Jinjun Chen

Clinical and Virological Characteristics of Hepatitis B Virus Subgenotypes Ba, C1, and C2 in China

ABSTRACT Hepatitis B virus (HBV) subgenotypes Ba, C1 (Cs), and C2 (Ce) are the most prevalent HBV variants in China. To investigate the virological characteristics of these subgenotypes and their clinical implications, we enrolled a cohort of 211 patients in the Guangdong Province of China, including 132 with chronic hepatitis B virus infection (CH), 32 with liver cirrhosis (LC), and 47 with hepatocellular carcinoma (HCC) according to clinical examination, liver function test, and ultrasonograph results. Overall, HBV Ba was found in 51.2% (108/211), HBV C1 in 33.6% (71/211), and HBV C2 in 15.2% (32/211) of the cases. The distribution of HBV genotype C was greater among patients in the LC and HCC groups than among patients in the CH group, while the distribution of HBV genotype B was greater among the CH patients than among the LC and HCC patients. No significant differences in clinical features were found among patients with HBV Ba, C1, and C2. Virologically, HBV C1 had the strongest association with the A1762T G1764A double mutation, while the mutation at position 1896 resulting in A (1896A) was uncommon. In contrast, HBV Ba had the highest frequency of 1896A but the lowest of A1762T G1764A, and HBV C2 had intermediate frequencies of these mutations. Mutations of 1653T and 1753V were specifically associated with HBV C2 and C1, respectively. Multivariate analyses showed that the 1653T, 1753V, and A1762T G1764A mutations and patient age significantly increased the risk of HCC development. In conclusion, HBV Ba, C1, and C2 have different mutation patterns in the enhancer II/core promoter/precore region. Therefore, genotyping and detecting the 1653T and 1753V mutations, in addition to the A1762T G1764A double mutation, might have important clinical implications as predictive risk factors for hepatocarcinogenesis.

Detection and significance of a G1862T variant of hepatitis B virus in Chinese patients with fulminant hepatitis

The prevalence of a G1862T variant of hepatitis B virus (HBV) has been investigated in patients with fulminant hepatitis and chronic liver disease, using primer mismatch amplification, followed by restriction fragment length polymorphism analysis. This variant was five times more common in patients with fulminant hepatitis (13.7%, 7 of 52) than in chronic carriers (2.5%, 2 of 81). The G-->T substitution at position 1862 leads to an amino acid change in codon 17 of the precore protein of the virus, which is part of a signal peptidase recognition motif. Variants with this mutation were only seen in patients infected with genotype B. In vitro translation experiments showed that this variant has greatly reduced capacity to produce hepatitis B e antigen (HBeAg) from its precore protein precursor. Furthermore, 88.5% of patients with fulminant hepatitis had mutations that are known to be associated with abrogated or reduced production of HBeAg. This suggests that, following HBV infection, the absence or reduced amounts of HBeAg may be a contributing factor in fulminant disease.

Hepatitis B virus genotype B with G1896A and A1762T/G1764A mutations is associated with hepatitis B related acute-on-chronic liver failure.

The existence of statistical associations between hepatitis B‐related acute‐on‐chronic liver failure and both hepatitis B virus (HBV) genotype and mutations in the basal core promoter (BCP) and precore (PC) regions needs to be confirmed. A total of 322 patients with a chronic HBV infection, including 77 with hepatitis B‐related acute‐on‐chronic liver failure, 109 with hepatocellular carcinoma (HCC) and 136 with chronic hepatitis B (CHB) were enrolled. The HBV genotype and the presence of mutations in the BCP/PC regions were determined by direct sequencing, and the frequencies were compared in the three patient groups. Overall, 198/322 (61.5%) were infected with genotype B and 124/322 (38.5%) with genotype C. Genotype B was significantly more frequent in patients with acute‐on‐chronic liver failure than CHB (92.2% vs. 60.3%, P < 0.001). As a contrast, genotype C was more common in patients with HCC than CHB (58.7% vs. 39.7%, P = 0.003). In genotype B patients, the A1762T/G1764A, A1846T, and G1896A mutations were significantly more prevalent in patients with acute‐on‐chronic liver failure than CHB (50.7% vs. 28.0%, P = 0.004; 59.2% vs. 34.1%, P = 0.002; 69.0% vs. 41.5%, P = 0.001, respectively). In multivariate analysis, the risk factors for acute‐on‐chronic liver failure were genotype B, A1762T/G1764A, and G1896A. In conclusion, CHB patients with genotype B, G1896A, and A1762T/G1764A had a higher tendency to develop liver failure than patients with genotype C. Therefore, HBV genotyping and detecting G1896A and A1762T/G1764A mutations might have important clinical implications as predictive risk factors for hepatitis B‐related acute‐on‐chronic liver failure. J. Med. Virol. 83:1544–1550, 2011.

Cellular immune responses in patients with hepatitis B surface antigen seroclearance induced by antiviral therapy.

