RESUMO
BACKGROUND: One mechanism by which alcoholic liver disease (ALD) progresses is oxidative stress and the generation of reactive oxygen species, among others due to the induction of cytochrome P-4502E1 (CYP2E1). Experimental data underline the key role of CYP2E1 because ALD could be partially prevented in rats by the administration of the specific CYP2E1 inhibitor chlormethiazole. As CYP2E1 is linked to the formation of carcinogenic etheno DNA adducts in ALD patients, a causal role of alcohol-induced CYP2E1 in hepatocarcinogenesis is implicated. The purpose of this study was to investigate CYP2E1 induction in ALD, and its correlation with oxidative DNA lesions and with hepatic histology. METHODS: Hepatic biopsies from 97 patients diagnosed with ALD were histologically scored for steatosis, inflammation, and fibrosis. CYP2E1 and the exocyclic etheno DNA adduct 1,N6 -etheno-2'deoxyadenosine (εdA) were determined immunohistochemically. In addition, in 42 patients, 8-hydroxydeoxyguanosine (8-OHdG) was also evaluated using immunohistochemistry. RESULTS: A significant positive correlation was found between CYP2E1 and εdA (p < 0.0001) as well as between CYP2E1 and 8-OHdG (p = 0.039). Both CYP2E1 (p = 0.0094) and ÉdA (p < 0.0001) also correlated significantly with the stage of hepatic fibrosis. Furthermore, a significant correlation between the fibrosis stage and the grade of lobular inflammation (p < 0.0001) was observed. However, the amount of alcohol consumed did not correlate with any of the parameters determined. CONCLUSIONS: These data suggest an important role of CYP2E1 in the generation of εdA, in the fibrotic progression of ALD, and thus in alcohol-mediated hepatocarcinogenesis. CYP2E1 may be a target in the treatment of ALD and a potential prognostic marker for disease progression.
Assuntos
Carcinogênese , Citocromo P-450 CYP2E1/metabolismo , Desoxiadenosinas/metabolismo , Hepatopatias Alcoólicas/metabolismo , Fígado/metabolismo , 8-Hidroxi-2'-Desoxiguanosina , Carcinoma Hepatocelular , Desoxiguanosina/análogos & derivados , Desoxiguanosina/metabolismo , Feminino , Fibrose , Humanos , Imuno-Histoquímica , Inflamação/metabolismo , Inflamação/patologia , Gordura Intra-Abdominal/metabolismo , Gordura Intra-Abdominal/patologia , Fígado/patologia , Cirrose Hepática Alcoólica/metabolismo , Cirrose Hepática Alcoólica/patologia , Hepatopatias Alcoólicas/patologia , Neoplasias Hepáticas , Masculino , Pessoa de Meia-IdadeRESUMO
The peptide hormone hepcidin regulates mammalian iron homeostasis by blocking ferroportin-mediated iron export from macrophages and the duodenum. During inflammation, hepcidin is strongly induced by interleukin 6, eventually leading to the anemia of chronic disease. Here we show that hepatoma cells and primary hepatocytes strongly up-regulate hepcidin when exposed to low concentrations of H(2)O(2) (0.3-6 µM), concentrations that are comparable with levels of H(2)O(2) released by inflammatory cells. In contrast, bolus treatment of H(2)O(2) has no effect at low concentrations and even suppresses hepcidin at concentrations of >50 µM. H(2)O(2) treatment synergistically stimulates hepcidin promoter activity in combination with recombinant interleukin-6 or bone morphogenetic protein-6 and in a manner that requires a functional STAT3-responsive element. The H(2)O(2)-mediated hepcidin induction requires STAT3 phosphorylation and is effectively blocked by siRNA-mediated STAT3 silencing, overexpression of SOCS3 (suppressor of cytokine signaling 3), and antioxidants such as N-acetylcysteine. Glycoprotein 130 (gp130) is required for H(2)O(2) responsiveness, and Janus kinase 1 (JAK1) is required for adequate basal signaling, whereas Janus kinase 2 (JAK2) is dispensable upstream of STAT3. Importantly, hepcidin levels are also increased by intracellular H(2)O(2) released from the respiratory chain in the presence of rotenone or antimycin A. Our results suggest a novel mechanism of hepcidin regulation by nanomolar levels of sustained H(2)O(2). Thus, similar to cytokines, H(2)O(2) provides an important regulatory link between inflammation and iron metabolism.
