Obésité et grossesse (étude de l'influence d'un marqueur de l'obésité sur les mécanismes cellulaires et tissulaires de l'accouchement dans un modèle d'explants myométriaux humains)

L obésité maternelle, dont la prévalence ne cesse d augmenter, est associée à de nombreux troubles de l accouchement, tels que des dépassements de terme à l origine d une augmentation du taux de césariennes. Ces troubles pourraient, en partie, s expliquer par une concentration plasmatique de leptine plus élevée chez les femmes enceintes obèses ainsi que par les effets inhibiteurs, démontrés in vitro, de cette adipokine sur la contractilité myométriale. Au moment de l accouchement, la transition phénotypique du myomètre d un état de quiescence utérine à un état contractile est une étape clé indispensable à la mise en route du travail. Elle est associée à une activation de l apoptose des cellules myométriales ainsi qu à un remodelage de la matrice extracellulaire utérine. Le but de notre travail était d étudier la capacité de la leptine à moduler l apoptose et le remodelage myométriaux induits par le lipopolysaccharide (LPS).Les échantillons de myomètre ont été prélevés lors de césariennes réalisées avant la mise en route du travail, à la maternité du CHU de Dijon. Les effets de la leptine ont été évalués après incubation des explants myométriaux pendant 48 heures avec du LPS (10 g/ml) avec ou sans leptine (de 10-10 à 10-8 M).Nos résultats ont démontré la capacité de la leptine à inhiber, de façon concentration-dépendante, l apoptose induite par le LPS en diminuant l expression des protéines pro-apoptotiques (caspase-3 clivée, BAX) et en augmentant celle du médiateur anti-apoptotique BCL2. Cet effet anti-apoptotique de la leptine dans le myomètre gestant était associé à l activation de la voie de signalisation ERK1/2. De plus, nos résultats ont montré que la leptine était également capable de s opposer, de façon concentration-dépendante, à la dégradation du collagène de la matrice extracellulaire myométriale induite par le LPS. Cet effet était associé à l inhibition de l activation et de la surexpression des métalloprotéinases MMP2 et MMP9 induites par le LPS.Ce travail a permis d approfondir les connaissances sur le rôle de la leptine dans la régulation de l'activité du myomètre. Nos résultats suggèrent que les troubles de l accouchement observés chez les femmes obèses résulteraient de l inhibition de l apoptose et du remodelage myométriaux par la leptine, en plus de l inhibition de la contractilité utérine déjà décrite.Maternal obesity is associated with a wide spectrum of delivery disorders, such as delayed or post-term delivery, that might be explained partly by the increase in plasma leptin levels in obese women, as leptin inhibits in vitro myometrial contractility. Delivery involves uterine apoptosis and remodelling of the extracellular matrix, via the activation of matrix metalloproteinases (MMP). This study was aimed to assess the role of leptin on human myometrium, by studying the interaction of leptin with lipopolysaccharide (LPS)-induced apoptosis and degradation of myometrial collagen.Myometrial biopsies were obtained from women undergoing caesarean delivery before labour onset. The effects of leptin on myometrial apoptosis and remodelling were assessed by incubating the strips for 48h with LPS (10 g/ml) alone or with leptin (from 10-10 to 10-8 M).Leptin prevented LPS-induced apoptosis, in a concentration-dependent manner, by down-regulating cleaved caspase-3, BAX and up-regulating BCL2 expression. This effect was specifically mediated through leptin receptors stimulation followed by ERK1/2 signalling pathway activation. Leptin prevented, in a concentration-dependent manner, an LPS-induced decrease in myometrial collagen content, and this effect was associated with a decrease in MMP2 and MMP9 activity and overexpression. These effects of leptin were abolished by pre-treatment with a selective leptin receptor antagonist. These results suggest new potential pathways involved in delivery disorders of obese women and propose a role for leptin-induced inhibition of myometrial apoptosis and extracellular matrix remodelling in the development of such disorders.DIJON-BU Doc.électronique (212319901) / SudocSudocFranceF

Obesity and pregnancy : study of the influence of a marker of obesity on the cellular and tissular mechanisms of delivery in an in vitro human myometrial model

