Journal of Viral Hepatitis

Published by Wiley
Online ISSN: 1365-2893
Print ISSN: 1352-0504
The aim of this work was to assess the effect of a high-dose (10 million units, MU) short-duration (14 weeks) interferon-alpha2b (IFN-alpha2b) regimen in relapsers compared with the standard IFN regimen of 3 MU three times weekly (t.i.w.) for 6 months. Fifty-eight non-cirrhotic patients (who had relapsed after previous treatment with IFN) with chronic hepatitis were randomized: 29 to the high-dose, short-duration regimen and 29 to the standard regimen. By the end of IFN therapy, in the high-dose, short-duration group alanine aminotransferase (ALT) normalization was observed in 23 (79%) of 29 patients, and undetectable hepatitis C virus (HCV) RNA in eight (28%) vs 25 (86%) and 11 (38%) of the 29 patients in the standard group, respectively (P = NS). At the end of the 72-week follow-up, in the high-dose, short-duration group a sustained ALT normalization was observed in two (7%) patients, and undetectable HCV RNA in 0 (0%) vs five (17%) and four (14%) patients in the standard group (P = NS). There was less fibrosis improvement in the high-dose, short-duration group (two of 26 patients, 8%) than in the standard group (eight of 25 patients, 32%) (P = 0.04). Tolerance to IFN was good and similar in the two groups. In conclusion, in IFN relapsers, high-dose, short-duration treatment with IFN-alpha has no advantage when compared to a 6-month treatment with 3 MU IFN t.i.w.
  A new hepatitis B virus (HBV) protein, hepatitis B splice-generated protein (HBSP), has been detected in liver biopsy specimens from patients with chronic active hepatitis. The aim of this study was to characterize the phenotype and functions of peripheral HBSP-specific T cells and to determine whether these T-cell responses may be implicated in liver damage or viral control. Two groups of patients were studied: HBV-infected patients with chronic active hepatitis and HBV-infected patients who were inactive carriers of hepatitis B surface antigen. HBSP-specific T-cell responses were analysed ex vivo and after in vitro stimulation of peripheral blood mononuclear cells. Soluble cytokines and chemokines were analysed in sera and in cell culture supernatants. Few HBSP- or capsid-specific T-cell responses were detected in patients with chronic active hepatitis whereas frequency of HBV-specific T cells was significantly higher in inactive carrier patients. HBSP activated CD8+ and CD4+ T cells that recognized multiple epitopes and secreted inflammatory cytokines. The IL-12 level was significantly lower in sera from asymptomatic carrier patients compared to patients with chronic active hepatitis. IL-12 and IP-10 levels in the sera were significantly and independently correlated with both alanine amino transferase and HBV DNA levels. Our results show that the HBSP protein activates cellular immune responses in HBV-infected patients but has probably no prominent role in liver damage. The pattern of cytokines and chemokines in sera was linked to HBV viral load and was consistent with the level of inflammation during chronic hepatitis.
Sofosbuvir and GS-0938 are distinct nucleotide analogues with activity against hepatitis C virus (HCV) in vitro. We evaluated the antiviral activity and safety of sofosbuvir and GS-0938 alone and in combination in HCV genotype 1 patients. In this double-blind study, 40 treatment-naïve patients were randomly assigned to 4 treatment cohorts: (i) GS-0938 for 14 days, (ii) GS-0938 for 7 days followed by GS-0938 plus sofosbuvir for 7 days, (iii) sofosbuvir for 7 days followed by GS-0938 plus sofosbuvir for 7 days and (iv) GS-0938 plus sofosbuvir for 14 days. In each arm, 8 patients received active drug and 2 placebo. After 7 days of dosing, patients in all 4 dose groups experienced substantial reductions in HCV RNA, with median declines (Q1, Q3) of -4.50 (-4.66, -4.24) in Cohort 1, -4.55 (-4.97, -4.13) in Cohort 2, -4.65 (-4.78, -4.17) in Cohort 3 and -4.43 (-4.81, -4.13) in Cohort 4; patients receiving placebo had essentially no change in HCV RNA (+0.07 log10 IU/mL). Seven days after the end of treatment, the proportions of patients with HCV RNA <15 IU/mL were 4 (50%), 8 (100%), 7 (88%) and 5 (63%) for Cohorts 1-4, respectively, vs 0 for placebo. No viral breakthrough or resistance mutations were observed. No serious adverse events or Grade 3 or 4 adverse events were reported. Sofosbuvir and GS-0938-alone and in combination-were well tolerated and led to substantial reductions in viral load. Sofosbuvir is undergoing further investigation as a possible backbone of an all-oral regimen for chronic HCV.
Health regulatory approval of the 1.5 microg/kg body weight dose of pegylated interferon (PEG-I) alpha-2b in combination with ribavirin for the treatment of chronic hepatitis C was based on a study using PEG-I alpha-2b at doses of only 0.5 and 1.5 microg/kg body weight (BW), in spite of the previously shown flat dose-response curve at doses of > or =1.0 microg/kg. Our aim was to compare PEG-I alpha-2b 1.0 microg/kg with 1.5 microg/kg, both in combination with ribavirin. Open-label, randomized study in 227 patients with biopsy-proven chronic hepatitis C (Metavir < or =F2), receiving oral ribavirin (400 mg, twice daily) in combination with subcutaneous PEG-I alpha-2b (1.0 or 1.5 microg/kg, once weekly) for 24 weeks (genotype 2 or 3), or 48 weeks (other genotypes), followed by a 24-week drug-free period. Virologic response rates did not differ between the two doses of PEG-I alpha-2b: in patients infected with hepatitis C virus (HCV) genotype 1 or 4 treated with PEG-I 1.0 microg/kg BW, 38% (22/58) had a sustained virologic response compared with 39% (27/70) in the PEG-I 1.5 microg/kg BW dose group (P = ns). The corresponding values in patients infected with HCV genotype 2 or 3 were 71% (39/55) and 81% (29/36) respectively (P = ns). Adverse events led to transient or permanent dose reductions in fewer patients in the 1.0 microg/kg BW dose group (48/113 patients; 42%) than in the 1.5 microg/kg BW dose group (63/106 patients; 59%, P = 0.015). Furthermore, 89% of patients treated for 24 weeks but only 58% of patients treated for 48 weeks (P < 0.001) tolerated the treatment without relevant dose reduction or premature termination. In combination with ribavirin, PEG-I alpha-2b 1.0 microg/kg was as effective as 1.5 microg/kg but was better tolerated in patients with chronic hepatitis C and up to moderate fibrosis.
Increasing evidence suggests that interleukin-10 (IL-10) gene promoter polymorphisms may be associated with chronic hepatitis C virus (HCV) infection and HCV clearance. To more precisely estimate the association between these variants and the risk of HCV infection, we performed a meta-analysis of 26 studies describing the IL-10-1082A/G, -819C/T, -592C/A genotypes, including 4039 chronic HCV infection cases and 2902 controls. When compared with a healthy population, the -1082GG allele had a 43% increased risk of chronic HCV infection in combined populations (GG vs GA + AA: odds ratio (OR) = 1.433, 95% confidence interval (CI) = 1.052-1.952, P = 0.023). In subgroup analysis by ethnicity, a significant increased risk was associated with the -1082GG genotype in the Caucasian population (GG vs AA: OR = 1.390, 95% CI: 1.108-1.744, P = 0.004; GG vs GA + AA: OR = 1.621, 95% CI: 1.267-2.075, P = 0.000). However, no significant association was found in Asian, African or Chinese populations. Moreover, a higher distribution of -592A was found in the spontaneously recovered population (AA vs CC: OR = 0.585, 95% CI = 0.387-0.884, P = 0.011; AA + AC vs CC: OR = 0.738, 95% CI = 0.551-0.988, P = 0.041; AA vs AC + CC: OR = 0.788, 95% CI = 0.664-0.935, P = 0.006) than that in the chronic HCV infection population. In conclusion, the IL-10-1082GG allele may increase the risk of chronic HCV infection in Caucasian population, and people carrying the IL-10-592A allele are more likely to clear HCV spontaneously.
Clinical trial design for pooled phase III studies. SMV, simeprevir; PBO, placebo; PR, peginterferon and ribavirin; QD, once daily.
Mean (±SE) changes from baseline in PRO scores over time by treatment group; ITT (primary pool). The WPAI Productivity Impairment data shown correspond to impairment during the hours actually worked (question 5 from the WPAI questionnaire). CES-D, Center for Epidemiologic Studies Depression scale; EQ-5D, EuroQoL 5 dimensions; FSS, Fatigue Severity Scale; HCV, hepatitis C virus; PBO, placebo; PR, peginterferon/ribavirin; PRO, patient-reported outcome; SE, standard error; SMV, simeprevir; WPAI, Work Productivity and Activity Impairment questionnaire.
Mean (±SE) change from baseline in the Fatigue Severity Score (FSS) over time in the SMV/PR arm and PBO/PR arm, respectively, by SVR12, METAVIR fibrosis score, and by RGT (SMV/PR arm only); ITT (primary pool). PBO, placebo; PR, peginterferon/ribavirin; PRO, patient-reported outcome; RGT, response-guided therapy; SE, standard error; SMV, simeprevir.