Assuntos
Peptídeos Catiônicos Antimicrobianos/metabolismo , Peróxido de Hidrogênio/farmacologia , Mediadores da Inflamação/farmacologia , Fator de Transcrição STAT3/metabolismo , Regulação para Cima , Acetilcisteína/farmacologia , Peptídeos Catiônicos Antimicrobianos/genética , Sítios de Ligação , Proteína Morfogenética Óssea 6/fisiologia , Linhagem Celular Tumoral , Sequestradores de Radicais Livres/farmacologia , Hepcidinas , Humanos , Interleucina-6/fisiologia , Fosforilação , Regiões Promotoras Genéticas , Processamento de Proteína Pós-Traducional , Transdução de Sinais , Transcrição GênicaRESUMO
BACKGROUND: Enhanced drug elimination in alcoholics remains largely indefinable. In contrast, the reduced elimination of drugs in patients with advanced alcoholic liver disease (ALD) is normally owing to hepatic end-stage disease such as cirrhosis. We here study the mRNA expression of various hepatic drug metabolizing enzymes and transporters in association with liver stiffness (LS) being a novel noninvasive parameter for the assessment of cirrhosis to unravel the dynamic relationship between ALD and determinants of pharmacokinetics such as drug metabolizing enzymes and transporters. METHODS: We quantified mRNA expression levels of various cytochrome P-450 isoenzymes (CYPs) and drug transporters in 26 liver specimens of chronic alcoholics and 5 controls by quantitative polymerase chain reaction. In addition, liver histology, clinical data, and LS evaluated by transient elastography (Fibroscan) were obtained. RESULTS: Eighteen patients had a normal or moderate LS < 8 kPa (69.2%), while in the remaining 8 patients (30.7%) advanced F3 or F4 fibrosis could be established with an LS > 8 kPa. Overall, CYP3A4, CYP2E1, and solute carrier organic anion transporter 1B1 (SLCO1B1) were negatively correlated with increasing LS. CYPs and drug transporters tended to be up-regulated in alcoholics without advanced fibrosis (LS < 8.0 kPa) compared to healthy controls supporting data of boosted drug elimination in alcoholics without advanced ALD. However, in alcoholics with severely increased LS (>8 kPa), expression levels of CYP2E1, SLC22A2, and SLCO1B1 were significantly lower. CONCLUSIONS: In conclusion, CYPs and drug transporters seem to be induced in chronic alcoholics without irreversible liver damage but decline in case of manifest cirrhosis. Our study also suggests that noninvasive measurements of LS could be useful for pharmacokinetic predictions and individualized pharmacotherapy.
Assuntos
Sistema Enzimático do Citocromo P-450/farmacocinética , Cirrose Hepática/metabolismo , Hepatopatias Alcoólicas/metabolismo , Fígado/metabolismo , Transportadores de Ânions Orgânicos/farmacocinética , Adulto , Sistema Enzimático do Citocromo P-450/biossíntese , Sistema Enzimático do Citocromo P-450/genética , Técnicas de Imagem por Elasticidade/métodos , Feminino , Regulação da Expressão Gênica , Humanos , Fígado/patologia , Cirrose Hepática/genética , Cirrose Hepática/patologia , Hepatopatias Alcoólicas/genética , Hepatopatias Alcoólicas/patologia , Masculino , Pessoa de Meia-Idade , Transportadores de Ânions Orgânicos/biossíntese , Transportadores de Ânions Orgânicos/genética , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , RNA Mensageiro/farmacocinéticaRESUMO
Transient elastography (TE, Fibroscan) has been established as a noninvasive assessment tool of liver fibrosis. We evaluated potentials and limitations of TE for identifying renal allograft fibrosis. The technical possibility of kidney examination by TE was assessed in two 10-week-old German landrace pigs and kidney stiffness (KS) was evaluated in 164 renal transplant patients. KS could be determined in all animals at the pole and pars media (29 ± 10 kPa vs. 31 ± 17 kPa). In human renal allografts KS was successfully performed in 94.5% of the test series with reliable results in 72% of the measurements. Mean KS at the pole or pars media were comparable (35.0 ± 19.9 kPa vs. 33.2 ± 18.6 kPa). Significantly higher KS was detected in renal allografts with histologically confirmed advanced fibrosis. Body-mass-index, skin-allograft distance, and peri or intrarenal fluid accumulation were important confounders of successful KS measurements (BMI: r = -0.31; P < 0.001; distance: r = -0.50; P < 0.001). Notably, KS did not correlate with renal function. TE represents a noninvasive approach in selected transplant recipients to identify allografts with severe fibrosis. The heterogeneous kidney morphology and several other confounding factors negatively affect measurability of KS by TE. Further technical modifications are required to improve applicability of TE for kidney assessment.
Assuntos
Técnicas de Imagem por Elasticidade/métodos , Transplante de Rim , Rim/patologia , Adulto , Animais , Creatinina/sangue , Técnicas de Imagem por Elasticidade/estatística & dados numéricos , Feminino , Fibrose , Humanos , Rim/diagnóstico por imagem , Masculino , Pessoa de Meia-Idade , Variações Dependentes do Observador , Sus scrofa , Transplante HomólogoRESUMO
AIMS: Results of several animal studies suggest that similar to humans, female rodents are more susceptible to chronic alcohol-induced liver disease (ALD). The aim of the present study was to determine whether female mice are more susceptible to acute alcohol-induced steatosis than male mice and to investigate possible mechanisms involved. METHODS: Male and female C57BL/6J mice received one single dose of ethanol (6 g/kg bodyweight) or isocaloric maltose-dextrin solution intragastrically. Plasma alcohol concentration, markers of hepatic steatosis, activation of the TLR-4 signaling cascade and triglyceride export as well as lipid peroxidation and of iron metabolism were measured 12 h after acute alcohol intake. RESULTS: In male and female ethanol-treated mice, plasma alcohol concentrations were still markedly increased 12 h after the alcohol challenge, which was associated with a significant accumulation of lipids in the liver and increase of transaminases in plasma; however, lipid accumulation was â¼3-fold higher in females in comparison with male animals. Expression of MyD88 was only found to be significantly induced in livers of female alcohol-exposed mice, whereas protein levels of ApoB were found to be significantly lower only in livers of female mice exposed to ethanol. Levels of 4-HNE protein adducts and ferritin were induced in livers of male and female ethanol-treated mice. CONCLUSION: Taken together, these data suggest that female mice are also more susceptible to acute alcohol-induced liver steatosis and that this involves an increased activation of TLR-4-dependent signaling pathways in the liver.
