RESUMEN
Interactions between the gut microbiota, diet, and the host potentially contribute to the development of metabolic diseases. Here, we identify imidazole propionate as a microbially produced histidine-derived metabolite that is present at higher concentrations in subjects with versus without type 2 diabetes. We show that imidazole propionate is produced from histidine in a gut simulator at higher concentrations when using fecal microbiota from subjects with versus without type 2 diabetes and that it impairs glucose tolerance when administered to mice. We further show that imidazole propionate impairs insulin signaling at the level of insulin receptor substrate through the activation of p38γ MAPK, which promotes p62 phosphorylation and, subsequently, activation of mechanistic target of rapamycin complex 1 (mTORC1). We also demonstrate increased activation of p62 and mTORC1 in liver from subjects with type 2 diabetes. Our findings indicate that the microbial metabolite imidazole propionate may contribute to the pathogenesis of type 2 diabetes.
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Diabetes Mellitus Tipo 2/metabolismo , Microbioma Gastrointestinal , Imidazoles/metabolismo , Insulina/metabolismo , Diana Mecanicista del Complejo 1 de la Rapamicina/metabolismo , Transducción de Señal , Animales , Células Cultivadas , Diabetes Mellitus Tipo 2/microbiología , Células HEK293 , Histidina/metabolismo , Humanos , Hígado/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Proteína Sequestosoma-1/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
The C. elegans proteins PAQR-2 (a homolog of the human seven-transmembrane domain AdipoR1 and AdipoR2 proteins) and IGLR-2 (a homolog of the mammalian LRIG proteins characterized by a single transmembrane domain and the presence of immunoglobulin domains and leucine-rich repeats in their extracellular portion) form a complex that protects against plasma membrane rigidification by promoting the expression of fatty acid desaturases and the incorporation of polyunsaturated fatty acids into phospholipids, hence increasing membrane fluidity. In the present study, we leveraged a novel gain-of-function allele of PAQR-1, a PAQR-2 paralog, to carry out structure-function studies. We found that the transmembrane domains of PAQR-2 are responsible for its functional requirement for IGLR-2, that PAQR-1 does not require IGLR-2 but acts via the same pathway as PAQR-2, and that the divergent N-terminal cytoplasmic domains of the PAQR-1 and PAQR-2 proteins serve a regulatory function and may regulate access to the catalytic site of these proteins. We also show that overexpression of human AdipoR1 or AdipoR2 alone is sufficient to confer increased palmitic acid resistance in HEK293 cells, and thus act in a manner analogous to the PAQR-1 gain-of-function allele.
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Proteínas de Caenorhabditis elegans/genética , Caenorhabditis elegans/genética , Proteínas de la Membrana/genética , Receptores de Adiponectina/genética , Alelos , Animales , Caenorhabditis elegans/metabolismo , Membrana Celular/genética , Membrana Celular/metabolismo , Mutación con Ganancia de Función/genética , Células HEK293 , Humanos , Fluidez de la Membrana/genética , Fenotipo , Fosfolípidos/genética , Fosfolípidos/metabolismoRESUMEN
Nonalcoholic fatty liver disease (NAFLD) is emerging as a leading cause of chronic liver disease worldwide. Despite intensive nonclinical and clinical research in this field, no specific pharmacological therapy is currently approved to treat NAFLD, which has been recognized as one of the major unmet medical needs of the 21st century. Our recent studies have identified STE20-type kinase MST3, which localizes to intracellular lipid droplets, as a critical regulator of ectopic fat accumulation in human hepatocytes. Here, we explored whether treatment with Mst3-targeting antisense oligonucleotides (ASOs) can promote hepatic lipid clearance and mitigate NAFLD progression in mice in the context of obesity. We found that administration of Mst3-targeting ASOs in mice effectively ameliorated the full spectrum of high-fat diet-induced NAFLD including liver steatosis, inflammation, fibrosis, and hepatocellular damage. Mechanistically, Mst3 ASOs suppressed lipogenic gene expression, as well as acetyl-CoA carboxylase (ACC) protein abundance, and substantially reduced lipotoxicity-mediated oxidative and endoplasmic reticulum stress in the livers of obese mice. Furthermore, we found that MST3 protein levels correlated positively with the severity of NAFLD in human liver biopsies. In summary, this study provides the first in vivo evidence that antagonizing MST3 signaling is sufficient to mitigate NAFLD progression in conditions of excess dietary fuels and warrants future investigations to assess whether MST3 inhibitors may provide a new strategy for the treatment of patients with NAFLD.
