RESUMO
Findings about chronic complex diseases are difficult to extrapolate from animal models to humans. We reason that organs may have core network modules that are preserved between species and are predictably altered when homeostasis is disrupted. To test this idea, we perturbed hepatic homeostasis in mice by dietary challenge and compared the liver transcriptome with that in human fatty liver disease and liver cancer. Co-expression module preservation analysis pointed to alterations in immune responses and metabolism (core modules) in both human and mouse datasets. The extent of derailment in core modules was predictive of survival in the cancer genome atlas (TCGA) liver cancer dataset. We identified module eigengene quantitative trait loci (module-eQTL) for these predictive co-expression modules, targeting of which may resolve homeostatic perturbations and improve patient outcomes. The framework presented can be used to understand homeostasis at systems levels in pre-clinical models and in humans. A record of this paper's transparent peer review process is included in the supplemental information.
Assuntos
Redes Reguladoras de Genes , Neoplasias Hepáticas , Animais , Redes Reguladoras de Genes/genética , Homeostase , Neoplasias Hepáticas/genética , Camundongos , Locos de Características Quantitativas/genéticaRESUMO
Lambda interferons (IFN-λs) are a major component of the innate immune defense to viruses, bacteria, and fungi. In human liver, IFN-λ not only drives antiviral responses, but also promotes inflammation and fibrosis in viral and non-viral diseases. Here we demonstrate that macrophages are primary responders to IFN-λ, uniquely positioned to bridge the gap between IFN-λ producing cells and lymphocyte populations that are not intrinsically responsive to IFN-λ. While CD14+ monocytes do not express the IFN-λ receptor, IFNLR1, sensitivity is quickly gained upon differentiation to macrophages in vitro. IFN-λ stimulates macrophage cytotoxicity and phagocytosis as well as the secretion of pro-inflammatory cytokines and interferon stimulated genes that mediate immune cell chemotaxis and effector functions. In particular, IFN-λ induced CCR5 and CXCR3 chemokines, stimulating T and NK cell migration, as well as subsequent NK cell cytotoxicity. Using immunofluorescence and cell sorting techniques, we confirmed that human liver macrophages expressing CD14 and CD68 are highly responsive to IFN-λ ex vivo. Together, these data highlight a novel role for macrophages in shaping IFN-λ dependent immune responses both directly through pro-inflammatory activity and indirectly by recruiting and activating IFN-λ unresponsive lymphocytes.
Assuntos
Interferons/imunologia , Macrófagos/imunologia , Degranulação Celular , Diferenciação Celular , Movimento Celular , Células Cultivadas , Hepatite C/imunologia , Humanos , Interferons/genética , Células Matadoras Naturais/imunologia , Fígado/imunologia , Monócitos/imunologia , FagocitoseRESUMO
Adiponectin demonstrates beneficial effects in various metabolic diseases, including diabetes, and in bowel cancer. Recent data also suggest a protective role in colitis. However, the precise molecular mechanisms by which adiponectin and its receptors modulate colitis and the nature of the adaptive immune response in murine models are yet to be elucidated. Adiponectin knock-out mice were orally administered dextran sulfate sodium for 7 days and were compared with wild-type mice. The severity of disease was analyzed histopathologically and through cytokine profiling. HCT116 colonic epithelial cells were employed to analyze the in vitro effects of adiponectin and AdipoR1 interactions in colonic injury following dextran sulfate sodium treatment. Adiponectin knock-out mice receiving dextran sulfate sodium exhibited severe colitis, had greater inflammatory cell infiltration, and an increased presence of activated B cells compared with controls. This was accompanied by an exaggerated proinflammatory cytokine profile and increased STAT3 signaling. Adiponectin knock-out mouse colons had markedly reduced proliferation and increased epithelial apoptosis and cellular stress. In vitro, adiponectin reduced apoptotic, anti-proliferative, and stress signals and restored STAT3 signaling. Following the abrogation of AdipoR1 in vitro, these protective effects of adiponectin were abolished. In summary, adiponectin maintains intestinal homeostasis and protects against murine colitis through interactions with its receptor AdipoR1 and by modulating adaptive immunity.
