RESUMEN
The immune response is an energy-demanding process that must be coordinated with systemic metabolic changes redirecting nutrients from stores to the immune system. Although this interplay is fundamental for the function of the immune system, the underlying mechanisms remain elusive. Our data show that the pro-inflammatory polarization of Drosophila macrophages is coupled to the production of the insulin antagonist ImpL2 through the activity of the transcription factor HIF1α. ImpL2 production, reflecting nutritional demands of activated macrophages, subsequently impairs insulin signaling in the fat body, thereby triggering FOXO-driven mobilization of lipoproteins. This metabolic adaptation is fundamental for the function of the immune system and an individual's resistance to infection. We demonstrated that analogically to Drosophila, mammalian immune-activated macrophages produce ImpL2 homolog IGFBP7 in a HIF1α-dependent manner and that enhanced IGFBP7 production by these cells induces mobilization of lipoproteins from hepatocytes. Hence, the production of ImpL2/IGFBP7 by macrophages represents an evolutionarily conserved mechanism by which macrophages alleviate insulin signaling in the central metabolic organ to secure nutrients necessary for their function upon bacterial infection.
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Infecciones Bacterianas , Proteínas de Drosophila , Resistencia a la Insulina , Animales , Antagonistas de Insulina/metabolismo , Antagonistas de Insulina/farmacología , Drosophila/metabolismo , Insulina/metabolismo , Macrófagos/metabolismo , Infecciones Bacterianas/metabolismo , Mamíferos , Proteínas de Unión a Factor de Crecimiento Similar a la Insulina/metabolismo , Proteínas de Drosophila/metabolismoRESUMEN
Although multiple populations of macrophages have been described in the human liver, their function and turnover in patients with obesity at high risk of developing non-alcoholic fatty liver disease (NAFLD) and cirrhosis are currently unknown. Herein, we identify a specific human population of resident liver myeloid cells that protects against the metabolic impairment associated with obesity. By studying the turnover of liver myeloid cells in individuals undergoing liver transplantation, we find that liver myeloid cell turnover differs between humans and mice. Using single-cell techniques and flow cytometry, we determine that the proportion of the protective resident liver myeloid cells, denoted liver myeloid cells 2 (LM2), decreases during obesity. Functional validation approaches using human 2D and 3D cultures reveal that the presence of LM2 ameliorates the oxidative stress associated with obese conditions. Our study indicates that resident myeloid cells could be a therapeutic target to decrease the oxidative stress associated with NAFLD.
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Enfermedad del Hígado Graso no Alcohólico , Humanos , Ratones , Animales , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Obesidad/metabolismo , Células Mieloides/metabolismo , Estrés FisiológicoRESUMEN
Adipocyte hypertrophy and expression of adipokines in subcutaneous adipose tissue (SAT) have been linked to steatosis, nonalcoholic steatohepatitis (NASH) and fibrosis in morbidly obese (BMI ≥ 40 kg/m2) subjects. It is unknown if this is also true for subjects with NAFLD with lesser degrees of obesity (BMI < 35 kg/m2). Thirty-two subjects with biopsy-proven NAFLD and 15 non-diabetic controls matched for BMI underwent fine-needle biopsies of SAT. Adipocyte volume was calculated. RNA-sequencing of SAT was performed in a subset of 20 NAFLD patients. Adipocyte volume and gene expression levels were correlated to the presence of NASH or significant fibrosis. Subjects with NAFLD had larger adipocyte volume compared with controls, (1939 pL, 95% CI 1130-1662 vs. 854 pL, 95% CI 781-926, p < 0.001). There was no association between adipocyte volume and the presence of NASH. Gene expression of adipokines previously described to correlate with NASH in morbid obesity, was not associated with NASH or fibrosis. Our results suggest that persons with NAFLD have larger SAT adipocytes compared with controls and that adipocytes are involved in the pathophysiology of hepatic steatosis in NAFLD. However, adipocyte volume was not associated with NASH or fibrosis in NAFLD subjects with varying degrees of obesity.
