Asymptomatic Hepatitis B Virus Carriers Research Articles

Phenotypes of peripheral CD4+ T helper cell subsets in pregnant women with HBeAg-negative chronic asymptomatic HBV carriers.

Chronic hepatitis B virus (HBV) infection is a major public health problem worldwide, and mother-to-child transmission is the key mode of HBV infection. CD4+ T helper (Th) cells play a critical role in the immune microenvironment of specific maternal tolerance to the foetus during pregnancy. However, the roles of Th cell subsets in pregnant women (PW) with chronic asymptomatic HBV carriers (ASCs) remain completely unclear. Here, we aimed to characterize CD4+ T-cell immunity in PW with hepatitis Be antigen (HBeAg)-negative chronic ASCs. Human peripheral blood mononuclear cells (PBMCs) from PW without HBV infection or with chronic ASCs and healthy controls (HC) were isolated, and CD4+ Th cell subsets were detected by flow cytometry in addition to serum cytokines. Serological HBV markers, liver function and hormone levels of these individuals were also tested. The frequencies of circulating T follicular helper (Tfh) type 2 (Tfh2) cells were significantly evaluated, but Tfh1 cell frequencies were notably decreased in PW compared to HC. Moreover, the frequencies of Th22 cells were only notably increased in PW with chronic ASCs in comparison with PW. Additionally, increased levels of serum IL-4 were positively correlated with Tfh2 cell frequencies in healthy PW. Interestingly, serum P4 levels were positively associated with the frequencies of circulating Tfh2 or Th2 cells but were negatively related to the frequencies of circulating Tfh17 or Th17 cells in healthy PW. Although there were some changes in the other CD4+ Th cell frequencies and cytokine levels or other references, significant differences were not found among HC, healthy PW, PW with HBeAg-negative chronic ASCs. CD4+ Th cell subsets played a critical role in the immune microenvironment of PW, and these findings provided potential evidence for why PW with chronic ASCs did not receive antenatal antiviral prophylaxis.

Immune Checkpoint Molecules Expressed on CD4+ T Cell Subsets in Chronic Asymptomatic Hepatitis B Virus Carriers With Hepatitis B e Antigen-Negative.

BackgroundChronic hepatitis B virus (HBV) infection remains a major public health problem worldwide. Immune checkpoint molecules expressed on CD4+ T cells play critical roles in chronic HBV infection. However, their roles in chronic asymptomatic HBV carriers (ASCs) with hepatitis B e antigen (HBeAg)-negative remain unclear. In this study, we explored the role of immune checkpoint molecules expressed on CD4+ T cell subsets in chronic ASCs with HBeAg-negative.MethodsHuman peripheral blood mononuclear cells (PBMCs) from the ASCs with HBeAg-negative and healthy controls (HC) were isolated, and immune checkpoint molecules expressed on CD4+ T cell subsets and serum cytokines were detected by flow cytometry. Moreover, the mRNA expressions of immune checkpoint molecules were analyzed by a real-time quantitative PCR assay.ResultsIn comparison with HC, CD4+ T cells highly expressed LAG-3, TIM-3, and PD-1 in PBMCs from chronic ASCs with HBeAg-negative. Interestingly, the expressions of TIM-3 and PD-1 on circulating follicular helper T (Tfh) cells in ASCs were significantly high. Moreover, high expressions of LAG-3, TIM-3, and PD-1 were different among Treg, Th1, Th2, and Th17 cells. In addition, the expressions of TIM-3 and CTLA-4 mRNA in PBMCs from ASCs were significantly elevated. However, the frequency of CTLA-4+CD4+ T cell subsets in PBMCs from ASCs was not different from HC. The levels of six cytokines in serum from ASCs were not clearly different from HC.ConclusionImmune checkpoint molecules highly expressed on CD4+ T cell subsets indicated an important role in chronic ASCs with HBeAg-negative, which provided potential therapeutic targets in the pathogenesis of chronic HBV infection.

Vagaries of the Host Response to Chronic Hepatitis B Virus Infection: What is the Ultimate Outcome of So-called “Asymptomatic HBV Carriers” Observed Over Several Decades?

