BACKGROUND/AIMS: We aimed to clarify the association of hepatitis B surface antigen (HBsAg)/hepatitis B core antigen (HBcAg) with the disease status and treatment response in patients with chronic hepatitis B (CHB). METHODS: We investigated 171 biopsy-proven entecavir-treated CHB patients (109 hepatitis B e antigen [HBeAg]-positive, 62 HBeAg-negative). HBcAg expression was positive when ≥10% of hepatocytes stained, and classified into nuclear, mixed, and cytoplasmic patterns. HBsAg expressions were intracytoplasmic (diffuse, globular, and submembranous) and membranous. The histologic activity index (HAI) and fibrosis stage followed Ishak system. RESULTS: In HBeAg-positive patients, older age, increased HAI score, advanced fibrosis, and reduced viral load were observed when HBcAg expression shifted from nucleus to cytoplasm in HBcAg-positive patients, and HBsAg expression from non-submembranous to submembranous in HBcAg-negative patients (all, p<0.05). In HBeAg-negative patients, only intracytoplasmic HBsAg expression patterns had clinical relevance with decreased ALT levels and viremia. In HBeAg-positive patients without favorable predictors of virologic response, negative HBcAg and membranous HBsAg expression predicted greater virologic response (both, p<0.05). The probability of HBeAg seroclearance was higher in patients with increased HAI or lacking HBcAg expression (both, p<0.05). Higher serum HBsAg levels and hepatocyte HBcAg positivity were associated with reduced serum HBsAg during first and post-first year treatment, respectively (both, p<0.05). CONCLUSIONS: Hepatocyte HBcAg/HBsAg expression is a good marker for disease status and predicting treatment response.
Cytoplasm ; Fibrosis ; Hepatitis B Core Antigens* ; Hepatitis B e Antigens ; Hepatitis B Surface Antigens* ; Hepatitis B* ; Hepatitis B, Chronic* ; Hepatitis* ; Hepatitis, Chronic* ; Hepatocytes* ; Humans ; Viral Load ; Viremia

Alanine Transaminase ; blood ; DNA, Viral ; blood ; Hepatitis B Antibodies ; blood ; Hepatitis B Core Antigens ; blood ; Hepatitis B Surface Antigens ; blood ; Hepatitis B e Antigens ; blood ; Hepatitis B virus ; genetics ; immunology ; Hepatitis B, Chronic ; virology ; Humans

BACKGROUND: Hepatitis B core-related antigen (HBcrAg) is a promising disease-monitoring marker for chronic hepatitis B (CHB). We investigated correlations between HBcrAg with antiviral efficacy and virological and histological variables. METHODS: One hundred and forty-five CHB patients from the mainland of China between August 2013 and September 2016 who underwent liver biopsy received entecavir therapy and had paired liver biopsy at 78 weeks. We analyzed correlations between HBcrAg and virological and histological variables in hepatitis B e antigen (HBeAg)-positive and HBeAg-negative patients. We also explored the predictors of HBeAg loss after 78 weeks of antiviral therapy. Pearson correlation analysis and logistic forward stepwise regression were the main statistic methods. RESULTS: HBeAg-positive patients (n = 93) had higher baseline HBcrAg (median 7.4 vs. 5.3 log10 U/mL P < 0.001) and greater HBcrAg declines (median 1.6 vs. 0.9 log10 U/mL P = 0.007) than HBeAg-negative patients after 78 weeks of therapy. At baseline, HBcrAg correlated with hepatitis B virus (HBV) DNA in both HBeAg-positive (r = 0.641, P < 0.001) and -negative patients (r = 0.616, P < 0.001), with hepatitis B surface antigen (HBsAg) in HBeAg-positive patients (r = 0.495, P < 0.001), but not with anti-hepatitis B virus core antibody (anti-HBc). Weak correlations existed between HBcrAg, histology activity index (HAI; r = 0.232, P = 0.025), and Ishak fibrosis score (r = -0.292, P = 0.005) in HBeAg-positive patients. At 78 weeks, significant correlations existed only between HBcrAg and anti-HBc in HBeAg-positive (r = -0.263, P = 0.014) and HBeAg-negative patients (r = -0.291, P = 0.045). Decreased HBcrAg significantly correlated with reduced HBV DNA (r = 0.366, P = 0.001; r = 0.626, P < 0.001) and HBsAg (r = 0.526, P = 0.001; r = 0.289, P = 0.044) in HBeAg-positive and -negative patients, respectively, and with reduced HAI in HBeAg-positive patients (r = 0.329, P = 0.001). Patients with HBeAg loss (n = 29) showed a larger reduction in HBcrAg than those without (median 2.3 vs. 1.3 log10 U/mL, P = 0.001). In multivariate analysis, decreased HBcrAg was an independent predictor of HBeAg loss (P = 0.005). CONCLUSIONS: HBcrAg reflects viral replication and protein production. Decreased HBcrAg could predict HBeAg loss after antiviral therapy. TRIAL REGISTRATION Clinical Trials.gov: NCT01962155; https://www.clinicaltrials.gov/ct2/show/NCT01962155?term=NCT01962155&draw=2&rank=1.
Antiviral Agents/therapeutic use* ; Biomarkers ; China ; DNA, Viral ; Hepatitis B Core Antigens/therapeutic use* ; Hepatitis B Surface Antigens ; Hepatitis B e Antigens ; Hepatitis B virus/genetics* ; Hepatitis B, Chronic/drug therapy* ; Humans ; Virus Replication

