RESUMO
Maternal obesity and consumption of a high-fat diet significantly elevate risk for pediatric nonalcoholic fatty liver disease (NAFLD), affecting 10% of children in the US. Almost half of these children are diagnosed with nonalcoholic steatohepatitis (NASH), a leading etiology for liver transplant. Animal models show that signs of liver injury and perturbed lipid metabolism associated with NAFLD begin in utero; however, safe dietary therapeutics to blunt developmental programming of NAFLD are unavailable. Using a mouse model of maternal Western-style diet (WD), we previously showed that pyrroloquinoline quinone (PQQ), a potent dietary antioxidant, protected offspring of WD-fed dams from development of NAFLD and NASH. Here, we used untargeted mass spectrometry-based lipidomics to delineate lipotoxic effects of WD on offspring liver and identify lipid targets of PQQ. PQQ exposure during pregnancy altered hepatic lipid profiles of WD-exposed offspring, upregulating peroxisome proliferator-activated receptor (PPAR) α signaling and mitochondrial fatty acid oxidation to markedly attenuate triglyceride accumulation beginning in utero. Surprisingly, the abundance of very long-chain ceramides, important in promoting gut barrier and hepatic function, was significantly elevated in PQQ-treated offspring. PQQ exposure reduced the hepatic phosphatidylcholine/phosphatidylethanolamine (PC/PE) ratio in WD-fed offspring and improved glucose tolerance. Notably, levels of protective n - 3 polyunsaturated fatty acids (PUFAs) were elevated in offspring exposed to PQQ, beginning in utero, and the increase in n - 3 PUFAs persisted into adulthood. Our findings suggest that PQQ supplementation during gestation and lactation augments pathways involved in the biosynthesis of long-chain fatty acids and plays a unique role in modifying specific bioactive lipid species critical for protection against NAFLD risk in later life.
Assuntos
Ácidos Graxos Ômega-3 , Hepatopatia Gordurosa não Alcoólica , Adulto , Animais , Criança , Dieta Hiperlipídica/efeitos adversos , Suplementos Nutricionais , Ácidos Graxos Ômega-3/metabolismo , Feminino , Humanos , Metabolismo dos Lipídeos , Fígado/metabolismo , Longevidade , Hepatopatia Gordurosa não Alcoólica/metabolismo , Estresse Oxidativo , PPAR alfa/metabolismo , Cofator PQQ/farmacologia , GravidezRESUMO
The epigenetic marker 5-methylcytosine (5mC) is an important factor in DNA modification and epigenetics. It can be modified through a three-step oxidation performed by ten-eleven-translocation (TET) enzymes and we have previously reported that the iron(IV)-oxo complex [Fe(O)(Py5 Me2 H)]2+ (1) can oxidize 5mC. Here, we report the reactivity of this iron(IV)-oxo complex towards a wider scope of methylated cytosine and uracil derivatives relevant for synthetic DNA applications, such as 1-methylcytosine (1mC), 5-methyl-iso-cytosine (5miC) and thymine (T/5mU). The observed kinetic parameters are corroborated by calculation of the C-H bond energies at the reactive sites which was found to be an efficient tool for reaction rate prediction of 1 towards methylated DNA bases. We identified oxidation products of methylated cytosine derivatives using HPLC-MS and GC-MS. Thereby, we shed light on the impact of the methyl group position and resulting C-H bond dissociation energies on reactivity towards TET-like oxidation.
Assuntos
5-Metilcitosina/química , DNA/síntese química , Compostos de Ferro/química , DNA/química , Humanos , Cinética , Modelos Moleculares , Estrutura Molecular , Oxirredução , Termodinâmica , Uracila/químicaRESUMO
Nonalcoholic fatty liver disease (NAFLD) is widespread in adults and children. Early exposure to maternal obesity or Western-style diet (WD) increases steatosis and oxidative stress in fetal liver and is associated with lifetime disease risk in the offspring. Pyrroloquinoline quinone (PQQ) is a natural antioxidant found in soil, enriched in human breast milk, and essential for development in mammals. We investigated whether a supplemental dose of PQQ, provided prenatally in a mouse model of diet-induced obesity during pregnancy, could protect obese offspring from progression of NAFLD. PQQ treatment given pre- and postnatally in WD-fed offspring had no effect on weight gain but increased metabolic flexibility while reducing body fat and liver lipids, compared with untreated obese offspring. Indices of NAFLD, including hepatic ceramide levels, oxidative stress, and expression of proinflammatory genes (Nos2, Nlrp3, Il6, and Ptgs2), were decreased in WD PQQ-fed mice, concomitant with increased expression of fatty acid oxidation genes and decreased Pparg expression. Notably, these changes persisted even after PQQ withdrawal at weaning. Our results suggest that supplementation with PQQ, particularly during pregnancy and lactation, protects offspring from WD-induced developmental programming of hepatic lipotoxicity and may help slow the advancing epidemic of NAFLD in the next generation.-Jonscher, K. R., Stewart, M. S., Alfonso-Garcia, A., DeFelice, B. C., Wang, X. X., Luo, Y., Levi, M., Heerwagen, M. J. R., Janssen, R. C., de la Houssaye, B. A., Wiitala, E., Florey, G., Jonscher, R. L., Potma, E. O., Fiehn, O. Friedman, J. E. Early PQQ supplementation has persistent long-term protective effects on developmental programming of hepatic lipotoxicity and inflammation in obese mice.