Assuntos
Fígado Gorduroso Alcoólico/patologia , Alanina Transaminase/metabolismo , Aldeídos/metabolismo , Animais , Aspartato Aminotransferases/metabolismo , Proteínas de Transporte/metabolismo , Depressores do Sistema Nervoso Central/sangue , Etanol/sangue , Feminino , Absorção Intestinal/fisiologia , Ferro/metabolismo , Células de Kupffer/metabolismo , Metabolismo dos Lipídeos/efeitos dos fármacos , Testes de Função Hepática , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Ocludina/metabolismo , Permeabilidade , RNA/biossíntese , RNA/isolamento & purificação , Reação em Cadeia da Polimerase em Tempo Real , Caracteres Sexuais , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Chronic alcohol consumption is a major risk factor for esophageal cancer. Various mechanisms may mediate carcinogenesis including the genotoxic effect of acetaldehyde and oxidative stress. Ethanol exerts its carcinogenic effect in the liver among others via the induction of cytochrome P450 2E1 (CYP2E1) and the generation of carcinogenic etheno-DNA adducts. Here we investigated if such effects can also be observed in the human esophagus. We studied nontumorous esophageal biopsies of 37 patients with upper aerodigestive tract cancer and alcohol consumption of 102.3 ± 131.4 g/day (range: 15-600 g) as well as 16 controls without tumors (12 teetotalers and 4 subjects with a maximum of 25 g ethanol/day). CYP2E1, etheno-DNA adducts and Ki67 as a marker for cell proliferation were determined immunohistologically. Chronic alcohol ingestion resulted in a significant induction of CYP2E1 (p = 0.015) which correlated with the amount of alcohol consumed (r = 0.6, p < 0.001). Furthermore, a significant correlation between CYP2E1 and the generation of the carcinogenic exocyclic etheno-DNA adducts 1,N(6)-ethenodeoxyadenosine (r = 0.93, p < 0.001) and 3,N(4)-ethenodeoxycytidine (r = 0.92, p < 0.001) was observed. Etheno-DNA adducts also correlated significantly with cell proliferation (p < 0.01), which was especially enhanced in patients who both drank and smoked (p < 0.001). Nonsmokers and nondrinkers had the lowest rate of cell proliferation, CYP2E1 expression and DNA lesions. Our data demonstrate for the first time an induction of CYP2E1 in the esophageal mucosa by ethanol in a dose dependent manner in man and may explain, at least in part, the generation of carcinogenic DNA lesions in this target organ.
Assuntos
Consumo de Bebidas Alcoólicas/efeitos adversos , Citocromo P-450 CYP2E1/genética , Neoplasias Esofágicas/induzido quimicamente , Neoplasias Esofágicas/genética , Etanol/toxicidade , Idoso , Consumo de Bebidas Alcoólicas/genética , Testes de Carcinogenicidade , Citocromo P-450 CYP2E1/metabolismo , Adutos de DNA/metabolismo , Neoplasias Esofágicas/enzimologia , Feminino , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Valores de Referência , Fumar/epidemiologia , Fumar/genéticaRESUMO
The cannabinoid system (CS) is implicated in the regulation of hepatic fibrosis, steatosis and inflammation, with cannabinoid receptors 1 and 2 (CB1 and CB2) being involved in regulation of pro- and antifibrogenic effects. Daily cannabis smoking is an independent risk factor for the progression of fibrosis in chronic hepatitis C and a mediator of experimental alcoholic steatosis. However, the role and function of CS in alcoholic liver fibrosis (ALF) is unknown so far. Thus, human liver samples from patients with alcoholic liver disease (ALD) were collected for analysis of CB1 expression. In vitro, hepatic stellate cells (HSC) underwent treatment with acetaldehyde, Δ9-tetrahydrocannabinol H2O2, endo- and exocannabinoids (2-arachidonoylglycerol (2-AG) and [THC]), and CB1 antagonist SR141716 (rimonabant). In vivo, CB1 knockout (KO) mice received thioacetamide (TAA)/ethanol (EtOH) to induce fibrosis. As a result, in human ALD, CB1 expression was restricted to areas with advanced fibrosis only. In vitro, acetaldehyde, H2O2, as well as 2-AG and THC, alone or in combination with acetaldehyde, induced CB1 mRNA expression, whereas CB1 blockage with SR141716 dose-dependently inhibited HSC proliferation and downregulated mRNA expression of fibrosis-mediated genes PCα1(I), TIMP-1 and MMP-13. This was paralleled by marked cytotoxicity of SR141716 at high doses (5-10 µmol/L). In vivo, CB1 knockout mice showed marked resistance to alcoholic liver fibrosis. In conclusion, CB1 expression is upregulated in human ALF, which is at least in part triggered by acetaldehyde (AA) and oxidative stress. Inhibition of CB1 by SR141716, or via genetic knock-out protects against alcoholic-induced fibrosis in vitro and in vivo.
