RESUMO
BACKGROUND & AIMS: Patients with alcoholic liver disease (ALD) have vitamin A (VA) deficiency and an enhanced immune response associated with disease severity. All-trans retinoic acid (ATRA), a VA-active metabolite, has anti-inflammatory effects and its deficiency could contribute to the exacerbated proinflammatory reaction. The aim of this study was to investigate the effects of ATRA/VA deficiency and supplementation on the monocyte response in ALD. METHODS: Vitamin A and ATRA plasma levels were quantified in ALD patients and healthy subjects (HS). The in vitro effect of ATRA on lipopolysaccharide (LPS)-induced TNF-α production by human peripheral blood mononuclear cells (PBMC) was assessed by ELISA and RT-PCR. The activation pattern of peritoneal macrophages (PerMΦ) and circulating monocytes isolated from VA-deficient mice and ALD patients, respectively, was evaluated by flow cytometry, quantification of TNF-α and NO2 production. RESULTS: Alcoholic liver disease patients (n = 85) showed plasmatic VA deficiency that was correlated with scores of severity and with the hepatic venous pressure gradient. ATRA levels correlated significantly with VA levels. In vitro, ATRA pretreatment decreased the overproduction of TNF-α by LPS-stimulated PBMC of ALD patients. In vivo, VA deficiency in mice was associated with increased activation of PerMΦ, while oral ATRA supplementation normalized it. CONCLUSION: For the first time, we show that VA/ATRA deficiencies in ALD patients are associated with disease severity. Furthermore, our data strongly suggest that the VA deficiency observed in ALD patients might participate in the pathophysiology of the disease by priming immune cells, and that ATRA supplementation could downregulate the deleterious proinflammatory state in cirrhosis and might thus be of therapeutic use.
Assuntos
Cirrose Hepática Alcoólica/imunologia , Monócitos/imunologia , Tretinoína/sangue , Tretinoína/farmacologia , Fator de Necrose Tumoral alfa/metabolismo , Deficiência de Vitamina A/complicações , Adulto , Idoso , Animais , Estudos de Casos e Controles , Células Cultivadas , Modelos Animais de Doenças , Regulação para Baixo , Feminino , Humanos , Lipopolissacarídeos/farmacologia , Ativação de Macrófagos/imunologia , Macrófagos Peritoneais/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , RNA Mensageiro/genética , Vitamina A/sangueRESUMO
BACKGROUND & AIMS: Vitamin D deficiency has been frequently reported in advanced liver disease. However, its influence on alcoholic liver disease (ALD) has been poorly elucidated. We investigated the association of vitamin D with clinical, biological, and histological parameters and survival in ALD patients. Furthermore, we explored the effect of vitamin D treatment on ALD patient peripheral blood mononuclear cells (PBMCs), and in a murine experimental model of ALD. METHODS: Serum levels of 25-hydroxyvitamin D [25(OH)D] were determined in 324 Caucasian ALD patients and 201 healthy controls. In vitro experiments on vitamin D pre-treated PBMCs evaluated TNFα production by ELISA in culture supernatants. Mice were submitted to an ethanol-fed diet and some of them were orally supplemented three times per week with 1,25(OH)2D. RESULTS: Severe deficiency in 25(OH)D (<10 ng/ml) was significantly associated with higher aspartate aminotransferase levels (p=1.00 × 10(-3)), increased hepatic venous pressure gradient (p=5.80 × 10(-6)), MELD (p=2.50 × 10(-4)), and Child-Pugh scores (p=8.50 × 10(-7)). Furthermore, in multivariable analysis, a low 25(OH)D concentration was associated with cirrhosis (OR=2.13, 95% CI=1.18-3.84, p=0.013) and mortality (HR=4.33, 95% CI=1.47-12.78, p=7.94 × 10(-3)) at one year. In addition, in vitro, 1,25(OH)2D pretreatment decreased TNFα production by stimulated PBMCs of ALD patients (p=3.00 × 10(-3)), while in vivo, it decreased hepatic TNFα expression in ethanol-fed mice (p=0.04). CONCLUSIONS: Low 25(OH)D levels are associated with increased liver damage and mortality in ALD. Our results suggest that vitamin D might be both a biomarker of severity and a potential therapeutic target in ALD.