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Dieta Alta en Grasa/efectos adversos , Enfermedad del Hígado Graso no Alcohólico/prevención & control , Obesidad/complicaciones , Oligonucleótidos Antisentido/genética , Estrés Oxidativo , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Animales , Lipogénesis , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Obesos , Enfermedad del Hígado Graso no Alcohólico/etiología , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Enfermedad del Hígado Graso no Alcohólico/patología , Proteínas Serina-Treonina Quinasas/genética , Transducción de SeñalRESUMEN
The prevalence of non-alcoholic fatty liver disease (NAFLD) continues to increase dramatically, and there is no approved medication for its treatment. Recently, we predicted the underlying molecular mechanisms involved in the progression of NAFLD using network analysis and identified metabolic cofactors that might be beneficial as supplements to decrease human liver fat. Here, we first assessed the tolerability of the combined metabolic cofactors including l-serine, N-acetyl-l-cysteine (NAC), nicotinamide riboside (NR), and l-carnitine by performing a 7-day rat toxicology study. Second, we performed a human calibration study by supplementing combined metabolic cofactors and a control study to study the kinetics of these metabolites in the plasma of healthy subjects with and without supplementation. We measured clinical parameters and observed no immediate side effects. Next, we generated plasma metabolomics and inflammatory protein markers data to reveal the acute changes associated with the supplementation of the metabolic cofactors. We also integrated metabolomics data using personalized genome-scale metabolic modeling and observed that such supplementation significantly affects the global human lipid, amino acid, and antioxidant metabolism. Finally, we predicted blood concentrations of these compounds during daily long-term supplementation by generating an ordinary differential equation model and liver concentrations of serine by generating a pharmacokinetic model and finally adjusted the doses of individual metabolic cofactors for future human clinical trials.
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Acetilcisteína/administración & dosificación , Carnitina/administración & dosificación , Metabolómica/métodos , Niacinamida/análogos & derivados , Serina/administración & dosificación , Acetilcisteína/sangre , Adulto , Animales , Carnitina/sangre , Suplementos Dietéticos , Quimioterapia Combinada , Voluntarios Sanos , Humanos , Masculino , Modelos Animales , Niacinamida/administración & dosificación , Niacinamida/sangre , Enfermedad del Hígado Graso no Alcohólico/dietoterapia , Medicina de Precisión , Compuestos de Piridinio , Ratas , Serina/sangreRESUMEN
OBJECTIVE: Bariatric surgery improves glucose metabolism. Recent data suggest that faecal microbiota transplantation (FMT) using faeces from postbariatric surgery diet-induced obese mice in germ-free mice improves glucose metabolism and intestinal homeostasis. We here investigated whether allogenic FMT using faeces from post-Roux-en-Y gastric bypass donors (RYGB-D) compared with using faeces from metabolic syndrome donors (METS-D) has short-term effects on glucose metabolism, intestinal transit time and adipose tissue inflammation in treatment-naïve, obese, insulin-resistant male subjects. DESIGN: Subjects with metabolic syndrome (n=22) received allogenic FMT either from RYGB-D or METS-D. Hepatic and peripheral insulin sensitivity as well as lipolysis were measured at baseline and 2 weeks after FMT by hyperinsulinaemic euglycaemic stable isotope (2H2-glucose and 2H5-glycerol) clamp. Secondary outcome parameters were changes in resting energy expenditure, intestinal transit time, faecal short-chain fatty acids (SCFA) and bile acids, and inflammatory markers in subcutaneous adipose tissue related to intestinal microbiota composition. Faecal SCFA, bile acids, glycaemic control and inflammatory parameters were also evaluated at 8 weeks. RESULTS: We observed a significant decrease in insulin sensitivity 2 weeks after allogenic METS-D FMT (median rate of glucose disappearance: from 40.6 to 34.0 µmol/kg/min; p<0.01). Moreover, a trend (p=0.052) towards faster intestinal transit time following RYGB-D FMT was seen. Finally, we observed changes in faecal bile acids (increased lithocholic, deoxycholic and (iso)lithocholic acid after METS-D FMT), inflammatory markers (decreased adipose tissue chemokine ligand 2 (CCL2) gene expression and plasma CCL2 after RYGB-D FMT) and changes in several intestinal microbiota taxa. CONCLUSION: Allogenic FMT using METS-D decreases insulin sensitivity in metabolic syndrome recipients when compared with using post-RYGB-D. Further research is needed to delineate the role of donor characteristics in FMT efficacy in human insulin-resistant subjects. TRIAL REGISTRATION NUMBER: NTR4327.
