RESUMO
Primary biliary cholangitis (PBC) is an autoimmune disease characterized by chronic destruction of the bile ducts. A major unanswered question regarding the pathogenesis of PBC is the precise mechanisms of small bile duct injury. Emperipolesis is one of cell-in-cell structures that is a potential histological hallmark associated with chronic hepatitis B. This study aimed to clarify the pathogenesis and characteristics of emperipolesis in PBC liver injury. Sixty-six PBC patients, diagnosed by liver biopsy combined with laboratory test, were divided into early-stage PBC (stages I and II, n = 39) and late-stage PBC (stages III and IV, n = 27). Emperipolesis was measured in liver sections stained with haematoxylin-eosin. The expressions of CK19, CD3, CD4, CD8, CD20, Ki67 and apoptosis of BECs were evaluated by immunohistochemistry or immunofluorescence double labelling. Emperipolesis was observed in 62.1% of patients with PBC, and BECs were predominantly host cells. The number of infiltrating CD3+ and CD8+ T cells correlated with the advancement of emperipolesis (R2 = 0.318, P < .001; R2 = 0.060, P < .05). The cell numbers of TUNEL-positive BECs and double staining for CK19 and Ki67 showed a significant positive correlation with emperipolesis degree (R2 = 0.236, P < .001; R2 = 0.267, P < .001). We conclude that emperipolesis mediated by CD8+ T cells appears to be relevant to apoptosis of BEC and thus may aggravate the further injury of interlobular bile ducts.
Assuntos
Apoptose , Ductos Biliares/patologia , Linfócitos T CD8-Positivos/imunologia , Emperipolese , Células Epiteliais/patologia , Cirrose Hepática Biliar/fisiopatologia , Ductos Biliares/imunologia , Ductos Biliares/lesões , Estudos de Casos e Controles , Proliferação de Células , Células Epiteliais/imunologia , Feminino , Humanos , Masculino , Pessoa de Meia-IdadeRESUMO
BACKGROUND: Peroxisome proliferator activated receptor alpha (PPARα) ameliorates ethanol induced hepatic steatohepatitis. However, its role in alcoholic liver fibrosis has not been fully clarified. The aim of this study was to elucidate the effect and the molecular basis of PPARα in ethanol induced liver fibrosis in mice. METHODS: C57BL/6J mice were fed with 4% ethanol-containing Lieber-DeCarli liquid diet for eight weeks, and intraperitoneal injected with 5% carbon tetrachloride (CCl4) for the last four weeks to induce alcoholic liver fibrosis. PPARα agonist WY14643 was administered to mice during the last couple of weeks. The effects of PPARα induction on liver histology, activation of hepatic stellate cells (HSCs), as well as hepatic expression of inflammatory and fibrogenic factors were assessed. RESULTS: The ethanol plus CCl4 treated mice exhibited progressive liver injury including piecemeal necrosis of hepatocytes, severe inflammatory cells infiltration and bridging fibrosis. This was accompanied by down-regulated hepatic expression of PPARα and the protective cytokines adiponectin, heme oxygenase-1 and interleukin-10. Additionally, up-regulation of the proinflammatory cytokine tumor necrosis factor-alpha, as well as the profibrogenic genes osteopontin, transforming growth factor-beta 1, visfatin, phosphatidylinositol 3-kinase, matrix metalloproteinase-2 (MMP-2) and MMP-9 was observed. WY14643 treatment restored expression of cytokines altered by ethanol plus CCl4 treatment and concomitantly ameliorated the liver injury. CONCLUSIONS: The present study provides evidence for the protective role of PPARα induction in ameliorating ethanol mediated fibrosis through mediation of inflammatory and fibrogenic factors.
Assuntos
Anticolesterolemiantes/farmacologia , Células Estreladas do Fígado/efeitos dos fármacos , Hepatócitos/efeitos dos fármacos , Cirrose Hepática/tratamento farmacológico , PPAR alfa/genética , Pirimidinas/farmacologia , Adiponectina/genética , Adiponectina/metabolismo , Animais , Tetracloreto de Carbono , Citocinas/genética , Citocinas/metabolismo , Etanol , Regulação da Expressão Gênica/efeitos dos fármacos , Heme Oxigenase-1/genética , Heme Oxigenase-1/metabolismo , Células Estreladas do Fígado/metabolismo , Células Estreladas do Fígado/patologia , Hepatócitos/metabolismo , Hepatócitos/patologia , Interleucina-10/genética , Interleucina-10/metabolismo , Fígado/efeitos dos fármacos , Fígado/metabolismo , Fígado/patologia , Cirrose Hepática/induzido quimicamente , Cirrose Hepática/metabolismo , Cirrose Hepática/patologia , Masculino , Metaloproteinases da Matriz/genética , Metaloproteinases da Matriz/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Nicotinamida Fosforribosiltransferase/genética , Nicotinamida Fosforribosiltransferase/metabolismo , Osteopontina/genética , Osteopontina/metabolismo , PPAR alfa/metabolismo , Transdução de Sinais/efeitos dos fármacos , Fator de Crescimento Transformador beta1/genética , Fator de Crescimento Transformador beta1/metabolismo , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismoRESUMO
OBJECTIVE: To summarize the clinicopathological manifestations of Wilson disease(WD) so as to improve its recognition. METHODS: A total of 29 WD cases were retrospectively analyzed, including clinical presentations, liver function test, serum ceruloplasmin, 24 hour urinary copper excretion, ATP7B gene analysis and liver histology. All cases were diagnosed from January 2007 to October 2012 at Third Hospital of Hebei Medical University and China-Japan Friendship Hospital. RESULTS: There were 18 males and 11 females with an average age of 25.9 years. The major clinical symptoms included fatigue (n = 18, 62.1%), abdominal distension (n = 4,13.8%) and pruritus (n = 4, 13.8%). The common physical signs were hepatomegaly (n = 11, 37.9%), splenomegaly(n = 15, 51.7%) and ascites (n = 4, 13.8%). The laboratory examinations included abnormal liver function (n = 29, 100%), high level of 24-hour urinary copper excretion (n = 29, 100.0%), low serum ceruloplasmin (n = 24, 82.8%) and Kayser-Fleischer ring (n = 8, 27.6%). ATP7B gene mutations were at exons 5, 8, 11, 12, 16 and 18. The earliest histologic abnormalities of liver included steatosis (both microvesicular and macrovesicular). Timm's stain showed positive or negative staining. There was no or focal hepatocellular necrosis in liver. During chronic hepatitis phase, the major changes included inflammatory cells infiltration in portal area with biliary epithelium degeneration. The periportal area hepatic cells were swollen, cytoplasm slightly stained and accompanied with some copper particles deposition and cholestic changes. There were many spotty or focal lesion of necrosis in liver. During cirrhotic phase, portal area became enlarged by fibrotic tissue, numerous copper particles deposited in wide fibrous septa and small bile ducts were damaged and became proliferative. Hepatocytes around fibrous interval showed cholestatic changes and contained many copper particles. They diagnosed on the basis of clinical presentation(n = 6), clinical presentation and liver histology (n = 4) and clinical presentation, liver histology and gene analysis (n = 19). CONCLUSIONS: There is a high misdiagnosis rate of WD based solely on clinical presentation. Cholestic changes around fibrous interval are common histologic features. The most common ATP7B gene mutations are compound heterozygotes in exons 16. Comprehensive evaluations of clinical presentation, liver histology and gene analysis are helpful for early diagnosis and timely treatment so that it helps to reduce the misdiagnosis and missed diagnosis rate of WD.