Assuntos
Hepatopatias Alcoólicas/complicações , Deficiência de Vitamina D/complicações , Vitamina D/análogos & derivados , Adulto , Animais , Biomarcadores/sangue , Estudos de Casos e Controles , Modelos Animais de Doenças , Feminino , Humanos , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/fisiologia , Cirrose Hepática Alcoólica/complicações , Cirrose Hepática Alcoólica/patologia , Cirrose Hepática Alcoólica/fisiopatologia , Hepatopatias Alcoólicas/patologia , Hepatopatias Alcoólicas/fisiopatologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , Prognóstico , Estudos Prospectivos , Fatores de Risco , Fator de Necrose Tumoral alfa/biossíntese , Vitamina D/sangue , Vitamina D/farmacologia , Deficiência de Vitamina D/sangueRESUMO
Acute pancreatitis (AP) is an inflammatory disease in which the regulatory pathways are not clearly elucidated. Activation of interleukin 1ß (IL-1ß) and immunomodulation via MyD88, the first signaling molecule in the ST2 pathway, seem to be involved. Because IL-33, the ST2 ligand, is an IL-1 family member and acts as an alarmin, we explored the ST2 pathway in human and mouse AP. Soluble ST2 was assayed by enzyme-linked immunosorbent assay (ELISA) in plasma of 44 patients admitted for AP. The levels of soluble ST2 increased early during AP and correlated with parameters of severity. Under two different experimental models of AP (ie, choline-deficient-ethionine-supplemented diet and cerulein injections), ST2-deficient mice (Il1rl1(-/-)) presented with more severe disease than wild-type mice, with increased activation of mast cells. In vitro, Il1rl1(-/-) bone-marrow-derived mast cells exhibited exacerbated degranulation, compared with the wild type. Flow cytometry identified mast cells as the main peritoneal population expressing ST2. Using immunohistochemistry and ELISA, we showed constitutive expression of IL-33 in murine pancreas and its release during experimental AP. Correlated with AP severity, increased soluble ST2 levels evoke involvement of the ST2 pathway in human AP. Furthermore, our experimental data suggest a protective role for ST2 during AP, highlighting the potential regulatory role of mast cells and the possibility of the ST2 pathway as a new therapeutic target in AP.
Assuntos
Pancreatite/metabolismo , Receptores de Superfície Celular/sangue , Doença Aguda , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Degranulação Celular/fisiologia , Feminino , Humanos , Proteína 1 Semelhante a Receptor de Interleucina-1 , Interleucina-33 , Interleucinas/metabolismo , Masculino , Mastócitos/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Pessoa de Meia-Idade , Pâncreas/metabolismo , Pancreatite/patologia , Cavidade Peritoneal/citologia , Receptores de Superfície Celular/fisiologia , Receptores de Interleucina/deficiência , Receptores de Interleucina/fisiologia , Índice de Gravidade de Doença , Transdução de Sinais/fisiologia , Adulto JovemRESUMO
BACKGROUND & AIMS: In decompensated cirrhosis, the early innate immune response to the Toll-like receptor 4 (TLR4) agonist, lipopolysaccharides (LPS), is characterized by a hyper-production of pro-inflammatory cytokines and hypo-production of the anti-inflammatory cytokine IL-10. In LPS-stimulated non-cirrhotic immune cells, the constitutively active glycogen synthase kinase (GSK) 3 favors pro- vs. anti-inflammatory cytokines, by acting on gene induction. However, in these cells, TLR4 dampens its own pro-inflammatory response by inducing early (within minutes) AKT-mediated phosphorylation of GSK3ß (one of two GSK3 isoforms) on Ser9. Phosphorylation of GSK3ß (Ser9) inhibits its activity, decreases pro-inflammatory cytokines, and increases IL-10. Thus, we investigated the role of GSK3 in LPS-induced cytokine production by peripheral blood mononuclear cells (PBMCs) or monocytes from patients with advanced cirrhosis and normal subjects. METHODS: Cells were pre-incubated with or without GSK3 inhibitor (SB216763 or lithium chloride) for 1h and then stimulated with LPS. Cytokine production was assessed at mRNA and secreted proteins levels, by real-time RT-PCR at 1h and ELISA at 20 h, respectively. GSK3ß phosphorylation was assessed using Western blotting. RESULTS: In cirrhotic and normal PBMCs pretreated with GSK3 inhibitors, LPS-induced production of pro-inflammatory proteins TNF-α and IL-12p40 was significantly decreased while that of IL-10 was increased. LPS-induced, AKT-mediated phosphorylation of GSK3ß on Ser9 found in normal monocytes, was abolished in cirrhotic cells. CONCLUSIONS: GSK3 is involved in the early TLR4-mediated pro-inflammatory response in patients with decompensated cirrhosis. This was associated with a defect in AKT-mediated GSK3ß phosphorylation resulting in unrestricted 'pro-inflammatory' activity of the enzyme.
