Lanyi Lin

Patients with chronic hepatitis B infection display deficiency of plasmacytoid dendritic cells with reduced expression of TLR9.

Chronic hepatitis B virus (HBV) infection is a complex interaction between replicating noncytopathic virus and dysregulatory host antiviral immunity. Plasmacytoid dendritic cells (pDCs) contribute to innate antiviral immunity via secreting type I interferons. Toll-like receptor (TLR) 9 is involved in major pattern recognition receptors expressed in pDCs. The frequency of pDCs and TLR9 expression in peripheral blood mononuclear cells (PBMC) was determined, using flow cytometry. IFN-alpha production by PBMC was evaluated in vitro in the presence of cytidine phosphate guanosine (CpG) with/without pDCs. The correlation between TLR9, pDCs frequency and viral load was also evaluated. TLR9 expression in pDCs in chronic HBV patients was significantly ( approximately 50%) reduced, supported by approximately 70% reduction of TLR9 mRNA, in comparison to healthy controls, correlating with the impairment of IFN-alpha production in vitro. Furthermore, pDCs frequency in these patients was substantially reduced ( approximately 30%), inversely correlating with serum ALT levels and HBV viral load. HBsAg and HBcAg were detected by immunohistochemistry in pDCs in chronic HBV patients. We conclude that HBV infection results in reduced frequency of circulating pDCs and their functional impairment via inhibiting the expression of TLR9. These data may provide useful information in both basic research and clinical treatment of chronic HBV infection.

Relationship Between Serum Hepatitis B Virus DNA and Surface Antigen With Covalently Closed Circular DNA in HBeAg-Negative Patients

Hepatitis B virus (HBV) covalently closed circular DNA (cccDNA) is responsible for viral persistence. This study aimed to investigate the serum surrogate markers for cccDNA and to evaluate the intrahepatic viral events associated with disease activity in HBeAg‐negative chronic hepatitis B patients. Thirty‐three treatment‐naïve patients with a negative HBeAg who had a liver biopsy were studied. Active disease was defined as a serum alanine aminotransferase >40 IU/L and a serum HBV DNA >10,000 copies/ml. This study showed significant correlation between serum HBV DNA and both log cccDNA (r = 0.41, P = 0.018) and log total intrahepatic HBV DNA (r = 0.71, P < 0.0001). No significant correlation was observed between serum HBsAg and log cccDNA (P = 0.15) or log total intrahepatic HBV DNA (P = 0.97). Fourteen and 19 patients had inactive and active disease, respectively. The median log cccDNA and log total intrahepatic HBV DNA (copies/106 cells) were significantly higher in patients with active disease compared with those with inactive disease (4.11 vs. 3.53, P = 0.03 and 5.46 vs. 4.64, P < 0.001, respectively). The HBV replicative efficiency, defined as the ratio of serum HBV DNA to cccDNA, was approximately 20% higher in patients with active disease. No significant difference was observed in the HBsAg levels and the ratio of serum HBsAg to cccDNA between the two groups. In conclusion, serum HBV DNA, but not HBsAg, reflects the amount of cccDNA and the replication efficiency of HBV in patients with HBeAg‐negative chronic hepatitis B. J. Med. Virol. 82:1494–1500, 2010.

Common variants of the TLR9 gene influence the clinical course of HBV infection

Hepatitis B virus (HBV) infection leads to the development of liver inflammation, causing morbidity and mortality. Multiple factors influence HBV progression, including genetic factors. Toll-like receptor (TLR)9 plays a key role in innate immunity, and mutations in the genes encoding this receptor have been associated with liver damage progression. Our study aimed to investigate one-tag single nucleotide polymorphisms (rs187084) representing the majority of common variations in TLR9 in a population-based study of Chinese patients. A total of 209 Chinese patients with HBV infection (130 with chronic hepatitis and 79 with liver cirrhosis) and 193 healthy individuals were studied. Our results showed that the frequencies of the C/C genotype and C allele were statistically higher in patients with HBV-related liver cirrhosis than in the healthy controls (26.6 vs. 15.5%; OR=1.97, 95% CI 1.05-3.71, χ2=4.483, P=0.034/43.1 vs. 37.8%; OR=1.49, 95% CI 1.02-2.16, χ2=4.323, P=0.038). No significant differences in the frequencies of alleles or genotypes were found between patients with chronic hepatitis B and the control subjects. In conclusion, this study is the first to show that small effects within TLR9 contribute towards the development of HBV, supporting the hypothesis that little is currently known regarding the contribution of genetic factors to HBV.

