RESUMEN
Cyp2c70 is the liver enzyme in rodents responsible for synthesis of the primary 6-hydroxylated muricholate bile acid (BA) species. Cyp2c70 KO mice are devoid of protective, hydrophilic muricholic acids, leading to a more human-like BA composition and subsequent cholestatic liver injury. Pharmacological inhibition of the ileal BA transporter (IBAT) has been shown to be therapeutic in cholestatic models. Here, we aimed to determine if IBAT inhibition with SC-435 is protective in Cyp2c70 KO mice. As compared to WT mice, we found male and female Cyp2c70 KO mice exhibited increased levels of serum liver injury markers, and our evaluation of liver histology revealed increased hepatic inflammation, macrophage infiltration, and biliary cell proliferation. We demonstrate serum and histologic markers of liver damage were markedly reduced with SC-435 treatment. Additionally, we show hepatic gene expression in pathways related to immune cell activation and inflammation were significantly upregulated in Cyp2c70 KO mice and reduced to levels indistinguishable from WT with IBAT inhibition. In Cyp2c70 KO mice, the liver BA content was significantly increased, enriched in chenodeoxycholic acid, and more hydrophobic, exhibiting a hydrophobicity index value and red blood cell lysis properties similar to human liver BAs. Furthermore, we determined IBAT inhibition reduced the total hepatic BA levels but did not affect overall hydrophobicity of the liver BAs. These findings suggest that there may be a threshold in the liver for pathological accretion of hydrophobic BAs and reducing hepatic BA accumulation can be sufficient to alleviate liver injury, independent of BA pool hydrophobicity.
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Colestasis , Hígado , Animales , Ácidos y Sales Biliares/metabolismo , Proteínas Portadoras , Ácido Quenodesoxicólico/metabolismo , Colestasis/metabolismo , Óxidos N-Cíclicos , Femenino , Humanos , Inflamación/metabolismo , Hígado/metabolismo , Masculino , Glicoproteínas de Membrana , Ratones , TropanosRESUMEN
Multidrug resistance protein 4 (Mrp4) is an efflux transporter known to transport several xenobiotics and endogenous molecules. We recently identified that the lack of Mrp4 increases adipose tissue and body weights in mice. However, the role of Mrp4 in adipose tissue physiology are unknown. The current study aimed at characterizing these specific roles of Mrp4 using wild-type (WT) and knockout (Mrp4-/- ) mice. Our studies determined that Mrp4 is expressed in mouse adipose tissue and that the lack of Mrp4 expression is associated with adipocyte hypertrophy. Furthermore, the lack of Mrp4 increased blood glucose and leptin levels, and impaired glucose tolerance. Additionally, in 3T3-L1 cells and human pre-adipocytes, pharmacological inhibition of Mrp4 increased adipogenesis and altered expression of adipogenic genes. Lack of Mrp4 activity in both of our in vivo and in vitro models leads to increased activation of adipose tissue cAMP response element-binding protein (Creb) and decreased plasma prostaglandin E (PGE) metabolite levels. These changes in Creb activation, coupled with decreased PGE levels, together promoted the observed metabolic phenotype in Mrp4-/- mice. In conclusion, our results indicate that Mrp4 as a novel genetic factor involved in the pathogenesis of metabolic diseases, such as obesity and diabetes.
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Diabetes Mellitus/metabolismo , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/metabolismo , Obesidad/metabolismo , Adipocitos/metabolismo , Adipogénesis/genética , Adipogénesis/fisiología , Animales , Western Blotting , Calorimetría , Diferenciación Celular/genética , Diferenciación Celular/fisiología , Diabetes Mellitus/genética , Humanos , Ratones , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/genética , Obesidad/genética , RNA-SeqRESUMEN
Multidrug resistance-associated protein 4 (Mrp4) is an efflux transporter involved in the active transport of several endogenous and exogenous chemicals. Previously, we have shown that hepatic Mrp4 expression increases following acetaminophen overdose. In mice, these increases in Mrp4 expression are observed specifically in hepatocytes undergoing active proliferation. From this, we hypothesized that Mrp4 plays a key role in hepatocyte proliferation and that lack of Mrp4 impedes liver regeneration following liver injury and/or tissue loss. To evaluate the role of Mrp4 in these processes, we employed two-third partial hepatectomy (PH) as an experimental liver regeneration model. In this study, we performed PH-surgery on male wildtype (C57BL/6J) and Mrp4 knockout mice. Plasma and liver tissues were collected at 24, 48, and 72 h postsurgery and evaluated for liver injury and liver regeneration endpoints, and for PH-induced hepatic lipid accumulation. Our results show that lack of Mrp4 did not alter hepatocyte proliferation and liver injury following PH as evaluated by Ki-67 antigen staining and plasma alanine aminotransferase levels. To our surprise, Mrp4 knockout mice exhibited increased hepatic lipid content, in particular, di- and triglyceride levels. Gene expression analysis showed that lack of Mrp4 upregulated hepatic lipin1 and diacylglycerol O-acyltransferase 1 and 2 gene expression, which are involved in the synthesis of di- and triglycerides. Our observations indicate that lack of Mrp4 prolonged PH-induced hepatic steatosis in mice and suggest that Mrp4 may be a novel genetic factor in the development of hepatic steatosis.