The value of adding simeprevir (SMV) vs placebo (PBO) to peginterferon and ribavirin (PR) for treatment of chronic hepatitis C virus infection was examined using patient-reported outcomes (PROs); further, concordance of PROs with virology endpoints and adverse events (AEs) was explored. Patients (n = 768 SMV/PR, n = 393 PBO/PR) rated fatigue (FSS), depressive symptoms (CES-D) and functional impairment (WPAI: Hepatitis C Productivity, Daily Activity and Absenteeism) at baseline and throughout treatment in three randomised, double-blind trials comparing the addition of SMV or PBO during initial 12 weeks of PR. PR was administered for 48 weeks (PBO group) and 24/48 weeks (SMV group) using a response-guided therapy (RGT) approach. Mean PRO scores (except Absenteeism) worsened from baseline to Week 4 to the same extent in both groups but reverted after Week 24 for SMV/PR and only after Week 48 for PBO/PR. Accordingly, there was a significantly lower area under the curve (baseline-Week 60, AUC60 ) and fewer weeks with clinically important worsening of scores in the SMV/PR group at any time point. Incidences of patients with fatigue and anaemia AEs were similar in both groups, but FSS scores showed that clinically important increases in fatigue lasted a mean of 6.9 weeks longer with PBO/PR (P < 0.001). PRO score subgroup analysis indicated better outcomes for patients who met the criteria for RGT or achieved sustained virological response 12 weeks post-treatment (SVR12); differences in mean PRO scores associated with fibrosis level were only observed with PBO/PR. Greater efficacy of SMV/PR enabled reduced treatment duration and reduced time with PR-related AEs without adding to AE severity. © 2014 The Authors. Journal of Viral Hepatitis published by John Wiley & Sons Ltd.
Reduction of the window period of hepatitis C virus (HCV) infection represents an important goal in the transfusional and diagnostic setting. A prototype assay designed to simultaneously detect circulating HCV antigen and anti-HCV, has been developed. Aim of this study was to evaluate the performance of this new assay in terms of specificity and sensitivity and to compare its efficacy with commercial assays. To evaluate the specificity of the assay, 400 samples from the general population and 100 'difficult' sera, negative for anti-HCV, were tested. To assess sensitivity, the new test was used on 76 PCR-positive and anti-HCV negative sera, seven natural or commercial seroconversion panels that included 17 RNA-positive and anti-HCV negative sera and 31 anti-HCV positive sera, 20 weak anti-HCV positive sera, 80 viraemic and anti-HCV-positive sera from patients infected with different subtypes and 10 sera from patients with HBV-HCV or HIV-HCV co-infections. Of 500 anti-HCV negative samples, 499 (99.8%) were negative with a cut-off index <0.5, while one sample was within the grey zone. Of the 93 HCV-RNA positive and anti-HCV negative sera from patients and panels, 85 (91.4%) resulted positive, and one had the cut-off index in the grey zone. The reduction in the diagnostic window period observed with the new test and HCV-RNA assays were equal, on average, to 24 and 34.4 days respectively. All anti-HCV positive sera were positive. The new assay shows high sensitivity and specificity and could be a useful tool not only in the diagnostic setting, where procedures to reduce the window period, such as antigen or HCV-RNA detection, are not currently recommended, but also in the screening of blood donations, when nucleic acid technologies is not feasible because of costs, organization, emergency and/or logistic difficulties.
The levels of the liver-specific microRNA-122 (miR-122) circulating extracellularly in the blood have been shown to be increased upon liver damage. However, it is unknown if the levels of serum miR-122 are altered during antiviral therapy and reflect the therapeutic success. Here, we investigated miR-122 serum levels in patients with chronic hepatitis C virus (HCV) genotype 1 infection during antiviral therapy with pegylated interferon and ribavirin. Therefore, sera from 60 patients with chronic HCV infection genotype 1 showing sustained virological response (SVR), non-response or relapse to therapy obtained at baseline, 4, 12, 24 weeks, end of treatment and follow-up were analysed retrospectively for miR-122 content by quantitative real-time reverse transcription PCR. The time courses of miR-122 were correlated with HCV RNA as well as standard liver parameters. We found that while there was no relation between serum miR-122 and HCV RNA levels at baseline, the decline in HCV RNA upon beginning of the therapy closely correlated with the reduction of serum miR-122 in the three different patient groups. Moreover, the serum miR-122 level correlated well with alanine aminotransaminase, a marker of ongoing liver damage. At follow-up serum miR-122 levels remained low in SVR, but increased to baseline levels in patients not responding or showing relapse to therapy. In contrast, the serum concentration of the ubiquitously expressed miR-16 did not change during therapy. We conclude that the serum level of miR-122 well reflects the success of interferon/ribavirin therapy in patients with chronic HCV infection.
As chronic hepatitis C patients with progressive disease can present themselves with normal ALT levels, more sensitive biomarkers are needed. MicroRNAs are newly discovered small noncoding RNAs that are stable and detectable in the circulation. We aimed to investigate the association between hepatocyte-derived microRNAs in serum and liver injury in patients with chronic hepatitis C. The hepatocyte-derived miR-122 and miR-192 were analysed in sera of 102 chronic HCV-infected patients and 24 healthy controls. Serum levels of miR-122 and miR-192 correlated strongly with ALT (R = 0.67 and R = 0.65, respectively, P < 0.001 for both). Median levels of miR-122 and miR-192 in HCV-infected patients were 23 times and 8 times higher as in healthy controls (P < 0.001 for both). Even within the HCV-infected patients with a normal ALT (n = 38), the levels of miR-122 and miR-192 were 12 times and 4 times higher compared with healthy controls (P < 0.001 for both). Multivariate logistic regression analyses showed that only miR-122 was a significant predictor of the presence of chronic HCV infection (P = 0.026). Importantly, miR-122 was also superior in discriminating chronic HCV-infected patients with a normal ALT from healthy controls compared with the ALT level (AUC = 0.97 vs AUC = 0.78, P = 0.007). In conclusion, our study confirmed that liver injury is associated with high levels of hepatocyte-derived microRNAs in circulation and demonstrated that in particular miR-122 is a sensitive marker to distinguish chronic hepatitis C patients from healthy controls. More sensitive blood markers would benefit especially those patients with minor levels of hepatocellular injury, who are not identified by current screening with ALT testing.
miR-122 is a liver-specific microRNA, which also circulates in the blood. The levels of miR-122 in serum and plasma correlate with hepatic necroinflammation in patients with hepatitis B virus (HBV) infection. Here, we investigated whether miR-122 levels correlate with surrogate markers for viral replication and translation. Furthermore, we examined whether miR-122 levels differ in the different groups of HBV-infected patients and whether miR-122 levels may be useful to identify patients with higher or lower risk for liver disease progression. Therefore, RNA was extracted from sera of therapy-naïve patients with HBV infection (n = 89) and from healthy volunteers (n = 19). The concentration of miR-122 was assessed by quantitative real-time reverse-transcription PCR. HBs antigen and HBV DNA levels were quantified as surrogate parameters for HBV replication and translation. Liver biopsies were examined according to the histological activity index and the degree of fibrosis was assessed. We found that the miR-122 serum concentration correlated with the level of ALT, HBV DNA and HBs antigen (r = 0.259, P < 0.05; r = 0.225, P < 0.05; r = 0.508, P < 0.001, respectively). The miR-122 serum levels discriminated the different patient groups infected with HBV from healthy subjects (P < 0.001), and inactive carrier patients with high (>3500 IU/mL) or low (<3500 IU/mL) levels of HBs antigen could be differentiated by the miR-122 serum concentration (P < 0.05). As serum miR-122 levels strongly correlated with HBs antigen, it might be an indicator for viral translation. Furthermore, serum miR-122 levels discriminated HBV carrier patients with high or low risk for disease progression and may, thus, be an additional marker for risk stratification.
Interferon-alpha (IFN-alpha) is the major treatment for chronic hepatitis C virus (HCV) infection. Drug resistance is problematic, particularly among African-Americans who typically show poorer clinical outcomes than Caucasians. The reasons for ethnic variation in IFN-alpha sensitivity are not clear. We speculated that African-American insensitivity to IFN-alpha may be mediated by reduced density of the IFN-alpha receptor (IFN-alphaR) or reduced internalization of the IFN-alpha/IFN-alphaR complex. This speculation was evaluated by comparing binding, uptake and release of 125iodine-labelled IFN-alpha (125I-IFN-alpha) to peripheral blood cells from African-Americans and Caucasians with HCV infection and ethnically matched healthy volunteers. Under various in vitro conditions, binding of 125IFN-alpha to surface receptors was equivalent (P = ns) between African-Americans and Caucasians with HCV infection as well as healthy volunteers (P = ns). Similarly, internalization and release of the 125I-IFN-alpha/IFN-alphaR complex was equivalent (P = ns) between African-Americans and Caucasians with HCV infection and healthy volunteers (P = ns). In addition, ethnicity did not influence (P = ns) IFN-alpha suppression of phytohaemagluttinen induced proliferation. However, IFN-alpha therapy of the same patients showed that African-Americans had lower response rates than Caucasians (14%vs 54%, P < 0.0001). In summary, IFN-alpha resistance among African-Americans is not mediated by intrinsic differences in IFN-alpha receptor density or internalization.