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Trasplante de Microbiota Fecal , Derivación Gástrica , Glucosa/metabolismo , Resistencia a la Insulina , Síndrome Metabólico/metabolismo , Adulto , Anciano , Ácidos y Sales Biliares/análisis , Quimiocina CCL2/sangre , Quimiocina CCL2/genética , Metabolismo Energético , Ácidos Grasos Volátiles/análisis , Heces/química , Microbioma Gastrointestinal , Tránsito Gastrointestinal , Expresión Génica , Humanos , Lipólisis , Masculino , Síndrome Metabólico/fisiopatología , Síndrome Metabólico/terapia , Metabolómica , Persona de Mediana Edad , Grasa Subcutánea/metabolismo , Donantes de Tejidos , Adulto JovenRESUMEN
Nonalcoholic fatty liver disease (NAFLD) and nonalcoholic steatohepatitis (NASH) are emerging as leading causes of liver disease worldwide and have been recognized as one of the major unmet medical needs of the 21st century. Our recent translational studies in mouse models, human cell lines, and well-characterized patient cohorts have identified serine/threonine kinase (STK)25 as a protein that coats intrahepatocellular lipid droplets (LDs) and critically regulates liver lipid homeostasis and progression of NAFLD/NASH. Here, we studied the mechanism-of-action of STK25 in steatotic liver by relative quantification of the hepatic LD-associated phosphoproteome from high-fat diet-fed Stk25 knockout mice compared with their wild-type littermates. We observed a total of 131 proteins and 60 phosphoproteins that were differentially represented in STK25-deficient livers. Most notably, a number of proteins involved in peroxisomal function, ubiquitination-mediated proteolysis, and antioxidant defense were coordinately regulated in Stk25-/- versus wild-type livers. We confirmed attenuated peroxisomal biogenesis and protection against oxidative and ER stress in STK25-deficient human liver cells, demonstrating the hepatocyte-autonomous manner of STK25's action. In summary, our results suggest that regulation of peroxisomal function and metabolic stress response may be important molecular mechanisms by which STK25 controls the development and progression of NAFLD/NASH.
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Hígado Graso/metabolismo , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Gotas Lipídicas/enzimología , Peroxisomas/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Estrés Fisiológico , Animales , Dieta Alta en Grasa/efectos adversos , Modelos Animales de Enfermedad , Humanos , Péptidos y Proteínas de Señalización Intracelular/deficiencia , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Proteínas Serina-Treonina Quinasas/deficienciaRESUMEN
Ectopic lipid storage in the liver is considered the main risk factor for nonalcoholic steatohepatitis (NASH). Understanding the molecular networks controlling hepatocellular lipid deposition is therefore essential for developing new strategies to effectively prevent and treat this complex disease. Here, we describe a new regulator of lipid partitioning in human hepatocytes: mammalian sterile 20-like (MST) 3. We found that MST3 protein coats lipid droplets in mouse and human liver cells. Knockdown of MST3 attenuated lipid accumulation in human hepatocytes by stimulating ß-oxidation and triacylglycerol secretion while inhibiting fatty acid influx and lipid synthesis. We also observed that lipogenic gene expression and acetyl-coenzyme A carboxylase protein abundance were reduced in MST3-deficient hepatocytes, providing insight into the molecular mechanisms underlying the decreased lipid storage. Furthermore, MST3 expression was positively correlated with key features of NASH (i.e., hepatic lipid content, lobular inflammation, and hepatocellular ballooning) in human liver biopsies. In summary, our results reveal a role of MST3 in controlling the dynamic metabolic balance of liver lipid catabolism vs. lipid anabolism. Our findings highlight MST3 as a potential drug target for the prevention and treatment of NASH and related complex metabolic diseases.-Cansby, E., Kulkarni, N. M., Magnusson, E., Kurhe, Y., Amrutkar, M., Nerstedt, A., Ståhlman, M., Sihlbom, C., Marschall, H.-U., Borén, J., Blüher, M., Mahlapuu, M. Protein kinase MST3 modulates lipid homeostasis in hepatocytes and correlates with nonalcoholic steatohepatitis in humans.
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Hepatocitos/metabolismo , Proteínas Asociadas a Gotas Lipídicas/fisiología , Gotas Lipídicas/metabolismo , Metabolismo de los Lípidos , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Proteínas Serina-Treonina Quinasas/fisiología , Animales , Compartimento Celular , Células Cultivadas , Dieta Alta en Grasa/efectos adversos , Grasas de la Dieta/farmacocinética , Femenino , Técnicas de Silenciamiento del Gen , Homeostasis , Humanos , Hígado/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Mitocondrias Hepáticas/metabolismo , Especificidad de Órganos , Oxidación-Reducción , Estrés Oxidativo , Proteínas Serina-Treonina Quinasas/análisis , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Proteínas Serina-Treonina Quinasas/genética , Interferencia de ARN , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/farmacología , Ratas , Triglicéridos/metabolismoRESUMEN
BACKGROUND: Bile acids (BAs) regulate hepatic lipid metabolism and inflammation. Bile salt export pump (BSEP) KO mice are metabolically preconditioned with a hydrophilic BA composition protecting them from cholestasis. We hypothesize that changes in hepatic BA profile and subsequent changes in BA signalling may critically determine the susceptibility to steatohepatitis. METHODS: Wild-type (WT) and BSEP KO mice were challenged with methionine choline-deficient (MCD) diet to induce steatohepatitis. Serum biochemistry, lipid profiling as well as intestinal lipid absorption were assessed. Markers of inflammation, fibrosis, lipid and BA metabolism were analysed. Hepatic and faecal BA profile as well as serum levels of the BA synthesis intermediate 7-hydroxy-4-cholesten-3-one (C4) were also investigated. RESULTS: Bile salt export pump KO MCD-fed mice developed less steatosis but more inflammation than WT mice. Intestinal neutral lipid levels were reduced in BSEP KO mice at baseline and under MCD conditions. Faecal non-esterified fatty acid concentrations at baseline and under MCD diet were markedly elevated in BSEP KO compared to WT mice. Serum liver enzymes and hepatic expression of inflammatory markers were increased in MCD-fed BSEP KO animals. PPARα protein levels were reduced in BSEP KO mice. Accordingly, PPARα downstream targets Fabp1 and Fatp5 were repressed, while NFκB subunits were increased in MCD-fed BSEP KO mice. Farnesoid X receptor (FXR) protein levels were reduced in MCD-fed BSEP KO vs WT mice. Hepatic BA profile revealed elevated levels of TßMCA, exerting FXR antagonistic action, while concentrations of TCA (FXR agonistic function) were reduced. CONCLUSION: Presence of hydroxylated BAs result in increased faecal FA excretion and reduced hepatic lipid accumulation. This aggravates development of MCD diet-induced hepatitis potentially by decreasing FXR and PPARα signalling.
