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1.
Mutat Res ; 850-851: 503161, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32247561

RESUMO

Through diet, people are chronically exposed to low doses of a large number of contaminants that could exhibit adverse health effects. Toxicological evaluation of food contaminants increases in complexity when the exposure involves chemical mixtures. The aim of this study is to investigate the genotoxic potential, through measuring ©H2AX induction, of six common mixtures of food contaminants to which French adult consumers are chronically exposed. Mixtures were identified by combining information from consumption surveys and contaminant concentration levels in foods. Both single and repeated exposures were evaluated in human liver-derived HepaRG cells. Our results indicated that after a single 24-h exposure, only one mixture induced genotoxicity, and that response occurred at the highest concentration tested. In contrast, we observed after repeated exposures over 3 or 7 days, induction of ©H2AX for all mixtures except one, and a time- and concentration-dependent manner toxicity for four mixtures. Interestingly, we also observed a non-monotonic cytotoxicity concentration-response for one mixture, which might reflect cellular adaptation to the exposure. In conclusion, our study demonstrated that longer-term treatments for in vitro toxicological evaluation, instead of the classical 24 h treatment, may be more relevant regarding human toxicology assessment.


Assuntos
Dano ao DNA/efeitos dos fármacos , Dieta/efeitos adversos , Contaminação de Alimentos/análise , Histonas/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Hepatócitos/efeitos dos fármacos , Humanos , Fígado/efeitos dos fármacos , Testes de Mutagenicidade
2.
Int J Mol Sci ; 21(7)2020 Apr 08.
Artigo em Inglês | MEDLINE | ID: mdl-32276399

RESUMO

When interferons (IFNs) bind to their receptors, they upregulate numerous IFN-stimulated genes (ISGs) with antiviral and immune regulatory activities. Hepatitis C virus (HCV) is a single-stranded, positive-sense RNA virus that affects over 71 million people in the global population. Hepatocytes infected with HCV produce types I and III IFNs. These endogenous IFNs upregulate a set of ISGs that negatively impact the outcome of pegylated IFN-α and ribavirin treatments, which were previously used to treat HCV. In addition, the IFNL4 genotype was the primary polymorphism responsible for a suboptimal treatment response to pegylated IFN-α and ribavirin. However, recently developed direct-acting antivirals have demonstrated a high rate of sustained virological response without pegylated IFN-α. Herein, we review recent studies on types I and III IFN responses to in HCV-infected hepatocytes. In particular, we focused on open issues related to IFN responses in the direct-acting antiviral era.


Assuntos
Antivirais/farmacologia , Hepatite C/tratamento farmacológico , Hepatócitos/metabolismo , Imunidade Inata , Interferons/genética , Antivirais/uso terapêutico , Regulação da Expressão Gênica , Hepatite C/genética , Hepatite C/imunologia , Hepatócitos/imunologia , Humanos , Interferon Tipo I/genética
3.
Mutat Res ; 852: 503169, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32265043

RESUMO

The phycotoxins, okadaic acid (OA) and dinophysistoxins 1 and 2 (DTX-1 and -2), are protein phosphatase PP2A and PP1 inhibitors involved in diarrhetic shellfish poisoning (DSP) in humans. Data on the in vivo acute toxicity of the OA-group toxins show some differences and the European Food Safety Authority (EFSA) has determined toxicity equivalent factors (TEFs) of one for the reference toxin, OA, as well as for DTX-1 and 0.6 for DTX-2. However, recent in vitro studies indicated that DTX-1 seems to be more toxic than OA. As OA was described as apoptotic and aneugenic compound, we analyzed the DNA damage responses induced by the 3 toxins through γH2AX and pH3 biomarkers on proliferative HepaRG cells using High Content Analysis. We quantitatively examined the responses for γH2AX and pH3 by benchmark dose analyzing (BMD) using PROAST software. We found that the three toxins increased both γH2AX- and pH3-positive cells populations in a concentration-dependent manner. The 3 toxins induced mitotic arrest, characteristic of aneugenic compounds, as well as DNA strand-breaks concomitantly to cytotoxicity. BMD analysis showed that DTX-1 is the most potent inducer of DNA damage, followed by OA and DTX-2. The quantitative genotoxic data provided in this study are additional findings for reconsidering the estimated TEFs of this group of phycotoxins.


Assuntos
Inibidores Enzimáticos/toxicidade , Histonas/genética , Mutagênicos/toxicidade , Ácido Okadáico/toxicidade , Piranos/toxicidade , Benchmarking , Biomarcadores/metabolismo , Linhagem Celular Transformada , Proliferação de Células/efeitos dos fármacos , Dano ao DNA , Relação Dose-Resposta a Droga , Hepatócitos/citologia , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Histonas/metabolismo , Humanos , Interações Hidrofóbicas e Hidrofílicas , Mitose/efeitos dos fármacos , Testes de Mutagenicidade , Fosforilação/efeitos dos fármacos , Software
4.
Life Sci ; 250: 117599, 2020 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-32234491