Considerable evidence suggests that host genetic factor play an important role in the pathogenesis and clinical outcome of chronic hepatitis B virus (HBV) infection in several ethic groups. Association study was performed included 150 chronic HBV patients, 100 resolved hepatitis B and 150 healthy individuals with similar ethic background. Interestingly, human leucocyte antigen (HLA)-DR13 show a strong association with the clearance of HBV [odds ratio (OR) = 0.04, 95% confidence interval (CI) = 0.00-0.26, corrected P-value (P(c)) = 0.0008] similar to reports from several ethic groups. TNF-alpha promoter polymorphisms (-863, -308 and -238) were also analysed. Only -863 C allele was found to be significantly decreased in chronic HBV patients compared with healthy control (P(c) = 0.03, OR = 0.54, 95% CI = 0.35-0.84 respectively). This -863C allele was not in linkage disequilibrium with HLA-DR13 suggesting that other genetic markers linked with -863C might influence clearance of chronic HBV infection in Thai. When stratified chronic HBV patients into patients without hepatocellular carcinoma (HCC) and with HCC, the -863 A allele was significantly increased in the HCC group compared to healthy control (P(c) = 0.003, OR = 2.61, 95% CI = 1.49-4.60). Haplotype analysis (-863/-308/-238) revealed that the homozygosity of the haplotype (CGG/CGG) was a protective marker for HCC (OR = 0.37, 95% CI = 0.17-0.79, P(c) = 0.02). One can propose that carriers of -863A genotype are associated with increased levels of TNF-alpha in the liver in response to HBV infection and induce hapatocyte damage that may finally lead to HCC development. Additional study is needed to validate these finding and to further explore the genetic pathogenesis of HBV infection.
The pathogenic role of core promoter (CP) mutations (T1762/A1764) of hepatitis B virus (HBV) in hepatitis B e antigen (HBeAg) seroconversion or disease progression remains unclear. We investigated the clinical relevance of these mutants over a long-term follow-up period of up to 15 years. In this longitudinal cohort study, 29 HBeAg-positive patients with biopsy-proved chronic active hepatitis without cirrhosis were regularly monitored for >10 years. The viral isolates were characterized, using the frozen liver tissue obtained on the day of biopsy. Long-term outcomes were compared between patients with and without CP mutations of HBV at baseline. HBV genotyping showed that 100% of study subjects were infected with genotype C HBV. During a median follow-up period of 12.5 years, patients without double CP mutations of HBV at baseline showed a tendency towards achieving an earlier HBeAg seroconversion than those with (6.9 vs 9.4 years, P = 0.062) double CP mutations. Double CP mutations at baseline were also significantly associated with the eventual development of cirrhosis or hepatocellular carcinoma (P = 0.013), whereas the absence of double CP mutations predicted inactive carrier status at the last follow-up (P = 0.027). At 10 years, liver-related tests were also significantly better in patients without double CP mutations of HBV than in those with these mutations, as reflected by higher platelet counts and albumin levels (P = 0.036 and P = 0.044, respectively). Double T1762/A1764 mutations are significantly related to liver deterioration in HBeAg-positive genotype C active hepatitis patients. A longer period of immune clearance coupled with delayed HBeAg seroconversion appears to contribute to disease progression in patients harbouring these mutations in the CP region of HBV.
R-130a inhibits HCV replication in both replicon (1b) and J6/JFH1 culture systems. (a) miR-130a overexpression after transfection of miR-130a mimic in HCV Con1b replicon cells. (b) miR-130a overexpression in JFH-1 HCVcc system. (c) Overexpression of miR-130a inhibited HCV RNA replication in Con1b replicon cells. (d) Overexpression of miR-130a inhibited HCV RNA replication in JFH-1 HCVcc system. Cells were transfected for 48 h with 2 nM miR-130a mimic or microRNA mimic negative control (MMNC) by DharmaFECT4 (Dharmacon), after which cells were harvested and total RNA was extracted. The levels of miR-130a or HCV RNA and host cell U6 or GAPDH mRNA were measured by quantitative RT-PCR as described in 'Materials and Methods'. Data are presented as means ± SEM, n = 3. Error bars indicate standard error of mean (SEM). *P < 0.05; **P < 0.01 vs control and mock and MMNC (microRNA mimic negative control).
R-130a restores innate immune system to produce type I IFNs. (a) Overexpression of miR-130a increased endogenous IFN-α expression in HCV Con1b replicon. (b) Overexpression of miR-130a increased endogenous IFN-β expression in HCV Con1b replicon. (c) Overexpression of miR-130a increased endogenous IFN-α expression in JFH-1 HCVcc. (d) Overexpression of miR-130a increased endogenous IFN-β expression in JFH-1 HCVcc. Cells were transfected for 48 h with 2 nm miR-130a mimic or microRNA mimic negative control (MMNC) by DharmaFECT4 (Dharmacon), after which cells were harvested and total RNA was extracted. The levels of IFN-α or IFN-β and host cell GAPDH mRNA were measured by quantitative RT-PCR as described in 'Materials and Methods'. Data are presented as means ± SEM, n = 3. Error bars indicate standard error of mean (SEM). *P < 0.05; **P < 0.01 vs control and mock and MMNC (microRNA mimic negative control).
Overexpression of miR-130a decreased miR-122 expression in HCV Con1b replicon and JFH-1 HCVcc. Cells were transfected for 48 h with 2 nm miR-130a mimic or microRNA mimic negative control (MMNC) by DharmaFECT4 (Dharmacon), after which cells were harvested and total RNA was extracted. The levels of miR-122 and host cell U6 were measured by quantitative RT-PCR as described in 'Materials and Methods'. Data are presented as means ± SEM, n = 3. Error bars indicate standard error of mean (SEM). **P < 0.01 vs control and mock and MMNC (microRNA mimic negative control).
Hepatitis C virus (HCV) infection is a major cause of chronic hepatitis and hepatocellular carcinoma. Currently pegylated interferon (IFN) combined with ribavirin remains the best therapeutic approach, although patients infected with HCV genotype I may benefit from adding protease inhibitors as 'triple therapy'. MicroRNAs (miRNAs) are endogenous small noncoding RNAs that regulate gene expression and have recently been shown to play an important role in human innate immune response and as an antiviral in chimpanzees. We studied the effect of miR-130a on the HCV replication. We found that miR-130a significantly inhibits HCV replication in both HCV replicon and J6-/JFH1-infected cells. Over expression of miR-130a upregulated the expression of type I IFN (IFN-α/IFN -β), ISG15, USP18 and MxA, which are involved in innate immune response and decreased expression of miR-122, a well-defined miRNA promoting HCV production. In conclusion, miR-130a inhibits HCV replication/production by restoring host innate immune responses and/or downregulating pro-HCV miR-122. miR-130a might be a potential drug target by modulating host innate immune responses to combat HCV infection.
  Hepatitis C virus (HCV) nonstructural protein 5B (NS5B) is an RNA-dependent RNA polymerase (RdRp) that is involved in genome replication and virus assembly. NS5B contains a distinct loop (loop Λ2) at the beginning of the nucleoside triphosphate tunnel with a highly conserved lysine (K151). In this study, reverse genetic analysis revealed that substitution of Jc1 NS5B K151 for alanine (K151A) and aspartic acid (K151D) affected genome replication and infectious virus production. However, genome replication and virus production by Jc1 containing NS5B K151R remained unaltered. A major deletion in loop Λ2 abolished RNA replication, suggesting a role for this structural domain in NS5B polymerase activity. In conclusion, this study demonstrated that the conserved K151 modulates infectious virus production; and loop Λ2 is essential for the polymerase activity of NS5B.
The hepatitis C virus (HCV) establishes persistent infections despite strong activation of the innate immune system through TLR3 and other sensors. Therefore, we analysed regulatory mechanisms of TLR3-induced immune responses in nonparenchymal liver cells (NPCs). Effects of Interleukin-10 (IL-10), transforming growth factor beta (TGF-β) and immunoregulatory miR-155 on poly I:C-activated murine (C57BL/6) Kupffer cells (KC) and sinusoidal endothelial cells (LSEC) were assessed in vitro. NPCs were assayed for inflammatory and antiviral cytokines and T-cell (Balb/c)-activating factors. Gene expression of miR-155, IL-10, TGF-β and interferon sensitive genes (ISGs) in biopsies of patients with HCV was determined by qrt-PCR. TLR3-induced antiviral activity in murine NPCs was potently suppressed by IL-10 and TGF-β which correlated with decreased TLR3 expression and inhibition of NF-κB and IRF-3 activation. T-cell activation, induced by TLR3-activated NPCs, was also suppressed by IL-10 and TGF-β, which was associated with a down-regulation of CD80 and CD86. Pretreatment with IL-10 or TGF-β suppressed TLR3-induced miR-155 expression, which itself positively regulated poly I:C-mediated immune responses, thus counteracting IL-10 or TGF-β-induced immunosuppression. In addition, hepatic expression of miR-155 was elevated in chronically infected patients with HCV, was associated with an IL-28B SNP (rs12979860) and was inversely correlated with HCV serum load and ISG expression levels. As miR-155 is a key regulator of anti-inflammatory mechanisms that control innate and adaptive hepatic immune responses during HCV infection, miR-155 based therapies may represent a novel mechanism to control HCV in the future.