BackgroundThe mechanisms by which chronic hepatitis B is completely resolved through antiviral therapy are unknown, and the contribution of acquired T cell immunity to hepatitis B surface antigen (HBsAg) seroclearance has not been investigated. Therefore, we measured the T-cell responses to core and envelope antigens in patients with HBsAg seroclearance.MethodsFourteen subjects with HBsAg seroclearance following antiviral treatment for chronic hepatitis B, 7 HBeAg-positive immunotolerant HBV carriers and 9 HBeAg-negative inactive HBsAg carriers were recruited. HBV-specific T-cell responses to recombinant HBV core (rHBcAg) and envelope (rHBsAg) proteins and pools of core and envelope peptides were measured using an ELISPOT assay detecting interferon-gamma and intracellular cytokine staining (ICS) assays detecting interferon-gamma or interleukin 2.ResultsInterferon-gamma ELISPOT assays showed a low frequency of weak responses to the rHBsAg and S peptide pool in the HBsAg seroclearance group, and the response frequency to the rHBcAg and the C peptide pool was higher than to the rHBsAg (P < 0.001) and S peptide pool (P = 0.001) respectively. A higher response frequency to C than S peptide pools was confirmed in the interferon-gamma ICS assays for both CD4+ (P = 0.033) and CD8+ (P = 0.040) T cells in the HBsAg seroclearance group. The responses to C and S antigens in the inactive carriers were similar.ConclusionsThere was a low frequency of CD4+ and CD8+ T cell immune responses to envelope antigens in Chinese subjects with HBsAg seroclearance following antiviral therapy. It is unlikely that these immune responses are responsible for HBsAg seroclearance in these subjects.

Hepatitis B virus genotypes, subgenotypes, precore, and basal core promoter mutations in the two largest provinces of Pakistan

Background and Aim:  Hepatitis B virus (HBV) genotyping has been done in most countries, but unfortunately, in Pakistan, HBV genotypic distribution is still unclear. The aim of the present study was to determine the prevalent genotype and subgenotype in the two most populated provinces in Pakistan: Punjab and Sind.

Serum HBsAg changes in HBeAg positive chronic hepatitis B patients with continuous viral load reductions during treatment with adefovir or peg-interferon-α-2a.

Hepatitis B surface antigen (HBsAg) loss under antiviral therapy is rare in chronic hepatitis B patients and the dynamics of serum HBsAg in these patients are not available. The changes in serum HBsAg following treatment with adefovir (n=31) or peg-interferon-alpha-2a (n=23) were studied in hepatitis B e-antigen (HBeAg) positive chronic hepatitis B patients. Abbott Architect HBsAg assay was used to quantify serum HBsAg. HBsAg levels were significantly decreased during the first 12 weeks of treatment with median change of -397.0 IU/ml and -555.4 IU/ml, respectively for adefovir and peg-interferon-alpha-2a (p=0.005 and 0.001, respectively). Beyond 12 weeks, no further significant HBsAg reductions were found even in patients with sustained viral replication inhibition in either group. Three distinct patterns of HBsAg changes were observed in most patients in both treatment groups: biphasic pattern (rapid HBsAg reduction from baseline to week 12); assurgent pattern (higher HBsAg level at week 12 than at baseline); and wavy pattern (HBsAg reduction from baseline to week 12, followed by relapse at week 24 or week 28). These results might offer insights into the possible mechanism(s) underlying the unusual occurrences of HBsAg loss under antiviral therapy.

GTPase activity is not essential for the interferon-inducible MxA protein to inhibit the replication of hepatitis B virus

Multiple studies have established that GTPase activity is critical for MxA to act against RNA viruses. Recently, it was shown that MxA can also restrict the replication of hepatitis B virus (HBV), a DNA virus, but the requirements for GTPase activity in inhibition of HBV by MxA remain unknown. Here, we report that GTPase-defective mutants (K83A, T103A, and L612K) can downregulate extracellullar HBsAg and HBeAg and reduce the expression of extra- and intracellular HBV DNA in HepG2 cells to levels similar to that achieved by wild-type MxA. Furthermore, TMxA and T103, two nuclear forms of wild-type MxA and a GTPase-defective mutant (T103A) could only slightly decrease the expression of extra- and intracellular HBV DNA in HepG2 cells. In conclusion, GTPase activity is not essential for MxA protein to inhibit HBV replication, and MxA may have only a minimal effect on the replicative cycle of HBV in the nucleus.

