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The inhabitants of our intestines, collectively called the gut microbiome, comprise fungi, viruses, and bacterial strains. These microorganisms are involved in the fermentation of dietary compounds and the regulation of our adaptive and innate immune systems. Less known is the reciprocal interaction between the gut microbiota and type 2 diabetes mellitus (T2DM), as well as their role in modifying therapies to reduce associated morbidity and mortality. In this review, we aim to discuss the existing literature on gut microbial strains and their diet-derived metabolites involved in T2DM. We also explore the potential diagnostics and therapeutic avenues the gut microbiota presents for targeted T2DM management. Personalized treatment plans, driven by diet and medication based on the patient's microbiome and clinical markers, could optimize therapy.
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Diabetes Mellitus Tipo 2 , Microbioma Gastrointestinal , Diabetes Mellitus Tipo 2/microbiología , Humanos , Dieta , Bacterias/metabolismo , Bacterias/clasificación , Animales , Disbiosis/microbiologíaRESUMEN
OBJECTIVE: Patients with alcohol-associated hepatitis (AH) have a high mortality. Alcohol exacerbates liver damage by inducing gut dysbiosis, bacterial translocation and inflammation, which is characterised by increased numbers of circulating and hepatic neutrophils. DESIGN: In this study, we performed tandem mass tag (TMT) proteomics to analyse proteins in the faeces of controls (n=19), patients with alcohol-use disorder (AUD; n=20) and AH (n=80) from a multicentre cohort (InTeam). To identify protein groups that are disproportionately represented, we conducted over-representation analysis using Reactome pathway analysis and Gene Ontology to determine the proteins with the most significant impact. A faecal biomarker and its prognostic effect were validated by ELISA in faecal samples from patients with AH (n=70), who were recruited in a second and independent multicentre cohort (AlcHepNet). RESULT: Faecal proteomic profiles were overall significantly different between controls, patients with AUD and AH (principal component analysis p=0.001, dissimilarity index calculated by the method of Bray-Curtis). Proteins that showed notable differences across all three groups and displayed a progressive increase in accordance with the severity of alcohol-associated liver disease were predominantly those located in neutrophil granules. Over-representation and Reactome analyses confirmed that differentially regulated proteins are part of granules in neutrophils and the neutrophil degranulation pathway. Myeloperoxidase (MPO), the marker protein of neutrophil granules, correlates with disease severity and predicts 60-day mortality. Using an independent validation cohort, we confirmed that faecal MPO levels can predict short-term survival at 60 days. CONCLUSIONS: We found an increased abundance of faecal proteins linked to neutrophil degranulation in patients with AH, which is predictive of short-term survival and could serve as a prognostic non-invasive marker.
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INTRODUCTION: Ammonia is a pathogenic factor implicated in the progression of metabolic-associated steatotic liver disease (MASLD). The contribution of the glutaminase 1 (GLS) isoform, an enzyme converting glutamine to glutamate and ammonia, to hepatic ammonia build-up and the mechanisms underlying its upregulation in metabolic-associated steatohepatitis (MASH) remain elusive. METHODS: Multiplex transcriptomics and targeted metabolomics analysis of liver biopsies in dietary mouse models representing the whole spectra of MASLD were carried out to characterize the relevance of hepatic GLS during disease pathological progression. In addition, the acute effect of liver-specific GLS inhibition in hepatic ammonia content was evaluated in cultured hepatocytes and in in vivo mouse models of diet-induced MASLD. Finally, the regulatory mechanisms of hepatic GLS overexpression related to the lipopolysaccharide (LPS)/Toll-like receptor 4 (TLR4) axis were explored in the context of MASH. RESULTS: In mouse models of diet-induced MASLD, we found that augmented liver GLS expression is closely associated with the build-up of hepatic ammonia as the disease progresses from steatosis to steatohepatitis. Importantly, the acute silencing/pharmacological inhibition of GLS diminishes the ammonia burden in cultured primary mouse hepatocytes undergoing dedifferentiation, in steatotic hepatocytes, and in a mouse model of diet-induced steatohepatitis, irrespective of changes in ureagenesis and gut permeability. Under these conditions, GLS upregulation in the liver correlates positively with the hepatic expression of TLR4 that recognizes LPS. In agreement, the pharmacological inhibition of TLR4 reduces GLS and hepatic ammonia content in LPS-stimulated mouse hepatocytes and hyperammonemia animal models of endotoxemia. CONCLUSIONS: Overall, our results suggest that the LPS/TLR4 axis regulates hepatic GLS expression promoting liver ammonia build-up as steatotic liver disease progresses to steatohepatitis.
