RESUMO
BACKGROUND AND AIMS: Peroxisome proliferator-activated receptor α (PPARα, NR1C1) is a ligand-activated nuclear receptor involved in the regulation of lipid catabolism and energy homeostasis. PPARα activation induces hepatomegaly and plays an important role in liver regeneration, but the underlying mechanisms remain unclear. APPROACH AND RESULTS: In this study, the effect of PPARα activation on liver enlargement and regeneration was investigated in several strains of genetically modified mice. PPARα activation by the specific agonist WY-14643 significantly induced hepatomegaly and accelerated liver regeneration after 70% partial hepatectomy (PHx) in wild-type mice and Pparafl/fl mice, while these effects were abolished in hepatocyte-specific Ppara-deficient (PparaΔHep ) mice. Moreover, PPARα activation promoted hepatocyte hypertrophy around the central vein area and hepatocyte proliferation around the portal vein area. Mechanistically, PPARα activation regulated expression of yes-associated protein (YAP) and its downstream targets (connective tissue growth factor, cysteine-rich angiogenic inducer 61, and ankyrin repeat domain 1) as well as proliferation-related proteins (cyclins A1, D1, and E1). Binding of YAP with the PPARα E domain was critical for the interaction between YAP and PPARα. PPARα activation further induced nuclear translocation of YAP. Disruption of the YAP-transcriptional enhancer factor domain family member (TEAD) association significantly suppressed PPARα-induced hepatomegaly and hepatocyte enlargement and proliferation. In addition, PPARα failed to induce hepatomegaly in adeno-associated virus-Yap short hairpin RNA-treated mice and liver-specific Yap-deficient mice. Blockade of YAP signaling abolished PPARα-induced hepatocyte hypertrophy around the central vein area and hepatocyte proliferation around the portal vein area. CONCLUSIONS: This study revealed a function of PPARα in regulating liver size and liver regeneration through activation of the YAP-TEAD signaling pathway. These findings have implications for understanding the physiological functions of PPARα and suggest its potential for manipulation of liver size and liver regeneration.
Assuntos
Hepatomegalia/genética , Regeneração Hepática/genética , PPAR alfa/metabolismo , Fatores de Transcrição de Domínio TEA/metabolismo , Proteínas de Sinalização YAP/metabolismo , Animais , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Modelos Animais de Doenças , Regulação da Expressão Gênica/efeitos dos fármacos , Técnicas de Silenciamento de Genes , Hepatectomia/efeitos adversos , Hepatócitos/patologia , Hepatomegalia/patologia , Humanos , Fígado/patologia , Fígado/cirurgia , Regeneração Hepática/efeitos dos fármacos , Masculino , Camundongos , Camundongos Transgênicos , PPAR alfa/agonistas , Pirimidinas/administração & dosagem , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Proteínas de Sinalização YAP/genéticaRESUMO
Peroxisome proliferator-activated receptor α (PPARα) is a key nuclear receptor involved in the control of lipid homeostasis. In rodents, PPARα is also a potent hepatic mitogen. Hepatocyte-specific disruption of PPARα inhibits agonist-induced hepatocyte proliferation; however, little is known about the exact role of PPARα in partial hepatectomy (PHx)-induced liver regeneration. Herein, using hepatocyte-specific PPARα-deficient (PparaΔHep) mice, the function of hepatocyte PPARα in PHx-induced liver regeneration was investigated. PPARα protein level and transcriptional activity were increased in the liver after PHx. Compared with the Pparafl/fl mice, PparaΔHep mice exhibited significantly reduced hepatocyte proliferation at 32 hours after PHx. Consistently, reduced Ccnd1 and Pcna mRNA and CYCD1 and proliferating cell nuclear antigen protein were observed at 32 hours after PHx in PparaΔHep mice. Furthermore, PparaΔHep mice showed increased hepatic lipid accumulation and enhanced hepatic triglyceride contents because of impaired hepatic fatty acid ß-oxidation when compared with that observed in Pparafl/fl mice. These results indicate that PPARα promotes liver regeneration after PHx, at least partially via regulating the cell cycle and lipid metabolism.