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Hígado Graso , Metionina , Miembro 11 de la Subfamilia B de Transportador de Casetes de Unión al ATP , Animales , Ácidos y Sales Biliares , Colina , Dieta , Proteínas de Unión a Ácidos Grasos , Inflamación , Hígado , Ratones , Ratones Endogámicos C57BL , Ratones NoqueadosRESUMEN
Perfluorooctanoic acid (PFOA) and perfluorooctanesulfonic acid (PFOS) are man-made chemicals that are used for the fabrication of many products with water- and dirt-repellent properties. The toxicological potential of both substances is currently under debate. In a recent Scientific Opinion, the European Food Safety Authority (EFSA) has identified increased serum total cholesterol levels in humans as one major critical effect being associated with exposure to PFOA or PFOS. In animal studies, both substances induced a decrease of serum cholesterol levels, and the underlying molecular mechanism(s) for these opposed effects are unclear so far. In the present study, we examined the impact of PFOA and PFOS on cholesterol homoeostasis in the human HepaRG cell line as a model for human hepatocytes. Cholesterol levels in HepaRG cells were not affected by PFOA or PFOS, but both substances strongly decreased synthesis of a number of bile acids. The expression of numerous genes whose products are involved in synthesis, metabolism and transport of cholesterol and bile acids was strongly affected by PFOA and PFOS at concentrations above 10 µM. Notably, both substances led to a strong decrease of CYP7A1, the key enzyme catalyzing the rate-limiting step in the synthesis of bile acids from cholesterol, both at the protein level and at the level of gene expression. Moreover, both substances led to a dilatation of bile canaliculi that are formed by differentiated HepaRG cells in vitro. Similar morphological changes are known to be induced by cholestatic agents in vivo. Thus, the strong impact of PFOA and PFOS on bile acid synthesis and bile canalicular morphology in our in vitro experiments may allow the notion that both substances have a cholestatic potential that is connected to the observed increased serum cholesterol levels in humans in epidemiological studies.
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Ácidos Alcanesulfónicos/toxicidad , Ácidos y Sales Biliares/metabolismo , Caprilatos/toxicidad , Fluorocarburos/toxicidad , Animales , Carcinoma Hepatocelular , Colesterol , Expresión Génica , Hepatocitos , Homeostasis , Humanos , Metabolismo de los Lípidos/efectos de los fármacos , Neoplasias HepáticasRESUMEN
Dietary fatty acids can be incorporated directly into phospholipids. This poses a specific challenge to cellular membranes since their composition, hence properties, could greatly vary with different diets. That vast variations in diets are tolerated therefore implies the existence of regulatory mechanisms that monitor and regulate membrane compositions. Here we show that the adiponectin receptor AdipoR2, and its C. elegans homolog PAQR-2, are essential to counter the membrane rigidifying effects of exogenously provided saturated fatty acids. In particular, we use dietary supplements or mutated E. coli as food, together with direct measurements of membrane fluidity and composition, to show that diets containing a high ratio of saturated to monounsaturated fatty acids cause membrane rigidity and lethality in the paqr-2 mutant. We also show that mammalian cells in which AdipoR2 has been knocked-down by siRNA are unable to prevent the membrane-rigidifying effects of palmitic acid. We conclude that the PAQR-2 and AdipoR2 proteins share an evolutionarily conserved function that maintains membrane fluidity in the presence of exogenous saturated fatty acids.
