RESUMEN
Genetic polymorphisms that impair very low-density lipoprotein (VLDL) secretion are linked to hepatic steatosis, fibrosis, and hepatocellular cancer. Liver-specific deletion of microsomal triglyceride transfer protein (Mttp-LKO) impairs VLDL assembly, promoting hepatic steatosis and fibrosis, which are attenuated in Mttp-LKO X Fabp1-null [Fabp1/Mttp double knockout (DKO)] mice. The current study examined the impact of impaired VLDL secretion in Mttp-LKO mice on hepatocellular cancer incidence and progression in comparison to Fabp1/Mttp DKO mice. Diethylnitrosamine-treated Mttp-LKO mice exhibited steatosis with increased tumor burden compared with flox controls, whereas diethylnitrosamine-treated Fabp1/Mttp DKO mice exhibited a paradoxical increase in tumor burden and >50% mortality by 50 weeks. Serum high-density lipoprotein cholesterol was elevated in both Mttp-LKO and Fabp1/Mttp DKO mice, with increased intratumoral expression of apolipoprotein A1 and apolipoprotein E. Lipidomic surveys revealed progressive enrichment in distinct triglyceride species in livers from Mttp-LKO mice with further enrichment in Fabp1/Mttp DKO mice. RNA sequencing revealed mRNA changes suggesting altered monocarboxylic acid use and increased aerobic glycolysis, whereas hepatocytes from Fabp1/Mttp DKO mice exhibited increased capacity to use glucose and glutamine. These metabolic shifts were accompanied by reduced expression of HNF1 homeobox A (HNF1a), which correlated with tumor burden. Taken together, these findings demonstrate that hepatic tumorigenesis is increased in mice with impaired VLDL secretion and further accelerated via pathways including altered fatty acid compartmentalization and shifts in hepatic energy use.
Asunto(s)
Carcinogénesis , Proteínas de Unión a Ácidos Grasos , Lipoproteínas VLDL , Neoplasias Hepáticas , Animales , Ratones , Carcinogénesis/genética , Carcinogénesis/patología , Carcinogénesis/metabolismo , Carcinoma Hepatocelular/patología , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Proteínas Portadoras/metabolismo , Proteínas Portadoras/genética , Proteínas de Unión a Ácidos Grasos/genética , Proteínas de Unión a Ácidos Grasos/metabolismo , Hígado Graso/metabolismo , Hígado Graso/patología , Hígado Graso/genética , Eliminación de Gen , Lipoproteínas VLDL/metabolismo , Hígado/metabolismo , Hígado/patología , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patología , Ratones NoqueadosRESUMEN
Obesity and the metabolic syndrome are complex disorders resulting from multiple factors including genetics, diet, activity, inflammation, and gut microbes. Animal studies have identified roles for each of these, however the contribution(s) specifically attributed to the gut microbiota remain unclear, as studies have used combinations of genetically altered mice, high fat diet, and/or colonization of germ-free mice, which have an underdeveloped immune system. We investigated the role(s) of the gut microbiota driving obesity and inflammation independent of manipulations in diet and genetics in mice with fully developed immune systems. We demonstrate that the human obese gut microbiota alone was sufficient to drive weight gain, systemic, adipose tissue, and intestinal inflammation, but did not promote intestinal barrier leak. The obese microbiota induced gene expression promoting caloric uptake/harvest but was less effective at inducing genes associated with mucosal immune responses. Thus, the obese gut microbiota is sufficient to induce weight gain and inflammation.
Asunto(s)
Microbioma Gastrointestinal , Humanos , Animales , Ratones , Obesidad/metabolismo , Aumento de Peso , Inflamación/metabolismo , Dieta Alta en Grasa/efectos adversos , Tejido Adiposo/metabolismo , Ratones Endogámicos C57BLRESUMEN
BACKGROUND AND AIMS: We previously identified subsets of patients with NAFLD with different metabolic phenotypes. Here we align metabolomic signatures with cardiovascular disease (CVD) and genetic risk factors. APPROACH AND RESULTS: We analyzed serum metabolome from 1154 individuals with biopsy-proven NAFLD, and from four mouse models of NAFLD with impaired VLDL-triglyceride (TG) secretion, and one with normal VLDL-TG secretion. We identified three metabolic subtypes: A (47%), B (27%), and C (26%). Subtype A phenocopied the metabolome of mice with impaired VLDL-TG secretion; subtype C phenocopied the metabolome of mice with normal VLDL-TG; and subtype B showed an intermediate signature. The percent of patients with NASH and fibrosis was comparable among subtypes, although subtypes B and C exhibited higher liver enzymes. Serum VLDL-TG levels and secretion rate were lower among subtype A compared with subtypes B and C. Subtype A VLDL-TG and VLDL-apolipoprotein B concentrations were independent of steatosis, whereas subtypes B and C showed an association with these parameters. Serum TG, cholesterol, VLDL, small dense LDL5,6 , and remnant lipoprotein cholesterol were lower among subtype A compared with subtypes B and C. The 10-year high risk of CVD, measured with the Framingham risk score, and the frequency of patatin-like phospholipase domain-containing protein 3 NAFLD risk allele were lower in subtype A. CONCLUSIONS: Metabolomic signatures identify three NAFLD subgroups, independent of histological disease severity. These signatures align with known CVD and genetic risk factors, with subtype A exhibiting a lower CVD risk profile. This may account for the variation in hepatic versus cardiovascular outcomes, offering clinically relevant risk stratification.