Assuntos
Ciclo Celular , Metabolismo dos Lipídeos , Regeneração Hepática , Fígado/metabolismo , PPAR alfa/metabolismo , Animais , Ciclina D1/genética , Ciclina D1/metabolismo , Ácidos Graxos/genética , Ácidos Graxos/metabolismo , Hepatectomia , Masculino , Camundongos , Camundongos Transgênicos , Oxirredução , PPAR alfa/genética , Antígeno Nuclear de Célula em Proliferação/genética , Antígeno Nuclear de Célula em Proliferação/metabolismo , Fatores de Tempo , Triglicerídeos/genética , Triglicerídeos/metabolismoRESUMO
Chronic activation of the nuclear receptor peroxisome proliferator-activated receptor alpha (PPARA) promotes MYC-linked hepatocellular carcinoma (HCC) in mice. Recent studies have shown that MYC can function as an amplifier of transcription where MYC does not act as an "on-off" switch for gene expression but rather accelerates transcription rates at active promoters by stimulating transcript elongation. Considering the possibility that MYC may amplify the expression of PPARA target genes to potentiate cell proliferation and liver cancer, gene expression was analyzed from livers of wild-type and liver-specific Myc knockout (MycΔHep ) mice treated with the PPARA agonist pirinixic acid. A subset of PPARA target genes was amplified in the presence of MYC, including keratin 23 (Krt23). The induction of Krt23 was significantly attenuated in MycΔHep mice and completely abolished in Ppara-null mice. Reporter gene assays and chromatin immunoprecipitation confirmed direct binding of both PPARA and MYC to sites within the Krt23 promoter. Forced expression of KRT23 in primary hepatocytes induced cell cycle-related genes. These data indicate that PPARA activation elevates MYC expression, which in turn potentiates the expression of select PPARA target genes involved in cell proliferation. Finally, KRT23 protein is highly elevated in human HCCs. Conclusion: These results revealed that MYC-mediated transcriptional potentiation of select PPARA target genes, such as Krt23, may remove rate-limiting constraints on hepatocyte growth and proliferation leading to liver cancer.
Assuntos
Regulação da Expressão Gênica , Hepatócitos/fisiologia , Queratinas/metabolismo , Proteína Oncogênica p55(v-myc)/metabolismo , PPAR alfa/metabolismo , Animais , Carcinoma Hepatocelular/sangue , Carcinoma Hepatocelular/etiologia , Proliferação de Células , Feminino , Humanos , Queratinas/genética , Queratinas Tipo I/sangue , Neoplasias Hepáticas/sangue , Neoplasias Hepáticas/etiologia , Masculino , CamundongosRESUMO
Nonalcoholic steatohepatitis (NASH) is the progressive stage of nonalcoholic fatty liver disease that may ultimately lead to cirrhosis and liver cancer, and there are few therapeutic options for its treatment. Glycyrrhizin (GL), extracted from the traditional Chinese medicine liquorice, has potent hepatoprotective effects in both preclinical animal models and in humans. However, little is currently known about its effects and mechanisms in treating NASH. To explore the effects of GL on NASH, GL or its active metabolite glycyrrhetinic acid (GA) was administered to mice treated with a methionine- and choline-deficient (MCD) diet-induced NASH model, and histologic and biochemical analyses were used to measure the degree of lipid disruption, liver inflammation, and fibrosis. GL significantly improved MCD diet-induced hepatic steatosis, inflammation, and fibrosis and inhibited activation of the NLR family pyrin domain-containing 3 (NLRP3) inflammasome. GL significantly attenuated serum bile acid accumulation in MCD diet-fed mice partially by restoring inflammation-mediated hepatic farnesoid X receptor inhibition. In Raw 264.7 macrophage cells, both GL and GA inhibited deoxycholic acid-induced NLRP3 inflammasome-associated inflammation. Notably, both intraperitoneal injection of GL's active metabolite GA and oral administration of GL prevented NASH in mice, indicating that GL may attenuate NASH via its active metabolite GA. These results reveal that GL, via restoration of bile acid homeostasis and inhibition of inflammatory injury, can be a therapeutic option for treatment of NASH.