Assuntos
Adiponectina/metabolismo , Linfócitos B/imunologia , Colite/metabolismo , Receptores de Adiponectina/metabolismo , Fator de Transcrição STAT3/metabolismo , Doença Aguda , Adiponectina/genética , Animais , Apoptose , Proliferação de Células , Colite/prevenção & controle , Citocinas/metabolismo , Sulfato de Dextrana , Modelos Animais de Doenças , Células HCT116 , Homeostase , Humanos , Sistema Imunitário , Inflamação , Mucosa Intestinal/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fator de Necrose Tumoral alfa/metabolismoRESUMO
UNLABELLED: Adiponectin protects against liver fibrosis, but the mechanisms have not been fully elucidated. Here, we showed that adiponectin upregulated inducible nitric oxide synthase (iNOS) messenger RNA (mRNA) and protein expression in hepatic non-parenchymal cells, particularly in hepatic stellate cells (HSCs), and increased nitric oxide (NO2-/NO3-) concentration in HSC-conditioned medium. Adiponectin attenuated HSC proliferation and migration but promoted apoptosis in a NO-dependent manner. More advanced liver fibrosis with decreased iNOS/NO levels was observed in adiponectin knockout mice comparing to wide-type mice when administered with CCI4 while NO donor supplementation rescued the phenotype. Further experiments demonstrated that adiponectin-induced iNOS/NO system activation is mediated through adipoR2-AMPK-JNK/Erk1/2-NF-κB signaling. These data suggest that adiponectin inhibits HSC function, further limiting the development of liver fibrosis at least in part through adiponectin-induced NO release. Therefore, adiponectin-mediated NO signaling may be a novel target for the treatment of liver fibrosis. KEY MESSAGES: ⢠Adiponectin activates HSC iNOS/NO and SEC eNOS/NO systems. ⢠Adiponectin inhibits HSC proliferation and migration but promotes its apoptosis. ⢠Adiponectin inhibits CCL4-induced liver fibrosis by modulation of liver iNOS/NO. ⢠Adiponectin stimulates HSC iNOS/NO via adipoR2-AMPK-JNK/ErK1/2-NF-κB pathway.
Assuntos
Adiponectina/genética , Adiponectina/metabolismo , Células Estreladas do Fígado/metabolismo , Cirrose Hepática/genética , Cirrose Hepática/metabolismo , Óxido Nítrico/metabolismo , Adiponectina/farmacologia , Animais , Apoptose/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Modelos Animais de Doenças , Regulação da Expressão Gênica/efeitos dos fármacos , Células Estreladas do Fígado/efeitos dos fármacos , Cirrose Hepática/induzido quimicamente , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Camundongos , Camundongos Knockout , Óxido Nítrico Sintase Tipo II/genética , Óxido Nítrico Sintase Tipo II/metabolismo , Ratos , Receptores de Adiponectina/metabolismo , Fator de Transcrição RelA/metabolismoRESUMO
Non-alcoholic fatty liver disease and non-alcoholic steatohepatitis (NASH) are increasing clinical problems for which effective treatments are required. The polyphenol resveratrol prevents the development of fatty liver disease in a number of experimental studies. We hypothesized that it could revert steatohepatitis, including hepatic inflammation and fibrosis, in an experimental NASH model. To induce hepatic steatohepatitis, a 65% fat, 2% cholesterol and 0.5% cholate (HFC) diet was fed to rats for 1 or 16 weeks, prior to treatment. Subsequently, the diet was supplemented with resveratrol (approx. 100mg/rat/day) to three intervention groups; week 2-4, 2-7 or 17-22. Treated animals were sacrificed at the end of each intervention period with appropriate control and HFC diet controls. Blood and liver were harvested for analysis. When commenced early, resveratrol treatment partially mitigated transaminase elevations, hepatic enlargement and TNFα induced protein-3 protein expression, but generally resveratrol treatment had no effect on elevated hepatic triglyceride levels, histological steatohepatitis or fibrosis. We observed a slight reduction in Collagen1α1 mRNA expression and no reduction in the mRNA expression of other markers of fibrosis, inflammation or steatosis (TGFß, TNFα, α2-MG, or SREBP-1c). Resveratrol metabolites were detected in serum, including trans-resveratrol-3-O-sulphate/trans-resveratrol-4'-O-sulphate (mean concentration 7.9 µg/ml). Contrary to the findings in experimental steatosis, resveratrol treatment had no consistent therapeutic effect in alleviating manifest experimental steatohepatitis.
