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OBJECTIVE: We evaluated the influence of sex on the pathophysiology of non-alcoholic fatty liver disease (NAFLD). We investigated diet-induced phenotypic responses to define sex-specific regulation between healthy liver and NAFLD to identify influential pathways in different preclinical murine models and their relevance in humans. DESIGN: Different models of diet-induced NAFLD (high-fat diet, choline-deficient high-fat diet, Western diet or Western diet supplemented with fructose and glucose in drinking water) were compared with a control diet in male and female mice. We performed metabolic phenotyping, including plasma biochemistry and liver histology, untargeted large-scale approaches (liver metabolome, lipidome and transcriptome), gene expression profiling and network analysis to identify sex-specific pathways in the mouse liver. RESULTS: The different diets induced sex-specific responses that illustrated an increased susceptibility to NAFLD in male mice. The most severe lipid accumulation and inflammation/fibrosis occurred in males receiving the high-fat diet and Western diet, respectively. Sex-biased hepatic gene signatures were identified for these different dietary challenges. The peroxisome proliferator-activated receptor α (PPARα) co-expression network was identified as sexually dimorphic, and in vivo experiments in mice demonstrated that hepatocyte PPARα determines a sex-specific response to fasting and treatment with pemafibrate, a selective PPARα agonist. Liver molecular signatures in humans also provided evidence of sexually dimorphic gene expression profiles and the sex-specific co-expression network for PPARα. CONCLUSIONS: These findings underscore the sex specificity of NAFLD pathophysiology in preclinical studies and identify PPARα as a pivotal, sexually dimorphic, pharmacological target. TRIAL REGISTRATION NUMBER: NCT02390232.
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Enfermedad del Hígado Graso no Alcohólico , Animales , Dieta Alta en Grasa/efectos adversos , Modelos Animales de Enfermedad , Femenino , Humanos , Metabolismo de los Lípidos , Hígado/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Enfermedad del Hígado Graso no Alcohólico/metabolismo , PPAR alfa/metabolismoRESUMEN
Glial fibrillary acidic protein (GFAP), ubiquitin carboxy-terminal hydrolase-L1 (UCH-L1), and matrix metalloproteinase 9 (MMP-9) are potential biomarkers of traumatic brain injury (TBI) but also of secondary insults to the brain. The aim of this study was to describe the cerebral distribution of GFAP, UCH-L1, and MMP-9 in a rat model of diffuse TBI associated with standardized hypoxia-hypotension (HH). Adult male Sprague-Dawley rats were allocated to Sham (n = 10), TBI (n = 10), HH (n = 10), and TBI+HH (n = 10) groups. After 4 hours, brains were rapidly removed and immunostaining of GFAP, UCH-L1, and MMP-9 was performed. Areas of interest that have been described as particularly sensitive to hypoxic insults were analyzed. For GFAP, in the neocortex, immunostaining revealed a significant decrease in strong staining for HH and TBI+HH groups compared with TBI group (P < .0001). For UCH-L1, the total immunostaining (6 regions of interest) reported a significant increase in strong staining (P < .0001) and decrease in weak staining (P < .0001) for the HH and TBI+HH groups compared with the Sham and TBI groups. For MMP-9, for the HH and TBI+HH groups, a significant increase in moderate (P < .0001) and weak staining (P < .0001) and a decrease in negative staining (P < .0001) compared with the Sham and TBI groups were observed. UCH-L1 and MMP-9 immunostainings increased after HH alone or HH combined with TBI compared with TBI alone. GFAP immunostaining decreased particularly in the neocortex after HH alone or HH combined with TBI compared with TBI alone. These three biomarkers could therefore be considered as potential biomarkers of HH insults independently of TBI.
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The tuberculosis (TB) bacillus, Mycobacterium tuberculosis (Mtb), and HIV-1 act synergistically; however, the mechanisms by which Mtb exacerbates HIV-1 pathogenesis are not well known. Using in vitro and ex vivo cell culture systems, we show that human M(IL-10) anti-inflammatory macrophages, present in TB-associated microenvironment, produce high levels of HIV-1. In vivo, M(IL-10) macrophages are expanded in lungs of co-infected non-human primates, which correlates with disease severity. Furthermore, HIV-1/Mtb co-infected patients display an accumulation of M(IL-10) macrophage markers (soluble CD163 and MerTK). These M(IL-10) macrophages form direct cell-to-cell bridges, which we identified as tunneling nanotubes (TNTs) involved in viral transfer. TNT formation requires the IL-10/STAT3 signaling pathway, and targeted inhibition of TNTs substantially reduces the enhancement of HIV-1 cell-to-cell transfer and overproduction in M(IL-10) macrophages. Our study reveals that TNTs facilitate viral transfer and amplification, thereby promoting TNT formation as a mechanism to be explored in TB/AIDS potential therapeutics.