Thirty to 40% of patients with chronic hepatitis C have persistently normal alanine aminotransferase (PNALT). Even though traditionally considered as healthy people, most PNALT carriers actually have some degree of clinical progression and histological liver damage. We evaluated the clinical and histological outcome of a 17-year follow-up on a cohort of patients with chronic HCV infection and PNALT. Between 1994 and 2011, 70 PNALTs and 55 Hyper-alanine aminotransferase (ALT) subjects underwent a clinical, biochemical, virological and histological follow-up. At the end of the follow-up, all patients were alive. In the PNALT group, none of the patients developed hepatic decompensation, while 14.5% of Hyper-ALTs were diagnosed as affected by decompensated cirrhosis. No significant variation of the Metavir grading and staging scores was observed among PNALTs by comparing pre- and post-follow-up liver specimens. On the contrary, a significant increase in both Metavir grading and staging scores was noticed within the Hyper-ALT group. Finally, the analysis of IL28B single-nucleotide polymorphism rs12979860 revealed no difference between Hyper-ALTs and PNALTs in terms of frequency of C/C genotype. In conclusion, progression of chronic hepatitis C among PNALTs is slow or even absent, because at the end of the 17-year follow-up histological and clinical parameters had not worsened significantly.
High-mobility group box 1 (HMGB1) proteins are substantially up-regulated in acute and chronic hepatitis. However, the immunopathogenic role of HMGB1 in patients with chronic hepatitis B (CHB) has not been elucidated. In this study, using a cohort of 36 CHB patients, we demonstrated a crucial role for HMGB1 to modulate balance between regulatory T (Treg) and T helper 17 (Th17) cells via the toll-like receptor (TLR)-4-interleukin (IL)-6 pathway. Serum HMGB1 levels were dramatically higher in CHB patients and increased along with liver injury, inflammation and fibrosis. Notably, HMGB1 increased along with decreased Treg/Th17 cells ratios in the periphery or intrahepatic microenvironment, which provides a clue for HMGB1 to favour Th17 responses whereas inhibit Treg responses. For in vitro studies, serum pools were constructed with serum from CHB patients at an advanced stage, whereas peripheral blood mononuclear cells (PBMC) pools were constructed with cells from those at an early stage. CHB-serum significantly enhanced retinoic acid-related orphan receptor-γt (RORγt), whereas they inhibited forkhead box P3 (Foxp3) expression in CHB-PBMC, which could be reversed by blocking of HMGB1, TLR4, or IL-6. Besides, recombinant HMGB1 (rHMGB1) dose-dependently up-regulated RORγt whereas down-regulated Foxp3 expression in CHB-PBMC, and meanwhile, rHMGB1 enhanced TLR4 and IL-6 expression in CHB-PBMC. Moreover, the axis of HMGB1-TLR4-IL-6-Treg/Th17 required noncontact interactions between CD4 and non-CD4 cells. In addition, rHMGB1 down-regulated anti-inflammatory proteins on CD4(+) CD25(+) cells whereas up-regulated pro-inflammatory cytokines in CD4(+) CD25(-) cells. In summary, enriched HMGB1 in CHB patients shifts Treg/Th17 balance to Th17 dominance via the TLR4-IL-6 pathway, which exacerbates liver injury and inflammation.
Hepatitis B virus (HBV) infection and its sequelae remain a major health problem for Taiwan. The national hepatitis B (HB) vaccination programme was first launched in 1984 to combat the spread of this infection. This study examined the status of HBV infection amongst students at a Taiwanese university in 2005, 18 years after the implementation of a nation-wide mass HB vaccination programme. In 2005, 5875 new university entrants, who were born during the period 1 July 1976 to 30 June 1988, were subdivided into one of 12 one-year-interval birth-year cohorts. Each student was individually tested for serum hepatitis B surface antigen (HBsAg), Antibody to hepatitis B surface antigen (anti-HBs) and antibody to hepatitis B core antigen (anti-HBc) status. We observed a declining trend of past exposure to HB infection from 48.7% (1976 birth-year cohort) to 5.2% (1987 birth-year cohort). The prevalence of chronic HB infection also declined from 14.5% (1976 birth-year cohort) to 1.9% (1987 birth-year cohort). The prevalence of persistent HB immunity through (earlier) active vaccination declined from 72% (1984 birth-year cohort) to 41.6% (1987 birth-year cohort). The prevalence of HB infection-naïve individuals increased from 18.2% (1984 birth-year cohort) to 53.1% (1987 birth-year cohort). This study demonstrates that as the implementation of the mass HB vaccination programme in 1984, the incidence of HB infection in Taiwan has declined, although a 'waning-off' effect of serum anti-HBs to low or undetectable levels, which may not provide protection, amongst this student population has arisen, 18 years following the implementation of the nation-wide HB vaccination programme. Such a situation may mean that these individuals may not be effectively protected against future HB infection. A booster dose of HB vaccine, given 18 years following HB vaccination, perhaps even earlier, should be considered.
Thirty-seven chronic hepatitis C patients with virological relapse (VR) after previous interferon-alpha (IFN) or IFN/ribavirin (Riba) therapy, were re-treated. Patients were randomized for either IFN/Riba and amantadine (Ama) including a 2-week initial high IFN induction course (18 MU IFN daily) (group A) or the same 2-week IFN induction course combined with Riba/Ama, followed by Riba/Ama without IFN (group B). Treatment duration for both groups was 24 weeks with a 24-week follow-up thereafter. The inclusion in group B was prematurely stopped because all patients (n = 10) relapsed within 2 weeks after stopping IFN. Therefore, all subsequent patients were included in group A (n = 27). In group A, 44% achieved a sustained virological response (SVR) and 29% of the patients with an end-of-treatment virological response had a VR again. Of all pretreatment characteristics, only genotype non-1 patients had a significantly higher chance of achieving SVR (P < 0.001). Of the characteristics during treatment only a negative hepatitis C virus (HCV)-RNA test result in transcription-mediated amplification (TMA) at week 6 had a high predictive value for SVR, 80% in all patients and 92% in genotype non-1 patients. In conclusion, hepatitis C patients with a VR to previous antiviral treatment can be successfully re-treated with IFN induction combined with Riba/Ama for only 6 months, when they have genotype non-1 and a negative HCV-RNA test result in TMA 6 weeks after the start of therapy. Riba/Ama combination therapy without IFN does not prevent VR after 2 weeks high IFN induction.
To describe the clinical and immunologic patterns of disease expression of patients with chronic hepatitis C virus (HCV) infection and positive antimitochondrial antibodies (AMA). We investigated the presence of AMA in 237 consecutive HCV patients with extrahepatic manifestations from an International Registry. AMA were detected by indirect immunofluorescence in triple rat tissue (liver, stomach and kidney), aceton-fixed criosections and FITC-conjugated rabbit anti-human immunoglobulins. We found positive AMA in 18 (8%) out of 237 HCV patients. All patients were female with a mean age at protocol inclusion of 65.8 years (ranging from 37 to 87 years). Twelve (67%) patients fulfilled classification criteria for systemic autoimmune diseases (SAD), including Sjögren's syndrome (n = 7), systemic sclerosis (n = 3) and systemic lupus erythematosus (n = 2). Fourteen (78%) of the HCV-AMA patients presented at least one of the highly suggestive characteristics of primary biliary cirrhosis (PBC): 9 (50%) had a specific M2 pattern, 6 (33%) had more than twice normal levels of alkaline phosphatase, 5 (28%) had raised IgM levels and 4 (22%) a histological pattern compatible with PBC. Five (28%) patients developed neoplasia after detection of AMA. Seven (39%) patients died, due to neoplasia (n = 4), cirrhotic complications (n = 2) and hepatopulmonary syndrome (n = 1). We describe a subset of HCV patients with positive AMA who presented a broad spectrum of clinical features, including liver, autoimmune and neoplasic manifestations. Two-thirds of these patients presented an associated SAD, mainly Sjögren's syndrome or systemic sclerosis, together with a high frequency of multiple autoantibodies and an increased prevalence of cirrhosis and neoplasia.