Assuntos
Adenosina Trifosfatases/genética , Proteínas de Transporte de Cátions/genética , Degeneração Hepatolenticular/genética , Degeneração Hepatolenticular/patologia , Adolescente , Adulto , Idoso , Ceruloplasmina , Criança , Pré-Escolar , ATPases Transportadoras de Cobre , Éxons , Feminino , Degeneração Hepatolenticular/sangue , Humanos , Masculino , Pessoa de Meia-Idade , Mutação , Estudos Retrospectivos , Adulto JovemRESUMO
OBJECTIVE: To explore the role and mechanism of the Fas/Fas ligand (FasL) system and its downstream signaling pathway related to the progression of alcoholic steatohepatitis and liver fibrosis. METHODS: Eighteen C57BL/6J mice were randomly divided into three groups: controls; alcoholic steatohepatitis model, given four-weeks of a 4% ethanol-containing Lieber-DeCarli liquid diet; alcoholic steatohepatitis and liver fibrosis model, given the four-week alcohol diet followed by twice weekly intraperitoneal injections of carbon tetrachloride (5% olive oil solution; 2 mL/kg dose) during the fifth to eighth weeks. Mice in the model groups were sacrificed at the end of week 4 and 8, respectively, along with control mice for comparative analyses. Liver tissue sections were evaluated for hepatocellular apoptosis by terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay. The mRNA expression of Fas, FasL, cysteine aspartate-specific proteases 3 (caspase 3), and cytochrome P450 2E1 (CYP 2E1) in liver tissues was detected by reverse transcription (RT)-PCR, visualized by ethidium bromide staining, and normalized to the gray-value of GAPDH expression. The protein expression of Fas and caspase 3 were detected by western blotting (b-actin normalized), and of FasL and CYP 2E1 by immunohistochemistry staining. Intergroup differences and statistical significance were evaluated by single factor analysis of variance and the least squares difference-t test or the Kruskal-Wallis H test and the Mann-Whitney U test. RESULTS: The number of apoptotic cells in the liver sections was significantly higher in both model groups with alcoholic steatohepatitis (vs. controls) and the amount in the alcoholic steatohepatitis plus liver fibrosis model was significantly higher than that in the model with only alcoholic steatohepatitis. In addition, activation of Fas, FasL and its downstream signaling pathway showed an increasing trend with extent of liver injury. The hepatic mRNA (by RT-PCR) and protein (by western blotting) normalized expression levels in the controls, alcoholic steatohepatitis models, and alcoholic steatohepatitis plus liver fibrosis models were, respectively: Fas mRNA: 0.50+/-0.05, 0.61+/-0.10, 0.76+/-0.03 (H=12.137, P less than 0.05), protein: 0.52+/-0.14, 0.86+/-0.10, 0.99+/-0.09 (F=12.758, P less than 0.01); FasL mRNA: 0.31+/-0.03, 0.53+/-0.02, 1.02+/-0.04 (F=153.260, P less than 0.01); caspase 3 mRNA: 0.86+/-0.11, 0.85+/-0.05, 1.33+/-0.16 (F=8.740, P less than 0.01), protein: 0.40+/-0.03, 0.69+/-0.06, 1.02+/-0.10 (F=90.785, P less than 0.01); CYP 2E1 mRNA: 0.72+/-0.14, 1.00+/-0.15, 1.30+/-0.20 (H=4.713, P less than 0.01). The changes in hepatic FasL and CYP 2E1 expression detected by immunohistochemistry were consistent with the mRNA expression. CONCLUSION: Activation of Fas/FasL and its downstream signaling pathway, which induces hepatocellular apoptosis, contributes to the development of alcoholic steatohepatitis and liver fibrosis.
Assuntos
Proteína Ligante Fas/metabolismo , Fígado Gorduroso Alcoólico/metabolismo , Cirrose Hepática/metabolismo , Transdução de Sinais , Receptor fas/metabolismo , Animais , Apoptose , Citocromo P-450 CYP2E1/metabolismo , Fígado Gorduroso Alcoólico/patologia , Cirrose Hepática/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BLRESUMO
OBJECTIVE: To create a convenient method to establish an alcoholic liver fibrosis model in mice and use it to explore the putative pathogenic mechanisms involving the immunomodulatory proteins osteopontin (OPN) and transforming growth factor-betal (TGF-beta1). METHODS: Forty C57BLI6J mice were fed the Lieber-DeCarli 4% ethanol-containing liquid diet for four weeks, followed by an additional four weeks of the 4% ethanol diet combined with intraperitoneal injection of carbon tetrachloride (CC14 5% solution in olive oil; 2ml/ kg body weight, 2 times/week) to induce alcoholic liver fibrosis. Control groups (n = 6 each) included: normal diet; normal diet plus CCl4 injections; ethanol diet alone; ethanol diet plus solvent (olive oil) injections. Model establishment was monitored by sacrificing six mice at model inception (week 0), and weeks 4, 5, 6, 7, and 8 of modeling to collect liver tissues and blood for histological and biochemical analyses. Extent of hepatic steatosis, inflammation, and fibrosis was assessed by hematoxylin-eosin and Masson staining. Liver function markers, serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels, were tested by automated enzymatic assays. Alpha-smooth muscle actin (alpha-SMA) expression was detected by immunohistochemistry. The mRNA and protein expression of OPN and TGF-beta1 was detected by real-time quantitative reverse transcription-PCR and western blotting, respectively. Significance of differences between multiple groups was assessed by one-way ANOVA analysis followed by least significant difference t-test or Kruskal-Wallis H test followed by the Mann-Whitney U test. RESULTS: Compared to the control groups, the group of mice administrated ethanol and CCl4 developed mild to moderate hepatic steatosis at week 4 of modeling, progressive necroinflammation and perisinusoidal and portal fibrosis from weeks 5-8, and irregular necrosis and bridging fibrosis at week 8. In addition, the model group showed progressive up-regulation of a-SMA expression in the activated hepatic stellate cells (HSCs) and fibrotic areas from weeks 5-8. Both hepatic OPN and TGF-beta1 showed significantly increasing trends in mRNA and protein expressions from weeks 5-8 (OPN mRNA: 1.83 +/- 0.25, 2.94 +/- 0.19, 3.45 +/- 0.31, and 5.99 +/- 0.17 (F= 476.27, P < 0.001); OPN protein: 0.52 +/- 0.06, 1.02 +/- 0.10, 1.52 +/- 0.11 and 1.50 +/- 0.08 (F= 298.03, P< 0.001); TGF-beta1 mRNA: 13.19 +/- 0.40, 3.31 +/- 0.28, 1.58 +/- 0.18 and 2.08 +/- 0.26 (F= 85.55, P < 0.001); TGF-P31 protein: 1.26 +/- 0.16, 0.96 +/- 0.12, 1.09 +/- 0.25 and 1.10 +/- 0.20 (F = 43.64, P < 0.001). CONCLUSION: Feeding C57BL/6J mice the Lieber-DeCarli ethanol-containing liquid diet combined with CCl4 intraperitoneal injection is a convenient method to establish a model of alcoholic liver fibrosis within a relatively short amount of time (eight weeks). Progression of alcoholic liver fibrosis is accompanied by increased hepatic expression of OPN and TGF-beta1, which may contribute to the pathogenic mechanism of this disease and may be targets of future molecular therapies.