As part of the immunoserology program of the Korean Association of External Quality Assessment Service, we organized two trials on the external quality assessment of hepatitis viral markers in 2016 and 2017. The hepatitis viral antigens and antibodies program consisted of 10 test items. We delivered two and three types of pooled sera specimens to 965 and 965 institutions for the first and second trials of external proficiency testing in 2016, respectively. The number of participating laboratories was 915 (94.8%) and 913 (95.0%) in the first and second trials in 2016, respectively. We also delivered three kinds of pooled sera specimens to 936 and 1,015 institutions for the first and second trials of external proficiency testing in 2017, respectively. The number of participating laboratories was 920 (98.3%) and 996 (98.1%) in the first and second trials in 2017, respectively. The most commonly tested items were hepatitis B surface antigen, followed by the antibodies to hepatitis B surface antigen, anti-hepatitis C virus, hepatitis B envelope antigen, antibodies to hepatitis B envelope antigen, anti-hepatitis A virus and antibodies to hepatitis B core antigen. The most frequently used methods for detecting viral markers were the chemiluminescence immunoassay and the electrochemiluminescence immunoassay, but they yielded a few-false positive results due to the matrix effect. The immunochromatographic assay yielded false-negative results for anti-hepatitis A virus due to low sensitivity. Continuous improvement in the quality of viral hepatitis testing through participation in the survey seems necessary.
Antibodies ; Antigens, Viral ; Biomarkers* ; Hepatitis A ; Hepatitis B ; Hepatitis B Core Antigens ; Hepatitis B Surface Antigens ; Hepatitis B virus ; Hepatitis C ; Hepatitis* ; Immunoassay ; Immunochromatography ; Korea* ; Laboratory Proficiency Testing ; Luminescence

<b>OBJECTIVEb>To establish an in vitro model of hepatitis B virus (HBV) replication and expression in primary rat hepatocytes (PRH) transfected with circular viral DNA for further study on the interaction of HBV with hepatocytes.

<b>METHODSb>Circular viral DNA containing complete HBV genome were transfected into PRH by electroporation (transfected group, about 4mug of circular viral DNA/1 10(7)cells). From day 1 to day 10 after transfection, HBsAg and HBeAg in the supernatants and lysates of PRH were measured with IMX system. HBcAg was assayed with western blotting, immunol dot blotting and immunocytochemistry. Meanwhile, HBV S-mRNA and X-mRNA were tested with RT-PCR, and replicative intermediates of HBV DNA were analyzed by southern blotting and dot blotting. Moreover, Transmission electron microscopy was used if viral particles were produced in transfected rat hepatocytes. PRH electroporated only was used as control group.

<b>RESULTSb>(1) Viral antigen production in transfected rat hepatocytes: HBsAg in cell lysates was positive. P/N values ranged from 4.83 to 85.69, and could be maintained for 10 days after transfection. The average P/N values was 18.239 27.459. Whereas, HBsAg was negative in the supernatants of transfected group (P/N values, negative<2.1). HBeAg in the supernatants and lysates of transfected hepatocytes all was negative (P/N values<2.1) during 10 days following transfection. HBcAg was only found positive in transfected hepatocytes by immunol dot blotting. (2) Detection of viral transcripts: transcription of HBV DNA was investigated by preparing total RNA from rat hepatocytes 2 days after transfection and looking for S-mRNA and X-mRNA by RT-PCR. Results showed S-mRNA positive, X-mRNA negative. (3) HBV DNA replication analysis: intracellular total DNA was extracted 2 days after transfection and analysed by southern blotting. All replicative DNA intermediates, including relaxed circular (rcDNA), covalently closed circular (cccDNA), and single-stranded (ssDNA) linear HBV DNA forms, were indicated. Dot blotting showed intracellular HBV DNA positive in transfected group during 10 days after transfection. However, viral particles were not found in transfected hepatocytes during 3 days after transfection.

