Biomedical subjects

F Zoulim

Publications and source records attributed to F Zoulim.

At least 19 recordsLinked to original sources

PubMed2006

Etanercept treatment for three months is safe in patients with rheumatological manifestations associated with hepatitis C virus.

OBJECTIVE: The treatment of the rheumatological manifestations associated with hepatitis C virus (HCV) remains difficult. To examine the safety of anti-tumour necrosis factor-alpha treatment, nine patients having rheumatological manifestations associated with HCV were treated with etanercept 25 mg twice a week for 3 months. METHODS: Five patients had a positive viral load at study entry (Group I), four were negative (Group II). Clinical data recorded were: disease duration, painful and swollen joint count, patient global and physician global assessment, the number of 18 specified fibromyalgia tender points and the Health Assessment Questionnaire score. Laboratory studies included checking for the presence of cryoglobulinaemia and transaminase levels. Quantitative HCV viral RNA was performed by real-time polymerase chain reaction (PCR). RESULTS: At 3 months, no patient was found to have evidence of increased hepatic inflammation based on serial serum transaminase levels. In the five patients from Group I with detectable HCV RNA, no significant viral load increase was observed. No reactivation was observed in the four patients from Group II with undetectable HCV RNA. The effect on the clinical rheumatological manifestations was more heterogeneous but appears to be lower than that observed in rheumatoid arthritis. CONCLUSION: In this phase II open short-term study, etanercept appeared to be safe in patients with articular manifestations associated with HCV.

Antirheumatic Agents↗

PubMed2006

A prospective study of the evolution of lamivudine resistance mutations in patients with chronic hepatitis B treated with lamivudine.

Lamivudine resistance has been described in subjects with chronic hepatitis B infections, associated with mutations in the viral polymerase gene. The objective of this study was to estimate the emergence rate of lamivudine-resistant viral strains and their consequences over a 2-year period. We evaluated 283 lamivudine-naïve subjects with chronic hepatitis B. Clinical and virological features were assessed at inclusion and every 6 months thereafter. Viral DNA was characterized using polymerase chain reaction (PCR)-based sequencing. Potential risk factors for the emergence of lamivudine resistance mutations were assessed using logistic regression analysis. The annualized incidence rate for viral polymerase mutations was 22%. The only independent risk factor identified was high viral load, at inclusion. Detectable viral DNA and elevated transaminases were more frequent in subjects harbouring mutant viral strains, and these underwent a lower rate of hepatitis B e seroconversion. All subjects responded favourably to treatment, with no difference in symptoms between the two groups. This prospective cohort study identified lamivudine-resistant mutations emerging in 22% of subjects, yearly, which were apparently not associated with clinical aggravation over the study period.

Adolescent↗

PubMed2006

Mutations affecting the replication capacity of the hepatitis B virus.

The genetic variability of the hepatitis B virus (HBV) encounters two compounding forces: a high viral copy number produced during active replication and the lack of proofreading activity in the HBV polymerase, resulting in a high mutational rate. A large pool of quasispecies is generated in which the fittest virus, i.e. the virus that replicates best, becomes the dominant species. Immune and antiviral selection pressures result in vaccine/immunoglobulin escape mutants and antiviral resistant variants. Viruses encoding changes associated with antiviral resistance often have reduced replication in vitro, but the accumulation of additional mutations helps restore viral fitness. These compensatory mutations may occur not only in the polymerase gene but also in other genes such as the overlapping envelope gene, the precore gene, or in regulatory regions such as the basal core promoter. In this report we aim to review the new findings that have appeared in recent months.

Hepatitis B↗

PubMed2006

In vitro characterization of the anti-hepatitis B virus activity and cross-resistance profile of 2',3'-dideoxy-3'-fluoroguanosine.