Assuntos
Modelos Animais de Doenças , Cirrose Hepática Alcoólica/metabolismo , Fígado/metabolismo , Osteopontina/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Actinas/metabolismo , Animais , Masculino , Camundongos , Camundongos Endogâmicos C57BLRESUMO
OBJECTIVE: To determine the role of IGF-1/PI3K pathway and investigate the molecular mechanism of Fuzhenghuayu (FZHY) therapy in a spontaneous recovery rat model of liver fibrosis. METHODS: The liver fibrosis model was induced in male Wistar rats by administering 8 weeks of twice weekly CCL4 intraperitoneal injections without (untreated model) or with once daily FZHY (treated model). Normal, untreated rats served as the control group. At weeks 4, 6 and 8 (fibrosis) and 10, 12 and 14 (spontaneous recovery) after modeling initiation, effects on protein (a-SMA, IGF-1, PI3K) and mRNA (IGF-1, PI3K) expression levels were evaluated by immunohistochemistry and RT-PCR, respectively. Serum markers of liver function (alanine aminotransferase (ALT) and aspartate aminotransferase (AST)) and liver cell damage (alkaline hydrolysis, HYP) were measured. Histology was performed to assess the degree of inflammation and fibrosis (Ishak scoring system). RESULTS: In the untreated model group, progression of liver fibrosis (weeks 4, 6 and 8) was accompanied by gradual increases in inflammation, necrosis, serum ALT and AST, and hepatic expression of a-SMA protein and IGF-1 and PI3K protein and mRNA; however, during the spontaneous recovery period (weeks 10, 12 and 14) the IGF-1 and PI3K protein and mRNA levels rapidly decreased and the HYP level, Ishak score, and a-SMA hepatic expression also decreased. The FZHY-treated model group showed significantly lower fibrosis-related up-regulation of IGF-1 and PI3K protein and mRNA expression, HYP level, Ishak score, and a-SMA hepatic expression at each time point (vs. untreated model group). CONCLUSION: The IGF-1/PI3K pathway may contribute to progression of liver fibrosis. The mechanism by which FZHY prevents liver fibrosis in a rat model may involve blocking of the IGF/PI3K pathway and inhibiting HSC activation.
Assuntos
Medicamentos de Ervas Chinesas/farmacologia , Cirrose Hepática Experimental/metabolismo , Fígado/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Animais , Fator de Crescimento Insulin-Like I/metabolismo , Fígado/metabolismo , Fígado/patologia , Cirrose Hepática Experimental/patologia , Masculino , Fosfatidilinositol 3-Quinases/metabolismo , Ratos , Ratos WistarRESUMO
OBJECTIVE: To investigate the outcomes of chronic hepatitis C (CHC) patients treated with antiviral regimens of interferon (IFN) plus ribavirin (RBV) using individualized doses and durations. METHODS: This study was designed as an open-label, prospective clinical trial to analyze the virological responses of 169 CHC patients who received individualized dosages of IFNa-2b or pegylated (Peg)IFNa-2a combined with RBV based on their weight ( less than 60 kg or more than or equal to 60 kg), age (less than 65 years or 65-75 years), morbid state (liver cirrhosis or not), and complications (such as heart disease, diabetes, thyroid disorder). Treatment duration was calculated using the time required to induce HCV RNA negativity. The rates of virological response and adverse effects among the different groups were compared. RESULTS: The IFNa-2b treatment was given to 116 patients, and PegIFNa-2a was given to 53 patients. Compared to the IFNa-2b group, the PegIFNa-2a group showed significantly higher rates of complete early virological response (cEVR; 76.7% vs. 92.5%, P less than 0.05) and sustained virological response (SVR; 53.6% vs. 92.3%, P less than 0.05) among the patients who had completed their course of treatment; the rapid virological response (RVR) rate was also higher for the PegIFNa-2a group but the difference did not reach statistical significance (48.7% vs. 60.4%, P more than 0.05). Seventy-eight patients received the routine dose, and 91 patients received the low dose; there were no significant differences between these two groups for RVR (53.8% vs. 58.9%, P more than 0.05), cEVR (78.0% vs. 80.8%, P more than 0.05), or SVR (65.5% vs. 58.3%, P more than 0.05). CONCLUSION: Use of an individualized antiviral treatment strategy designed according to the patient's baseline condition, early viral kinetics, and tolerability to adverse reactions can achieve a high rate of SVR, as well as improve the safety, prognosis, and cost-effectiveness associated with treating CHC patients.