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Proteínas de Caenorhabditis elegans/genética , Membrana Celular/genética , Fluidez de la Membrana/genética , Proteínas de la Membrana/genética , Receptores de Adiponectina/genética , Animales , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Membrana Celular/química , Membrana Celular/metabolismo , Ácidos Grasos/química , Ácidos Grasos/metabolismo , Células HEK293 , Humanos , Proteínas de la Membrana/metabolismo , Fosfolípidos/química , Fosfolípidos/genética , ARN Interferente Pequeño , Receptores de Adiponectina/metabolismoRESUMEN
AIMS: The microsomal triglyceride transport protein (MTTP) is critical for assembly and secretion of apolipoprotein B (apoB)-containing lipoproteins and is most abundant in the liver and intestine. Surprisingly, MTTP is also expressed in the heart. Here we tested the functional relevance of cardiac MTTP expression. MATERIALS AND METHODS: We combined clinical studies, advanced expression analysis of human heart biopsies and analyses in genetically modified mice lacking cardiac expression of the MTTP-A isoform of MTTP. RESULTS: Our results indicate that lower cardiac MTTP expression in humans is associated with structural and perfusion abnormalities in patients with ischemic heart disease. MTTP-A deficiency in mice heart does not affect total MTTP expression, activity or lipid concentration in the heart. Despite this, MTTP-A deficient mice displayed impaired cardiac function after a myocardial infarction. Expression analysis of MTTP indicates that MTTP expression is linked to cardiac function and responses in the heart. CONCLUSIONS: Our results indicate that MTTP may play an important role for the heart function in conjunction to ischemic events.
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Cardiotónicos/metabolismo , Proteínas Portadoras/metabolismo , Corazón/fisiopatología , Isquemia Miocárdica/fisiopatología , Animales , Proteínas Portadoras/genética , Femenino , Regulación de la Expresión Génica , Humanos , Metabolismo de los Lípidos , Hígado/metabolismo , Masculino , Ratones Noqueados , Persona de Mediana Edad , Infarto del Miocardio/diagnóstico por imagen , Infarto del Miocardio/genética , Infarto del Miocardio/fisiopatología , Isquemia Miocárdica/genética , Polimorfismo de Nucleótido Simple/genética , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismoRESUMEN
The FA composition of phospholipids must be tightly regulated to maintain optimal cell membrane properties and compensate for a highly variable supply of dietary FAs. Previous studies have shown that AdipoR2 and its homologue PAQR-2 are important regulators of phospholipid FA composition in HEK293 cells and Caenorhabditiselegans, respectively. Here we show that both AdipoR1 and AdipoR2 are essential for sustaining desaturase expression and high levels of unsaturated FAs in membrane phospholipids of many human cell types, including primary human umbilical vein endothelial cells, and for preventing membrane rigidification in cells challenged with exogenous palmitate, a saturated FA. Three independent methods confirm the role of the AdipoRs as regulators of membrane composition and fluidity: fluorescence recovery after photobleaching, measurements of Laurdan dye generalized polarization, and mass spectrometry to determine the FA composition of phospholipids. Furthermore, we show that the AdipoRs can prevent lipotoxicity in the complete absence of adiponectin, their putative ligand. We propose that the primary cellular function of AdipoR1 and AdipoR2 is to maintain membrane fluidity in most human cell types and that adiponectin is not required for this function.
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Adiponectina/metabolismo , Fluidez de la Membrana , Receptores de Adiponectina/metabolismo , Células Cultivadas , Células HEK293 , Células Hep G2 , Homeostasis , HumanosRESUMEN
BACKGROUND & AIMS: The nuclear farnesoid X receptor (FXR) agonist obeticholic acid (OCA) has been developed for the treatment of liver diseases. We aimed to determine whether OCA treatment increases the risk of gallstone formation. METHODS: Twenty patients awaiting laparoscopic cholecystectomy were randomized to treatment with OCA (25â¯mg/day) or placebo for 3â¯weeks until the day before surgery. Serum bile acids (BAs), the BA synthesis marker C4 (7α-hydroxy-4-cholesten-3-one), and fibroblast growth factor 19 (FGF19) were measured before and after treatment. During surgery, biopsies from the liver and the whole bile-filled gallbladder were collected for analyses of gene expression, biliary lipids and FGF19. RESULTS: In serum, OCA increased FGF19 (from 95.0⯱â¯8.5 to 234.4⯱â¯35.6â¯ng/L) and decreased C4 (from 31.4⯱â¯22.8 to 2.8⯱â¯4.0â¯nmol/L) and endogenous BAs (from 1,312.2⯱â¯236.2 to 517.7⯱â¯178.9â¯nmol/L; all p <0.05). At surgery, BAs in gallbladder bile were lower in patients that received OCA than in controls (OCA, 77.9⯱â¯53.6â¯mmol/L; placebo, 196.4⯱â¯99.3â¯mmol/L; p <0.01), resulting in a higher cholesterol saturation index (OCA, 2.8⯱â¯1.1; placebo, 1.8⯱â¯0.8; p <0.05). In addition, hydrophobic OCA conjugates accounted for 13.6⯱â¯5.0% of gallbladder BAs after OCA treatment, resulting in a higher hydrophobicity index (OCA, 0.43⯱â¯0.09; placebo, 0.34⯱â¯0.07, p <0.05). Gallbladder FGF19 levels were 3-fold higher in OCA patients than in controls (OCA, 40.3⯱â¯16.5â¯ng/L; placebo, 13.5⯱â¯13.1â¯ng/ml; p <0.005). Gene expression analysis indicated that FGF19 mainly originated from the gallbladder epithelium. CONCLUSIONS: Our results show for the first time an enrichment of FGF19 in human bile after OCA treatment. In accordance with its murine homolog FGF15, FGF19 might trigger relaxation and filling of the gallbladder which, in combination with increased cholesterol saturation and BA hydrophobicity, would enhance the risk of gallstone development. LAY SUMMARY: Obeticholic acid increased human gallbladder cholesterol saturation and bile acid hydrophobicity, both decreasing cholesterol solubility in bile. Together with increased hepatobiliary levels of fibroblast growth factor 19, our findings suggest that pharmacological activation of the farnesoid X receptor increases the risk of gallstone formation. Clinical trial number: NCT01625026.