L’obésité maternelle, dont la prévalence ne cesse d’augmenter, est associée à de nombreux troubles de l’accouchement, tels que des dépassements de terme à l’origine d’une augmentation du taux de césariennes. Ces troubles pourraient, en partie, s’expliquer par une concentration plasmatique de leptine plus élevée chez les femmes enceintes obèses ainsi que par les effets inhibiteurs, démontrés in vitro, de cette adipokine sur la contractilité myométriale. Au moment de l’accouchement, la transition phénotypique du myomètre d’un état de quiescence utérine à un état contractile est une étape clé indispensable à la mise en route du travail. Elle est associée à une activation de l’apoptose des cellules myométriales ainsi qu’à un remodelage de la matrice extracellulaire utérine. Le but de notre travail était d’étudier la capacité de la leptine à moduler l’apoptose et le remodelage myométriaux induits par le lipopolysaccharide (LPS).Les échantillons de myomètre ont été prélevés lors de césariennes réalisées avant la mise en route du travail, à la maternité du CHU de Dijon. Les effets de la leptine ont été évalués après incubation des explants myométriaux pendant 48 heures avec du LPS (10 µg/ml) avec ou sans leptine (de 10-10 à 10-8 M).Nos résultats ont démontré la capacité de la leptine à inhiber, de façon concentration-dépendante, l’apoptose induite par le LPS en diminuant l’expression des protéines pro-apoptotiques (caspase-3 clivée, BAX) et en augmentant celle du médiateur anti-apoptotique BCL2. Cet effet anti-apoptotique de la leptine dans le myomètre gestant était associé à l’activation de la voie de signalisation ERK1/2. De plus, nos résultats ont montré que la leptine était également capable de s’opposer, de façon concentration-dépendante, à la dégradation du collagène de la matrice extracellulaire myométriale induite par le LPS. Cet effet était associé à l’inhibition de l’activation et de la surexpression des métalloprotéinases MMP2 et MMP9 induites par le LPS.Ce travail a permis d’approfondir les connaissances sur le rôle de la leptine dans la régulation de l'activité du myomètre. Nos résultats suggèrent que les troubles de l’accouchement observés chez les femmes obèses résulteraient de l’inhibition de l’apoptose et du remodelage myométriaux par la leptine, en plus de l’inhibition de la contractilité utérine déjà décrite.Maternal obesity is associated with a wide spectrum of delivery disorders, such as delayed or post-term delivery, that might be explained partly by the increase in plasma leptin levels in obese women, as leptin inhibits in vitro myometrial contractility. Delivery involves uterine apoptosis and remodelling of the extracellular matrix, via the activation of matrix metalloproteinases (MMP). This study was aimed to assess the role of leptin on human myometrium, by studying the interaction of leptin with lipopolysaccharide (LPS)-induced apoptosis and degradation of myometrial collagen.Myometrial biopsies were obtained from women undergoing caesarean delivery before labour onset. The effects of leptin on myometrial apoptosis and remodelling were assessed by incubating the strips for 48h with LPS (10 µg/ml) alone or with leptin (from 10-10 to 10-8 M).Leptin prevented LPS-induced apoptosis, in a concentration-dependent manner, by down-regulating cleaved caspase-3, BAX and up-regulating BCL2 expression. This effect was specifically mediated through leptin receptors stimulation followed by ERK1/2 signalling pathway activation. Leptin prevented, in a concentration-dependent manner, an LPS-induced decrease in myometrial collagen content, and this effect was associated with a decrease in MMP2 and MMP9 activity and overexpression. These effects of leptin were abolished by pre-treatment with a selective leptin receptor antagonist. These results suggest new potential pathways involved in delivery disorders of obese women and propose a role for leptin-induced inhibition of myometrial apoptosis and extracellular matrix remodelling in the development of such disorders

Obésité et grossesse : étude de l'influence d'un marqueur de l'obésité sur les mécanismes cellulaires et tissulaires de l'accouchement dans un modèle d'explants myométriaux humains