Hepatocellular apoptosis plays a major role in the pathogenesis of chronic hepatitis C. It can be measured noninvasively by determining the circulating levels of cytokeratin-18 fragments. We hypothesized that the effect of antiviral therapy on this parameter will be different in patients with a sustained virological response, relapse (REL) and nonresponse (NR). We quantified cytokeratin-18 fragments in plasma of patients participating in the Swiss Hepatitis C cohort, who received antiviral therapy without stopping because of sides effects. A total of 315 patients were included, 183 with a sustained response, 64 with NR and 68 who relapsed. Mean levels ±SD of circulating cytokeratin-18 fragments before therapy were 174 ± 172 U/L for responsders, 188 ± 145 for nonresponders and 269 ± 158 U/L for patients who relapsed. The values were significantly higher in the REL group (ANOVA P < 0.006). A sustained response was associated with a significant improvement of the plasma levels (94 ± 92 U/L, paired test P < 0.000001), whereas there was no improvement in the nonresponder group (183 ± 158 U/L) and in the relapser group (158 ± 148 U/L). There was a weak correlation between alanine aminotransferase (ALT) and cytokeratin-18 fragment levels (r² = 0.35, P < 0.000001) before therapy but not after therapy and none with hepatitis C virus (HCV) viremia. Successful antiviral therapy results in a significant decrease in circulating levels of cytokeratin-18 fragments arguing for a reduction in hepatocellular apoptosis after clearance of the HCV. Baseline cytokeratin-18 fragment levels are higher in relapsers. Correlations with ALT are weak, suggesting that these two tests measure different but related processes.
In patients with chronic hepatitis C, alanine aminotransferase (ALT) levels do not accurately reflect the extent of liver inflammation. The discrepancy between ALT level and liver damage could be related to the mode of cell death. In the present study, we quantified serum levels of apoptotic cytokeratin 18 (CK-18) neoepitopes that are generated by activated caspases during apoptosis. Apoptotic CK-18 neoepitopes were quantified by enzyme linked immunosorbent assay in sera from patients with chronic hepatitis C and elevated ALT levels (n = 72), patients with chronic hepatitis C and persistently normal ALT levels (n = 27) and healthy controls (n = 19). Serum CK-18 neoepitope levels were strongly correlated with ALT (r = 0.659, P < 0.0001) and the histology activity index (r = 0.374, P < 0.001). Patients with chronic hepatitis C and persistently normal ALT levels had higher apoptotic CK-18 neoepitope levels than healthy controls (P = 0.03) but lower levels than patients with chronic hepatitis C and elevated ALT levels (P < 0.001). Highest serum CK-18 neoepitope levels were observed in patients with cirrhosis (P = 0.002). Hence apoptotic CK-18 neoepitopes in serum of patients with chronic hepatitis C are associated with ALT level and histological liver damage. Serum apoptotic CK-18 neoepitope levels are elevated both in patients with chronic hepatitis C and elevated ALT levels as well as in patients with normal ALT levels indicating that also patients with chronic hepatitis C and normal ALT have an increased hepatocyte loss by apoptosis.
The aim of this work was to analyse apoptosis rate, measured by the serum levels of proapoptotic interleukin (IL)-18 and of soluble Fas (sFas), as well as of anti-inflammatory IL-10, in patients with chronic hepatitis C, at baseline and after treatment with interferon alpha and ribavirin. Twenty-seven patients with biopsy-proven chronic hepatitis C were studied, at baseline and after treatment with interferon alpha (21 cases) or pegylated interferon (6 cases) plus ribavirin. A group of 15 healthy sex- and age-matched individuals was selected as control. Serum concentrations of sFas, IL-10 and IL-18 were determined by ELISA in sandwich. The relationship of these molecules to necro-inflammatory and fibrotic activity was evaluated. Evolution of the serum concentrations of these molecules was analysed after treatment. Significantly increased serum concentrations of sFas were detected in patients with chronic hepatitis, compared with controls. Levels of this molecule were significantly correlated with necroinflammatory activity. Likewise, concentrations of IL-10 were significantly increased in the group of patients, compared with controls. Treatment with interferon and ribavirin induced a significant decrease of IL-18 concentration independently of the viral response. In contrast, levels of sFas decreased only in those patients with sustained response to therapy. Finally, baseline levels of IL-10 were significantly increased in patients without response to treatment, compared with those with sustained response, but the concentration did not change with the treatment. Increased serum levels of IL-10 are a negative prognostic marker of response to hepatitis C treatment. A significant decrease of apoptotic rate, as determined by sFas, can be expected in patients with a response to therapy.
We present a patient with an unusual course of hepatitis B e antigen (HBeAg)-negative chronic hepatitis B who had repeated reactivations of his disease progressing to cirrhosis with terminal liver failure. Each flare up presented like an acute hepatitis with very high titres of hepatitis B virus (HBV) and high inflammatory activity followed by rapid clearance of viraemia. The pre-core genome of HBV isolated from sera during 5 years of follow up was analysed. Direct sequencing of polymerase chain reaction (PCR) products derived from consecutive sera showed a rare pre-core stop-codon mutation at nucleotide (nt.) 1897 G --> A with an accompanying mutation nt. 1857 C --> T as well as a stop-codon mutation nt. 1896 G --> A. By cloning and sequencing of PCR products the mutant strain with mutation nt. 1897 was shown to predominate over viral strains with a mutation nt. 1896 during the course of disease, although the stop-codon mutation nt. 1896 in general is observed more frequently. Both mutations allow viral replication by stabilizing the encapsidation signal 'epsilon'. This allowed HBV replication at a very high level as observed during flare ups. The absence of HBeAg may be responsible for the massive cytotoxic T-cell response towards hepatocytes which might explain the rapid progression to liver cirrhosis although no, or very little, HBV replication was observed for long periods. However, there is no clear explanation as to why the nt. 1897 mutant strain overwhelmed the other virus strains.
MicroRNAs (miRNAs) are an abundant class of small nonprotein-coding RNAs with posttranscriptional regulatory functions as tumour suppressors and oncogenes. Aberrant expression and structural alteration of miRNAs are thought to participate in tumourigenesis and cancer development. It has been suggested that the presence of single-nucleotide polymorphisms in precursor miRNAs (pre-miRNAs) can alter miRNA processing, expression, and/or binding to target mRNA and represent another type of genetic variability that can contribute to the development of human cancers. Recent studies have indicated that the miR-196a-2 rs11614913 (C→T) polymorphism could alter mature miR-196a-2 expression and target mRNA binding. To determine the association of the miR-196a-2 rs11614913 polymorphism with the risk of hepatocellular carcinoma (HCC) development in a Turkish population, a hospital-based case-control study was designed consisting of 185 subjects with HCC and 185 cancer-free control subjects matched for age, gender, smoking and alcohol status. The genotype frequency of the miR-196a-2 rs11614913 polymorphism was determined by using a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay. Our data shows that the CC genotype of the miR-196a-2 rs11614913 polymorphism is associated with increased risk of HCC development in this Turkish population (OR = 2.41, 95% CI: 1.30-4.50, P = 0.005). Furthermore, according to stratified analysis, a significant association was observed between the homozygote CC genotype and HCC risk in the subgroups of male gender (OR = 3.12, 95% CI: 1.53-6.34, P = 0.002) and patients with hepatitis B virus (HBV)-related HCC (OR = 2.88, 95% CI: 1.33-6.22, P = 0.007). Because our results suggest for the first time that the miR-196a-2 rs11614913 polymorphism may be a genetic susceptibility factor for HCC (especially in the male gender and HBV-infected patients) in the Turkish population, further independent studies are required to validate our findings in a larger series, as well as in patients of different ethnic origins.
Out of the 15 hepatitis E (HEV) epidemics that occurred during the years 1976-1995 in the Gujarat and Maharashtra states of India, 45.78% (76/166) stool samples showed the presence of HEV RNA. HEV RNA was found significantly more often in samples that were transported in liquid nitrogen (50.9%) compared with samples that were transported in wet ice (37.0%) (P < 0.05). Stool samples collected within 7 days after the onset of the disease (59.2%) were more often positive for HEV RNA when compared with samples that were collected 7-20 days after the onset of the disease (28.5%) (P < 0.01). It has been observed in experimentally infected Rhesus monkeys that they excrete HEV throughout the incubation period and for a variable length of time after the elevation of serum ALT levels. A similar situation is found in humans.
The presence or absence of antibodies to the second envelope protein (anti-E2) of hepatitis C virus (HCV) was determined in stored sera taken from a cohort of 87 Irish women with antibodies to HCV (anti-HCV) who were all infected by HCV genotype 1b from contaminated anti-D immunoglobulin given in 1977. Anti-E2 was found in 16 patients (100%) who were HCV RNA positive but only in 31 of 50 patients (62%) who were HCV antibody positive by recombinant immunoblot assay (RIBA) but HCV RNA negative. In the remaining 21 sera taken from women who had indeterminate recombinant immunoblot assays and who were repeatedly negative on testing for HCV RNA, anti-E2 was found in only three cases (14%). This suggests that loss or absence of anti-E2 may be useful in confirming clearance of HCV.