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Ácido Quenodesoxicólico/análogos & derivados , Cálculos Biliares/inducido químicamente , Cálculos Biliares/cirugía , Hepatopatías/tratamiento farmacológico , Receptores Citoplasmáticos y Nucleares/agonistas , Adulto , Ácidos y Sales Biliares/sangre , Ácidos y Sales Biliares/genética , Biopsia , Proteínas Portadoras/genética , Ácido Quenodesoxicólico/efectos adversos , Ácido Quenodesoxicólico/farmacología , Colestenonas/sangre , Método Doble Ciego , Femenino , Factores de Crecimiento de Fibroblastos/sangre , Vesícula Biliar/patología , Vesícula Biliar/cirugía , Cálculos Biliares/sangre , Expresión Génica , Humanos , Hígado/patología , Hepatopatías/sangre , Masculino , Persona de Mediana Edad , Resultado del TratamientoRESUMEN
The pathogenesis of non-alcoholic fatty liver disease (NAFLD) and hepatocellular carcinoma (HCC) has been associated with altered expression of liver-specific genes including pyruvate kinase liver and red blood cell (PKLR), patatin-like phospholipase domain containing 3 (PNPLA3) and proprotein convertase subtilisin/kexin type 9 (PCSK9). Here, we inhibited and overexpressed the expression of these three genes in HepG2 cells, generated RNA-seq data before and after perturbation and revealed the altered global biological functions with the modulation of these genes using integrated network (IN) analysis. We found that modulation of these genes effects the total triglycerides levels within the cells and viability of the cells. Next, we generated IN for HepG2 cells, identified reporter transcription factors based on IN and found that the modulation of these genes affects key metabolic pathways associated with lipid metabolism (steroid biosynthesis, PPAR signalling pathway, fatty acid synthesis and oxidation) and cancer development (DNA replication, cell cycle and p53 signalling) involved in the progression of NAFLD and HCC. Finally, we observed that inhibition of PKLR lead to decreased glucose uptake and decreased mitochondrial activity in HepG2 cells. Hence, our systems level analysis indicated that PKLR can be targeted for development efficient treatment strategy for NAFLD and HCC.
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Metabolismo de los Lípidos/genética , Mitocondrias/metabolismo , Piruvato Quinasa/metabolismo , Células Hep G2 , Humanos , Lipogénesis , Hígado/enzimología , Redes y Vías Metabólicas/genética , Piruvato Quinasa/genética , Triglicéridos/biosíntesisRESUMEN
Objective- Recent cohort studies have shown that nonalcoholic fatty liver disease (NAFLD), and especially nonalcoholic steatohepatitis (NASH), associate with atherosclerosis and cardiovascular disease, independently of conventional cardiometabolic risk factors. However, the mechanisms underlying the pathophysiological link between NAFLD/NASH and cardiovascular disease still remain unclear. Our previous studies have identified STK25 (serine/threonine protein kinase 25) as a critical determinant in ectopic lipid storage, meta-inflammation, and progression of NAFLD/NASH. The aim of this study was to assess whether STK25 is also one of the mediators in the complex molecular network controlling the cardiovascular disease risk. Approach and Results- Atherosclerosis was induced in Stk25 knockout and transgenic mice, and their wild-type littermates, by gene transfer of gain-of-function mutant of PCSK9 (proprotein convertase subtilisin/kexin type 9), which induces the downregulation of hepatic LDLR (low-density lipoprotein receptor), combined with an atherogenic western-type diet. We found that Stk25-/- mice displayed reduced atherosclerosis lesion area as well as decreased lipid accumulation, macrophage infiltration, collagen formation, and oxidative stress in aortic lesions compared with wild-type littermates, independently from alterations in dyslipidemia. Reciprocally, Stk25 transgenic mice presented aggravated plaque formation and maturation compared with wild-type littermates despite similar levels of fasting plasma cholesterol. We also found that STK25 protein was expressed in all layers of the aorta, suggesting a possible direct role in cardiovascular disease. Conclusions- This study provides the first evidence that STK25 plays a critical role in regulation of cardiovascular disease risk and suggests that pharmacological inhibition of STK25 function may provide new possibilities for prevention/treatment of atherosclerosis.