Assuntos
Antioxidantes/uso terapêutico , Fígado/efeitos dos fármacos , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Estilbenos/uso terapêutico , Animais , Antioxidantes/administração & dosagem , Antioxidantes/metabolismo , Modelos Animais de Doenças , Feminino , Fígado/metabolismo , Fígado/patologia , Testes de Função Hepática , Hepatopatia Gordurosa não Alcoólica/metabolismo , Hepatopatia Gordurosa não Alcoólica/patologia , Tamanho do Órgão/efeitos dos fármacos , Ratos Wistar , Resveratrol , Estilbenos/administração & dosagem , Estilbenos/metabolismo , Resultado do Tratamento , Triglicerídeos/metabolismoRESUMO
Hepatic stellate cells (HSC) are central players in liver fibrosis that when activated, proliferate, migrate to sites of liver injury, and secrete extracellular matrix. Obesity, a known risk factor for liver fibrosis is associated with reduced levels of adiponectin, a protein that inhibits liver fibrosis in vivo and limits HSC proliferation and migration in vitro. Adiponectin-mediated activation of adenosine monophosphate-activated kinase (AMPK) inhibits HSC proliferation, but the mechanism by which it limits HSC migration to sites of injury is unknown. Here we sought to elucidate how adiponectin regulates HSC motility. Primary rat HSCs were isolated and treated with adiponectin in migration assays. The in vivo actions of adiponectin were examined by treating mice with carbon tetrachloride for 12 weeks and then injecting them with adiponectin. Cell and tissue samples were collected and analyzed for gene expression, signaling, and histology. Serum from patients with liver fibrosis was examined for adiponectin and tissue inhibitor of metalloproteinase-1 (TIMP-1) protein. Adiponectin administration into mice increased TIMP-1 gene and protein expression. In cultured HSCs, adiponectin promoted TIMP-1 expression and through binding of TIMP-1 to the CD63/ß1-integrin complex reduced phosphorylation of focal adhesion kinase to limit HSC migration. In mice with liver fibrosis, adiponectin had similar effects and limited focal adhesion kinase phosphorylation. Finally, in patients with advanced fibrosis, there was a positive correlation between serum adiponectin and TIMP-1 levels. In sum, these data show that adiponectin stimulates TIMP-1 secretion by HSCs to retard their migration and contributes to the anti-fibrotic effects of adiponectin.