Introduction: Chronic hepatitis B virus (HBV) infection is prevalent worldwide and up to 40% is known to progress to serious complications including cirrhosis and hepatocellular carcinoma (HCC). The outcome of the remaining infected individuals is not well documented. Our case series describes a longer cohort of chronic HBV infections that have remained asymptomatic with no progression of liver disease. Case Series: Thirty-three patients (ages 31-84) with chronic HBV infection were identified. All patients had no significant elevations in transaminase levels and were followed over 32 years, collectively. 18/33 had a fluctuating greater magnitude of HBV viral load with no elevations in tumor marker or significant radiographic changes to their liver. Discussion/Conclusion: Chronic HBV infection can lead to serious complications over time, the mechanism of which are not well understood. The variation in patients that do and do not develop these complications stresses the importance of the individual response to the virus and may highlight host immune response differences.

Interleukin-18 Plays a Positive Feedback Activity to Natural Killer-Like B Cells in Hepatitis B Virus-Associated Acute-on-Chronic Liver Failure.

Natural killer-like B (NKB) cells are a newly identified immune subset, which are separated from NK cells and B cells. NKB cells demonstrated immunoregulatory functions in elimination of microbial infection and inflammation through secretion of interleukin (IL)-12 and IL-18. However, the role of NKB cells in hepatitis B virus (HBV)-related diseases has not been reported. In this study, peripheral T cells, B cells, NK cells, and NKB cells in HBV-associated acute-on-chronic liver failure (ACLF), chronic hepatitis B (CHB), asymptomatic HBV carriers (AsC), and controls were investigated by flow cytometry. Plasma IL-12 and IL-18 levels were measured by enzyme-linked immunosorbent assay. Peripheral blood mononuclear cells from HBV-ACLF patients were stimulated with recombinant IL-12 or IL-18. Changes of immune cell percentage and nuclear factor-kappa B (NF-κB) phosphorylation were assessed. There were no statistical differences of T cell percentage, B cell percentage, or NK cell percentage among groups. NKB cell percentage within lymphocytes, and plasma IL-12 and IL-18 levels in HBV-ACLF patients were significantly elevated compared with in CHB, AsC, and controls. NKB cell percentage and IL-18, but not IL-12, had a better prognosis function for the 28-day survival status in HBV-ACLF patients. Recombinant IL-12 enhanced T cell and NK cell percentage, while only high concentrations (10 ng/mL) of IL-18 promoted NKB cell percentage in HBV-ACLF patients. High concentrations (10 ng/mL) of IL-18 induced NF-κB phosphorylation in NKB cells probably through suppression of IL-18 binding protein in HBV-ACLF patients. The current data indicated that elevated NKB cells and IL-18 might be important indicators for poor prognosis of HBV-ACLF patients. Increased IL-18 might play a positive feedback activity to NKB cells in HBV-ACLF patients.

Imbalance Between Soluble and Membrane-Bound CD100 Regulates Monocytes Activity in Hepatitis B Virus-Associated Acute-on-Chronic Liver Failure

CD100 is an important immune semaphorin that is a secreted and membrane bound protein involved in infectious diseases. However, CD100 expression profile and the regulation to innate immune system in hepatitis B virus (HBV)-associated acute-on-chronic liver failure (ACLF) was not previously reported. The aim of this study was to investigate CD100 level and modulatory function of CD100 to CD14+ monocytes in HBV-ACLF patients. Plasma-soluble CD100 (sCD100) level and membrane-bound CD100 (mCD100) expression on peripheral CD14+ monocytes was analyzed in HBV-ACLF patients. CD14+ monocytes-induced cytotoxicity and CD14+ monocytes-mediated T cell activation in response to CD100 stimulation was also assessed in direct and indirect contact coculture culture systems. HBV-ACLF patients had lower plasma sCD100 and higher mCD100 level on CD14+ monocytes compared with asymptomatic HBV carriers, chronic hepatitis B patients, and controls. CD14+ monocytes from HBV-ACLF patients induced limited target Huh7.5 cell death and secreted less interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α), and granzyme B in both direct and indirect contact coculture systems compared with controls. Recombinant sCD100 not only enhanced CD14+ monocytes-mediated Huh7.5 cell death and granzyme B secretion, but it also elevated CD14+ monocytes-induced IFN-γ/interleukin-17 production by CD4+ T cells as well as IFN-γ/TNF-α secretion by CD8+ T cells in HBV-ACLF patients. The current data indicated that severe inflammation induced sCD100/mCD100 imbalance to inactivate CD14+ monocytes response, which might be beneficial for the survival of HBV-ACLF patients.