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Infecciones por VIH/complicaciones , Interleucina-10/metabolismo , Macrófagos/patología , Nanotubos , Factor de Transcripción STAT3/metabolismo , Tuberculosis Pulmonar/complicaciones , Adulto , Anciano , Animales , Células Cultivadas , Coinfección/patología , Coinfección/virología , Femenino , Infecciones por VIH/inmunología , Infecciones por VIH/patología , Infecciones por VIH/virología , Humanos , Macaca mulatta , Activación de Macrófagos , Macrófagos/virología , Masculino , Persona de Mediana Edad , Mycobacterium tuberculosis , Transducción de Señal , Tuberculosis Pulmonar/inmunología , Tuberculosis Pulmonar/patología , Replicación Viral , Adulto JovenRESUMEN
Fumonisin B1 (FB1), a congener of fumonisins produced by Fusarium species, is the most abundant and most toxicologically active fumonisin. FB1 causes severe mycotoxicosis in animals, including nephrotoxicity, hepatotoxicity, and disruption of the intestinal barrier. However, mechanisms associated with FB1 toxicity are still unclear. Preliminary studies have highlighted the role of liver X receptors (LXRs) during FB1 exposure. LXRs belong to the nuclear receptor family and control the expression of genes involved in cholesterol and lipid homeostasis. In this context, the toxicity of FB1 was compared in female wild-type (LXR+/+) and LXRα,ß double knockout (LXR-/-) mice in the absence or presence of FB1 (10 mg/kg body weight/day) for 28 days. Exposure to FB1 supplemented in the mice's drinking water resulted in more pronounced hepatotoxicity in LXR-/- mice compared to LXR+/+ mice, as indicated by hepatic transaminase levels (ALT, AST) and hepatic inflammatory and fibrotic lesions. Next, the effect of FB1 exposure on the liver transcriptome was investigated. FB1 exposure led to a specific transcriptional response in LXR-/- mice that included altered cholesterol and bile acid homeostasis. ELISA showed that these effects were associated with an elevated FB1 concentration in the plasma of LXR-/- mice, suggesting that LXRs participate in intestinal absorption and/or clearance of the toxin. In summary, this study demonstrates an important role of LXRs in protecting the liver against FB1-induced toxicity, suggesting an alternative mechanism not related to the inhibition of sphingolipid synthesis for mycotoxin toxicity.
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Enfermedad Hepática Inducida por Sustancias y Drogas/metabolismo , Fumonisinas/toxicidad , Receptores X del Hígado/metabolismo , Alanina Transaminasa/metabolismo , Animales , Aspartato Aminotransferasas/metabolismo , Enfermedad Hepática Inducida por Sustancias y Drogas/etiología , Enfermedad Hepática Inducida por Sustancias y Drogas/patología , Femenino , Fumonisinas/sangre , Regulación de la Expresión Génica/efectos de los fármacos , Hígado/efectos de los fármacos , Hígado/fisiología , Receptores X del Hígado/genética , Ratones Endogámicos C57BL , Ratones Noqueados , Esfingolípidos/metabolismoRESUMEN
DC-SIGN (CD209/CLEC4L) is a C-type lectin receptor (CLR) that serves as a reliable cell-surface marker of interleukin 4 (IL-4)-activated human macrophages [M(IL-4)], which historically represent the most studied subset within the M2 spectrum of macrophage activation. Although DC-SIGN plays important roles in Mycobacterium tuberculosis (Mtb) interactions with dendritic cells, its contribution to the Mtb-macrophage interaction remains poorly understood. Since high levels of IL-4 are correlated with tuberculosis (TB) susceptibility and progression, we investigated the role of DC-SIGN in M(IL-4) macrophages in the TB context. First, we demonstrate that DC-SIGN expression is present both in CD68+ macrophages found in tuberculous pulmonary lesions of non-human primates, and in the CD14+ cell population isolated from pleural effusions obtained from TB patients (TB-PE). Likewise, we show that DC-SIGN expression is accentuated in M(IL-4) macrophages derived from peripheral blood CD14+ monocytes isolated from TB patients, or in macrophages stimulated with acellular TB-PE, arguing for the pertinence of DC-SIGN-expressing macrophages in TB. Second, using a siRNA-mediated gene silencing approach, we performed a transcriptomic analysis of DC-SIGN-depleted M(IL-4) macrophages and revealed the upregulation of pro-inflammatory signals in response to challenge with Mtb, as compared to control cells. This pro-inflammatory gene signature was confirmed by RT-qPCR, cytokine/chemokine-based protein array, and ELISA analyses. We also found that inactivation of DC-SIGN renders M(IL-4) macrophages less permissive to Mtb intracellular growth compared to control cells, despite the equal level of bacteria uptake. Last, at the molecular level, we show that DC-SIGN interferes negatively with the pro-inflammatory response and control of Mtb intracellular growth mediated by another CLR, Dectin-1 (CLEC7A). Collectively, this study highlights a dual role for DC-SIGN as, on the one hand, being a host factor granting advantage for Mtb to parasitize macrophages and, on the other hand, representing a molecular switch to turn off the pro-inflammatory response in these cells to prevent potential immunopathology associated to TB.