Although tattooing is recognized as a risk factor for transmission of hepatitis C, the efficiency with which transmission occurs is unknown. Sera stored from a serosurvey of tattooists undertaken in 1984 to test for human immunodeficiency virus (HIV) provided the opportunity to determine the prevalence of serological markers of hepatitis B virus (HBV) and hepatitis C virus (HCV) in tattooists at that time. The stored sera had been obtained from five unregistered and 36 of 37 (97%) of the registered tattooists operating in 1984. Serological status for hepatitis B (hepatitis B surface antigen (HBsAg), antibody to hepatitis B surface antigen (HBsAb) and antibody to hepatitis B core antigen (HBcAb) in standard assays) or hepatitis C (HCV antibody reactivity in second and third generation tests, confirmed by recombinant immunoblot assay) was determined. No sera was HIV positive or HBsAg positive. Of 35 specimens tested for HCV specific antibody, only two (5.6%) were positive despite markers of HBV in 48.6% of the same sera. As acute HBV infection was common amongst tattooists prior to 1984, it is clear that hepatitis B vaccination would be of benefit to this group. Despite frequent needlestick injuries reported by tattooists at the time, the low seroprevalence of HCV in this group suggests that HCV may not be transmitted efficiently by intradermal inoculation using solid-bore tattooing needles.
The Andaman and Nicobar Islands, Union Territory of India, are home to six primitive tribes. Studies carried out earlier among these tribes revealed very high rates of hepatitis B infection. We have now studied hepatitis A and E infection among them. A total of 951 serum samples were collected from four accessible tribes (Nicobarese, Shompens, Onges and Great Andamanese) and tested for antibodies against hepatitis A and E viruses. In addition, 240 serum samples collected a decade earlier from age-stratified Nicobarese were also screened. Hepatitis A virus (HAV) infection was found to be highly endemic among all the tribes, whereas hepatitis E virus (HEV) infection was common among the Nicobarese and Shompens. The age group-wise prevalence of these infections among the Nicobarese showed different patterns, HAV prevalence rising significantly from those aged 10 years and thereafter reaching a plateau, whereas HEV prevalence was found to be more evenly distributed over all age groups, but rising somewhat after 30 years of age. Over the last decade, the prevalence of HAV among the Nicobarese has declined slightly, particularly in those aged 10 years or less whereas HEV infection has more than doubled over all age ranges. Different HEV prevalence observed among the tribes could not be attributed to differences in sanitation or water supply. This fact and the different age-wise patterns of HAV and HEV prevalences is suggestive of different modes of transmission of HEV that are not shared. The highest rates for HEV were among those tribes which reared pigs suggesting that pigs might serve as reservoir of HEV. Further studies are needed, however, to validate these findings.
A retrospective study of 139188 blood donor records for the period 1991-99 was conducted in the largest blood bank in the federal state of Santa Catarina, in southern Brazil. The incidence/window method, based on 11286 repeat donors with 8917 person-years of follow-up, was used to estimate the residual risk for transfusing hepatitis B and C due to infectious window periods for early, mid and late years of the decade. The residual risk for transfusing HBsAg contaminated blood decreased almost three times over the 1990 decade but still remains very high at 1 : 2077 (95% confidence limits 1 : 1075-1 : 4624), with a corresponding incidence of 3.00 (1.35-5.77) per 1000 person-years. Similarly, although residual risk for hepatitis C was reduced more than 30 times in late 1990s, compared with the earlier period, the risk of 1 : 13721 (1 : 7102-1 : 30820) and corresponding incidence of 0.51 (0.23-0.99) per 1000 person-years are still very high compared to developed countries. In addition to vaccination against hepatitis B and health promotion efforts aimed at reduction of hepatitis transmission, special measures such as PCR screening of pooled blood donations might be needed to rapidly achieve a further residual risk reduction in high prevalence areas.
Crude incidence rates of hepatocellular carcinoma by age, sex and year of viral hepatitis notification, 1992-2007 HBV monoinfection N = 45 642 HCV monoinfection N = 87 980 HBV/HCV co-infection N = 3532
Age-period-cohort model* of hepatocellular carcinoma incidence
Chronic hepatitis B virus (HBV) and hepatitis C virus (HCV) infections are the major risk factors for hepatocellular carcinoma (HCC). We examined trends in the incidence of HCC among a population-based cohort of people infected with HBV or HCV. HBV and HCV cases notified to the New South Wales Health Department between 1992 and 2007 were linked to the Central Cancer Registry, Registry of Births, Deaths and Marriages, and National HIV/AIDS Registries. Crude HCC incidence rates were estimated using person-time methodology. Age-standardized incidence rates were calculated using the 2001 Australian population. Trends in incidence were examined using join point regression models. Between 1992 and 2007, 1201 people had a linked HCC record: 556 of those with HBV; 592 with HCV; 45 with HBV/HCV co-infection; and 8 with HIV co-infection. The overall age-standardized HCC incidence rates declined non-significantly from 148.0 (95% confidence intervals (CI) 63.7, 287.4) per 100,000 population in 1995 to 101.2 (95% CI 67.3, 144.6) in 2007 among the HBV monoinfected group and significantly from 151.8 (95% CI 62.4, 299.8) per 100,000 population to 75.3 (95% CI 50.8, 105.5) among the HCV monoinfected group. However, incidence rates in the HCV monoinfected group progressively increased from the period 1992-1997 to 2004-2007 when adjusted for age, sex, and birth cohort, and the total number of cases per annum continued to increase. Despite declines in the age-adjusted incidence rates of HCC over time, the absolute number of cases increased likely due to the ageing cohort and an increasing prevalence of both hepatitis B and C in Australia.
To report the prevalence and the risk factors for hepatitis C virus (HCV) infection in a hospital cohort of 2691 sexually human immunodeficiency virus (HIV)-infected patients. The patients were enrolled in the Lyon section of the French Hospital Database on HIV between 1992 and 2002. Baseline characteristics were analysed. The detection of HCV-antibodies (Ab) was used for diagnosis. The HCV-Ab prevalence rate was 5.7 and 12.89% for individuals infected by HIV after homosexual intercourse or heterosexual intercourse, respectively. HCV-Ab was three times more frequently found among patients infected with HIV after heterosexual intercourse compared with patients infected with HIV after homosexual intercourse (adjusted OR: 3.2, 95% CI: 2.28-4.62, multiple logistic regression). The risk of HCV infection among HIV-infected individuals differed according to sexual behaviour. The determinants associated with HCV transmission through the sexual route needs to be explored further.
Individuals infected with hepatitis C virus (HCV) need to be diagnosed well before developing end-stage liver disease to benefit from treatment. We aimed to ascertain what proportion of cases had been tested for HCV to inform on the effectiveness of current guidelines. Record linkage between national databases of HCV tests, hospital discharges and deaths identified 10,645 persons who were hospitalized or had died with mention of end-stage liver disease in Glasgow, Scotland, between 1993 and 2007. We estimated HCV test uptake and prevalence of HCV infection within the study population. The associations between both HCV test uptake and HCV-antibody status and sex, age group and deprivation quintile were estimated using logistic regression. We found that 43% of those hospitalized (n = 9153) and 23% of those who otherwise died (n = 1492) with first-time mention of end-stage liver disease had been tested for HCV during this period. Test uptake in those hospitalized increased from 13 (95% CI: 12-14%) in 1993-1997 to 58% (56-59%) in 2003-2007. The adjusted odds of being tested for HCV were significantly higher for men (OR=1.3, 95% CI: 1.2-1.5), for ages 25-54 (25-34 years: 2.7, 95% CI: 2.1-3.4; 35-44 years: 2.3, 95% CI: 2.0-2.6; 45-54 years: 1.5, 95% CI: 1.4-1.7) compared with 55+ years, and for those residing in the two most deprived quintiles (1.1, 95% CI: 1.0-1.2). Twenty-eight per cent of the HCV testees aged 25-44 years were HCV infected. These results highlight the continuing need for raising awareness among medical professionals for comprehensive HCV testing in patients with liver disease.
Hepatitis B (HB) is thought to be an expanding health problem in Russia. The incidence of infection was estimated from mandatorily reported HB cases in St Petersburg. The two-sided t-test for independent samples and the LOESS (locally-weighted regression) smoother were used to compare the age at infection for symptomatic, asymptomatic and chronic infections, by gender. The force of infection was estimated from seroprevalence data (907 sera taken in 1999) using a newly developed nonparametric method based on local polynomials, as well as an earlier method based on isotonic regression and kernel smoothers. With the local polynomial method, pointwise confidence intervals (95%) were constructed by bootstrapping. On average, men contracted HB infection at a significantly younger age than women (in 1999, 21.8 vs 22.7 years, respectively). The overall male to female ratio was 1.92. In 1999 the overall incidence almost doubled compared with the preceding years and tripled among the age groups with highest incidence (15-29-year olds: 85% of cases in 1999). The incidence increase was associated with a lower average age at infection (24.1 years in 1994 vs 22.1 years in 1999). The age and gender-specific force of infection estimates generally confirmed the incidence estimates and emphasized the usefulness of local polynomials to do this. Hence HB transmission in St Petersburg occurs mainly in young adults. The dramatic increase of infections in 1999 was probably due to injecting drug use. Without intervention, HB virus is expected to continue to spread rapidly with a greater proportion of female infections caused by sexual transmission. These trends may also provide an indication for HIV transmission.