<b>CONCLUSIONSb>Circular HBV DNA transfected into primary adult rat hepatocytes could obtain continuous replication and stable expression of HBV surface antigen. This in vitro model has high reproducibility and stability, and is useful for directly studying the interaction of HBV with hepatocytes.

Animals ; DNA Replication ; DNA, Circular ; genetics ; DNA, Viral ; genetics ; Gene Expression ; Hepatitis B Core Antigens ; analysis ; Hepatitis B Surface Antigens ; analysis ; Hepatitis B e Antigens ; analysis ; Hepatitis B virus ; genetics ; Hepatocytes ; virology ; Male ; Rats ; Rats, Wistar ; Transfection ; Virus Replication

BACKGROUND: Cutaneous vasculitis associated with viral hepatitis seems to occur as a hypersensitivity reaction against the circulating viral antigens. Hepatitis B virus(HBV)-encoded X antigen(HBxAg) is known to participate in the carcinogenesis of hepatocellular carcinoma(HCC) by the inactivation of p53. However, HBxAg has been found in chronic infiammatory lesions without the overexpression of p53. Accordingly, not only EBsAg and HBcAg but also HBxAg may be involved in HCC-associated cutaneous vasculitis, regardless of the alteration of p53. OBJECTIVE: This study was conducted to investigate the expression of HBV-encoded antigens in cutaneous vasculitis accompanied by HBV hepatopathy. Additionally, we have compared the expression of 3 HBV antigens and p53 between vasculitic patients with HCC and in others showing HCC-non-associated vasculitis. METHODS: Immunohistochemically, we examined the expression of HBsAg, HBcAg, and HBxAg in the tissue specimens taken from the vasculitic lesions of the 33 HBsAg-positive enrolled patients with cutaneous vasculitis proven by skin biopsy. RESULTS: 1. The immunohistochemical positivity rate to HBsAg in vasculitic patients with HBV hepatopathy was 66.7% overall. It was 90% in HCC-associated vasculitic subjects and 56.5% in the vasculitic subjects without HCC, respectively. 2. We found the expression of HBxAg in 80% of the vasculitic subjects showing HCC. The vasculitic patients without HCC showed 17,3% of the positivity rate to HBxAg. 3. We could not find the overexpression of p53 in the vasculitic tissue specimens of the HCC patients without the cutaneous metastasis from primary HCC. CONCLUSION: HBsAg, HBcAg and HBxAg may participate in the pathogenesis of cutaneous vasculitis with HBV hepatopathy, regardless of tumorigenesis.
Antigens, Viral ; Biopsy ; Carcinogenesis ; Hepatitis B Core Antigens ; Hepatitis B Surface Antigens ; Hepatitis B virus* ; Hepatitis B* ; Hepatitis* ; Humans ; Hypersensitivity ; Neoplasm Metastasis ; Skin ; Vasculitis*

Biomarkers ; blood ; DNA, Viral ; blood ; Female ; Hepatitis B ; genetics ; immunology ; prevention & control ; Hepatitis B Antibodies ; biosynthesis ; Hepatitis B Core Antigens ; immunology ; Hepatitis B Surface Antigens ; immunology ; Hepatitis B Vaccines ; immunology ; Hepatitis B e Antigens ; blood ; immunology ; Hepatitis B virus ; isolation & purification ; Humans ; Male ; Vaccination