The fluorinated guanosine analog 2',3'-dideoxy-3'-fluoroguanosine (FLG) was shown to inhibit wild-type (wt) hepatitis B virus (HBV) replication in a human hepatoma cell line permanently expressing HBV. Experiments performed in the duck model of HBV infection also showed its in vivo antiviral activity. In this study, we investigated the mechanism of inhibition of FLG on HBV replication and its profile of antiviral activity against different HBV or duck hepatitis B virus (DHBV) drug-resistant mutants. We found that FLG-triphosphate inhibits weakly the priming of the reverse transcription compared to adefovir-diphosphate in a cell-free system assay allowing the expression of an enzymatically active DHBV reverse transcriptase. It inhibits more potently wt DHBV minus-strand DNA synthesis compared to lamivudine-triphosphate and shows a similar activity compared to adefovir-diphosphate. FLG-triphosphate was most likely a competitive inhibitor of dGTP incorporation and a DNA chain terminator. In Huh7 cells transiently transfected with different HBV constructs, FLG inhibited similarly the replication of wt, lamivudine-resistant, adefovir-resistant, and lamivudine-plus-adefovir-resistant HBV mutants. These results were consistent with those obtained in the DHBV polymerase assay using the same drug-resistant polymerase mutants. In conclusion, our data provide new insights in the mechanism of action of FLG-triphosphate on HBV replication and demonstrate its inhibitory activity on drug-resistant mutant reverse transcriptases in vitro. Furthermore, our results provide the rationale for further clinical evaluation of FLG in the treatment of drug-resistant virus infection and in the setting of combination therapy to prevent or delay drug resistance.

Adenine↗

PubMed2006

European multicenter evaluation of high-density DNA probe arrays for detection of hepatitis B virus resistance mutations and identification of genotypes.

Polymorphisms along the hepatitis B virus (HBV) genome have an impact on disease outcome, sensitivity to antiviral treatment, escape from vaccination, and laboratory diagnosis. We have designed a diagnostic tool based on duplex amplification of the whole HBV genome and a high-density DNA chip designed to detect 245 mutations, 20 deletions, and 2 insertions at 151 positions and to determine the genotype of the virus in serum. Assay performances were evaluated with 170 samples, characterized by determination of viral load and sequencing of the Pol, S, and precore genes and the basal core promoter. One hundred fifty-three samples (90%) could be amplified and analyzed by the chip. Only two samples with more than 10(3) genome copies/ml could not be analyzed. Genotype had no impact on analytical sensitivity. Reproducibility studies showed no difference between repeats for codon and genotype determination. Genotype determination by sequencing and the chip were concordant in 148 of 151 samples. Twelve thousand one hundred sixty-one codons were analyzed by both techniques. Only 89.4% could be determined by sequencing, and among the remaining 11,335 codons, 92.8% were identical by sequencing and the chip. Failures to identify an amino acid by the chip were mainly due to reduced hybridization efficiency attributed to unexpected polymorphisms. Optimization of the chip-based reagent for the analysis of the HBV genome is ongoing. This first evaluation showed that DNA chip technology can provide important information in relation to the clinical management of chronic hepatitis B.

DNA, Viral↗

PubMed2005

Rheumatological manifestations of hepatitis C: incidence in a rheumatology and non-rheumatology setting and the effect of methotrexate and interferon.

OBJECTIVES: To evaluate hepatitis C virus (HCV)-positive patients followed in a rheumatology department and to compare them with a similar population of HCV-positive patients who had never seen a rheumatologist, in order to describe the rheumatological symptoms present and the effects of methotrexate and interferon-alpha therapy. METHODS: We performed a retrospective study of clinical, radiological and biological data on 21 rheumatology patients (Group I) presenting symptoms consistent with a chronic inflammatory arthritis with a known HCV infection and compared them with 41 members of an HCV support association (Group II). RESULTS: Symptoms of myalgia, sicca syndrome, Raynaud's phenomenon or paraesthesias were similarly frequent in the two groups. However, inflammatory joint pain and joint swelling were more common in Group I. In this group rheumatoid factor was positive in 48%, antinuclear antibodies in 26%, cryoglobulin in 44% and a reduced complement level in 63%. The majority of patients from Group I treated with methotrexate demonstrated an amelioration of the rheumatological symptoms with few negative outcomes. Regarding interferon-alpha therapy and rheumatological symptoms-in Groups I and II respectively 50 and 66% demonstrated a deterioration, 33 and 30% showed no change and 17 and 4% showed an amelioration. CONCLUSION: Rheumatological symptoms are common in patients chronically infected with HCV. It is essential to individualize the role of treatment with interferon-alpha and to consider the use of methotrexate for difficult cases.