The objective of the present study was to determine mortality because of end-stage liver disease (ESLD) in a nationwide population of HIV-infected patients, 7 years following the introduction of highly active antiretroviral therapy (HAART). All departments of internal medicine and infectious diseases from the GERMIVIC Study Group prospectively recorded all deaths in HIV-infected patients during 2003. Fifty-nine departments, following a total of 20 940 HIV-infected patients, participated in the study. Results were compared with those of previous surveys conducted using similar methodology in 1995, 1997 and 2001. Among 215 deaths observed during 2003, 101 (46.9%) were related to AIDS, 27 (12.6%) to ESLD and 87 (40.5%) to other causes. Mortality because of ESLD represented 23.7% of non-AIDS-related deaths. Patients dying from ESLD had chronic hepatitis because of hepatitis C virus (HCV) in 92.6% of cases and moderate (30-60 g) or high (>60 g) alcohol consumption (43.5% and 26.0%, respectively). In this population, deaths because of ESLD were 1.5% in 1995, 6.6% in 1997, 14.3% in 2001 and 12.6% in 2003. The prevalence of hepatocellular carcinoma as a cause of death remained high in 2003 but stable when compared with 2001 (25%vs 14.8%). Treatment of hepatitis C in patients who died from ESLD was more frequent in 2003 (44.4%) than in 2001 (26.3%). Seven years after the introduction of HAART, ESLD associated with HCV infections is a leading cause of mortality in HIV-infected patients, which did not increase between the years 2001 and 2003.
Coinfection with hepatitis B virus (HBV) is an important and preventable cause of chronic liver disease among HIV-infected patients. We calculated the prevalence of chronic HBV infection annually from 1996 to 2007 by age, gender, race/ethnicity, and HIV transmission risk in a multisite observational cohort study of HIV-infected patients. Prevalence of chronic HBV infection was calculated as the number of patients with a positive HBsAg or detectable HBV DNA divided by the number of patients tested using either one of these assays. Among 4467 (59%) patients tested for chronic HBV infection from a total of 7618 patients active during 1996-2007, median age was 38.5 years, 77% were men, 49% were white, 35% were black, 13% were Hispanic, and 53% were men who had sex with men (MSM). Overall, 8.4% tested positive for HBsAg or detectable HBV DNA. Annual chronic HBV prevalence during 1996-2007 ranged from 7.8% to 8.6% without a statistically significant trend. Overall, prevalence was greater among men compared with women; among whites, blacks, and persons of other race compared with Hispanics; among MSM compared with injection drug users and high-risk heterosexuals; and among patients aged 35-44 years compared with younger or older patients. MSM constituted the greatest fraction (63-72%) of all HBV-infected patients in the HIV Outpatient Study (HOPS) over the period. Of eligible patients, 5.8%, 23.4%, and 31.6% had received at least one dose of HBV vaccine by years 1996, 2002, and 2007, respectively. Despite the availability of an effective HBV vaccine for over two decades and long-standing recommendations for immunization of persons (with or without HIV infection) at risk for HBV, the prevalence of chronic HBV infection in this study has been largely unchanged over the past decade among patients in all groups, and overall was 20 times as high as the national population prevalence.
The authors present the results of an International Symposium on 'Viral Hepatitis', held in Warsaw on 24-25 October 1997 and dedicated to the scientific activity of Professor Adam Nowoslawski, the founder of the Polish school of Immunopathology, with many contributions to the viral hepatitis research. The symposium was divided into main sessions and poster reports which covered most of the topics in this field. This successful meeting has gathered many distinguished speakers from different countries and was attended by ca. 350 participants, mainly from Poland, but also from the neighbouring countries.
Autoimmune hepatitis is a disease of unknown cause. Apart from genetic markers such as HLA DR3 and HLA DR4, female predominance, hypergammaglobulinaemia and characteristic autoantibodies are diagnostic hallmarks. Several viruses have been discussed to induce autoimmune hepatitis, among them all major hepatotropic viruses, Epstein-Barr virus and herpes simplex virus. It seems that herpes viruses may be responsible in at least some cases of patients with autoimmune hepatitis type 2. Furthermore, hepatotropic viruses like hepatitis C and hepatitis D virus may cause autoimmune phenomena which are similar to those in idiopathic autoimmune hepatitis. LKM-1 antibodies in hepatitis C and LKM-3 antibodies in hepatitis D may cause diagnostic problems. LKM-1 antibodies in hepatitis C are directed either against cytochrome P450 2D6 or other yet unidentified microsomal antigens. As in hepatitis C the antimicrosomal autoantibody response in hepatitis D is more heterogeneous. These LKM-3 antibodies react with several epitopes on proteins of family 1 and 2 UDP-glucuronosyltransferases (UGT). Additional autoantibodies are seen in hepatitis D virus infection. Liver diseases are models to study autoimmune disease, drug-induced and virus-induced autoimmunity in humans.
In 1999, the Department of Health allocated additional funding to Health Authorities in England to expand hepatitis B immunization among injecting drug users (IDUs), with the aim of increasing coverage by 20%. In 2001, a vaccination programme for prison inmates in England was also instigated. Between 1998 and 2004 current IDUs participated in a series of annual unlinked anonymous surveys that recorded vaccine uptake (n = 11 383). The proportion self-reporting vaccine uptake rose significantly from 27% in 1998 to 59% in 2004 [adjusted odds ratio: 3.7 (95% CI 3.2-4.3); increase in uptake of 25% per annum (95% CI 22-27%)]. A second survey, which recruited 852 current IDUs from community settings in 2003/04, found that prisons were the most common source (38%) of vaccine doses, followed by drug services (28%) and general practitioners (17%), with only 14% receiving doses through needle exchanges. These data suggest that the 20% target of improving vaccination coverage has been met, with the prison vaccination programme likely to have made a substantive contribution in recent years. However, prevalence of antibodies to the hepatitis B core antigen was stable (21%) and is currently similar among the vaccinated and unvaccinated. Consideration needs to be given to improving community vaccination provision for IDUs, targeting recent initiates, and determining when surveillance data should indicate reductions in infection so that the effectiveness of the targeted strategy can be assessed.
Little is currently known about hepatitis C virus (HCV) test seeking behaviours at the population level. Given the centralized nature of testing for HCV infection in the province of Alberta, Canada, we had an opportunity to examine HCV testing behaviour at the population level on all newly diagnosed HCV-positive cases using laboratory data to validate the time and number of prior tests for each case. Record linkage identified 3323, 2937, 2660 and 2703 newly diagnosed cases of HCV infections in Alberta during 1998, 1999, 2000 and 2001, respectively, corresponding to age-adjusted rates of 149.8, 129, 114.3 and 113.7 per 100,000 population during these years, respectively. Results from secondary analyses of laboratory data suggest that the majority of HCV cases (95.3%) who were newly diagnosed between 1998 and 2001 were first-time testers for HCV infection. Among repeat testers, analysis of a negative test result within 1 year prior to a first of a positive test report suggests that 211 (38.4%) may be seroconvertors. These findings suggest that 339 or 61.7% of repeat testers may not have discovered their serostatus within 1 year of infection. Among this group, HCV testing was sought infrequently, with a median interval of 2.3 years between the last negative and first positive test. This finding is of concern given the risks for HCV transmission, particularly if risk-taking behaviours are not reduced because of unknown serostatus. These findings also reinforce the need to make the most of each test-seeking event with proper counselling and other appropriate support services.
The prevalence of hepatitis B virus (HBV) infection among persons with diabetes has not been assessed among the US population, despite increasing reports of HBV transmission in institutional care settings. Using national survey data, we found a 60% higher prevalence of HBV infection among persons with (vs without) diagnosed diabetes.
The majority of new and existing cases of hepatitis C virus (HCV) infection occur among people who inject drugs (PWID). Despite safe and efficacious HCV antiviral therapy, uptake remains low in this population. This study examined trends in HCV treatment uptake among a large national sample of PWID attending Australian Needle and Syringe Programs between 1999 and 2011. Annual cross-sectional sero-surveys conducted among PWID since 1995 involve completion of a self-administered questionnaire and provision of a dried blood spot for HCV antibody testing. Multivariate logistic regression identified variables independently associated with HCV treatment uptake among 9478 participants with both self-reported and serologically confirmed prior HCV infection. Between 1999 and 2011, the proportion currently receiving treatment increased from 1.1% to 2.1% (P < 0.001), while the proportion having ever received treatment increased from 3.4% to 8.6% (P < 0.001). Men were significantly more likely than women to have undertaken HCV treatment (P = 0.002). Among men, independent predictors of HCV treatment uptake were homosexual identity and older age; among women, independent predictors included homosexual identity and an incarceration history. Despite increases in HCV treatment among Australian PWID between 1999 and 2011, uptake remains low. Strategies are required to increase the proportion of PWID assessed and treated for HCV infection to address the increasing burden of disease. Specific approaches that target women may also be warranted. Continued surveillance of HCV treatment uptake among PWID will be important to monitor the roll-out of simple, safe and more effective HCV treatments expected to be available in the future.