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Proliferación Celular/efectos de los fármacos , Hígado Graso/fisiopatología , Hepatectomía/efectos adversos , Hepatocitos/efectos de los fármacos , Hepatocitos/metabolismo , Regeneración Hepática/efectos de los fármacos , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/metabolismo , Animales , Modelos Animales de Enfermedad , Hígado Graso/etiología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones NoqueadosRESUMEN
Acetaminophen (APAP) overdose is the most frequent cause of drug-induced liver injury in humans and a common chemical model to investigate genetic determinants of susceptibility to drug-induced liver injury (DILI). Previous studies performed in our laboratory identified the efflux transporter multidrug resistance-associated protein 4 (Mrp4) as an inducible gene in the liver following toxic APAP exposure in both humans and rodents. In mice, blockade of hepatic Mrp4 induction following APAP administration increases susceptibility towards APAP hepatotoxicity. Collectively, these findings suggest that Mrp4 plays an important role in tolerance to APAP-induced liver injury. To further study the role of Mrp4 in APAP-induced hepatotoxicity, we treated 10-12 weeks old male wild type (WT, C57BL/6J) and Mrp4 knockout (Mrp4-/-) mice with APAP (400 mg/Kg in saline, i.p.) or vehicle. Liver injury endpoints and hepatic gene expression were analyzed at 12, 24 and 48 h post-APAP injections. Unexpectedly, the kinetics of histologically measured liver damage and plasma ALT revealed that Mrp4-/ mice had decreased ALT levels and hepatic necrosis compared to WT mice only at 12 h. Notably, hepatic non-protein sulfhydryl (NPSH) levels were increased in the APAP treated Mrp4-/- mice at intervals less than 24 h, consistent with the capability of Mrp4 to export glutathione. Further gene expression analysis revealed that hepatic drug metabolism genes were downregulated in Mrp4-/- mice at earlier time points post-APAP administration. However, despite significant decreases in endpoints of liver injury detected at an early time point after APAP treatment, these changes were not sustained at later time points as Mrp4-/- mice ultimately had hepatic toxicity at levels comparable to WT mice. In conclusion, our data indicate that lack of Mrp4 by itself in mice does not alter susceptibility to APAP toxicity.
RESUMEN
Alcoholic fatty liver disease (AFLD) is a major risk factor for cirrhosis-associated liver diseases. Studies demonstrate that alcohol increases serum bile acids in humans and rodents. AFLD has been linked to cholestasis, although the physiologic relevance of increased bile acids in AFLD and the underlying mechanism of increasing the bile acid pool by alcohol feeding are still unclear. In this study, we used mouse models either deficient of or overexpressing cholesterol 7α-hydroxylase (Cyp7a1), the rate-limiting and key regulatory enzyme in bile acid synthesis, to study the effect of alcohol drinking in liver metabolism and inflammation. Mice were challenged with chronic ethanol feeding (10 days) plus a binge dose of alcohol by oral gavage (5 g/kg body weight). Alcohol feeding reduced bile acid synthesis gene expression but increased the bile acid pool size, hepatic triglycerides and cholesterol, and inflammation and injury in wild-type mice and aggravated liver inflammation and injury in Cyp7a1-deficient mice. Interestingly, alcohol-induced hepatic inflammation and injury were ameliorated in Cyp7a1 transgenic mice. Conclusion: Alcohol feeding alters hepatic bile acid and cholesterol metabolism to cause liver inflammation and injury, while maintenance of bile acid and cholesterol homeostasis protect against alcohol-induced hepatic inflammation and injury. Our findings indicate that CYP7A1 plays a key role in protection against alcohol-induced steatohepatitis. (Hepatology Communications 2018;2:99-112).