Assuntos
Cirrose Hepática Alcoólica/metabolismo , Receptor CB1 de Canabinoide/metabolismo , Acetaldeído/farmacologia , Animais , Apoptose/efeitos dos fármacos , Canabinoides/farmacologia , Hipóxia Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Colágeno/metabolismo , Feminino , Células Estreladas do Fígado/efeitos dos fármacos , Células Estreladas do Fígado/metabolismo , Células Estreladas do Fígado/patologia , Humanos , Peróxido de Hidrogênio/farmacologia , Inflamação/complicações , Inflamação/patologia , Fígado/efeitos dos fármacos , Fígado/metabolismo , Fígado/patologia , Cirrose Hepática Alcoólica/complicações , Cirrose Hepática Alcoólica/enzimologia , Cirrose Hepática Alcoólica/patologia , Masculino , Metaloproteinases da Matriz/genética , Metaloproteinases da Matriz/metabolismo , Camundongos , Pessoa de Meia-Idade , Piperidinas/toxicidade , Pirazóis/toxicidade , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptor CB1 de Canabinoide/deficiência , Receptor CB2 de Canabinoide/metabolismo , Rimonabanto , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/genéticaRESUMO
The hereditary hyperferritinaemia-cataract syndrome (HHCS) is characterised by an autosomal dominant cataract and high levels of serum ferritin without iron overload. The cataract develops due to L-ferritin deposits in the lens and its pulverulent aspect is pathognomonic. The syndrome is caused by mutations within the iron-responsive element of L-ferritin. These mutations prevent efficient binding of iron regulatory proteins 1 and 2 to the IRE in L-ferritin mRNA, resulting in an unleashed ferritin translation. This paper reviews all 31 mutations (27 single nucleotide transitions and four deletions) that have been described since 1995. Laboratory test showing hyperferritinaemia, normal serum iron and normal transferrin saturation are indicative for HHCS after exclusion of other causes of increased ferritin levels (inflammation, malignancy, alcoholic liver disease) and should prompt an ophthalmological consultation for diagnostic confirmation. Invasive diagnostics such as liver biopsy are not indicated. HHCS is an important differential diagnosis of hyperferritinaemia. Haematologists, gastroenterologists and ophthalmologists should be aware of this syndrome to spare patients from further invasive diagnosis (liver biopsy), and also from a false diagnosis of hereditary haemochromatosis followed by venesections. Patients diagnosed with HHCS should be counselled regarding the relative harmlessness of this genetic disease, with early cataract surgery as the only clinical consequence.
Assuntos
Catarata/genética , Ferritinas/metabolismo , Distúrbios do Metabolismo do Ferro/genética , Mutação , Fenótipo , Sequência de Bases , Catarata/epidemiologia , Ferritinas/genética , Predisposição Genética para Doença , Humanos , Distúrbios do Metabolismo do Ferro/diagnóstico , Distúrbios do Metabolismo do Ferro/epidemiologia , Modelos Biológicos , Dados de Sequência Molecular , Conformação de Ácido Nucleico , SíndromeRESUMO
BACKGROUND & AIMS: Liver stiffness (LS) as measured by transient elastography [Fibroscan] offers a novel non-invasive approach to assess liver cirrhosis. Since Fibroscan seems to be unreliable in patients with congestive heart failure, it remains to be determined whether hemodynamic changes affect LS irrespective of fibrosis. METHODS & RESULTS: Using landrace pigs, we studied the direct relationship between the central venous pressure and LS measured by Fibroscan. Clamping of the inferior caval vein increased LS from 3.1 to 27.8kPa while reopening reversed LS within 5min to almost normal values of 5.1kPa. We then studied LS as a function of venous pressure in the isolated pig liver by clamping the upper and lower caval, portal vein and hepatic artery. The stepwise increase of intravenous pressure to 36cm of water column (3.5kPa) linearly and reversibly increased LS to the upper detection limit of 75kPa. We finally measured LS in 10 patients with decompensated congestive heart failure before and after recompensation. Initial LS was elevated in all patients, in 8 of them to a degree that suggested liver cirrhosis (median 40.7kPa). Upon recompensation with a median weight loss of 3.0kg, LS decreased in all 10 patients down to a median LS of 17.8kPa. Inflammation could not account for increased LS since initial liver enzyme counts were only slightly elevated and did not change significantly. CONCLUSION: LS is a direct function of central venous pressure which should be considered when assessing the degree of fibrosis.