Assuntos
Adiponectina/farmacologia , Movimento Celular/efeitos dos fármacos , Células Estreladas do Fígado/efeitos dos fármacos , Inibidor Tecidual de Metaloproteinase-1/metabolismo , Adiponectina/sangue , Animais , Apoptose/efeitos dos fármacos , Western Blotting , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Proteína-Tirosina Quinases de Adesão Focal/metabolismo , Expressão Gênica/efeitos dos fármacos , Células Estreladas do Fígado/citologia , Células Estreladas do Fígado/metabolismo , Humanos , Integrina beta1/genética , Integrina beta1/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Microscopia Confocal , Hepatopatia Gordurosa não Alcoólica/sangue , Fosforilação/efeitos dos fármacos , Ligação Proteica/efeitos dos fármacos , Interferência de RNA , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Tetraspanina 30/genética , Tetraspanina 30/metabolismo , Inibidor Tecidual de Metaloproteinase-1/sangue , Inibidor Tecidual de Metaloproteinase-1/genéticaRESUMO
UNLABELLED: Advanced liver fibrosis in nonalcoholic steatohepatitis (NASH) is often accompanied by a reduction in hepatic fat to the point of complete fat loss (burnt-out NASH), but the mechanisms behind this phenomenon have not been elucidated. Adiponectin is raised in cirrhosis of any cause and has potent antisteatotic activity. In this study we examined 65 patients with advanced biopsy-proven NASH (fibrosis stage 3-4) and 54 with mild disease (fibrosis stage 0-1) to determine if disappearance of steatosis correlated with changes in serum adiponectin. All patents had fasting blood tests and anthropometric measures at the time of liver biopsy. Liver fat was accurately quantitated by morphometry. Serum adiponectin was measured by immunoassay. When compared to those with early disease, patients with advanced NASH were more insulin-resistant, viscerally obese, and older, but there was no difference in liver fat content or adiponectin levels. Adiponectin had a significant negative correlation with liver fat percentage in the whole cohort (r = -0.28, P < 0.01), driven by patients with advanced NASH (r = -0.40, P < 0.01). In advanced NASH, for each 4 µg/L increase in adiponectin there was an odds ratio OR of 2.0 (95% confidence interval [CI]: 1.3-3.0, P < 0.01) for a 5% reduction in hepatic fat. Adiponectin was highly and significantly associated with almost complete hepatic fat loss or burnt-out NASH (12.1 versus 7.4 µg/L, P = 0.001) on multivariate analysis. A relationship between adiponectin, bile acids, and adipocyte fexaramine activation was demonstrated in vivo and in vitro, suggestive of hepatocyte-adipocyte crosstalk. CONCLUSION: Serum adiponectin levels in advanced NASH are independently associated with hepatic fat loss. Adiponectin may in part be responsible for the paradox of burnt-out NASH. (HEPATOLOGY 2012).
Assuntos
Adiponectina/sangue , Fígado Gorduroso/sangue , Metabolismo dos Lipídeos , Fígado/metabolismo , Fígado/patologia , Adulto , Idoso , Ácidos e Sais Biliares/sangue , Biópsia , Estudos Transversais , Feminino , Fibrose , Humanos , Masculino , Pessoa de Meia-Idade , Hepatopatia Gordurosa não AlcoólicaRESUMO
BACKGROUND: The mechanisms whereby patients with obesity/overweight are more susceptible to alcohol-associated liver fibrosis are unclear. Leptin, a peptide hormone secreted by white adipose tissue is increased in association with overweight/obesity and is recognized as mediator of liver fibrosis. We sought to assess whether leptin contributes to alcoholic liver fibrosis by in vitro studies in hepatic stellate cells (HSC). METHODS: Rat HSCs in second or third passage were utilised. Leptin, Acetaldehyde or combination with leptin and acetaldehyde were incubated for specific periods in cultured HSCs. Profibrogenic gene and protein expression were determined and associated-signalling pathways were assessed. Interleukin 6 (IL-6) antibody neutralization was used to evaluate the role of IL-6. RESULTS: Leptin did not promote acetaldehyde-induced gene expression of collagen I, transforming growth factor ß1 (TGFß1) and tissue inhibitor of metalloproteinase 1 (TIMP1) in vitro. However, combined treatment of leptin with acetaldehyde synergistically enhanced the protein expression of smooth muscle actin (αSMA), an activation marker of HSCs, and of Interleukin-6 (IL-6). The combination of leptin and acetaldehyde also augmented MAPK/p38 and MAPK/ERK1/2 phosphoprotein expression. IL-6 neutralization down-regulated protein expression of pp38, pERK1/2 and αSMA, while exogenous rat recombinant IL-6 administration up-regulated αSMA. Similarly, MAPK/p38 and MAPK/ERK1/2 inhibition attenuated αSMA expression. H(2)O(2) induction by acetaldehyde was not potentiated by co-treatment with leptin nor did IL-6 neutralization reduce acetaldehyde-induced H(2)O(2) production. CONCLUSIONS: We conclude that leptin potentiates acetaldehyde-induced HSC activation and αSMA expression by an IL-6-dependent mechanism.