RESUMEN
BACKGROUND: Exposure to chemicals during critical windows of development may re-program liver for increased risk of nonalcoholic fatty liver disease (NAFLD). Bisphenol A (BPA), a plastics component, has been described to impart adverse effects during gestational and lactational exposure. Our work has pointed to nuclear factor E2-related factor 2 (Nrf2) being a modulator of hepatic lipid accumulation in models of NAFLD. OBJECTIVES: To determine if chemical exposure can prime liver for steatosis via modulation of NRF2 and epigenetic mechanisms. METHODS: Utilizing BPA as a model exposure, pregnant CD-1 mice were administered 25µg/kg/day BPA via osmotic minipumps from gestational day 8 through postnatal day (PND)16. The offspring were weaned on PND21 and exposed to same dose of BPA via their drinking water through PND35. Tissues were collected from pups at week 5 (W5), and their littermates at week 39 (W39). RESULTS: BPA increased hepatic lipid content concomitant with increased Nrf2 and pro-lipogenic enzyme expression at W5 and W39 in female offspring. BPA exposure increased Nrf2 binding to a putative antioxidant response element consensus sequence in the sterol regulatory-element binding protein-1c (Srebp-1c) promoter. Known Nrf2 activators increased SREBP-1C promoter reporter activity in HepG2 cells. Methylated DNA immunoprecipitation-PCR and pyrosequencing revealed that developmental BPA exposure induced hypomethylation of the Nrf2 and Srebp-1c promoters in livers of W5 mice, which was more prominent in W39 mice than in others. CONCLUSION: Exposure to a xenobiotic during early development induced persistent fat accumulation via hypomethylation of lipogenic genes. Moreover, increased Nrf2 recruitment to the Srebp-1c promoter in livers of BPA-exposed mice was observed. Overall, the underlying mechanisms described a broader impact beyond BPA exposure and can be applied to understand other models of NAFLD. https://doi.org/10.1289/EHP664.
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Compuestos de Bencidrilo/toxicidad , Contaminantes Ambientales/toxicidad , Epigénesis Genética/efectos de los fármacos , Factor 2 Relacionado con NF-E2/genética , Enfermedad del Hígado Graso no Alcohólico/inducido químicamente , Fenoles/toxicidad , Efectos Tardíos de la Exposición Prenatal/metabolismo , Proteína 1 de Unión a los Elementos Reguladores de Esteroles/genética , Animales , Metilación de ADN/efectos de los fármacos , Modelos Animales de Enfermedad , Femenino , Humanos , Metabolismo de los Lípidos/efectos de los fármacos , Hígado/efectos de los fármacos , Masculino , Ratones , Factor 2 Relacionado con NF-E2/metabolismo , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Embarazo , Efectos Tardíos de la Exposición Prenatal/inducido químicamente , Pubertad/efectos de los fármacos , Proteína 1 de Unión a los Elementos Reguladores de Esteroles/metabolismoRESUMEN
Bile acids are signaling molecules that play a critical role in regulation of hepatic metabolic homeostasis by activating nuclear farnesoid X receptor (Fxr) and membrane G-protein-coupled receptor (Takeda G-protein-coupled receptor 5; Tgr5). The role of FXR in regulation of bile acid synthesis and hepatic metabolism has been studied extensively. However, the role of TGR5 in hepatic metabolism has not been explored. The liver plays a central role in lipid metabolism, and impaired response to fasting and feeding contributes to steatosis and nonalcoholic fatty liver and obesity. We have performed a detailed analysis of gallbladder bile acid and lipid metabolism in Tgr5-/- mice in both free-fed and fasted conditions. Lipid profiles of serum, liver and adipose tissues, bile acid composition, energy metabolism, and messenger RNA and protein expression of the genes involved in lipid metabolism were analyzed. Results showed that deficiency of the Tgr5 gene in mice alleviated fasting-induced hepatic lipid accumulation. Expression of liver oxysterol 7α-hydroxylase in the alternative bile acid synthesis pathway was reduced. Analysis of gallbladder bile acid composition showed marked increase of taurocholic acid and decrease of tauro-α and ß-muricholic acid in Tgr5-/- mice. Tgr5-/- mice had increased hepatic fatty acid oxidation rate and decreased hepatic fatty acid uptake. Interestingly, fasting induction of fibroblast growth factor 21 in liver was attenuated. In addition, fasted Tgr5-/- mice had increased activation of hepatic growth hormone-signal transducer and activator of transcription 5 (GH-Stat5) signaling compared to wild-type mice. CONCLUSION: TGR5 may play a role in determining bile acid composition and in fasting-induced hepatic steatosis through a novel mechanism involving activation of the GH-Stat5 signaling pathway. (Hepatology 2017;65:813-827).
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Ácidos y Sales Biliares/metabolismo , Hígado Graso/metabolismo , Regulación de la Expresión Génica , Proteínas de Unión al ARN/metabolismo , Receptores Acoplados a Proteínas G/genética , Análisis de Varianza , Animales , Modelos Animales de Enfermedad , Metabolismo Energético/fisiología , Ayuno , Hígado Graso/patología , Homeostasis/genética , Metabolismo de los Lípidos/genética , Masculino , Ratones , Ratones Endogámicos C57BL , Consumo de Oxígeno/fisiología , Distribución Aleatoria , Transducción de SeñalRESUMEN
Metabolic syndrome is a multifactorial disease associated with obesity, insulin resistance, diabetes, and the alteration of multiple metabolic hormones. Obesity rates have been rising worldwide, which increases our need to understand how this population will respond to drugs and exposure to other chemicals. The purpose of this study was to determine in lean and obese mice the ontogeny of clinical biomarkers such as serum hormone and blood glucose levels as well as the physiologic markers that correlate with nuclear receptor- and transporter-related pathways. Livers from male and female wild-type (WT) (C57BL/6) and ob/ob mice littermates were collected before, during, and after the onset of obesity. Serum hormone and mRNA levels were analyzed. Physiologic changes and gene expression during maturation and progression to obesity were performed and correlation analysis was performed using canonical correlations. Significant ontogenic changes in both WT and ob/ob mice were observed and these ontogenic changes differ in ob/ob mice with the development of obesity. In males and females, the ontogenic pattern of the expression of genes such as Abcc3, 4, Abcg2, Cyp2b10, and 4a14 started to differ from week 3, and became significant at weeks 4 and 8 in ob/ob mice compared with WT mice. In obese males, serum resistin, glucagon, and glucose levels correlated with the expression of most hepatic ATP-binding cassette (Abc) transporters, whereas in obese females, serum glucagon-like peptide 1 levels were correlated with most hepatic uptake transporters and P450 enzymes. Overall, the correlation between physiologic changes and gene expression indicate that metabolism-related hormones may play a role in regulating the genes involved in drug metabolism and transport.