Copy number gain of pro-inflammatory genes in patients with HBV-related acute-on-chronic liver failure

BackgroundHost genetic factors such as single nucleotide variations may play a crucial role in the onset and progression of HBV-related acute-on-chronic liver failure (ACLF). However, the underlying genomic copy number variations (CNVs) involved in the pathology are currently unclear.MethodsWe genotyped two cohorts with 389 HBV-related ACLF patients and 391 asymptomatic HBV carriers (AsCs), and then carried out CNV-based global burden analysis and a genome-wide association study (GWAS).ResultsFor 1874 rare CNVs, HBV-related ACLF patients exhibited a high burden of deletion segments with a size of 100–200 kb (P value = 0.04), and the related genes were significantly enriched in leukocyte transendothelial migration pathway (P value = 4.68 × 10–3). For 352 common CNVs, GWAS predicted 17 significant association signals, and the peak one was a duplication segment located on 1p36.13 (~ 38 Kb, P value = 1.99 × 10–4, OR = 2.66). The associated CNVs resulted in more copy number of pro-inflammatory genes (MST1L, DEFB, and HCG4B) in HBV-related ACLF patients than in AsC controls.ConclusionsOur results suggested that the impact of host CNV on HBV-related ACLF may be through decreasing natural immunity and enhancing host inflammatory response during HBV infection. The findings highlighted the potential importance of gene dosage on excessive hepatic inflammation of this disease.

Potential role of\xa0intestinal microflora\xa0in disease progression among patients with different stages of Hepatitis B

BackgroundIncreasing evidence demonstrate that the gut microbiota is involved in the pathogenesis of liver diseases, and faecal microbiota transplantation is considered to be a promising new treatment option. However, there are no reports on the intestinal flora of asymptomatic HBV carriers using next-generation sequencing. This study intends to investigate the potential role of the intestinal microflora in predicting the progression of Hepatitis B patients in different non-cancerous stages.ResultsA total of 266 patients with different stages of Hepatitis B and 31 healthy controls were included in this study. Some of the subjects (217 cases) underwent 16S rRNA gene sequencing. Compared with the control group (CK), the α diversity of patients in Group A (HBV carrier) slightly increased, while that of patients in the other three groups decreased. Each group of patients, especially those in Group C (cirrhosis) and Group D (acute-on-chronic liver failure), could be separated from the CK using weighted UniFrac PCoA and ANOSIM. LEfSe revealed that 40 taxa belonging to three phyla had an LDA larger than 4. In addition to the comparison between Group B (chronic Hepatitis B) and Group C, the specific flora and potential taxonomic function were also identified. Different microbial communities were found to be highly correlated with clinical indicators and the Child-Pugh scores. Changes in the microbial community were highly related to the alternations of host metabolism, which in turn, was related to the development of Hepatitis B. Our analysis identified a total of 47 strains with potential biomarker functions at all levels except for the phylum level.ConclusionsFaecal microbiota transplantation of some potential beneficial bacteria can change with the occurrence of disease, and HBV carriers might be the most suitable donors.

Circulating CTLA-4 levels and CTLA4 polymorphisms associate with disease condition and progression and hepatocellular carcinoma patients' survival in chronic hepatitis B virus infection