Adult↗

PubMed2004

Analysis of HCV co-infection with occult hepatitis B virus in patients undergoing IFN therapy.

BACKGROUND/AIM: Occult hepatitis B virus (HBV) infection is characterized by the presence of HBV DNA in the absence of hepatitis B surface antigen (HBsAg) in the patient serum. Although such infections have been identified in patients with chronic hepatitis C, the clinical significance of those co-infections is still not understood. Our aim was, therefore, to assess the prevalence and clinical consequences of occult HBV infection in chronic hepatitis C patients undergoing antiviral therapy. METHODS: The study population consisted of 53 HBsAg-negative patients with chronic hepatitis C treated with IFN/ribavirin or IFN/ribavirin/amantadine. Nine patients experienced a viral breakthrough (BT), 30 were non-responders (NR) and 14 were responders (R). HBV-DNA detection by PCR was performed using primers specific for the S region of the HBV genome and HCV-RNA detection by PCR with primers localised in both the 5'NC and core region of HCV genome, before, during and after treatment. Viral genome sequences were also studied. RESULTS: Occult HBV genomes were found in the serum of four of 53 (7.5%) patients, unrelated to anti-HBc status. No significant differences in biochemical, virological, or histological markers, age, duration of infection, were observed in patients with or without HBV DNA. There was an inverse correlation in the evolution of HBV DNA and HCV RNA levels. Direct sequencing showed that S gene of occult HBV presented mutations in the "a" determinant while no specific mutation in the core region of HCV was observed. None of the four patients co-infected with HBV and HCV were responders to anti-HCV therapy. CONCLUSION: In our clinical setting, the prevalence of occult HBV co-infection among patients with chronic hepatitis C was low and independent of the presence of markers of previous HBV infection. Further studies in larger cohort of patients are warranted to determine if occult HBV co-infection may be involved in HCV resistance to combination therapy.

Adult↗

PubMed2004

Evolution of hepatitis B viral load and viral genome sequence during adefovir dipivoxil therapy.

Phase II and III clinical trials of adefovir dipivoxil (ADV) for the treatment of chronic hepatitis B have shown that this hepadnavirus polymerase inhibitor is well tolerated and effectively suppresses hepatitis B virus (HBV) replication. We therefore analysed the evolution of viral load and the emergence of HBV polymerase mutants in a 22-patient subgroup from a phase III clinical trial of ADV for the treatment of HBeAg-positive chronic hepatitis B. HBV DNA serum titres were quantified using a real-time polymerase chain reaction (PCR) assay with molecular hybridization probes. Emergence of polymerase mutants was assessed by direct sequencing of the viral reverse transcriptase domain after PCR amplification of HBV DNA isolated from serum. Our results indicated that ADV therapy effectively suppressed HBV replication in these patients (median serum HBV decrease at week 48 of treatment = 4.3 log10 copies/mL). The initial drop of HBV DNA titres in serum at week 12 of ADV therapy seemed to be predictive of subsequent HBe seroconversion (P = 0.059). Neither viral breakthrough nor the selection of drug resistant mutants were observed during the study period. Our results showed that ADV administration for 48-72 weeks effectively suppresses HBV replication without the emergence of resistant viral mutants.

Adenine↗

PubMed2004

Effect of a combination of clevudine and emtricitabine with adenovirus-mediated delivery of gamma interferon in the woodchuck model of hepatitis B virus infection.