Assuntos
Anti-Inflamatórios/uso terapêutico , Ácidos e Sais Biliares/fisiologia , Ácido Glicirrízico/uso terapêutico , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Hepatopatia Gordurosa não Alcoólica/metabolismo , Animais , Anti-Inflamatórios/farmacologia , Ácido Glicirrízico/farmacologia , Células Hep G2 , Humanos , Inflamação/tratamento farmacológico , Inflamação/metabolismo , Inflamação/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Hepatopatia Gordurosa não Alcoólica/patologia , Células RAW 264.7 , Distribuição AleatóriaRESUMO
Rutaecarpine (RUT), evodiamine (EOD), and dehydroevodiamine (DHED) are the three main bioactive indoloquinazoline alkaloids isolated from Euodia rutaecarpa, a widely prescribed traditional Chinese medicine. Here, the structure-activity relationships of these analogs for aryl hydrocarbon receptor (AHR) activation were explored by use of Ahr-deficient (Ahr-/-) mice, primary hepatocyte cultures, luciferase reporter gene assays, in silico ligand-docking studies, and metabolomics. In vitro, both mRNA analysis of AHR target genes in mouse primary hepatocytes and luciferase reporter assays in hepatocarcinoma cell lines demonstrated that RUT, EOD, and DHED significantly activated AHR, with an efficacy order of RUT > DHED > EOD. Ligand-docking analysis predicted that the methyl substitute at the N-14 atom was a key factor affecting AHR activation. In vivo, EOD was poorly orally absorbed and failed to activate AHR, whereas RUT and DHED markedly upregulated expression of the hepatic AHR gene battery in wild-type mice, but not in Ahr-/- mice. Furthermore, RUT, EOD, and DHED were not hepatotoxic at the doses used; however, RUT and DHED disrupted bile acid homeostasis in an AHR-dependent manner. These findings revealed that the methyl group at the N-14 atom of these analogs and their pharmacokinetic behaviors were the main determinants for AHR activation, and suggest that attention should be given to monitoring bile acid metabolism in the clinical use of E. rutaecarpa.
Assuntos
Ácidos e Sais Biliares/metabolismo , Medicamentos de Ervas Chinesas/farmacologia , Evodia/química , Homeostase/efeitos dos fármacos , Receptores de Hidrocarboneto Arílico/metabolismo , Alcaloides/farmacologia , Animais , Linhagem Celular Tumoral , Genes Reporter/efeitos dos fármacos , Células Hep G2 , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Humanos , Alcaloides Indólicos/farmacologia , Fígado/diagnóstico por imagem , Fígado/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Quinazolinas/farmacologia , RNA Mensageiro/metabolismo , Relação Estrutura-Atividade , Regulação para Cima/efeitos dos fármacosRESUMO
Acetaminophen (APAP) overdose is the leading cause of drug-induced acute liver failure in Western countries. Glycyrrhizin (GL), a potent hepatoprotective constituent extracted from the traditional Chinese medicine liquorice, has potential clinical use in treating APAP-induced liver failure. The present study determined the hepatoprotective effects and underlying mechanisms of action of GL and its active metabolite glycyrrhetinic acid (GA). Various administration routes and pharmacokinetics-pharmacodynamics analyses were used to differentiate the effects of GL and GA on APAP toxicity in mice. Mice deficient in cytochrome P450 2E1 enzyme (CYP2E1) or receptor interacting protein 3 (RIPK3) and their relative wild-type littermates were subjected to histologic and biochemical analyses to determine the potential mechanisms. Hepatocyte death mediated by tumor necrosis factorα(TNFα)/caspase was analyzed by use of human liver-derived LO2 cells. The pharmacokinetics-pharmacodynamics analysis using various administration routes revealed that GL but not GA potently attenuated APAP-induced liver injury. The protective effect of GL was found only with intraperitoneal and intravenous administration and not with gastric administration. CYP2E1-mediated metabolic activation and RIPK3-mediated necroptosis were unrelated to GL's protective effect. However, GL inhibited hepatocyte apoptosis via interference with TNFα-induced apoptotic hepatocyte death. These results demonstrate that GL rapidly attenuates APAP-induced liver injury by directly inhibiting TNFα-induced hepatocyte apoptosis. The protective effect against APAP-induced liver toxicity by GL in mice suggests the therapeutic potential of GL for the treatment of APAP overdose.