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Amoníaco , Hígado Graso , Glutaminasa , Lipopolisacáridos , Hígado , Receptor Toll-Like 4 , Animales , Glutaminasa/metabolismo , Receptor Toll-Like 4/metabolismo , Amoníaco/metabolismo , Ratones , Hígado/metabolismo , Hígado/patología , Hígado Graso/metabolismo , Hígado Graso/patología , Progresión de la Enfermedad , Masculino , Ratones Endogámicos C57BL , Hepatocitos/metabolismoRESUMEN
BACKGROUND AND AIMS: Mitochondrial antiviral signaling protein (MAVS) is a critical regulator that activates the host's innate immunity against RNA viruses, and its signaling pathway has been linked to the secretion of proinflammatory cytokines. However, the actions of MAVS on inflammatory pathways during the development of metabolic dysfunction-associated steatotic liver disease (MASLD) have been little studied. APPROACH AND RESULTS: Liver proteomic analysis of mice with genetically manipulated hepatic p63, a transcription factor that induces liver steatosis, revealed MAVS as a target downstream of p63. MAVS was thus further evaluated in liver samples from patients and in animal models with MASLD. Genetic inhibition of MAVS was performed in hepatocyte cell lines, primary hepatocytes, spheroids, and mice. MAVS expression is induced in the liver of both animal models and people with MASLD as compared with those without liver disease. Using genetic knockdown of MAVS in adult mice ameliorates diet-induced MASLD. In vitro, silencing MAVS blunts oleic and palmitic acid-induced lipid content, while its overexpression increases the lipid load in hepatocytes. Inhibiting hepatic MAVS reduces circulating levels of the proinflammatory cytokine TNFα and the hepatic expression of both TNFα and NFκß. Moreover, the inhibition of ERK abolished the activation of TNFα induced by MAVS. The posttranslational modification O -GlcNAcylation of MAVS is required to activate inflammation and to promote the high lipid content in hepatocytes. CONCLUSIONS: MAVS is involved in the development of steatosis, and its inhibition in previously damaged hepatocytes can ameliorate MASLD.
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OBJECTIVE: p63 is a transcription factor involved in multiple biological functions. In the liver, the TAp63 isoform induces lipid accumulation in hepatocytes. However, the role of liver TAp63 in the progression of metabolic dysfunction-associated steatohepatitis (MASH) with fibrosis is unknown. METHODS: We evaluated the hepatic p63 levels in different mouse models of steatohepatitis with fibrosis induced by diet. Next, we used virogenetic approaches to manipulate the expression of TAp63 in adult mice under diet-induced steatohepatitis with fibrosis and characterized the disease condition. Finally, we performed proteomics analysis in mice with overexpression and knockdown of hepatic TAp63. RESULTS: Levels of TAp63, but not of ΔN isoform, are increased in the liver of mice with diet-induced steatohepatitis with fibrosis. Both preventive and interventional strategies for the knockdown of hepatic TAp63 significantly ameliorated diet-induced steatohepatitis with fibrosis in mice fed a methionine- and choline-deficient diet (MCDD) and choline deficient and high fat diet (CDHFD). The overexpression of hepatic TAp63 in mice aggravated the liver condition in mice fed a CDHFD. Proteomic analysis in the liver of these mice revealed alteration in multiple proteins and pathways, such as oxidative phosphorylation, antioxidant activity, peroxisome function and LDL clearance. CONCLUSIONS: These results indicate that liver TAp63 plays a critical role in the progression of diet-induced steatohepatitis with fibrosis, and its inhibition ameliorates the disease.
