Alcohol consumption analysis among patients with liver disease in China / 中华医学杂志(英文版)

BACKGROUND@#Alcohol consumption has been observed to be a contributing factor in liver damage. However, very few studies have tried to decipher the correlation between patients with liver disease and alcohol consumption. Therefore, this study was planned to determine the prevalence of alcohol consumption among patients with liver disease, and to evaluate the risk factors, liver diseases, and chronic medical conditions associated with alcohol drinking.@*METHODS@#A cross-sectional study was conducted among patients with liver disease in 30 provinces, autonomous regions, and municipalities across China. All participants answered the questionnaire, which led to the calculation of Alcohol Use Disorders Inventory Test (AUDIT) score for each patient. Based on this score, low-risk drinkers, hazardous drinkers, and harmful drinkers were defined as having AUDIT score of <8, between 8 and 15, and ≥16, respectively.@*RESULTS@#A total of 1489 participants completed the questionnaire. Based on this information, 900 (60.44%) participants were classified as alcohol drinkers. Among these, 8.66% were ex-drinkers, 22.10% were low-risk drinkers, 17.13% were hazardous drinkers, and 12.56% were harmful drinkers. Further investigation of the association between alcohol consumption and other baseline characteristics of patients with liver disease revealed that usually men <40 years old, participants having higher family annual income, having college degree or higher education, living alone, having higher body mass index (BMI), current smokers, and ex-smokers had significant association with higher risk of alcohol consumption. In addition, among the 18.07% of the participants with cirrhosis, it was observed that risk of cirrhosis increased with higher alcohol consumption. Furthermore, harmful drinkers showed greater odds of hypertension and heart diseases, while hazardous drinkers and harmful drinkers, both had greater odds of hyperlipidemia.@*CONCLUSIONS@#Overall our analyses indicated that among the patients with liver disease in China, there was high rate of alcohol consumption and dependence. Alcohol consumption usually associated with men <40 years old, higher family income, education level, living alone, high BMI, and smoking. Increased alcohol consumption not only increased the risk of cirrhosis, but also enhanced the risk of hypertension, heart diseases, and hyperlipidemia.

Drug Sensitivity and Resistance Gene of Carbapenem Resistant Enterobacteriaceae / 现代检验医学杂志

Objective To analyze the resistance genes of carbapenem resistant Enterohacteriaceae (CRE) from Zhongnan Hospital of Wuhan University,so as to provide evidence for CRE infection.Methods WHONET 5.6 software was used to analyze the resistance of carbapenem resistant Enterobacteriaceae isolated.Phenotype screening of 53 CRE strains taken with the modified Hodge test by three drug susceptibility slip to imipenem,meropenem and ertapenem,and analysed resistance gene analysis of 31 CRE strains phenotype screening.Results In 53 CRE strains,34 strains of Klebsiella pneumoniae,ac counting for 64.15 ％,12 strains of Escherichia coli,accounting for 22.64 ％,7 strains of Enterobacter cloacae accounted for 13.21％,and there were 31 strains of Hodge test positive strains,accounting for 58.49％ (31/53).The Hodge test results of three kinds of drug sensitive paper of imipenem,meropenem and ertapenem were consistent.In 31 strains of CRE,29 strains of blaKPC gone were amplified,accounting for 93.55 ％,9 strains of blaNDM gene.accounting for 29.03 ％,contained simaltaneous and two genes of blaKPC and blaNDM were 8 strains,accounting for 25.81 ％,the resistance genes of BlaIMP,blaOXA,and blaVIM were not amplified.Conclusion Resistance to carbapenems was mainly due to the production of KPC and NDM in Enterobacteriaceae of 31 strains.The CRE was detected in the Intensive Care Unit and Respiratory ward for mainly Klebsiella pneumoniae,and the main genotype was KPC.The CRE was detected in the Cadre ward mainly Escherich ia coli,and the drug resistant genotype was mainly a mixed type of KPC and NDM.