Assuntos
Antioxidantes/uso terapêutico , Hepatopatia Gordurosa não Alcoólica/prevenção & controle , Obesidade/complicações , Cofator PQQ/uso terapêutico , Efeitos Tardios da Exposição Pré-Natal/prevenção & controle , Animais , Antioxidantes/administração & dosagem , Antioxidantes/farmacologia , Ceramidas/metabolismo , Ciclo-Oxigenase 2/genética , Ciclo-Oxigenase 2/metabolismo , Dieta Hiperlipídica/efeitos adversos , Suplementos Nutricionais , Feminino , Interleucina-6/genética , Interleucina-6/metabolismo , Fígado/efeitos dos fármacos , Fígado/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Óxido Nítrico Sintase Tipo II/genética , Óxido Nítrico Sintase Tipo II/metabolismo , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Hepatopatia Gordurosa não Alcoólica/etiologia , Obesidade/tratamento farmacológico , Obesidade/etiologia , Estresse Oxidativo , PPAR gama/metabolismo , Cofator PQQ/administração & dosagem , Cofator PQQ/farmacologia , Gravidez , Efeitos Tardios da Exposição Pré-Natal/tratamento farmacológico , Efeitos Tardios da Exposição Pré-Natal/etiologiaRESUMO
This study was designed to investigate mechanisms of lipid metabolic inflexibility in human obesity and the ability of fenofibrate (FENO) to increase skeletal muscle fatty acid oxidation (FAO) in primary human skeletal muscle cell cultures (HSkMC) exhibiting metabolic inflexibility. HSkMC from 10 lean and 10 obese, insulin resistant subjects were treated with excess fatty acid for 24 h (24hFA) to gauge lipid-related metabolic flexibility. Metabolically inflexible HSkMC from obese individuals were then treated with 24hFA in combination with FENO to determine effectiveness for increasing FAO. Mitochondrial enzyme activity and FAO were measured in skeletal muscle from subjects with prediabetes (n=11) before and after 10 weeks of fenofibrate in vivo. 24hFA increased FAO to a greater extent in HSkMC from lean versus obese subjects (+49% vs. +9%, for lean vs. obese, respectively; p<0.05) indicating metabolic inflexibility with obesity. Metabolic inflexibility was not observed for measures of cellular respiration in permeabilized cells using carbohydrate substrate. Fenofibrate co-incubation with 24hFA, increased FAO in a subset of HSkMC from metabolically inflexible, obese subjects (p<0.05), which was eliminated by PPARα antagonist. In vivo, fenofibrate treatment increased skeletal muscle FAO in a subset of subjects with prediabetes but did not affect gene transcription or mitochondrial enzyme activity. Lipid metabolic inflexibility observed in HSkMC from obese subjects is not due to differences in electron transport flux, but rather upstream decrements in lipid metabolism. Fenofibrate increases the capacity for FAO in human skeletal muscle cells, though its role in skeletal muscle metabolism in vivo remains unclear.
Assuntos
Ácidos Graxos/metabolismo , Fenofibrato/farmacologia , Metabolismo dos Lipídeos/efeitos dos fármacos , Músculo Esquelético/efeitos dos fármacos , Músculo Esquelético/metabolismo , Obesidade/metabolismo , Adolescente , Adulto , Idoso , Células Cultivadas , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Músculo Esquelético/patologia , Obesidade/patologia , Oxirredução/efeitos dos fármacos , Estado Pré-Diabético/metabolismo , Estado Pré-Diabético/patologia , Adulto JovemRESUMO
BACKGROUND: Non-alcoholic fatty liver disease (NAFLD) is on the rise among youth. Identifying biomarkers of NAFLD progression/risk can aid in prevention efforts. AIMS: This pilot study investigated associations of two endotoxin biomarkers-lipopolysaccharide-binding protein (LBP) and anti-endotoxin core immunoglobulin G (EndoCab)-with markers of NAFLD among 99 Latino/Latina adolescents (11-19 years) with obesity. MATERIALS & METHODS: We used linear regression to examine associations of each endotoxin biomarker (per 1-SD) with hepatic fat fraction (HFF), liver volume, and liver stiffness. RESULTS: We found positive associations of LBP with HFF and liver volume. Each 1-SD increment in LBP corresponded with 2.35% (95% CI: 0.46%, 4.23%) higher HFF and 0.14 (0.06, 0.23) L greater liver volume after adjusting for age, sex, and maternal education. Accounting for abdominal adiposity and Tanner stage did not change results. Excluding 72 participants with NAFLD attenuated associations of LBP with HFF but associations with liver volume persisted (0.11 [0.01, 0.21] L). EndoCab was not associated with any liver outcomes. Neither endotoxin biomarker predicted liver stiffness. DISCUSSION/CONCLUSION: While additional research is warranted, our results support LBP as a biomarker of NAFLD risk/progression in high-risk youth.