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Hígado Graso , Cirrosis Hepática , Hígado , Ratones Endogámicos C57BL , Animales , Ratones , Hígado/metabolismo , Hígado/patología , Masculino , Cirrosis Hepática/metabolismo , Cirrosis Hepática/patología , Hígado Graso/metabolismo , Hígado Graso/patología , Modelos Animales de Enfermedad , Dieta Alta en Grasa/efectos adversos , Transactivadores/metabolismo , Transactivadores/genética , Proteómica , Metionina/deficiencia , Metionina/metabolismoRESUMEN
The p63 protein has pleiotropic functions and, in the liver, participates in the progression of nonalcoholic fatty liver disease (NAFLD). However, its functions in hepatic stellate cells (HSCs) have not yet been explored. TAp63 is induced in HSCs from animal models and patients with liver fibrosis and its levels positively correlate with NAFLD activity score and fibrosis stage. In mice, genetic depletion of TAp63 in HSCs reduces the diet-induced liver fibrosis. In vitro silencing of p63 blunts TGF-ß1-induced HSCs activation by reducing mitochondrial respiration and glycolysis, as well as decreasing acetyl CoA carboxylase 1 (ACC1). Ectopic expression of TAp63 induces the activation of HSCs and increases the expression and activity of ACC1 by promoting the transcriptional activity of HER2. Genetic inhibition of both HER2 and ACC1 blunt TAp63-induced activation of HSCs. Thus, TAp63 induces HSC activation by stimulating the HER2-ACC1 axis and participates in the development of liver fibrosis.
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Células Estrelladas Hepáticas , Enfermedad del Hígado Graso no Alcohólico , Humanos , Ratones , Animales , Células Estrelladas Hepáticas/metabolismo , Células Estrelladas Hepáticas/patología , Enfermedad del Hígado Graso no Alcohólico/patología , Activación Metabólica , Cirrosis Hepática/genética , Cirrosis Hepática/inducido químicamente , Cirrosis Hepática/metabolismo , Fibrosis , Acetil-CoA Carboxilasa/genética , Acetil-CoA Carboxilasa/metabolismoRESUMEN
There has been an intense focus to uncover the molecular mechanisms by which fasting triggers the adaptive cellular responses in the major organs of the body. Here, we show that in mice, hepatic S-adenosylmethionine (SAMe)-the principal methyl donor-acts as a metabolic sensor of nutrition to fine-tune the catabolic-fasting response by modulating phosphatidylethanolamine N-methyltransferase (PEMT) activity, endoplasmic reticulum-mitochondria contacts, ß-oxidation, and ATP production in the liver, together with FGF21-mediated lipolysis and thermogenesis in adipose tissues. Notably, we show that glucagon induces the expression of the hepatic SAMe-synthesizing enzyme methionine adenosyltransferase α1 (MAT1A), which translocates to mitochondria-associated membranes. This leads to the production of this metabolite at these sites, which acts as a brake to prevent excessive ß-oxidation and mitochondrial ATP synthesis and thereby endoplasmic reticulum stress and liver injury. This work provides important insights into the previously undescribed function of SAMe as a new arm of the metabolic adaptation to fasting.
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Neoplasias Hepáticas , S-Adenosilmetionina , Ratones , Animales , S-Adenosilmetionina/metabolismo , Hígado/metabolismo , Neoplasias Hepáticas/metabolismo , Ayuno , Adenosina Trifosfato/metabolismo , Metionina Adenosiltransferasa/metabolismo , Fosfatidiletanolamina N-Metiltransferasa/metabolismoRESUMEN
Neddylation is a post-translational mechanism that adds a ubiquitin-like protein, namely neural precursor cell expressed developmentally downregulated protein 8 (NEDD8). Here, we show that neddylation in mouse liver is modulated by nutrient availability. Inhibition of neddylation in mouse liver reduces gluconeogenic capacity and the hyperglycemic actions of counter-regulatory hormones. Furthermore, people with type 2 diabetes display elevated hepatic neddylation levels. Mechanistically, fasting or caloric restriction of mice leads to neddylation of phosphoenolpyruvate carboxykinase 1 (PCK1) at three lysine residues-K278, K342, and K387. We find that mutating the three PCK1 lysines that are neddylated reduces their gluconeogenic activity rate. Molecular dynamics simulations show that neddylation of PCK1 could re-position two loops surrounding the catalytic center into an open configuration, rendering the catalytic center more accessible. Our study reveals that neddylation of PCK1 provides a finely tuned mechanism of controlling glucose metabolism by linking whole nutrient availability to metabolic homeostasis.