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Enfermedades Cardiovasculares , Enfermedad del Hígado Graso no Alcohólico , Animales , Apolipoproteínas B , Enfermedades Cardiovasculares/epidemiología , Enfermedades Cardiovasculares/etiología , VLDL-Colesterol/metabolismo , Factores de Riesgo de Enfermedad Cardiaca , Lipoproteínas VLDL , Hígado/patología , Ratones , Enfermedad del Hígado Graso no Alcohólico/patología , Fosfolipasas/metabolismo , Factores de Riesgo , Triglicéridos/metabolismoRESUMEN
Regulating dietary fat absorption may impact progression of nonalcoholic fatty liver disease (NAFLD). Here, we asked if inducible inhibition of chylomicron assembly, as observed in intestine-specific microsomal triglyceride (TG) transfer protein knockout mice (Mttp-IKO), could retard NAFLD progression and/or reverse established fibrosis in two dietary models. Mttp-IKO mice fed a methionine/choline-deficient (MCD) diet exhibited reduced hepatic TGs, inflammation, and fibrosis, associated with reduced oxidative stress and downstream activation of c-Jun N-terminal kinase and nuclear factor kappa B signaling pathways. However, when Mttpflox mice were fed an MCD for 5 weeks and then administered tamoxifen to induce Mttp-IKO, hepatic TG was reduced, but inflammation and fibrosis were increased after 10 days of reversal along with adaptive changes in hepatic lipogenic mRNAs. Extending the reversal time, following 5 weeks of MCD feeding to 30 days led to sustained reductions in hepatic TG, but neither inflammation nor fibrosis was decreased, and both intestinal permeability and hepatic lipogenesis were increased. In a second model, similar reductions in hepatic TG were observed when mice were fed a high-fat/high-fructose/high-cholesterol (HFFC) diet for 10 weeks, then switched to chow ± tamoxifen (HFFC â chow) or (HFFC â Mttp-IKO chow), but again neither inflammation nor fibrosis was affected. In conclusion, we found that blocking chylomicron assembly attenuates MCD-induced NAFLD progression by reducing steatosis, oxidative stress, and inflammation. In contrast, blocking chylomicron assembly in the setting of established hepatic steatosis and fibrosis caused increased intestinal permeability and compensatory shifts in hepatic lipogenesis that mitigate resolution of inflammation and fibrogenic signaling despite 50-90-fold reductions in hepatic TG.
Asunto(s)
Quilomicrones/metabolismo , Hígado Graso/metabolismo , Fibrosis/metabolismo , Inflamación/metabolismo , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Animales , Deficiencia de Colina , Quilomicrones/antagonistas & inhibidores , Dieta/efectos adversos , Femenino , Intestinos/efectos de los fármacos , Intestinos/metabolismo , Masculino , Metionina/deficiencia , Ratones , Ratones Noqueados , Ratones TransgénicosRESUMEN
BACKGROUND AND AIMS: Human transmembrane 6 superfamily 2 (TM6SF2) variant rs58542926 is associated with NAFLD and HCC. However, conflicting reports in germline Tm6sf2 knockout mice suggest no change or decreased very low density lipoprotein (VLDL) secretion and either unchanged or increased hepatic steatosis, with no increased fibrosis. We generated liver-specific Tm6Sf2 knockout mice (Tm6 LKO) to study VLDL secretion and the impact on development and progression of NAFLD. APPROACH AND RESULTS: Two independent lines of Tm6 LKO mice exhibited spontaneous hepatic steatosis. Targeted lipidomic analyses showed increased triglyceride species whose distribution and abundance phenocopied findings in mice with liver-specific deletion of microsomal triglyceride transfer protein. The VLDL triglyceride secretion was reduced with small, underlipidated particles and unchanged or increased apolipoprotein B. Liver-specific adeno-associated viral, serotype 8 (AAV8) rescue using either wild-type or mutant E167K-Tm6 reduced hepatic steatosis and improved VLDL secretion. The Tm6 LKO mice fed a high milk-fat diet for 3 weeks exhibited increased steatosis and fibrosis, and those phenotypes were further exacerbated when mice were fed fibrogenic, high fat/fructose diets for 20 weeks. In two models of HCC, either neonatal mice injected with streptozotocin (NASH/STAM) and high-fat fed or with diethylnitrosamine injection plus fibrogenic diet feeding, Tm6 LKO mice exhibited increased steatosis, greater tumor burden, and increased tumor area versus Tm6 flox controls. Additionally, diethylnitrosamine-injected and fibrogenic diet-fed Tm6 LKO mice administered wild-type Tm6 or E167K-mutant Tm6 AAV8 revealed significant tumor attenuation, with tumor burden inversely correlated with Tm6 protein levels. CONCLUSIONS: Liver-specific Tm6sf2 deletion impairs VLDL secretion, promoting hepatic steatosis, fibrosis, and accelerated development of HCC, which was mitigated with AAV8- mediated rescue.
