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Chronic oxidative stress plays a critical role in the development of brain malignancies due to the high rate of brain oxygen utilization and concomitant production of reactive oxygen species. The nuclear factor-erythroid-2-related factor 2 (NRF2), a master regulator of antioxidant signaling, is a key factor in regulating brain physiology and the development of age-related neurodegenerative diseases. Also, NRF2 is known to exert a protective antioxidant effect against the onset of oxidative stress-induced diseases, including cancer, along with its pro-oncogenic activities through regulating various signaling pathways and downstream target genes. In glioblastoma (GB), grade 4 glioma, tumor resistance, and recurrence are caused by the glioblastoma stem cell population constituting a small bulk of the tumor core. The persistence and self-renewal capacity of these cell populations is enhanced by NRF2 expression in GB tissues. This review outlines NRF2's dual involvement in cancer and highlights its regulatory role in human brain physiology and diseases, in addition to the development of primary brain tumors and therapeutic potential, with a focus on GB.
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BACKGROUND: Microtubes (MTs), cytoplasmic extensions of glioma cells, are important cell communication structures promoting invasion and treatment resistance through network formation. MTs are abundant in chemoresistant gliomas, in particular, glioblastomas (GBMs), while they are uncommon in chemosensitive IDH-mutant and 1p/19q co-deleted oligodendrogliomas. The aim of this study was to identify potential signaling pathways involved in MT formation. METHODS: Bioinformatics analysis of TCGA was performed to analyze differences between GBM and oligodendroglioma. Patient-derived GBM stem cell lines were used to investigate MT formation under transforming growth factor-beta (TGF-ß) stimulation and inhibition in vitro and in vivo in an orthotopic xenograft model. RNA sequencing and proteomics were performed to detect commonalities and differences between GBM cell lines stimulated with TGF-ß. RESULTS: Analysis of TCGA data showed that the TGF-ß pathway is highly activated in GBMs compared to oligodendroglial tumors. We demonstrated that TGF-ß1 stimulation of GBM cell lines promotes enhanced MT formation and communication via calcium signaling. Inhibition of the TGF-ß pathway significantly reduced MT formation and its associated invasion in vitro and in vivo. Downstream of TGF-ß, we identified thrombospondin 1 (TSP1) as a potential mediator of MT formation in GBM through SMAD activation. TSP1 was upregulated upon TGF-ß stimulation and enhanced MT formation, which was inhibited by TSP1 shRNAs in vitro and in vivo. CONCLUSION: TGF-ß and its downstream mediator TSP1 are important mediators of the MT network in GBM and blocking this pathway could potentially help to break the complex MT-driven invasion/resistance network.
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Glioblastoma , Glioma , Oligodendroglioma , Glioblastoma/patología , Humanos , Trombospondina 1/genética , Trombospondina 1/metabolismo , Factor de Crecimiento Transformador beta/genética , Factor de Crecimiento Transformador beta/metabolismoRESUMEN
Mitochondrial ATP-synthesis is catalyzed by a F1Fo-ATP synthase, an enzyme of dual genetic origin enriched at the edge of cristae where it plays a key role in their structure/stability. The enzyme's biogenesis remains poorly understood, both from a mechanistic and a compartmentalization point of view. The present study provides novel molecular insights into this process through investigations on a human protein called TMEM70 with an unclear role in the assembly of ATP synthase. A recent study has revealed the existence of physical interactions between TMEM70 and the subunit c (Su.c), a protein present in 8 identical copies forming a transmembrane oligomeric ring (c-ring) within the ATP synthase proton translocating domain (Fo). Herein we analyzed the ATP-synthase assembly in cells lacking TMEM70, mitochondrial DNA or F1 subunits and observe a direct correlation between TMEM70 and Su.c levels, regardless of the status of other ATP synthase subunits or of mitochondrial bioenergetics. Immunoprecipitation, two-dimensional blue-native/SDS-PAGE, and pulse-chase experiments reveal that TMEM70 forms large oligomers that interact with Su.c not yet incorporated into ATP synthase complexes. Moreover, discrete TMEM70-Su.c complexes with increasing Su.c contents can be detected, suggesting a role for TMEM70 oligomers in the gradual assembly of the c-ring. Furthermore, we demonstrate using expansion super-resolution microscopy the specific localization of TMEM70 at the inner cristae membrane, distinct from the MICOS component MIC60. Taken together, our results show that TMEM70 oligomers provide a scaffold for c-ring assembly and that mammalian ATP synthase is assembled within inner cristae membranes.