Assuntos
Hepatopatia Gordurosa não Alcoólica , Adolescente , Humanos , Hepatopatia Gordurosa não Alcoólica/epidemiologia , Hepatopatia Gordurosa não Alcoólica/metabolismo , Endotoxinas/metabolismo , Projetos Piloto , Fígado/diagnóstico por imagem , Fígado/metabolismo , Biomarcadores/metabolismoRESUMO
Maternal obesity and/or Western diet (WD) is associated with an increased risk of metabolic dysfunction-associated steatotic liver disease (MASLD) in offspring, driven, in part, by the dysregulation of the early life microbiome. Here, using a mouse model of WD-induced maternal obesity, we demonstrate that exposure to a disordered microbiome from WD-fed dams suppressed circulating levels of endogenous ligands of the aryl hydrocarbon receptor (AHR; indole, indole-3-acetate) and TMAO (a product of AHR-mediated transcription), as well as hepatic expression of Il10 (an AHR target), in offspring at 3 weeks of age. This signature was recapitulated by fecal microbial transfer from WD-fed pregnant dams to chow-fed germ-free (GF) lactating dams following parturition and was associated with a reduced abundance of Lactobacillus in GF offspring. Further, the expression of Il10 was downregulated in liver myeloid cells and in LPS-stimulated bone marrow-derived macrophages (BMDM) in adult offspring, suggestive of a hypo-responsive, or tolerant, innate immune response. BMDMs from adult mice lacking AHR in macrophages exhibited a similar tolerogenic response, including diminished expression of Il10. Overall, our study shows that exposure to maternal WD alters microbial metabolites in the offspring that affect AHR signaling, potentially contributing to innate immune hypo-responsiveness and progression of MASLD, highlighting the impact of early life gut dysbiosis on offspring metabolism. Further investigations are warranted to elucidate the complex interplay between maternal diet, gut microbial function, and the development of neonatal innate immune tolerance and potential therapeutic interventions targeting these pathways.
Assuntos
Dieta Ocidental , Microbioma Gastrointestinal , Imunidade Inata , Receptores de Hidrocarboneto Arílico , Triptofano , Animais , Feminino , Gravidez , Dieta Ocidental/efeitos adversos , Triptofano/metabolismo , Camundongos , Receptores de Hidrocarboneto Arílico/metabolismo , Camundongos Endogâmicos C57BL , Interleucina-10/metabolismo , Efeitos Tardios da Exposição Pré-Natal , Obesidade Materna/metabolismo , Fígado/metabolismo , Fenômenos Fisiológicos da Nutrição Materna , Masculino , Macrófagos/metabolismo , Macrófagos/imunologia , Modelos Animais de DoençasRESUMO
Studying fetal hematopoiesis is challenging as hematopoiesis transitions from the liver to bone marrow. Obtaining human samples is not possible, and small animal models may not provide sufficient biological material. Here, we present a protocol for isolating hematopoietic cells from the nonhuman primate fetal liver and bone. We describe steps for using cells from the same fetus for fluorescence lifetime imaging microscopy to measure metabolism, assessing cellular function, and flow cytometry for immunophenotyping at the single-cell level. For complete details on the use and execution of this protocol, please refer to Nash et al. (2023).1.
Assuntos
Leucócitos , Fígado , Animais , Humanos , Imunofenotipagem , Feto , PrimatasRESUMO
Strong evidence exists for a link between chronic low level inflammation and dietary-induced insulin resistance; however, little is known about the transcriptional networks involved. Here we show that high fat diet (HFD) or saturated fatty acid exposure directly activates CCAAT/enhancer-binding protein ß (C/EBPß) protein expression in liver, adipocytes, and macrophages. Global C/EBPß deletion prevented HFD-induced inflammation and surprisingly increased mitochondrial gene expression in white adipose tissue along with brown adipose tissue markers PRDM16, CIDEa, and UCP1, consistent with a resistance to HFD-induced obesity. In isolated peritoneal macrophages from C/EBPß(-/-) mice, the anti-inflammatory gene LXRα and its targets SCD1 and DGAT2 were strikingly up-regulated along with IL-10, while NLRP3, a gene important for activating the inflammasome, was suppressed in response to palmitate. Using RAW 264.7 macrophage cells or 3T3-L1 adipocytes, C/EBPß knockdown prevented palmitate-induced inflammation and p65-NFκB DNA binding activity, while C/EBPß overexpression induced NFκB binding, JNK activation, and pro-inflammatory cytokine gene expression directly. Finally, chimeric bone marrow mice transplanted with bone marrow lacking C/EBPß(-/-) demonstrated reduced systemic and adipose tissue inflammatory markers, macrophage content, and maintained insulin sensitivity on HFD. Taken together, these results demonstrate that HFD or palmitate exposure triggers C/EBPß expression that controls expression of distinct aspects of alternative macrophage activation. Reducing C/EBPß in macrophages confers protection from HFD-induced systemic inflammation and insulin resistance, suggesting it may be an attractive therapeutic target for ameliorating obesity-induced inflammatory responses.
