Objective To analyze the effect of N-acetyl-L-cysteine(NAC)on endoplasmic reticulum stress mediated HepG2 cells apoptosis and evaluate the role of NAC in the treatment of liver injury.Methods HepG2 cells were treated with thapsigargin(TG)to establish the model of oxidative endoplasmic reticulum stress mediated apoptosis,and NAC was used to intervene in apoptosis.To evaluate the apoptosis,various methods such as MTT assay,flow cytometry,DNA ladder and Western blot were performed.Results After treated with 2 μmol/L TG for 0,24,36 and 48 hours,the vitality of HepG2 cells decreased.The ratio of apoptotic cells increased along with the prolonged treatment duration of TG(0.7%±0.5%,27.6%±6.3%,29.7%±3.3%,47.9%±3.5% respectively,P<0.05),and the production of reactive oxygen species(ROS)also increased in time-dependent manner(14.0%±0.5%,36.1%±3.0%,38.2%±6.0%,48.3%±12.4%,P<0.05).The HepG2 cells showed typical morphologic change of endoplasmic retieulum stress induced by 2 μmol/L TG after 36 h and 48 h.DNA ladder was observed at the same concentration and time point correspondingly.Endoplasmic reticulum stress mediated-apoptosis was confirmed by Western blot.Both 10 mmol/L and 20 mmol/L NAC could protect ceils from apoptosis.The ratio of apoptotic cells decreased to 14.0%±1.3% and 11.0%±0.3%,respectively.The production of ROS decreased to 34.7%±0.8% and 31.5%±2.9%,respectively.The effect was related to the concentration of NAC.Conclusions As a Ca2+-adenosine triphoshatase inhibitor,TG may disrupt intracellular calcium homeostasis,which can induce endoplasmie reticulum stress and apoptosis.NAC,the precursor of the synthesis of-SH,can directly inhibit the ROS reaction and alleviate liver damage,which may play a role in the treatment of liver failure.

Objective To evaluate the clinical and histological outcomes in a cohort of chronic hepatitis B (CHB) patients who had histologically confirmed severe liver fibrosis and received lamivudine (LAM) therapy for up to 10 years. Methods Thirty-nine CHB patients with severe liver fibrosis (Ishak fibrosis score≥4) were treated with LAM for up to 10 years. Disease progression liver histological improvement, virological and biochemical responses were evaluated during follow-up. Data were analyzed using paired t test, Fisher exact test and Willcoxon test. Results Twenty-eight patients completed the 10-year follow-up. There were 5 (17.9% ) patients with disease progression.At the end of follow up, 16 patients received a second liver biopsy, which showed significant improvement of histological activity index (1.1 ± 1.4 vs 7. 1 ± 3.2, t =- 0.82, P＜0.01 ) and Ishak fibrosis score (3.6±2.2 vs 5.3±0.7, t= -2.89, P＜0.05) compared to baseline. There were 3 cases with Ishak fibrosis score improved from F5 to F0. Among 27 patients, 3(11% ) cases achieved hepatitis B surface antigen (HBsAg) loss and 2 (7 % ) achieved HBsAg seroconversion. At the end of follow-up, 19 out of 23 (83% ) hepatitis B e antigen (HBeAg) positive patients obtained HBeAg loss and 9 (39 % ) obtained HBeAg seroconversion. During LAM treatment, 11 patients experienced virological breakthrough or detected documented LAM-related resistance mutation. The viral loads of all patients were below 1 ×103 copy/mL at the end of follow-up after rescued by add-on or switch to another nucleotide analog.Conclusions Long-term LAM therapy can delay the disease progression in CHB patients with severe liver fibrosis, increase HBsAg and HBeAg loss rates, sustain suppression of HBV replication at a low level and even totally reverse the liver fibrosis in some patients. The effect of LAM resistance mutation on disease outcomes would be reduced by rescue therapy.