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Proteínas de la Membrana/química , Proteínas de la Membrana/metabolismo , Proteínas Mitocondriales/química , Proteínas Mitocondriales/metabolismo , ATPasas de Translocación de Protón Mitocondriales/química , ATPasas de Translocación de Protón Mitocondriales/metabolismo , Línea Celular , Metabolismo Energético , Técnicas de Inactivación de Genes , Células HEK293 , Humanos , Proteínas de la Membrana/genética , Microscopía Electrónica , Membranas Mitocondriales/metabolismo , Proteínas Mitocondriales/genética , Dominios Proteicos , Multimerización de ProteínaRESUMEN
O. Warburg conducted one of the first studies on tumor energy metabolism. His early discoveries pointed out that cancer cells display a decreased respiration and an increased glycolysis proportional to the increase in their growth rate, suggesting that they mainly depend on fermentative metabolism for ATP generation. Warburg's results and hypothesis generated controversies that are persistent to this day. It is thus of great importance to understand the mechanisms by which cancer cells can reversibly regulate the two pathways of their energy metabolism as well as the functioning of this metabolism in cell proliferation. Here, we made use of yeast as a model to study the Warburg effect and its eventual function in allowing an increased ATP synthesis to support cell proliferation. The role of oxidative phosphorylation repression in this effect was investigated. We show that yeast is a good model to study the Warburg effect, where all parameters and their modulation in the presence of glucose can be reconstituted. Moreover, we show that in this model, mitochondria are not dysfunctional, but that there are fewer mitochondria respiratory chain units per cell. Identification of the molecular mechanisms involved in this process allowed us to dissociate the parameters involved in the Warburg effect and show that oxidative phosphorylation repression is not mandatory to promote cell growth. Last but not least, we were able to show that neither cellular ATP synthesis flux nor glucose consumption flux controls cellular growth rate.
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Iron is required for the oxidative response of neutrophils to allow the production of reactive oxygen species (ROS). However, neutrophil function may be severely altered in conditions of iron overload, as observed in chronically transfused patients. Therefore, a tight regulation of neutrophil iron homeostasis seems to be critical for avoiding iron toxicity. Hepcidin is the key iron regulator in organisms; however, no studies have investigated its role in maintaining neutrophil iron homeostasis or characterized neutrophil function in patients with hereditary hemochromatosis (HH), a common iron overload genetic disorder that results from a defect in hepcidin production. To explore these issues, we studied 2 mouse models of iron overload: an experimentally induced iron overload model (EIO), in which hepcidin is increased, and a genetic HH model of iron overload with a deletion of hepatic hepcidin. We found that iron-dependent increase of hepatic hepcidin results in neutrophil intracellular iron trapping and consecutive defects in oxidative burst activity. In contrast, in both HH mouse models and HH patients, the lack of hepcidin expression protects neutrophils from toxic iron accumulation. Moreover, systemic iron overload correlated with a surprising neutrophil priming and resulted in a more powerful oxidative burst. Indeed, important factors in neutrophil priming and activation, such as tumor necrosis factor α (TNF-α), VCAM-1, and ICAM-1 are increased in the plasma of HH patients and are associated with an increase in HH neutrophil phagocytosis capacity and a decrease in L-selectin surface expression. This is the first study to characterize neutrophil iron homeostasis and associated functions in patients with HH.