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Moléculas de Adhesión Celular/metabolismo , Lectinas Tipo C/metabolismo , Macrófagos/inmunología , Macrófagos/metabolismo , Mycobacterium tuberculosis/inmunología , Receptores de Superficie Celular/metabolismo , Tuberculosis/inmunología , Tuberculosis/metabolismo , Animales , Moléculas de Adhesión Celular/genética , Supervivencia Celular/genética , Supervivencia Celular/inmunología , Citocinas/metabolismo , Femenino , Expresión Génica , Interacciones Huésped-Patógeno/genética , Interacciones Huésped-Patógeno/inmunología , Humanos , Mediadores de Inflamación/metabolismo , Lectinas Tipo C/genética , Macaca mulatta , Macrófagos/microbiología , Monocitos/inmunología , Monocitos/metabolismo , Fagocitosis/inmunología , Receptores de Superficie Celular/genética , Tuberculosis/genética , Tuberculosis/microbiologíaRESUMEN
BACKGROUND: Epidemiological evidence suggests a link between pesticide exposure and the development of metabolic diseases. However, most experimental studies have evaluated the metabolic effects of pesticides using individual molecules, often at nonrelevant doses or in combination with other risk factors such as high-fat diets. OBJECTIVES: We aimed to evaluate, in mice, the metabolic consequences of chronic dietary exposure to a pesticide mixture at nontoxic doses, relevant to consumers' risk assessment. METHODS: A mixture of six pesticides commonly used in France, i.e., boscalid, captan, chlorpyrifos, thiofanate, thiacloprid, and ziram, was incorporated in a standard chow at doses exposing mice to the tolerable daily intake (TDI) of each pesticide. Wild-type (WT) and constitutive androstane receptor-deficient (CAR-/-) male and female mice were exposed for 52 wk. We assessed metabolic parameters [body weight (BW), food and water consumption, glucose tolerance, urinary metabolome] throughout the experiment. At the end of the experiment, we evaluated liver metabolism (histology, transcriptomics, metabolomics, lipidomics) and pesticide detoxification using liquid chromatography-mass spectrometry (LC-MS). RESULTS: Compared to those fed control chow, WT male mice fed pesticide chow had greater BW gain and more adiposity. Moreover, these WT males fed pesticide chow exhibited characteristics of hepatic steatosis and glucose intolerance, which were not observed in those fed control chow. WT exposed female mice exhibited fasting hyperglycemia, higher reduced glutathione (GSH):oxidized glutathione (GSSG) liver ratio and perturbations of gut microbiota-related urinary metabolites compared to WT mice fed control chow. When we performed these experiments on CAR-/- mice, pesticide-exposed CAR-/- males did not exhibit BW gain or changes in glucose metabolism compared to the CAR-/- males fed control chow. Moreover, CAR-/- females fed pesticide chow exhibited pesticide toxicity with higher BWs and mortality rate compared to the CAR-/- females fed control chow. CONCLUSIONS: To our knowledge, we are the first to demonstrate a sexually dimorphic obesogenic and diabetogenic effect of chronic dietary exposure to a common mixture of pesticides at TDI levels, and to provide evidence for a partial role for CAR in an in vivo mouse model. This raises questions about the relevance of TDI for individual pesticides when present in a mixture. https://doi.org/10.1289/EHP2877.
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Fungicidas Industriales/toxicidad , Trastornos del Metabolismo de la Glucosa/inducido químicamente , Insecticidas/toxicidad , Receptores Citoplasmáticos y Nucleares/genética , Animales , Animales Modificados Genéticamente , Peso Corporal/efectos de los fármacos , Receptor de Androstano Constitutivo , Exposición Dietética , Hígado Graso/inducido químicamente , Femenino , Glutatión/metabolismo , Inactivación Metabólica , Hígado/efectos de los fármacos , Hígado/metabolismo , Masculino , Metaboloma/efectos de los fármacos , Ratones , Ratones Endogámicos C57BL , Factores Sexuales , Pruebas de Toxicidad CrónicaRESUMEN
Bone deficits are frequent in HIV-1-infected patients. We report here that osteoclasts, the cells specialized in bone resorption, are infected by HIV-1 in vivo in humanized mice and ex vivo in human joint biopsies. In vitro, infection of human osteoclasts occurs at different stages of osteoclastogenesis via cell-free viruses and, more efficiently, by transfer from infected T cells. HIV-1 infection markedly enhances adhesion and osteolytic activity of human osteoclasts by modifying the structure and function of the sealing zone, the osteoclast-specific bone degradation machinery. Indeed, the sealing zone is broader due to F-actin enrichment of its basal units (i.e., the podosomes). The viral protein Nef is involved in all HIV-1-induced effects partly through the activation of Src, a regulator of podosomes and of their assembly as a sealing zone. Supporting these results, Nef-transgenic mice exhibit an increased osteoclast density and bone defects, and osteoclasts derived from these animals display high osteolytic activity. Altogether, our study evidences osteoclasts as host cells for HIV-1 and their pathological contribution to bone disorders induced by this virus, in part via Nef.