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Transportadoras de Casetes de Unión a ATP/biosíntesis , Hormonas/sangre , Hígado/metabolismo , Síndrome Metabólico/metabolismo , Fenotipo , Factores de Transcripción/biosíntesis , Transportadoras de Casetes de Unión a ATP/genética , Animales , Femenino , Regulación de la Expresión Génica , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones ObesosRESUMEN
OBJECTIVE: To evaluate whether Nrf2 deficiency impacts insulin resistance and lipid accumulation in liver and white adipose tissue. METHODS: Lep(ob/ob) mice (OB) with targeted Nrf2 deletion (OB-Nrf2KO) were generated. Pathogenesis of obesity and type 2 diabetes was measured in C57BL/6J, Nrf2KO, OB, and OB-Nrf2KO mice. Hepatic lipid content, lipid clearance, and very low-density lipoprotein (VLDL) secretion were determined between OB and OB-Nrf2KO mice. RESULTS: OB-Nrf2KO mice exhibited decreased white adipose tissue mass and decreased adipogenic and lipogenic gene expression compared with OB mice. Nrf2 deficiency prolonged hyperglycemia in response to glucose challenge, which was paralleled by reduced insulin-stimulated Akt phosphorylation. In OB mice, Nrf2 deficiency decreased hepatic lipid accumulation, decreased peroxisome proliferator-activated receptor γ expression and nicotinamide adenine dinucleotide phosphate (NADPH) content, and enhanced VLDL secretion. However, this observation was opposite in lean mice. Additionally, OB-Nrf2KO mice exhibited increased plasma triglyceride content, decreased HDL-cholesterol content, and enhanced apolipoprotein B expression, suggesting Nrf2 deficiency caused dyslipidemia in these mice. CONCLUSIONS: Nrf2 deficiency in Lep(ob/ob) mice reduced white adipose tissue mass and prevented hepatic lipid accumulation but induced insulin resistance and dyslipidemia. This study indicates a dual role of Nrf2 during metabolic dysregulation-increasing lipid accumulation in liver and white adipose tissue but preventing lipid accumulation in obese mice.
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Tejido Adiposo/metabolismo , Resistencia a la Insulina , Leptina/deficiencia , Hígado/metabolismo , Factor 2 Relacionado con NF-E2/metabolismo , Obesidad/metabolismo , Tejido Adiposo/citología , Animales , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Obesos , Factores de Transcripción/metabolismoRESUMEN
Flavin-containing monooxygenase-3 (FMO3) catalyzes metabolic reactions similar to cytochrome P450 monooxygenase, however, most metabolites of FMO3 are considered non-toxic. Recent findings in our laboratory demonstrated Fmo3 gene induction following toxic acetaminophen (APAP) treatment in mice. The goal of this study was to evaluate Fmo3 gene expression in other diverse mouse models of hepatic oxidative stress and injury. Fmo3 gene regulation by Nrf2 was also investigated using Nrf2 knockout (Nrf2 KO) mice. In our studies, male C57BL/6J mice were treated with toxic doses of hepatotoxicants or underwent bile duct ligation (BDL, 10 days). Hepatotoxicants included APAP (400 mg/kg, 24-72 h), alpha-naphthyl isothiocyanate (ANIT; 50 mg/kg, 2-48 h), carbon tetrachloride (CCl4; 10 or 30 µL/kg, 24 and 48 h) and allyl alcohol (AlOH; 30 or 60 mg/kg, 6 and 24 h). Because oxidative stress activates nuclear factor (erythroid-derived 2)-like 2 (Nrf2), additional studies investigated Fmo3 gene regulation by Nrf2 using Nrf2 knockout (Nrf2 KO) mice. At appropriate time-points, blood and liver samples were collected for assessment of plasma alanine aminotransferase (ALT) activity, plasma and hepatic bile acid levels, as well as liver Fmo3 mRNA and protein expression. Fmo3 mRNA expression increased significantly by 43-fold at 12 h after ANIT treatment, and this increase translates to a 4-fold change in protein levels. BDL also increased Fmo3 mRNA expression by 1899-fold, but with no change in protein levels. Treatment of mice with CCl4 decreased liver Fmo3 gene expression, while no change in expression was detected with AlOH treatment. Nrf2 KO mice are more susceptible to APAP (400mg/kg, 72 h) treatment compared to their wild-type (WT) counterparts, which is evidenced by greater plasma ALT activity. The Fmo3 mRNA and protein expression increased in Nrf2 KO mice after APAP treatment. Collectively, not all hepatotoxicants that produce oxidative stress alter Fmo3 gene expression. Along with APAP, toxic ANIT treatment in mice markedly increased Fmo3 gene expression. While BDL increased the Fmo3 mRNA expression, the protein level did not change. The discrepancy with Fmo3 induction in cholestatic models, ANIT and BDL, is not entirely clear. Results from Nrf2 KO mice with APAP suggest that the transcriptional regulation of Fmo3 during liver injury may not involve Nrf2.