Aberrant expression and dysfunction of TLR2 and its soluble form in chronic HBV infection and its regulation by antiviral therapy

Toll-like receptor 2 (TLR2) plays an important role in the immunopathogenesis of hepatitis B virus (HBV) infection. The relationship between TLR2 expression and clinical outcome of chronic HBV infection is not yet elucidated in details so far. Here, we employed clinical cohorts to investigate TLR2 expression and function in different phases of HBV infection and dynamic changes of TLR2 expression in HBeAg-positive chronic hepatitis B (CHB) patients during antiviral therapy. TLR2 was mainly expressed in monocytes and its ligand stimulation resulted in TNF-α, IL-6 and IL-10 production. Serum soluble TLR2 (sTLR2) levels were negatively correlated with TLR2 mRNA in PBMCs. As compared with immunotolerant carriers and inactive carriers, CHB patients showed an elevated TLR2 expression and TNF-α, IL-6 induction in PBMC, but had a decreased level of sTLR2 in serum. However, TLR2 expression and TNF-α induction in monocytes of CHB patients remained lower than healthy controls. Furthermore, higher TLR2 expression in PBMCs and lower level of sTLR2 in serum at baseline were predictive of a complete response to 52 weeks of telbivudine (LdT) therapy. Temporal dynamic analysis showed that TLR2 expression was restored with viral suppression and ALT normalization from week 12 to 24. However, peg-IFN-α-2a therapy induced a slightly decline in TLR2 expression. In conclusion, TLR2 expression and function in monocytes were impaired by chronic HBV infection. Higher TLR2 expression in PBMC and lower level of sTLR2 in serum at baseline were associated with a complete response to LdT therapy, and dynamic TLR2 expression was differently regulated by LdT and peg-IFN-α-2a therapy.

Factors associated with serum hepatitis B surface antigen levels and its on-treatment changes in patients under lamivudine therapy.

BACKGROUND We aimed to investigate factors associated with serum hepatitis B surface antigen (HBsAg) levels and its kinetics under lamivudine treatment in chronic hepatitis B patients. METHODS HBsAg levels were measured with the Architect HBsAg assay (Abbott laboratories, Abbott Park, IL, USA) in genotype B (HBV/B) or C (HBV/C) patients (n=218). Early HBsAg kinetics in 86 hepatitis B e antigen (HBeAg)-positive patients and long-term HBsAg changes in 45 patients with rapid and sustained viral suppression were further analysed. RESULTS Mean HBsAg levels were higher in male (n=181) than in female (n=37) patients (3.59 versus 3.23 log(10) IU/ml; P=0.036), and higher in 121 HBV/B than in 97 HBV/C patients (3.68 versus 3.34 log(10) IU/ml; P=0.006). In addition to HBV DNA loads (P<0.001), male gender (P=0.012) and HBV/B infection (P=0.035) were independently associated with higher HBsAg levels in antiviral-naive patients. HBsAg increases (0.00-0.87 log(10)) were found in 28/86 patients who obtained viral suppression under 12 weeks of lamivudine therapy. Higher baseline HBsAg levels (P=0.046), HBV/B infection (P=0.007) and faster HBV DNA declines (P=0.006) independently contributed to greater HBsAg decreases under 12 weeks treatment. An apparent dissociation between HBsAg and HBV DNA changes were found in 14/45 patients with rapid and sustained viral suppression, who had low baseline HBsAg levels and predominant HBV/C infection. CONCLUSIONS HBV/B and male gender were associated with higher HBsAg levels in antiviral-naive patients. Higher baseline HBsAg levels and HBV/B infection contributed to greater early HBsAg declines in HBeAg-positive patients, and might correlate with discordance between HBsAg and HBeAg or HBV DNA under long-term lamivudine treatment.

Clinical and virological characteristics of chronic hepatitis B with concurrent hepatitis B E antigen and antibody detection.

Summary.  The concurrent detection of hepatitis B e antigen (HBeAg) and its corresponding antibody (anti‐HBe) in patients with chronic hepatitis B virus (HBV) infection is well established but the clinical features remain poorly understood. Demographic information, clinical and laboratory data were collected from 1624 consecutive inpatient records of patients with chronic hepatitis B. Viral genotype, basic core promoter and precore mutations were determined by direct sequencing. In vitro HBeAg and anti‐HBe binding experiments were conducted with three pairs of HBeAg‐positive and anti‐HBe‐positive serum samples, which were mixed at variable ratios and incubated at 37 °C for 3–24 h. Of the 1624 chronic patients, 169 (10.4%) had concurrent HBeAg and anti‐HBe positivity, and this was associated with intermediate age and HBV‐DNA load, higher alanine aminotransferase level and more pronounced liver damage compared with HBeAg‐positive or anti‐HBe‐positive patients alone. HBeAg and anti‐HBe titres (median and interquartile range, S/CO) in the concurrent positive group were 4.2 (1.8–9.6) and 0.54 (0.27–0.72), which were closer to their respective cut‐off values than those of HBeAg‐positive or anti‐HBe‐positive groups alone. For the cases successfully sequenced, 110/134 (82.1%) harboured T1762/A1764 or/and A1896 mutants. The binding experiments showed that HBeAg and anti‐HBe could be concurrently observed provided an optimal ratio (HBeAg to anti‐HBe) was chosen. In antiviral treatment‐naive patients, concurrence of HBeAg and anti‐HBe was not uncommon, and such patients had profound liver disease. An optimal ratio between HBeAg and anti‐HBe led to their concurrent detection when sera were tested by sensitive assays.