Long-term lamivudine treatment achieves regression of advanced liver fibrosis/cirrhosis in patients with chronic hepatitis B.

Antiviral therapy is important in advanced liver fibrosis/cirrhosis with chronic hepatitis B (AdLF‐CHB) patients, but complete regression of cirrhosis remains to be the challenge. We aimed to investigate whether up to 10 years lamivudine treatment achieves liver fibrosis/cirrhosis regression in AdLF‐CHB patients.

Circulating cell death biomarker: good candidates of prognostic indicator for patients with hepatitis B virus related acute-on-chronic liver failure

Investigations on survival of patients with hepatitis B virus related acute-on-chronic liver failure (HBV-ACLF) are sparse and urgently needed. The current study aimed to evaluate the prognostic value of circulating cell death biomarkers (M30-anigen, M65-antigen and HMGB1) for HBV ACLF. In this prospective study (2/2013–8/2014), 94 patients including 54 HBV-ACLF and 40 chronic hepatitis B (CHB) patients were recruited. 40 healthy controls (HC) were also recruited. HBV-ACLF were followed up for 3 months for short-term mortality. All three biomarkers were significantly elevated in HBV-ACLF compared with CHB or HC. M30- and M65-antigens could significantly discriminate between non-survivors and survivors in HBV-ACLF. However, HMGB1 showed no prognostic value. By Cox regression analysis, M30- and M65-antigens and MELD were identified as independent predictors for short-term mortality. A novel prognostic model, MELD-CD (MELD-cell death) was established based on the multivariate results. The adjusted Harrell’s C-index of MELD-CD was 0.86 (P < 0.001) and was significantly higher (P < 0.001 for all) than the currently used models, MELD (C-index, 0.71, P < 0.001), MELD-NA (0.67, P < 0.001), CTPs (0.61, P < 0.05). Dynamic analyses further confirmed the prognostic utility of M30- and M65-antigen. Future studies are warranted to validate the results.

Persistently elevated circulating Th22 reversely correlates with prognosis in HBV-related acute-on-chronic liver failure.

Hepatitis B virus‐related acute‐on‐chronic liver failure (HBV‐ACLF) is an acute deterioration of liver function on chronic liver disease with immune disorder. Th22 cells and IL‐22 were correlated with inflammatory and autoimmune diseases. However, Th22 cells and IL‐22 in the pathogenesis of HBV‐ACLF remains to be elucidated. It was investigated the correlation between Th22 and prognosis in HBV‐ACLF.

Identification of acetyltransferase genes (HAT1 and KAT8) regulating HBV replication by RNAi screening

BackgroundThe initiation of hepatitis B virus (HBV) replication involves the formation of covalently closed circular DNA (cccDNA) and its transcription into pregenomic RNA (pgRNA) in hepatocyte nuclei. The regulatory mechanism of HBV replication by acetyltransferase is thus far not well understood, but human acetyltransferase has been reported as being involved in the regulation of HBV replication.ResultsDepletion of KAT8 or HAT1 via RNA interference (RNAi) markedly down-regulated HBV-DNA and pgRNA levels in HepG2.2.15 cells, with KAT8 knockdown reducing both HBsAg and HBeAg more than HAT1 knockdown. Consistent with these observations, HBV replication regulators hepatocyte nuclear factor-4-α (HNF4α) and peroxisome proliferator-activated receptor gamma coactivator- (PPARGC-) 1-α were decreased following knockdown of HAT1 or KAT8.ConclusionsThese data suggest that KAT8 or HAT1 regulate HBV replication and may be potential drug targets of anti-HBV therapy.

Randomized clinical trial: Nucleos(t)ide analogues improved survival of CHB-related HCC patients via reducing severity and progression of malignancy