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Enfermedad del Hígado Graso no Alcohólico , Obesidad Mórbida , Humanos , Enfermedad del Hígado Graso no Alcohólico/complicaciones , Obesidad Mórbida/complicaciones , Adipoquinas , Adipocitos , Biopsia con Aguja Fina , Fibrosis , HipertrofiaRESUMEN
Macrophages have diverse phenotypes and functions due to differences in their origin, location and pathophysiological context. Although their main role in the liver has been described as immunoregulatory and detoxifying, changes in macrophage phenotypes, diversity, dynamics and function have been reported during obesity-related complications such as non-alcoholic fatty liver disease (NAFLD). NAFLD encompasses multiple disease states from hepatic steatosis to non-alcoholic steatohepatitis (NASH), fibrosis, cirrhosis and hepatocarcinoma. Obesity and insulin resistance are prominent risk factors for NASH, a disease with a high worldwide prevalence and no approved treatment. In this Review, we discuss the turnover and function of liver-resident macrophages (Kupffer cells) and monocyte-derived hepatic macrophages. We examine these populations in both steady state and during NAFLD, with an emphasis on NASH. The explosion in high-throughput gene expression analysis using single-cell RNA sequencing (scRNA-seq) within the last 5 years has revolutionized the study of macrophage heterogeneity, substantially increasing our understanding of the composition and diversity of tissue macrophages, including in the liver. Here, we highlight scRNA-seq findings from the last 5 years on the diversity of liver macrophages in homeostasis and metabolic disease, and reveal hepatic macrophage function beyond their classically described inflammatory role in the progression of NAFLD and NASH pathogenesis.
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Enfermedad del Hígado Graso no Alcohólico , Animales , Humanos , Inflamación/metabolismo , Hígado/metabolismo , Macrófagos/metabolismo , Ratones , Ratones Endogámicos C57BL , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Obesidad/metabolismoRESUMEN
Natural killer (NK) cells are innate immune cells that contribute to host defense against virus infections. NK cells respond to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in vitro and are activated in patients with acute coronavirus disease 2019 (COVID-19). However, by which mechanisms NK cells detect SARS-CoV-2-infected cells remains largely unknown. Here, we show that the Non-structural protein 13 of SARS-CoV-2 encodes for a peptide that is presented by human leukocyte antigen E (HLA-E). In contrast with self-peptides, the viral peptide prevents binding of HLA-E to the inhibitory receptor NKG2A, thereby rendering target cells susceptible to NK cell attack. In line with these observations, NKG2A-expressing NK cells are particularly activated in patients with COVID-19 and proficiently limit SARS-CoV-2 replication in infected lung epithelial cells in vitro. Thus, these data suggest that a viral peptide presented by HLA-E abrogates inhibition of NKG2A+ NK cells, resulting in missing self-recognition.
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COVID-19 , Antígenos de Histocompatibilidad Clase I , Células Asesinas Naturales , Metiltransferasas , Subfamília C de Receptores Similares a Lectina de Células NK , ARN Helicasas , SARS-CoV-2 , Proteínas no Estructurales Virales , COVID-19/inmunología , Antígenos de Histocompatibilidad Clase I/inmunología , Humanos , Células Asesinas Naturales/inmunología , Metiltransferasas/inmunología , Subfamília C de Receptores Similares a Lectina de Células NK/inmunología , Subfamília C de Receptores Similares a Lectina de Células NK/metabolismo , Péptidos/metabolismo , ARN Helicasas/inmunología , Proteínas no Estructurales Virales/inmunología , Antígenos HLA-ERESUMEN
The mechanisms promoting disturbed white adipocyte function in obesity remain largely unclear. Herein, we integrate white adipose tissue (WAT) metabolomic and transcriptomic data from clinical cohorts and find that the WAT phosphocreatine/creatine ratio is increased and creatine kinase-B expression and activity is decreased in the obese state. In human in vitro and murine in vivo models, we demonstrate that decreased phosphocreatine metabolism in white adipocytes alters adenosine monophosphate-activated protein kinase activity via effects on adenosine triphosphate/adenosine diphosphate levels, independently of WAT beigeing. This disturbance promotes a pro-inflammatory profile characterized, in part, by increased chemokine (C-C motif) ligand 2 (CCL2) production. These data suggest that the phosphocreatine/creatine system links cellular energy shuttling with pro-inflammatory responses in human and murine white adipocytes. Our findings provide unexpected perspectives on the mechanisms driving WAT inflammation in obesity and may present avenues to target adipocyte dysfunction.