Objective To explore the relationship among cytokine levels and Toll-like receptor (TLR) 2,4 in hepatitis B virus (HBV) related cirrhosis. Methods Heparin anticoagulated venous blood of 35 randomly selected HBV related cirrhosis and 35 healthy volunteers were collected aseptically. Plasma tumor necrosis factor (TNF)-α concentration was determined using enzyme-linked immunosorbent assay (ELISA). Peripheral blood mononuclear cells (PBMC) were separated and stained with anti-TLR2,4 monoclonal antibodies, then analysed by flow cytometry. Total RNA was extracted from PBMC and TLR2,4 mRNA expression levels were evaluated by real-time quantitative polymerase chain reaction (PCR) using SYBR Green detection. Means of normal distribution were compared by t test and one factor analysis of variance. The data of abnormal distribution were analyzed using Mann-Whithey U test, Kruskal-Wallis H test and Spearman correlation analysis. Results The plasma concentration of TNF-α in the cirrhotic patients was significantly higher than that in the healthy controls (33.52 ng/L vs 6.07 ng/L, Z=-6.584, P<0. 01), which was parellel with Child-Pugh score. TLR2 positive rate in PBMC from the cirrhotic patients was significantly higher than that from the healthy controls (20.65% vs 12.04%, Z=-4.458, P<0.01), which was positively correlated with plasma TNF-α level (r= 0.448 3, P<0.05), and parellel with Child-Pugh score. Between the cirrhotic and healthy groups, there was no significant difference of TLR4 positive rate in PBMC. The mRNA expression level of TLR2/GAPDH in PBMC from the cirrhotic patients was significantly higher than the controls (0.234 2 vs 0.043 1, Z=-6.83, P<0.01), which was positively correlated with plasma TNF-α level (r=0.411 1, P<0.05), and parellel with Child-Pugh score. Between the two groups, there was no significant difference of TLR4 mRNA expression level in PBMC. Conclusions The expression of TLR2 in PBMC from cirrhotic patients is significantly elevated, which is positively correlated with plasma TNF-α level and the severity of liver disease. The expression of TLR4 in PBMC from cirrhotic patients is unchanged. It suggests that TLR2 but not TLR4, plays an important role in the progression of liver cirrhosis.

Objective To compare the diagnostic values of γ-glutamyl transpeptidase-to-platelet ratio index (GPRI) ,aspartate aminotransferase-to-platelet ratio index (APRI) and fibrosis index based on the four factors (Fib-4) for chronic hepatitis B (CHB) fibrosis and cirrhosis .Methods According to the Metavir score ,262 CHB patients were divided into F0 — F1 group (n= 131) ,F2 — F3 group (n= 102) and F4 group (n = 29 ) . The age , gender , alanine aminotransferase (ALT ) , aspartate aminotransferase (AST ) ,γ-glutamyl transpeptidas (γ-GT ) and platelet count were recorded .GPRI ,APRI ,Fib-4 scores were calculated separately .Statistical analysis was performed by t test ,Kruskal-Wallis H test ,and χ2 test .The correlations between serum models and liver fibrosis stages were analyzed using the Spearman test .Results The scores of GPRI in F0 — F1 group ,F2 — F3 group and F4 group were 0 .39 (0 .21 , 0 .95) ,1 .05 (0 .38 ,2 .39) and 2 .11 (1 .12 ,3 .33) ,respectively .The difference was statistically significant (χ2 = 40 .645 ,P< 0 .01) .APRI scores in the three groups were 0 .49 (0 .32 ,0 .97) ,0 .77 (0 .52 ,1 .52) and 1 .12 (0 .77 ,2 .50) ,respectively .The difference was statistically significant (χ2 = 32 .636 , P < 0 .01) . Fib-4 scores in the three groups were 1 .36 (0 .92 ,2 .05) ,2 .34 (1 .28 ,4 .35) and 3 .86 (3 .03 ,8 .99) , respectively .The difference was statistically significant (χ2 = 48 .943 , P< 0 .01) .Age ,γ-GT ,AST and liver fibrosis were all positively correlated with liver fibrosis (r = 0 .322 ,0 .301 and 0 .199 ,respectively , all P< 0 .05) .Platelet was negatively correlated with liver fibrosis (r = — 0 .455 , P< 0 .05) ,while ALT was not significantly associated with hepatic fibrosis (r= 0 .111 ,P= 0 .073) .GPRI ,APRI and Fib-4 were positively correlated with liver fibrosis (r = 0 .625 ,0 .417 and 0 .399 ,respectively ,all P < 0 .05) .The areas under the operating characteristic curve of GPRI for significant hepatic fibrosis , advanced liver fibrosis and cirrhosis were 0 .818 ,0 .864 and 0 .837 ,respectively .APRI for significant hepatic fibrosis , advanced liver fibrosis and cirrhosis were 0 .694 ,0 .766 and 0 .722 ,respectively ,while Fib-4 were 0 .696 , 0 .770 and 0 .724 ,respectively .The low cutoff values of GPRI for various stages of liver fibrosis were 0 .99 ,1 .04 and 1 .06 ,respectively ,and the sensitivity ,specificity ,positive predictive value (PPV ) and negative predictive value (NPV) were all higher than those of APRI and Fib-4 .The high cutoff values of GPRI for liver fibrosis at each stage were 2 .49 ,3 .69 and 6 .77 , respectively , and the sensitivity , specificity ,PPV and NPV of the diagnosis were all higher than those of APRI and Fib-4 .Conclusion The diagnostic value of GPRI for CHB liver fibrosis is higher than those of APRI and Fib-4 .