Asunto(s)
Carcinoma Hepatocelular/genética , Hígado Graso/genética , Lipoproteínas VLDL/metabolismo , Cirrosis Hepática/genética , Neoplasias Hepáticas/genética , Hígado/metabolismo , Proteínas de la Membrana/genética , Animales , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patología , Hígado Graso/metabolismo , Lipidómica , Hígado/patología , Cirrosis Hepática/metabolismo , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patología , Ratones , Ratones Noqueados , Enfermedad del Hígado Graso no Alcohólico/genética , Triglicéridos/metabolismoRESUMEN
We previously established that global deletion of the enhancer of trithorax and polycomb (ETP) gene, Asxl2, prevents weight gain. Because proinflammatory macrophages recruited to adipose tissue are central to the metabolic complications of obesity, we explored the role of ASXL2 in myeloid lineage cells. Unexpectedly, mice without Asxl2 only in myeloid cells (Asxl2ΔLysM) were completely resistant to diet-induced weight gain and metabolically normal despite increased food intake, comparable activity, and equivalent fecal fat. Asxl2ΔLysM mice resisted HFD-induced adipose tissue macrophage infiltration and inflammatory cytokine gene expression. Energy expenditure and brown adipose tissue metabolism in Asxl2ΔLysM mice were protected from the suppressive effects of HFD, a phenomenon associated with relatively increased catecholamines likely due to their suppressed degradation by macrophages. White adipose tissue of HFD-fed Asxl2ΔLysM mice also exhibited none of the pathological remodeling extant in their control counterparts. Suppression of macrophage Asxl2 expression, via nanoparticle-based siRNA delivery, prevented HFD-induced obesity. Thus, ASXL2 controlled the response of macrophages to dietary factors to regulate metabolic homeostasis, suggesting modulation of the cells' inflammatory phenotype may impact obesity and its complications.
Asunto(s)
Metabolismo Energético , Células Mieloides/metabolismo , Obesidad/prevención & control , Proteínas Represoras/deficiencia , Tejido Adiposo Pardo/metabolismo , Tejido Adiposo Pardo/patología , Tejido Adiposo Blanco/metabolismo , Tejido Adiposo Blanco/patología , Animales , Dieta Alta en Grasa/efectos adversos , Femenino , Técnicas de Silenciamiento del Gen , Inflamación/metabolismo , Inflamación/patología , Macrófagos/metabolismo , Macrófagos/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Células Mieloides/patología , Obesidad/metabolismo , Obesidad/patología , Especificidad de Órganos , ARN Interferente Pequeño/genética , Proteínas Represoras/genética , Proteínas Represoras/metabolismo , Aumento de Peso/genética , Aumento de Peso/fisiologíaRESUMEN
The mechanisms by which alterations in intestinal bile acid (BA) metabolism improve systemic glucose tolerance and hepatic metabolic homeostasis are incompletely understood. We examined metabolic adaptations in mice with conditional intestinal deletion of the abetalipoproteinemia (ABL) gene microsomal triglyceride transfer protein (Mttp-IKO), which blocks chylomicron assembly and impairs intestinal lipid transport. Mttp-IKO mice exhibit improved hepatic glucose metabolism and augmented insulin signaling, without weight loss. These adaptations included decreased BA excretion, increased pool size, altered BA composition, and increased fibroblast growth factor 15 production. Mttp-IKO mice absorb fructose normally but are protected against dietary fructose-induced hepatic steatosis, without weight loss or changes in energy expenditure. In addition, Mttp-IKO mice exhibit altered cecal microbial communities, both at baseline and following fructose feeding, including increased abundance of Bacteroides and Lactobacillus genera. Transplantation of cecal microbiota from chow-fed Mttp-IKO mice into antibiotic-treated wild-type recipients conferred transmissible protection against fructose-induced hepatic steatosis in association with a bloom in Akkermansia and increased Clostridium XIVa genera, whose abundance was positively correlated with fecal coprostanol and total neutral sterol excretion in recipient mice. However, antibiotic-treated Mttp-IKO mice were still protected against fructose-induced hepatic steatosis, suggesting that changes in microbiota are not required for this phenotype. Nevertheless, we found increased abundance of fecal Akkermansia from two adult ABL subjects with MTTP mutations compared to their heterozygous parents and within the range noted in six healthy control subjects. Furthermore, Akkermansia abundance across all subjects was positively correlated with fecal coprostanol excretion. Conclusion: The findings collectively suggest multiple adaptive pathways of metabolic regulation following blocked chylomicron assembly, including shifts in BA signaling and altered microbial composition that confer a transmissible phenotype.
