of calcineurin specifically stimulates the expression of slow myosin heavy chain isoform. These data demonstrate that, partly via calcineurin expression, mitochondrial activity not only regulates myoblast differentiation, but also the contractile type of muscle fibres.

Mitochondrial dysfunctions are frequently reported in cancer cells, but their direct involvement in tumorigenesis remains unclear. To understand this relation, we stimulated mitochondrial activity by overexpression of the mitochondrial triiodothyronine receptor (p43) in human dermal fibroblasts. In all clones, this stimulation induced morphologic changes and cell fusion in myotube-like structures associated with the expression of several muscle-specific genes (Myf5, desmin, connectin, myosin, AchRα). In addition, these clones displayed all the in vivo and in vitro features of cell transformation. This phenotype was related to an increase in c-Jun and c-Fos expression and extinction of tumor suppressor gene expression (p53, p21WAF1, Rb3). Lastly, reactive oxygen species (ROS) production was increased in positive correlation to the stimulation of mitochondrial activity. The direct involvement of mitochondrial activity in this cell behavior was studied by adding chloramphenicol, an inhibitor of mitochondrial protein synthesis, to the culture medium. This inhibition resulted in partial restoration of the normal phenotype, with the loss of the ability to fuse, a strong decrease in muscle-specific gene expression, and potent inhibition of the transformed phenotype. However, expression of tumor suppressor genes was not restored. Similar results were obtained by using N-acetylcysteine, an inhibitor of ROS production. These data indicate that stimulation of mitochondrial activity in human dermal fibroblasts induces cell transformation through events involving ROS production.

The btg1 (B-cell translocation gene 1) gene coding sequence was isolated from a translocation break point in a case of B-cell chronic lymphocytic leukaemia. We have already shown that BTG1, considered as an antiproliferative protein, strongly stimulates myoblast differentiation. However, the mechanisms involved in this influence remained unknown. In cultured myoblasts, we found that BTG1 stimulates the transcriptional activity of nuclear receptors (T3 and all-trans retinoic acid receptors but not RXRalpha and PPARgamma), c-Jun and myogenic factors (CMD1, Myf5, myogenin). Immunoprecipitation experiments performed in cells or using in vitro-synthesized proteins and GST pull-down assays established that BTG1 directly interacts with T3 and all-trans retinoic acid receptors and with avian MyoD (CMD1). These interactions are mediated by the transactivation domain of each transcription factor and the A box and C-terminal part of BTG1. NCoR presence induces the ligand dependency of the interaction with nuclear receptors. Lastly, deletion of BTG1 interacting domains abrogates its ability to stimulate nuclear receptors and CMD1 activity, and its myogenic influence. In conclusion, BTG1 is a novel important coactivator involved in the regulation of myoblast differentiation. It not only stimulates the activity of myogenic factors, but also of nuclear receptors already known as positive myogenic regulators.