Assuntos
Pressão Venosa Central/fisiologia , Elasticidade/fisiologia , Cirrose Hepática/diagnóstico , Fígado/irrigação sanguínea , Fígado/fisiologia , Idoso , Idoso de 80 Anos ou mais , Animais , Técnicas de Imagem por Elasticidade , Feminino , Insuficiência Cardíaca/fisiopatologia , Humanos , Cirrose Hepática/fisiopatologia , Masculino , Pessoa de Meia-Idade , Sus scrofaRESUMO
Studies have suggested the reversibility of liver fibrosis, but the mechanisms of fibrosis reversal are poorly understood. We investigated the possible functional link between apoptosis, macrophages, and matrix turnover in rat liver during reversal of fibrosis secondary to bile duct ligation (BDL). Biliary fibrosis was induced by BDL for 4 wk. After Roux-en-Y (RY)-bilio-jejunal-anastomosis, resolution of fibrosis was monitored for up to 12 wk by hepatic collagen content, matrix metalloproteinase (MMP) expression and activities, and fibrosis-related gene expression. MMP expression and activities were studied in macrophages after engulfment of apoptotic cholangiocytes in vitro. Hepatic collagen decreased to near normal at 12 wk after RY-anastomosis. During reversal, profibrogenic mRNA declined, whereas expression of several profibrolytic MMPs increased. Fibrotic septa showed fragmentation at week 4 and disappeared at week 12. Peak histological remodeling at week 4 was characterized by massive apoptosis of cytokeratin 19+ cholangiocytes, >90% in colocalization with CD68+ macrophages, and a 2- to 7.5-fold increase in matrix-degrading activities. In vitro, phagocytosis of apoptotic cholangiocytes induced matrix-degrading activities and MMP-3, -8, and -9 in rat peritoneal macrophages. We concluded that reconstruction of bile flow after BDL leads to an orchestrated fibrolytic program that results in near complete reversal of advanced fibrosis. The peak of connective tissue remodeling and fibrolytic activity is associated with massive apoptosis of cholangiocytes and their phagocytic clearance by macrophages in vivo. Macrophages upregulate MMPs and become fibrolytic effector cells upon apoptotic cholangiocyte engulfment in vitro, suggesting that phagocytosis-associated MMP induction in macrophages significantly contributes to biliary fibrosis reversal.
Assuntos
Apoptose/fisiologia , Ductos Biliares Intra-Hepáticos/patologia , Cirrose Hepática Biliar/patologia , Cirrose Hepática Experimental/patologia , Macrófagos/fisiologia , Fagocitose/fisiologia , Anastomose em-Y de Roux , Animais , Ductos Biliares Extra-Hepáticos/cirurgia , Linhagem Celular , Movimento Celular/fisiologia , Células Cultivadas , Colágeno/metabolismo , Colágeno Tipo I/genética , Cadeia alfa 1 do Colágeno Tipo I , Colagenases/metabolismo , Regulação para Baixo/genética , Gelatinases/metabolismo , Expressão Gênica/genética , Cadeias beta de Integrinas/genética , Ligadura , Fígado/metabolismo , Fígado/patologia , Cirrose Hepática Biliar/metabolismo , Cirrose Hepática Biliar/cirurgia , Cirrose Hepática Experimental/metabolismo , Cirrose Hepática Experimental/cirurgia , Macrófagos/enzimologia , Macrófagos/patologia , Masculino , Metaloproteinases da Matriz/metabolismo , Camundongos , Modelos Biológicos , Inibidor 1 de Ativador de Plasminogênio/genética , Ratos , Ratos Sprague-Dawley , Inibidor Tecidual de Metaloproteinase-1/genética , Fator de Crescimento Transformador beta/genéticaRESUMO
UNLABELLED: Iron in association with reactive oxygen species (ROS) is highly toxic, aggravating oxidative stress reactions. Increased iron not only plays an important role in the progression of hereditary hemochromatosis (HH) but also in common liver diseases such as chronic hepatitis C. The underlying mechanisms of hepatitis C virus (HCV)-mediated iron accumulation, however, are poorly understood. We introduce an in vitro-targeted approach to identify ROS/iron-regulated genes in patients with HCV using a genome-wide DNA microarray. The sensitivity of the 32,231 complementary DNA clone-carrying microarray was approximately 20% as estimated by detecting target genes of the genome-wide transcription factor hypoxia inducible factor 1alpha. Upon in vitro challenge to iron and oxidative stress, 265 iron-related and 1326 ROS-related genes could be identified in HepG2 cells; 233 significantly regulated genes were found in patients with mild (HCV) or severe (HH) iron deposition. Notably, 17 of the in vitro-selected genes corresponded to the genes identified in patients with HCV or HH. Among them, natriuretic peptide precursor B (NPPB) was the only iron-regulated gene identified in vitro that was differentially regulated between HCV and HH. Reverse-transcription polymerase chain reaction confirmed most of the microarray-identified genes in an even larger group of patients (n = 12). In patients with HCV, these included genes that are associated with RNA processing (MED9/NFAT, NSUN2), proliferation, differentiation, hypoxia, or iron metabolism (ISG20, MIG6, HIG2, CA9, NDRG1), whereas none of the nine known iron-related genes showed significant differences between HCV and HH. CONCLUSION: Although high-density microarray technology is less suitable for routine liver diagnosis, its use in combination with prior in vitro selection is a powerful approach to identify candidate genes relevant for liver disease.