Asunto(s)
Adaptación Fisiológica/genética , Quilomicrones/genética , Hígado Graso/metabolismo , Microbioma Gastrointestinal/genética , Metabolismo de los Lípidos/genética , Akkermansia , Animales , Ácidos y Sales Biliares/metabolismo , Transporte Biológico/genética , Proteínas Portadoras/metabolismo , Modelos Animales de Enfermedad , Hígado Graso/patología , Fructosa/farmacología , Prueba de Tolerancia a la Glucosa , Humanos , Mucosa Intestinal/metabolismo , Ratones , Ratones Endogámicos C57BL , Distribución Aleatoria , Sensibilidad y Especificidad , Transducción de Señal , Verrucomicrobia/patogenicidadRESUMEN
Liver fatty acid binding protein (L-Fabp) modulates lipid trafficking in enterocytes, hepatocytes, and hepatic stellate cells (HSCs). We examined hepatocyte vs. HSC L-Fabp deletion in hepatic metabolic adaptation and fibrotic injury. Floxed L-Fabp mice were bred to different transgenic Cre mice or injected with adeno-associated virus type 8 (AAV8) Cre and fed diets to promote steatosis and fibrosis or were subjected to either bile duct ligation or CCl4 injury. Albumin-Cre-mediated L-Fabp deletion revealed recombination in hepatocytes and HSCs; these findings were confirmed with 2 other floxed alleles. Glial fibrillary acid protein-Cre and platelet-derived growth factor receptor ß-Cre-mediated L-Fabp deletion demonstrated recombination only in HSCs. Mice with albumin promoter-driven Cre recombinase (Alb-Cre)-mediated or AAV8-mediated L-Fabp deletion were protected against food withdrawal-induced steatosis. Mice with Alb-Cre-mediated L-Fabp deletion were protected against high saturated fat-induced steatosis and fibrosis, phenocopying germline L-Fabp-/- mice. Mice with HSC-specific L-Fabp deletion exhibited retinyl ester depletion yet demonstrated no alterations in fibrosis. On the other hand, fibrogenic resolution after CCl4 administration was impaired in mice with Alb-Cre-mediated L-Fabp deletion. These findings suggest cell type-specific roles for L-Fabp in mitigating hepatic steatosis and in modulating fibrogenic injury and reversal.-Newberry, E. P., Xie, Y., Lodeiro, C., Solis, R., Moritz, W., Kennedy, S., Barron, L., Onufer, E., Alpini, G., Zhou, T., Blaner, W. S., Chen, A., Davidson, N. O. Hepatocyte and stellate cell deletion of liver fatty acid binding protein reveal distinct roles in fibrogenic injury.
Asunto(s)
Intoxicación por Tetracloruro de Carbono/metabolismo , Proteínas de Unión a Ácidos Grasos/fisiología , Hígado Graso/metabolismo , Células Estrelladas Hepáticas/metabolismo , Hepatocitos/metabolismo , Cirrosis Hepática/metabolismo , Albúminas/genética , Animales , Conductos Biliares , Intoxicación por Tetracloruro de Carbono/patología , Cruzamientos Genéticos , Dependovirus/genética , Grasas de la Dieta/toxicidad , Proteínas de Unión a Ácidos Grasos/deficiencia , Ácidos Grasos/toxicidad , Hígado Graso/etiología , Hígado Graso/patología , Femenino , Fibrosis , Privación de Alimentos , Eliminación de Gen , Genes Sintéticos , Células Estrelladas Hepáticas/patología , Hepatocitos/patología , Integrasas , Ligadura , Cirrosis Hepática/inducido químicamente , Cirrosis Hepática/patología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Especificidad de Órganos , Regiones Promotoras GenéticasRESUMEN
Hepatic stellate cell (HSC) activation occurs along with decreased Perilipin5 (Plin5) and liver fatty acid-binding protein (L-Fabp) expression and coincident lipid droplet (LD) depletion. Conversely, the activated phenotype is reversible in WT HSCs upon forced expression of Plin5. Here, we asked if L-Fabp expression is required for Plin5-mediated rescue of the quiescent phenotype. Lentiviral Plin5 transduction of passaged L-Fabp-/- HSCs failed to reverse activation markers or restore lipogenic gene expression and LD formation. However, adenoviral L-Fabp infection of lentiviral Plin5 transduced L-Fabp-/- HSCs restored both the quiescent phenotype and LD formation, an effect also mediated by adenoviral intestine-Fabp or adipocyte-Fabp. Expression of exogenous Plin5 in activated WT HSCs induced a transcriptional program of lipogenic gene expression including endogenous L-Fabp, but none of the other FABPs. We further demonstrated that selective, small molecule inhibition of endogenous L-Fabp also eliminated the ability of exogenous Plin5 to rescue LD formation and reverse activation of WT HSCs. This functional coordination of L-Fabp with Plin5 was 5'-AMP-activated protein kinase (AMPK)-dependent and was eliminated by AMPK inhibition. Taken together, our results indicate that L-Fabp is required for Plin5 to activate a transcriptional program that restores LD formation and reverses HSC activation.