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Hemocromatosis , Sobrecarga de Hierro , Animales , Hemocromatosis/genética , Hepcidinas/genética , Humanos , Hierro , Ratones , NeutrófilosRESUMEN
In Duchenne muscular dystrophy (DMD) patients, absence of dystrophin causes muscle wasting by impacting both the myofiber integrity and the properties of muscle stem cells (MuSCs). Investigation of DMD encompasses the use of MuSCs issued from human skeletal muscle. However, DMD-derived MuSC usage is restricted by the limited number of divisions that human MuSCs can undertake in vitro before losing their myogenic characteristics and by the scarcity of human material available from DMD muscle. To overcome these limitations, immortalization of MuSCs appears as a strategy. Here, we used CDK4/hTERT expression in primary MuSCs and we derived MuSC clones from a series of clinically and genetically characterized patients, including eight DMD patients with various mutations, four congenital muscular dystrophies and three age-matched control muscles. Immortalized cultures were sorted into single cells and expanded as clones into homogeneous populations. Myogenic characteristics and differentiation potential were tested for each clone. Finally, we screened various promoters to identify the preferred gene regulatory unit that should be used to ensure stable expression in the human MuSC clones. The 38 clonal immortalized myogenic cell clones provide a large collection of controls and DMD clones with various genetic defects and are available to the academic community.
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Distrofia Muscular de Duchenne/fisiopatología , Células Satélite del Músculo Esquelético/metabolismo , Animales , Estudios de Casos y Controles , Diferenciación Celular , Niño , Preescolar , Modelos Animales de Enfermedad , Femenino , Humanos , Lactante , MasculinoRESUMEN
Heme (iron protoporphyrin IX) is a well-known prosthetic group for enzymes involved in metabolic pathways such as oxygen transport and electron transfer through the mitochondrial respiratory chain. However, heme has also been shown to be an important regulatory molecule (as "labile" heme) for diverse processes such as translation, kinase activity, and transcription in mammals, yeast, and bacteria. Taking advantage of a yeast strain deficient for heme production that enabled controlled modulation and monitoring of labile heme levels, here we investigated the role of labile heme in the regulation of mitochondrial biogenesis. This process is regulated by the HAP complex in yeast. Using several biochemical assays along with EM and epifluorescence microscopy, to the best of our knowledge, we show for the first time that cellular labile heme is critical for the post-translational regulation of HAP complex activity, most likely through the stability of the transcriptional co-activator Hap4p. Consequently, we found that labile heme regulates mitochondrial biogenesis and cell growth. The findings of our work highlight a new mechanism in the regulation of mitochondrial biogenesis by cellular metabolites.
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Factor de Unión a CCAAT/metabolismo , Hemina/metabolismo , Mitocondrias/metabolismo , Biogénesis de Organelos , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/metabolismo , Factor de Unión a CCAAT/genética , Consumo de Oxígeno , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/genética , Transducción de SeñalRESUMEN
Novel approaches for adjunctive therapy are urgently needed for complicated infections and patients with compromised immunity. Necrotizing fasciitis (NF) is a destructive skin and soft tissue infection. Despite treatment with systemic antibiotics and radical debridement of necrotic tissue, lethality remains high. The key iron regulatory hormone hepcidin was originally identified as a cationic antimicrobial peptide (AMP), but its putative expression and role in the skin, a major site of AMP production, have never been investigated. We report here that hepcidin production is induced in the skin of patients with group A Streptococcus (GAS) NF. In a GAS-induced NF model, mice lacking hepcidin in keratinocytes failed to restrict systemic spread of infection from an initial tissue focus. Unexpectedly, this effect was due to its ability to promote production of the CXCL1 chemokine by keratinocytes, resulting in neutrophil recruitment. Unlike CXCL1, hepcidin is resistant to degradation by major GAS proteases and could therefore serve as a reservoir to maintain steady-state levels of CXCL1 in infected tissue. Finally, injection of synthetic hepcidin at the site of infection can limit or completely prevent systemic spread of GAS infection, suggesting that hepcidin agonists could have a therapeutic role in NF.