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Enfermedad Hepática Inducida por Sustancias y Drogas/enzimología , Colestasis/enzimología , Hígado/enzimología , Estrés Oxidativo , Oxigenasas/metabolismo , Alanina Transaminasa/sangre , Animales , Ácidos y Sales Biliares/sangre , Conductos Biliares/cirugía , Biomarcadores/sangre , Enfermedad Hepática Inducida por Sustancias y Drogas/sangre , Enfermedad Hepática Inducida por Sustancias y Drogas/genética , Enfermedad Hepática Inducida por Sustancias y Drogas/patología , Colestasis/sangre , Colestasis/genética , Colestasis/patología , Modelos Animales de Enfermedad , Regulación Enzimológica de la Expresión Génica , Ligadura , Hígado/patología , Masculino , Ratones Endogámicos C57BL , Ratones Noqueados , Factor 2 Relacionado con NF-E2/deficiencia , Factor 2 Relacionado con NF-E2/genética , Oxigenasas/genética , ARN Mensajero/metabolismo , Factores de TiempoRESUMEN
AIMS: The purpose of this study was to determine whether 3'-5'-cyclic adenosine monophosphate (cAMP)-protein kinase A (PKA) and Sirtuin-1 (SIRT1) dependent mechanisms modulate ATP-binding Cassette (ABC) transport protein expression. ABC transport proteins (ABCC2-4) are essential for chemical elimination from hepatocytes and biliary excretion. Nuclear factor-E2 related-factor 2 (NRF2) is a transcription factor that mediates ABCC induction in response to chemical inducers and liver injury. However, a role for NRF2 in the regulation of transporter expression in nonchemical models of liver perturbation is largely undescribed. RESULTS: Here we show that fasting increased NRF2 target gene expression through NRF2- and SIRT1-dependent mechanisms. In intact mouse liver, fasting induces NRF2 target gene expression by at least 1.5 to 5-fold. In mouse and human hepatocytes, treatment with 8-Bromoadenosine-cAMP, a cAMP analogue, increased NRF2 target gene expression and antioxidant response element activity, which was decreased by the PKA inhibitor, H-89. Moreover, fasting induced NRF2 target gene expression was decreased in liver and hepatocytes of SIRT1 liver-specific null mice and NRF2-null mice. Lastly, NRF2 and SIRT1 were recruited to MAREs and Antioxidant Response Elements (AREs) in the human ABCC2 promoter. INNOVATION: Oxidative stress mediated NRF2 activation is well described, yet the influence of basic metabolic processes on NRF2 activation is just emerging. CONCLUSION: The current data point toward a novel role of nutrient status in regulation of NRF2 activity and the antioxidant response, and indicates that cAMP/PKA and SIRT1 are upstream regulators for fasting-induced activation of the NRF2-ARE pathway.