Asunto(s)
Proteínas de Unión a Ácidos Grasos/metabolismo , Células Estrelladas Hepáticas/citología , Células Estrelladas Hepáticas/metabolismo , Perilipina-5/metabolismo , Animales , Células Cultivadas , Proteínas de Unión a Ácidos Grasos/deficiencia , Femenino , Gotas Lipídicas/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Perilipina-5/antagonistas & inhibidores , Perilipina-5/genética , Bibliotecas de Moléculas Pequeñas/farmacologíaRESUMEN
The scavenger receptor and multiligand transporter CD36 functions to promote cellular free fatty acid uptake and regulates aspects of both hepatic and intestinal cholesterol metabolism. However, the role of CD36 in regulating canalicular and biliary cholesterol transport and secretion is unknown. Here, we show that germline Cd36 knockout (KO) mice are protected against lithogenic diet (LD)-induced gallstones compared with congenic (C57BL6/J) controls. Cd36 KO mice crossed into congenic L-Fabp KO mice (DKO mice) demonstrated protection against LD-induced gallstones, reversing the susceptibility phenotype observed in L-Fabp KO mice. DKO mice demonstrated reduced biliary cholesterol secretion and a shift into more hydrophophilic bile acid species, without changes in either BA pool size or fecal excretion. In addition, we found that the mean and maximum force of gallbladder contraction was increased in germline Cd36 KO mice, and gallbladder lipid content was reduced compared with wild-type controls. Finally, whereas germline Cd36 KO mice were protected against LD-induced gallstones, neither liver- nor intestine-specific Cd36 KO mice were protected. Taken together, our findings show that CD36 plays an important role in modifying gallstone susceptibility in mice, at least in part by altering biliary lipid composition, but also by promoting gallbladder contractility.
Asunto(s)
Antígenos CD36/deficiencia , Antígenos CD36/genética , Dieta/efectos adversos , Cálculos Biliares/genética , Animales , Ácidos y Sales Biliares/química , Ácidos y Sales Biliares/metabolismo , Colesterol/metabolismo , Vesícula Biliar/metabolismo , Vesícula Biliar/fisiopatología , Cálculos Biliares/etiología , Cálculos Biliares/metabolismo , Cálculos Biliares/fisiopatología , Técnicas de Inactivación de Genes , Hígado/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Contracción Muscular/genéticaRESUMEN
Blocking hepatic very low-density lipoprotein secretion through genetic or pharmacologic inhibition of microsomal triglyceride transfer protein (Mttp) causes hepatic steatosis, yet the risks for developing hepatic fibrosis are poorly understood. We report that liver-specific Mttp knockout mice (Mttp-LKO) exhibit both steatosis and fibrosis, which is exacerbated by a high-transfat/fructose diet. When crossed into germline liver fatty acid (FA) binding protein null mice (Mttp-LKO, i.e., double knockout mice) hepatic steatosis was greatly diminished and fibrosis prevented, on both low-fat and high-fat diets. The mechanisms underlying protection include reduced long chain FA uptake, shifts in FA distribution (lipidomic profiling), and metabolic turnover, specifically decreased hepatic 18:2 FA and triglyceride species and a shift in 18:2 FA use for oxidation versus incorporation into newly synthesized triglyceride. Double knockout mice were protected against fasting-induced hepatic steatosis (a model of enhanced exogenous FA delivery) yet developed steatosis upon induction of hepatic de novo lipogenesis with fructose feeding. Mttp-LKO mice, on either the liver FA binding protein null or Apobec-1 null background (i.e., apolipoprotein B100 only) exhibited only subtle increases in endoplasmic reticulum stress, suggesting that an altered unfolded protein response is unlikely to account for the attenuated phenotype in double knockout mice. Acute, antisense-mediated liver FA binding protein knockdown in Mttp-LKO mice also reduced FA uptake, increased oxidation versus incorporation of 18:2 species with complete reversal of hepatic steatosis, increased hepatic injury, and worsened fibrosis. CONCLUSION: Perturbing exogenous hepatic FA use modulates both hepatic steatosis and fibrosis in the setting of hepatic Mttp deletion, adding new insight into the pathophysiological mechanisms and consequences of defective very low-density lipoprotein secretion. (Hepatology 2017;65:836-852).
Asunto(s)
Proteínas Portadoras/genética , Proteínas de Unión a Ácidos Grasos/metabolismo , Hígado Graso/metabolismo , Metabolismo de los Lípidos/genética , Cirrosis Hepática/prevención & control , Animales , Biopsia con Aguja , Células Cultivadas , VLDL-Colesterol/metabolismo , Modelos Animales de Enfermedad , Hígado Graso/patología , Hepatocitos/metabolismo , Inmunohistoquímica , Metabolismo de los Lípidos/fisiología , Lipogénesis/fisiología , Cirrosis Hepática/genética , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Oxidación-Reducción , Distribución Aleatoria , Valores de ReferenciaRESUMEN
Trehalose is a naturally occurring disaccharide that has gained attention for its ability to induce cellular autophagy and mitigate diseases related to pathological protein aggregation. Despite decades of ubiquitous use as a nutraceutical, preservative, and humectant, its mechanism of action remains elusive. We showed that trehalose inhibited members of the SLC2A (also known as GLUT) family of glucose transporters. Trehalose-mediated inhibition of glucose transport induced AMPK (adenosine 5'-monophosphate-activated protein kinase)-dependent autophagy and regression of hepatic steatosis in vivo and a reduction in the accumulation of lipid droplets in primary murine hepatocyte cultures. Our data indicated that trehalose triggers beneficial cellular autophagy by inhibiting glucose transport.