RESUMO

Chemotherapeutic antibiotic doxorubicin belongs to the anthracycline class, slaughters not only the cancer cells but also non-cancerous cells even in the non-targeted organs thereby resulting in the toxicity. The liver is primarily involved in the process of detoxification and this mini-review we focused mainly to investigate the molecular mechanisms heading hepatotoxicity caused due to doxorubicin administration. The alterations in the doxorubicin treated liver tissue include vacuolation of hepatocytes, degeneration of hepatocyte cords, bile duct hyperplasia and focal necrosis. About the literature conducted, hepatotoxicity caused by doxorubicin has been explained by estimating the levels of liver serum biomarkers, ROS production, antioxidant enzymes, lipid peroxidation, and mitochondrial dysfunction. The liver serum biomarkers such as ALT and AST, elated levels of free radicals inducing oxidative stress characterized by a surge in Nrf-2, FOXO-1 and HO-1 genes and diminution of anti-oxidant activity characterized by a decline in SOD, GPx, and CAT genes. The augmented levels of SGOT, SGPT, LDH, creatine kinase, direct and total bilirubin levels also reveal the toxicity in the hepatic tissue due to doxorubicin treatment. The molecular insight of hepatotoxicity is mainly due to the production of ROS, ameliorated oxidative stress and inflammation, deteriorated mitochondrial production and functioning, and enhanced apoptosis. Certain substances such as extracts from medicinal plants, natural products, and chemical substances have been shown to produce an alleviating effect against the doxorubicin-induced hepatotoxicity are also discussed.


Assuntos
Doença Hepática Induzida por Substâncias e Drogas/fisiopatologia , Doxorrubicina/efeitos adversos , Fígado/efeitos dos fármacos , Trifosfato de Adenosina/metabolismo , Animais , Antioxidantes/metabolismo , Apoptose , Fragmentação do DNA , Células Hep G2 , Hepatócitos/efeitos dos fármacos , Humanos , Inflamação , Peroxidação de Lipídeos/efeitos dos fármacos , Malondialdeído/metabolismo , Mitocôndrias/metabolismo , Estresse Oxidativo , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Extratos Vegetais/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais
5.
Chin J Nat Med ; 18(3): 186-195, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-32245588

RESUMO

Alcoholic liver disease (ALD) has become one of the leading causes of death in the world. Berbamine (BM), a natural product mainly derived from Berberis vulgaris L, possesses multiple bioactivities as a traditional medicine. However, the protective effect of BM on ALD remains unknown. In this study, we investigated the effect of BM on ethanol-induced hepatic injury in mice and its underlying mechanism. It was shown that BM at 0.3125-40 µmol·L-1had no effect on macrophages and hepatocytes proliferation. BM at 5-20 µmol·L-1 significantly inhibited lipopolysaccharide (LPS) or acetate-induced IL-1ß and IL-6 mRNA expression in RAW264.7 cells. Moreover, BM treatment significantly inhibited LPS-induced p65 and STAT3 phosphorylation in RAW264.7 cells. Hepatic histopathology analysis showed that inflammatory cells infiltration and lipid accumulation were suppressed by 25 and 50 mg·kg-1 BM administration in ethanol-induced hepatic injury mouse model. Meanwhile, BM treatment significantly inhibited serum ALT and AST levels in ethanol-fed mice. Oil red O staining results showed that BM administration ameliorated hepatic lipid accumulation in ethanol-fed mice. Preventions of ethanol-induced hepatic injury by BM were reflected by markedly decreased serum and hepatic triglyceride (TG) and total cholesterol (TC) contents. Real-time PCR results showed that BM treatment significantly inhibited pro-inflammatory cytokines mRNA expression in ethanol-fed mouse liver. Remarkably, the mechanism of action of BM was related to the reduction of ethanol-induced NF-κB and STAT3 phosphorylation levels in liver. In addition, BM treatment significantly inhibited ERK phosphorylation but not JNK and p38 of MAPK pathway. Taken together, our results demonstrate a beneficial effect of BM on ethanol-induced liver injury via a mechanism associated with inactivation of NF-κB, STAT3 and ERK pathway, which gives insight into the further evaluation of the therapeutic potential of BM for ALD.


Assuntos
Benzilisoquinolinas/farmacologia , Doença Hepática Crônica Induzida por Substâncias e Drogas/tratamento farmacológico , Inflamação/tratamento farmacológico , Fígado/efeitos dos fármacos , Animais , Colesterol/sangue , Citocinas/metabolismo , Etanol/efeitos adversos , Feminino , Hepatócitos/efeitos dos fármacos , Metabolismo dos Lipídeos , Sistema de Sinalização das MAP Quinases , Camundongos , Camundongos Endogâmicos C57BL , NF-kappa B/metabolismo , Células RAW 264.7 , Fator de Transcrição STAT3/metabolismo , Triglicerídeos/sangue
6.
Pestic Biochem Physiol ; 164: 183-190, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-32284125