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Transportadoras de Casetes de Unión a ATP/genética , Proteínas Quinasas Dependientes de AMP Cíclico/metabolismo , Ayuno/fisiología , Regulación de la Expresión Génica , Factor 2 Relacionado con NF-E2/genética , Sirtuina 1/metabolismo , Región de Flanqueo 5' , 8-Bromo Monofosfato de Adenosina Cíclica/farmacología , Transportadoras de Casetes de Unión a ATP/metabolismo , Animales , Elementos de Respuesta Antioxidante , Línea Celular , AMP Cíclico/metabolismo , Hepatocitos/metabolismo , Humanos , Hígado/metabolismo , Masculino , Ratones , Proteína 2 Asociada a Resistencia a Múltiples Medicamentos , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/genética , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/metabolismo , NAD(P)H Deshidrogenasa (Quinona)/genética , NAD(P)H Deshidrogenasa (Quinona)/metabolismo , Factor 2 Relacionado con NF-E2/metabolismo , Coactivador 1-alfa del Receptor Activado por Proliferadores de Peroxisomas gamma , Unión Proteica , Transducción de Señal/efectos de los fármacos , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
AIMS: The purpose of this study was to determine whether Nrf2 activation, via Keap1-knockdown (Keap1-KD), regulates lipid metabolism and mobilization induced by food deprivation (e.g. fasting). METHODS AND RESULTS: Male C57BL/6 (WT) and Keap1-KD mice were either fed ad libitum or food deprived for 24 hours. After fasting, WT mice exhibited a marked increase in hepatic lipid accumulation, but Keap1-KD mice had an attenuated increase of lipid accumulation, along with reduced expression of lipogenic genes (acetyl-coA carboxylase, stearoyl-CoA desaturase-1, and fatty acid synthase) and reduced expression of genes related to fatty acid transport, such as fatty acid translocase/CD36 (CD36) and Fatty acid transport protein (FATP) 2, which may attribute to the reduced induction of Peroxisome proliferator-activated receptor (Ppar) α signaling in the liver. Additionally, enhanced Nrf2 activity by Keap1-KD increased AMP-activated protein kinase (AMPK) phosphorylation in liver. In white adipose tissue, enhanced Nrf2 activity did not change the lipolysis rate by fasting, but reduced expression of fatty acid transporters--CD36 and FATP1, via a PPARα-dependent mechanism, which impaired fatty acid transport from white adipose tissue to periphery circulation system, and resulted in increased white adipose tissue fatty acid content. Moreover, enhanced Nrf2 activity increased glucose tolerance and Akt phosphorylation levels upon insulin administration, suggesting Nrf2 signaling pathway plays a key role in regulating insulin signaling and enhanced insulin sensitivity in skeletal muscle. CONCLUSION: Enhanced Nrf2 activity via Keap1-KD decreased fasting-induced steatosis, pointing to an important function of Nrf2 on lipid metabolism under the condition of nutrient deprivation.
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Proteínas Adaptadoras Transductoras de Señales/deficiencia , Tejido Adiposo/metabolismo , Proteínas del Citoesqueleto/deficiencia , Ayuno/metabolismo , Ácidos Grasos/metabolismo , Hígado Graso/metabolismo , Metabolismo de los Lípidos/genética , Proteínas Adaptadoras Transductoras de Señales/genética , Animales , Proteínas del Citoesqueleto/genética , Proteína 1 Asociada A ECH Tipo Kelch , Metabolismo de los Lípidos/fisiología , Masculino , Ratones , Ratones Endogámicos C57BLRESUMEN
PURPOSE: Fatty liver alters liver transporter expression. Caloric restriction (CR), the recommended therapy to reverse fatty liver, increases Sirtuin1 deacetylase activity in liver. This study evaluated whether CR and CR mimetics reversed obesity-induced transporter expression in liver and hepatocytes. METHODS: mRNA and protein expression was determined in adult lean (lean) and leptin-deficient obese (OB) mice fed ad libitum or placed on 40% (kCal) reduced diet. Hepatocytes were isolated from lean and OB mice, treated with AMP Kinase activators, and gene expression was determined. RESULTS: CR decreased Oatp1a1, Oatp1b2, and Abcb11 mRNA expression in lean, but not OB mice. CR increased Abcc2 mRNA OB livers, whereas protein expression increased in both genotypes. CR increased Abcc3 protein expression increased in OB livers. CR did not alter Abcc1, 4 and 5 mRNA expression in lean mice but decreased expression in livers of OB mice. CR increased Abcc4 protein in lean, but not OB mice. CONCLUSIONS: CR restriction reversed the expression of some, but not all transporters in livers of OB mice. Overall, these data indicate a potential for CR to restore some hepatic transporter changes in OB mice, but suggest a functional leptin axis is needed for reversal of expression for some transporters.
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Proteínas Quinasas Activadas por AMP/metabolismo , Restricción Calórica , Regulación de la Expresión Génica , Hígado/metabolismo , Obesidad/dietoterapia , Obesidad/genética , Miembro 11 de la Subfamilia B de Transportador de Casetes de Unión al ATP , Transportadoras de Casetes de Unión a ATP/genética , Animales , Peso Corporal , Células Cultivadas , Activación Enzimática , Lípidos/análisis , Hígado/patología , Transportador 1 de Anión Orgánico Específico del Hígado , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Obesos , Proteína 2 Asociada a Resistencia a Múltiples Medicamentos , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/genética , Obesidad/enzimología , Obesidad/patología , Transportadores de Anión Orgánico Sodio-Independiente/genética , Proteínas de Transporte de Catión Orgánico/genética , ARN Mensajero/genética , Receptores Citoplasmáticos y Nucleares/genéticaRESUMEN
Unsafe use of alcohol results in approximately 2.5 million deaths worldwide, with cirrhosis contributing to 16.6% of reported deaths. Serum insulin levels are often elevated in alcoholism and may result in diabetes, which is why alcoholic liver disease and diabetes often are present together. Because there is a sizable population with these diseases alone or in combination, the purpose of this study was to determine whether transporter expression in human liver is affected by alcoholic cirrhosis, diabetes, and alcoholic cirrhosis coexisting with diabetes. Transporters aid in hepatobiliary excretion of many drugs and toxic chemicals and can be determinants of drug-induced liver injury. Drug transporter expression and transcription factor-relative mRNA and protein expression in normal, diabetic, cirrhotic, and cirrhosis with diabetes human livers were quantified. Cirrhosis significantly increased ABCC4, 5, ABCG2, and solute carrier organic anion (SLCO) 2B1 mRNA expression and decreased SLCO1B3 mRNA expression in the liver. ABCC1, 3-5, and ABCG2 protein expression was also upregulated by alcoholic cirrhosis. ABCC3-5 and ABCG2 protein expression was also upregulated in diabetic cirrhosis. Cirrhosis increased nuclear factor E2-related factor 2 mRNA expression, whereas it decreased pregnane-X-receptor and farnesoid-X-receptor mRNA expression in comparison with normal livers. Hierarchical cluster analysis indicated that expressions of ABCC2, 3, and 6; SLCO1B1 and 1B3; and ABCC4 and 5 were more closely related in the livers from this cohort. Overall, alcoholic cirrhosis altered transporter expression in human liver.