Assuntos
Genoma Humano , Hepatite C/genética , Análise de Sequência com Séries de Oligonucleotídeos , Adulto , DNA Complementar/genética , Progressão da Doença , Hemocromatose/genética , Hepatite C/complicações , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Ferro/metabolismo , Cirrose Hepática/epidemiologia , Pessoa de Meia-Idade , RNA Mensageiro/genética , Espécies Reativas de Oxigênio/metabolismo , Receptores do Fator Natriurético Atrial/genética , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
UNLABELLED: Oxidative stress is thought to play a major role in the pathogenesis of hepatocellular cancer (HCC), a frequent complication of alcoholic liver disease (ALD). However, the underlying mechanisms are poorly understood. In hepatocytes of ALD patients, we recently detected by immunohistochemistry significantly increased levels of carcinogenic etheno-DNA adducts that are formed by the reaction of the major lipid peroxidation product, 4-hydroxynonenal (4-HNE) with nucleobases. In the current study, we show that protein-bound 4-HNE and etheno-DNA adducts both strongly correlate with cytochrome P450 2E1 (CYP2E1) expression in patients with ALD (r = 0.9, P < 0.01). Increased levels of etheno-DNA adducts were also detected in the liver of alcohol-fed lean (Fa/?) and obese (fa/fa) Zucker rats. The number of nuclei in hepatocytes stained positively for etheno-DNA adducts correlated significantly with CYP2E1 expression (r = 0.6, P = 0.03). To further assess the role of CYP2E1 in the formation of etheno-DNA adducts, HepG2 cells stably transfected with human CYP2E1 were exposed to ethanol with or without chlormethiazole (CMZ), a specific CYP2E1 inhibitor. Ethanol increased etheno-DNA adducts in the nuclei of CYP2E1-transfected HepG2 cells in a concentration-dependent and time-dependent manner, but not in vector mock-transfected control cells. CMZ blocked the generation of etheno-DNA adducts by 70%-90% (P < 0.01). CONCLUSION: Our data support the assumption that ethanol-mediated induction of hepatic CYP2E1 leading inter alia to highly miscoding lipid peroxidation-derived DNA lesions may play a central role in hepatocarcinogenesis in patients with ALD.
Assuntos
Aldeídos/metabolismo , Citocromo P-450 CYP2E1/metabolismo , Adutos de DNA/metabolismo , Hepatopatias Alcoólicas/metabolismo , Adulto , Idoso , Animais , Biópsia , Linhagem Celular Tumoral , Etanol/efeitos adversos , Feminino , Humanos , Fígado/metabolismo , Masculino , Pessoa de Meia-Idade , Ratos , Ratos ZuckerRESUMO
Breath composition is altered in liver diseases. We tested if ion-molecule-reaction mass spectrometry (IMR-MS) combined with a new statistical modality improves the diagnostic accuracy of breath analysis in liver diseases. We analysed 114 molecules in the breath of 126 individuals (healthy controls, and patients with non-alcoholic and alcoholic fatty liver disease and liver cirrhosis) by IMR-MS. Characteristic exhalation patterns were identified for each group. Combining two to seven molecules in the new stacked feature ranking model reached a diagnostic accuracy (area under the curve) for individual liver diseases between 0.88 and 0.97. IMR-MS followed by sophisticated statistical analysis is a promising tool for liver diagnostics by breath analysis.
Assuntos
Testes Respiratórios , Hepatopatias/diagnóstico , Espectrometria de Massas , Acetaldeído/análise , Adulto , Idoso , Biomarcadores , Butadienos/análise , Etanol/análise , Fígado Gorduroso/diagnóstico , Fígado Gorduroso Alcoólico/diagnóstico , Feminino , Hemiterpenos/análise , Humanos , Cirrose Hepática/diagnóstico , Hepatopatias/classificação , Masculino , Pessoa de Meia-Idade , Pentanos/análise , Projetos PilotoRESUMO
UNLABELLED: Transient elastography (FibroScan [FS]) is a novel non-invasive tool to assess liver fibrosis/cirrhosis. However, it remains to be determined if other liver diseases such as extrahepatic cholestasis interfere with fibrosis assessment because liver stiffness is indirectly measured by the propagation velocity of an ultrasound wave within the liver. In this study, we measured liver stiffness immediately before endoscopic retrograde cholangiopancreatography and 3 to 12 days after successful biliary drainage in patients with extrahepatic cholestasis mostly due to neoplastic invasion of the biliary tree. Initially elevated liver stiffness decreased in 13 of 15 patients after intervention, in 10 of them markedly. In three patients, liver stiffness was elevated to a degree that suggested advanced liver cirrhosis (mean, 15.2 kPa). Successful drainage led to a drop of bilirubin by 2.8 to 9.8 mg/dL whereas liver stiffness almost normalized (mean, 7.1 kPa). In all patients with successful biliary drainage, the decrease of liver stiffness highly correlated with decreasing bilirubin (Spearman's rho = 0.67, P < 0.05) with a mean decrease of liver stiffness of 1.2 +/- 0.56 kPa per 1 g/dL bilirubin. Two patients, in whom liver stiffness did not decrease despite successful biliary drainage, had advanced liver cirrhosis and multiple liver metastases, respectively. The relationship between extrahepatic cholestasis and liver stiffness was reproduced in an animal model of bile duct ligation in landrace pigs where liver stiffness increased from 4.6 kPa to 8.8 kPa during 120 minutes of bile duct ligation and decreased to 6.1 kPa within 30 minutes after decompression. CONCLUSION: Extrahepatic cholestasis increases liver stiffness irrespective of fibrosis. Once extrahepatic cholestasis is excluded (e.g., by liver imaging and laboratory parameters) transient elastography is a valuable tool to assess liver fibrosis in chronic liver diseases.