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Resorción Ósea/etiología , Infecciones por VIH/complicaciones , VIH-1/fisiología , Osteoclastos/virología , Actinas/metabolismo , Animales , Resorción Ósea/metabolismo , Resorción Ósea/patología , Resorción Ósea/fisiopatología , Huesos/metabolismo , Adhesión Celular , Femenino , Infecciones por VIH/metabolismo , Infecciones por VIH/patología , Infecciones por VIH/virología , VIH-1/genética , Humanos , Ratones , Osteoclastos/citología , Osteoclastos/metabolismo , Productos del Gen nef del Virus de la Inmunodeficiencia Humana/genética , Productos del Gen nef del Virus de la Inmunodeficiencia Humana/metabolismoRESUMEN
RATIONALE: In addition to their well-known function as antibody-producing cells, B lymphocytes can markedly influence the course of infectious or noninfectious diseases via antibody-independent mechanisms. In tuberculosis (TB), B cells accumulate in lungs, yet their functional contribution to the host response remains poorly understood. OBJECTIVES: To document the role of B cells in TB in an unbiased manner. METHODS: We generated the transcriptome of B cells isolated from Mycobacterium tuberculosis (Mtb)-infected mice and validated the identified key pathways using in vitro and in vivo assays. The obtained data were substantiated using B cells from pleural effusion of patients with TB. MEASUREMENTS AND MAIN RESULTS: B cells isolated from Mtb-infected mice displayed a STAT1 (signal transducer and activator of transcription 1)-centered signature, suggesting a role for IFNs in B-cell response to infection. B cells stimulated in vitro with Mtb produced type I IFN, via a mechanism involving the innate sensor STING (stimulator of interferon genes), and antagonized by MyD88 (myeloid differentiation primary response 88) signaling. In vivo, B cells expressed type I IFN in the lungs of Mtb-infected mice and, of clinical relevance, in pleural fluid from patients with TB. Type I IFN expression by B cells induced an altered polarization of macrophages toward a regulatory/antiinflammatory profile in vitro. In vivo, increased provision of type I IFN by B cells in a murine model of B cell-restricted Myd88 deficiency correlated with an enhanced accumulation of regulatory/antiinflammatory macrophages in Mtb-infected lungs. CONCLUSIONS: Type I IFN produced by Mtb-stimulated B cells favors macrophage polarization toward a regulatory/antiinflammatory phenotype during Mtb infection.
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Linfocitos B/metabolismo , Interferón Tipo I/metabolismo , Macrófagos/metabolismo , Tuberculosis/metabolismo , Animales , Modelos Animales de Enfermedad , Humanos , Pulmón/metabolismo , Pulmón/microbiología , Ratones , Ratones Endogámicos C57BL , Mycobacterium tuberculosis , Transducción de Señal , Bazo/metabolismo , Bazo/microbiologíaRESUMEN
Breast cancer (BC) remains the primary cause of death from cancer among women worldwide. Cholesterol-5,6-epoxide (5,6-EC) metabolism is deregulated in BC but the molecular origin of this is unknown. Here, we have identified an oncometabolism downstream of 5,6-EC that promotes BC progression independently of estrogen receptor α expression. We show that cholesterol epoxide hydrolase (ChEH) metabolizes 5,6-EC into cholestane-3ß,5α,6ß-triol, which is transformed into the oncometabolite 6-oxo-cholestan-3ß,5α-diol (OCDO) by 11ß-hydroxysteroid-dehydrogenase-type-2 (11ßHSD2). 11ßHSD2 is known to regulate glucocorticoid metabolism by converting active cortisol into inactive cortisone. ChEH inhibition and 11ßHSD2 silencing inhibited OCDO production and tumor growth. Patient BC samples showed significant increased OCDO levels and greater ChEH and 11ßHSD2 protein expression compared with normal tissues. The analysis of several human BC mRNA databases indicated that 11ßHSD2 and ChEH overexpression correlated with a higher risk of patient death, highlighting that the biosynthetic pathway producing OCDO is of major importance to BC pathology. OCDO stimulates BC cell growth by binding to the glucocorticoid receptor (GR), the nuclear receptor of endogenous cortisol. Interestingly, high GR expression or activation correlates with poor therapeutic response or prognosis in many solid tumors, including BC. Targeting the enzymes involved in cholesterol epoxide and glucocorticoid metabolism or GR may be novel strategies to prevent and treat BC.
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Neoplasias de la Mama/metabolismo , Carcinógenos/metabolismo , Colesterol/metabolismo , Receptores de Glucocorticoides/metabolismo , 11-beta-Hidroxiesteroide Deshidrogenasa de Tipo 2/metabolismo , Animales , Línea Celular , Línea Celular Tumoral , Colesterol/análogos & derivados , Epóxido Hidrolasas/metabolismo , Receptor alfa de Estrógeno/metabolismo , Femenino , Células HEK293 , Humanos , Células MCF-7 , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Desnudos , ARN Mensajero/metabolismoRESUMEN
HLA-A*0201/DRB1*0101 transgenic mice (A2/DR1 mice) have been developed to study the immunogenicity of tumor antigen-derived T cell epitopes. To extend the use and application of this mouse model in the field of antitumor immunotherapy, we described a tumor cell line generated from a naturally occurring tumor in A2/DR1 mouse named SARC-L1. Histological and genes signature analysis supported the sarcoma origin of this cell line. While SARC-L1 tumor cells lack HLA-DRB1*0101 expression, a very low expression of HLA-A*0201 molecules was found on these cells. Furthermore they also weakly but constitutively expressed the programmed death-ligand 1 (PD-L1). Interestingly both HLA-A*0201 and PD-L1 expressions can be increased on SARC-L1 after IFN-γ exposure in vitro. We also obtained two genetically modified cell lines highly expressing either HLA-A*0201 or both HLA-A*0201/ HLA-DRB1*0101 molecules referred as SARC-A2 and SARC-A2DR1 respectively. All the SARC-L1-derived cell lines induced aggressive subcutaneous tumors in A2DR1 mice in vivo. The analysis of SARC-L1 tumor microenvironment revealed a strong infiltration by T cells expressing inhibitory receptors such as PD-1 and TIM-3. Finally, we found that SARC-L1 is sensitive to several drugs commonly used to treat sarcoma and also susceptible to anti-PD-L1 monoclonal antibody therapy in vivo. Collectively, we described a novel syngeneic tumor model A2/DR1 mice that could be used as preclinical tool for the evaluation of antitumor immunotherapies.