Maternal obesity is associated with a wide spectrum of delivery disorders, such as delayed or post-term delivery, that might be explained partly by the increase in plasma leptin levels in obese women, as leptin inhibits in vitro myometrial contractility. Delivery involves uterine apoptosis and remodelling of the extracellular matrix, via the activation of matrix metalloproteinases (MMP). This study was aimed to assess the role of leptin on human myometrium, by studying the interaction of leptin with lipopolysaccharide (LPS)-induced apoptosis and degradation of myometrial collagen.Myometrial biopsies were obtained from women undergoing caesarean delivery before labour onset. The effects of leptin on myometrial apoptosis and remodelling were assessed by incubating the strips for 48h with LPS (10 µg/ml) alone or with leptin (from 10-10 to 10-8 M).Leptin prevented LPS-induced apoptosis, in a concentration-dependent manner, by down-regulating cleaved caspase-3, BAX and up-regulating BCL2 expression. This effect was specifically mediated through leptin receptors stimulation followed by ERK1/2 signalling pathway activation. Leptin prevented, in a concentration-dependent manner, an LPS-induced decrease in myometrial collagen content, and this effect was associated with a decrease in MMP2 and MMP9 activity and overexpression. These effects of leptin were abolished by pre-treatment with a selective leptin receptor antagonist. These results suggest new potential pathways involved in delivery disorders of obese women and propose a role for leptin-induced inhibition of myometrial apoptosis and extracellular matrix remodelling in the development of such disorders.L’obésité maternelle, dont la prévalence ne cesse d’augmenter, est associée à de nombreux troubles de l’accouchement, tels que des dépassements de terme à l’origine d’une augmentation du taux de césariennes. Ces troubles pourraient, en partie, s’expliquer par une concentration plasmatique de leptine plus élevée chez les femmes enceintes obèses ainsi que par les effets inhibiteurs, démontrés in vitro, de cette adipokine sur la contractilité myométriale. Au moment de l’accouchement, la transition phénotypique du myomètre d’un état de quiescence utérine à un état contractile est une étape clé indispensable à la mise en route du travail. Elle est associée à une activation de l’apoptose des cellules myométriales ainsi qu’à un remodelage de la matrice extracellulaire utérine. Le but de notre travail était d’étudier la capacité de la leptine à moduler l’apoptose et le remodelage myométriaux induits par le lipopolysaccharide (LPS).Les échantillons de myomètre ont été prélevés lors de césariennes réalisées avant la mise en route du travail, à la maternité du CHU de Dijon. Les effets de la leptine ont été évalués après incubation des explants myométriaux pendant 48 heures avec du LPS (10 µg/ml) avec ou sans leptine (de 10-10 à 10-8 M).Nos résultats ont démontré la capacité de la leptine à inhiber, de façon concentration-dépendante, l’apoptose induite par le LPS en diminuant l’expression des protéines pro-apoptotiques (caspase-3 clivée, BAX) et en augmentant celle du médiateur anti-apoptotique BCL2. Cet effet anti-apoptotique de la leptine dans le myomètre gestant était associé à l’activation de la voie de signalisation ERK1/2. De plus, nos résultats ont montré que la leptine était également capable de s’opposer, de façon concentration-dépendante, à la dégradation du collagène de la matrice extracellulaire myométriale induite par le LPS. Cet effet était associé à l’inhibition de l’activation et de la surexpression des métalloprotéinases MMP2 et MMP9 induites par le LPS.Ce travail a permis d’approfondir les connaissances sur le rôle de la leptine dans la régulation de l'activité du myomètre. Nos résultats suggèrent que les troubles de l’accouchement observés chez les femmes obèses résulteraient de l’inhibition de l’apoptose et du remodelage myométriaux par la leptine, en plus de l’inhibition de la contractilité utérine déjà décrite

ADRB3 stimulation induced M2-like phenotype in in vitro primary human macrophages

International audienc

Wound Healing versus Metastasis: Role of Oxidative Stress

Many signaling pathways, molecular and cellular actors which are critical for wound healing have been implicated in cancer metastasis. These two conditions are a complex succession of cellular biological events and accurate regulation of these events is essential. Apart from inflammation, macrophages-released ROS arise as major regulators of these processes. But, whatever the pathology concerned, oxidative stress is a complicated phenomenon to control and requires a finely tuned balance over the different stages and responding cells. This review provides an overview of the pivotal role of oxidative stress in both wound healing and metastasis, encompassing the contribution of macrophages. Indeed, macrophages are major ROS producers but also appear as their targets since ROS interfere with their differentiation and function. Elucidating ROS functions in wound healing and metastatic spread may allow the development of innovative therapeutic strategies involving redox modulators

Macrophage-induced reactive oxygen species promote myometrial contraction and labor-associated mechanisms

At labor, the myometrium is infiltrated by a massive influx of macrophages that secrete high levels of pro-inflammatory cytokines inducing the expression of specific labor-associated markers. However, the interactions between myocytes and macrophages and the role of macrophages in the myometrium at labor remain to be elucidated. In this work, we studied the role of myometrium-infiltrated macrophages and their interaction with myocytes in lipopolysaccharide-induced preterm labor. A co-culture model of human primary myometrial cells and macrophages was developed and validated. Collagen lattices were used to evaluate myocyte contraction. Differentiation steps were assessed by (i) phalloidin and vinculin staining for cytoskeleton reorganization, (ii) gap junction protein alpha 1 expression and scrape loading/dye transfer with Lucifer Yellow for gap junction intercellular communication, and (iii) calcium imaging for cell excitability. We demonstrated that macrophages favored lipopolysaccharide-induced contraction and early differentiation of myometrial cells. Transwell assays showed that previous activation of macrophages by lipopolysaccharide was essential for this differentiation and that macrophage/myocyte interactions involved macrophage release of reactive oxygen species (ROS). The effects of macrophage-released ROS in myometrial cell transactivation were mimicked by H2O2, suggesting that superoxide anion is a major intermediate messenger in macrophage/myocyte crosstalk during labor. These novel findings provide the foundation for innovative approaches to managing preterm labor, specifically the use of antioxidants to inhibit the initial stages of labor before the contractile phenotype has been acquired. In addition, the co-culture model developed by our team could be used in future research to decipher pathophysiological signaling pathways or screen/develop new tocolytics