Assuntos
Ductos Biliares/patologia , Colestase Extra-Hepática/patologia , Icterícia Obstrutiva/patologia , Cirrose Hepática/patologia , Fígado/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Colestase Extra-Hepática/diagnóstico por imagem , Técnicas de Imagem por Elasticidade/métodos , Feminino , Neoplasias Gastrointestinais/patologia , Humanos , Icterícia Obstrutiva/etiologia , Cirrose Hepática/diagnóstico por imagem , Masculino , Pessoa de Meia-Idade , Neoplasias Pancreáticas/patologia , Neoplasias Gástricas/patologia , Tomografia Computadorizada por Raios XRESUMO
BACKGROUND: Chronic alcohol consumption is a risk factor for colorectal cancer. Animal experiments as well as genetic linkage studies in Japanese individuals with inactive acetaldehyde dehydrogenase leading to elevated acetaldehyde concentrations following ethanol ingestion support the hypothesis that acetaldehyde may be responsible for this carcinogenic effect of alcohol. In Caucasians, a polymorphism of alcohol dehydrogenase 1C (ADH1C) exists resulting in different acetaldehyde concentrations following ethanol oxidation. METHODS: To evaluate whether the association between alcohol consumption and colorectal tumor development is modified by ADH1C polymorphism, we recruited 173 individuals with colorectal tumors diagnosed by colonoscopy and 788 control individuals without colorectal tumors. Genotyping was performed using genomic DNA extracted from whole blood followed by polymerase chain reaction. RESULTS: Genotype ADH1C*1/1 was more frequent in patients with alcohol-associated colorectal neoplasia compared to patients without cancers in the multivariate model controlling for age, gender, and alcohol intake (odds ratio = 1.674, 95% confidence interval = 1.110-2.524, 2-sided p from Wald test = 0.0139). In addition, the joint test of the genetic effect and interaction between ADH1C genotype and alcohol intake (2-sided p = 0.0007) indicated that the difference in ADH1C*1 polymorphisms between controls and colorectal neoplasia is strongly influenced by the alcohol consumption and that only individuals drinking more than 30 g ethanol per day with the genotype ADH1C*1/1 had an increased risk for colorectal tumors. CONCLUSIONS: These data identify ADH1C homozygosity as a genetic risk marker for colorectal tumors in individuals consuming more than 30 g alcohol per day and emphasize the role of acetaldehyde as a carcinogenic agent in alcohol-related colorectal carcinogenesis.
Assuntos
Adenoma/genética , Álcool Desidrogenase/genética , Consumo de Bebidas Alcoólicas/efeitos adversos , Carcinoma/genética , Neoplasias Colorretais/genética , Acetaldeído/metabolismo , Adulto , Idoso , Álcool Desidrogenase/metabolismo , Neoplasias Colorretais/metabolismo , Feminino , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Polimorfismo Genético , Fatores de RiscoRESUMO
The exact regulation of the liver-secreted peptide hepcidin, the key regulator of systemic iron homeostasis, is still poorly understood. It is potently induced by iron, inflammation, cytokines or H2O2 but conflicting results have been reported on hypoxia. In our current study, we first show that pronounced (1%) and mild (5%) hypoxia strongly induces hepcidin in human Huh7 hepatoma and primary liver cells predominantly at the transcriptional level via STAT3 using two hypoxia systems (hypoxia chamber and enzymatic hypoxia by the GOX/CAT system). SiRNA silencing of JAK1, STAT3 and NOX4 diminished the hypoxia-mediated effect while a role of HIF1α could be clearly ruled out by the response to hypoxia-mimetics and competition experiments with a plasmid harboring the oxygen-dependent degradation domain of HIF1α. Specifically, hypoxia drastically enhances the H2O2-mediated induction of hepcidin strongly pointing towards an oxidase as powerful upstream control of hepcidin. We finally provide evidences for an efficient regulation of hepcidin expression by NADPH-dependent oxidase 4 (NOX4) in liver cells. In summary, our data demonstrate that hypoxia strongly potentiates the peroxide-mediated induction of hepcidin via STAT3 signaling pathway. Moreover, oxidases such as NOX4 or artificially overexpressed urate oxidase (UOX) can induce hepcidin. It remains to be studied whether the peroxide-STAT3-hepcidin axis simply acts to continuously compensate for oxygen fluctuations or is directly involved in iron sensing per se.
Assuntos
Hepcidinas/genética , NADPH Oxidase 4/genética , Fator de Transcrição STAT3/genética , Hipóxia Tumoral/genética , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica , Humanos , Peróxido de Hidrogênio/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Ferro/metabolismo , Janus Quinase 1/genética , Oxigênio/metabolismo , Peróxidos/metabolismo , Transdução de Sinais/genética , Urato Oxidase/genéticaRESUMO
Infections are a major adverse effect during the treatment with anti-TNF-alpha. While exclusion of any bacterial infection and screening for tuberculosis are mandatory before initiating a therapy with anti-TNF-alpha-antibodies, there are no guidelines whether to screen for or how to deal with chronic viral infections such as hepatitis B. In this case report, we have described a patient with Crohn's disease who developed subfulminant hepatitis B after the fourth infusion of infliximab due to an unrecognized HBs-antigen carrier state. He recovered completely after lamivudine therapy was started, but this severe adverse event could have been prevented if screening for HBV and pre-emptive therapy with lamivudine would have been started prior to infliximab. We therefore strongly argue in favor of extended screening recommendations for infectious diseases including viral infections before considering a therapy with infliximab.