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Diabetes Mellitus Tipo 2 , Ratones , Animales , Fosfoenolpiruvato/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Proteínas/metabolismo , Hígado/metabolismo , Lisina/metabolismo , Glucosa/metabolismoRESUMEN
OBJECTIVE: O-GlcNAcylation is a post-translational modification that directly couples the processes of nutrient sensing, metabolism, and signal transduction, affecting protein function and localization, since the O-linked N-acetylglucosamine moiety comes directly from the metabolism of glucose, lipids, and amino acids. The addition and removal of O-GlcNAc of target proteins are mediated by two highly conserved enzymes: O-linked N-acetylglucosamine (O-GlcNAc) transferase (OGT) and O-GlcNAcase (OGA), respectively. Deregulation of O-GlcNAcylation has been reported to be associated with various human diseases such as cancer, diabetes, and cardiovascular diseases. The contribution of deregulated O-GlcNAcylation to the progression and pathogenesis of NAFLD remains intriguing, and a better understanding of its roles in this pathophysiological context is required to uncover novel avenues for therapeutic intervention. By using a translational approach, our aim is to describe the role of OGT and O-GlcNAcylation in the pathogenesis of NAFLD. METHODS: We used primary mouse hepatocytes, human hepatic cell lines and in vivo mouse models of steatohepatitis to manipulate O-GlcNAc transferase (OGT). We also studied OGT and O-GlcNAcylation in liver samples from different cohorts of people with NAFLD. RESULTS: O-GlcNAcylation was upregulated in the liver of people and animal models with steatohepatitis. Downregulation of OGT in NAFLD-hepatocytes improved diet-induced liver injury in both in vivo and in vitro models. Proteomics studies revealed that mitochondrial proteins were hyper-O-GlcNAcylated in the liver of mice with steatohepatitis. Inhibition of OGT is able to restore mitochondrial oxidation and decrease hepatic lipid content in in vitro and in vivo models of NAFLD. CONCLUSIONS: These results demonstrate that deregulated hyper-O-GlcNAcylation favors NAFLD progression by reducing mitochondrial oxidation and promoting hepatic lipid accumulation.
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Enfermedad del Hígado Graso no Alcohólico , Humanos , Ratones , Animales , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Regulación hacia Abajo , Acetilglucosamina/metabolismo , Mitocondrias/metabolismo , Hepatocitos/metabolismo , LípidosRESUMEN
OBJECTIVE: p63 is a transcription factor within the p53 protein family that has key roles in development, differentiation and prevention of senescence, but its metabolic actions remain largely unknown. Herein, we investigated the physiological role of p63 in glucose metabolism. DESIGN: We used cell lines and mouse models to genetically manipulate p63 in hepatocytes. We also measured p63 in the liver of patients with obesity with or without type 2 diabetes (T2D). RESULTS: We show that hepatic p63 expression is reduced on fasting. Mice lacking the specific isoform TAp63 in the liver (p63LKO) display higher postprandial and pyruvate-induced glucose excursions. These mice have elevated SIRT1 levels, while SIRT1 knockdown in p63LKO mice normalises glycaemia. Overexpression of TAp63 in wild-type mice reduces postprandial, pyruvate-induced blood glucose and SIRT1 levels. Studies carried out in hepatocyte cell lines show that TAp63 regulates SIRT1 promoter by repressing its transcriptional activation. TAp63 also mediates the inhibitory effect of insulin on hepatic glucose production, as silencing TAp63 impairs insulin sensitivity. Finally, protein levels of TAp63 are reduced in obese persons with T2D and are negatively correlated with fasting glucose and homeostasis model assessment index. CONCLUSIONS: These results demonstrate that p63 physiologically regulates glucose homeostasis.