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Epidermis/metabolismo , Fascitis Necrotizante/metabolismo , Hepcidinas/metabolismo , Neutrófilos/metabolismo , Infecciones Estreptocócicas/metabolismo , Streptococcus pyogenes/metabolismo , Animales , Quimiocina CXCL1/metabolismo , Modelos Animales de Enfermedad , Epidermis/microbiología , Epidermis/patología , Fascitis Necrotizante/patología , Humanos , Queratinocitos/metabolismo , Queratinocitos/patología , Ratones , Ratones Noqueados , Neutrófilos/patología , Infecciones Estreptocócicas/patologíaRESUMEN
Chronic inflammation and fibrosis characterize Duchenne muscular dystrophy (DMD). We show that pro-inflammatory macrophages are associated with fibrosis in mouse and human DMD muscle. DMD-derived Ly6Cpos macrophages exhibit a profibrotic activity by sustaining fibroblast production of collagen I. This is mediated by the high production of latent-TGF-ß1 due to the higher expression of LTBP4, for which polymorphisms are associated with the progression of fibrosis in DMD patients. Skewing macrophage phenotype via AMPK activation decreases ltbp4 expression by Ly6Cpos macrophages, blunts the production of latent-TGF-ß1, and eventually reduces fibrosis and improves DMD muscle force. Moreover, fibro-adipogenic progenitors are the main providers of TGF-ß-activating enzymes in mouse and human DMD, leading to collagen production by fibroblasts. In vivo pharmacological inhibition of TGF-ß-activating enzymes improves the dystrophic phenotype. Thus, an AMPK-LTBP4 axis in inflammatory macrophages controls the production of TGF-ß1, which is further activated by and acts on fibroblastic cells, leading to fibrosis in DMD.
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Proteínas Quinasas Activadas por AMP/metabolismo , Proteínas de Unión a TGF-beta Latente/metabolismo , Distrofia Muscular de Duchenne/metabolismo , Factor de Crecimiento Transformador beta1/metabolismo , Animales , Biomarcadores/metabolismo , Modelos Animales de Enfermedad , Activación Enzimática , Fibroblastos/metabolismo , Fibrosis , Inflamación/patología , Macrófagos/metabolismo , Ratones , Ratones Endogámicos C57BL , Músculo Esquelético/metabolismo , Músculo Esquelético/patología , Distrofia Muscular de Duchenne/patología , Células 3T3 NIHRESUMEN
Inflammation and tissue regeneration follow tissue damage, but little is known about how these processes are coordinated. High Mobility Group Box 1 (HMGB1) is a nuclear protein that, when released on injury, triggers inflammation. We previously showed that HMGB1 with reduced cysteines is a chemoattractant, whereas a disulfide bond makes it a proinflammatory cytokine. Here we report that fully reduced HMGB1 orchestrates muscle and liver regeneration via CXCR4, whereas disulfide HMGB1 and its receptors TLR4/MD-2 and RAGE (receptor for advanced glycation end products) are not involved. Injection of HMGB1 accelerates tissue repair by acting on resident muscle stem cells, hepatocytes, and infiltrating cells. The nonoxidizable HMGB1 mutant 3S, in which serines replace cysteines, promotes muscle and liver regeneration more efficiently than the wild-type protein and without exacerbating inflammation by selectively interacting with CXCR4. Overall, our results show that the reduced form of HMGB1 coordinates tissue regeneration and suggest that 3S may be used to safely accelerate healing after injury in diverse clinical contexts.
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Proteína HMGB1/metabolismo , Regeneración Hepática/fisiología , Músculos/metabolismo , Músculos/fisiología , Receptores CXCR4/metabolismo , Animales , Línea Celular , Factores Quimiotácticos/metabolismo , Citocinas/metabolismo , Células HEK293 , Hepatocitos/metabolismo , Hepatocitos/fisiología , Humanos , Inflamación/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Cicatrización de Heridas/fisiologíaRESUMEN
Tissue regeneration requires inflammatory and reparatory activity of macrophages. Macrophages detect and eliminate the damaged tissue and subsequently promote regeneration. This dichotomy requires the switch of effector functions of macrophages coordinated with other cell types inside the injured tissue. The gene regulatory events supporting the sensory and effector functions of macrophages involved in tissue repair are not well understood. Here we show that the lipid activated transcription factor, PPARγ, is required for proper skeletal muscle regeneration, acting in repair macrophages. PPARγ controls the expression of the transforming growth factor-ß (TGF-ß) family member, GDF3, which in turn regulates the restoration of skeletal muscle integrity by promoting muscle progenitor cell fusion. This work establishes PPARγ as a required metabolic sensor and transcriptional regulator of repair macrophages. Moreover, this work also establishes GDF3 as a secreted extrinsic effector protein acting on myoblasts and serving as an exclusively macrophage-derived regeneration factor in tissue repair.