Liver disease due to hepatitis C virus (HCV) infection is a leading cause of non-AIDS-related morbidity and mortality in patients infected with HIV. We assessed the frequency of and predictors for initiation of treatment for HCV infection among patients coinfected with HCV/HIV enrolled in the HIV Outpatient Study (HOPS) during 1999-2007. We included patients with confirmed HCV infection, at least 1 year of subsequent follow-up, and no evidence of prior HCV treatment. We assessed predictors of HCV treatment initiation using Cox proportional hazards analyses. During 1999-2007, 103 (20%) HOPS patients coinfected with HCV/HIV initiated HCV treatment during a median of 4.3 years of follow-up (interquartile range: 2.7, 6.7). In multivariable analysis, non-Hispanic black race/ethnicity (hazard ratio HR] 0.3; 95% confidence interval [CI] = 0.2, 0.6) was independently associated with a lower likelihood of HCV treatment. Elevated alanine aminotransferase (ALT; HR 3.5; 95% CI = 2.2, 5.6) and CD4+ cell count ≥500 cells/mm(3) (HR 1.8; 95% CI = 1.2, 2.8) at the start of observation were independently associated with higher likelihood of HCV treatment. For patients starting observation in 1999-2001, 2002-2004 and 2005-2007, 5%, 11% and 21% of patients initiated treatment during the first year of follow-up, respectively. Between 1999 and 2007, despite a stable low fraction of patients coinfected with HCV/HIV initiating treatment for HCV infection, an increasing proportion initiated treatment within the first year after the infection was confirmed. Treatment of HCV infection in patients coinfected with HCV/HIV should be considered a priority, given the increased risk of accelerated end-stage liver disease.
The aim of this study was to update our previous meta-analysis of interferon (IFN) in the treatment of hepatitis C and to analyse new factors, namely, HCV RNA end-point, patients with cirrhosis and patients with normal ALT. We use the Der Simonian and Laird method, with heterogeneity and sensitivity analyses. Seventy-six randomized control trials (RCTs) in naive patients were found but we focused our analysis on 59 RCTs with chronic hepatitis C (26 vs. controls and 33 comparing different regimens) and on seven RCTs in acute hepatitis. Interferon-alpha (IFN-alpha) at 3 MU thrice weekly (TIW) for 12 months exhibited 39% of virological end-of-treatment response (ETR) and 17% of virological sustained response (SR), respectively, vs. 1% and 3% in untreated controls (all P < 0.001). There was a significant dose effect (in favour of 6 vs. 3 MU TIW): the virological SR at 6 months were 35% in the 6 MU group (95% CI: 24-47) and 16% in the 3 MU group (95% CI: 8-27) and were at 12 months 43% in the 6 MU group (95%CI: 31-56) and 25% in the 3 MU group (95% CI: 16-37). There was a significant duration effect (12 vs. 6 months) upon the virological SR rate both at 3 and 6 MU: 3 MU provided 14% of virological SR (95% CI: 11-19) in the 12 months group vs. 7% (95% CI: 5-11) in the 6 months group and 6 MU provided 22% (95% CI: 17-29) and 16% (95% CI: 11-22) virological SR in the 12 and 6 months groups, respectively. Cirrhotic treated patients had 17% of virological SR (95 CI: 9-24%; P < 0.001) vs. 0% in controls and provided a 20% reduction rate (95 CI: -2% to -37%, P=0.03) in hepatocellular carcinoma incidence. In acute hepatitis C, a 3-month treatment with IFN-alpha showed significant efficacy vs. controls upon the virological SR rate (32% vs. 4%, P < 0.001). In conclusion, we confirm the dose and duration effect of IFN in chronic hepatitis C, and the efficacy of IFN-alpha in the treatment of acute hepatitis and in cirrhotic patients.
Re-treatment with interferon-alpha alone for chronic hepatitis C nonresponders to interferon-alpha monotherapy is almost ineffective. This multicentre, randomized, parallel-group, dose-finding study evaluated the efficacy of interferon-beta-1a in the treatment of chronic hepatitis C patients unresponsive to interferon-alpha. A total of 267 patients were randomized to one of four groups: subcutaneous interferon-beta-1a 12 MIU (44 microg) or 24 MIU (88 microg) administered three times weekly or daily. Patients were treated for 48 weeks and then followed up for an additional 24 weeks. There was a trend towards a dose-response relationship regarding virological [loss of detectable serum hepatitis C virus (HCV) RNA] and biochemical response (normalization of serum alanine aminotransferase). Overall, 22 patients (8.3%) had a virological response at the end of treatment; nine patients (3.4%) had a sustained virological response (SVR). Strikingly, 21.7% (5/23) of Chinese patients achieved SVR. Univariate analysis revealed that race was the only variable related to SVR [odds ratio (OR) 16.6; 95% CI 4.1-67.3; P < 0.0001]. Multiple logistic regression analysis also confirmed that more Chinese patients achieved SVR than non-Chinese patients (OR 12.3; 95% CI 2.6-59.3; P = 0.0017). In addition, complete clearance of HCV-RNA occurred earlier in Chinese than in non-Chinese responders (median 2 vs 30 weeks; P = 0.020). Thirty-six patients were withdrawn from treatment because of adverse events. Most adverse events were mild or moderate in severity. In conclusion, interferon-beta-1a provided considerable clinical benefit in Chinese patients with chronic hepatitis C unresponsive to interferon-alpha. The evaluation of interferon-beta-1a in this setting is progressing.
Several studies have reported correlation between mutations in core and NS5A proteins of hepatitis C virus (HCV) and response to interferon (IFN) therapy. In particular, mutations in NS5A protein have been shown to correlate with responsiveness to IFN treatment of HCV-1b in Japanese patients. This study investigated whether amino acid (aa) mutations in the core and NS5A proteins of HCV-1a, 1b, 3a, 3b and 6f correlated with the response to pegylated interferon (Peg-IFN) plus ribavirin (RBV) therapy in Thai patients. The entire sequences of core and NS5A of HCV from 76 HCV-infected patients were analysed in comparison with corresponding reference sequences. The data revealed that the number of aa mutations in full-length NS5A, its C-terminus, IFN sensitivity-determining region, variable region 3 (V3) and V3 plus flanking region of HCV-1b NS5A protein were significantly higher in responders than in the treatment failure group (P = 0.010, 0.031, 0.046, 0.020 and 0.006, respectively). Similar results were found in a putative protein kinase R binding domain region in HCV-6f NS5A protein (P = 0.022). Moreover, specific aa substitutions in NS5A that appeared to be associated with responders or the treatment failure group were observed at positions 78 and 305 for HCV-1b (P = 0.028), 64 and 52 for HCV-1a (P = 0.033) and 6f (P = 0.045). Nevertheless, analysis of aa sequences of core protein revealed highly conserved sequences among HCV genotypes and no significant differences between the viruses from responders and the treatment failure group. Our findings indicate that mutations in aa residues of NS5A of HCV-1a, 1b and 6f correlated well with responsiveness to Peg-IFN and RBV combination therapy.
The chimpanzee is the only recognized animal model for the study of hepatitis C virus (HCV). However, recently it was reported that rhesus monkeys were susceptible to HCV and developed hepatitis during infection. In the present study, we inoculated two rhesus monkeys each with HCV strain H77 (genotype 1a), strain HC-J6 (genotype 2a) or strain S52 (genotype 3a). Weekly serum samples were tested for liver enzyme values, HCV antibodies and HCV RNA. We did not find evidence of HCV infection in any of the monkeys during 24 weeks of follow-up. Our study demonstrates that rhesus monkeys are not readily infected with HCV and apparently do not represent a useful animal model for the study of HCV.
The nucleotide sequence diversity present among hepatitis C virus (HCV) isolates allows rapid adjustment to exterior forces including host immunity and drug therapy. This viral response reflects a combination of a high rate of replication together with an error-prone RNA-dependent RNA polymerase, providing for the selection and proliferation of the viruses with the highest fitness. We examined HCV subtype 1a whole-genome sequences to identify positions contributing to genotypic and phenotypic diversity. Phylogenetic tree reconstructions showed two distinct clades existing within the 1a subtype with each clade having a star-like tree topology and lacking definite correlation between time or place of isolation and phylogeny. Identification of significant phylogenetically informative sites at the nucleotide level revealed positions not only contributing to clade differentiation, but which are located at or proximal to codons associated with resistance to protease inhibitors (NS3 Q41) or polymerase inhibitors (NS5B S368). Synonymous/nonsynonymous substitution mutation analyses revealed that the majority of nucleotide mutations yielded synonymous amino acids, indicating the presence of purifying selection pressure across the polyprotein with pockets of positive selection also being detected. Despite evidence for divergence at several loci, certain 1a characteristics were preserved including the length of the alternative reading frame/F protein (ARF/F) gene, and a subtype 1a-specific phosphorylation site in NS5A (S349). Our analysis suggests that there may be strain-specific differences in the development of antiviral resistance to viruses infecting patients who are dependent on the genetic variation separating these two clades.
Top-cited authors
Christoph Sarrazin
  • Goethe-Universität Frankfurt am Main
Gamal Esmat
  • Cairo University
Heiner Wedemeyer
  • Hannover Medical School
Maria Buti
  • University Hospital Vall d'Hebron
Thomas Berg
  • University of Leipzig