Assuntos
Hepatite C Crônica , Polietilenoglicóis , Hepatite C Crônica/tratamento farmacológico , Humanos , Interferon-alfa/uso terapêutico , Polietilenoglicóis/uso terapêutico , Estudos Prospectivos , Ribavirina/uso terapêutico , Resultado do TratamentoRESUMO
OBJECTIVE: To investigate the association of single nucleotide polymorphisms (SNPs) in the interleukin 17 (IL-17) gene and serum protein levels in patients with chronic hepatitis C virus (HCV) infection. METHODS: A total of 228 patients with chronic HCV infection and 81 healthy controls were enrolled in the study. The frequencies of IL-17 rs8193036 and rs2275913 polymorphisms were detected by the TaqMan SNP genotyping assay. Serum levels of IL-17 protein were detected by ELISA. Pairwise comparisons were made by the Chi-square test, and the significance of between-group differences was assessed by the Student's t-test with P less than 0.05. RESULTS: The patients with chronic HCV infection and the healthy controls showed similar frequencies of the rs8193036 C/T allele (x2 = 1.428, P = 0.232) and the rs2275913 A/G allele (x2 = 0.106, P = 0.744). In addition, the two groups showed similar distribution of the rs8193036 CC (chronic HCV infection: 46.49% vs. healthy controls: 41.98%), CT (45.61% vs. 44.44%) and TT (7.89% vs. 13.58%) genotypes (x2 = 2.346, P = 0.309), and of the rs2275913 AA (16.23% vs. 13.58%), AG (48.25% vs. 50.62%) and GG (35.53% vs. 35.80%) genotypes (x2 = 0.340, P = 0.844). Subgroup analysis of chronic HCV infection patients stratified according to HCV genotypes 1 and 2 showed no differences in the distribution of rs8193036 and rs2275913 alleles (x2 = 1.127, P = 0.288; x2 = 1.088, P = 0.297) and genotypes (x2 = 2.825, P = 0.246; x2 = 0.970, P = 0.616). However, the chronic HCV infection group did show significantly higher levels of serum IL-17 than the controls (97.67+/-39.68 vs. 71.60+/-19.78 pg/ml, t = 2.414, P = 0.033). CONCLUSION: Chronic HCV infection is associated with increased serum IL-17; however, the IL-17 polymorphisms rs8193036 and rs2275913 were not associated with chronic HCV infection susceptibility in this study's Chinese cohort.
Assuntos
Hepatite C Crônica/genética , Interleucina-17/sangue , Interleucina-17/genética , Polimorfismo de Nucleotídeo Único , Adolescente , Adulto , Idoso , Alelos , Estudos de Casos e Controles , Predisposição Genética para Doença , Genótipo , Hepacivirus , Hepatite C Crônica/sangue , Hepatite C Crônica/virologia , Humanos , Pessoa de Meia-Idade , Adulto JovemRESUMO
BACKGROUND: Fuzheng Huayu recipe (FZHY), a compound of Chinese herbal medicine, was reported to improve liver function and fibrosis in patients with hepatitis B virus infection. However, its effect on nutritional fibrosing steatohepatitis is unclear. We aimed to elucidate the role and molecular mechanism of FZHY on this disorder in mice. METHODS: C57BL/6 J mice were fed with methionine-choline deficient (MCD) diet for 8 weeks to induce fibrosing steatohepatitis. FZHY and/or heme oxygenase-1 (HO-1) chemical inducer (hemin) were administered to mice, respectively. The effect of FZHY was assessed by comparing the severity of hepatic injury, levels of hepatic lipid peroxides, activation of hepatic stellate cells (HSCs) and the expression of oxidative stress, inflammatory and fibrogenic related genes. RESULTS: Mice fed with MCD diet for 8 weeks showed severe hepatic injury including hepatic steatosis, necro-inflammation and fibrosis. Administration of FZHY or hemin significantly lowered serum levels of alanine aminotransferase, aspartate aminotransferase, reduced hepatic oxidative stress and ameliorated hepatic inflammation and fibrosis. An additive effect was observed in mice fed MCD supplemented with FZHY or/and hemin. These effects were associated with down-regulation of pro-oxidative stress gene cytochrome P450 2E1, up-regulation of anti-oxidative gene HO-1; suppression of pro-inflammation genes tumor necrosis factor alpha and interleukin-6; and inhibition of pro-fibrotic genes including α-smooth muscle actin, transforming growth factor beta 1, collagen type I (Col-1) and Col-3. CONCLUSIONS: Our study demonstrated the protective role of FZHY in ameliorating nutritional fibrosing steatohepatitis. The effect was mediated through regulating key genes related to oxidative stress, inflammation and fibrogenesis.
Assuntos
Antioxidantes/uso terapêutico , Medicamentos de Ervas Chinesas/uso terapêutico , Fígado Gorduroso/prevenção & controle , Fígado/efeitos dos fármacos , Actinas/genética , Actinas/metabolismo , Animais , Citocromo P-450 CYP2E1/genética , Citocromo P-450 CYP2E1/metabolismo , Citocinas/genética , Citocinas/metabolismo , Dieta/efeitos adversos , Quimioterapia Combinada , Inibidores Enzimáticos/uso terapêutico , Fígado Gorduroso/etiologia , Fígado Gorduroso/metabolismo , Fígado Gorduroso/patologia , Fibrose , Regulação da Expressão Gênica/efeitos dos fármacos , Heme Oxigenase-1/antagonistas & inibidores , Heme Oxigenase-1/genética , Heme Oxigenase-1/metabolismo , Hemina/uso terapêutico , Células Estreladas do Fígado/efeitos dos fármacos , Células Estreladas do Fígado/metabolismo , Células Estreladas do Fígado/patologia , Fígado/metabolismo , Fígado/patologia , Fígado/fisiopatologia , Masculino , Proteínas de Membrana/antagonistas & inibidores , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Necrose , RNA Mensageiro/metabolismo , Distribuição AleatóriaRESUMO
OBJECTIVE: The pathogenesis of non-alcoholic steatohepatitis is still unclear. We have demonstrated previously that peroxisome proliferator activated receptor gamma (PPARγ) ligand protects against inflammation and fibrogenesis in experimental non-alcoholic steatohepatitis. We aim to elucidate the effect and the mechanism of PPARγ itself on nutritional fibrotic steatohepatitis in mice. METHODS: C57BL/6J mice were fed with methionine-choline deficient (MCD) diet for 8 weeks to induce fibrotic steatohepatitis. Mice fed the MCD diet were treated with adenovirus carrying PPARγ (Ad-PPARγ), Ad-PPARγ plus PPARγ agonist rosiglitazone, or PPARγ antagonist 2-chloro-5-nitrobenzaniliden (GW9662), respectively. The effects of up-regulation of PPARγ in the presence or absence of its agonist/or antagonist were assessed by comparing the severity of hepatic injury, activation of hepatic stellate cells and the expression of adiponectin, heme oxygenase-1, and fibrogenic related genes. RESULTS: Mice fed with MCD diet for 8 weeks showed severe hepatic injury including hepatic steatosis, inflammatory infiltration, and fibrosis. Administration of Ad-PPARγ significantly lowered serum alanine aminotransferase level and ameliorated hepatic steatosis, necroinflammation, and fibrosis. These effects were associated with enhanced expression of PPARγ, up-regulated expression of adiponectin and heme oxygenase-1, and down-regulated expression of tumor necrosis factor alpha, interleukin-6, α-smooth muscle actin, transforming growth factor beta 1, matrix metallopeptidase-2, and -9. Administration of GW9662 promoted the severity of liver histology. CONCLUSIONS: The present study provided evidences for the protective role of overexpressing PPARγ in ameliorating hepatic fibrosing steatohepatitis in mice. Modulation of PPARγ expression might serve as a therapeutic approach for fibrotic steatohepatitis.