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Adipocitos Blancos , Creatina , Adipocitos Blancos/metabolismo , Animales , Humanos , Inflamación/metabolismo , Ratones , Obesidad/metabolismo , FosfocreatinaRESUMEN
While single-cell analyses have improved our understanding of liver macrophage heterogeneity, their localization and cellular interactions remain unclear. In a recent issue of Cell, Guilliams et al. provide strategies to localize liver macrophage populations and their communication with neighboring cells during health and disease.
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Macrófagos del Hígado , Hígado , Comunicación CelularRESUMEN
OBJECTIVE: To better comprehend transcriptional phenotypes of cancer cells, we globally characterised RNA-binding proteins (RBPs) to identify altered RNAs, including long non-coding RNAs (lncRNAs). DESIGN: To unravel RBP-lncRNA interactions in cancer, we curated a list of ~2300 highly expressed RBPs in human cells, tested effects of RBPs and lncRNAs on patient survival in multiple cohorts, altered expression levels, integrated various sequencing, molecular and cell-based data. RESULTS: High expression of RBPs negatively affected patient survival in 21 cancer types, especially hepatocellular carcinoma (HCC). After knockdown of the top 10 upregulated RBPs and subsequent transcriptome analysis, we identified 88 differentially expressed lncRNAs, including 34 novel transcripts. CRISPRa-mediated overexpression of four lncRNAs had major effects on the HCC cell phenotype and transcriptome. Further investigation of four RBP-lncRNA pairs revealed involvement in distinct regulatory processes. The most noticeable RBP-lncRNA connection affected lipid metabolism, whereby the non-canonical RBP CCT3 regulated LINC00326 in a chaperonin-independent manner. Perturbation of the CCT3-LINC00326 regulatory network led to decreased lipid accumulation and increased lipid degradation in cellulo as well as diminished tumour growth in vivo. CONCLUSIONS: We revealed that RBP gene expression is perturbed in HCC and identified that RBPs exerted additional functions beyond their tasks under normal physiological conditions, which can be stimulated or intensified via lncRNAs and affected tumour growth.
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We have determined the lipid, protein and miRNA composition of skeletal muscle (SkM)-released extracellular vesicles (ELVs) from Ob/ob (OB) vs wild-type (WT) mice. The results showed that atrophic insulin-resistant OB-SkM released less ELVs than WT-SkM, highlighted by a RAB35 decrease and an increase in intramuscular cholesterol content. Proteomic analyses of OB-ELVs revealed a group of 37 proteins functionally connected, involved in lipid oxidation and with catalytic activities. OB-ELVs had modified contents for phosphatidylcholine (PC 34-4, PC 40-3 and PC 34-0), sphingomyelin (Sm d18:1/18:1) and ceramides (Cer d18:1/18:0) and were enriched in cholesterol, likely to alleviated intracellular accumulation. Surprisingly many ELV miRNAs had a nuclear addressing sequence, and targeted genes encoding proteins with nuclear activities. Interestingly, SkM-ELV miRNA did not target mitochondria. The most significant function targeted by the 7 miRNAs altered in OB-ELVs was lipid metabolism. In agreement, OB-ELVs induced lipid storage in recipient adipocytes and increased lipid up-take and fatty acid oxidation in recipient muscle cells. In addition, OB-ELVs altered insulin-sensitivity and induced atrophy in muscle cells, reproducing the phenotype of the releasing OB muscles. These data suggest for the first time, a cross-talk between muscle cells and adipocytes, through the SkM-ELV route, in favor of adipose tissue expansion.