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Antígeno B7-H1/genética , Antígeno HLA-A2/genética , Cadenas HLA-DRB1/genética , Neoplasias/genética , Animales , Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Apoptosis/genética , Apoptosis/inmunología , Antígeno B7-H1/metabolismo , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Modelos Animales de Enfermedad , Femenino , Expresión Génica , Perfilación de la Expresión Génica , Antígeno HLA-A2/inmunología , Cadenas HLA-DRB1/inmunología , Humanos , Linfocitos Infiltrantes de Tumor/efectos de los fármacos , Linfocitos Infiltrantes de Tumor/inmunología , Linfocitos Infiltrantes de Tumor/metabolismo , Masculino , Ratones , Ratones Transgénicos , Neoplasias/inmunología , Neoplasias/metabolismo , Neoplasias/patología , Sarcoma/genética , Sarcoma/metabolismo , Sarcoma/patologíaRESUMEN
Immune response against pathogens is a tightly regulated process that must ensure microbial control while preserving integrity of the infected organs. Tuberculosis (TB) is a paramount example of a chronic infection in which antimicrobial immunity is protective in the vast majority of infected individuals but can become detrimental if not finely tuned. Here, we report that C-type lectin dendritic cell (DC) immunoreceptor (DCIR), a key component in DC homeostasis, is required to modulate lung inflammation and bacterial burden in TB. DCIR is abundantly expressed in pulmonary lesions in Mycobacterium tuberculosis-infected nonhuman primates during both latent and active disease. In mice, we found that DCIR deficiency impairs STAT1-mediated type I IFN signaling in DCs, leading to increased production of IL-12 and increased differentiation of T lymphocytes toward Th1 during infection. As a consequence, DCIR-deficient mice control M. tuberculosis better than WT animals but also develop more inflammation characterized by an increased production of TNF and inducible NOS (iNOS) in the lungs. Altogether, our results reveal a pathway by which a C-type lectin modulates the equilibrium between infection-driven inflammation and pathogen's control through sustaining type I IFN signaling in DCs.
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Células Dendríticas/inmunología , Interferón Tipo I/inmunología , Lectinas Tipo C/inmunología , Tuberculosis/inmunología , Animales , Femenino , Lectinas Tipo C/genética , Macaca mulatta , Ratones Endogámicos C57BL , Ratones Noqueados , Fosforilación , Factor de Transcripción STAT1/inmunología , Transducción de SeñalRESUMEN
BACKGROUND: Intestinal absorption of dietary lipids involves their hydrolysis in the lumen of proximal intestine as well as uptake, intracellular transport and re-assembly of hydrolyzed lipids in enterocytes, leading to the formation and secretion of the lipoproteins chylomicrons and HDL. In this study, we examined the potential involvement of cytosolic lipid droplets (CLD) whose function in the process of lipid absorption is poorly understood. METHODS: Intestinal lipid absorption was studied in mouse after gavage. Three populations of CLD were purified by density ultracentrifugations, as well as the brush border membranes, which were analyzed by western-blots. Immunofluorescent localization of membranes transporters or metabolic enzymes, as well as kinetics of CLD production, were also studied in intestine or Caco-2 cells. RESULTS: We isolated three populations of CLD (ranging from 15 to 1000 nm) which showed differential expression of the major lipid transporters scavenger receptor BI (SR-BI), cluster of differentiation 36 (CD-36), Niemann Pick C-like 1 (NPC1L1), and the ATP-binding cassette transporters ABCG5/G8 but also caveolin 2 and fatty acid binding proteins. The enzyme monoacylglycerol acyltransferase 2 (MGAT2) was identified in the brush border membrane (BBM) in addition to the endoplasmic reticulum, suggesting local synthesis of triglycerides and CLD at both places. CONCLUSIONS: We show a very fast production of CLD by enterocytes associated with a transfer of apical constituents as lipid transporters. Our findings suggest that following their uptake by enterocytes, lipids can be partially metabolized at the BBM and packaged into CLD for their transportation to the ER.
