| Background & AimHepatitis B virus (HBV) infection is one of the most common chronic viral infection and a serious threat to health in the world. After acute HBV infection, about 10% adults will develop into chronic hepatitis B (CHB), which is strongly associated with the development of cirrhosis, liver failure and hepatocellular carcinoma. It is generally accepted that the characteristics of the host immune response is the key factor affecting the prognosis of HBV infection. As the inhibitor of NF-κB, zinc finger protein A20 mediated the crucial steps in inflammation and immunity. Emerging related articles suggested that A20 played a critical role in inflammatory and immunological diseases, such as systemic lupus erythematosus and chronic hepatitis C virus infection, however, the expression and significance of A20 in patients with CHB is rarely studied. In this study, we aimed to detect the expression level of A20 in the peripheral blood and liver tissue from patients with CHB and explore the potential role and mechanism of A20 in the progression of chronic hepatitis B virus infection.Patients and MethodsThis retrospective study contained 136 patients with CHB and 30 healthy volunteers as controls. According to different immune stages, patients with chronic hepatitis B were classified into 4 groups:16 subjects were in the immune tolerant (IT) phase; 80 subjects were in the immune clearance (IC) phase; 15 were in the low-replicative (LR) phase; and 25 were in the HBeAg negative hepatitis (ENH) phase. The mRNA level of A20, TNF-a, NF-kB p65 and toll-like receptor (TLR) 4 in peripheral blood mononuclear cells (PBMCs) was determined using quantitative real-time polymerase chain reaction(qRT-PCR). The localization and expression of A20 in liver tissues from 26 patients with CHB and 6 healthy liver transplantation donor were detected by immunohistochemistry method. All of the statistical analyses were performed using the SPSS 19.0 software. The Kolmogorov-Smirnov test was performed to identify whether the data was appropriate for normal distribution. Variables were expressed as median (centile 25;centile 75) or number. Comparisons between or among different groups were performed using the Mann-Whitney U test, Kxuskal-Wallis test or chi-square test. The correlation analysis was calculated by Spearman’s rank correlation test. P value<0.05 was set as statistical significance.Results1. The A20 mRNA level in patients with CHB was significantly higher than that in healthy controls (P<0.001). Further, no significant difference of A20 mRNA between hepatitis B e antigen (HBeAg)(+) and HBeAg(-) was found in patients with chronic hepatitis B.2. A20 mRNA expression levels in patients with CHB were positively correlated with alanine aminotransferase (ALT) (r=0.680, P< 0.001), aspartate aminotransferase (AST) (r= 0.656, P< 0.001), and total bilirubin (TBIL) (r= 0.326, P< 0.001), respectively. No statistically significant relationships were found between A20 mRNA expression levels and other clinical manifestations for the patients with CHB, such as HBV-DNA, hepatitis B surface antigen (HBsAg) and albumin (ALB).3. The relative expression of A20 mRNA in IC/ENH immune phases were significantly higher than LR/IT phases and healthy controls (both P< 0.05, respectively). No significant differences of A20 mRNA in ENH and IC phases (P> 0.05), as well as LR and IT phases (P> 0.05) were found in this study. A cut-off of 4.19 for A20 mRNA was optimal with a sensitivity of 71.25%, a specificity of 81.25% to discriminate the IC phase from the IT phase. The area under the receiver operating characteristic curve (AUROC) of A20 mRNA for the diagnosis of IC from IT individuals was 0.831(95% confidence interval:0.741-0.900, P< 0.001). Meanwhile, A cut-off of 3.97 for A20 mRNA was optimal with a sensitivity of 92.0%, a specificity of 86.67% to discriminate ENH phase from LR phase. The AUROC of A20 mRNA for the diagnosis of ENH from LR individuals was 0.927 (95% confidence interval: 0.798-0.985, P< 0.001).4. The apparent intrahepatic A20 expression was in the cytoplasm from patients with chronic hepatitis B. In addition, the intrahepatic expression of A20 protein was significantly increased in the CHB patients compared with normal controls in accordance with the raised grade of liver inflammation.5. The mRNA levels of TNF-α NF-κB p65 were upregulated in CHB patients compared with HCs (both P< 0.001, respectively), inversely, the expression of TLR4 mRNA was significantly reduced in CHB patients than HCs (P< 0.001). Further analysis indicated significantly positive correlations between A20 and TNF-α、NF-κB p65 mRNA levels (r= 0.245, P< 0.05; r= 0.446, P< 0.001, respectively). A significantly negative correlation was also found between A20 and TLR4 mRNA levels (r=-0.191, P< 0.05).ConclusionPeripheral and intrahepatic A20 expression levels were dramatically increased in patients with CHB compared with healthy controls, which contributed to disease progression of chronic hepatitis B virus infection. It indicated that A20 might significantly participate in the chronically progression of hepatitis B virus infection. |
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