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Aorta/enzimología , Enfermedades de la Aorta/enzimología , Aterosclerosis/enzimología , Hipercolesterolemia/enzimología , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Placa Aterosclerótica , Proteínas Serina-Treonina Quinasas/metabolismo , Animales , Aorta/patología , Enfermedades de la Aorta/genética , Enfermedades de la Aorta/patología , Enfermedades de la Aorta/prevención & control , Aterosclerosis/genética , Aterosclerosis/patología , Aterosclerosis/prevención & control , Dieta Alta en Grasa , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Técnicas de Transferencia de Gen , Hipercolesterolemia/genética , Péptidos y Proteínas de Señalización Intracelular/deficiencia , Péptidos y Proteínas de Señalización Intracelular/genética , Hígado/enzimología , Hígado/patología , Masculino , Ratones Endogámicos C57BL , Ratones Noqueados , Mutación , Enfermedad del Hígado Graso no Alcohólico/enzimología , Enfermedad del Hígado Graso no Alcohólico/genética , Enfermedad del Hígado Graso no Alcohólico/patología , Proproteína Convertasa 9/genética , Proproteína Convertasa 9/metabolismo , Proteínas Serina-Treonina Quinasas/deficiencia , Proteínas Serina-Treonina Quinasas/genética , Receptores de LDL/deficiencia , Receptores de LDL/genética , Transducción de SeñalRESUMEN
In spite of the worldwide impact of diabetes on human health, the mechanisms behind glucose toxicity remain elusive. Here we show that C. elegans mutants lacking paqr-2, the worm homolog of the adiponectin receptors AdipoR1/2, or its newly identified functional partner iglr-2, are glucose intolerant and die in the presence of as little as 20 mM glucose. Using FRAP (Fluorescence Recovery After Photobleaching) on living worms, we found that cultivation in the presence of glucose causes a decrease in membrane fluidity in paqr-2 and iglr-2 mutants and that genetic suppressors of this sensitivity act to restore membrane fluidity by promoting fatty acid desaturation. The essential roles of paqr-2 and iglr-2 in the presence of glucose are completely independent from daf-2 and daf-16, the C. elegans homologs of the insulin receptor and its downstream target FoxO, respectively. Using bimolecular fluorescence complementation, we also show that PAQR-2 and IGLR-2 interact on plasma membranes and thus may act together as a fluidity sensor that controls membrane lipid composition.
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Proteínas de Caenorhabditis elegans/genética , Caenorhabditis elegans/genética , Glucosa/toxicidad , Fluidez de la Membrana/genética , Lípidos de la Membrana/metabolismo , Proteínas de la Membrana/genética , Animales , Proteínas de Caenorhabditis elegans/metabolismo , Recuperación de Fluorescencia tras Fotoblanqueo , Factores de Transcripción Forkhead/genética , Fluidez de la Membrana/fisiología , Proteínas de la Membrana/metabolismo , Receptor de Insulina/genéticaRESUMEN
[This corrects the article DOI: 10.1371/journal.pgen.1005982.].
RESUMEN
BACKGROUND AND AIMS: Interruption of the enterohepatic circulation of bile acids (BAs) may protect against BA-mediated cholestatic liver and bile duct injury. BA sequestrants are established to treat cholestatic pruritus, but their impact on the underlying cholestasis is still unclear. We aimed to explore the therapeutic effects and mechanisms of the BA sequestrant colesevelam in a mouse model of sclerosing cholangitis. METHODS: Mdr2-/- mice received colesevelam for 8 weeks. Gene expression profiles of BA homeostasis, inflammation and fibrosis were explored in liver, intestine and colon. Hepatic and faecal BA profiles and gut microbiome were analysed. Glucagon-like peptide 1 (GLP-1) levels in portal blood were measured by ELISA. Furthermore, Mdr2-/- mice as well as wild-type 3,5-diethoxy-carbonyl-1,4-dihydrocollidine-fed mice were treated with GLP-1-receptor agonist exendin-4 for 2 weeks prior to analysis. RESULTS: Colesevelam reduced serum liver enzymes, BAs and expression of proinflammatory and profibrogenic markers. Faecal BA profiling revealed increased levels of secondary BAs after resin treatment, while hepatic and biliary BA composition showed a shift towards more hydrophilic BAs. Colonic GLP-1 secretion, portal venous GLP-1 levels and intestinal messenger RNA expression of gut hormone Proglucagon were increased, while ileal Fgf15 expression was abolished by colesevelam. Exendin-4 treatment increased bile duct mass without promoting a reactive cholangiocyte phenotype in mouse models of sclerosing cholangitis. Microbiota analysis showed an increase of the phylum δ-Proteobacteria after colesevelam treatment and a shift within the phyla Firmicutes from Clostridiales to Lactobacillus. CONCLUSION: Colesevelam increases faecal BA excretion and enhances BA conversion towards secondary BAs, thereby stimulating secretion of GLP-1 from enteroendocrine L-cells and attenuates liver and bile duct injury in Mdr2-/- mice.