Assuntos
Acetaminofen/efeitos adversos , Apoptose/efeitos dos fármacos , Doença Hepática Induzida por Substâncias e Drogas/tratamento farmacológico , Ácido Glicirrízico/farmacologia , Fígado/efeitos dos fármacos , Substâncias Protetoras/farmacologia , Fator de Necrose Tumoral alfa/metabolismo , Ativação Metabólica/efeitos dos fármacos , Animais , Linhagem Celular , Citocromo P-450 CYP2E1/metabolismo , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Humanos , Fígado/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Mitocôndrias Hepáticas/efeitos dos fármacos , Mitocôndrias Hepáticas/metabolismo , Proteína Serina-Treonina Quinases de Interação com Receptores/metabolismoRESUMO
The liver has a remarkable capacity to regenerate after injury. Although the regulatory mechanisms of hepatocytic regeneration have been a subject of intense study, the dynamism of the sinusoids, specialized blood vessels in the liver, remains largely unknown. Transient activation of hepatic stellate cells and hepatic sinusoidal endothelial cells, which constitute the sinusoids, contributes to liver regeneration during acute injury, whereas their sustained activation causes liver fibrosis during chronic injury. We focused on understanding the association between damaged hepatocytes and sinusoidal regeneration or liver fibrogenesis using a carbon tetrachloride-induced liver injury mouse model. Damaged hepatocytes rapidly expressed semaphorin 3E (Sema3e), which induced contraction of sinusoidal endothelial cells and thereby contributed to activating hepatic stellate cells for wound healing. In addition, ectopic and consecutive expression of Sema3e in hepatocytes by the hydrodynamic tail-vein injection method resulted in disorganized regeneration of sinusoids and sustained activation of hepatic stellate cells. In contrast, liver fibrosis ameliorated in Sema3e-knockout mice compared with wild-type mice in a chronic liver injury model. Our results indicate that Sema3e, secreted by damaged hepatocytes, affects sinusoidal regeneration in a paracrine manner during liver regeneration, suggesting that Sema3e is a novel therapeutic target in liver fibrogenesis.
Assuntos
Hepatócitos/metabolismo , Cirrose Hepática/metabolismo , Regeneração Hepática/fisiologia , Semaforinas/metabolismo , Animais , Modelos Animais de Doenças , Células Estreladas do Fígado/metabolismo , Imuno-Histoquímica , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Nephronectin (Npnt) is an extracellular matrix protein known to play a critical role in kidney development; however, its physiological role in the liver remains elusive. Here we show that Npnt expression is upregulated in mouse models of both acute and chronic hepatitis induced by Concanavalin A (Con A) and 3,5-diethocarbonyl-1,4-dihydrocollidine (DDC), respectively. In both models, Npnt was localized in inflammatory foci and was mainly secreted from mesenchymal cells and in part by cholangiocytes. Interestingly, ectopic expression of Npnt in hepatocytes induced the development of granuloma-like cell clusters mainly composed of CD4(+) T cells or NKT cells but did not induce apparent hepatitis. Furthermore, we found that Npnt exacerbated the Con A-induced acute hepatitis. These results indicate that Npnt plays an important role in the initiation of hepatitis by recruiting CD4(+) T cells or NKT cells into the foci of inflammation. In addition, we reveal that Npnt expression is also upregulated in human hepatitis. Therefore, Npnt may be a potential therapeutic target for acute and chronic hepatitis.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Proteínas da Matriz Extracelular/fisiologia , Hepatite/imunologia , Hepatite/patologia , Fígado/imunologia , Fígado/patologia , Doença Aguda , Animais , Movimento Celular , Concanavalina A/toxicidade , Modelos Animais de Doenças , Progressão da Doença , Proteínas da Matriz Extracelular/genética , Regulação da Expressão Gênica , Granuloma/imunologia , Granuloma/patologia , Hepatite/genética , Hepatite Crônica/genética , Hepatite Crônica/imunologia , Hepatite Crônica/patologia , Fígado/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Regulação para CimaRESUMO
Elasmobranchs (sharks and rays) exhibit unique reproductive characteristics and, in contrast to the situation in teleosts, very little is known about the identity, structure and physical characteristics of their egg yolk proteins. The aims of this study were to (1) detect and purify the vitellogenin (Vtg; egg yolk precursor) and yolk proteins (YPs) of the cloudy catshark (Scyliorhinus torazame), (2) examine the relationships between Vtg and YPs and (3) characterize and classify the deduced primary structure of the Vtg transcript (vtg). The apparent molecular weights of purified Vtg and putative Vtg-related YPs (lipovitellin: Lv, phosvitin: Pv) were determined by gel filtration and were ~560, >669 and ~58 kDa, respectively. Following SDS-PAGE, these purified products (i.e., Vtg, Lv and Pv) appeared as bands of ~210, ~110 and ~22 kDa, respectively. On Western blots, antisera against purified Vtg, Lv and Pv recognized the ~210 kDa Vtg band. Catshark Pv, in contrast to teleost Pvs, had a very low serine content. The catshark Vtg cDNA sequence (vtg) appeared to contain an open-reading frame consisting of domains encoding Lv, Pv and ß'-component (ß'-c). A phylogenetic analysis, with a consideration of genome duplication events, placed catshark vtg into the 'vtgAB type.' It is concluded that at least a single major type of Vtg protein, which is transcribed and translated from catshark vtgAB gene, is the precursor of three egg yolk proteins (Lv, Pv and ß'-c) in catshark.