Our aim was to evaluate the antiviral effect of a combination of two nucleoside reverse transcriptase inhibitors, emtricitabine (FTC) and clevudine (L-FMAU), with the addition of an adenovirus-driven delivery of recombinant gamma interferon (IFN-gamma) in the woodchuck model of hepatitis B virus infection. Six woodchuck hepatitis virus (WHV)-infected woodchucks received L-FMAU (10 mg/kg) plus FTC (30 mg/kg) intraperitoneally for 8 weeks; six other animals received in addition an intravenous injection of a recombinant adenovirus vector expressing woodchuck IFN-gamma (Ad-IFN) at weeks 4 and 8. In the control group, two animals received Ad-IFN alone, two received adenovirus vector expressing the green fluorescent protein reporter gene, and one remained untreated. In less than 2 weeks, all woodchucks that received L-FMAU plus FTC showed a rapid and marked inhibition of viral replication, with a 4-log(10) drop in serum WHV DNA. In two animals, viremia remained suppressed for several months after the end of treatment. Similarly, a dramatic decrease in intrahepatic replicative intermediates of viral DNA was observed in the L-FMAU/FTC-treated groups. The additional administration of Ad-IFN led to increased inflammation in the liver but did not enhance the antiviral effect of the L-FMAU/FTC combination. In conclusion, therapies combining L-FMAU and FTC in WHV-infected woodchucks resulted in a potent and sustained antihepadnaviral effect both in the liver and in the blood circulation. However, no extra benefit of adding IFN-gamma gene transduction to the L-FMAU/FTC combination could be detected.

Adenoviridae↗

PubMed2003

Long term histological improvement and clearance of intrahepatic hepatitis C virus RNA following sustained response to interferon-ribavirin combination therapy in liver transplanted patients with hepatitis C virus recurrence.

BACKGROUND AND AIMS: A proportion of liver transplanted patients with recurrent chronic hepatitis have a sustained virological response to combination therapy with interferon plus ribavirin. However, the long term benefit of antiviral therapy with regard to hepatitis C virus (HCV) RNA clearance remains unknown in patients with HCV recurrence. This study examined the long term biochemical, virological, and histological outcome in transplanted patients with recurrent chronic hepatitis who had a sustained virological response to antiviral therapy. PATIENTS AND METHODS: Fifty four patients with recurrent hepatitis C were treated with antiviral therapy involving induction by combination therapy (interferon (IFN) plus ribavirin) for six months and maintenance ribavirin therapy for 12 months. Fourteen patients who had recurrent chronic hepatitis and sustained virological response to antiviral therapy were followed for three years after the end of antiviral therapy. Serum alanine aminotransferases were assessed every three months during the observation period. Serum hepatitis C RNA detected by polymerase chain reaction was evaluated every six months during follow up, and protocol biopsy procedures were performed routinely every year. Semiquantitative histopathological assessment of allograft hepatitis was performed using the Knodell score and HCV was also detected by polymerase chain reaction on frozen graft tissue samples. RESULTS: At the end of antiviral therapy, the sustained response rate was 26%. A complete response (normal serum alanine aminotransferase level and undetectable serum HCV RNA) was achieved in 13/14 (93%) patients three years after the end of treatment. A comparison of liver histology findings before and after a mean of three years after antiviral therapy showed a clear improvement in 12/14 (86%) patients. In 5/14 (36%) patients, the last biopsy showed normal or near normal histological findings. After three years of follow up, the total Knodell score was 3.2 (range 1-8) versus 8.3 (range 5-12) before treatment (p=0.001). Graft HCV RNA was detectable before treatment in all 14 patients and was undetectable at the end of follow up in 13/14 (93%) patients tested. CONCLUSION: In patients with biochemical and virological responses induced by ribavirin and interferon, a complete response was sustained in 93% for at least three years after cessation of therapy. This long term response was associated with absence of detectable intrahepatic hepatitis C RNA and marked histological improvement.

Adult↗

PubMed2002

Animal models for the study of HBV infection and the evaluation of new anti-HBV strategies.