BackgroundCTLA-4 is involved in the immune dysfunction of hepatitis B virus (HBV) infection and hepatocellular carcinoma (HCC). This study analyzed the association of circulating CTLA-4 levels and CTLA4 polymorphisms with disease condition and progression in chronic HBV infection. MethodsSerum CTLA-4 levels and CTLA4 rs231775 and rs5742909 polymorphisms were determined in patients with various HBV-related diseases [53 asymptomatic HBV carrier status (ASC), 147 chronic hepatitis, 130 cirrhosis and 102 HCC] and nearly a 10-year follow-up. ResultsSerum CTLA-4 levels were stepwisely increased from ASC, chronic hepatitis, cirrhosis to HCC and independently associated with HCC (OR 2.628, P < 0.001). HCC patients had lower frequencies of rs231775 genotype GA, genotype AA and allele A than ASC, chronic hepatitis and cirrhosis patients. Rs231775 genotype GG was independently associated with HCC (OR 2.324, P = 0.010) and higher CTLA-4 levels in patients with HBV infection. In the follow-up, higher baseline CTLA-4 levels and CTLA4 rs231775 genotype GG significantly associated with disease progression from chronic hepatitis to cirrhosis (OR 2.561, P = 0.011 and OR 2.799, P = 0.015, respectively) or from cirrhosis to HCC (OR 2.673, P = 0.008 and OR 2.097, P = 0.023, respectively) and with a shorter overall survival in HCC patients (HR 0.317, P = 0.018 and HR 0.682, P = 0.026, respectively). Rs5742909 had no significant association with CTLA-4 levels and disease progression. ConclusionCTLA-4 levels and CTLA4 rs231775 polymorphism associate with the disease condition and progression and HCC development in chronic HBV infection and their determination may be used for monitoring disease progression and predicting patient prognosis.

Hepatitis B Virus (HBV) Subviral Particles as Protective Vaccines and Vaccine Platforms

Hepatitis B remains one of the major global health problems more than 40 years after the identification of human hepatitis B virus (HBV) as the causative agent. A critical turning point in combating this virus was the development of a preventative vaccine composed of the HBV surface (envelope) protein (HBsAg) to reduce the risk of new infections. The isolation of HBsAg sub-viral particles (SVPs) from the blood of asymptomatic HBV carriers as antigens for the first-generation vaccines, followed by the development of recombinant HBsAg SVPs produced in yeast as the antigenic components of the second-generation vaccines, represent landmark advancements in biotechnology and medicine. The ability of the HBsAg SVPs to accept and present foreign antigenic sequences provides the basis of a chimeric particulate delivery platform, and resulted in the development of a vaccine against malaria (RTS,S/AS01, MosquirixTM), and various preclinical vaccine candidates to overcome infectious diseases for which there are no effective vaccines. Biomedical modifications of the HBsAg subunits allowed the identification of strategies to enhance the HBsAg SVP immunogenicity to build potent vaccines for preventative and possibly therapeutic applications. The review provides an overview of the formation and assembly of the HBsAg SVPs and highlights the utilization of the particles in key effective vaccines.

Genetic Polymorphisms Predisposing the Interleukin 6-Induced APOBEC3B-UNG Imbalance Increase HCC Risk via Promoting the Generation of APOBEC-Signature HBV Mutations.

APOBEC3-UNG imbalance contributes to hepatitis B virus (HBV) inhibition and somatic mutations. We aimed to explore the associations between hepatocellular carcinoma (HCC) risk and genetic polymorphisms predisposing the imbalance.Experimental Design: Genetic polymorphisms at APOBEC3 promoter and UNG enhancer regions were genotyped in 5,621 participants using quantitative PCR. HBV mutations (nt.1600-nt.1945, nt.2848-nt.155) were determined by Sanger sequencing. Dual-luciferase reporter assay was applied to detect the transcriptional activity. Effects of APOBEC3B/UNG SNPs and expression levels on HCC prognosis were evaluated with a cohort of 400 patients with HCC and public databases, respectively. APOBEC3B rs2267401-G allele and UNG rs3890995-C allele significantly increased HCC risk. rs2267401-G allele was significantly associated with the generation of APOBEC-signature HBV mutation whose frequency consecutively increased from asymptomatic HBV carriers to patients with HCC. Multiplicative interaction of rs2267401-G allele with rs3890995-C allele increased HCC risk, with an adjusted OR (95% confidence interval) of 1.90 (1.34-2.81). rs2267401 T-to-G and rs3890995 T-to-C conferred increased activities of APOBEC3B promoter and UNG enhancer, respectively. IL6 significantly increased APOBEC3B promoter activity and inhibited UNG enhancer activity, and these effects were more evident in those carrying rs2267401-G and rs3890995-C, respectively. APOBEC3B rs2267401-GG genotype, higher APOBEC3B expression, and higher APOBEC3B/UNG expression ratio in HCCs indicated poor prognosis. APOBEC-signature somatic mutation predicts poor prognosis in HBV-free HCCs rather than in HBV-positive ones. Polymorphic genotypes predisposing the APOBEC3B-UNG imbalance in IL6-presenting microenvironment promote HCC development, possibly via promoting the generation of high-risk HBV mutations. This can be transformed into specific prophylaxis of HBV-caused HCC.