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Autofagia , Hígado Graso/metabolismo , Proteínas Facilitadoras del Transporte de la Glucosa/metabolismo , Trehalosa/metabolismo , Proteínas Quinasas Activadas por AMP/genética , Proteínas Quinasas Activadas por AMP/metabolismo , Animales , Hígado Graso/genética , Hígado Graso/patología , Proteínas Facilitadoras del Transporte de la Glucosa/genética , Células HEK293 , Células Hep G2 , Humanos , Ratones , Ratones NoqueadosRESUMEN
Phenotypic divergence in diet-induced obesity (DIO) and hepatic steatosis has been reported in two independently generated lines of L-Fabp(-/-) mice [New Jersey (NJ) L-Fabp(-/-) vs. Washington University (WU) L-Fabp(-/-) mice]. We performed side-by-side studies to examine differences between the lines and investigate the role of genetic background, intestinal microbiota, sex, and diet in the divergent phenotypes. Fasting-induced steatosis was attenuated in both L-Fabp(-/-) lines compared with C57BL/6J controls, with restoration of hepatic triglyceride levels following adenoviral L-Fabp rescue. Both lines were protected against DIO after high-saturated-fat diet feeding. Hepatic steatosis was attenuated in WU but not NJ L-Fabp(-/-) mice, although this difference between the lines disappeared upon antibiotic treatment and cohousing. In contrast, there was phenotypic divergence in L-Fabp(-/-) mice fed a high cocoa butter fat diet, with WU L-Fabp(-/-) mice, but not NJ L-Fabp(-/-) mice, showing protection against both DIO and hepatic steatosis, with some sex-dependent (female > male) differences. Dense mapping revealed no evidence of unintended targeting, duplications, or deletions surrounding the Fabp1 locus in either line and only minor differences in mRNA expression of genes located near the targeted allele. However, a C57BL/6 substrain screen showed that the NJ L-Fabp(-/-) line contains â¼40% C57BL/6N genomic DNA, despite reports that these mice were backcrossed six generations. Overall, these findings suggest that some of the phenotypic divergence between the two L-Fabp(-/-) lines may reflect unanticipated differences in genetic background, underscoring the importance of genetic background in phenotypic characterization.
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Proteínas de Unión a Ácidos Grasos/metabolismo , Regulación de la Expresión Génica/fisiología , Animales , Antibacterianos/administración & dosificación , Antibacterianos/farmacología , Grasas de la Dieta , Proteínas de Unión a Ácidos Grasos/genética , Hígado Graso/etiología , Hígado Graso/genética , Hígado Graso/prevención & control , Femenino , Privación de Alimentos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Obesidad/genética , Obesidad/metabolismo , Obesidad/prevención & control , Fenotipo , ARN Mensajero/genética , ARN Mensajero/metabolismoRESUMEN
Previous studies demonstrated that L-Fabp KO mice are more susceptible to lithogenic diet (LD)-induced gallstones because of altered hepatic cholesterol metabolism and increased canalicular cholesterol secretion. Other studies demonstrated that liver-specific deletion of microsomal triglyceride transfer protein (Mttp-LKO) reduced LD-induced gallstone formation by increasing biliary phospholipid secretion. Here we show that mice with combined deletion (i.e., DKO mice) are protected from LD-induced gallstone formation. Following 2 weeks of LD feeding, 73% of WT and 100% of L-Fabp KO mice developed gallstones versus 18% of Mttp-LKO and 23% of DKO mice. This phenotype was recapitulated in both WT and L-Fabp KO mice treated with an Mttp antisense oligonucleotide (M-ASO). Biliary cholesterol secretion was increased in LD-fed L-Fabp KO mice and decreased in DKO mice. However, phospholipid secretion was unchanged in LD-fed Mttp-LKO and DKO mice as well as in M-ASO-treated mice. Expression of the canalicular export pump ABCG5/G8 was reduced in LD-fed DKO mice and in M-ASO-treated L-Fabp KO mice. We conclude that liver-specific Mttp deletion not only eliminates apical lipoprotein secretion from hepatocytes but also attenuates canalicular cholesterol secretion, which in turn decreases LD-induced gallstone susceptibility.