BACKGROUND: Our aim was to evaluate the anti-HBV activity of a novel L-nucleoside analog, 2',3'-dideoxy-2',3'-didehydro-beta-L-5-fluorocytidine (beta-L-Fd4C), in study models of HBV infection. METHOD: Its mechanism of action was evaluated on the in vitro expressed duck HBV (DHBV) reverse transcriptase and in primary hepatocyte cultures of duck and human origin. The capacity of antiviral therapy to clear viral infection was analyzed in vivo in the duck and woodchuck models. RESULTS: beta-L-Fd4C-TP exhibited a more potent inhibitory effect on the RT activity of the DHBV polymerase than other cytidine analogs (lamivudine-TP, ddC-TP, beta-L-FddC-TP). In primary duck hepatocyte cultures, beta-L-Fd4C exhibited a long-lasting inhibitory effect on viral DNA synthesis but could not clear viral cccDNA. In vivo treatment with beta-L-Fd4C in infected ducklings and woodchucks, induced a greater suppression of viremia and intrahepatic viral DNA synthesis than with lamivudine. However, covalently closed circular DNA persistence explained the relapse of viral replication after treatment withdrawal. Viral spread was strongly reduced in the case of early therapeutical intervention, but the number of infected cells did not decline when therapy was started during chronic infection. Liver histology analysis showed a decrease in the inflammatory activity of chronic hepatitis while no ultrastructural modification of liver cells was observed in electron microscopy studies. Furthermore, in human primary hepatocyte cultures, beta-L-Fd4C induced a significant inhibition of HBV DNA synthesis. CONCLUSION: beta-L-Fd4C is a potent inhibitor of hepadnavirus RT and inhibits viral DNA synthesis in hepatocytes both in vitro and in vivo. These experimental studies allowed as to show that beta-L-Fd4C is a promising anti-HBV agent. Combination therapy should be evaluated to eradicate viral infection.

Animals↗

PubMed2001

Duck hepatitis B virus polymerase gene mutants associated with resistance to lamivudine have a decreased replication capacity in vitro and in vivo.

BACKGROUND/AIMS: Hepatitis B virus mutants of the polymerase gene are frequently selected during lamivudine therapy for chronic hepatitis B. To study the biology of these mutants, we analyzed their replication capacity in the duck hepatitis B virus (DHBV) infection. METHODS: The B and C domain polymerase mutants corresponding to the clinical isolates were engineered by site directed mutagenesis in the DHBV genome in different expression vectors. RESULTS: The study of the enzymatic activity of the mutated viral polymerase polypeptides analyzed in a cell free system demonstrated a lower priming activity and a decreased capacity of elongation of viral minus strand DNA that was consistent with the lower replication capacity of these mutants in transfected leghorn male hepatoma cells compared to wild type genome. These mutants had a lower replication capacity in primary hepatocytes and in in vivo transfected ducklings. Although resistant to lamivudine, these mutants remained sensitive to PMEA. CONCLUSION: YMDD mutants of the DHBV reverse transcriptase have a decreased replication capacity both in vitro and in vivo, and are not cross-resistant to PMEA. These results may be important to design new antiviral strategies to combat the replication of the lamivudine resistant viral strains.

Adenine↗

PubMed2001

Successful combination therapy of polyarteritis nodosa associated with a pre-core promoter mutant hepatitis B virus infection.

BACKGROUND/AIMS: To describe the clinical and virological evolution of a polyarteritis nodosa (PAN) case associated with a hepatitis B virus (HBV) pre-core promoter mutant infection that was successfully treated with plasma exchanges, corticosteroids, and interferon alpha (IFN-alpha). METHODS: Viral markers were used, including HBV DNA quantified by the branched DNA assay and detected by PCR, the HBV genome sequence, pre-S1Ag and anti-HBC IgM which were studied throughout the treatment period and the entire follow-up in the serum, while the presence of virus in extrahepatic sites was detected by immuno-staining. RESULTS: The patient was infected with a typical pre-core promoter mutant harboring four point mutations. Pre-S1Ag was cleared rapidly from serum, most likely via the formation of immune complexes since HBV DNA declined more progressively. Viral infection was then cleared after a second episode of hepatocyte lysis. This was accompanied by a recovery from all clinical manifestations. CONCLUSIONS: The favorable treatment outcome observed in this first case of pre-core promoter HBV mutant associated PAN underlines that combination therapy based on IFN-alpha can clear pre-core promoter HBV infection and cure PAN. It also provides new insight in the pathogenesis of HBV associated PAN.

Adrenal Cortex Hormones↗

PubMed2001

High incidence of hepatitis B infections among chronic hepatitis cases of unknown aetiology.