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Factor 3 de Diferenciación de Crecimiento/metabolismo , Músculo Esquelético/fisiología , Mioblastos/metabolismo , PPAR gamma/metabolismo , Regeneración/fisiología , Animales , Western Blotting , Separación Celular , Inmunoprecipitación de Cromatina , Modelos Animales de Enfermedad , Regulación de la Expresión Génica/fisiología , Ratones , Ratones Endogámicos C57BL , Músculo Esquelético/lesiones , Análisis de Secuencia por Matrices de Oligonucleótidos , Cicatrización de Heridas/fisiologíaRESUMEN
Macrophage gene expression determines phagocyte responses and effector functions. Macrophage plasticity has been mainly addressed in in vitro models that do not account for the environmental complexity observed in vivo. In this study, we show that microarray gene expression profiling revealed a highly dynamic landscape of transcriptomic changes of Ly6C(pos)CX3CR1(lo) and Ly6C(neg)CX3CR1(hi) macrophage populations during skeletal muscle regeneration after a sterile damage. Systematic gene expression analysis revealed that the time elapsed, much more than Ly6C status, was correlated with the largest differential gene expression, indicating that the time course of inflammation was the predominant driving force of macrophage gene expression. Moreover, Ly6C(pos)/Ly6C(neg) subsets could not have been aligned to canonical M1/M2 profiles. Instead, a combination of analyses suggested the existence of four main features of muscle-derived macrophages specifying important steps of regeneration: 1) infiltrating Ly6C(pos) macrophages expressed acute-phase proteins and exhibited an inflammatory profile independent of IFN-γ, making them damage-associated macrophages; 2) metabolic changes of macrophages, characterized by a decreased glycolysis and an increased tricarboxylic acid cycle/oxidative pathway, preceded the switch to and sustained their anti-inflammatory profile; 3) Ly6C(neg) macrophages, originating from skewed Ly6C(pos) cells, actively proliferated; and 4) later on, restorative Ly6C(neg) macrophages were characterized by a novel profile, indicative of secretion of molecules involved in intercellular communications, notably matrix-related molecules. These results show the highly dynamic nature of the macrophage response at the molecular level after an acute tissue injury and subsequent repair, and associate a specific signature of macrophages to predictive specialized functions of macrophages at each step of tissue injury/repair.
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Macrófagos/citología , Macrófagos/metabolismo , Activación Transcripcional/genética , Cicatrización de Heridas/genética , Animales , Receptor 1 de Quimiocinas CX3C , Inflamación/genética , Inflamación/inmunología , Macrófagos/inmunología , Macrófagos/fisiología , Ratones , Receptores de Quimiocina/genética , Receptores de Quimiocina/inmunología , Activación Transcripcional/inmunología , Cicatrización de Heridas/inmunologíaRESUMEN
MicroRNAs (miRNAs) in the AGO-containing RISC complex control messenger RNA (mRNA) translation by binding to mRNA 3' untranslated region (3'UTR). The relationship between miRNAs and other regulatory factors that also bind to mRNA 3'UTR, such as CPEB1 (cytoplasmic polyadenylation element-binding protein), remains elusive. We found that both CPEB1 and miR-15b control the expression of WEE1, a key mammalian cell cycle regulator. Together, they repress WEE1 protein expression during G1 and S-phase. Interestingly, the 2 factors lose their inhibitory activity at the G2/M transition, at the time of the cell cycle when WEE1 expression is maximal, and, moreover, rather activate WEE1 translation in a synergistic manner. Our data show that translational regulation by RISC and CPEB1 is essential in cell cycle control and, most importantly, is coordinated, and can be switched from inhibition to activation during the cell cycle.