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Diabetes Mellitus Tipo 2 , Sirtuina 1 , Transactivadores , Animales , Ratones , Glucosa/metabolismo , Hígado/metabolismo , Piruvatos/metabolismo , Sirtuina 1/metabolismo , Transactivadores/metabolismoRESUMEN
O-GlcNAcylation is a posttranslational modification ruled by the activity of a single pair of enzymes, O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA). These two enzymes carry out the dynamic cycling of O-GlcNAcylation on a wide range of cytosolic, nuclear, and mitochondrial proteins in a nutrient- and stress-responsive manner. To maintain proper glucose homeostasis, a precise mechanism to sense blood glucose levels is required, to adapt cell physiology to fluctuations in nutrient intake to maintain glycemia within a narrow range. Disruptions in glucose homeostasis generates metabolic syndrome and type 2 diabetes. In this review we will discuss and summarize emerging findings that points O-GlcNAcylation as a hub in the control of systemic glucose homeostasis, and its involvement in the generation of insulin resistance and type 2 diabetes.
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Diabetes Mellitus Tipo 2 , Resistencia a la Insulina , Glucosa , Homeostasis , Humanos , Procesamiento Proteico-PostraduccionalRESUMEN
BACKGROUND & AIMS: The pathogenesis of liver fibrosis requires activation of hepatic stellate cells (HSCs); once activated, HSCs lose intracellular fatty acids but the role of fatty acid oxidation and carnitine palmitoyltransferase 1A (CPT1A) in this process remains largely unexplored. METHODS: CPT1A was found in HSCs of patients with fibrosis. Pharmacological and genetic manipulation of CPT1A were performed in human HSC cell lines and primary HCSs. Finally, we induced fibrosis in mice lacking CPT1A specifically in HSCs. RESULTS: Herein, we show that CPT1A expression is elevated in HSCs of patients with non-alcoholic steatohepatitis, showing a positive correlation with the fibrosis score. This was corroborated in rodents with fibrosis, as well as in primary human HSCs and LX-2 cells activated by transforming growth factor ß1 (TGFß1) and fetal bovine serum (FBS). Furthermore, both pharmacological and genetic silencing of CPT1A prevent TGFß1- and FBS-induced HSC activation by reducing mitochondrial activity. The overexpression of CPT1A, induced by saturated fatty acids and reactive oxygen species, triggers mitochondrial activity and the expression of fibrogenic markers. Finally, mice lacking CPT1A specifically in HSCs are protected against fibrosis induced by a choline-deficient high-fat diet, a methionine- and choline-deficient diet, or treatment with carbon tetrachloride. CONCLUSIONS: These results indicate that CPT1A plays a critical role in the activation of HSCs and is implicated in the development of liver fibrosis, making it a potentially actionable target for fibrosis treatment. LAY SUMMARY: We show that the enzyme carnitine palmitoyltransferase 1A (CPT1A) is elevated in hepatic stellate cells (HSCs) in patients with fibrosis and mouse models of fibrosis, and that CPT1A induces the activation of these cells. Inhibition of CPT1A ameliorates fibrosis by preventing the activation of HSCs.