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OBJECTIVE: Peroxisome proliferator-activated receptor α (PPARα) is a nuclear receptor expressed in tissues with high oxidative activity that plays a central role in metabolism. In this work, we investigated the effect of hepatocyte PPARα on non-alcoholic fatty liver disease (NAFLD). DESIGN: We constructed a novel hepatocyte-specific PPARα knockout (Pparα(hep-/-)) mouse model. Using this novel model, we performed transcriptomic analysis following fenofibrate treatment. Next, we investigated which physiological challenges impact on PPARα. Moreover, we measured the contribution of hepatocytic PPARα activity to whole-body metabolism and fibroblast growth factor 21 production during fasting. Finally, we determined the influence of hepatocyte-specific PPARα deficiency in different models of steatosis and during ageing. RESULTS: Hepatocyte PPARα deletion impaired fatty acid catabolism, resulting in hepatic lipid accumulation during fasting and in two preclinical models of steatosis. Fasting mice showed acute PPARα-dependent hepatocyte activity during early night, with correspondingly increased circulating free fatty acids, which could be further stimulated by adipocyte lipolysis. Fasting led to mild hypoglycaemia and hypothermia in Pparα(hep-/-) mice when compared with Pparα(-/-) mice implying a role of PPARα activity in non-hepatic tissues. In agreement with this observation, Pparα(-/-) mice became overweight during ageing while Pparα(hep-/-) remained lean. However, like Pparα(-/-) mice, Pparα(hep-/-) fed a standard diet developed hepatic steatosis in ageing. CONCLUSIONS: Altogether, these findings underscore the potential of hepatocyte PPARα as a drug target for NAFLD.
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Envejecimiento , Ácidos Grasos/metabolismo , Factores de Crecimiento de Fibroblastos/genética , Hepatocitos , Enfermedad del Hígado Graso no Alcohólico/genética , PPAR alfa/genética , Adipocitos , Envejecimiento/fisiología , Animales , Sistema Enzimático del Citocromo P-450/genética , Familia 4 del Citocromo P450/genética , Modelos Animales de Enfermedad , Ayuno , Fenofibrato/farmacología , Factores de Crecimiento de Fibroblastos/biosíntesis , Expresión Génica/efectos de los fármacos , Perfilación de la Expresión Génica , Homeostasis/genética , Hipoglucemia/genética , Hipolipemiantes/farmacología , Hipotermia/genética , Metabolismo de los Lípidos/genética , Lipólisis/genética , Masculino , Ratones Endogámicos C57BL , Ratones Noqueados , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Sobrepeso/genética , PPAR alfa/metabolismo , ARN Mensajero/metabolismo , Triglicéridos/metabolismoRESUMEN
The human CD14(+) monocyte compartment is composed by two subsets based on CD16 expression. We previously reported that this compartment is perturbed in tuberculosis (TB) patients, as reflected by the expansion of CD16(+) monocytes along with disease severity. Whether this unbalance is beneficial or detrimental to host defense remains to be elucidated. Here in the context of active TB, we demonstrate that human monocytes are predisposed to differentiate towards an anti-inflammatory (M2-like) macrophage activation program characterized by the CD16(+)CD163(+)MerTK(+)pSTAT3(+) phenotype and functional properties such as enhanced protease-dependent motility, pathogen permissivity and immunomodulation. This process is dependent on STAT3 activation, and loss-of-function experiments point towards a detrimental role in host defense against TB. Importantly, we provide a critical correlation between the abundance of the CD16(+)CD163(+)MerTK(+)pSTAT3(+) cells and the progression of the disease either at the local level in a non-human primate tuberculous granuloma context, or at the systemic level through the detection of the soluble form of CD163 in human sera. Collectively, this study argues for the pathogenic role of the CD16(+)CD163(+)MerTK(+)pSTAT3(+) monocyte-to-macrophage differentiation program and its potential as a target for TB therapy, and promotes the detection of circulating CD163 as a potential biomarker for disease progression and monitoring of treatment efficacy.
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Inmunomodulación , Interleucina-10/metabolismo , Monocitos/inmunología , Monocitos/patología , Receptores de IgG/metabolismo , Factor de Transcripción STAT3/metabolismo , Tuberculosis/inmunología , Tuberculosis/patología , HumanosRESUMEN
BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal malignancies with a mortality that is almost identical to incidence. Because early detected PDAC is potentially curable, blood-based biomarkers that could detect currently developing neoplasia would improve patient survival and management. PDAC develops from pancreatic intraepithelial neoplasia (PanIN) lesions, graded from low grade (PanIN1) to high grade (PanIN3). We made the hypothesis that specific proteomic signatures from each precancerous stage exist and are detectable in plasma. METHODS: We explored the peptide profiles of microdissected PanIN cells and of plasma samples corresponding to the different PanIN grade from genetically engineered mouse models of PDAC using capillary electrophoresis coupled to mass spectrometry (CE-MS) and Chip-MS/MS. RESULTS: We successfully characterised differential peptides profiles from PanIN microdissected cells. We found that plasma from tumor-bearing mice and age-matched controls exhibit discriminative peptide signatures. We also determined plasma peptide signatures corresponding to low- and high-grade precancerous step present in the mice pancreas using the two mass spectrometry technologies. Importantly, we identified biomarkers specific of PanIN3. CONCLUSIONS: We demonstrate that benign and advanced PanIN lesions display distinct plasma peptide patterns. This strongly supports the perspectives of developing a non-invasive screening test for prediction and early detection of PDAC.