Assuntos
Tecido Adiposo Marrom/metabolismo , Proteína beta Intensificadora de Ligação a CCAAT/biossíntese , Gorduras na Dieta/efeitos adversos , Regulação da Expressão Gênica/efeitos dos fármacos , Ativação de Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Obesidade/metabolismo , Células 3T3-L1 , Tecido Adiposo Marrom/patologia , Animais , Antígenos de Diferenciação/genética , Antígenos de Diferenciação/metabolismo , Proteína beta Intensificadora de Ligação a CCAAT/genética , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Gorduras na Dieta/farmacologia , Ativação Enzimática/efeitos dos fármacos , Ativação Enzimática/genética , Inflamassomos/genética , Inflamassomos/metabolismo , Inflamação/genética , Inflamação/metabolismo , Inflamação/patologia , Interleucina-10/genética , Interleucina-10/metabolismo , MAP Quinase Quinase 4/genética , MAP Quinase Quinase 4/metabolismo , Ativação de Macrófagos/genética , Macrófagos/patologia , Camundongos , Camundongos Knockout , Mitocôndrias/genética , Mitocôndrias/metabolismo , Mitocôndrias/patologia , Proteína 3 que Contém Domínio de Pirina da Família NLR , Obesidade/induzido quimicamente , Obesidade/genética , Obesidade/patologia , Fator de Transcrição RelA/genética , Fator de Transcrição RelA/metabolismoRESUMO
Chronic hepatitis C virus (HCV) infection greatly increases the risk for type 2 diabetes and nonalcoholic steatohepatitis; however, the pathogenic mechanisms remain incompletely understood. Here we report gluconeogenic enzyme phosphoenolpyruvate carboxykinase (PEPCK) transcription and associated transcription factors are dramatically up-regulated in Huh.8 cells, which stably express an HCV subgenome replicon. HCV increased activation of cAMP response element-binding protein (CREB), CCAAT/enhancer-binding protein (C/EBPß), forkhead box protein O1 (FOXO1), and peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α) and involved activation of the cAMP response element in the PEPCK promoter. Infection with dominant-negative CREB or C/EBPß-shRNA significantly reduced or normalized PEPCK expression, with no change in PGC-1α or FOXO1 levels. Notably, expression of HCV nonstructural component NS5A in Huh7 or primary hepatocytes stimulated PEPCK gene expression and glucose output in HepG2 cells, whereas a deletion in NS5A reduced PEPCK expression and lowered cellular lipids but was without effect on insulin resistance, as demonstrated by the inability of insulin to stimulate mobilization of a pool of insulin-responsive vesicles to the plasma membrane. HCV-replicating cells demonstrated increases in cellular lipids with insulin resistance at the level of the insulin receptor, increased insulin receptor substrate 1 (Ser-312), and decreased Akt (Ser-473) activation in response to insulin. C/EBPß-RNAi normalized lipogenic genes sterol regulatory element-binding protein-1c, peroxisome proliferator-activated receptor γ, and liver X receptor α but was unable to reduce accumulation of triglycerides in Huh.8 cells or reverse the increase in ApoB expression, suggesting a role for increased lipid retention in steatotic hepatocytes. Collectively, these data reveal an important role of NS5A, C/EBPß, and pCREB in promoting HCV-induced gluconeogenic gene expression and suggest that increased C/EBPß and NS5A may be essential components leading to increased gluconeogenesis associated with HCV infection.
Assuntos
Proteína beta Intensificadora de Ligação a CCAAT/fisiologia , Fígado Gorduroso/virologia , Genoma Viral , Hepacivirus/genética , Fosfoenolpiruvato Carboxiquinase (GTP)/genética , Proteínas não Estruturais Virais/fisiologia , Animais , Proteína beta Intensificadora de Ligação a CCAAT/genética , Proteína beta Intensificadora de Ligação a CCAAT/metabolismo , Linhagem Celular Tumoral , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Diabetes Mellitus Tipo 2/virologia , Indução Enzimática , Fígado Gorduroso/enzimologia , Proteína Forkhead Box O1 , Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/metabolismo , Genes Reporter , Gluconeogênese/genética , Glucose/metabolismo , Glucose-6-Fosfatase/genética , Glucose-6-Fosfatase/metabolismo , Hepacivirus/fisiologia , Humanos , Insulina/fisiologia , Metabolismo dos Lipídeos/genética , Luciferases/biossíntese , Luciferases/genética , Fosfoenolpiruvato Carboxiquinase (GTP)/metabolismo , Regiões Promotoras Genéticas , Ratos , Vesículas Secretórias/metabolismo , Transdução de Sinais , Replicação ViralRESUMO
Drugs designed specifically to activate liver X receptors (LXRs) have beneficial effects on lowering cholesterol metabolism and inflammation but unfortunately lead to severe hepatic steatosis. The transcription factor CCAAT/enhancer binding protein beta (C/EBPß) is an important regulator of liver gene expression but little is known about its involvement in LXR-based steatosis and cholesterol metabolism. The present study investigated the role of C/EBPß expression in LXR agonist (T0901317)-mediated alteration of hepatic triglyceride (TG) and lipogenesis in mice. C/EBPß deletion in mice prevented LXR agonist-mediated induction of lipogenic gene expression in liver in conjunction with significant reduction of liver TG accumulation. Surprisingly, C/EBPß(-/-) mice showed a major increase in liver mitochondrial electron chain function compared to WT mice. Furthermore, LXR activation in C/EBPß(-/-) mice increased the expression of liver ATP-binding cassette transporter ABCG1, a gene implicated in cholesterol efflux and reducing blood levels of total and LDL-cholesterol. Together, these findings establish a central role for C/EBPß in the LXR-mediated steatosis and mitochondrial function, without impairing the influence of LXR activation on lowering LDL and increasing HDL-cholesterol. Inactivation of C/EBPß might therefore be an important therapeutic strategy to prevent LXR activation-mediated adverse effects on liver TG metabolism without disrupting its beneficial effects on cholesterol metabolism.