Assuntos
Fígado Gorduroso/prevenção & controle , Vetores Genéticos/administração & dosagem , PPAR gama/biossíntese , PPAR gama/uso terapêutico , Adenoviridae/genética , Anilidas/administração & dosagem , Animais , Colina , Dieta , Fígado Gorduroso/etiologia , Fígado Gorduroso/genética , Fígado Gorduroso/metabolismo , Inflamação/genética , Inflamação/fisiopatologia , Cirrose Hepática/etiologia , Cirrose Hepática/genética , Cirrose Hepática/metabolismo , Cirrose Hepática/prevenção & controle , Cirrose Hepática Experimental , Masculino , Metionina/deficiência , Camundongos , Camundongos Endogâmicos C57BL , PPAR gama/administração & dosagem , PPAR gama/genética , Distribuição Aleatória , Rosiglitazona , Tiazolidinedionas/administração & dosagem , Transfecção , beta-Galactosidase/administração & dosagem , beta-Galactosidase/genéticaRESUMO
BACKGROUND: Heme oxygenase-1 (HO-1), an antioxidant defense enzyme, has been shown to protect against oxidant-induced liver injury. However, its role on liver fibrosis remains unclear. This study aims to elucidate the effect and the mechanism of HO-1 in nutritional fibrosing steatohepatitis in mice. METHODS: Male C57BL/6J mice were fed with a methionine-choline deficient (MCD) diet for eight weeks to induce hepatic fibrosis. HO-1 chemical inducer (hemin), HO-1 chemical inhibitor zinc protoporphyrin IX (ZnPP-IX) and/or adenovirus carrying HO-1 gene (Ad-HO-1) were administered to mice, respectively. Liver injury was assessed by serum ALT, AST levels and histological examination; hepatic lipid peroxides levels were determined; the expression levels of several fibrogenic related genes were assayed by real-time quantitative PCR and Western blot. RESULTS: MCD feeding mice showed progressive hepatic injury including hepatic steatosis, inflammatory infiltration and fibrosis. Induction of HO-1 by hemin or Ad-HO-1 significantly attenuated the severity of liver injury. This effect was associated with the up-regulation of HO-1, reduction of hepatic lipid peroxides levels, down-regulation of inflammatory factors tumor necrosis factor-alpha, interleukin-6 and suppressor of cytokine signaling-1 as well as the pro-fibrotic genes alpha-smooth muscle actin, transforming growth factor-ß1, matrix metallopeptidase-2 and matrix metallopeptidase-9. A contrary effect was observed in mice treated with ZnPP-IX. CONCLUSIONS: The present study provided the evidence for the protective role of HO-1 in ameliorating MCD diet-induced fibrosing steatohepatitis. Modulation of HO-1 expression might serve as a therapeutic approach for fibrotic steatohepatitis.
Assuntos
Fígado Gorduroso/prevenção & controle , Heme Oxigenase-1/biossíntese , Proteínas de Membrana/biossíntese , Actinas/biossíntese , Alanina Transaminase/sangue , Animais , Aspartato Aminotransferases/sangue , Deficiência de Colina/complicações , Indução Enzimática , Fígado Gorduroso/patologia , Hemina/farmacologia , Peróxidos Lipídicos/metabolismo , Fígado/efeitos dos fármacos , Fígado/patologia , Cirrose Hepática/complicações , Cirrose Hepática/etiologia , Masculino , Malondialdeído/metabolismo , Metaloproteinase 2 da Matriz/biossíntese , Metaloproteinase 9 da Matriz/biossíntese , Metionina/deficiência , Camundongos , Camundongos Endogâmicos C57BL , Protoporfirinas/farmacologia , Fator de Crescimento Transformador beta1/biossíntese , Fator de Necrose Tumoral alfa/biossíntese , Regulação para CimaRESUMO
OBJECTIVE: To elucidate the effect of targeted gene modulation of peroxisome proliferator activated receptor gamma (PPARg) on hepatocellular apoptosis in nutritional fibrotic steatohepatitis in mice. C57BL/6J mice were fed with high fat, methionine-choline deficient (MCD) diet for 8 weeks to induce fibrotic steatohepatitis. Mice fed the MCD diet were treated with adenovirus carrying PPARg (Ad-PPARg), adenovirus-beta-galactosidase (Ad-LacZ), Ad-PPARg plus PPARg agonist rosiglitazone, or PPARg antagonist 2-chloro-5-nitro- benzanilide (GW9662), respectively. H and E stain was performed for observation of hepatocellular apoptosis, hepatic steatosis, inflammation and fibrosis in the liver sections. The expression levels of mRNA and protein of PPARg and apoptosis related genes, Fas, Fas Ligand (FasL), B cell lymphoma/leukemia-2 (Bcl-2), Bcl-2 associated X protein (Bax) and cysteine-containing aspartate-specific proteases-3 (caspase-3) were detected by real-time RT-PCR and Western blot assay, respectively. RESULTS: Mice fed with MCD diet for 8 weeks showed severe hepatic injury including steatosis, hepatocellular apoptosis, inflammatory infiltration and fibrosis, concomitancy with enhanced expression of pro-apoptosis genes, Fas, FasL, Bax and caspase-3 and increased expression of anti-apoptosis gene Bcl-2, by comparing with the control group. The mRNA expression levels of these genes were 3.59+/-0.35 vs 1.11+/-0.37, 4.37+/-1.03 vs 1.09+/-0.33, 4.27+/-0.48 vs 1.03+/-0.10, 4.93+/-0.67 vs 1.12+/-0.24 and 3.95+/-0.34 vs 1.20+/-0.19, and LSD-t values were 2.49, 3.28, 3.25, 3.80 and 2.75, as compared with the control group, P is less than 0.01; the protein expression levels were 1.96+/-0.07 vs 0.45+/-0.07, 0.53+/-0.07 vs 0.22+/-0.02, 1.32+/-0.06 vs 0.59+/-0.03, 1.51+/-0.23 vs 0.36+/-0.09 and 0.57+/-0.01 vs 0.29+/-0.01, and LSD-t values were 1.51, 0.31, 0.73, 1.14 and 0.28, P is less than 0.01. Administration of PPARg agonist rosiglitazone and/or Ad-PPARg significantly ameliorated hepatic steatosis, hepatocellular apoptosis, necro inflammation and fibrosis. These effects were associated with repressed expression of pro-apoptosis genes and up-regulated expression of anti-apoptosis gene. After rosiglitazone treatment, the mRNA expression levels were 3.78+/-0.58, 3.66+/-0.83, 3.04+/-0.37, 2.54+/-0.62 and 4.42+/-0.42, and LSD-t values were 0.18, 0.71, 1.23, 2.39 and 0.46, as compared with MCD group, the P values were 0.627, 0.241, less than 0.01, less than 0.01 and 0.278, the protein expression levels were 1.06+/-0.03, 0.30+/-0.01, 0.70+/-0.05, 1.19+/-0.30 and 0.90+/-0.01, and LSD-t values were 0.90, 0.23, 0.62, 0.31 and 0.34, the P values were less than 0.01, less than 0.01, less than 0.01, 0.122, less than 0.01. After Ad-PPARg treatment, the mRNA expression levels were 2.31+/-0.16, 2.71+/-0.23, 2.52+/-0.27, 1.79+/-0.32 and 5.97+/-0.72, and LSD-t values were 1.28, 1.66, 1.75, 3.13 and 2.02, as compared with MCD group, P is less than 0.05; the protein expression levels were 1.73+/-0.07, 0.43+/-0.04, 1.01+/-0.08, 1.31+/-0.10 and 1.56+/-0.04, and LSD-t values were 0.23, 0.10, 0.30, 0.20 and 0.99, with P values equal 0.009, 0.01, less than 0.01, 0.322 and less than 0.01. CONCLUSIONS: This study provided evidences for the protective role of activation and overexpression of PPARg in ameliorating hepatocellular apoptosis in mice with hepatic fibrosing steatohepatitis.