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Biomarcadores de Tumor/sangre , Carcinoma in Situ/sangre , Carcinoma Ductal Pancreático/sangre , Proteínas de Neoplasias/sangre , Neoplasias Pancreáticas/sangre , Péptidos/sangre , Lesiones Precancerosas/sangre , Animales , Biomarcadores de Tumor/análisis , Carcinoma in Situ/química , Carcinoma in Situ/patología , Carcinoma Ductal Pancreático/química , Modelos Animales de Enfermedad , Ratones , Proteínas de Neoplasias/análisis , Neoplasias Pancreáticas/química , Péptidos/análisis , Lesiones Precancerosas/química , Lesiones Precancerosas/patología , Análisis por Matrices de Proteínas , Proteoma/análisisRESUMEN
Models of microbe-elicited peritonitis have been invaluable to identify mechanisms underlying inflammation resolution, but whether resolution mechanisms differ from an inflammatory agent to another has not been determined. Thus, we analyzed the cellular and molecular components of the resolution phase of non-microbe-induced inflammation. In thioglycollate (TG)-induced peritonitis, resolution started at 12 h (Tmax) and displayed a 22 h resolution interval (Ri). During resolution, lipoxin A4, resolvin (Rv) D1 and RvD2, protectin D1 (PD1), and maresin 1 (MaR1) were transiently produced while RvD5 was continually generated. In addition, docosahexaenoic acid (DHA)-derived mediators were produced to a higher extent than in microbial peritonitis. We also investigated leukocyte infiltration and clearance in peritoneal tissues surrounding the inflammatory site. In the omentum, resolution parameters, neutrophil apoptosis, and efferocytosis were similar to those of the peritoneal cavity. However, we noticed long-term persistence of M2-polarized macrophages and B-lymphocytes in the omentum after TG administration, whereas zymosan injection caused M1/M2-macrophage and T-lymphocyte persistence regardless of the magnitude of the inflammatory response. Our study indicates that some aspects of resolution are shaped in a stimulus-specific manner, and it ultimately argues that the tissues surrounding the inflammatory site must also be considered to address the inflammatory response globally.
Asunto(s)
Linfocitos B/inmunología , Inflamación/inmunología , Leucocitos/inmunología , Macrófagos/inmunología , Peritonitis/inmunología , Peritonitis/metabolismo , Tioglicolatos/toxicidad , Animales , Apoptosis/inmunología , Linfocitos B/metabolismo , Linfocitos B/patología , Western Blotting , Células Cultivadas , Ácidos Docosahexaenoicos/genética , Ácidos Docosahexaenoicos/metabolismo , Femenino , Citometría de Flujo , Técnicas para Inmunoenzimas , Inflamación/metabolismo , Inflamación/patología , Leucocitos/metabolismo , Leucocitos/patología , Lípidos/análisis , Macrófagos/metabolismo , Macrófagos/patología , Ratones , Ratones Endogámicos C57BL , Epiplón/inmunología , Epiplón/metabolismo , Epiplón/patología , Peritonitis/inducido químicamente , Fagocitosis/fisiología , ARN Mensajero/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Zimosan/toxicidadRESUMEN
In osteoclasts, Src controls podosome organization and bone degradation, which leads to an osteopetrotic phenotype in src(-/-) mice. Since this phenotype was even more severe in src(-/-)hck(-/-) mice, we examined the individual contribution of Hck in bone homeostasis. Compared to wt mice, hck(-/-) mice exhibited an osteopetrotic phenotype characterized by an increased density of trabecular bone and decreased bone degradation, although osteoclastogenesis was not impaired. Podosome organization and matrix degradation were found to be defective in hck(-/-) osteoclast precursors (preosteoclast) but were normal in mature hck(-/-) osteoclasts, probably through compensation by Src, which was specifically overexpressed in mature osteoclasts. As a consequence of podosome defects, the 3-dimensional migration of hck(-/-) preosteoclasts was strongly affected in vitro. In vivo, this translated by altered bone homing of preosteoclasts in hck(-/-) mice: in metatarsals of 1-wk-old mice, when bone formation strongly depends on the recruitment of these cells, reduced numbers of osteoclasts and abnormal developing trabecular bone were observed. This phenotype was still detectable in adults. In summmary, Hck is one of the very few effectors of preosteoclast recruitment described to date and thereby plays a critical role in bone remodeling.