Interleukin-35 modulates the balance between viral specific CD4+CD25+CD127dim/- regulatory T cells and T helper 17 cells in chronic hepatitis B virus infection

BackgroundInterleukin (IL)-35 regulates imbalance between regulatory T cells (Tregs) and T helper (Th) 17 cells, leading to an important modulator in autoimmune disorder, cancer, and infectious diseases. Our previous study revealed an immunosuppressive activity of IL-35 in chronic hepatitis B virus (HBV) infection. Thus, the aim of the current study was to investigate the role of regulatory function of IL-35 to viral specific Tregs/Th17 cells balance in chronic HBV infection.MethodsA total of 44 HLA-A2 restricted chronic HBV infected patients, including 21 of chronic hepatitis B (CHB) and 23 of asymptomatic HBV carriers (ASC) were enrolled. Purified CD4+ T cells or CD4+CD25+CD127dim/− Tregs were stimulated with recombinant IL-35. HBV core antigen specific Tregs and Th17 cells were determined by flow cytometry. FoxP3 and RORγt mRNA was measured by real-time PCR. Cytokines production (IL-10 and IL-17) was investigated by ELISA.ResultsPeripheral viral specific Tregs was comparable between CHB and ASC. However, increased percentage of viral specific Th17 cells was found in CHB, leading to the reduction of Tregs/Th17 ratio in CHB patients. IL-35 stimulation elevated viral specific Tregs, but not Th17 cells frequency, in both CHB and ASC, resulting in the elevation of Tregs/Th17 ratio in both groups. This process was accompanied by increased expression of FoxP3 mRNA and IL-10 production, and decreased IL-17 secretion and STAT3 phosphorylation in purified CD4+ T cells. Moreover, IL-35 stimulation inhibited viral specific Th17-like phenotype differentiation from Tregs in CHB patients. Effective anti-HBV therapy did not affect viral specific Tregs/Th17 cells frequency or IL-35 expression in CHB patients, however, reduced responsiveness of CD4+ T cells or Tregs to IL-35 stimulation in vitro.ConclusionOur findings indicated that IL-35 regulation to viral specific Tregs/Th17 balance may contribute to viral persistence in chronic HBV infection.

Corrigendum] Sequence analysis of the Pre‑S gene in chronic asymptomatic HBV carriers with low‑level HBsAg.

After the publication of the article, the authors realize that the grant funding number for the support that they received from Natural Science Foundation of Zhejiang Province appeared incorrectly in the Funding section of the paper: This should have been featured as grant no. LY15H200001 (i.e., with an 'L' at the start of the number). The authors regret that this error was not corrected prior to the publication of their paper, and apologize to the funders for any inconvenience caused. [the original article was published in International Journal of Molecular Medicine 42: 2689-2699, 2018; DOI:10.3892/ijmm.2018.3831].

Importance of Prolidase Enzyme Activity and Serum Cytokeratin 18 Levels for Differential Diagnosis between Asymptomatic Hepatitis B Carriers and HBeAg Negative Chronic Hepatitis B Patients

Background: In this study, it was aimed to evaluate the relationship between the severity of necroinflammation in the liver and the stage of fibrosis and the serum levels of Serum Prolidase Activity (SPA) and Cytokeratin (CK)-18 in patients with active Chronic Hepatitis B (CHB) and asymptomatic Hepatitis B Virus (HBV) carriers. Methods: Biochemical analyses, serological parameters associated with HBV and serum prolidase activity and CK-18 levels were measured in asymptomatic HBV carriers (n=65), active CHB patients (n=60) and healthy controls (n=27). Liver biopsies were performed on asymptomatic HBV carriers and active CHB patients. Findings: SPA level was significantly higher in active CHB patients (819.92 ± 123.74 IU/L) compared to asymptomatic HBV carriers (732.99 ± 124.70 IU/L) and was higher in asymptomatic HBV carriers compared to healthy controls (529.4 ± 74.73 IU/L) (p=0.001). The diagnostic cut-off value of SPA level was found 751.15 U/L. When this cut-off value was taken to differentiate HBe-Ag negative CHB in asymptomatic HBV carriers, sensitivity and specificity of efficacy were 72% and 63% respectively (c-statistics: 0.707). A strong positive correlation was observed between serum prolidase level and the severity of fibrosis in asymptomatic HBV carriers (r=0.603, p=0.000). A positive correlation was determined between SPA level and Histological Activity Index (HAI) scores in patients with active CHB and asymptomatic HBV carriers. The serum CK-18 levels were significantly lower in the healthy control group when compared to the asymptomatic HBV carriers and active CHB patients (p=0.001). Conclusion: Prolidase enzyme may be beneficial in differentiating asymptomatic HBV carriers from HBeAg-negative CHB patients when used in combination with ALT and HBV-DNA levels.