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Homeostasis , Resistencia a la Insulina , Lípidos/análisis , MicroARNs/genética , Proteínas Musculares/metabolismo , Músculo Esquelético/metabolismo , Atrofia Muscular/patología , Tejido Adiposo , Animales , Exosomas/genética , Exosomas/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Obesos , Proteínas Musculares/genética , Atrofia Muscular/genética , Atrofia Muscular/metabolismo , Proteoma/análisis , Proteoma/metabolismoRESUMEN
Obesity is considered an important factor for many chronic diseases, including diabetes, cardiovascular disease and cancer. The expansion of adipose tissue in obesity is due to an increase in both adipocyte progenitor differentiation and mature adipocyte cell size. Adipocytes, however, are thought to be unable to divide or enter the cell cycle. We demonstrate that mature human adipocytes unexpectedly display a gene and protein signature indicative of an active cell cycle program. Adipocyte cell cycle progression associates with obesity and hyperinsulinemia, with a concomitant increase in cell size, nuclear size and nuclear DNA content. Chronic hyperinsulinemia in vitro or in humans, however, is associated with subsequent cell cycle exit, leading to a premature senescent transcriptomic and secretory profile in adipocytes. Premature senescence is rapidly becoming recognized as an important mediator of stress-induced tissue dysfunction. By demonstrating that adipocytes can activate a cell cycle program, we define a mechanism whereby mature human adipocytes senesce. We further show that by targeting the adipocyte cell cycle program using metformin, it is possible to influence adipocyte senescence and obesity-associated adipose tissue inflammation.
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Adipocitos/metabolismo , Ciclo Celular/fisiología , Senescencia Celular/fisiología , Hiperinsulinismo/patología , Obesidad/patología , Tejido Adiposo/metabolismo , Diferenciación Celular/fisiología , Ciclina D1/metabolismo , Humanos , Hipoglucemiantes/farmacología , Metformina/farmacologíaRESUMEN
Tissue macrophages are immune cells whose phenotypes and functions are dictated by origin and niches. However, tissues are complex environments, and macrophage heterogeneity within the same organ has been overlooked so far. Here, we used high-dimensional approaches to characterize macrophage populations in the murine liver. We identified two distinct populations among embryonically derived Kupffer cells (KCs) sharing a core signature while differentially expressing numerous genes and proteins: a major CD206loESAM- population (KC1) and a minor CD206hiESAM+ population (KC2). KC2 expressed genes involved in metabolic processes, including fatty acid metabolism both in steady-state and in diet-induced obesity and hepatic steatosis. Functional characterization by depletion of KC2 or targeted silencing of the fatty acid transporter Cd36 highlighted a crucial contribution of KC2 in the liver oxidative stress associated with obesity. In summary, our study reveals that KCs are more heterogeneous than anticipated, notably describing a subpopulation wired with metabolic functions.
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Antígenos CD36/metabolismo , Macrófagos del Hígado/metabolismo , Hígado/metabolismo , Obesidad/metabolismo , Estrés Oxidativo/fisiología , Animales , RatonesRESUMEN
BACKGROUND AND AIMS: Oxidative stress plays a key role in the development of metabolic complications associated with obesity, including insulin resistance and the most common chronic liver disease worldwide, nonalcoholic fatty liver disease. We have recently discovered that the microRNA miR-144 regulates protein levels of the master mediator of the antioxidant response, nuclear factor erythroid 2-related factor 2 (NRF2). On miR-144 silencing, the expression of NRF2 target genes was significantly upregulated, suggesting that miR-144 controls NRF2 at the level of both protein expression and activity. Here we explored a mechanism whereby hepatic miR-144 inhibited NRF2 activity upon obesity via the regulation of the tricarboxylic acid (TCA) metabolite, fumarate, a potent activator of NRF2. METHODS: We performed transcriptomic analysis in liver macrophages (LMs) of obese mice and identified the immuno-responsive gene 1 (Irg1) as a target of miR-144. IRG1 catalyzes the production of a TCA derivative, itaconate, an inhibitor of succinate dehydrogenase (SDH). TCA enzyme activities and kinetics were analyzed after miR-144 silencing in obese mice and human liver organoids using single-cell activity assays in situ and molecular dynamic simulations. RESULTS: Increased levels of miR-144 in obesity were associated with reduced expression of Irg1, which was restored on miR-144 silencing in vitro and in vivo. Furthermore, miR-144 overexpression reduces Irg1 expression and the production of itaconate in vitro. In alignment with the reduction in IRG1 levels and itaconate production, we observed an upregulation of SDH activity during obesity. Surprisingly, however, fumarate hydratase (FH) activity was also upregulated in obese livers, leading to the depletion of its substrate fumarate. miR-144 silencing selectively reduced the activities of both SDH and FH resulting in the accumulation of their related substrates succinate and fumarate. Moreover, molecular dynamics analyses revealed the potential role of itaconate as a competitive inhibitor of not only SDH but also FH. Combined, these results demonstrate that silencing of miR-144 inhibits the activity of NRF2 through decreased fumarate production in obesity. CONCLUSIONS: Herein we unravel a novel mechanism whereby miR-144 inhibits NRF2 activity through the consumption of fumarate by activation of FH. Our study demonstrates that hepatic miR-144 triggers a hyperactive FH in the TCA cycle leading to an impaired antioxidant response in obesity.