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Proteínas de Ciclo Celular/metabolismo , MicroARNs/fisiología , Proteínas Nucleares/metabolismo , Proteínas Tirosina Quinasas/metabolismo , Factores de Transcripción/fisiología , Factores de Escisión y Poliadenilación de ARNm/fisiología , Secuencia de Bases , Proteínas de Ciclo Celular/genética , Puntos de Control de la Fase G2 del Ciclo Celular , Células HeLa , Humanos , Proteínas Nucleares/genética , Proteínas Tirosina Quinasas/genética , Interferencia de ARNRESUMEN
Besides their role in cellular responses to hypoxia, hypoxia-inducible factors (HIFs) are involved in innate immunity and also have anti-inflammatory (M2) functions, such as resolution of inflammation preceding healing. Whereas the first steps of the inflammatory response are associated with proinflammatory (M1) macrophages (MPs), resolution of inflammation is associated with anti-inflammatory MPs exhibiting an M2 phenotype. This M1 to M2 sequence is observed during postinjury muscle regeneration, which provides an excellent paradigm to study the resolution of sterile inflammation. In this study, using in vitro and in vivo approaches in murine models, we demonstrated that deletion of hif1a or hif2a in MPs has no impact on the acquisition of an M2 phenotype. Furthermore, using a multiscale methodological approach, we showed that muscles did not require macrophagic hif1a or hif2a to regenerate. These results indicate that macrophagic HIFs do not play a crucial role during skeletal muscle regeneration induced by sterile tissue damage.
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Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Inflamación/genética , Inflamación/metabolismo , Músculo Esquelético/fisiología , Células Mieloides/metabolismo , Regeneración , Animales , Animales Modificados Genéticamente , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Modelos Animales de Enfermedad , Técnicas de Inactivación de Genes , Hipoxia/genética , Hipoxia/metabolismo , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Inflamación/diagnóstico , Inflamación/inmunología , Macrófagos/inmunología , Macrófagos/metabolismo , Imagen por Resonancia Magnética , Masculino , Ratones , Imagen Molecular , Músculo Esquelético/patología , Fagocitosis , FenotipoRESUMEN
Helicobacter pylori infection triggers chronic inflammation of the gastric mucosa that may progress to gastric cancer. The hypoxia-inducible factors (HIFs) are the central mediators of cellular adaptation to low oxygen levels (hypoxia), but they have emerged recently as major transcriptional regulators of immunity and inflammation. No studies have investigated whether H. pylori affects HIF signaling in immune cells and a potential role for HIF in H. pylori-mediated gastritis. HIF-1 and HIF-2 expression was examined in human H. pylori-positive gastritis biopsies. Subsequent experiments were performed in naive and polarized bone marrow-derived macrophages from wild-type (WT) and myeloid HIF-1α-null mice (HIF-1(Δmyel)). WT and HIF-1(Δmyel) mice were inoculated with H. pylori by oral gavage and sacrificed 6 mo postinfection. HIF-1 was specifically expressed in macrophages of human H. pylori-positive gastritis biopsies. Macrophage HIF-1 strongly contributed to the induction of proinflammatory genes (IL-6, IL-1ß) and inducible NO synthase in response to H. pylori. HIF-2 expression and markers of M2 macrophage differentiation were decreased in response to H. pylori. HIF-1(Δmyel) mice inoculated with H. pylori for 6 mo presented with a similar bacterial colonization than WT mice but, surprisingly, a global increase of inflammation, leading to a worsening of the gastritis, measured by an increased epithelial cell proliferation. In conclusion, myeloid HIF-1 is protective in H. pylori-mediated gastritis, pointing to the complex counterbalancing roles of innate immune and inflammatory phenotypes in driving this pathology.