RESUMO

Imidacloprid (IMD) is a neonicotinoid insecticide widely used in crops, pets, and on farm animals for pest control, which can cause hepatotoxicity in animals and humans. In a previous study using isolated rat liver mitochondria, we observed that IMD inhibited the activity of FoF1-ATP synthase. The aim of this study was to evaluate the effects of IMD on rat isolated hepatocytes and perfused rat liver, besides the influence of its biotransformation on the toxicological potential. For the latter goal, rats were pretreated with dexamethasone or phenobarbital, two classical cytochrome P-450 stimulators, before hepatocytes isolation or liver perfusion. IMD (150 and 200 µM) reduced state 3 mitochondrial respiration in digitonin-permeabilized cells that were energized with glutamate plus malate but did not dissipate the mitochondrial membrane potential. In intact (non-permeabilized) hepatocytes, the intracellular ATP concentration and cell viability were reduced when high IMD concentrations were used (1.5-3.0 mM), and only in cells isolated from dexamethasone-pretreated rats, revealing that IMD biotransformation increases its toxicity and that IMD itself affects isolated mitochondria or mitochondria in permeabilized hepatocytes in concentrations that do not affect mitochondrial function in intact hepatocytes. Coherently, in the prefused liver, IMD (150 and 250 µM) inhibited gluconeogenesis from alanine, but without affecting oxygen consumption and urea production, indicating that such effect was not of mitochondrial origin. The gluconeogenesis inhibition was incomplete and occurred only when the rats were pretreated with phenobarbital, signs that IMD biotransformation was involved in the observed effect. Our findings reveal that changes in hepatic energy metabolism may be acutely implicated in the hepatotoxicity of IMD only when animals and humans are exposed to high levels of this compound, and that IMD metabolites seem to be the main cause for its toxicity.


Assuntos
Hepatócitos , Fígado , Animais , Biotransformação , Neonicotinoides , Nitrocompostos , Ratos
7.
Chemosphere ; 249: 126420, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32208215

RESUMO

Since the DeepWater Horizon oil spill and the use at 1450 m depth of dispersant as a technical response, the need of relevant ecotoxicological data on deep-sea ecosystems becomes crucial. In this context, this study focused on the effect of high hydrostatic pressure (10.1 MPa) on turbot hepatocytes isolated from fish exposed either to chemically dispersed oil, mechanically dispersed oil or dispersant alone. Potential combined effects of oil/dispersant and hydrostatic pressure, were assessed on cell mortality (total cell death, necrosis and apoptosis), cell viability and on hepatocyte oxygen consumption (MO2). No change in cell mortality was observed in any of the experimental conditions, whereas, the results of cell viability showed a strong and significant increase in the two oil groups independently of the pressure exposure. Finally, oil exposure and hydrostatic pressure have additive effects on oxygen consumption at a cellular level. Presence of dispersant prevent any MO2 increase in our experimental conditions. These mechanistic effects leading to this increased energetic demand and its eventual inhibition by dispersant must be investigated in further experiments.


Assuntos
Linguados/fisiologia , Poluição por Petróleo , Petróleo/toxicidade , Poluentes Químicos da Água/toxicidade , Animais , Ecossistema , Ecotoxicologia , Hepatócitos , Pressão Hidrostática , Alimentos Marinhos , Poluentes Químicos da Água/análise
8.
Life Sci ; 250: 117561, 2020 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-32198052

RESUMO

AIMS: Pyruvate kinase M2 (PKM2), a unique isoform of the pyruvate kinases, not only acts as a crucial metabolic enzyme when it locates in the cytoplasm, but also plays important roles in tumor formation and growth when it accumulates in the nuclei. Our aim was to investigate the potential role of PKM2 in liver regeneration in mice insulted with carbon tetrachloride (CCl4). MATERIAL AND METHODS: The liver regeneration model was established by intraperitoneal injection of CCl4 for 48 h in male BALB/c mice. The expression of PKM2, phospho-STAT3, STAT3, proliferating cell nuclear antigen (PCNA) and Cyclin D1 were evaluated by western blot. The distribution of PKM2 was verified by immunofluorescence staining. The degree of injured region was assessed by hematoxylin and eosin (HE) staining. The proliferation of liver cells was tested by Immunohistochemistry. KEY FINDINGS: The nuclear accumulation of PKM2 increased in the liver treated with CCl4, but treatment with ML-265 significantly suppressed CCl4-induced nuclear accumulation of PKM2. In addition, treatment with ML-265 suppressed the level of cyclin D1 and proliferating cell nuclear antigen (PCNA), reduced the count of Ki67-positive hepatocytes, and expanded the damaged region in histological examination. Meanwhile, treatment with ML-265 suppressed the phosphorylation of nuclear signal transducer and activator of transcription 3 (STAT3). Inhibition of STAT3 by stattic made the same effects as ML-265. SIGNIFICANCE: These data uncovered the role of nuclear PKM2 in liver regeneration and the pro-proliferation effects of nuclear PKM2 may be through targeting its downstream transcription factor STAT3.


