Investigating the real role of HIF-1 and HIF-2 in iron recycling by macrophages

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The gut in iron homeostasis: role of HIF-2 under normal and pathological conditions.

International audienceAlthough earlier, seminal studies demonstrated that the gut per se has the intrinsic ability to regulate the rates of iron absorption, the spotlight in the past decade has been placed on the systemic regulation of iron homeostasis by the hepatic hormone hepcidin and the molecular mechanisms that regulate its expression. Recently, however, attention has returned to the gut based on the finding that hypoxia inducible factor-2 (HIF-2α) regulates the expression of key genes that contribute to iron absorption. Here we review the current understanding of the molecular mechanisms that regulate iron homeostasis in the gut by focusing on the role of HIF-2 under physiological steady-state conditions and in the pathogenesis of iron-related diseases. We also discuss implications for adapting HIF-2-based therapeutic strategies in iron-related pathological conditions

Hepatic hypoxia-inducible factor-2 down-regulates hepcidin expression in mice through an erythropoietin-mediated increase in erythropoiesis

The online version of this article has a Supplementary Appendix. Background Iron metabolism, regulated by the iron hormone hepcidin, and oxygen homeostasis, dependent on hypoxia-inducible factors, are strongly interconnected. We previously reported that in mice in which both liver hypoxia-inducible factors-1 and -2 are stabilized (the hepatocyte von Hippel-Lindau knockout mouse model), hepcidin expression was strongly repressed and we hypothesized that hypoxia-inducible factor-2 could be the major regulatory component contributing to the hepcidin down-regulation. Design and Methods We generated and analyzed hepatocyte-specific knockout mice harboring either hypoxiainducible factor-2α deficiency (Hif2a knockout) or constitutive hypoxia-inducible factor-2α stabilization (Vhlh/Hif1a knockout) and ex vivo systems (primary hepatocyte cultures). Hif2a knockout mice were fed an iron-deficient diet for 2 months and Vhlh/Hif1a knockout mice were treated with neutralizing erythropoietin antibody. Results We demonstrated that hypoxia-inducible factor-2 is dispensable in hepcidin gene regulation in the context of an adaptive response to iron-deficiency anemia. However, its overexpression in the double Vhlh/Hif1a hepatocyte-specific knockout mice indirectly down-regulates hepcidin expression through increased erythropoiesis and erythropoietin production. Experiments in primary hepatocytes confirmed the non-autonomous role of hypoxia-inducible factor-2 in hepcidin regulation. Conclusions While our results indicate that hypoxia-inducible factor-2 is not directly involved in hepcidin repression, they highlight the contribution of hepatic hypoxia-inducible factor-2 to the repression of hepcidin through erythropoietin-mediated increased erythropoiesis, a result of potential clinical interest

Etude des voies de signalisation en aval de Ras impliquées dans le contrôle de la division et de la différenciation de cellules nerveuses

L'oncoprotéine Ras est une GTPase membranaire retrouvée fréquemment mutée dans les tumeurs humaines. La transformation de fibroblastes induite par Ras est médiée par différentes voies de signalisation. Plusieurs effecteurs de Ras participant aux processus de transformation cellulaire ont été mis en évidence chez les vertébrés. Il s'agit des kinases de la famille Raf, responsables de l'activation de la voie MAPK, de la sous-unité catalytique de la PI-3 Kinase (p110) et des facteurs d'échange de la GTPase Ra1. En utilisant des mutants de Ras générés pour leur capacité à n'interagir qu'avec un seul de ces effecteurs, nous avons montré que l'activation constitutive de chacune de ces voies en aval de Ras était capable d'induire la division de cellules de neurorétine (NR) en voie de différenciation. Ces mutants sont également capables de réprimer le promoteur du gène QR1, un gène exprimé spécifiquement dans les cellules NR quiescentes. De plus, en utilisant des versions constitutivement actives ou dominantes négatives de molécules situées en aval de Ras, nous avons démontré la nécessité d'une coopération entre les voies Raf/MAPK et PI-3K/Rac pour l'induction de la prolifération cellulaire. Enfin, nos résultats suggèrent également la mise en place d'une boucle autocrine/paracrine requise pour ce processus. La régulation de B-Raf, l'un des effecteurs majeurs de Ras a également été étudiée. B-Raf code de multiples isoformes générées par épissage alternatif. La présence des séquences alternatives module les propriétés biochimiques et biologiques de B-Raf. En particulier, la présence de l'exon alternatif 10 augmente l'affinité de B-Raf pour son substrat MEK et sa capacité à le phosphoryler. En utilisant les cellules PC12, un modèle de différenciation neuronale in vitro, nous avons mis en évidence un effet de l'exon 10 sur la régulation de l'activité de B-Raf par phosphorylation.The oncoprotein Ras is a membrane-associated GTPase frequently mutated in human cancers. Transformation of fibroblastic cell lines by Ras is mediated through distinct downstream signaling pathways. Several effectors of Ras have been described in vertebrates, including Raf family kinases, responsible for MAPK pathway activation, the catalytic subunit of phosphatidylinositol 3-kinase (p110) and the exchange factors of the Ra1 GTPase. By using Ras mutants, which interact preferentially with only one Ras effector, we have shown that constitutive activation of each Ras downstream signaling pathway was able to induce division of postmitotic and differentiating neuroretina (NR) cells. In addition, these mutants can repress the promoter activity of QR1, a gene specifically expressed in quiescent NR cells. Moreover, by using constitutive and dominant negative mutants of components lying in the downstream signaling pathways, we established the requirement of a cooperation between Raf/MAPK and PI3K/Rac pathways. Finally, the activation of these two pathways possibly relies on an autocrine or paracrine loop, involving endogenous Ras activity. We also studied the regulation of B-Raf, a major Ras effector. B-Raf encodes several isoforms resulting tram complex alternative splicing. The presence of alternative sequences modulates the biochemical and biological properties of B-Raf. In particular, the presence of alternative exon 10 increases the affinity of B-Raf for its substrate MEK and its ability to phopshorylate it. By using the PC12 cell line, an in vitro model of neuronal differentiation, we have disclosed an effect of exon 10 on the regulation of B-Raf activity by phosphorylation.ORSAY-PARIS 11-BU Sciences (914712101) / SudocSudocFranceF