Assuntos
Apoptose , Cirrose Hepática/patologia , Fígado/patologia , PPAR gama/metabolismo , Animais , Fígado/metabolismo , Cirrose Hepática/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BLRESUMO
OBJECTIVE: Our previous study indicated that the death receptor Fas played a key role on hepatocyte apoptosis in nutritional steatohepatitis in mice. This study aimed to explore whether Fas mutation accelerated hepatic steatosis and inflammatory infiltration in methionine-choline deficient (MCD) diet feeding mice. METHODS: Mice homozygous for the lymphoproliferation spontaneous mutation (C57BL/6J-Faslpr) and wild type C57BL/6J mice were fed with MCD diet for three weeks to induce non-alcoholic steatohepatitis (NASH). Serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), triglyceride (TG) and total cholesterol (TC) levels were detected by an Olympus AU5400 automatic chemical analyzer. The role of Fas gene mutation on NASH was assessed by comparing the severity of hepatic steatosis and inflammation in the liver sections, the mRNA and protein expressions of hepatic inflammatory and fibrogenesis related factors, proliferating cell nuclear antigen (PCNA) and transforming growth factor beta 1 (TGFb1). RESULTS: The serum ALT levels of the wild type and Faslpr mice fed with MCD were significant higher than that of the control mice (126.33+/-10.50 U/L vs (25.00+/-10.14) U/L, (160.33+/-48.29) U/L vs (18.33+/-9.08) U/L, with the LSD-t value 12.02, 5.08 respectively, the P value<0.001, 0.007 respectively. The serum ALT levels showed no significant difference between the Faslpr and wild type mice fed with MCD, with the LSD-t value 1.19, the P value 0.229. The serum AST, TG and TC levels showed neithere significant difference among the four groups. MCD diet induced hepatic steatosis and inflammatory infiltration in both of the wild type and Faslpr mice. Especially, severer hepatic injury was observed in Faslpr mice as compared with wild type mice. The mRNA expression levels of cell proliferation factor PCNA and fibrogenesis growth factor TGF b1 in wild type mice fed with MCD were significantly higher than that of the control mice (2.84+/-0.73, 2.77+/-0.54 vs 1.31+/-0.18, 0.89+/-0.18), with the LSD-t value 4.99, 8.08 respectively, the P value 0.001, <0.001 respectively. The mRNA expression levels of PCNA and TGFb1 in Faslpr mice fed with MCD were significantly higher than that of the Faslpr control mice and the wild type mice fed with MCD (5.57+/-1.13, 5.73+/-0.89 vs 1.04+/-0.16, 0.85+/-0.11 and 2.84+/-0.73, 2.77+/-0.54), with the LSD-t value 10.15, 13.19 and 5.33, 6.91 respectively, the P value<0.001. The protein expressions levels of PCNA and TGFb1 were concordant with the mRNA. CONCLUSIONS: Faslpr promoted hepatic steatosis and inflammatory infiltration in mice fed with MCD diet, which might associated with excessive release of cell proliferative, inflammatory and fibrogenesis factors.
Assuntos
Fígado Gorduroso/genética , Mutação , Receptor fas/genética , Animais , Fígado Gorduroso/induzido quimicamente , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Hepatopatia Gordurosa não Alcoólica , Antígeno Nuclear de Célula em Proliferação/metabolismo , Fator de Crescimento Transformador beta1/metabolismoRESUMO
BACKGROUND: Inflammatory activation of hepatic macrophages plays a primary role in drug-induced liver injury (DILI). However, the exact mechanism underlying DILI remains unclear. METHODS: A total of 328 DILI patients and 80 healthy individuals were prospectively enrolled in this study. The DILI patients were categorized into subgroups based on either disease severity or histopathological patterns. Plasma soluble CD163 (sCD163) and hepatic CD163 were examined to determine hepatic macrophage activation, and CD8, CD20, and MUM-1 were assessed to determine cellular immunity using immunohistochemistry. The lipopolysaccharide (LPS) pathway proteins [e.g. LPS, soluble CD14 (sCD14), and LPS-binding protein (LBP)] were measured using enzyme-linked immunosorbent assay. RESULTS: Plasma sCD163 levels were nine-fold higher in DILI patients than in healthy controls at the baseline, but significantly decreased at the 4-week follow-up visit after treatment. The numbers of hepatic macrophages, B cells, and plasma cells were significantly higher in the liver tissues from DILI patients than those from healthy controls. Furthermore, the baseline levels of LPS pathway proteins in the DILI patients were significantly higher than those in the controls. Notably, these proteins significantly decreased at the 4-week follow-up visit but remained significantly higher than the levels for the controls. CONCLUSIONS: Hepatic inflammation in DILI involves the activation of hepatic macrophages and cellular immunity, in which the LPS pathway likely plays a role, at least in part. As such, this study has improved our understanding of the pathological mechanisms for DILI and may facilitate the development of better treatments for patients with DILI.