Assuntos
Anticolesterolemiantes/efeitos adversos , Proteína beta Intensificadora de Ligação a CCAAT/genética , Fígado Gorduroso/induzido quimicamente , Fígado Gorduroso/genética , Hidrocarbonetos Fluorados/efeitos adversos , Mitocôndrias Hepáticas/metabolismo , Receptores Nucleares Órfãos/agonistas , Sulfonamidas/efeitos adversos , Animais , Anticolesterolemiantes/administração & dosagem , HDL-Colesterol/metabolismo , Deleção de Genes , Hidrocarbonetos Fluorados/administração & dosagem , Lipogênese/efeitos dos fármacos , Receptores X do Fígado , Masculino , Camundongos , Camundongos Mutantes , Sulfonamidas/administração & dosagem , Triglicerídeos/metabolismoRESUMO
It is generally assumed that white adipocytes arise from resident adipose tissue mesenchymal progenitor cells. We challenge this paradigm by defining a hematopoietic origin for both the de novo development of a subset of white adipocytes in adults and a previously uncharacterized adipose tissue resident mesenchymal progenitor population. Lineage and cytogenetic analysis revealed that bone marrow progenitor (BMP)-derived adipocytes and adipocyte progenitors arise from hematopoietic cells via the myeloid lineage in the absence of cell fusion. Global gene expression analysis indicated that the BMP-derived fat cells are bona fide adipocytes but differ from conventional white or brown adipocytes in decreased expression of genes involved in mitochondrial biogenesis and lipid oxidation, and increased inflammatory gene expression. The BMP-derived adipocytes accumulate with age, occur in higher numbers in visceral than in subcutaneous fat, and in female versus male mice. BMP-derived adipocytes may, therefore, account in part for adipose depot heterogeneity and detrimental changes in adipose metabolism and inflammation with aging and adiposity.
Assuntos
Adipócitos Brancos/citologia , Tecido Adiposo/citologia , Mesoderma/citologia , Células Mieloides/citologia , Adipócitos Marrons/citologia , Adipócitos Marrons/metabolismo , Adipócitos Brancos/metabolismo , Tecido Adiposo/metabolismo , Fatores Etários , Animais , Células da Medula Óssea/citologia , Células da Medula Óssea/metabolismo , Diferenciação Celular , Linhagem da Célula , Células Cultivadas , Análise Citogenética , Feminino , Perfilação da Expressão Gênica , Masculino , Mesoderma/metabolismo , Camundongos , Modelos Biológicos , Células Mieloides/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos , Fatores SexuaisRESUMO
Pediatric NAFLD has distinct and variable pathology, yet causation remains unclear. We have shown that maternal Western-style diet (mWSD) compared with maternal chow diet (CD) consumption in nonhuman primates produces hepatic injury and steatosis in fetal offspring. Here, we define the role of mWSD and postweaning Western-style diet (pwWSD) exposures on molecular mechanisms linked to NAFLD development in a cohort of 3-year-old juvenile nonhuman primates offspring exposed to maternal CD or mWSD followed by CD or Western-style diet after weaning. We used histologic, transcriptomic, and metabolomic analyses to identify hepatic pathways regulating NAFLD. Offspring exposed to mWSD showed increased hepatic periportal collagen deposition but unchanged hepatic triglyceride levels and body weight. mWSD was associated with a downregulation of gene expression pathways underlying HNF4α activity and protein, and downregulation of antioxidant signaling, mitochondrial biogenesis, and PPAR signaling pathways. In offspring exposed to both mWSD and pwWSD, liver RNA profiles showed upregulation of pathways promoting fibrosis and endoplasmic reticulum stress and increased BiP protein expression with pwWSD. pwWSD increased acylcarnitines and decreased anti-inflammatory fatty acids, which was more pronounced when coupled with mWSD exposure. Further, mWSD shifted liver metabolites towards decreased purine catabolism in favor of synthesis, suggesting a mitochondrial DNA repair response. Our findings demonstrate that 3-year-old offspring exposed to mWSD but weaned to a CD have periportal collagen deposition, with transcriptional and metabolic pathways underlying hepatic oxidative stress, compromised mitochondrial lipid sensing, and decreased antioxidant response. Exposure to pwWSD worsens these phenotypes, triggers endoplasmic reticulum stress, and increases fibrosis. Overall, mWSD exposure is associated with altered expression of candidate genes and metabolites related to NAFLD that persist in juvenile offspring preceding clinical presentation of NAFLD.