Background The influence of nucleos(t)ide analogues (NAs) to treat Chronic hepatitis B (CHB) related hepatocellular carcinoma (HCC) remains to be explored. Aim To investigate if NAs reduce the severity and progression of CHB-related HCC. Results Among 532 patients, there were 118 or 414 CHB-related HCC with or without NAs therapy, respectively. BCLC scores, serum level of ALT/AST and HBV DNA were compared. During follow-up, the survival period of CHB-related HCC patients with sustained NAs is significantly longer than that with NAs post-HCC and NAs naïve (p < 0.05). Factors significantly associated with the poor overall survival of CHB-related HCC include BCLC scores (hazard ratio, 1.84 [95% confidence interval, 1.57−2.15], p < 0.001), NAs post-HCC or NAs naïve (1.33 [1.07−1.65], p < 0.01), serum AST ≥ 40 IU/L (1.48 [1.03−2.12], p < 0.05) and HBV DNA ≥ 104 copies/ml (1.36 [1.01−1.83], p < 0.001). Methods Outcomes of 532 CHB-related HCC patients with/without NAs were investigated. Overall survival of CHB-related HCC patients, NAs naïve (n = 156), NAs received post-HCC (n = 258) and NAs sustained (n = 118) were determined. Conclusions NAs reduced severity of CHB-related HCC patients. Sustained NAs is an important factor associated with the extended survival of CHB-related HCC patients.

Serum hepatocyte apoptosis biomarker predicts the presence of significant histological lesion in chronic hepatitis B virus infection.

BACKGROUND Hepatocyte death, either apoptosis or necrosis, is closely associated with hepatic inflammation and fibrosis. AIMS To investigate the potential values of hepatocytes death biomarker, M30 (apoptosis) and M65 (total death) in predicting histological lesions in chronic hepatitis B virus (HBV) infection. METHODS Total 201 treatment-naïve patients were prospectively recruited. Liver biopsies were performed prior to antiviral treatments for treatments starting evaluation. Sera were collected on the day of liver biopsy for biomarker measurements. Sera from 200 age-matched healthy volunteers served as healthy controls (HCs). RESULTS Significant histological lesions (SHL, i.e. significant inflammation and/or significant fibrosis) were confirmed in 150 (74.63%) patients. There were significantly higher serum M30 and M65 in patients with SHL than those without SHL (p<0.001) or than HCs (p<0.001). Serum M30, but not M65, independently predicted SHL [odds ratio:3.4 (95% CI, 1.8-6.2) per increase of 50U/L, p<0.001] after adjusting other potential confounding factors. A novel model based on M30 provided good diagnostic performance in predicting SHL [AUC, 0.87 (0.81-0.92)]. Cut-off value of >0 to confirm or ≤-0.5 to exclude SHL has ∼12% misclassification rate. CONCLUSION Hepatocyte apoptosis biomarker, M30 is a promising non-invasive alternative to liver biopsy in chronic HBV infection upon treatment evaluation.

Serum soluble ST2 is a promising prognostic biomarker in HBV-related acute-on-chronic liver failure

BACKGROUND The IL-33/ST2 axis is involved in the pathogenesis of many diseases such as autoimmune diseases, cancer, and heart failure. However, studies of the IL-33/ST2 pathway in HBV-related acute-on-chronic liver failure (HBV-ACLF) are lacking. The present study aimed to determine the prognostic role of serum IL-33/soluble ST2 (sST2) in HBV-ACLF. METHODS Serum levels of IL-33 and sST2 in healthy controls (HC, n=18), chronic hepatitis B (CHB, n=27) and HBV-ACLF (n=51) patients at the 1st and 4th week after enrollment were detected using ELISA, and clinical data were collected. The follow-up of HBV-ACLF patients lasted for 6 months at least. RESULTS There was no significant difference of serum IL-33 level among HC, CHB and HBV-ACLF patients at week 1. However, serum sST2 level differed significantly among the three groups: highest in the HBV-ACLF group, moderate in the CHB group and lowest in the HC group. There was a reverse correlation between serum sST2 level and the survival of HBV-ACLF patients. The level of serum sST2 in HBV-ACLF survivors was significantly declined from week 1 to week 4 following the treatment, whereas that in HBV-ACLF non-survivors remained at a high level during the same period. Furthermore, serum sST2 level was significantly correlated with laboratory parameters and the most updated prognostic scores (CLIF-C OF score, CLIF-C ACLF score and ACLF grades). The receiver operating characteristics curves demonstrated that serum sST2 level was a good diagnostic marker for predicting the 6-month mortality in HBV-ACLF patients, comparable to the most updated prognostic scores. Serum sST2 cut-off points for predicting prognosis in HBV-ACLF patients were 76 ng/mL at week 1 or 53 ng/mL at week 4, respectively. HBV-ACLF patients with serum sST2 level above the cut-off point often had a worse prognosis than those below the cut-off point. CONCLUSION Serum sST2 may act as a promising biomarker to assess severity and predict prognosis of patients with HBV-ACLF and help for the early identification and optimal treatment of HBV-ACLF patients at high risk of mortality.