Asunto(s)
Proteínas Portadoras/metabolismo , Proteínas de Unión a Ácidos Grasos/deficiencia , Cálculos Biliares/metabolismo , Hígado/metabolismo , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 5 , Transportador de Casete de Unión a ATP, Subfamilia G, Miembro 8 , Transportadoras de Casetes de Unión a ATP/genética , Transportadoras de Casetes de Unión a ATP/metabolismo , Animales , Bilis/efectos de los fármacos , Bilis/metabolismo , Proteínas Portadoras/genética , Colesterol/sangre , Colesterol/metabolismo , Dieta/efectos adversos , Proteínas de Unión a Ácidos Grasos/genética , Cálculos Biliares/etiología , Cálculos Biliares/genética , Expresión Génica/efectos de los fármacos , Predisposición Genética a la Enfermedad/etiología , Predisposición Genética a la Enfermedad/genética , Immunoblotting , Lípidos/análisis , Lípidos/sangre , Lipoproteínas/genética , Lipoproteínas/metabolismo , Hígado/efectos de los fármacos , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Oligonucleótidos Antisentido/administración & dosificación , Oligonucleótidos Antisentido/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Triglicéridos/sangre , Triglicéridos/metabolismoRESUMEN
Evidence suggests a relationship between dietary fat intake, obesity, and colorectal cancer, implying a role for fatty acid metabolism in intestinal tumorigenesis that is incompletely understood. Liver fatty acid-binding protein (L-Fabp), a dominant intestinal fatty acid-binding protein, regulates intestinal fatty acid trafficking and metabolism, and L-Fabp deletion attenuates diet-induced obesity. Here, we examined whether changes in intestinal fatty acid metabolism following L-Fabp deletion modify adenoma development in Apc(Min)(/+) mice. Compound L-Fabp(-/-)Apc(Min)(/+) mice were generated and fed a 10% fat diet balanced equally between saturated, monounsaturated, and polyunsaturated fat. L-Fabp(-/-)Apc(Min)(/+) mice displayed significant reductions in adenoma number and total polyp area compared with Apc(Min)(/+)controls, reflecting a significant shift in distribution toward smaller polyps. Adenomas from L-Fabp(-/-)Apc(Min)(/+) mice exhibited reductions in cellular proliferation, high-grade dysplasia, and nuclear ß-catenin translocation. Intestinal fatty acid content was increased in L-Fabp(-/-)Apc(Min)(/+) mice, and lipidomic profiling of intestinal mucosa revealed significant shifts to polyunsaturated fatty acid species with reduced saturated fatty acid species. L-Fabp(-/-)Apc(Min)(/+) mice also showed corresponding changes in mRNA expression of enzymes involved in fatty acid elongation and desaturation. Furthermore, adenomas from L-Fabp(-/-)Apc(Min)(/+) mice displayed significant reductions in mRNA abundance of nuclear hormone receptors involved in cellular proliferation and in enzymes involved in lipogenesis. These findings collectively implicate L-Fabp as an important genetic modifier of intestinal tumorigenesis, and identify fatty acid trafficking and metabolic compartmentalization as an important pathway linking dietary fat intake, obesity, and intestinal tumor formation.
Asunto(s)
Adenoma/metabolismo , Proteínas de Unión a Ácidos Grasos/genética , Proteínas de Unión a Ácidos Grasos/metabolismo , Ácidos Grasos/metabolismo , Regulación Neoplásica de la Expresión Génica , Mucosa Intestinal/metabolismo , Animales , Proliferación Celular , Grasas de la Dieta , Dinoprostona/metabolismo , Femenino , Eliminación de Gen , Genotipo , Inmunohistoquímica , Lípidos/química , Ratones , Ratones Transgénicos , Reacción en Cadena de la Polimerasa , ARN Mensajero/metabolismo , Transducción de Señal , Factores de TiempoRESUMEN
UNLABELLED: Activation of hepatic stellate cells (HSCs) is crucial to the development of fibrosis in nonalcoholic fatty liver disease. Quiescent HSCs contain lipid droplets (LDs), whose depletion upon activation induces a fibrogenic gene program. Here we show that liver fatty acid-binding protein (L-Fabp), an abundant cytosolic protein that modulates fatty acid (FA) metabolism in enterocytes and hepatocytes, also modulates HSC FA utilization and in turn regulates the fibrogenic program. L-Fabp expression decreased 10-fold following HSC activation, concomitant with depletion of LDs. Primary HSCs isolated from L-FABP(-/-) mice contain fewer LDs than wild-type (WT) HSCs, and exhibit up-regulated expression of genes involved in HSC activation. Adenoviral L-Fabp transduction inhibited activation of passaged WT HSCs and increased both the expression of prolipogenic genes and also augmented intracellular lipid accumulation, including triglyceride and FA, predominantly palmitate. Freshly isolated HSCs from L-FABP(-/-) mice correspondingly exhibited decreased palmitate in the free FA pool. To investigate whether L-FABP deletion promotes HSC activation in vivo, we fed L-FABP(-/-) and WT mice a high-fat diet supplemented with trans-fatty acids and fructose (TFF). TFF-fed L-FABP(-/-) mice exhibited reduced hepatic steatosis along with decreased LD abundance and size compared to WT mice. In addition, TFF-fed L-FABP(-/-) mice exhibited decreased hepatic fibrosis, with reduced expression of fibrogenic genes, compared to WT mice. CONCLUSION: L-FABP deletion attenuates both diet-induced hepatic steatosis and fibrogenesis, despite the observation that L-Fabp paradoxically promotes FA and LD accumulation and inhibits HSC activation in vitro. These findings highlight the importance of cell-specific modulation of hepatic lipid metabolism in promoting fibrogenesis in nonalcoholic fatty liver disease. (Hepatology 2013).