Assuntos
Núcleo Celular/metabolismo , Regeneração Hepática , Piruvato Quinase/metabolismo , Fator de Transcrição STAT3/metabolismo , Animais , Tetracloreto de Carbono , Proliferação de Células , Citoplasma/metabolismo , Hepatócitos/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Fígado/metabolismo , Hepatopatias/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Fosforilação/efeitos dos fármacos , Antígeno Nuclear de Célula em Proliferação/metabolismo
9.
Nat Cell Biol ; 22(3): 321-331, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32123335

RESUMO

CRISPR-Cas9 technology has revolutionized genome editing and is applicable to the organoid field. However, precise integration of exogenous DNA sequences into human organoids is lacking robust knock-in approaches. Here, we describe CRISPR-Cas9-mediated homology-independent organoid transgenesis (CRISPR-HOT), which enables efficient generation of knock-in human organoids representing different tissues. CRISPR-HOT avoids extensive cloning and outperforms homology directed repair (HDR) in achieving precise integration of exogenous DNA sequences into desired loci, without the necessity to inactivate TP53 in untransformed cells, which was previously used to increase HDR-mediated knock-in. CRISPR-HOT was used to fluorescently tag and visualize subcellular structural molecules and to generate reporter lines for rare intestinal cell types. A double reporter-in which the mitotic spindle was labelled by endogenously tagged tubulin and the cell membrane by endogenously tagged E-cadherin-uncovered modes of human hepatocyte division. Combining tubulin tagging with TP53 knock-out revealed that TP53 is involved in controlling hepatocyte ploidy and mitotic spindle fidelity. CRISPR-HOT simplifies genome editing in human organoids.


Assuntos
Sistemas CRISPR-Cas , Edição de Genes , Técnicas de Introdução de Genes/métodos , Organoides/citologia , Hepatócitos/citologia , Hepatócitos/ultraestrutura , Humanos , Intestinos/citologia , Fígado/citologia , Organoides/ultraestrutura , Fuso Acromático/ultraestrutura , Proteína Supressora de Tumor p53/fisiologia
10.
Emerg Microbes Infect ; 9(1): 651-663, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32192415

RESUMO

Equine parvovirus-hepatitis (EqPV-H) has recently been associated with cases of Theiler's disease, a form of fulminant hepatic necrosis in horses. To assess whether EqPV-H is the cause of Theiler's disease, we first demonstrated hepatotropism by PCR on tissues from acutely infected horses. We then experimentally inoculated horses with EqPV-H and 8 of 10 horses developed hepatitis. One horse showed clinical signs of liver failure. The onset of hepatitis was temporally associated with seroconversion and a decline in viremia. Liver histology and in situ hybridization showed lymphocytic infiltrates and necrotic EqPV-H-infected hepatocytes. We next investigated potential modes of transmission. Iatrogenic transmission via allogeneic stem cell therapy for orthopedic injuries was previously suggested in a case series of Theiler's disease, and was demonstrated here for the first time. Vertical transmission and mechanical vectoring by horse fly bites could not be demonstrated in this study, potentially due to limited sample size. We found EqPV-H shedding in oral and nasal secretions, and in feces. Importantly, we could demonstrate EqPV-H transmission via oral inoculation with viremic serum. Together, our findings provide additional information that EqPV-H is the likely cause of Theiler's disease and that transmission of EqPV-H occurs via both iatrogenic and natural routes.


Assuntos
Hepatite Viral Animal/virologia , Doenças dos Cavalos/virologia , Fígado/virologia , Infecções por Parvoviridae/veterinária , Parvovirus/fisiologia , Animais , Dípteros/virologia , Fezes/virologia , Feminino , Hepatite Viral Animal/patologia , Hepatite Viral Animal/transmissão , Hepatócitos/patologia , Hepatócitos/virologia , Doenças dos Cavalos/patologia , Doenças dos Cavalos/transmissão , Cavalos , Transmissão Vertical de Doença Infecciosa , Insetos Vetores/virologia , Fígado/patologia , Linfócitos , Masculino , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais/virologia , Boca/virologia , Necrose , Infecções por Parvoviridae/patologia , Infecções por Parvoviridae/transmissão , Infecções por Parvoviridae/virologia , Parvovirus/isolamento & purificação , Parvovirus/patogenicidade , Tropismo Viral , Viremia , Eliminação de Partículas Virais
11.
Zhonghua Gan Zang Bing Za Zhi ; 28(2): 147-151, 2020 Feb 20.
Artigo em Chinês | MEDLINE | ID: mdl-32164066

RESUMO

Objective: To investigate the effect of knockdown of O-GlcNAc transferase (OGT) on hepatocyte fat synthesis. Methods: Liver cell line L02 were used to established the model of hepatic steatosis. The levels of OGT and O-GlcNAc protein were detected by Western blot. The OGT knockdown cell line of L02 cells was established, and its lipid formation ability was detected after induction of oleic acid (OA). Real-time quantitative PCR (qRT-PCR) and Western blot were used to detect mRNA and protein expression of enzymes related to fat synthesis. An independent sample t test was used. Results: Western blot showed that the expression of OGT and O-GlcNAc was increased in L02 cells after adipogenesis (P < 0.05). After shOGT lentivirus infects L02 cells, OGT mRNA levels were down-regulated (P < 0.01). Oil red O staining showed that the lipid in L02 shOGT cells decreased, qRT-PCR showed that the mRNA expressions of fat synthase (ACC1), (FASN) and (SCD1) were decreased, the difference was statistically significant (P < 0.05), protein Expression is consistent with mRNA expression. Conclusion: Knockdown of OGT can inhibit hepatocyte fat synthesis by reducing O-GlcNAc levels.