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Cirrosis Hepática Alcohólica/metabolismo , Hígado/metabolismo , Proteínas de Transporte de Membrana/metabolismo , Preparaciones Farmacéuticas/metabolismo , Receptores Citoplasmáticos y Nucleares/metabolismo , Análisis por Conglomerados , Glutatión Peroxidasa/metabolismo , Humanos , Mediadores de Inflamación/metabolismo , Proteínas de Transporte de Membrana/genética , Proteína 2 Asociada a Resistencia a Múltiples Medicamentos , NAD(P)H Deshidrogenasa (Quinona)/metabolismo , Factor 2 Relacionado con NF-E2/metabolismo , ARN Mensajero/genética , Factores de Transcripción/genéticaRESUMEN
Deltamethrin, a type II pyrethroid, is a widely used insecticide. The purpose of this study was to determine whether perinatal deltamethrin exposure altered the expression of adipogenic and lipogenic genes in white adipose tissue (WAT) in adult pups. C57BL/6 pregnant mice were administered 0, 1, or 3 mg/kg of deltamethrin orally every 3 days throughout gestation and lactation. Offspring were weaned on postnatal day 25, and WAT was collected from 5-month-old male mice. Perinatal deltamethrin exposure decreased the mRNA expression of adipogenesis-related transcription factors Pparγ, Cebpα, and lipogenic genes Srebp1c, Acc-1, Cd36, Lpl, Scd-1; along with Nrf2 and target genes Nqo1 and Gclc at the 1 mg/kg treatment. Cytokine expression of Fas/Tnf-R and Cd209e at the 1 mg/kg treatment was significantly decreased, and expression of Tnf, Cd11c, and Fas/Tnf-R was decreased at the 3 mg/kg treatment. Developmental deltamethrin exposure did not overtly affect body weight or adipose weight, but decreased mRNA expression of specific genes that may potentially disrupt normal adipogenesis and lipid and glucose metabolism if the offspring are challenged by changes in diet or environment.
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Tejido Adiposo Blanco/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Insecticidas/toxicidad , Nitrilos/toxicidad , Efectos Tardíos de la Exposición Prenatal/metabolismo , Piretrinas/toxicidad , Adipogénesis/efectos de los fármacos , Tejido Adiposo Blanco/patología , Animales , Femenino , Glucosa/metabolismo , Metabolismo de los Lípidos/efectos de los fármacos , Masculino , Ratones , Embarazo , Efectos Tardíos de la Exposición Prenatal/patologíaRESUMEN
Phase II enzymes, including Ugts, Sults, and Gsts, are critical for the disposition and detoxification of endo- and xenobiotics. In this study, the mRNA and protein expression of major phase II enzymes, as well as key regulatory transcription factors, were quantified in livers of time-matched pregnant and virgin control C57BL/6 mice on gestation days (GD) 7, 11, 14, 17, and postnatal days (PND) 1, 15, and 30. Compared with virgin controls, the mRNA expression of Ugt1a1, 1a6, 1a9, 2a3, 2b1, 2b34, and 2b35 decreased 40 to 80% in pregnant dams. Protein expression of Ugt1a6 also decreased and corresponded with reduced in vitro glucuronidation of bisphenol A in S9 fractions from livers of pregnant mice. Similar to Ugts levels, Gsta1 and a4 mRNAs were reduced in pregnant dams in mid to late gestation; however no change in protein expression was observed. Conversely, Sult1a1, 2a1/2, and 3a1 mRNAs increased 100 to 500% at various time points in pregnant and lactating mice and corresponded with enhanced in vitro sulfation of acetaminophen in liver S9 fractions. Coinciding with maximal decreases in Ugts as well as increases in Sults, the expression of transcription factors CAR, PPARα, and PXR and their target genes were downregulated, whereas ERα mRNA was upregulated. Collectively, these data demonstrate altered regulation of hepatic phase II metabolism in mice during pregnancy and suggest that CAR, PPARα, PXR, and ERα signaling pathways may be candidate signaling pathways responsible for these changes.