The Effect of ICOS Polymorphism Interactions with HBV Mutations on HBV Subtype Infection Outcomes

Introduction and aim. Hepatitis B virus (HBV) infection remains a public health problem worldwide. In addition, HBV infection results are influenced by various virological, immunological, and genetic factors. Inducible T-cell costimulator (ICOS) polymorphisms involving chronic HBV infection have been confirmed in previous studies. This study was to explore the effects of ICOS single nucleotide polymorphisms in HBV subtypes and their interactions with viral mutations on HBV infection outcomes.Material and methods. A total of 1,636 Han Chinese individuals were recruited, including 47 asymptomatic HBV carriers (ASC), 353 chronic hepatitis B (CHB) patients, 327 HBV-related liver cirrhosis (LC) patients, 193 HBV-related hepatocellular carcinoma (HCC) patients, 464 patients with spontaneous recovery from HBV infection (SR), and 252 healthy controls (HC). DNA samples from these subjects were genotyped for four ICOS SNPs (rs11883722, rs10932029, rs1559931, and rs4675379). Direct sequencing was used to determine the HBV mutations in the enhancer II, basal core promoter, and pre-core regions.Results. We found that the genotype “TC” of ICOS rs10932029 SNP was associated with decreased HBV-related LC risk in the genotype C group. Additionally, the A1762T, G1764A and A1762T/G1764A mutations were associated with an increased risk of LC in the genotype C group. Further study indicated that interactions between ICOS rs10932029 genotype “TC” and A1762T or A1762T/G1764A mutations significantly decreased the LC risk in the genotype C group.Conclusion. The rs10932029 genotype “TC” might be an LC-protective factor for HBV genotype C infection. The interactions between the rs10932029 genotype “TC” and A1762T or A1762T/G1764A mutations could decrease the risk of LC.

Analysis of clinical characteristics and S gene sequences in chronic asymptomatic HBV carriers with low-level HBsAg

During the natural hepatitis B virus (HBV) infection process, some infected subjects are characterized by a sustained low serum HBV surface antigen (HBsAg) expression level. Most members in this population are chronic asymptomatic HBV carriers (ASCs). To elucidate the mechanism underlying low-level HBsAg expression in ASCs, we sequenced the HBV S gene in these patients to reveal specific sequence characteristics. Overall, 1308 cases of chronic ASCs were grouped according to their HBsAg serum expression levels (10 IU/mL). The clinical characteristics of the population were analysed in detail. The HBV S gene was sequenced from 276 ASC cases with low-level HBsAg expression. Additionally, 100 of 1032 ASC cases with high-level HBsAg expression were randomly selected for HBV S gene sequencing based on age matching according to the low-level HBsAg group. A comparative analysis was conducted with the HBV S gene sequences from ASCs with low HBsAg expression and the HBV reference S gene sequences from ASCs with high HBsAg expression. The population with low-level HBsAg expression displayed the following primary clinical characteristics: mostly chronic asymptomatic HBV carriers, older age (mean age 55.09 years), HBsAg/anti-HBe/anti-HBc (core) positivity as the main serological pattern (97.1%), low HBV DNA replication (1.32 ± 1.60 log10 IU/mL), a low HBV-DNA positive rate (45.65%) and primarily genotype B (82.54%) and serotype adw (84.13%). The comparative analysis of the HBV S gene sequences from ASCs with low-level HBsAg showed significant mutations (including co-mutations) on both sides of the main hydrophilic region (MHR). Significant mutations in multiple regions and at multiple sites (including co-mutations) on both sides of the MHR may be one cause of the low HBsAg expression level in this population.