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Hígado Graso/enzimología , Fumarato Hidratasa/metabolismo , Resistencia a la Insulina , Hígado/enzimología , Macrófagos/enzimología , MicroARNs/metabolismo , Factor 2 Relacionado con NF-E2/metabolismo , Obesidad/enzimología , Animales , Carboxiliasas/genética , Carboxiliasas/metabolismo , Ciclo del Ácido Cítrico , Modelos Animales de Enfermedad , Hígado Graso/genética , Fumarato Hidratasa/genética , Fumaratos/metabolismo , Humanos , Hidroliasas/genética , Hidroliasas/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , MicroARNs/genética , Factor 2 Relacionado con NF-E2/genética , Obesidad/genética , Estrés Oxidativo , Células RAW 264.7 , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal , Succinatos/metabolismoRESUMEN
While the role of transcription factors and coactivators in controlling enhancer activity and chromatin structure linked to gene expression is well established, the involvement of corepressors is not. Using inflammatory macrophage activation as a model, we investigate here a corepressor complex containing GPS2 and SMRT both genome-wide and at the Ccl2 locus, encoding the chemokine CCL2 (MCP-1). We report that corepressors co-occupy candidate enhancers along with the coactivators CBP (H3K27 acetylase) and MED1 (mediator) but act antagonistically by repressing eRNA transcription-coupled H3K27 acetylation. Genome editing, transcriptional interference, and cistrome analysis reveals that apparently related enhancer and silencer elements control Ccl2 transcription in opposite ways. 4C-seq indicates that corepressor depletion or inflammatory signaling functions mechanistically similarly to trigger enhancer activation. In ob/ob mice, adipose tissue macrophage-selective depletion of the Ccl2 enhancer-transcribed eRNA reduces metaflammation. Thus, the identified corepressor-eRNA-chemokine pathway operates in vivo and suggests therapeutic opportunities by targeting eRNAs in immuno-metabolic diseases.