Assuntos
Proteínas do Ovo/genética , Fosvitina/genética , Tubarões/metabolismo , Vitelogeninas/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Western Blotting/veterinária , Cromatografia/veterinária , Cromatografia em Gel/veterinária , Análise por Conglomerados , Biologia Computacional , Primers do DNA/genética , DNA Complementar/genética , Proteínas do Ovo/química , Eletroforese em Gel de Poliacrilamida/veterinária , Dados de Sequência Molecular , Peso Molecular , Fosvitina/química , Filogenia , Análise de Sequência de DNA/veterinária , Vitelogeninas/análise , Vitelogeninas/químicaRESUMO
Non-alcoholic fatty liver disease (NAFLD) is one of the most prevalent hepatic disorders that 20-30% of the world population suffers from. The feature of NAFLD is excess lipid accumulation in the liver, exacerbating multiple metabolic syndromes such as hyperlipidemia, hypercholesterolemia, hypertension, and type 2 diabetes. Approximately 20-30% of NAFLD cases progress to more severe chronic hepatitis, known as non-alcoholic steatohepatitis (NASH), showing deterioration of hepatic functions and liver fibrosis followed by cirrhosis and cancer. Previous studies uncovered that several metabolic regulators had roles in disease progression as key factors. Peroxisome proliferator-activated receptor alpha (PPARα) has been identified as one of the main players in hepatic lipid homeostasis. PPARα is abundantly expressed in hepatocytes, and is a ligand-dependent nuclear receptor belonging to the NR1C nuclear receptor subfamily, orchestrating lipid/glucose metabolism, inflammation, cell proliferation, and carcinogenesis. PPARα agonists are expected to be novel prescription drugs for NASH treatment, and some of them (e.g., Lanifibranor) are currently under clinical trials. These potential novel drugs are developed based on the knowledge of PPARα-activating target genes related to NAFLD and NASH. Intriguingly, PPARα is known to suppress the expression of subsets of target genes under agonist treatment; however, the mechanisms of PPARα-mediated gene suppression and functions of these genes are not well understood. In this review, we summarize and discuss the mechanisms of target gene repression by PPARα and the roles of repressed target genes on hepatic lipid metabolism, fibrosis and carcinogenesis related to NALFD and NASH, and provide future perspectives for PPARα pharmaceutical potentials.
RESUMO
Peroxisome proliferator-activated receptor α (PPARA) is a key mediator of lipid metabolism and inflammation. Activation of PPARA in rodents causes hepatocyte proliferation, but the underlying mechanism is poorly understood. This study focused on genes repressed by PPARA and analyzed the mechanism by which PPARA promotes hepatocyte proliferation in mice. Activation of PPARA by agonist treatment was autoregulated, and induced expression of the epigenetic regulator UHRF1 via activation of the newly described PPARA target gene E2f8, which, in turn, regulates Uhrf1. UHRF1 strongly repressed the expression of CDH1 via methylation of the Cdh1 promoter marked with H3K9me3. Repression of CDH1 by PPARA activation was reversed by PPARA deficiency or knockdown of E2F8 or UHRF1. Furthermore, a forced expression of CDH1 inhibited expression of the Wnt signaling target genes such as Myc after PPARA activation, and suppressed hepatocyte hyperproliferation. These results demonstrate that the PPARA-E2F8-UHRF1-CDH1 axis causes epigenetic regulation of hepatocyte proliferation.