L’hepcidine. Fer de lance de la guérison des maladies chroniques inflammatoires de l’intestin

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Iron: Innocent bystander or vicious culprit in COVID-19 pathogenesis?

International audienceLa pandémie de coronavirus 2 (SARS-CoV-2) se propage violemment à travers les continents avec des taux de mortalité en augmentation rapide. La prise en charge actuelle de la COVID-19 repose sur la prémisse que l’insuffisance respiratoire est la principale cause de mortalité. Cependant, de plus en plus de preuves établissent un lien entre la pathogenèse accélérée chez les patients gravement malades atteints de COVID-19 et un état hyper-inflammatoire impliquant une tempête de cytokines. Plusieurs composantes de l’état inflammatoire accru ont été traitées comme des cibles thérapeutiques. Un autre élément clé de l’état inflammatoire accru est l’hyper-ferritinémie qui identifierait les patients présentant un risque de mortalité accru. Malgré sa forte association avec la mortalité, il n’est pas encore clair si l’hyper-ferritinémie chez les patients atteints de COVID-19 n’est qu’un marqueur systémique de la progression de la maladie ou un modulateur clé de la pathogenèse de la maladie. Nous abordons ici les implications d’un rôle possible pour l’hyper-ferritinémie et l’homéostasie du fer altérée dans la pathogenèse COVID-19, ainsi que les cibles thérapeutiques potentielles à cet égard

Mitochondria and microbiota dysfunction in COVID-19 pathogenesis

International audienceThe COVID-19 pandemic caused by the coronavirus (SARS-CoV-2) has taken the world by surprise into a major crisis of overwhelming morbidity and mortality. This highly infectious disease is associated with respiratory failure unusual in other coronavirus infections. Mounting evidence link the accelerated progression of the disease in COVID-19 patients to the hyper-inflammatory state termed as the "cytokine storm" involving major systemic perturbations. These include iron dysregulation manifested as hyperferritinemia associated with disease severity. Iron dysregulation induces reactive oxygen species (ROS) production and promotes oxidative stress. The mitochondria are the hub of cellular oxidative homeostasis. In addition, the mitochondria may circulate "cell-free" in non-nucleated platelets, in extracellular vesicles and mitochondrial DNA is found in the extracellular space. The heightened inflammatory/oxidative state may lead to mitochondrial dysfunction leading to platelet damage and apoptosis. The interaction of dysfunctional platelets with coagulation cascades aggravates clotting events and thrombus formation. Furthermore, mitochondrial oxidative stress may contribute to microbiota dysbiosis, altering coagulation pathways and fueling the inflammatory/oxidative response leading to the vicious cycle of events. Here, we discuss various cellular and systemic incidents caused by SARS-CoV-2 that may critically impact intra and extracellular mitochondrial function, and contribute to the progression and severity of the disease. It is crucial to understand how these key modulators impact COVID-19 pathogenesis in the quest to identify novel therapeutic targets that may reduce fatal outcomes of the disease