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Carnitina O-Palmitoiltransferasa , Células Estrelladas Hepáticas , Animales , Carnitina O-Palmitoiltransferasa/genética , Carnitina O-Palmitoiltransferasa/metabolismo , Colina , Ácidos Grasos/metabolismo , Fibrosis , Células Estrelladas Hepáticas/metabolismo , Humanos , Hígado/patología , Cirrosis Hepática/metabolismo , Cirrosis Hepática/prevención & control , RatonesRESUMEN
BACKGROUND & AIMS: Autophagy-related gene 3 (ATG3) is an enzyme mainly known for its actions in the LC3 lipidation process, which is essential for autophagy. Whether ATG3 plays a role in lipid metabolism or contributes to non-alcoholic fatty liver disease (NAFLD) remains unknown. METHODS: By performing proteomic analysis on livers from mice with genetic manipulation of hepatic p63, a regulator of fatty acid metabolism, we identified ATG3 as a new target downstream of p63. ATG3 was evaluated in liver samples from patients with NAFLD. Further, genetic manipulation of ATG3 was performed in human hepatocyte cell lines, primary hepatocytes and in the livers of mice. RESULTS: ATG3 expression is induced in the liver of animal models and patients with NAFLD (both steatosis and non-alcoholic steatohepatitis) compared with those without liver disease. Moreover, genetic knockdown of ATG3 in mice and human hepatocytes ameliorates p63- and diet-induced steatosis, while its overexpression increases the lipid load in hepatocytes. The inhibition of hepatic ATG3 improves fatty acid metabolism by reducing c-Jun N-terminal protein kinase 1 (JNK1), which increases sirtuin 1 (SIRT1), carnitine palmitoyltransferase 1a (CPT1a), and mitochondrial function. Hepatic knockdown of SIRT1 and CPT1a blunts the effects of ATG3 on mitochondrial activity. Unexpectedly, these effects are independent of an autophagic action. CONCLUSIONS: Collectively, these findings indicate that ATG3 is a novel protein implicated in the development of steatosis. LAY SUMMARY: We show that autophagy-related gene 3 (ATG3) contributes to the progression of non-alcoholic fatty liver disease in humans and mice. Hepatic knockdown of ATG3 ameliorates the development of NAFLD by stimulating mitochondrial function. Thus, ATG3 is an important factor implicated in steatosis.
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Proteínas Relacionadas con la Autofagia/antagonistas & inhibidores , Hígado Graso/prevención & control , Mitocondrias Hepáticas/metabolismo , Enzimas Ubiquitina-Conjugadoras/antagonistas & inhibidores , Animales , Proteínas Relacionadas con la Autofagia/farmacología , Modelos Animales de Enfermedad , Hígado Graso/fisiopatología , Metabolismo de los Lípidos/genética , Ratones , Mitocondrias Hepáticas/fisiología , Proteómica/métodos , Enzimas Ubiquitina-Conjugadoras/farmacologíaRESUMEN
p53 regulates several signaling pathways to maintain the metabolic homeostasis of cells and modulates the cellular response to stress. Deficiency or excess of nutrients causes cellular metabolic stress, and we hypothesized that p53 could be linked to glucose maintenance. We show here that upon starvation hepatic p53 is stabilized by O-GlcNAcylation and plays an essential role in the physiological regulation of glucose homeostasis. More specifically, p53 binds to PCK1 promoter and regulates its transcriptional activation, thereby controlling hepatic glucose production. Mice lacking p53 in the liver show a reduced gluconeogenic response during calorie restriction. Glucagon, adrenaline and glucocorticoids augment protein levels of p53, and administration of these hormones to p53 deficient human hepatocytes and to liver-specific p53 deficient mice fails to increase glucose levels. Moreover, insulin decreases p53 levels, and over-expression of p53 impairs insulin sensitivity. Finally, protein levels of p53, as well as genes responsible of O-GlcNAcylation are elevated in the liver of type 2 diabetic patients and positively correlate with glucose and HOMA-IR. Overall these results indicate that the O-GlcNAcylation of p53 plays an unsuspected key role regulating in vivo glucose homeostasis.