Assuntos
Antígenos de Neoplasias/metabolismo , Fígado Gorduroso , Hepatócitos/metabolismo , Histona Acetiltransferases/metabolismo , Hialuronoglucosaminidase/metabolismo , N-Acetilglucosaminiltransferases/metabolismo , Linhagem Celular , Humanos
12.
Med Sci Monit ; 26: e921887, 2020 Mar 19.
Artigo em Inglês | MEDLINE | ID: mdl-32191680

RESUMO

BACKGROUND Recent studies have suggested that hepatocyte senescence could contribute to hepatic steatosis and its progression in nonalcoholic fatty liver disease (NAFLD). However, the underlying mechanism causing hepatocyte senescence in this pathological condition is still unclear. A thorough understanding of the mechanism could provide a new target for therapeutic intervention. The purpose of this study was to investigate the role of p66shc in hepatocyte senescence and hepatocyte damage in NAFLD progression. MATERIAL AND METHODS We examined the expression levels of hepatic p66shc and senescence markers in rats and humans with NAFLD, and we assessed the effect of p66shc knockdown or overexpression on senescence and steatosis in human liver cells. RESULTS In this study, we showed that increased hepatic p66shc expression was consistent with upregulated expression of the following senescence markers in NAFLD rats: heterochromatin protein-1-beta (HP1ß), p16, p21, and p53. Furthermore, senescence and steatosis could be induced in hepatoblastoma cell line (HepG2) cells when cells were stimulated with a low concentration of H2O2, and this effect was significantly alleviated by knockdown of p66shc. However, overexpression of p66shc could promote senescence and steatosis in L02 cells. Finally, increased hepatic p66shc protein levels correlated with enhanced expression of the senescence marker p21 and mirrored the degree of disease severity in NAFLD patients. CONCLUSIONS Our findings indicated that the increase in hepatocyte senescence and steatosis in NAFLD may be caused by the upregulation of p66shc expression, implying that strategies for p66shc-mediated regulation of hepatocyte senescence may provide new therapeutic tools for NAFLD.


Assuntos
Senescência Celular , Fígado Gorduroso/patologia , Hepatopatia Gordurosa não Alcoólica/patologia , Animais , Senescência Celular/fisiologia , Progressão da Doença , Fígado Gorduroso/metabolismo , Células Hep G2 , Hepatócitos/metabolismo , Humanos , Hepatopatia Gordurosa não Alcoólica/metabolismo , Ratos , Transdução de Sinais , Proteína 1 de Transformação que Contém Domínio 2 de Homologia de Src
13.
PLoS One ; 15(2): e0227940, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32027657

RESUMO

Tumor necrosis factor alpha (TNF) is capable of inducing regression of solid tumors. However, TNF released in response to Toll-like receptor 4 (TLR4) activation by bacterial lipopolysaccharide (LPS) is the key mediator of cytokine storm and septic shock that can cause severe tissue damage limiting anticancer applications of this cytokine. In our previous studies, we demonstrated that activation of another Toll-like receptor, TLR5, could protect from tissue damage caused by a variety of stresses including radiation, chemotherapy, Fas-activating antibody and ischemia-reperfusion. In this study, we tested whether entolimod could counteract TNF-induced toxicity in mouse models. We found that entolimod pretreatment effectively protects livers and lungs from LPS- and TNF-induced toxicity and prevents mortality caused by combining either of these agents with the sensitizer, D-galactosamine. While LPS and TNF induced significant activation of apoptotic caspase 3/7, lipid tissue peroxidation and serum ALT accumulation in mice without entolimod treatment, these indicators of toxicity were reduced by entolimod pretreatment to the levels of untreated control mice. Entolimod was effective when injected 0.5-48 hours prior to, but not when injected simultaneously or after LPS or TNF. Using chimeric mice with hematopoiesis differing in its TLR5 status from the rest of tissues, we showed that this protective activity was dependent on TLR5 expression by non-hematopoietic cells. Gene expression analysis identified multiple genes upregulated by entolimod in the liver and cultured hepatocytes as possible mediators of its protective activity. Entolimod did not interfere with the antitumor activity of TNF in mouse hepatocellular and colorectal tumor models. These results support further development of TLR5 agonists to increase tissue resistance to cytotoxic cytokines, reduce the risk of septic shock and enable safe systemic application of TNF as an anticancer therapy.


Assuntos
Antineoplásicos/farmacologia , Peptídeos/farmacologia , Receptor 5 Toll-Like/agonistas , Fator de Necrose Tumoral alfa/toxicidade , Animais , Linhagem Celular Tumoral , Células Cultivadas , Galactosamina , Hematopoese/efeitos dos fármacos , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Lipopolissacarídeos/toxicidade , Fígado/efeitos dos fármacos , Fígado/patologia , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Modelos Biológicos , NF-kappa B/metabolismo , Substâncias Protetoras/farmacologia , Análise de Sobrevida , Receptor 5 Toll-Like/metabolismo , Fator de Necrose Tumoral alfa/sangue , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/genética
14.
Nat Commun ; 11(1): 807, 2020 02 10.
Artigo em Inglês | MEDLINE | ID: mdl-32042044