Assuntos
Quinase 3 da Glicogênio Sintase/imunologia , Quinase 3 da Glicogênio Sintase/metabolismo , Hepatite/imunologia , Cirrose Hepática/imunologia , Transdução de Sinais/imunologia , Adjuvantes Imunológicos/farmacologia , Adulto , Idoso , Células Cultivadas , Feminino , Regulação Enzimológica da Expressão Gênica/imunologia , Quinase 3 da Glicogênio Sintase/antagonistas & inibidores , Quinase 3 da Glicogênio Sintase/genética , Glicogênio Sintase Quinase 3 beta , Hepatite/metabolismo , Humanos , Indóis/farmacologia , Interleucina-10/imunologia , Interleucina-10/metabolismo , Interleucina-12/imunologia , Interleucina-12/metabolismo , Lipopolissacarídeos/farmacologia , Cloreto de Lítio/farmacologia , Cirrose Hepática/metabolismo , Masculino , Maleimidas/farmacologia , Pessoa de Meia-Idade , Monócitos/imunologia , Monócitos/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Índice de Gravidade de Doença , Receptor 4 Toll-Like/imunologia , Receptor 4 Toll-Like/metabolismoRESUMO
BACKGROUND & AIMS: A recent genome-wide association study identified genetic polymorphism (rs738409 C>G) in the PNPLA3/adiponutrin gene associated with liver steatosis. This variant has also been linked to increased risk of alcoholic liver disease (ALD) and cirrhosis in Mestizo Mexicans with excessive alcohol intake. Our aim was to study the influence of this polymorphism on European Caucasian patients with histologically suggestive ALD. METHODS: Three-hundred-and-twenty-eight healthy controls and 330 ALD patients, among whom 265 had cirrhosis, were genotyped for the rs738409 polymorphism. We studied the impact of rs738409 on clinical and biological parameters, together with histological staging of steatosis and fibrosis. PNPLA3 messenger RNA (mRNA) levels were measured by quantitative real-time PCR according to the patient's phenotype. RESULTS: The G-allele was significantly more frequent in ALD patients than in controls (odds ratio [OR] = 1.54, 95% confidence interval [CI] = 1.12-2.11 p = 0.008) and was, among ALD patients, significantly associated with steatosis (p = 0.048), fibrosis (p = 0.001), and greater risk of cirrhosis (p = 0.001). In multivariate analysis, rs738409 remained the strongest independent factor associated with risk of cirrhosis (OR = 2.08; 95% CI = 1.15-3.77; p = 0.02). Furthermore, the PNPLA3 mRNA liver expression level was significantly lower in patients with more advanced fibrosis (p = 0.03) and negatively correlated with the hepatic venous pressure gradient (r = -0.41, p = 0.006). CONCLUSIONS: In European Caucasians, the rs738409 variant is associated with increased risk of ALD, liver damage, and cirrhosis. Further prospective studies are required to confirm these results and to evaluate the potential of PNPLA3 as both a predictor and a therapeutic target in ALD.