BACKGROUND/AIMS: In approximately 5% of chronic liver disease cases, no aetiology can be identified. We selected sera from 50 patients with chronic hepatitis of unknown aetiology who were enrolled in this follow-up study whose aim is to gain insight into the possible role of viruses and to define potential clinical outcomes. METHODS: Patients' sera were screened with highly sensitive polymerase chain reaction assays for hepatitis B (HBV), C, D, and G viruses and TT virus. Sera were also retested for antibodies against the core antigen of HBV. RESULTS: Surprisingly, HBV DNA was detected in both serum and liver in 15/50 (30%) patients. Immunostaining for HBV antigens on biopsies from patients positive for HBV DNA showed HBcAg and/or HBsAg expression at low levels in 9/15 samples. Eleven of the fifteen patients were anti-HBc positive. With one exception, all patients carried HBV genomes at low levels (10(4) copies/ml or less). Histological signs of chronic liver disease were observed in all patients. CONCLUSION: Unrecognised HBV infections may account for a high proportion of chronic hepatitis cases of unknown aetiology. Improved HBV detection tests, which appear mandatory for the diagnosis and management of non-A non-E hepatitis as well as for improved safety of transfusions and transplantations are needed.

Adult↗

PubMed2001

Detection of hepatitis B virus resistance to antivirals.

The development of new nucleoside analogs, that inhibit the HBV reverse transcriptase activity, such as lamivudine, famciclovir and others, has provided recently an alternative to interferon therapy for chronic hepatitis B. However, due to the kinetics of viral replication with a high rate of virus production, a relatively long half-life of virus (CCC) DNA in the nucleus of infected hepatocytes, long-term antiviral therapy with a reverse transcriptase inhibitor is required to eradicate viral infection. Recently, it has been reported that lamivudine therapy for chronic hepatitis B in immune competent patients may be associated with the selection of resistant strains in aproximately 20% of the patients after 12 months of therapy. Sequence analysis of the reverse transcriptase domain of resistant viral strains, at the time of viral breakthrough, revealed the occurrence of mutations located in the YMDD motif within the C domain of the viral enzyme with a methionine to valine (M552V) or to isoleucine (M5521) change. Recent reports on larger series of patients pointed that other mutations residing outside of the C domain but mainly in the B domain of the viral polymerase (L528M) could be associated with these mutations in the YMDD motif. The lamivudine resistant mutants, selected in vivo, can be classified in 2 main groups: group I with a double mutation L528M and M552V, and group II with a single mutation M5521. In vitro studies performed in cell culture showed that these mutants have a decreased replication capacity and are indeed resistant to lamivudine. With the development of new antiviral options, genotyping assays and quantitative determination of viremia with highly sensitive assay are clearly warranted for an optimal monitoring of antiviral therapy of chronic hepatitis B. In view of the experimental and clinical data, the capacity of new antiviral strategies based on combination of new inhibitors, including adefavir and entecavir, with immune modulators needs to be further evaluated in animal models and clinical trials to prevent the emergence of resistant viral strains.

Animals↗

PubMed2001

Evolution of hepatitis C virus genome in chronically infected patients receiving ribavirin monotherapy.

Recent results of clinical trials suggest that combination of interferon and ribavirin exhibits an enhanced antiviral effect in the treatment of chronic hepatitis C. To investigate the effect of ribavirin on hepatitis C virus (HCV) infection, we analysed the evolution of the genetic heterogeneity of HCV in relation to the anti-HCV humoral response in patients treated by ribavirin alone. The study population included 35 patients with liver biopsy proven chronic hepatitis C infected with HCV genotype 1. Among them, 26 were treated with ribavirin for at least 12 months and nine untreated patients served as a control group. Serum samples were analysed before and at 6 and 12 months of therapy. Three regions of the HCV genome, i.e. HVR1, a domain of NS5A including part of the interferon sensitivity determining region (ISDR), and a segment of NS5B, were amplified by RT-PCR using specific primers. The PCR products were then studied using single-strand conformation polymorphism (SSCP) analysis followed by either direct sequencing, or cloning and sequencing. In parallel, the humoral anti-E1 response was studied using an ELISA (Innotest HCV E1Ab, Innogenetics). The results of HCV genome analysis showed no significant effect on the amino acid sequence evolution of the HVR1, NS5A and NS5B regions of HCV. Analysis of a phylogenetic tree from the major quasispecies variants showed the absence of correlation with ribavirin response, and the absence of selection of viral strains during ribavirin treatment. A trend towards a decrease in the anti-E1 Ab response was also observed. Altogether these results suggest that ribavirin may not exhibit a direct antiviral effect, but may trigger a favourable response to interferon by modulating the immune response against HCV.