RESUMO

Autophagy is essential for cellular survival and energy homeostasis under nutrient deprivation. Despite the emerging importance of nuclear events in autophagy regulation, epigenetic control of autophagy gene transcription remains unclear. Here, we report fasting-induced Fibroblast Growth Factor-21 (FGF21) signaling activates hepatic autophagy and lipid degradation via Jumonji-D3 (JMJD3/KDM6B) histone demethylase. Upon FGF21 signaling, JMJD3 epigenetically upregulates global autophagy-network genes, including Tfeb, Atg7, Atgl, and Fgf21, through demethylation of histone H3K27-me3, resulting in autophagy-mediated lipid degradation. Mechanistically, phosphorylation of JMJD3 at Thr-1044 by FGF21 signal-activated PKA increases its nuclear localization and interaction with the nuclear receptor PPARα to transcriptionally activate autophagy. Administration of FGF21 in obese mice improves defective autophagy and hepatosteatosis in a JMJD3-dependent manner. Remarkably, in non-alcoholic fatty liver disease patients, hepatic expression of JMJD3, ATG7, LC3, and ULK1 is substantially decreased. These findings demonstrate that FGF21-JMJD3 signaling epigenetically links nutrient deprivation with hepatic autophagy and lipid degradation in mammals.


Assuntos
Autofagia/genética , Jejum/metabolismo , Fatores de Crescimento de Fibroblastos/metabolismo , Histona Desmetilases com o Domínio Jumonji/metabolismo , Fígado/metabolismo , Animais , Autofagia/efeitos dos fármacos , Epigênese Genética , Fígado Gorduroso/metabolismo , Fígado Gorduroso/prevenção & controle , Fatores de Crescimento de Fibroblastos/administração & dosagem , Fatores de Crescimento de Fibroblastos/deficiência , Hepatócitos/metabolismo , Humanos , Histona Desmetilases com o Domínio Jumonji/genética , Lipólise , Masculino , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Knockout , Camundongos Obesos , PPAR alfa/metabolismo , Fosforilação , Ligação Proteica , Transdução de Sinais , Regulação para Cima
15.
PLoS One ; 15(2): e0224644, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32101552

RESUMO

Polybrominated diphenyl ethers (PBDEs) were formally used as flame-retardants and are chemically stable, lipophlic persistent organic pollutants which are known to bioaccumulate in humans. Although its toxicities are well characterized, little is known about the changes in transcriptional regulation caused by PBDE exposure. Long non-coding RNAs (lncRNAs) are increasingly recognized as key regulators of transcriptional and translational processes. It is hypothesized that lncRNAs can regulate nearby protein-coding genes (PCGs) and changes in the transcription of lncRNAs may act in cis to perturb gene expression of its neighboring PCGs. The goals of this study were to 1) characterize PCGs and lncRNAs that are differentially regulated from exposure to PBDEs; 2) identify PCG-lncRNA pairs through genome annotation and predictive binding tools; and 3) determine enriched canonical pathways caused by differentially expressed lncRNA-PCGs pairs. HepaRG cells, which are human-derived hepatic cells that accurately represent gene expression profiles of human liver tissue, were exposed to BDE-47 and BDE-99 at a dose of 25 µM for 24 hours. Differentially expressed lncRNA-PCG pairs were identified through DESeq2 and HOMER; significant canonical pathways were determined through Ingenuity Pathway Analysis (IPA). LncTar was used to predict the binding of 19 lncRNA-PCG pairs with known roles in drug-processing pathways. Genome annotation revealed that the majority of the differentially expressed lncRNAs map to PCG introns. PBDEs regulated overlapping pathways with PXR and CAR such as protein ubiqutination pathway and peroxisome proliferator-activated receptor alpha-retinoid X receptor alpha (PPARα-RXRα) activation but also regulate distinctive pathways involved in intermediary metabolism. PBDEs uniquely down-regulated GDP-L-fucose biosynthesis, suggesting its role in modifying important pathways involved in intermediary metabolism such as carbohydrate and lipid metabolism. In conclusion, we provide strong evidence that PBDEs regulate both PCGs and lncRNAs in a PXR/CAR ligand-dependent and independent manner.


Assuntos
Retardadores de Chama/farmacologia , Perfilação da Expressão Gênica/métodos , Éteres Difenil Halogenados/farmacologia , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , RNA Longo não Codificante/metabolismo , Metabolismo dos Carboidratos , Linhagem Celular , Retardadores de Chama/administração & dosagem , Regulação da Expressão Gênica , Éteres Difenil Halogenados/administração & dosagem , Humanos , Íntrons/genética , Metabolismo dos Lipídeos , PPAR alfa/metabolismo , Receptor de Pregnano X/genética , Receptor de Pregnano X/metabolismo , Receptores Citoplasmáticos e Nucleares/genética , Receptores Citoplasmáticos e Nucleares/metabolismo , Receptor X Retinoide alfa/metabolismo
16.
Artigo em Inglês | MEDLINE | ID: mdl-32014661