Assuntos
Hepatopatia Gordurosa não Alcoólica , Animais , Hepatopatia Gordurosa não Alcoólica/etiologia , Dieta Ocidental , Antioxidantes , Fibrose , Fenótipo , PrimatasRESUMO
Maternal overnutrition increases inflammatory and metabolic disease risk in postnatal offspring. This constitutes a major public health concern due to increasing prevalence of these diseases, yet mechanisms remain unclear. Here, using nonhuman primate models, we show that maternal Western-style diet (mWSD) exposure is associated with persistent pro-inflammatory phenotypes at the transcriptional, metabolic, and functional levels in bone marrow-derived macrophages (BMDMs) from 3-year-old juvenile offspring and in hematopoietic stem and progenitor cells (HSPCs) from fetal and juvenile bone marrow and fetal liver. mWSD exposure is also associated with increased oleic acid in fetal and juvenile bone marrow and fetal liver. Assay for transposase-accessible chromatin with sequencing (ATAC-seq) profiling of HSPCs and BMDMs from mWSD-exposed juveniles supports a model in which HSPCs transmit pro-inflammatory memory to myeloid cells beginning in utero. These findings show that maternal diet alters long-term immune cell developmental programming in HSPCs with proposed consequences for chronic diseases featuring altered immune/inflammatory activation across the lifespan.
Assuntos
Medula Óssea , Células-Tronco Hematopoéticas , Humanos , Animais , Feminino , Dieta Ocidental/efeitos adversos , Primatas , Imunidade InataRESUMO
The conversion of pyruvate to acetyl-CoA in mitochondria is catalyzed by the pyruvate dehydrogenase complex (PDC). Activity of PDC is inhibited by phosphorylation via the pyruvate dehydrogenase kinases (PDKs). Here, we examined the regulation of Pdk4 gene expression by the CCAAT/enhancer-binding protein ß (C/EBPß). C/EBPß modulates the expression of multiple hepatic genes including those involved in metabolism, development, and inflammation. We found that C/EBPß induced Pdk4 gene expression and decreased PDC activity. This transcriptional induction was mediated through two C/EBPß binding sites in the Pdk4 promoter. C/EBPß participates in the hormonal regulation of gluconeogenic genes. Previously, we reported that Pdk4 was induced by thyroid hormone (T(3)). Therefore, we investigated the role of C/EBPß in the T(3) regulation of Pdk4. T(3) increased C/EBPß abundance in primary rat hepatocytes. Knockdown of C/EBPß with siRNA diminished the T(3) induction of the Pdk4 and carnitine palmitoyltransferase (Cpt1a) genes. CPT1a is an initiating step in the mitochondrial oxidation of long chain fatty acids. Our results indicate that C/EBPß stimulates Pdk4 expression and participates in the T(3) induction of the Cpt1a and Pdk4 genes.
Assuntos
Proteína beta Intensificadora de Ligação a CCAAT/metabolismo , Regulação Enzimológica da Expressão Gênica/fisiologia , Hepatócitos/metabolismo , Proteínas Serina-Treonina Quinases/biossíntese , Elementos de Resposta/fisiologia , Animais , Proteína beta Intensificadora de Ligação a CCAAT/genética , Carnitina O-Palmitoiltransferase/biossíntese , Carnitina O-Palmitoiltransferase/genética , Gluconeogênese/fisiologia , Células Hep G2 , Hepatócitos/citologia , Humanos , Proteínas Serina-Treonina Quinases/genética , Piruvato Desidrogenase Quinase de Transferência de Acetil , Complexo Piruvato Desidrogenase/biossíntese , Complexo Piruvato Desidrogenase/genética , Ratos , Tri-Iodotironina/genética , Tri-Iodotironina/metabolismoRESUMO
CONTEXT: Metabolic endotoxemia may be a shared mechanism underlying childhood obesity and early-onset metabolic diseases (eg, type 2 diabetes, nonalcoholic fatty liver disease). OBJECTIVE: Examine prospective associations of serum endotoxin biomarkers lipopolysaccharide (LPS) and its binding protein, LPS binding protein (LBP), and anti-endotoxin core immunoglobulin G (EndoCab IgG) with adiposity and cardiometabolic risk in youth. DESIGN/SETTING: This prospective study included 393 youth in the Exploring Perinatal Outcomes Among Children cohort in Colorado. Participants were recruited from 2006 to 2009 at age 10 years (baseline) and followed for 6 years (follow-up). We examined associations of endotoxin biomarkers at baseline with adiposity [body mass index (BMI) z-score, visceral adipose tissue (VAT), subcutaneous adipose tissue (SAT), skinfolds, waist circumference] and cardiometabolic risk (insulin, glucose, adipokines, lipid profile, blood pressure) across both visits using mixed-effects regression, and with hepatic fat fraction (HFF) at follow-up using linear regression. RESULTS: Higher LPS and LBP predicted greater adiposity across follow-up. Each 1-unit log-transformed LPS corresponded with 0.23 (95% CI 0.03, 0.43) units BMI z-score, 5.66 (95% CI 1.99, 9.33) mm3 VAT, 30.7 (95% CI 8.0, 53.3) mm3 SAT, and 8.26 (95% CI 4.13, 12.40) mm skinfold sum. EndoCab IgG was associated with VAT only [3.03 (95% CI 0.34, 5.71) mm3]. LPS was associated with higher insulin [1.93 (95% CI 0.08, 3.70) µU/mL] and leptin [2.28 (95% CI 0.66, 3.90) ng/mL] and an adverse lipid profile. No association was observed with HFF. Accounting for pubertal status and lifestyle behaviors did not change findings. However, adjustment for prepregnancy BMI and gestational diabetes attenuated most associations. CONCLUSIONS: Serum endotoxin may be a marker of pathophysiological processes underlying development of childhood obesity and cardiometabolic conditions associated with exposure to fetal overnutrition.