PD-1 expression in HBcAg-specific CD8+ T cells of adolescents with chronic HBV infection / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To investigate the differential expression of programmed death-1 (PD-1) in the hepatitis B core antigen (HBcAg)17-28-specific CD8+ T cell subsets of adolescent patients with chronic hepatitis B virus (HBV) infection during the immune tolerant phase and the immune clearance phase.</p><p><b>METHODS</b>A total of 105 patients between the ages of 12-28 years old (mean age 17.20+/-6.35) with chronic HBV infection and 15 healthy age-matched individuals were enrolled in the study. The patients were divided into two groups according to their current status in immune clearance phase (n = 55) or immune tolerant phase (n = 50), as determined by hepatic biopsy pathology. Flow cytometry was used to detect HLA-A2 type and PD-1 expression on peripheral blood mononuclear cells (PBMC) and HBcAg17-28-specific CD8+ T cells. PD-1 mRNA levels in PBMCs were measured by reverse transcription-polymerase chain reaction (RT-PCR). Independent samples t-test was used to compare means between the two groups, and one-way ANOVA was used to compare means among multiple groups. Pearson's correlation coefficient was used to assess the significance of correlation.</p><p><b>RESULTS</b>The frequency of HBcAg18-27-specific CD8+ T cells was significantly higher in the immune clearance phase group than in the immune tolerant phase group (t = 18.08, P less than 0.01), but the expression of PD-1 on the HBcAg18-27 specific CD8+ T cells was significantly lower in the immune clearance phase group than in the immune tolerant phase group (t = 4.72, P less than 0.01). A negative correlation existed between the frequency of HBcAg18-27-specific CD8+ T cells and PD-1 expression (r = -0.463, P less than 0.01). A positive correlation existed between HBV viral load and PD-1 expression on the HBcAg18-27-specific CD8+ T cells in chronic HBV infection patients (r = 0.882, P less than 0.01), and there was a negative correlation between PD-1 expression levels on HBcAg18-27-specific CD8+ T cells and hepatic tissue inflammation score (r = -0.76, P less than 0.01). PD-1 mRNA in PBMCs was significantly higher in the immune tolerant phase group than in the immune clearance phase group (t = 30.89, P less than 0.01).</p><p><b>CONCLUSION</b>Up-regulated expression of PD-1 is associated with HBV-specific CD8+ T cells and may play a crucial role in inhibiting their function during the immune tolerance phase of chronic HBV infection in adolescents.</p>

Dynamic changes in programmed death-1 expression on the surface of T cells in chronic hepatitis C patients undergoing interferon therapy / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To investigate the dynamic changes that occur in T cell subsets, particularly involving the surface expression of programmed death 1 (PD-1), in response to pegylated (Peg)-interferon (IFN) a-2a therapy in patients with chronic hepatitis C virus (HCV) infection.</p><p><b>METHODS</b>Twenty-five patients with HCV genotype 1b chronic infection and 10 healthy controls were enrolled in the study. All the HCV patients received combination antiviral therapy of Peg-IFNa-2a (180 mug/week) plus ribavirin. At treatment weeks 0 (baseline), 4, 12, 24 and 48, the level of PD-1 protein expression on the surface of total peripheral CD8+ and CD4+ T cells was determined by flow cytometry and the level of PD-1 mRNA expression in peripheral blood mononuclear cells (PBMCs) was determined by reverse transcription-polymerase chain reaction. Independent student's t-test were used to compare mean values between the two groups, repeat measure variance analysis was used to compare mean values among multiple groups, and Pearson's correlation coefficient was used to assess correlation significance.</p><p><b>RESULTS</b>Over the course of antiviral therapy, the proportions of CD4+ T cells and CD8+ T cells, as well as the CD4+/CD8+ ratio, increased (F = 81.23, 39.28, and 7.01 respectively; all P less than 0.01). In contrast, the PD-1 protein expression frequency on CD4+ T cells and CD8+ T cells significantly declined (F = 100.11 and 158.40 respectively; all P less than 0.01). The PD-1-mRNA expression level in PBMCs was: 1.40+/-0.26 at baseline, 1.30+/-0.27 at week-4, 1.14+/-0.18 at week-12, 1.06+/-0.26 at week-24, and 0.83+/-0.25 at week-48 (F = 20.09; P less than 0.01). A positive correlation existed between the PD-1 protein expression frequencies on CD4+ T cells and CD8+ T cells and the HCV RNA load detected at baseline (r = 0.82 and 0.75 respectively; all P less than 0.01).</p><p><b>CONCLUSION</b>The ability of Peg-IFN-a-2a-based antiviral therapy to suppress HCV replication may involve reduction of PD-1 protein expression on the surface of CD8+ T cells and CD4+ T cells.</p>