RESUMO

In this study, we performed the metabolism of endosulfan sulfate in human liver preparations (human liver microsomes, S9 fractions and hepatocytes) to identify new metabolites using liquid chromatography-high resolution mass spectrometry (LC-HRMS). Endosulfan sulfate is a major oxidized metabolite of the organochlorine insecticide endosulfan, and it exhibits a similar toxicity to endosulfan. Six metabolites, including 5 novel metabolites of endosulfan sulfate, were identified in the three different human liver reaction mixtures and metabolic pathways of endosulfan sulfate were proposed. The phase I metabolites M1 and M2 were observed in human liver microsomes, S9 fractions and hepatocytes. M1 was suggested to be an endosulfan diol monosulfate and M2 was identified as (1,4,5,6,7,7-hexachloro-3-formylbicyclo[2,2,1]hept-5-en-2-yl)methyl hydrogen sulfate through the interpretation of the HRMS spectrum. The phase II metabolite M3 was produced as an endosulfan sulfate-GSH conjugate in those three liver preparations and transformed to M5 (dipeptide) in S9 fractions and hepatocytes. M3 was the most predominant metabolite identified in the three liver preparations. M4 was only detected in microsomes as an M2-GSH conjugate and was metabolized to M6 (monopeptide) in hepatocytes. These results are different from the metabolic pathway of endosulfan and suggest the possible detoxification metabolic reaction of endosulfan sulfate in living organisms.


Assuntos
Endossulfano/análogos & derivados , Cromatografia Líquida de Alta Pressão , Endossulfano/análise , Endossulfano/metabolismo , Hepatócitos/metabolismo , Humanos , Fígado/metabolismo , Metaboloma/fisiologia , Microssomos Hepáticos/metabolismo , Oxirredução , Ésteres do Ácido Sulfúrico/análise , Ésteres do Ácido Sulfúrico/metabolismo , Espectrometria de Massas em Tandem
17.
Phytomedicine ; 68: 153153, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-32018210

RESUMO

BACKGROUD: Cholestasis, accompanied by the accumulation of bile acids in body, may ultimately cause liver failure and cirrhosis. There have been limited therapies for cholesteric disorders. Therefore, development of appropriate therapeutic drugs for cholestasis is required. Picroside II is a bioactive component isolated from Picrorhiza scrophulariiflora Pennell, its mechanistic contributions to the anti-cholestasis effect have not been fully elucidated, especially the role of picroside II on bile acid homeostasis via nuclear receptors remains unclear. PURPOSE: This study was designed to investigate the hepatoprotective effect of picroside II against alpha-naphthylisothiocyanate (ANIT)-induced cholestatic liver injury and elucidate the mechanisms in vivo and in vitro. METHODS: The ANIT-induced cholestatic mouse model was used with or without picroside II treatment. Serum and bile biochemical indicators, as well as liver histopathological changes were examined. siRNA, Dual-luciferase reporter, quantitative real-time PCR and Western blot assay were used to demonstrate the farnesoid X receptor (FXR) pathway in the anti-cholestasis effects of picroside II in vivo and in vitro. RESULTS: Picroside II exerted hepatoprotective effect against ANIT-induced cholestasis by impaired hepatic function and tissue damage. Picroside II increased bile acid efflux transporter bile salt export pump (Bsep), uptake transporter sodium taurocholate cotransporting polypeptide (Ntcp), and bile acid metabolizing enzymes sulfate transferase 2a1 (Sult2a1) and UDP-glucuronosyltransferase 1a1 (Ugt1a1), whereas decreased the bile acid synthesis enzymes cholesterol 7α-hydroxylase (Cyp7a1) and oxysterol 12α-hydroxylase (Cyp8b1). In addition, expression of FXR and the target gene Bsep was increased, whereas aryl hydrocarbon receptor (AhR), pregnane X receptor (PXR), peroxisome proliferator-activated receptor alpha (PPARα) and their corresponding target genes were not significantly influenced by picroside II under cholestatic conditions. Furthermore, regulation of transporters and enzymes involved in bile acid homeostasis by picroside II were abrogated by FXR silencing in mouse primary cultured hepatocytes. Dual-luciferase reporter assay performed in HepG2 cells demonstrated FXR activation by picroside II. CONCLUSION: Our findings demonstrate that picroside II exerts protective effect on ANIT-induced cholestasis possibly through FXR activation that regulates the transporters and enzymes involved in bile acid homeostasis. Picroside II might be an effective approach for the prevention and treatment of cholestatic liver diseases.


Assuntos
Colestase/prevenção & controle , Cinamatos/farmacologia , Glucosídeos Iridoides/farmacologia , Hepatopatias/prevenção & controle , Receptores Citoplasmáticos e Nucleares/metabolismo , 1-Naftilisotiocianato/toxicidade , Membro 11 da Subfamília B de Transportadores de Cassetes de Ligação de ATP/genética , Membro 11 da Subfamília B de Transportadores de Cassetes de Ligação de ATP/metabolismo , Animais , Ácidos e Sais Biliares/genética , Ácidos e Sais Biliares/metabolismo , Colestase/fisiopatologia , Regulação da Expressão Gênica/efeitos dos fármacos , Células Hep G2 , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Humanos , Fígado/efeitos dos fármacos , Hepatopatias/metabolismo , Hepatopatias/patologia , Masculino , Camundongos Endogâmicos C57BL , Substâncias Protetoras/farmacologia
18.
Nat Commun ; 11(1): 719, 2020 02 05.
Artigo em Inglês | MEDLINE | ID: mdl-32024826

RESUMO

Lipid overload results in lipid redistribution among metabolic organs such as liver, adipose, and muscle; therefore, the interplay between liver and other organs is important to maintain lipid homeostasis. Here, we show that liver responds to lipid overload first and sends hepatocyte-derived extracellular vesicles (EVs) targeting adipocytes to regulate adipogenesis and lipogenesis. Geranylgeranyl diphosphate synthase (Ggpps) expression in liver is enhanced by lipid overload and regulates EV secretion through Rab27A geranylgeranylation. Consistently, liver-specific Ggpps deficient mice have reduced fat adipose deposition. The levels of several EV-derived miRNAs in the plasma of non-alcoholic fatty liver disease (NAFLD) patients are positively correlated with body mass index (BMI), and these miRNAs enhance adipocyte lipid accumulation. Thus, we highlight an inter-organ mechanism whereby the liver senses different metabolic states and sends corresponding signals to remodel adipose tissue to adapt to metabolic changes in response to lipid overload.