Asunto(s)
Proteínas de Unión a Ácidos Grasos/metabolismo , Hígado Graso/metabolismo , Células Estrelladas Hepáticas/fisiología , Hepatocitos/metabolismo , Metabolismo de los Lípidos , Animales , Grasas de la Dieta/efectos adversos , Modelos Animales de Enfermedad , Ácidos Grasos/metabolismo , Hígado Graso/etiología , Femenino , Fibrosis , Fructosa/efectos adversos , Técnicas de Transferencia de Gen , Lipogénesis , Hígado/patología , Ratones , Ratones Endogámicos C57BL , Perilipina-5 , Proteínas/metabolismo , Triglicéridos/metabolismoRESUMEN
The tissue-specific sources and regulated production of physiological signals that modulate food intake are incompletely understood. Previous work showed that L-Fabp(-/-) mice are protected against obesity and hepatic steatosis induced by a high-fat diet, findings at odds with an apparent obesity phenotype in a distinct line of aged L-Fabp(-/-) mice. Here we show that the lean phenotype in L-Fabp(-/-) mice is recapitulated in aged, chow-fed mice and correlates with alterations in hepatic, but not intestinal, fatty acid amide metabolism. L-Fabp(-/-) mice exhibited short-term changes in feeding behavior with decreased food intake, which was associated with reduced abundance of key signaling fatty acid ethanolamides, including oleoylethanolamide (OEA, an agonist of PPARα) and anandamide (AEA, an agonist of cannabinoid receptors), in the liver. These reductions were associated with increased expression and activity of hepatic fatty acid amide hydrolase-1, the enzyme that degrades both OEA and AEA. Moreover, L-Fabp(-/-) mice demonstrated attenuated responses to OEA administration, which was completely reversed with an enhanced response after administration of a nonhydrolyzable OEA analog. These findings demonstrate a role for L-Fabp in attenuating obesity and hepatic steatosis, and they suggest that hepatic fatty acid amide metabolism is altered in L-Fabp(-/-) mice.
Asunto(s)
Peso Corporal , Proteínas de Unión a Ácidos Grasos/metabolismo , Hígado Graso/patología , Ácidos Oléicos/farmacología , Adiposidad , Factores de Edad , Amidohidrolasas/genética , Amidohidrolasas/metabolismo , Animales , Ácidos Araquidónicos/administración & dosificación , Ácidos Araquidónicos/farmacología , Cromosomas/genética , Cromosomas/metabolismo , Dieta con Restricción de Grasas , Endocannabinoides , Activación Enzimática , Proteínas de Unión a Ácidos Grasos/genética , Hígado Graso/genética , Conducta Alimentaria , Femenino , Metabolismo de los Lípidos , Hígado/efectos de los fármacos , Hígado/metabolismo , Hígado/patología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Obesidad/metabolismo , Obesidad/patología , Ácidos Oléicos/administración & dosificación , PPAR gamma/genética , PPAR gamma/metabolismo , Alcamidas Poliinsaturadas/administración & dosificación , Alcamidas Poliinsaturadas/farmacología , Sitios de Carácter Cuantitativo , Transducción de SeñalAsunto(s)
Antígenos CD36/metabolismo , Péptido 2 Similar al Glucagón/fisiología , Absorción Intestinal/fisiología , Metabolismo de los Lípidos/efectos de los fármacos , Animales , Antígenos CD36/deficiencia , Quilomicrones/metabolismo , Ácidos Grasos/metabolismo , Péptido 2 Similar al Glucagón/farmacología , Humanos , Absorción Intestinal/efectos de los fármacosRESUMEN
Quantitative trait mapping in mice identified a susceptibility locus for gallstones (Lith6) spanning the Apobec-1 locus, the structural gene encoding the RNA-specific cytidine deaminase responsible for production of apolipoprotein B48 in mammalian small intestine and rodent liver. This observation prompted us to compare dietary gallstone susceptibility in Apobec-1(-/-) mice and congenic C57BL/6 wild type controls. When fed a lithogenic diet (LD) for 2 weeks, 90% Apobec-1(-/-) mice developed solid gallstones in comparison with 16% wild type controls. LD-fed Apobec-1(-/-) mice demonstrated increased biliary cholesterol secretion as well as increased cholesterol saturation and bile acid hydrophobicity indices. These changes occurred despite a relative decrease in cholesterol absorption in LD-fed Apobec-1(-/-) mice. Among the possible mechanisms to account for this phenotype, expression of Cyp7a1 mRNA and protein were significantly decreased in chow-fed Apobec-1(-/-) mice, decreasing further in LD-fed animals. Cyp7a1 transcription in hepatocyte nuclei, however, was unchanged in Apobec-1(-/-) mice, excluding transcriptional repression as a potential mechanism for decreased Cyp7a1 expression. We demonstrated that APOBEC-1 binds to AU-rich regions of the 3'-untranslated region of the Cyp7a1 transcript, containing the UUUN(A/U)U consensus motif, using both UV cross-linking to recombinant APOBEC-1 and in vivo RNA co-immunoprecipitation. In vivo Apobec-1-dependent modulation of Cyp7a1 expression was further confirmed following adenovirus-Apobec-1 administration to chow-fed Apobec-1(-/-) mice, which rescued Cyp7a1 gene expression. Taken together, the findings suggest that the AU-rich RNA binding-protein Apobec-1 mediates post-transcriptional regulation of murine Cyp7a1 expression and influences susceptibility to diet-induced gallstone formation.