Amino Acid Sequence↗

PubMed2001

Impaired allostimulatory function of dendritic cells in chronic hepatitis C infection.

BACKGROUND & AIMS: Dendritic cells (DC), which play an essential role in the triggering of primary antiviral immune reactions, may also contribute, in some viral models, to the propagation of viral infection and the pathogenesis of viral disease. During natural infection with hepatitis C virus (HCV), the interactions between the virus and DC may contribute to viral persistence, a general feature of HCV infection. METHODS: We compared the phenotypical and biological functions of monocyte-derived DC from patients with chronic hepatitis C (HCV-DC; n = 6), seronegative individuals (naive-DC; n = 8), long-term responders to antiviral therapy (LTR-DC; n = 8), and a group of patients with non-HCV-hepatic disorders (n = 11). The presence and the nature of HCV sequences during the DC cultures was assessed by reverse transcription-polymerase chain reaction and the analysis of the viral quasispecies distribution. RESULTS: Although HCV-DC displayed a normal morphology, phenotype, and capacity to capture antigen, their ability to stimulate the proliferation of allogeneic T cells was dramatically impaired in comparison with naive-DC (P = 0.0013). Mixing experiments revealed that HCV-DC did not affect the proliferation of T cells induced by naive-DC. Remarkably, the allostimulatory function of LTR-DC or DC from patients with non-HCV-hepatic disorders did not show any impairment. The presence of HCV genomic sequences could be documented for 5 of 6 HCV carriers either in the cells and/or the supernatants of the DC cultures. The presence of HCV sequences was found in the DC cultures from one patient showing a dramatic allostimulation defect. For that patient, extensive analysis of the viral quasispecies distribution revealed the presence, in the DC cultures, of genomic sequences of a unique nature, distinct from those identified in the patient's mononuclear cells, serum, or liver. CONCLUSIONS: Overall, these results indicate that chronic infection by HCV is associated with an allostimulatory defect of monocyte-derived DC, possibly because these cells constitute an extrahepatic reservoir for the virus. Although the exact mechanism responsible for such an alteration remains to be unraveled, our observations argue against an active immunosuppression-based mechanism.

Adenovirus E2 Proteins↗

PubMed2001

Initial amplification of duck hepatitis B virus covalently closed circular DNA after in vitro infection of embryonic duck hepatocytes is increased by cell cycle progression.

The relationship between the cell cycle and early amplification of duck hepatitis B virus covalently closed circular (CCC) DNA was studied after in vitro infection of fetal hepatocytes. We first showed that embryonic hepatocytes proliferated for at least 6 days after plating and that complete viral replication including CCC DNA amplification occurred in these proliferating cells. Addition of sodium butyrate or aphidicolin reversibly blocked cells in the G1 phase and diminished CCC DNA synthesis, which was restored after drug withdrawal, concomitantly with the entry of cells into S phase. Cell cycle progression of fetal hepatocytes can be triggered by stimulation with epidermal growth factor (EGF), hepatocyte growth factor (HGF), and tumor growth factor alpha (TGF-alpha). CCC DNA synthesis increased with progression to the S phase induced by EGF, HGF, and TGF-alpha alone or in combination. By contrast, tumor growth factor beta (TGF-beta) alone or in combination with EGF inhibited cell proliferation and viral DNA synthesis. By double labeling, viral nucleocapsids were found predominantly in bromodeoxyuridine-positive hepatocytes, indicating that high viral replication occurs preferentially in proliferating hepatocytes. CCC DNA was also detected mainly in cells in the S and G2/M phases separated from cells in the G1 phase by cell sorting. Taken together, these results show that hepatocyte proliferation may positively regulate the initial amplification of CCC DNA of avian hepadnaviruses, and may explain why mitosis is not necessarily associated with loss of CCC DNA.

Animals↗