Assuntos
Tecido Adiposo/metabolismo , Vesículas Extracelulares/metabolismo , Hepatócitos/metabolismo , Metabolismo dos Lipídeos/fisiologia , Fígado/metabolismo , Hepatopatia Gordurosa não Alcoólica/sangue , Adipócitos/metabolismo , Adipócitos/patologia , Tecido Adiposo/patologia , Animais , Índice de Massa Corporal , Dieta Hiperlipídica/efeitos adversos , Vesículas Extracelulares/genética , Farnesiltranstransferase/genética , Humanos , Lipogênese , Fígado/citologia , Fígado/efeitos dos fármacos , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , MicroRNAs/sangue , Complexos Multienzimáticos/genética , Hepatopatia Gordurosa não Alcoólica/genética , Proteínas rab27 de Ligação ao GTP/genética , Proteínas rab27 de Ligação ao GTP/metabolismo
19.
Chemosphere ; 248: 126036, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32045972

RESUMO

Aflatoxin B1 (AFB1) and microcystin-LR (MC-LR) co-existed in food and water, and were associated with hepatocellular carcinoma (HCC). AFB1 induced HCC by activating oxidative stress and generating AFB1-DNA adducts, while MC-LR could promote HCC progression. However, whether they have co-effects in HCC progression remains uncertain. In this study, we found the antagonistic effects of MC-LR on AFB1 induced HCC when they were exposed simultaneously. Compared with single exposure to AFB1, co-exposed to MC-LR significantly repressed the AFB1 induced malignant transformation of human hepatic cells and the glutathione S-transferase Pi positive foci formation in rat livers. MC-LR inhibited AFB1 induced upregulation of cytochrome P450 family 1 subfamily A member 2 (CYP1A2) and reduced the AFB1-DNA adducts generation in both human hepatic cells and rat livers. These results suggest that when co-exposure with AFB1, MC-LR might repress hepatocarcinogenicity of AFB1, which might be associated with its repression on AFB1 induced CYP1A2 upregulation and activation.


Assuntos
Aflatoxina B1/metabolismo , Citocromo P-450 CYP1A2/metabolismo , Adutos de DNA/metabolismo , Microcistinas/toxicidade , Animais , Carcinoma Hepatocelular/induzido quimicamente , Carcinoma Hepatocelular/metabolismo , Hepatócitos/metabolismo , Humanos , Fígado/metabolismo , Neoplasias Hepáticas/induzido quimicamente , Neoplasias Hepáticas/metabolismo , Masculino , Estresse Oxidativo , Ratos
20.
Biochem Soc Trans ; 48(1): 51-59, 2020 02 28.
Artigo em Inglês | MEDLINE | ID: mdl-32096539

RESUMO

Much of the world's prominent and burdensome chronic diseases, such as diabetes, Alzheimer's, and heart disease, are caused by impaired metabolism. By acting as both an efficient fuel and a powerful signalling molecule, the natural ketone body, d-ß-hydroxybutyrate (ßHB), may help circumvent the metabolic malfunctions that aggravate some diseases. Historically, dietary interventions that elevate ßHB production by the liver, such as high-fat diets and partial starvation, have been used to treat chronic disease with varying degrees of success, owing to the potential downsides of such diets. The recent development of an ingestible ßHB monoester provides a new tool to quickly and accurately raise blood ketone concentration, opening a myriad of potential health applications. The ßHB monoester is a salt-free ßHB precursor that yields only the biologically active d-isoform of the metabolite, the pharmacokinetics of which have been studied, as has safety for human consumption in athletes and healthy volunteers. This review describes fundamental concepts of endogenous and exogenous ketone body metabolism, the differences between the ßHB monoester and other exogenous ketones and summarises the disease-specific biochemical and physiological rationales behind its clinical use in diabetes, neurodegenerative diseases, heart failure, sepsis related muscle atrophy, migraine, and epilepsy. We also address the limitations of using the ßHB monoester as an adjunctive nutritional therapy and areas of uncertainty that could guide future research.


Assuntos
Ácido 3-Hidroxibutírico/metabolismo , Ácido 3-Hidroxibutírico/uso terapêutico , Diabetes Mellitus/dietoterapia , Dieta Cetogênica , Suplementos Nutricionais , Epilepsia/dietoterapia , Jejum/metabolismo , Insuficiência Cardíaca/dietoterapia , Hepatócitos/metabolismo , Humanos , Doenças Neurodegenerativas/dietoterapia , Sepse/dietoterapia
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