Assuntos
Doença Hepática Induzida por Substâncias e Drogas/patologia , Lipopolissacarídeos/metabolismo , Ativação de Macrófagos , Proteínas de Fase Aguda/metabolismo , Adulto , Antígenos CD/metabolismo , Antígenos de Diferenciação Mielomonocítica/metabolismo , Proteínas de Transporte/metabolismo , Feminino , Humanos , Imunidade Celular/fisiologia , Células de Kupffer/imunologia , Células de Kupffer/metabolismo , Células de Kupffer/patologia , Fígado/patologia , Ativação de Macrófagos/imunologia , Masculino , Glicoproteínas de Membrana/metabolismo , Pessoa de Meia-Idade , Receptores de Superfície Celular/metabolismoRESUMO
Non-alcoholic steatohepatitis (NASH) has no approved therapy. The farnesoid X nuclear receptor (FXR) agonist obeticholic acid (OCA) has shown promise as a drug for NASH, but can adversely affect plasma lipid profiles. Therefore, the present study aimed to investigate the effects and underlying mechanisms of OCA in combination with simvastatin (SIM) in a high-fat diet (HFD)-induced model of NASH. C57BL/6J mice were fed with a HFD for 16 weeks to establish the NASH model. The mice were randomly divided into the following five groups: HFD, HFD + OCA, HFD + SIM, HFD + OCA + SIM and control. After 16 weeks, the mice were sacrificed under anesthesia. The ratios of liver weight to body weight (Lw/Bw) and of abdominal adipose tissue weight to body weight were calculated. Serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), total cholesterol, triglycerides and low-density lipoprotein were measured. Liver sections were stained with hematoxylin and eosin. The protein levels of FXR, small heterodimeric partner (SHP) and cytochrome P450 family 7 subfamily A member 1 (CYP7A1) in the liver were detected by western blotting, while the mRNA levels of FXR, SHP, CYP7A1, bile salt export pump, interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), sterol regulatory element binding protein-1 (SREBP1) and fatty acid synthase (FASN) were examined by reverse transcription-quantitative polymerase chain reaction. The administration of OCA with or without SIM reduced the liver inflammation score compared with those of the HFD and HFD + SIM groups, with no significant difference between the HFD + OCA and HFD + OCA + SIM groups. The steatosis score followed similar trends to the inflammation score. In HFD-fed mice, OCA combined with SIM prevented body weight gain compared with that in HFD and HFD + OCA groups, and reduced the Lw/Bw ratio compared with that in the HFD and HFD + SIM groups. In addition to preventing HFD-induced increases of ALT and AST, the combination of OCA and SIM reduced the mRNA levels of IL-6, TNF-α, SREBP1 and FASN. On the basis of these results, it may be concluded that the strategy of combining OCA with SIM represents an effective pharmacotherapy for NASH.
RESUMO
OBJECTIVE: To investigate the potential role of heme oxygenase-1 on preventing non-alcoholic steatohepatitis (NASH) in mice. METHODS: Experimental models of NASH were established by feeding male C57BL/6J mice with choline-methionine deficient diet (MCD) for four weeks. Control animals were fed with choline-methionine supplemented diet. The treatment groups were fed with MCD diet combined with HO-1 inducer hemin or inhibitor zinc protoporphyrin IX (ZnPP-IX). Serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were tested by enzymic method with automatic biochemistry analyzer. The degree of hepatic steatosis, inflammation and fibrosis were examined under HE staining. The hepatic mRNA and protein expressions of HO-1, TNFalpha and IL-6 were analyzed by RT-PCR and Western blot respectively. MCD fed mice showed increased serum ALT and AST levels and moderate to severe hepatic steatosis with inflammatory infiltration, hepatic spot or focal necrosis, light portal and sinus hepaticus fibrosis in the liver sections, which associated with enhanced expression of HO-1, TNFalpha and IL-6 mRNA and protein (1.13+/-0.11, 1.74+/-0.05; 0.20+/-0.01, 1.92+/-0.10; 0.58+/-0.02, 2.06+/-0.05 vs 0.43+/-0.02, 0.75+/-0.05; 0.08+/-0.00, 0.59+/-0.02; 0.22+/-0.01, 0.91+/-0.02). Administration of hemin significantly decreased serum ALT and AST levels and attenuated hepatic steatosis and necroinflammation which associated with up-regulation of antioxidative gene HO-1 and down-regulation of pro-inflammatory cytokines TNFalpha and IL-6 (P < 0.01). A contrary effect on serum aminotransferase levels and liver histopathology was observed in mice injected with ZnPP-IX (P < 0.01). CONCLUSIONS: The effect was associated with suppressed HO-1 expression and increased TNFaLPHA and IL-6 expression. The data provided a biochemical, morphological and molecular biological evidence for the protective role of HO-1 in ameliorating hepatic steatosis, necroinflammation in experimental nutritional steatohepatitis.
Assuntos
Fígado Gorduroso/metabolismo , Fígado Gorduroso/patologia , Heme Oxigenase-1/metabolismo , Proteínas de Membrana/metabolismo , Animais , Interleucina-6/metabolismo , Fígado/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Hepatopatia Gordurosa não Alcoólica , Fator de Necrose Tumoral alfa/metabolismoRESUMO
The liver is the only visceral organ that exhibits a remarkable capability of regenerating in response to partial hepatectomy (PH) or chemical injury. Improving liver regeneration (LR) ability is the basis for the favourable treatment outcome of patients after PH, which can serve as a potential indicator for postoperative survival. The present study aimed to investigate the protective effects of Yiqi Huoxue recipe (YQHX) on LR after PH in rats and further elucidate its underlying mechanism. A two-thirds PH rat model was used in this study. Wistar rats were randomly divided into four groups: sham-operated, PH, YQHX + PH, and Fuzheng Huayu decoction (FZHY) + PH groups. All rats were sacrificed under anesthesia at 24 and 72 h after surgery. The rates of LR were calculated, and the expression levels of cyclin D1 and c-jun were determined by immunohistochemical staining. The protein levels of p-JNK1/2, JNK1/2, p-c-jun, c-jun, Bax, and Bcl-2 were detected by Western blotting, while the mRNA levels of JNK1, JNK2, c-jun, Bax, and Bcl-2 were examined by real-time polymerase chain reaction (RT-PCR). At the corresponding time points, YQHX and FZHY administration dramatically induced the protein levels of p-JNK1/2 compared to the PH group (p < 0.05), while FZHY + PH group showed prominently increase in p-JNK1/2 protein levels compared to the YQHX + PH group (p < 0.05). A similar trend was observed for the expression levels of p-c-jun. Compared to the PH group, YQHX and FZHY markedly reduced the mRNA and protein expression levels of Bax at 24 h after PH, while those in the FZHY + PH group decreased more obviously (p < 0.05). Besides, in comparison with the PH group, YQHX and FZHY administration predominantly upregulated the mRNA and protein expression levels of Bcl-2 at 24 and 72 h after PH (p < 0.05). In conclusion, YQHX improves LR in rats after PH by inhibiting hepatocyte apoptosis via the JNK signaling pathway.