Sequence analysis of the Pre-S gene in chronic asymptomatic HBV carriers with low-level HBsAg

In a hepatitis B virus (HBV)-infected population, persistently low expression levels of serum HBV serum antigen (HBsAg) are present, particularly in chronic asymptomatic HBV carriers (ASCs). The present study sequenced the HBV Pre-S gene, and aimed to elucidate its features in ASCs with low HBsAg expression compared with in the established HBV Pre-S reference gene sequences from ASCs with high HBsAg expression. A total of 1,308 ASCs were grouped according to HBsAg serum levels (cut-off value, 10 IU/ml), and clinical characteristics were analyzed in detail. The HBV Pre-S gene was sequenced in 276 ASCs with low-level HBsAg; in addition, 100 of the remaining 1,032 ASCs with high-level HBsAg were randomly selected for HBV Pre-S gene sequencing on the basis of age matching with the low-level HBsAg group. Comparative analysis of the gene sequences from these groups was subsequently conducted. The major clinical features of the population with low-level HBsAg were as follows: Most were ASCs with chronic HBV infection; 97.1% were HBsAg/anti-HBe/anti-HBc-positive; 82.54% carried the B genotype; and 84.13% displayed the adw serotype. The results indicated that there were novel and meaningful mutations, including co-mutations, at numerous loci and sites in the Pre-S gene, as well as deletion mutations in the Pre-S2 gene. These mutations in the Pre-S1 and Pre-S2 gene frag ments accounted for 65.38% (68/104) of the 104 B genotype cases in the low-level HBsAg group and 90.91% (20/22) of the 22 C genotype cases in the low-level HBsAg group, respectively. In conclusion, Pre-S gene mutations may be associated with HBV replication defects, which may be the cause of the observed low expression levels of HBsAg.

The Effect of ICOS Polymorphism Interactions with HBV Mutations on HBV Subtype Infection Outcomes

Introduction and aim. Hepatitis B virus (HBV) infection remains a public health problem worldwide. In addition, HBV infection results are influenced by various virological, immunological, and genetic factors. Inducible T-cell costimulator (ICOS) polymorphisms involving chronic HBV infection have been confirmed in previous studies. This study was to explore the effects of ICOS single nucleotide polymorphisms in HBV subtypes and their interactions with viral mutations on HBV infection outcomes. Material and methods. A total of 1,636 Han Chinese individuals were recruited, including 47 asymptomatic HBV carriers (ASC), 353 chronic hepatitis B (CHB) patients, 327 HBV-related liver cirrhosis (LC) patients, 193 HBV-related hepatocellular carcinoma (HCC) patients, 464 patients with spontaneous recovery from HBV infection (SR), and 252 healthy controls (HC). DNA samples from these subjects were genotyped for four ICOS SNPs (rs11883722, rs10932029, rs1559931, and rs4675379). Direct sequencing was used to determine the HBV mutations in the enhancer II, basal core promoter, and pre-core regions. Results. We found that the genotype “TC” of ICOS rs10932029 SNP was associated with decreased HBV-related LC risk in the genotype C group. Additionally, the A1762T, G1764A and A1762T/G1764A mutations were associated with an increased risk of LC in the genotype C group. Further study indicated that interactions between ICOS rs10932029 genotype “TC” and A1762T or A1762T/G1764A mutations significantly decreased the LC risk in the genotype C group. Conclusion. The rs10932029 genotype “TC” might be an LC-protective factor for HBV genotype C infection. The interactions between the rs10932029 genotype “TC” and A1762T or A1762T/G1764A mutations could decrease the risk of LC.

Highly elevated soluble Tim-3 levels correlate with increased hepatocellular carcinoma risk and poor survival of hepatocellular carcinoma patients in chronic hepatitis B virus infection.