Assuntos
Anticorpos Monoclonais/uso terapêutico , Doença de Crohn/tratamento farmacológico , Vírus da Hepatite B/isolamento & purificação , Hepatite B/diagnóstico , Anticorpos Monoclonais/efeitos adversos , Humanos , Infliximab , Masculino , Programas de Rastreamento , Pessoa de Meia-IdadeRESUMO
OBJECTIVE: Growing evidence suggests that immune reactions to heat shock protein 60 (HSP60) are involved in atherogenesis. Because of the high phylogenetic conservation between microbial and human HSP60, bacterial infections might be responsible for breaking the tolerance to self-HSP60, which is expressed on the surface of stressed arterial endothelial cells. METHODS AND RESULTS: We purified serum antibodies to Escherichia coli HSP60 (GroEL), the 60-kD chlamydial HSP, and HSP65 of Mycobacterium tuberculosis by affinity chromatography from clinically healthy subjects with sonographically proven carotid atherosclerosis. Reactivity of the purified antibodies with overlapping human HSP60 peptides was measured, and 8 shared common epitopes, recognized by all anti-bacterial HSP60/65 antibodies, were identified. Antisera specific for these cross-reactive epitopes were produced by immunizing rabbits with peptides derived from human HSP60. By immunohistochemistry, the epitopes were found to be present in the arterial wall of young subjects during the earliest stages of the disease. CONCLUSIONS: Antibodies to microbial HSP60/65 recognize specific epitopes on human HSP60. These cross-reactive epitopes were shown to serve as autoimmune targets in incipient atherosclerosis and might provide further insights into the mechanisms of early atherogenesis.
Assuntos
Arteriosclerose/imunologia , Linfócitos B/imunologia , Proteínas de Bactérias/imunologia , Chaperonina 60/imunologia , Chaperoninas/imunologia , Epitopos/imunologia , Sequência de Aminoácidos , Animais , Antígenos de Bactérias/imunologia , Proteínas de Bactérias/química , Chaperonina 60/química , Chaperoninas/química , Chlamydia trachomatis/imunologia , Reações Cruzadas , Epitopos/química , Escherichia coli/imunologia , Humanos , Masculino , Modelos Moleculares , Dados de Sequência Molecular , Mycobacterium tuberculosis/imunologia , Fragmentos de Peptídeos/síntese química , Fragmentos de Peptídeos/imunologia , Coelhos , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Especificidade da EspécieRESUMO
A 45-year-old patient was sent to our department because of highly elevated transaminases and elevated lactate dehydrogenase. His medical history was unremarkable and he took no medication on regular basis. Physical examination did not detect any abnormalities. There was no evidence for viral hepatitis, Epstein-Barr virus or cytomegalovirus, autoimmune hepatitis, Budd-Chiari syndrome, haemochromatosis or Wilson's disease. During the interview he admitted that for 'prophylactic reasons' he had been drinking the juice of Noni (Morinda citrifolia), a Polynesian herbal remedy made from a tropical fruit, during the preceding 3 weeks. This gave rise to the suspicion of herbal toxicity, which was confirmed by a liver biopsy. After ceasing the ingestion of Noni, transaminase levels normalized quickly and were within normal ranges 1 month after the first presentation. To our knowledge, this is the first report of hepatotoxicity caused by this herbal remedy, which has been highly praised in the tabloid press.
Assuntos
Doença Hepática Induzida por Substâncias e Drogas/etiologia , Morinda/efeitos adversos , Preparações de Plantas/efeitos adversos , Doença Aguda , Doença Hepática Induzida por Substâncias e Drogas/imunologia , Eosinófilos/imunologia , Humanos , Fígado/imunologia , Testes de Função Hepática , Masculino , Pessoa de Meia-IdadeRESUMO
BACKGROUND: Carcinogenic exocyclic-DNA adducts like 1,N(6)-etheno-2'-deoxyadenosine (εdA) are formed through reactive intermediates of 4-hydroxynonenal (4-HNE) or other lipid peroxidation (LPO) products with the DNA bases A, C, methyl-C and G. High levels of hepatic etheno-DNA adducts have been detected in cancer prone liver diseases including alcoholic liver disease (ALD). In ALD εdA levels correlated significantly with cytochrome P-450 2E1 (CYP2E1) expression which is also induced in non-alcoholic steatohepatitis (NASH). We investigated the occurrence of εdA adducts in children with NASH as a DNA damage marker. METHODS: Liver biopsies from 21 children/adolescents with histologically proven NASH were analysed for hepatic fat content, inflammation, and fibrosis. εdA levels in DNA, CYP2E1-expression and protein bound 4-hydroxynonenal (HNE) were semi-quantitatively evaluated by immunohistochemistry. RESULTS: Among 21 NASH children, εdA levels in the liver were high in 3, moderate in 5, weak in 9 and not elevated in 4 patients. There was a positive correlation between CYP2E1 and protein-bound 4-HNE (r=0.60; P=0.008) and a trend for a positive relationship for CYP2E1 vs. staining intensity of εdA (r=0.45; P=0.06). Inflammatory activity and fibrosis correlated significantly (r=0.49, P=0.023). CONCLUSIONS: Our results demonstrate for the first time the presence of elevated carcinogenic etheno-DNA lesions (εdA) in the majority (17/21) of liver biopsies from young NASH patients. Our data suggest that LPO-derived etheno-adducts are implicated in NASH. Whether these adducts may serve as predictive risk markers in NASH children to develop hepatocellular cancer later in life remains to be investigated.