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Gastritis/etiología , Gastritis/metabolismo , Infecciones por Helicobacter/complicaciones , Infecciones por Helicobacter/metabolismo , Helicobacter pylori , Factor 1 Inducible por Hipoxia/metabolismo , Células Mieloides/metabolismo , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Biopsia , Citocinas/genética , Citocinas/metabolismo , Modelos Animales de Enfermedad , Mucosa Gástrica/inmunología , Mucosa Gástrica/metabolismo , Mucosa Gástrica/microbiología , Mucosa Gástrica/patología , Gastritis/patología , Infecciones por Helicobacter/genética , Infecciones por Helicobacter/inmunología , Helicobacter pylori/inmunología , Humanos , Mediadores de Inflamación/metabolismo , Macrófagos/inmunología , Macrófagos/metabolismo , Macrófagos/microbiología , Ratones , Ratones Transgénicos , Células Mieloides/inmunología , Neutrófilos/inmunología , Neutrófilos/metabolismo , Neutrófilos/microbiologíaRESUMEN
Macrophages control the resolution of inflammation through the transition from a proinflammatory (M1) to an anti-inflammatory (M2) phenotype. Here, we present evidence for a role of AMPKα1, a master regulator of energy homeostasis, in macrophage skewing that occurs during skeletal muscle regeneration. Muscle regeneration was impaired in AMPKα1(-/-) mice. In vivo loss-of-function (LysM-Cre;AMPKα1(fl/fl) mouse) and rescue (bone marrow transplantation) experiments showed that macrophagic AMPKα1 was required for muscle regeneration. Cell-based experiments revealed that AMPKα1(-/-) macrophages did not fully acquire the phenotype or the functions of M2 cells. In vivo, AMPKα1(-/-) leukocytes did not acquire the expression of M2 markers during muscle regeneration. Skewing from M1 toward M2 phenotype upon phagocytosis of necrotic and apoptotic cells was impaired in AMPKα1(-/-) macrophages and when AMPK activation was prevented by the inhibition of its upstream activator, CaMKKß. In conclusion, AMPKα1 is crucial for phagocytosis-induced macrophage skewing from a pro- to anti-inflammatory phenotype at the time of resolution of inflammation.
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Proteínas Quinasas Activadas por AMP/metabolismo , Inflamación/inmunología , Macrófagos/inmunología , Músculo Esquelético/metabolismo , Fagocitosis/inmunología , Proteínas Quinasas Activadas por AMP/genética , Animales , Apoptosis/inmunología , Quinasa de la Proteína Quinasa Dependiente de Calcio-Calmodulina/antagonistas & inhibidores , Células Cultivadas , Leucocitos/metabolismo , Activación de Macrófagos/inmunología , Macrófagos/metabolismo , Masculino , Ratones , Ratones Noqueados , Músculo Esquelético/crecimiento & desarrollo , Fagocitosis/genética , Fenotipo , Regeneración/inmunología , Transducción de Señal/inmunologíaRESUMEN
Adult skeletal muscle has the remarkable property of regenerating after damage, owing to satellite cells and myogenic precursor cells becoming committed to adult myogenesis to rebuild the muscle. This process is accompanied by the continuing presence of macrophages, from the phagocytosis of damaged myofibres to the full re-formation of new myofibres. In recent years, there has been huge progress in our understanding of the roles of macrophages during skeletal muscle regeneration, notably concerning their effects on myogenic precursor cells. Here, we review the most recent knowledge acquired on monocyte entry into damaged muscle, the various macrophage subpopulations, and their respective roles during the sequential phases of muscle repair. We also discuss the role of macrophages after exercise-induced muscle damage, notably in humans.
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Macrófagos/metabolismo , Monocitos/metabolismo , Desarrollo de Músculos/fisiología , Músculo Esquelético/citología , Mioblastos/citología , Regeneración/fisiología , Diferenciación Celular , Humanos , Inflamación/metabolismo , Inflamación/patología , Macrófagos/citología , Monocitos/citología , Músculo Esquelético/metabolismo , Mioblastos/fisiología , Fagocitosis/fisiologíaRESUMEN
Transplantation of muscle precursor cells is of therapeutic interest for focal skeletal muscular diseases. However, major limitations of cell transplantation are the poor survival, expansion and migration of the injected cells. The massive and early death of transplanted myoblasts is not fully understood although several mechanisms have been suggested. Various attempts have been made to improve their survival or migration. Taking into account that muscle regeneration is associated with the presence of macrophages, which are helpful in repairing the muscle by both cleansing the debris and deliver trophic cues to myoblasts in a sequential way, we attempted in the present work to improve myoblast transplantation by coinjecting macrophages. The present data showed that in the 5 days following the transplantation, macrophages efficiently improved: i) myoblast survival by limiting their massive death, ii) myoblast expansion within the tissue and iii) myoblast migration in the dystrophic muscle. This was confirmed by in vitro analyses showing that macrophages stimulated myoblast adhesion and migration. As a result, myoblast contribution to regenerating host myofibres was increased by macrophages one month after transplantation. Altogether, these data demonstrate that macrophages are beneficial during the early steps of myoblast transplantation into skeletal muscle, showing that coinjecting these stromal cells may be used as a helper to improve the efficiency of parenchymal cell engraftment.