Background and objectiveUpregulated T-cell immunoglobulin and mucin domain containing molecule-3 (Tim-3) in hepatitis B virus (HBV)-specific CD8+ T-cells contributes to CD8+ T-cell exhaustion during chronic HBV infection. The membrane-bound Tim-3 can be cleaved from the cell surface by sheddase, yielding soluble Tim-3 (sTim-3). This study investigated serum sTim-3 levels in patients with chronic HBV infection of various liver diseases.MethodsSerum sTim-3 levels were quantitatively determined in 288 patients with chronic HBV infection of various liver diseases. The sTim-3 levels were analyzed in relation to liver diseases including HBV-related hepatocellular carcinoma (HCC) and overall survival of HCC patients.ResultsSerum sTim-3 levels in the patients with chronic HBV infection were significantly elevated compared with healthy controls (P<0.001) and the levels from asymptomatic HBV carrier status, chronic hepatitis, liver cirrhosis to HCC were progressively increased. Serum sTim-3 levels were closely associated with the severity of liver function abnormalities. Importantly, serum sTim-3 levels were independently associated with HCC risk (OR, 4.310; 95% CI, 2.141–8.676, P<0.001) in comparison to non-HCC diseases in chronic HBV infection and significantly associated with the overall survival of HCC patients, with a level >3000 pg/mL being related to shorter overall survival than a level ≤3000 pg/mL (P=0.019).ConclusionSerum sTim-3 is involved in disease progression and HCC development in chronic HBV infection and its quantitative determination may be potentially used as a marker for monitoring the disease progression and predicting the HCC prognosis in chronic HBV infection.

Serum levels of miRNA in patients with hepatitis B virus-associated acute-on-chronic liver failure

BackgroundHepatitis B virus (HBV)-associated acute-on-chronic liver failure (HBV-ACLF) is a life-threatening condition and its exact pathophysiology and progression remain unclear. The present study aimed to assess the role of serum miRNAs in the evaluation of HBV-ACLF and to develop a model to predict the outcomes for ACLF. MethodsSerum was collected from 41 chronic hepatitis B and 55 HBV-ACLF patients in addition to 30 chronic asymptomatic HBV carriers as controls. The miRNAs expressions were measured by real-time quantitative PCR (q-PCR). Statistical analyses were conducted to assess the ability of differentially expressed miRNAs and other prognostic factors in identifying ACLF prognosis and to develop a new predictive model. ResultsReal-time q-PCR indicated that serum miR-146a-5p, miR-122-3p and miR-328-3p levels were significantly upregulated in ACLF patients compared to chronic hepatitis B and chronic asymptomatic HBV carriers patients. In addition, multivariate regression analyses indicated that Na+, INR, gastrointestinal bleeding and miR-122-3p are all independent factors that are reliable and sensitive to the prognosis of HBV-ACLF. Therefore, we developed a new model for the prediction of HBV-ACLF disease state: Y = 0.402 × Na+ − 1.72 × INR − 4.963 × gastrointestinal bleeding (Yes = 0; No = 1)−0.278 × (miR-122-3p) + 50.449. The predictive accuracy of the model was 95.3% and the area under the receiver operating characteristic curve (AUROC) was 0.847. ConclusionsExpression levels of these miRNAs (miR-146a-5p, miR-122-3p and miR-328-3p) positively correlate with the severity of liver inflammation in patients with ACLF and may be useful to predict HBV-ACLF severity.

Decreased CD122 on CD56dim NK associated with its impairment in asymptomatic chronic HBV carriers with high levels of HBV DNA, HBsAg and HBeAg

AimsNK cells play important roles in inhibiting HBV replication and preventing HBV infection. However, NK-cell dysfunction has not been fully studied in asymptomatic chronic HBV carriers (ASC). This study aims to assess decreased expression of CD122 associated with impaired NK cells and the restoration of NK cells with IL-2 and IL-15 stimulation. Main methodsThe experiments were performed by flow cytometer, cytotoxicity assay, ELISA and western blotting. Key findingsThe reduced CD122 on CD56+ NK cells and CD56dim NK cells is associated with high levels of HBV DNA, HBsAg or HBeAg in ASCs, in which CD56dim NK-cell impairment is observed. Moreover, decreased IFN-γ and degranulation and low cytotoxicity by CD56dim NK cells after CD122 blockade were revealed. IL-2 and/or IL-15 can restore impaired CD56dim NK cells, together with increased p-STAT5, which can be reversed by CD122 blockade. Additionally, IL-2 or IL-15 can enhance IFN-α2-activated CD56dim NK-cell immune responses via up-regulating interferon alpha and beta receptor subunit 2 (IFNAR2). SignificanceDown-regulated CD122 on CD56dim NK cell in ASCs with massive viral antigens and high viremia is associated with its impairment, which can be restored by IL-2 and/or IL-15, or combined with IFN-α2.