RESUMO
Peroxisome proliferator-activated receptor α (PPARα) controls hepatic lipid homeostasis and is the target of lipid-lowering fibrate drugs. PPARα activation represses expression of let-7 microRNA (miRNA), but the function of let-7 in PPARα signaling and lipid metabolism is unknown. In the current study, a hepatocyte-specific let-7b/c2 knockout (let7b/c2ΔHep) mouse line is generated, and these mice are found to exhibit pronounced resistance to diet-induced obesity and fatty liver. Let-7 inhibition by hepatocyte-specific let-7 sponge expression shows similar phenotypes as let7b/c2ΔHep mice. RNA sequencing (RNA-seq) analysis reveals that hepatic PPARα signaling is repressed in let7b/c2ΔHep mice. Protein expression of the obligate PPARα heterodimer partner retinoid X receptor α (RXRα) is reduced in the livers of let7b/c2ΔHep mice. Ring finger protein 8 (Rnf8), which is a direct target of let-7, is elevated in let7b/c2ΔHep mouse liver and identified as a E3 ubiquitin ligase for RXRα. This study highlights a let-7-RNF8-RXRα regulatory axis that modulates hepatic lipid catabolism.
Assuntos
Retroalimentação Fisiológica , MicroRNAs/metabolismo , PPAR alfa/metabolismo , Transdução de Sinais , Animais , Sequência de Bases , Dependovirus/metabolismo , Regulação da Expressão Gênica , Hepatócitos/metabolismo , Fígado/metabolismo , Camundongos Knockout , MicroRNAs/genética , Obesidade/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptor X Retinoide alfa/metabolismo , Ubiquitina-Proteína Ligases/metabolismoRESUMO
The idiosyncratic characteristics and severity of acetaminophen (APAP) overdose-induced hepatotoxicity render identifying the predisposing factors and mechanisms of APAP-induced liver toxicity necessary and urgent. Farnesoid X receptor (FXR) controls bile acid homeostasis and modulates the progression of various liver diseases. Although global FXR deficiency in mice enhances APAP intoxication, the mechanism remains elusive. In this study, an increased sensitivity to APAP-induced toxicity was found in global Fxr-null (Fxr-/-) mice, but was not observed in hepatocyte-specific or macrophage-specific Fxr-null mice, suggesting that global FXR deficiency enhances APAP hepatotoxicity via disruption of systematic bile acid homeostasis. Indeed, more bile acid accumulation was found in global Fxr-/- mice, while 2% cholestyramine diet feeding decreased serum bile acids and alleviated APAP hepatotoxicity in global Fxr-/- mice, suggesting that bile acid accumulation contributes to APAP toxicity. Bile acids were suspected to induce macrophage to release tumor necrosis factor-α (TNF-α), which is known to enhance the APAP hepatotoxicity. In vitro, deoxycholic acid (DCA), a secondary bile acid metabolite, significantly induced Tnfa mRNA and dose-dependently enhanced TNF-α release from macrophage, while the same dose of DCA did not directly potentiate APAP toxicity in cultured primary hepatocytes. In vivo, DCA enhanced TNF-α release and potentiated APAP toxicity, both of which were abolished by the specific TNF-α antagonist infliximab. These results reveal an FXR-DCA-TNF-α axis that potentiates APAP hepatotoxicity, which could guide the clinical safe use of APAP.
Assuntos
Acetaminofen , Doença Hepática Induzida por Substâncias e Drogas , Acetaminofen/toxicidade , Animais , Doença Hepática Induzida por Substâncias e Drogas/etiologia , Ácido Desoxicólico , Hepatócitos , Fígado , Camundongos , Camundongos Endogâmicos C57BLRESUMO
Inflammatory bowel disease (IBD) represents a group of chronic relapsing intestinal disorders. Rutaecarpine (RUT), isolated from the Traditional Chinese Medicine (TCM) of Evodia rutaecarpa, was reported to suppress IBD. However, the mechanism by which RUT ameliorates dextran sulfate sodium (DSS)-induced IBD is largely unknown. By use of nuclear factor-erythroid 2-related factor 2 (NRF2) knockout mice, cell-based studies, surface plasmon resonance (SPR), western blotting analysis, and molecular docking studies, the mechanism by which RUT affects DSS-induced colitis was explored. In DSS-treated wild-type mice but not in Nrf2-null mice, RUT significantly improved colitis as revealed by rescued body weight loss, improved histology and inflammation, and induced expression of NRF2 target genes in colon and ileum. Cell-based studies showed that RUT significantly increased the LD50 for hydrogen peroxide (H2O2)-induced cell damage, activated NRF2 nuclear translocation, and suppressed the production of reactive oxygen species in H2O2-treated HCT116 cells, activated NRF2 luciferase reporter activities in HCT116 cells and HepG2 cells, and induced expression of NRF2 target genes in primary intestinal epithelial cells. Molecular docking in silico and SPR assays indicated that RUT interacted with kelch-like ECH-associated protein 1 (KEAP1), and extracellular incubation studies revealed that RUT bound to the KEAP1 kelch domain with a calculated equilibrium dissociation constant Kd of 19.6 µM. In conclusion, these results demonstrate that RUT ameliorates DSS-induced colitis, dependent on NRF2, and could be a potential therapeutic option for IBD patients. Mechanistically, RUT potentiates NRF2 nuclear translocation to upregulate NRF2-mediated antioxidant response by directly inhibiting KEAP1-NRF2 interaction.