Assuntos
Doenças Cardiovasculares , Diabetes Mellitus Tipo 2 , Insulinas , Obesidade Infantil , Adiposidade , Adolescente , Biomarcadores , Índice de Massa Corporal , Doenças Cardiovasculares/epidemiologia , Doenças Cardiovasculares/etiologia , Criança , Endotoxinas , Feminino , Seguimentos , Humanos , Imunoglobulina G , Lipopolissacarídeos , Obesidade Infantil/epidemiologia , Gravidez , Estudos Prospectivos , Fatores de RiscoRESUMO
Maternal consumption of a high-fat, Western-style diet (WD) disrupts the maternal/infant microbiome and contributes to developmental programming of the immune system and nonalcoholic fatty liver disease (NAFLD) in the offspring. Epigenetic changes, including non-coding miRNAs in the fetus and/or placenta may also underlie this risk. We previously showed that obese nonhuman primates fed a WD during pregnancy results in the loss of beneficial maternal gut microbes and dysregulation of cellular metabolism and mitochondrial dysfunction in the fetal liver, leading to a perturbed postnatal immune response with accelerated NAFLD in juvenile offspring. Here, we investigated associations between WD-induced maternal metabolic and microbiome changes, in the absence of obesity, and miRNA and gene expression changes in the placenta and fetal liver. After ~8-11 months of WD feeding, dams were similar in body weight but exhibited mild, systemic inflammation (elevated CRP and neutrophil count) and dyslipidemia (increased triglycerides and cholesterol) compared with dams fed a control diet. The maternal gut microbiome was mainly comprised of Lactobacillales and Clostridiales, with significantly decreased alpha diversity (P = 0.0163) in WD-fed dams but no community-wide differences (P = 0.26). At 0.9 gestation, mRNA expression of IL6 and TNF in maternal WD (mWD) exposed placentas trended higher, while increased triglycerides, expression of pro-inflammatory CCR2, and histological evidence for fibrosis were found in mWD-exposed fetal livers. In the mWD-exposed fetus, hepatic expression levels of miR-204-5p and miR-145-3p were significantly downregulated, whereas in mWD-exposed placentas, miR-182-5p and miR-183-5p were significantly decreased. Notably, miR-1285-3p expression in the liver and miR-183-5p in the placenta were significantly associated with inflammation and lipid synthesis pathway genes, respectively. Blautia and Ruminococcus were significantly associated with miR-122-5p in liver, while Coriobacteriaceae and Prevotellaceae were strongly associated with miR-1285-3p in the placenta; both miRNAs are implicated in pathways mediating postnatal growth and obesity. Our findings demonstrate that mWD shifts the maternal microbiome, lipid metabolism, and inflammation prior to obesity and are associated with epigenetic changes in the placenta and fetal liver. These changes may underlie inflammation, oxidative stress, and fibrosis patterns that drive NAFLD and metabolic disease risk in the next generation.
RESUMO
Maternal obesity affects nearly one-third of pregnancies and is a major risk factor for nonalcoholic fatty liver disease (NAFLD) in adolescent offspring, yet the mechanisms behind NAFLD remain poorly understood. Here, we demonstrate that nonhuman primate fetuses exposed to maternal Western-style diet (WSD) displayed increased fibrillar collagen deposition in the liver periportal region, with increased ACTA2 and TIMP1 staining, indicating localized hepatic stellate cell (HSC) and myofibroblast activation. This collagen deposition pattern persisted in 1-year-old offspring, despite weaning to a control diet (CD). Maternal WSD exposure increased the frequency of DCs and reduced memory CD4+ T cells in fetal liver without affecting systemic or hepatic inflammatory cytokines. Switching obese dams from WSD to CD before conception or supplementation of the WSD with resveratrol decreased fetal hepatic collagen deposition and reduced markers of portal triad fibrosis, oxidative stress, and fetal hypoxemia. These results demonstrate that HSCs and myofibroblasts are sensitive to maternal WSD-associated oxidative stress in the fetal liver, which is accompanied by increased periportal collagen deposition, indicative of early fibrogenesis beginning in utero. Alleviating maternal WSD-driven oxidative stress in the fetal liver holds promise for halting steatosis and fibrosis and preventing developmental programming of NAFLD.