BACKGROUND: Cutaneous vasculitis associated with viral hepatitis seems to occur as a hypersensitivity reaction against the circulating viral antigens. Hepatitis B virus(HBV)-encoded X antigen(HBxAg) is known to participate in the carcinogenesis of hepatocellular carcinoma(HCC) by the inactivation of p53. However, HBxAg has been found in chronic infiammatory lesions without the overexpression of p53. Accordingly, not only EBsAg and HBcAg but also HBxAg may be involved in HCC-associated cutaneous vasculitis, regardless of the alteration of p53. OBJECTIVE: This study was conducted to investigate the expression of HBV-encoded antigens in cutaneous vasculitis accompanied by HBV hepatopathy. Additionally, we have compared the expression of 3 HBV antigens and p53 between vasculitic patients with HCC and in others showing HCC-non-associated vasculitis. METHODS: Immunohistochemically, we examined the expression of HBsAg, HBcAg, and HBxAg in the tissue specimens taken from the vasculitic lesions of the 33 HBsAg-positive enrolled patients with cutaneous vasculitis proven by skin biopsy. RESULTS: 1. The immunohistochemical positivity rate to HBsAg in vasculitic patients with HBV hepatopathy was 66.7% overall. It was 90% in HCC-associated vasculitic subjects and 56.5% in the vasculitic subjects without HCC, respectively. 2. We found the expression of HBxAg in 80% of the vasculitic subjects showing HCC. The vasculitic patients without HCC showed 17,3% of the positivity rate to HBxAg. 3. We could not find the overexpression of p53 in the vasculitic tissue specimens of the HCC patients without the cutaneous metastasis from primary HCC. CONCLUSION: HBsAg, HBcAg and HBxAg may participate in the pathogenesis of cutaneous vasculitis with HBV hepatopathy, regardless of tumorigenesis.
Antigens, Viral ; Biopsy ; Carcinogenesis ; Hepatitis ; Hepatitis B ; Hepatitis B Core Antigens ; Hepatitis B Surface Antigens ; Humans* ; Hypersensitivity ; Malassezia* ; Neoplasm Metastasis ; Skin* ; Vasculitis ; Yeasts*

<b>OBJECTIVEb>To clarify the relationship between fatal severe form hepatitis occurred during chronic hepatitis B and superinfections of hepatitis A, C, D or E virus as well as hepatitis B e system status and to adopt corresponding measures to reduce the mortality of chronic hepatitis B.

<b>METHODSb>This study detected the superinfections with hepatitis A, C, D or E virus and hepatitis B e system status in 219 patients with fatal severe form hepatitis occurred during chronic hepatitis B by enzyme linked immunosorbent assay.

<b>RESULTSb>The superinfections with hepatitis A, C, D or E virus were found in 1.4% (3/219), 9.6% (21/219), 1.8% (4/219) and 30.1% (66/219) of the patients, respectively, altogether 42.9% (94/219); hepatitis E was prominent and steady in superinfection rate in recent ten years. The causes of 57.1% (125/219) patients were not clear. The positive rate of HBeAg and anti-HBe were 17.0% (16/94) and 54.2% (51/94) in the group of superinfections with hepatitis A, C, D or E virus; and were 27.2% (34/125) and 47.2% (59/125) in the group with unknown causes, respectively.

<b>CONCLUSIONb>These results suggested that the patients with superinfections reached 42.9% (94/219), and the superinfections may be a part of causes of fatal severe form hepatitis, and the mortality of chronic hepatitis B may be decreased by strict food sanitation and use of safe blood products. There were no significant relation between hepatitis B e antigen seroconversion and the fatal severe form hepatitis occurred during chronic hepatitis B.

Adult ; DNA, Viral ; blood ; genetics ; Female ; Hepacivirus ; genetics ; physiology ; Hepatitis A virus ; genetics ; physiology ; Hepatitis B Core Antigens ; blood ; Hepatitis B Surface Antigens ; blood ; Hepatitis B e Antigens ; blood ; Hepatitis B virus ; genetics ; immunology ; physiology ; Hepatitis B, Chronic ; blood ; mortality ; virology ; Hepatitis Delta Virus ; genetics ; physiology ; Hepatitis E virus ; genetics ; physiology ; Host-Pathogen Interactions ; Humans ; Male ; Middle Aged ; Superinfection ; virology ; Survival Rate

Occult HBV infection is characterized by the absence of serum HBsAg with persistence of low level of intrahepatic HBV DNA. Several suggested mechanisms for the origin of occult HBV infection include strong suppression of viral replication and gene expression, mutation in the regulatory regions of HBV genome, formation of immunoglobulin-bound HBsAg, viral interference, and blockage of HBsAg secretion from infected hepatocytes. Standardized assays are not yet available, and sensitive HBV DNA amplification assay is necessary for the diagnosis of cryptic infection. Detection rate of HBV DNA is highest in IgG anti-HBc positive population. However, neither anti-HBc nor anti-HBs can be detected in a significant proportion of infected persons. Occult HBV infection occurs in a number of clinical settings and is highly prevalent in HCV-infected patients as well as in patients with cryptogenic chronic liver disease including hepatocellular carcinoma.
DNA, Viral/analysis ; Hepatitis B/*diagnosis/*epidemiology/metabolism ; Hepatitis B Antibodies/blood ; Hepatitis B Core Antigens/immunology ; Hepatitis B Surface Antigens/blood ; Humans