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Macrófagos/citología , Músculo Esquelético/citología , Distrofias Musculares/terapia , Mioblastos/citología , Animales , Western Blotting , Movimiento Celular/fisiología , Proliferación Celular , Supervivencia Celular/fisiología , Células Cultivadas , Macrófagos/metabolismo , Ratones , Ratones Endogámicos C57BL , Músculo Esquelético/metabolismo , Mioblastos/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
INTRODUCTION: Duchenne Muscular Dystrophy (DMD) is characterized by the lack of dystrophin that leads to severe myofiber degeneration. We have shown that endomysial fibrosis is correlated with age at ambulation loss in DMD patients. However, the dystrophin-deficient mdx mouse does not have fibrotic lesions in adult limb muscles. Here, we describe a model of chronic mechanical muscle injury that triggers chronic lesions in mdx hindlimb muscle. METHODS: Micromechanical injuries were performed daily in tibialis anterior muscles for 2 weeks. RESULTS: Endomysial fibrosis appeared beginning 1 week post-injury, remained stable for 3 months and was associated with loss of specific maximal force. Fibrosis was associated with an increased expression of factors involved in fibrogenesis including α-smooth muscle actin, connective tissue growth factor, and lysyl oxidase, which colocalized with collagen deposits. CONCLUSIONS: This induced fibrotic dystrophic model may be useful to study mechanisms of fibrosis in dystrophinopathies and to evaluate antifibrotic treatments.
Asunto(s)
Modelos Animales de Enfermedad , Miembro Posterior , Músculo Esquelético/lesiones , Músculo Esquelético/patología , Distrofias Musculares/patología , Distrofia Muscular de Duchenne/patología , Actinas/metabolismo , Animales , Biomarcadores/metabolismo , Colágeno/metabolismo , Factor de Crecimiento del Tejido Conjuntivo/metabolismo , Distrofina/deficiencia , Distrofina/genética , Distrofina/metabolismo , Fibrosis , Masculino , Ratones , Ratones Endogámicos mdx , Músculo Esquelético/metabolismo , Distrofias Musculares/metabolismo , Distrofia Muscular de Duchenne/genética , Distrofia Muscular de Duchenne/metabolismo , Proteína-Lisina 6-Oxidasa/metabolismoRESUMEN
Insulin-like growth factor 2 (IGF-2) mRNA-binding proteins (IMPs) are a family of posttranscriptional regulatory factors with well-understood roles in embryonic development and cancer but with poorly characterized functions in normal adult cells and tissues. We now show that IMP-2, the most ubiquitously expressed member of the family, is abundant in human and mouse adult skeletal myoblasts, where it is indispensable for cell motility and for stabilization of microtubules. To explore the functions of IMP-2, we analyzed the transcripts that were differentially regulated in IMP-2-depleted myoblasts and bound to IMP-2 in normal myoblasts. Among them were the mRNAs of PINCH-2, an important mediator of cell adhesion and motility, and MURF-3, a microtubule-stabilizing protein. By gain- and loss-of-function assays and gel shift experiments, we show that IMP-2 regulates the expression of PINCH-2 and MURF-3 proteins via direct binding to their mRNAs. Upregulation of PINCH-2 in IMP-2-depleted myoblasts is the key event responsible for their decreased motility. Our data reveal how the posttranscriptional regulation of gene expression by IMP-2 contributes to the control of adhesion structures and stable microtubules and demonstrate an important function for IMP-2 in cellular motility.