Assuntos
Dieta Ocidental/efeitos adversos , Cirrose Hepática/fisiopatologia , Fenômenos Fisiológicos da Nutrição Materna/fisiologia , Animais , Feminino , Exposição Materna , Gravidez , Primatas , ÚteroRESUMO
Non-alcoholic fatty liver disease (NAFLD) is the number one chronic liver disease worldwide and is estimated to affect nearly 40% of obese youth and up to 10% of the general pediatric population without any obvious signs or symptoms. Although the early stages of NAFLD are reversible with diet and lifestyle modifications, detecting such stages is hindered by a lack of non-invasive methods of risk assessment and diagnosis. This absence of non-invasive means of diagnosis is directly related to the scarcity of long-term prospective studies of pediatric NAFLD in children and adolescents. In the majority of pediatric NAFLD cases, the mechanisms driving the origin and rapid progression of NAFLD remain unknown. The progression from NAFLD to non-alcoholic steatohepatitis (NASH) in youth is associated with unique histological features and possible immune processes and metabolic pathways that may reflect different mechanisms compared with adults. Recent data suggest that circulating microRNAs (miRNAs) are important new biomarkers underlying pathways of liver injury. Several factors may contribute to pediatric NAFLD development, including high-sugar diets, in utero exposures via epigenetic alterations, changes in the neonatal microbiome, and altered immune system development and mitochondrial function. This review focuses on the unique aspects of pediatric NAFLD and how nutritional exposures impact the immune system, mitochondria, and liver/gastrointestinal metabolic health. These factors highlight the need for answers to how NAFLD develops in children and for early stage-specific interventions.
Assuntos
Fenômenos Fisiológicos da Nutrição Infantil/fisiologia , Hepatopatia Gordurosa não Alcoólica/diagnóstico , Hepatopatia Gordurosa não Alcoólica/etiologia , Adolescente , Biomarcadores/sangue , Criança , Progressão da Doença , Feminino , Humanos , Sistema Imunitário , Fígado/imunologia , Fígado/metabolismo , Masculino , MicroRNAs/sangue , Mitocôndrias , Hepatopatia Gordurosa não Alcoólica/prevenção & controle , Hepatopatia Gordurosa não Alcoólica/terapia , Medição de RiscoRESUMO
Overweight and obesity accompanies up to 70% of pregnancies and is a strong risk factor for offspring metabolic disease. Maternal obesity-associated inflammation and lipid profile are hypothesized as important contributors to excess offspring liver and skeletal muscle lipid deposition and oxidative stress. Here, we tested whether dams expressing the fat-1 transgene, which endogenously converts omega-6 (n-6) to omega-3 (n-3) polyunsaturated fatty acid, could protect wild-type (WT) offspring against high-fat diet induced weight gain, oxidative stress, and disrupted mitochondrial fatty acid oxidation. Despite similar body mass at weaning, offspring from fat-1 high-fat-fed dams gained less weight compared with offspring from WT high-fat-fed dams. In particular, WT males from fat-1 high-fat-fed dams were protected from post-weaning high-fat diet induced weight gain, reduced fatty acid oxidation, or excess oxidative stress compared with offspring of WT high-fat-fed dams. Adult offspring of WT high-fat-fed dams exhibited greater skeletal muscle triglycerides and reduced skeletal muscle antioxidant defense and redox balance compared with offspring of WT dams on control diet. Fat-1 offspring were protected from the reduced fatty acid oxidation and excess oxidative stress observed in offspring of WT high-fat-fed dams. These results indicate that a maternal fat-1 transgene has protective effects against offspring liver and skeletal muscle lipotoxicity resulting from a maternal high-fat diet, particularly in males. Altering maternal fatty acid composition, without changing maternal dietary composition or weight gain with high-fat feeding, may highlight important strategies for n-3-based prevention of developmental programming of obesity and its complications.
Assuntos
Proteínas de Caenorhabditis elegans , Dieta Hiperlipídica/efeitos adversos , Ácidos Graxos Dessaturases , Exposição Materna , Obesidade , Estresse Oxidativo/genética , Transgenes , Animais , Caenorhabditis elegans/enzimologia , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Ácidos Graxos Dessaturases/biossíntese , Ácidos Graxos Dessaturases/genética , Feminino , Masculino , Camundongos , Camundongos Transgênicos , Obesidade/genética , Obesidade/metabolismo , Obesidade/patologia , Obesidade/prevenção & controle , GravidezRESUMO
Gestational diabetes mellitus (GDM) is a worldwide public health problem affecting up to 27% of pregnancies with high predictive values for childhood obesity and inflammatory diseases. Compromised seeding of the infant gut microbiota is a risk factor for immunologic and metabolic diseases in the offspring; however, how GDM along with maternal obesity interact to alter colonization remains unknown. We hypothesized that GDM individually and in combination with maternal overweight/obesity would alter gut microbial composition, diversity, and short-chain fatty acid (SCFA) levels in neonates. We investigated 46 full-term neonates born to normal-weight or overweight/obese mothers with and without GDM, accounting for confounders including cesarean delivery, lack of breastfeeding, and exposure to antibiotics. Gut microbiota in 2-week-old neonates born to mothers with GDM exhibited differences in abundance of 26 microbial taxa; 14 of which showed persistent differential abundance after adjusting for pre-pregnancy BMI. Key pioneering gut taxa, including potentially important taxa for establishing neonatal immunity, were reduced. Lactobacillus, Flavonifractor, Erysipelotrichaceae, and unspecified families in Gammaproteobacteria were significantly reduced in neonates from mothers with GDM. GDM was associated with an increase in microbes involved in suppressing early immune cell function (Phascolarctobacterium). No differences in infant stool SCFA levels by maternal phenotype were noted; however, significant correlations were found between microbial abundances and SCFA levels in neonates. Our results suggest that GDM alone and together with maternal overweight/obesity uniquely influences seeding of specific infant microbiota in patterns that set the stage for future risk of inflammatory and metabolic disease.