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Quimiocina CCL2/genética , Proteínas Co-Represoras/genética , Elementos de Facilitación Genéticos , Péptidos y Proteínas de Señalización Intracelular/genética , Co-Represor 2 de Receptor Nuclear/genética , Obesidad/genética , Elementos Silenciadores Transcripcionales , Tejido Adiposo/inmunología , Tejido Adiposo/patología , Animales , Sistemas CRISPR-Cas , Quimiocina CCL2/inmunología , Proteínas Co-Represoras/inmunología , Edición Génica , Regulación de la Expresión Génica/efectos de los fármacos , Células HEK293 , Histona Acetiltransferasas/genética , Histona Acetiltransferasas/inmunología , Histonas/genética , Histonas/inmunología , Humanos , Péptidos y Proteínas de Señalización Intracelular/inmunología , Lipopolisacáridos/farmacología , Activación de Macrófagos/efectos de los fármacos , Masculino , Subunidad 1 del Complejo Mediador/genética , Subunidad 1 del Complejo Mediador/inmunología , Ratones , Ratones Obesos , Co-Represor 2 de Receptor Nuclear/inmunología , Obesidad/inmunología , Obesidad/patología , Células RAW 264.7 , ARN no Traducido/genética , ARN no Traducido/inmunología , Transducción de SeñalRESUMEN
The myeloid cells infiltrating the heart early after acute myocardial infarction elaborate a secretome that largely orchestrates subsequent ventricular wall repair. Regulating this innate immune response could be a means to improve infarct healing. To pilot this concept, we utilized (ß1,3-d-) glucan-encapsulated small interfering RNA (siRNA)-containing particles (GeRPs), targeting mononuclear phagocytes, delivered to mice as a one-time intramyocardial injection immediately after acute infarction. Findings demonstrated that cardiac macrophages phagocytosed GeRPs in vivo and had little systemic dissemination, thus providing a means to deliver local therapeutics. Acute infarcts were then injected in vivo with phosphate-buffered saline (PBS; vehicle) or GeRPs loaded with siRNA to Map4k4, and excised hearts were examined at 3 and 7 days by quantitative polymerase chain reaction, flow cytometry, and histology. Compared with infarcted PBS-treated hearts, hearts with intrainfarct injections of siRNA-loaded GeRPs exhibited 69-89% reductions in transcripts for Map4k4 (mitogen-activated protein kinase kinase kinase kinase 4), interleukin (IL)-1ß, and tumor necrosis factor α at 3 days. Expression of other factors relevant to matrix remodeling-monocyte chemoattractant protein-1 (MCP-1), matrix metalloproteinases, hyaluronan synthases, matricellular proteins, and profibrotic factors transforming growth factor beta (TGF-ß), and connective tissue growth factor (CTGF)-were also decreased. Most effects peaked at 3 days, but, in some instances (Map4k4, IL-1ß, TGF-ß, CTGF, versican, and periostin), suppression persisted to 7 days. Thus, direct intramyocardial GeRP injection could serve as a novel and clinically translatable platform for in vivo RNA delivery to intracardiac macrophages for local and selective immunomodulation of the infarct microenvironment.
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Kupffer cells and hepatocytes maintain liver homeostasis. These cells could be separated based on their size and weight, by centrifugation using a density gradient after a liver perfusion. Here, we describe a methodology to isolate both Kupffer cells and hepatocytes from a single mouse, which provides the unique advantage of studying these two cell types from the same liver.
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Centrifugación/métodos , Hepatocitos/citología , Macrófagos del Hígado/citología , Hígado/citología , Animales , Separación Celular/métodos , Células Cultivadas , Masculino , Ratones , Ratones Endogámicos C57BL , Perfusión/métodosRESUMEN
In this issue of Molecular Cell, Toda et al. (2020) show that postprandial elevation of LPS and insulin induce the production of IL-10 by adipose tissue macrophages. Hepatic gluconeogenesis is then inhibited synergistically by insulin and IL-10 to facilitate glucose clearance.
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Insulina , Interleucina-10 , Tejido Adiposo , Lipopolisacáridos , Hígado , Macrófagos , Proteínas Proto-Oncogénicas c-akt , Serina-Treonina Quinasas TORRESUMEN
Obesity and insulin resistance are risk factors for nonalcoholic fatty liver disease (NAFLD), the most common chronic liver disease worldwide. Because no approved medication nor an accurate and noninvasive diagnosis is currently available for NAFLD, there is a clear need to better understand the link between obesity and NAFLD. Lipid accumulation during obesity is known to be associated with oxidative stress and inflammatory activation of liver macrophages (LMs). However, we show that although LMs do not become proinflammatory during obesity, they display signs of oxidative stress. In livers of both humans and mice, antioxidant nuclear factor erythroid 2-related factor 2 (NRF2) was down-regulated with obesity and insulin resistance, yielding an impaired response to lipid accumulation. At the molecular level, a microRNA-targeting NRF2 protein, miR-144, was elevated in the livers of obese insulin-resistant humans and mice, and specific silencing of miR-144 in murine and human LMs was sufficient to restore NRF2 protein expression and the antioxidant response. These results highlight the pathological role of LMs and their therapeutic potential to restore the impaired endogenous antioxidant response in obesity-associated NAFLD.