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Hepatic fibrosis is a crucial pathological process involved in the development of chronic hepatitis C (CHC) and may progress to liver cirrhosis and hepatocellular carcinoma. Activated peripheral blood monocytes and intrahepatic macrophages further promote hepatic fibrogenesis by releasing proinflammatory and profibrogenic cytokines. The present study aimed to investigate the role of peripheral CD14+ monocytes and intrahepatic CD163+ macrophages in hepatitis C virus (HCV)-associated liver fibrosis and clarify whether serum soluble CD163 (sCD163) may serve as a fibrosis marker in patients with CHC. A total of 87 patients with CHC and 20 healthy controls were recruited. Serum sCD163 levels were measured by ELISA. Frequencies of peripheral CD14+ monocytes and inflammatory cytokines expressed by CD14+ monocytes were analyzed by flow cytometry. The degree of fibrosis in human liver biopsies was graded using the Metavir scoring system and patients were stratified into two groups based on those results (F<2 vs. F≥2). Hepatic expression of CD163 was examined by immunohistochemical staining. The diagnostic values of sCD163, aspartate aminotransferase to platelet ratio index (APRI), fibrosis 4 score (FIB-4) and the aspartate aminotransferase to alanine aminotransferase ratio (AAR) in significant fibrosis (F≥2) were evaluated and compared using receiver operating characteristic (ROC) curves. The results indicated that the serum sCD163 levels and the frequency of CD14+ monocytes were significantly higher in the patients than that in the controls and positively correlated with liver fibrosis. The level of serum sCD163 was consistent with hepatic CD163 expression in the liver sections from patients. The frequencies of interleukin (IL)-8- and tumor necrosis factor-α-expressing monocytes were increased and that of IL-10-expressing monocytes was decreased in the patients. The area under the ROC curve (AUROC) for sCD163, APRI, FIB-4 and AAR was 0.876, 0.785, 0.825 and 0.488, respectively, and the AUROC for sCD163 was significantly higher than those for APRI and AAR. In conclusion, sCD163 may serve as a novel marker for assessing the degree of liver fibrosis in HCV-infected patients.
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OBJECTIVE: Hepatic oxidative stress plays a key role in the development of non-alcoholic steatohepatitis (NASH). However, the protective effects of antioxidants on NASH are largely unknown. The aim of this study was to elucidate the effect and mechanism of antioxidants on NASH in mice. MATERIAL AND METHODS: C57BL6/J mice were fed a methionine-choline-deficient (MCD) diet for 10 days or 3 weeks to induce steatohepatitis. Antioxidants (vitamin E, ABT, or vitamin E plus ABT) were supplemented in mice fed a MCD diet, respectively. The effect of antioxidants on oxidative stress and apoptosis was assessed, and activation of adiponectin and expressions of inflammatory factors, apoptosis-related genes, and fibrosis-related genes were assayed. RESULTS: MCD feeding in mice showed increasing serum alanine aminotransferase (ALAT) and aspartate aminotransferase (ASAT) levels, and progressive hepatic injury including hepatic steatosis and inflammatory infiltration. Administration of antioxidants vitamin E and/or ABT significantly lowered serum ALAT and ASAT levels (p<0.001) and ameliorated hepatic steatosis and necroinflammation. These effects were associated with repressed hepatic lipid peroxides through reducing hepatic MDA content and enhancing hepatic superoxide dismutase (SOD) activity; down-regulated inflammatory factor COX-2, lowered activity of NF-kappaB, up-regulated anti-apoptotic gene Bcl-2, and down-regulated pro-apoptotic gene Bax suppressed expression of the fibrotic genes TGF-beta1 and MMP2. Moreover, expression of the anti-inflammatory factor adiponectin was also induced by vitamin E or ABT. A combination of vitamin E and ABT showed an additive effect on preventing liver injury. CONCLUSIONS: The present study provides morphological and molecular biological evidence for the protective role of the antioxidant vitamins E and ABT in ameliorating oxidative stress, hepatic apoptosis, and necroinflammation in experimental nutritional steatohepatitis.
Assuntos
Antioxidantes/farmacologia , Fígado Gorduroso/prevenção & controle , Triazóis/farmacologia , Vitamina E/farmacologia , Alanina Transaminase/sangue , Análise de Variância , Animais , Apoptose/efeitos dos fármacos , Aspartato Aminotransferases/sangue , Western Blotting , Ciclo-Oxigenase 2/sangue , Fígado Gorduroso/sangue , Genes bcl-2/efeitos dos fármacos , Inflamação/prevenção & controle , Peroxidação de Lipídeos/efeitos dos fármacos , Masculino , Metaloproteinase 2 da Matriz/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , Subunidade p50 de NF-kappa B/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fator de Crescimento Transformador beta1/efeitos dos fármacos , Proteína X Associada a bcl-2/efeitos dos fármacosRESUMO
OBJECTIVE: Currently, no agent has been confirmed as preventing the fibrosing progression of non-alcoholic steatohepatitis (NASH). In this study, rosiglitazone was used in the clinical treatment of insulin resistance in patients with type 2 diabetes mellitus. However, its protective effect on non-alcoholic fibrosing steatohepatitis is not clear. The study aimed to elucidate the effect and the mechanism of rosiglitazone in inhibiting nutrition-related fibrosis in mice. METHODS: C57BL6/J mice were fed a high fat, methionine-choline deficient (MCD) diet for 8 weeks to induce hepatic fibrosis, and rosiglitazone was given in the treated group. The effect of rosiglitazone was assessed by comparing the severity of hepatic fibrosis in liver sections, the activation of hepatic stellate cells (HSCs) and the expression of TGF-beta1 and connective tissue growth factor (CTGF). RESULTS: At week 8, MCD-diet-induced fibrosing NASH models showed increased serum ALT and AST levels, severe hepatic steatosis, and infiltration of inflammation and fibrosis which, associated with down-regulated PPAR gamma mRNA and protein expression, up-regulated alpha-SMA protein expression and enhanced TGF-beta1, CTGF mRNA and protein expression. Rosiglitazone significantly lowered serum ALT and AST and it reduced MCD-induced fibrosis by repressing levels of alpha-SMA protein expression and pro-fibrosis factors TGF-beta1 and CTGF. It also restored expression of PPAR gamma. CONCLUSIONS: The present study provides clear morphological and molecular biological evidence of the protective role of rosiglitazone in ameliorating nutritional fibrosing steatohepatitis. Rosiglitazone may ameliorate hepatic fibrosis by activating PPAR gamma, which can inhibit HSC activation and suppress TGF-beta1 and CTGF expression.