Assuntos
Colite , Fator 2 Relacionado a NF-E2 , Animais , Colite/induzido quimicamente , Colite/tratamento farmacológico , Colite/genética , Sulfato de Dextrana/toxicidade , Humanos , Peróxido de Hidrogênio , Alcaloides Indólicos , Proteína 1 Associada a ECH Semelhante a Kelch/genética , Proteína 1 Associada a ECH Semelhante a Kelch/metabolismo , Camundongos , Simulação de Acoplamento Molecular , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , QuinazolinasRESUMO
Hepatocellular carcinoma (HCC) is a leading cause of cancer deaths worldwide, and an association between altered bile acid (BA) metabolism, down-regulation of farnesoid X receptor (FXR), which is a master regulator of BA metabolism, and hepatocarcinogenesis has been documented. While global FXR deficiency in mice results in spontaneous HCC with aging, the contribution of tissue-specific FXR deficiency to hepatocarcinogenesis remains unclear. In this study, the prevalence of hepatic tumors, expression of genes related to tumorigenesis, and serum/liver BA levels were compared among male whole-body Fxr-null, hepatocyte-specific Fxr-null (Fxr ∆Hep), and enterocyte-specific Fxr-null (Fxr ∆IE) mice at the age of 3, 14, and 20 months. More than 90% of 20-month-old whole-body Fxr-null mice had hepatic tumors with enhanced hepatic expression of myelocytomatosis oncogene (Myc) and cyclin-dependent kinase 4 (Cdk4) messenger RNAs (mRNAs) and elevated serum taurocholate (TCA) and tauromuricholate (TMCA) and their respective unconjugated derivatives. The incidence of hepatic tumors was significantly lower in Fxr ∆Hep and Fxr ∆IE mice (20% and 5%, respectively), and the increases in Myc and Cdk4 mRNA or serum BA concentrations were not detected in these mice compared to Fxr floxed [fl]/fl mice; a similar tendency was observed in 14-month-old mice. However, increased hepatic c-Myc protein expression was found only in Fxr-null mice at the age of 3, 14, and 20 months. Treatment with TCA induced Myc expression in Fxr-null cultured primary mouse hepatocytes but not in wild-type (WT) mouse hepatocytes, demonstrating that the combination of hepatocyte FXR disruption with elevated TCA is required for Myc induction and ensuing age-dependent hepatocarcinogenesis in Fxr-null mice. Conclusion: There is a relatively low risk of hepatic tumors by inhibition of FXR in enterocytes, likely due to the lack of increased TCA and Myc induction.