Asunto(s)
Hígado/metabolismo , Acetaminofén/metabolismo , Animales , Compuestos de Bencidrilo/metabolismo , Western Blotting , Cromatografía Líquida de Alta Presión , Femenino , Glucurónidos/metabolismo , Glucuronosiltransferasa/metabolismo , Glutatión Transferasa/metabolismo , Técnicas In Vitro , Inactivación Metabólica , Masculino , Ratones , Ratones Endogámicos C57BL , Fenoles/metabolismo , Embarazo , ARN/biosíntesis , ARN/aislamiento & purificación , ARN Mensajero/biosíntesis , ARN Mensajero/genética , Receptores de Estrógenos/biosíntesis , Espectrofotometría Ultravioleta , Fracciones Subcelulares/metabolismo , Sulfatos/metabolismo , Sulfotransferasas/metabolismoRESUMEN
The study herein determined the role of nuclear factor erythoid 2-related factor 2 (Nrf2) in the pathogenesis of hepatic steatosis, insulin resistance, obesity, and type 2 diabetes. Lep(ob/ob)-Keap1-knockdown (KD) mice, which have increased Nrf2 activity, were generated. Markers of obesity and type 2 diabetes were measured in C57Bl/6J, Keap1-KD, Lep(ob/ob), and Lep(ob/ob)-Keap1-KD mice. Lep(ob/ob)-Keap1-KD mice exhibited less lipid accumulation, smaller adipocytes, decreased food intake, and reduced lipogenic gene expression. Enhanced Nrf2 activity impaired insulin signaling, prolonged hyperglycemia in response to glucose challenge, and induced insulin resistance in Lep(ob/ob) background. Nrf2 augmented hepatic steatosis and increased lipid deposition in liver. Next, C57Bl/6J and Keap1-KD mice were fed a high-fat diet (HFD) to determine whether Keap1 and Nrf2 impact HFD-induced obesity. HFD-induced obesity and lipid accumulation in white adipose tissue was decreased in Keap1-KD mice. Nrf2 activation via Keap1-KD or sulforaphane suppressed hormone-induced differentiation and decreased peroxisome proliferator-activated receptor-γ, CCAAT/enhancer-binding protein α, and fatty acid-binding protein 4 expression in mouse embryonic fibroblasts. Constitutive Nrf2 activation inhibited lipid accumulation in white adipose tissue, suppressed adipogenesis, induced insulin resistance and glucose intolerance, and increased hepatic steatosis in Lep(ob/ob) mice.
Asunto(s)
Tejido Adiposo/metabolismo , Hígado Graso/metabolismo , Factor 2 Relacionado con NF-E2/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Animales , Proteínas del Citoesqueleto/genética , Proteínas del Citoesqueleto/metabolismo , Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/metabolismo , Dieta Alta en Grasa/efectos adversos , Hígado Graso/genética , Resistencia a la Insulina/genética , Resistencia a la Insulina/fisiología , Proteína 1 Asociada A ECH Tipo Kelch , Leptina/genética , Leptina/metabolismo , Ratones , Ratones Noqueados , Factor 2 Relacionado con NF-E2/genética , Obesidad/genética , Obesidad/metabolismoRESUMEN
BACKGROUND: Cholestasis is a common disease of the liver. Chronic cholestasis eventually leads to hepatic cirrhosis and fibrosis, and rodent chronic cholestasis models are used to study aspects of fibrosis and cirrhosis. Cholestasis-induced liver injury and fibrosis are associated with increased oxidative stress and inflammation. Few pharmacological therapies exist for treatment of cholestasis or cirrhosis, but it is known that humans with better nutritional intake are less likely to develop certain types of cirrhosis. Eugenia jambolana (Jamun) is a tropical berry fruit rich in antioxidant anthocyanin compounds. AIM: As anthocyanins decrease cellular lipid peroxidation and oxidative stress, it was hypothesized that Jamun fruit extract (JFE) administration could protect against cholestatic liver injury and inflammation in mice. METHOD: Starting 24 h after sham or bile-duct ligation (BDL) surgery, male C57Bl/6 mice were administered vehicle or JFE (100 mg/kg, po) for 10 days. RESULTS: Mice that underwent BDL had elevated serum ALT levels, which were reduced to 60% by JFE treatment. Likewise, BDL caused hepatic inflammation, macrophage infiltration, fibrosis and necrosis, all of which were largely improved by JFE. Interestingly, hepatoprotection was observed in JFE-treated BDL mice, despite suppressed transporter expression and increased hepatic bile acid concentrations. CONCLUSION: Jamun fruit phytochemicals decreased hepatic inflammation and oxidative stress, and protected against hepatocellular injury in mice. Jamun warrants further investigation as a potential antioxidant/anti-inflammatory therapy not only to treat cholestasis but also other liver diseases with an inflammatory component.