Role of endoplasmic reticulum stress in alcoholic liver disease-related hepatocyte apoptosis / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To investigate the role of endoplasmic reticulum stress (ERS) in alcoholic liver disease (ALD)-related hepatocyte apoptosis.</p><p><b>METHODS</b>A rat model of ALD was established by continuous intragastric administration of ethanol. At 4, 8, 12, and 16 weeks later, randomly selected rats were sacrificed for serum and liver sample collection. Serum levels of total homocysteine (tHcy) were examined by chemiluminescence analysis. Cystathionine beta-synthase (CBS) activity in liver tissue was measured by chromatometry. The mRNA and protein expressions of ERS-related factors, glucose-regulated protein (GRP)-78, calpain 2 and caspase-12, were analyzed by reverse transcription-polymerase chain reaction and immunohistochemistry, respectively. Hepatocyte apoptosis was detected by the TdT-mediated dUTP nick end labeling assay.</p><p><b>RESULTS</b>At 16 weeks, the ALD rats' livers exhibited diffuse microvesicular adipose degeneration and fibrosis in the liver sinus and portal septa. As the duration of ethanol administration extended, the tHcy levels gradually increased (P less than 0.01), CBS activity decreased (P less than 0.01), gene expression levels of GRP-78, calpain 2, and caspase-12 were up-regulated (P less than 0.01), and protein expression levels of GRP-78 and calpain 2 were gradually increased. However, the protein level of procaspase-12 was found to decrease with increased duration of ethanol administration. Finally, the hepatocyte apoptosis index showed an increasing trend over time (P less than 0.01).</p><p><b>CONCLUSION</b>In our experimental ALD rat model, hepatic apoptosis was detected with increasing frequency over the duration of ALD. Increased apoptosis was likely due to decreased CBS activity causing hyperhomocysteinemia, which further induced ERS and activated the calpain 2 and caspase-12 signaling pathway. These ethanol-induced molecular changes may provoke hepatic apoptosis and subsequently promote the pathogenic processes of alcoholic liver disease.</p>

Adiponectin inhibits the activation of hepatic stellate cells induced by TGFb1 via up-regulating the expression of eNOS / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To investigate the molecular mechanism of adiponectin inhibiting activation of hepatic stellate cells in non-alcoholic fatty liver fibrosis.</p><p><b>METHODS</b>The rat models of non-alcoholic fatty liver fibrosis were successfully established by fat-rich diet administration. The expression of adiponectin mRNA and protein were respectively detected by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot. LX-2 cells were cultured in an adipogenic differentiation mixture to induce quiescent adipocytic phenotypes, and then they were treated with TGFβ1, adiponectin and TGFβ1 + adiponectin, respectively. RT-PCR and Western blot were used to determine the expressions of mRNAs and proteins of a-smooth muscle actin (a-SMA), Collagen, adenosine monophosphate-activated protein kinase (AMPK), inducible nitric oxide synthase (iNOS), and endothelial NOS (eNOS). The results were analyzed using one-way ANOVA, Student-Newman-Keuls test, and linear correlation analysis. A P value of less than 0.05 was considered as statistically significant.</p><p><b>RESULTS</b>In vivo, with the progress of non-alcoholic fatty liver fibrosis, the model rats gradually showed hepatic steatosis, inflammation, necrosis and fibrosis. Compared with the control group, the level of serum adiponectin (2.49 ± 0.86 vs 5.81 ± 0.87, P < 0.05) and hepatic expressions of adiponectin mRNA and protein (0.26 ± 0.04 vs 0.72 ± 0.08; 0.64 ± 0.07 vs 0.21 ± 0.07, all P < 0.05) were all decreased in the 24th week group, and were negatively correlated with the level of Collagen which increased gradually. In vitro, TGFβ1 could activate quiescent LX-2 cells by decreasing mRNA and protein expression of eNOS (0.30 ± 0.10 vs 0.44 ± 0.08; 0.30 ± 0.09 vs 0.46 ± 0.07, all P < 0.05) and increasing the expression of iNOS (0.53 ± 0.07 vs 0.37 ± 0.04; 0.55 ± 0.07 vs 0.39 ± 0.05, all P < 0.05). Recombinant adiponectin not only maintained the quiescent phenotype of LX-2 cells but also inhibited LX-2 cells activation due to TGFβ1 by increasing the expression of eNOS (0.43 ± 0.08 vs 0.30 ± 0.10; 0.42 ± 0.07 vs 0.30 ± 0.09, all P < 0.05) and phosphorylation of AMPK (0.43 ± 0.07 vs 0.24 ± 0.04, P < 0.05) and decreasing the expression of iNOS (0.44 ± 0.05 vs 0.53 ± 0.07; 0.46 ± 0.07 vs 0.55 ± 0.07, all P < 0.05).</p><p><b>CONCLUSIONS</b>Data suggested that adiponectin could play a protective role on the pathogenesis of non-alcoholic fatty liver fibrosis by inhibiting the activation of hepatic stellate cells via up-regulating the expression of eNOS, which might associate with increased phosphorylation of AMPK.</p>