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Acetilglucosamina/metabolismo , Glucosa/metabolismo , Hígado/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Animales , Secuencia de Bases , Restricción Calórica , Línea Celular , Colforsina/farmacología , Diabetes Mellitus Tipo 2/complicaciones , Diabetes Mellitus Tipo 2/metabolismo , Epinefrina/metabolismo , Glucagón/metabolismo , Glucocorticoides/metabolismo , Gluconeogénesis/efectos de los fármacos , Glicosilación , Hepatocitos/efectos de los fármacos , Hepatocitos/metabolismo , Humanos , Hidrocortisona/metabolismo , Hiperglucemia/complicaciones , Hiperglucemia/metabolismo , Resistencia a la Insulina , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Hígado/efectos de los fármacos , Ratones Endogámicos C57BL , Ratones Noqueados , Obesidad/complicaciones , Obesidad/metabolismo , Fosfoenolpiruvato Carboxiquinasa (GTP)/metabolismo , Regiones Promotoras Genéticas/genética , Unión Proteica/efectos de los fármacos , Estabilidad Proteica/efectos de los fármacos , Ácido Pirúvico/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Transcripción Genética/efectos de los fármacos , Proteína p53 Supresora de Tumor/genéticaRESUMEN
BACKGROUND AND AIMS: G protein-coupled receptor (GPR) 55 is a putative cannabinoid receptor, and l-α-lysophosphatidylinositol (LPI) is its only known endogenous ligand. Although GPR55 has been linked to energy homeostasis in different organs, its specific role in lipid metabolism in the liver and its contribution to the pathophysiology of nonalcoholic fatty liver disease (NAFLD) remains unknown. APPROACH AND RESULTS: We measured (1) GPR55 expression in the liver of patients with NAFLD compared with individuals without obesity and without liver disease, as well as animal models with steatosis and nonalcoholic steatohepatitis (NASH), and (2) the effects of LPI and genetic disruption of GPR55 in mice, human hepatocytes, and human hepatic stellate cells. Notably, we found that circulating LPI and liver expression of GPR55 were up-regulated in patients with NASH. LPI induced adenosine monophosphate-activated protein kinase activation of acetyl-coenzyme A carboxylase (ACC) and increased lipid content in human hepatocytes and in the liver of treated mice by inducing de novo lipogenesis and decreasing ß-oxidation. The inhibition of GPR55 and ACCα blocked the effects of LPI, and the in vivo knockdown of GPR55 was sufficient to improve liver damage in mice fed a high-fat diet and in mice fed a methionine-choline-deficient diet. Finally, LPI promoted the initiation of hepatic stellate cell activation by stimulating GPR55 and activation of ACC. CONCLUSIONS: The LPI/GPR55 system plays a role in the development of NAFLD and NASH by activating ACC.
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Lisofosfolípidos/metabolismo , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Obesidad/complicaciones , Receptores de Cannabinoides/metabolismo , Acetil-CoA Carboxilasa/antagonistas & inhibidores , Acetil-CoA Carboxilasa/metabolismo , Adulto , Anciano , Animales , Biopsia , Agonistas de Receptores de Cannabinoides/farmacología , Línea Celular , Estudios de Cohortes , Dieta Alta en Grasa/efectos adversos , Modelos Animales de Enfermedad , Femenino , Técnicas de Silenciamiento del Gen , Células Estrelladas Hepáticas , Hepatocitos , Humanos , Lipogénesis/efectos de los fármacos , Hígado/patología , Lisofosfolípidos/sangre , Masculino , Ratones , Persona de Mediana Edad , Enfermedad del Hígado Graso no Alcohólico/sangre , Enfermedad del Hígado Graso no Alcohólico/etiología , Enfermedad del Hígado Graso no Alcohólico/patología , Obesidad/sangre , Obesidad/metabolismo , Receptores de Cannabinoides/genética , Regulación hacia ArribaRESUMEN
BACKGROUND AND AIMS: The liver plays a central role in all metabolic processes in the body. However, precise characterization of liver metabolism is often obscured by its inherent complexity. Phosphorylated metabolites occupy a prominent position in all anabolic and catabolic pathways. Here, we develop a 31 P nuclear magnetic resonance (NMR)-based method to study the liver "phosphorome" through the simultaneous identification and quantification of multiple hydrophilic and hydrophobic phosphorylated metabolites. APPROACH AND RESULTS: We applied this technique to define the metabolic landscape in livers from a mouse model of the rare disease disorder congenital erythropoietic porphyria (CEP) as well as two well-known murine models of nonalcoholic steatohepatitis: one genetic, methionine adenosyltransferase 1A knockout mice, and the other dietary, mice fed a high-fat choline-deficient diet. We report alterations in the concentrations of phosphorylated metabolites that are readouts of the balance between glycolysis, gluconeogenesis, the pentose phosphate pathway, the tricarboxylic acid cycle, and oxidative phosphorylation and of phospholipid metabolism and apoptosis. Moreover, these changes correlate with the main histological features: steatosis, apoptosis, iron deposits, and fibrosis. Strikingly, treatment with the repurposed drug ciclopirox improves the phosphoromic profile of CEP mice, an effect that was mirrored by the normalization of liver histology. CONCLUSIONS: In conclusion, these findings indicate that NMR-based phosphoromics may be used to unravel metabolic phenotypes of liver injury and to identify the mechanism of drug action.