Beta3 adrenergic receptor stimulation in human macrophages inhibits NADPHoxidase activity and induces catalase expression via PPARγ activation

IF 4.521International audienceThe beta3 adrenergic receptor (β3-AR) stimulation plays a protective role against preterm labor by blocking myometrial contraction, cytokine production, remodeling and apoptosis. We previously demonstrated that macrophage-induced ROS production in the myometrium was a key element leading to the induction of all these labor-associated features. We thus aimed to investigate if the β3-AR could be expressed in human macrophages and could trigger its protective role in the myometrium by directly inhibiting ROS production. Using lipopolysaccharide (LPS)-stimulated myometrial samples and cell co-culture experiments, we demonstrated that β3-AR stimulation inhibits the activation of the NADPH oxidase, leading to the subsequent inhibition of ROS production by macrophages. This antioxidant effect was associated with a potent anti-inflammatory response in macrophages. Furthermore, we observed that β3-AR leads to the expression of catalase not only in macrophages but also in myometrial cells, thereby preventing the transactivation of myometrial cells by hydrogen peroxide. Pharmacological experiments allowed us to demonstrate that these effects were driven by an Erk1/2-mediated activation of the antioxidant transcription factor PPARγ. These results suggest that β3-AR protective effects in the myometrium could be due to its dual antioxidant properties. Further, the effects observed in a macrophage could highlight new applications in chronic inflammatory diseases

Idarubicin-Loaded ONCOZENE Drug-Eluting Embolic Agents for Chemoembolization of Hepatocellular Carcinoma: In Vitro Loading and Release and In Vivo Pharmacokinetics

CERVOXY COLL ACL CLINInternational audienc

Leptin-Induced HLA-G Inhibits Myometrial Contraction and Differentiation

International audienceMaternal obesity is associated with a wide spectrum of labour disorders, including preterm birth. Leptin, a pro-inflammatory adipokine and a key factor of obesity, is suspected to play a major role in these disorders. OB-R, its receptor, is expressed on macrophages and myocytes, two cell types critical for labour onset. Macrophages secrete reactive oxygen species/pro-inflammatory cytokines, responsible for myometrial differentiation while myocytes control uterine contractions. In this study, we assessed the effect of leptin on myometrial contraction and differentiation using our validated co-culture model of human primary macrophages and myocytes. We demonstrated that leptin had a different effect on myocytes and macrophages depending on the dose. A low leptin concentration induced a tocolytic effect by preventing myocytes’ contraction, differentiation, and macrophage-induced ROS production. Additionally, leptin led to an increase in HLA-G expression, suggesting that the tocolytic effect of leptin may be driven by HLA-G, a tolerogenic molecule. Finally, we observed that recombinant HLA-G also prevented LPS-induced ROS production by macrophages. Altogether, these data provide a putative molecular mechanism by which leptin may induce immune tolerance and therefore interfere with labour-associated mechanisms. Therefore, HLA-G represents a potential innovative therapeutic target in the pharmacological management of preterm labour

Dose-dependent biphasic leptin-induced proliferation is caused by non-specific IL-6/NF-κB pathway activation in human myometrial cells.

BACKGROUND AND PURPOSE: Leptin, an adipokine synthesized by the placenta during pregnancy, has been proposed for the management of preterm labour (PTL), as it is able to prevent in vitro uterine contractility and remodelling associated with labour onset. Another common feature of labour onset is the phenotypic switch of myometrial smooth muscle cells from a proliferative to a hypertrophic state. As proliferative effects have been demonstrated for leptin in other tissues, we aimed to investigate its ability to induce myometrial proliferation and thus to maintain uterine quiescence. EXPERIMENTAL APPROACH: We stimulated human primary myometrial smooth muscle cells with leptin in the presence or absence of receptor antagonists or signalling pathway inhibitors. KEY RESULTS: Leptin induced myometrial cell proliferation in a biphasic manner. At 6.25 ng · mL(-1), leptin-induced proliferation was mediated by the leptin receptor and required the early activation of ERK1/2. At a concentration above 25 ng · mL(-1), leptin induced direct non-specific stimulation of the IL-6 receptor, leading to NF-κB activation, and exerted anti-proliferative effects. However, at 50 ng · mL(-1), leptin re-induces proliferation via IL-6 receptor stimulation that requires STAT3 and delayed ERK1/2 activation. CONCLUSIONS AND IMPLICATIONS: These data bring new insights into leptin signalling-induced myometrial proliferation and its interrelationship with the IL-6/IL-6 receptor axis. In the light of our previous work, the present study emphasizes the potential value of leptin in the pharmacological management of PTL and it also strengthens the hypothesis that leptin might be a contributory factor in the parturition-related disorders observed in obese women