The virological response in prolonged therapy of chronic hepatitis C with low-dose peginterferon alpha-2a / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To investigate the virological response in prolonged therapy of chronic hepatitis C (CHC) with low-dose peginterferon alpha-2a.</p><p><b>METHODS</b>The 92 cases of in-patients with chronic hepatitis C in September 2004 to September 2006 were divided to three groups according the endurance of interferon. The dose of peginterferon alpha-2a was 67.5 microg, 90 microg and 180 microg per week in group A, B and C respectively. The treatment duration of peginterferon alpha-2a was 96 or 48 weeks in HCV genotype 1b and 2a in group A and B, and in the group C the duration was 48 or 24 weeks in genotype 1b and 2a patients respectively. Meanwhile, ribavirin for 900-1200 mg per day combined treated with all patients. The quantitation of serum HCV RNA were conducted to determine the rapid virological response (RVR), early virological response (EVR) and sustained virological response (SVR) respectively.</p><p><b>RESULTS</b>There were no significant difference between the three groups in the rate of RVR, EVR and SVR (P > 0.05). There was a higer rate of RVR, EVR and SVR in the genotype 2a group than the genotype 1b group (P < 0.05). HCV genotype was the independent predictor (OR = 12.78, 95%, CI = 11.97-82.89, P = 0.0075) of SVR.</p><p><b>CONCLUSION</b>There was a similar virological response between prolonged therapy of chronic hepatitis C with low-dose peginterferon alpha-2a and the standard dose and duration. The genotype was the independent predictors of SVR in peginterferon alpha-2a antiviral therapy of chronic hepatitis C.</p>

Role of resistin in inflammation of hepatocytes in nonalcoholic steatohepatitis / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To investigate the role and molecular mechanism of resistin in inflammation of hepatocytes in nonalcoholic steatohepatitis.</p><p><b>METHODS</b>Rat models of NASH were established successfully. The expression of resistin mRNA and protein were examined by quantitative RT-PCR and immunohistolostaining, respectively. The murine hepatocytes AML-12 were incubated with recombinant resistin or LPS for 48 hours, and the concentration of TNF alpha, IL-6 in supernatant of AML-12 cells were quantified by enzyme linked immunosorbent assay (ELISA), the nuclear translocation NF- kappa B were observed by immunofluorescence.</p><p><b>RESULTS</b>The steatosis of hepatocytes, inflammation in the lobule and perisinusoidal fibrosis in livers were found, and the expression of resistin mRNA and protein were increased in livers of rat model of NASH. The expression of resistin mRNA was 2.5 and 4 time higher in 12 weeks and 16 weeks of rat models respectively than that in normal control. The positive staining of resistin protein can be found mainly around the central veins. The concentration of TNF alpha and IL-6 were (1.856 +/- 0.049) pg/ml and (9.463 +/- 1.216) pg/ml in supernantant of AML-12 cells 48 hours after recombinant resistin treatment, and (1.791 +/- 0.046) pg/ml, (8.738 +/- 1.101) pg/ml 48 hours after LPS treatment. There was no significant difference between them, but both were higher than that in normal control (P < 0.01). The NF- kappa B p65 nuclear translocation had been observed in AML-12 cells 3 hours after resistin or LPS treatment.</p><p><b>CONCLUSIONS</b>Resistin can induce the production of TNF alpha, IL-6 and other inflammatory factors by hepatocytes, and therefore is an important inflammatory factor in NASH.</p>

Molecular mechanism of ciglitazone inhibiting the expression of extracellular matrix in human hepatic stellate cells / 中华肝脏病杂志