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Anticolesterolemiantes/uso terapéutico , Conductos Biliares/efectos de los fármacos , Colangitis Esclerosante/tratamiento farmacológico , Clorhidrato de Colesevelam/uso terapéutico , Hígado/efectos de los fármacos , Animales , Colestasis/tratamiento farmacológico , Modelos Animales de Enfermedad , Péptido 1 Similar al Glucagón/efectos de los fármacos , Homeostasis/efectos de los fármacos , Ratones , Ratones Noqueados , Resultado del TratamientoRESUMEN
Human rhinovirus (RV) infections are a significant risk factor for exacerbations of asthma and chronic obstructive pulmonary disease. Thus, approaches to prevent RV infection in such patients would give significant benefit. Through RNA interference library screening, we identified lanosterol synthase (LSS), a component of the cholesterol biosynthetic pathway, as a novel regulator of RV replication in primary normal human bronchial epithelial cells. Selective knock down of LSS mRNA with short interfering RNA inhibited RV2 replication in normal human bronchial epithelial cells. Small molecule inhibitors of LSS mimicked the effect of LSS mRNA knockdown in a concentration-dependent manner. We further demonstrated that the antiviral effect is not dependent on a reduction in total cellular cholesterol but requires a 24-hour preincubation with the LSS inhibitor. The rank order of antiviral potency of the LSS inhibitors used was consistent with LSS inhibition potency; however, all compounds showed remarkably higher potency against RV compared with the LSS enzyme potency. We showed that LSS inhibition led to an induction of 24(S),25 epoxycholesterol, an important regulator of the sterol pathway. We also demonstrated that LSS inhibition led to a profound increase in expression of the innate antiviral defense protein, IFN-ß. We found LSS to be a novel regulator of RV replication and innate antiviral immunity and identified a potential molecular mechanism for this effect, via induction of 24(S),25 epoxycholesterol. Inhibition of LSS could therefore be a novel therapeutic target for prevention of RV-induced exacerbations.
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Antivirales/farmacología , Bronquios/inmunología , Células Epiteliales/inmunología , Inmunidad Innata/inmunología , Transferasas Intramoleculares/metabolismo , Infecciones por Picornaviridae/inmunología , Rhinovirus/inmunología , Replicación Viral/inmunología , Bronquios/efectos de los fármacos , Bronquios/virología , Células Cultivadas , Células Epiteliales/efectos de los fármacos , Células Epiteliales/virología , Humanos , Inmunidad Innata/efectos de los fármacos , Transferasas Intramoleculares/antagonistas & inhibidores , Transferasas Intramoleculares/genética , Infecciones por Picornaviridae/tratamiento farmacológico , Infecciones por Picornaviridae/virología , ARN Interferente Pequeño/genética , Rhinovirus/efectos de los fármacos , Bibliotecas de Moléculas Pequeñas/farmacología , Replicación Viral/efectos de los fármacosRESUMEN
Increasing evidence suggests that polyphenols have a significant potential in the prevention and treatment of risk factors associated with metabolic syndrome. The objective of this study was to assess the metabolic outcomes of two polyphenol-containing extracts from cinnamon bark (CBE) and grape pomace (GPE) on C57BL/6J mice fed a high-fat diet (HFD) for 8 wk. Both CBE and GPE were able to decrease fat mass gain and adipose tissue inflammation in mice fed a HFD without reducing food intake. This was associated with reduced liver steatosis and lower plasma nonesterified fatty acid levels. We also observed a beneficial effect on glucose homeostasis, as evidenced by an improved glucose tolerance and a lower insulin resistance index. These ameliorations of the overall metabolic profile were associated with a significant impact on the microbial composition, which was more profound for the GPE than for the CBE. At the genus level, Peptococcus were decreased in the CBE group. In the GPE-treated group, several key genera that have been previously found to be linked with HFD, metabolic effects, and gut barrier integrity were affected: we observed a decrease of Desulfovibrio, Lactococcus, whereas Allobaculum and Roseburia were increased. In addition, the expression of several antimicrobial peptides and tight junction proteins was increased in response to both CBE and GPE supplementation, indicating an improvement of the gut barrier function. Collectively, these data suggest that CBE and GPE can ameliorate the overall metabolic profile of mice on a high-fat diet, partly by acting on the gut microbiota.