Asunto(s)
Huesos/citología , Huesos/metabolismo , Movimiento Celular/fisiología , Osteoclastos/citología , Osteopetrosis/metabolismo , Proteínas Proto-Oncogénicas c-hck/metabolismo , Animales , Movimiento Celular/genética , Células Cultivadas , Femenino , Homeostasis/genética , Homeostasis/fisiología , Masculino , Ratones , Ratones Noqueados , Osteoclastos/metabolismo , Osteopetrosis/genética , Proteínas Proto-Oncogénicas c-hck/genética , Familia-src Quinasas/genética , Familia-src Quinasas/metabolismoRESUMEN
Tumor protein p53-induced nuclear protein 1 (TP53INP1) is involved in cell stress response. Its expression is lost at the pancreatic intraepithelial neoplasia 1b (PanIN1b)/PanIN2 stage of pancreatic carcinogenesis. Our objective was to determine whether TP53INP1 loss of expression contributes to pancreatic cancer formation in a conditional KrasG12D mouse model. We generated Kras-INP1KO mice using LSL-Kras(G12D/+);Pdx1-Cre(+/-) mice (Kras mice) and TP53INP1(-/-) mice. Analysis of pancreases during ageing shows that in the presence of activated Kras, TP53INP1 loss of expression accelerated PanIN formation and increased pancreatic injury and the number of high-grade lesions as compared with what occurs in Kras mice. Moreover, cystic lesions resembling intraductal papillary mucinous neoplasm (IPMN) were observed as early as 2 months of age. Remarkably, TP53INP1 is down-regulated in human IPMN. Activation of the small GTPase Rac1 shows that more oxidative stress is generated in Kras-INP1KO than in Kras mice pancreas despite elevated levels of the Nrf2 antioxidant regulator. We firmly establish the link between Kras-INP1KO pancreatic phenotype and oxidative stress with rescue of the phenotype by the antioxidant action of N-acetylcysteine. Our data provide in vivo functional demonstration that TP53INP1 deficiency accelerates progression of pancreatic cancer, underlining its role in the occurrence of IPMN and highlighting the importance of TP53INP1 in the control of oxidative status during development of pancreatic cancer.
Asunto(s)
Proteínas Nucleares/fisiología , Neoplasias Pancreáticas/metabolismo , Proteínas Proto-Oncogénicas p21(ras)/genética , Animales , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/metabolismo , Carcinoma Ductal Pancreático/patología , Transformación Celular Neoplásica/genética , Transformación Celular Neoplásica/metabolismo , Progresión de la Enfermedad , Regulación hacia Abajo/fisiología , Humanos , Metaplasia/genética , Metaplasia/metabolismo , Ratones , Ratones Noqueados , Proteínas Nucleares/deficiencia , Proteínas Nucleares/genética , Estrés Oxidativo/genética , Estrés Oxidativo/fisiología , Páncreas/patología , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patología , Lesiones Precancerosas/genética , Lesiones Precancerosas/metabolismoRESUMEN
BACKGROUND & AIMS: Nutrients influence non-alcoholic fatty liver disease. Essential fatty acids deficiency promotes various syndromes, including hepatic steatosis, through increased de novo lipogenesis. The mechanisms underlying such increased lipogenic response remain unidentified. METHODS: We used wild type mice and mice lacking Liver X Receptors to perform a nutrigenomic study that aimed at examining the role of these transcription factors. RESULTS: We showed that, in the absence of Liver X Receptors, essential fatty acids deficiency does not promote steatosis. Consistent with this, Liver X Receptors are required for the elevated expression of genes involved in lipogenesis in response to essential fatty acids deficiency. CONCLUSIONS: This work identifies, for the first time, the central role of Liver X Receptors in steatosis induced by essential fatty acids deficiency.
Asunto(s)
Ácidos Grasos Esenciales/deficiencia , Hígado Graso/fisiopatología , Expresión Génica/fisiología , Lipogénesis/genética , Lipogénesis/fisiología , Receptores Nucleares Huérfanos/fisiología , Animales , Colesterol/metabolismo , Enfermedades Carenciales/fisiopatología , Grasas de la Dieta/farmacología , Modelos Animales de Enfermedad , Femenino , Expresión Génica/efectos de los fármacos , Lipogénesis/efectos de los fármacos , Hígado/metabolismo , Receptores X del Hígado , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Transgénicos , Receptores Nucleares Huérfanos/deficiencia , Receptores Nucleares Huérfanos/genética , Factores de Transcripción/fisiología , Triglicéridos/metabolismo , Regulación hacia Arriba/fisiologíaRESUMEN
We have recently discovered the existence of 5α-Hydroxy-6ß-[2-(1H-imidazol-4-yl)ethylamino]cholestan-3ß-ol, called Dendrogenin A (DDA), as the first endogenous steroidal alkaloid ever described in mammals. We found that the DDA content of tumors and cancer cell lines was low or absent compared with normal cells showing that a deregulation in DDA biosynthesis was associated with cancer and therefore suggesting that DDA could represent a metabolomic cancer biomarker. This prompted us to produce antibodies that selectively recognize DDA. For this purpose, the hapten 5α-hydroxy-6ß-[2-(1H-imidazol-4-yl)ethylamino]cholestan-3ß-o-hemisuccinate with a carboxylic spacer arm attached to the 3ß-hydroxyl group of DDA was synthesized. The hapten was coupled to bovine serum albumin and keyhole limpet hemocyanin for antibody production to develop an enzyme-linked immunosorbent assay (ELISA). The protein conjugates were injected into BALB/c mice to raise antibodies. The monoclonal antibodies that were secreted from the hybridoma cell lines established were assessed with indirect ELISA by competitive assays using dilutions of a DDA standard. The antibodies from the selected hybridomas had an IC(50) value ranging from 0.8 to 425 ng/ml. Three antibodies showed no cross-reactivity with structurally related compounds including histamine, cholesterol, ring B oxysterols and a regio-isomer of DDA. In this study, high-affinity and selective antibodies against DDA were produced for the first time, and a competitive indirect ELISA was developed.