<p><b>OBJECTIVE</b>We tested a hypothesis that PPARgamma inhibits TGFbeta1-activation of TGFbeta receptor (TGFbetaR)-1 signaling in quiescent stellate cells, thereby abrogating Smad3 phosphorylation and inhibiting PAI-1 and collagen expressions.</p><p><b>METHODS</b>Human stellate cells (HSC) were cultured in a medium containing isobutylmethylxanthine, dexamethasone and insulin (MDI) to induce a quiescent adipocytic phenotype one, and then they were treated with TGFbeta1 with or without SB431542, a TGFbetaR1 kinase inhibitor, or the PPARgamma agonist ciglitazone. Effects on Smad 3 phosphorylation, TGFbeta-responsive transcriptional activity, and expressions of collagen and PAI-1 were assessed.</p><p><b>RESULTS</b>Culturing HSC in MDI induced an adipocytic phenotype characterized by lipid accumulation and increased PPARgamma expression and transcriptional activity. TGFbeta1 treatment caused dose- and time-dependent increases in ECM gene expression, increasing collagen and PAI-1 mRNAs by 3 fold within 3 h and increasing PAI-1 protein levels by 8 fold within 6 h. Treatment with the TGFbetaR1 kinase inhibitor, SB431542, inhibited all of these responses. The PPARbeta agonist ciglitazone also caused a dose-dependent inhibition of TGFbeta1's fibrogenic actions. 1 mmol/L ciglitazone blocked TGFbeta1-transcriptional activity and abolished TGFbeta-mediated induction of collagen and PAI-1 expressions.</p><p><b>CONCLUSION</b>The anti-fibrotic ability of PPARgamma agonist ciglitazone may be related to its ability to inhibit TGFbeta1-TGFbetaR1 signaling and blocking pSmad3-dependent induction of PAI-1 and collagen expression.</p>

An experimental study on the reverse mechanism of PPAR-gamma agonist rosiglitazone in rats with non-alcoholic steatohepatitis / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To investigate the influence and significance of peroxisome proliferator- activated receptor-gamma (PPAR-gamma) agonist rosiglitazone on the expression of I kappa B kinase-beta(IKK-beta) mRNA and protein induced by LPS in Kupffer cells (KCs) cultured in vitro and to investigate the activity of nuclear factor-kappa B (NF-kappa B) together with the expression of cyclooxygenase-2 (COX-2) in livers of rats with non-alcoholic steatohepatitis (NASH).</p><p><b>METHODS</b>(1) KCs from healthy Wistar rats were isolated and purified with IV collagenase digestion and gradient centrifugalization, and then were incubated in the presence or absence of LPS (1 microg/ml) together with two different concentrations of rosiglitazone (10 nmol/L and 50 nmol/L). (2) Thirty-eight healthy Wistar rats were randomly divided into a normal blank control group (10 rats) fed with a normal diet and a NASH model group (28 rats) fed with a fat-rich diet (10% lard + 2% cholesterol + 5% corn oil). After the NASH model was established successfully and confirmed by pathological examination of the livers of 4 rats, 24 rats that continued with the high fat-rich diet, were divided into three groups (8 rats in each group): a control group fed normal saline (NS), a lower dose rosiglitazone group (1 mg.kg(-1).d(-1)) and a higher dose rosiglitazone group (4 mg.kg(-1).d(-1)) for 12 weeks. The mRNA expression of IKK-beta in KCs and COX-2 in livers were measured using reverse transcription-polymerase chain reaction (RT-PCR). The IKK-beta protein in KCs and the NF-kappa B activity of hepatic tissues were determined respectively by Western blot and electrophoretic mobility shift assay (EMSA). The concentration of tumor necrosis factor alpha (TNF alpha) in the supernatant of KCs cultures and serum of the rats was quantified by enzyme linked immunosorbent assay (ELISA).</p><p><b>RESULTS</b>LPS significantly increased the expression of IKK-beta mRNA and protein in the KCs and the concentration of TNF alpha in the supernatant of the KCs cultures. The expressions of COX-2 mRNA and protein were more obvious in rats with NASH than those in the normal control group, and the binding activity of NF-kB correlated positively with the expression of COX-2 in the livers and the level of serum TNF alpha of model rats as well. Rosiglitazone blocked the expression of IKK-beta mRNA and protein induced by LPS in KCs, and also inhibited NF-kappa B activation and reduced COX-2 expression in the rats.</p><p><b>CONCLUSIONS</b>PPAR-gamma specific agonist rosiglitazone can play an anti-inflammatory role by IKK-beta/I kappa B/NF-kappa B/TNF alpha signal ways, and minimize inflammatory reaction at cellular and molecular levels. This may help to provide a new idea for treating NASH effectively.</p>