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Hígado/metabolismo , Metaboloma/fisiología , Enfermedad del Hígado Graso no Alcohólico/patología , Animales , Modelos Animales de Enfermedad , Estudios de Factibilidad , Femenino , Humanos , Interacciones Hidrofóbicas e Hidrofílicas , Hígado/efectos de los fármacos , Hígado/patología , Espectroscopía de Resonancia Magnética , Masculino , Metaboloma/efectos de los fármacos , Metabolómica/métodos , Ratones , Ratones Transgénicos , Modelos Animales , Enfermedad del Hígado Graso no Alcohólico/tratamiento farmacológico , Fósforo , Fosforilación/efectos de los fármacosRESUMEN
Splicing alterations represent an actionable cancer hallmark. Splicing factor 3B subunit 1 (SF3B1) is a crucial splicing factor that can be targeted pharmacologically (e.g. pladienolide-B). Here, we show that SF3B1 is overexpressed (RNA/protein) in hepatocellular carcinoma (HCC) in two retrospective (n = 154 and n = 172 samples) and in five in silico cohorts (n > 900 samples, including TCGA) and that its expression is associated with tumor aggressiveness, oncogenic splicing variants expression (KLF6-SV1, BCL-XL) and decreased overall survival. In vitro, SF3B1 silencing reduced cell viability, proliferation and migration and its pharmacological blockade with pladienolide-B inhibited proliferation, migration, and formation of tumorspheres and colonies in liver cancer cell lines (HepG2, Hep3B, SNU-387), whereas its effects on normal-like hepatocyte-derived THLE-2 proliferation were negligible. Pladienolide-B also reduced the in vivo growth and the expression of tumor-markers in Hep3B-induced xenograft tumors. Moreover, SF3B1 silencing and/or blockade markedly modulated the activation of key signaling pathways (PDK1, GSK3b, ERK, JNK, AMPK) and the expression of cancer-associated genes (CDK4, CD24) and oncogenic SVs (KLF6-SV1). Therefore, the genetic and/or pharmacological inhibition of SF3B1 may represent a promising novel therapeutic strategy worth to be explored through randomized controlled trials.
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Biomarcadores de Tumor/metabolismo , Carcinoma Hepatocelular/patología , Regulación Neoplásica de la Expresión Génica , Neoplasias Hepáticas/patología , Fosfoproteínas/metabolismo , Factores de Empalme de ARN/metabolismo , Adulto , Anciano , Animales , Apoptosis , Biomarcadores de Tumor/genética , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Movimiento Celular , Proliferación Celular , Femenino , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Masculino , Ratones , Ratones Desnudos , Persona de Mediana Edad , Fosfoproteínas/genética , Pronóstico , Factores de Empalme de ARN/genética , Estudios Retrospectivos , Tasa de Supervivencia , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Dopamine signaling is a crucial part of the brain reward system and can affect feeding behavior. Dopamine receptors are also expressed in the hypothalamus, which is known to control energy metabolism in peripheral tissues. Here we show that pharmacological or chemogenetic stimulation of dopamine receptor 2 (D2R) expressing cells in the lateral hypothalamic area (LHA) and the zona incerta (ZI) decreases body weight and stimulates brown fat activity in rodents in a feeding-independent manner. LHA/ZI D2R stimulation requires an intact sympathetic nervous system and orexin system to exert its action and involves inhibition of PI3K in the LHA/ZI. We further demonstrate that, as early as 3 months after onset of treatment, patients treated with the D2R agonist cabergoline experience an increase in energy expenditure that persists for one year, leading to total body weight and fat loss through a prolactin-independent mechanism. Our results may provide a mechanistic explanation for how clinically used D2R agonists act in the CNS to regulate energy balance.