RESUMO
Cholangiocarcinoma (CC) is a type of relatively rare neoplasm in adenocarcinoma. The characteristics of CCs as well as biliary epithelial cells are heterogeneous at the different portion of the biliary tree. There are two candidate stem/progenitor cells of the biliary tree, i.e., biliary tree stem/progenitor cell (BTSC) at the peribiliary gland (PBG) of large bile ducts and liver stem/progenitor cell (LPC) at the canals of Hering of peripheral small bile duct. Although previous reports suggest that intrahepatic CC (ICC) can arise from such stem/progenitor cells, the characteristic difference between BTSC and LPC in pathological process needs further investigation, and the etiology of CC remains poorly understood. Here we show that Sterile alpha motif domain containing 5 (SAMD5) is exclusively expressed in PBGs of large bile ducts in normal mice. Using a mouse model of cholestatic liver disease, we demonstrated that SAMD5 expression was upregulated in the large bile duct at the hepatic hilum, the extrahepatic bile duct and PBGs, but not in proliferating intrahepatic ductules, suggesting that SAMD5 is expressed in BTSC but not LPC. Intriguingly, human ICCs and extrahepatic CCs exhibited striking nuclear localization of SAMD5 while the normal hilar large bile duct displayed slight-to-moderate expression in cytoplasm. In vitro experiments using siRNA for SAMD5 revealed that SAMD5 expression was associated with the cell cycle regulation of CC cell lines. CONCLUSION: SAMD5 is a novel marker for PBG but not LPC in mice. In humans, the expression and location of SAMD5 could become a promising diagnostic marker for the cell type as well as malignancy of bile ducts and CCs.
Assuntos
Neoplasias do Sistema Biliar/metabolismo , Colangiocarcinoma/metabolismo , Motivo Estéril alfa , Animais , Neoplasias do Sistema Biliar/patologia , Núcleo Celular/metabolismo , Proliferação de Células , Colangiocarcinoma/patologia , Camundongos , Camundongos Endogâmicos C57BLRESUMO
Nonalcoholic fatty liver disease is becoming the most common chronic liver disease in Western countries, and limited therapeutic options are available. Here we uncovered a role for intestinal hypoxia-inducible factor (HIF) in hepatic steatosis. Human-intestine biopsies from individuals with or without obesity revealed that intestinal HIF-2α signaling was positively correlated with body-mass index and hepatic toxicity. The causality of this correlation was verified in mice with an intestine-specific disruption of Hif2a, in which high-fat-diet-induced hepatic steatosis and obesity were substantially lower as compared to control mice. PT2385, a HIF-2α-specific inhibitor, had preventive and therapeutic effects on metabolic disorders that were dependent on intestine HIF-2α. Intestine HIF-2α inhibition markedly reduced intestine and serum ceramide levels. Mechanistically, intestine HIF-2α regulates ceramide metabolism mainly from the salvage pathway, by positively regulating the expression of Neu3, the gene encoding neuraminidase 3. These results suggest that intestinal HIF-2α could be a viable target for hepatic steatosis therapy.
Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Mucosa Intestinal/metabolismo , Hepatopatia Gordurosa não Alcoólica/metabolismo , Obesidade/metabolismo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Ceramidas/metabolismo , Humanos , Camundongos , Camundongos Knockout , Neuraminidase/genética , Hepatopatia Gordurosa não Alcoólica/etiologia , Obesidade/complicações , Transdução de SinaisRESUMO
Flutamide (FLU), an oral, nonsteroidal antiandrogen drug used in the treatment of prostate cancer, is associated with idiosyncratic hepatotoxicity that sometimes causes severe liver damage, including cholestasis, jaundice, and liver necrosis. To understand the mechanism of toxicity, a combination of aryl hydrocarbon receptor (Ahr)-deficient (Ahr-/-) mice, primary hepatocytes, luciferase reporter gene assays, in silico ligand docking and ultra-performance chromatography-quadrupole time-of-flight mass spectrometry-based metabolomics was used. A significant increase of liver weights, and liver and serum bile acid levels was observed after FLU treatment, indicating hepatomegaly and disrupted bile acid homeostasis. Expression of the AhR gene battery was markedly increased in livers of wild-type mice Ahr+/+ treated with FLU, while no change was noted in Ahr-/- mice. Messenger RNAs encoded by AhR target genes were induced in primary mouse hepatocytes cultured with FLU, which confirmed the in vivo results. Ligand-docking analysis further predicted that FLU is an AhR agonist ligand which was confirmed by luciferase reporter gene assays. Multivariate data analysis showed that bile acids were responsible for the separation of vehicle- and FLU-treated Ahr+/+ mice, while there was no separation in Ahr-/- mice. Expression of mRNA encoding the bile acid transporter ABCC4 was increased and farnesoid X receptor signaling was inhibited in the livers of Ahr+/+ mice, but not in Ahr-/- mice treated with FLU, in agreement with the observed downstream metabolic alterations. These findings provide new insights into the mechanism of liver injury caused by FLU treatment involving activation of AhR and the alterations of bile acid homeostasis, which could guide clinical application.