Fibrosis progression under maintenance interferon in hepatitis C is better detected by blood test than liver morphometry.

We evaluated whether quantitative measurements of liver fibrosis with recently developed diagnostics outperform histological staging in detecting natural or interferon-induced changes. We compared Metavir staging, morphometry (area and fractal dimension) and six blood tests in 157 patients with chronic hepatitis C from two trials testing maintenance interferon for 96 weeks. Paired liver biopsies and blood tests were available for 101 patients, and there was a significant improvement in Metavir activity and a significant increase in blood tests reflecting fibrosis quantity in patients treated with interferon when compared with controls - all per cent changes in histological fibrosis measures were significantly increased in F1 vs F2-4 stages only in the interferon group. For the whole population studied between weeks 0 and 96, there was significant progression only in the area of fibrosis (AOF) (P = 0.026), FibroMeter (P = 0.020) and CirrhoMeter (P = 0.003). With regards to dynamic reproducibility, agreement was good (r(ic) ≥ 0.72) only for Metavir fibrosis score, FibroMeter and CirrhoMeter. The per cent change in AOF was significantly higher than that of fractal dimension (P = 0.003) or Metavir fibrosis score (P = 0.015). CirrhoMeter was the only blood test with a change significantly higher than that of AOF (P = 0.039). AOF and two blood tests, reflecting fibrosis quantity, have high sensitivity and/or reproducibility permitting the detection of a small progression in liver fibrosis over two years. A blood test reflecting fibrosis quantity is more sensitive and reproducible than morphometry. The study also shows that maintenance interferon does not improve fibrosis, whatever its stage.

[Influence of treatment on long-term evolution of chronic hepatitis B]. / Influence du traitement sur l'évolution à long terme de l'hépatite chronique B.

The treatment of chronic hepatitis B is essentially based on the use of nucleoside or nucleotide analogues, which lead to viral suppression in the majority of cases. Viral suppression is associated with normal ALT values and progressive histological improvement of not only necroinflammatory lesions but also fibrosis. Regression of cirrhosis can be observed in severe cases. With the use of second-generation nucleoside or nucleotide analogues, the risk of mutation resistance is rare or inexistent. Finally, negativation and seroconversion of the HBs antigen can be observed in the medium-term. This seroconversion is usually associated with a decreased risk of complications and morbidity-mortality improvement.

[Hepatitis C, cirrhosis and hepatocellular carcinoma]. / Hépatite C, cirrhose et carcinome hépatocellulaire.

The screening for the detection of hepatocellular carcinoma is based on ultrasound sonography which should be realised in patients with post-hepatitis C cirrhosis with a delay between 3 and 6 months according to the most identified risk factors, in particular age and sex male. In the case of discovery of hypoechogen nodule < or = 1cm, a follow-up is mandatory because it is usually untypical by ultrasound sonography and to propose a liver biopsy in the case of an increasing in size is shown. The ultrasound guided cutting biopsy can precise the histological characteristics of the nodule, the grade, and indicate prognostic factors. The liver biopsy is also mandatory in the case of a nodule > 2 cm and when the ultrasound sonography is not contributive, especially when the nodule is between 1 and 2 cm in size.

Longitudinal evaluation of a fibrosis index combining MMP-1 and PIIINP compared with MMP-9, TIMP-1 and hyaluronic acid in patients with chronic hepatitis C treated by interferon-alpha and ribavirin.

We have recently described a fibrosis index combining serum procollagen type III N-terminal peptide (PIIINP) and matrix metalloproteinase 1 (MMP-1) concentrations for evaluating the amount of liver fibrosis in chronic hepatitis C patients. The aims of the present study were to validate this score in another cohort of patients and to assess its variations along those of TIMP-1, hyaluronic acid (HA) and MMP-9 during antiviral treatment. Seventy-nine patients treated by interferon-alpha and ribavirin for 24 or 48 weeks were included. A liver biopsy was performed within the 6 months before the start of treatment. Serum markers were measured in serum collected the day of the liver biopsy, at start of treatment, and every 3 months during treatment and a 6-month follow-up period. The PIIINP/MMP-1 index was significantly correlated to the METAVIR fibrosis (r = 0.68, P < 0.001). Its overall diagnostic value defined by the area under the receiver operating characteristics curves was 0.77 for discriminating F1 vs F2F3F4, and 0.81 for discriminating F1F2 vs F3F4, and was better than that observed for HA and TIMP-1. At the end of follow-up, the PIIINP/MMP-1 index significantly decreased in responders and remained stable in nonresponder patients. This decrease occurred early and continued regularly during the treatment period. This variation was because of both a decrease of PIIINP and an increase of MMP-1 concentrations. HA and TIMP-1 serum concentrations were also significantly lower at the end of follow-up in responder patients, but early changes were minimal and not influenced by the response to treatment. Our study shows that a noninvasive index combining PIIINP and MMP-1 is a useful tool to follow-up fibrosis change during and after antiviral therapy chronic hepatitis C patients.

Changes in histological lesions and serum fibrogenesis markers in chronic hepatitis C patients non-responders to interferon alpha.

BACKGROUND/AIMS: The aim of this study was to evaluate the changes in histological lesions and serum N-terminal peptide of type III procollagen (PIIINP) and hyaluronate (HA) levels in virologic non-responder patients treated by Interferon alpha (IFNalpha). METHODS: We enrolled 183 patients treated by IFNalpha and 56 controls, all with paired biopsy specimens. Yearly liver fibrosis progression was estimated before and during a follow-up of 1 year. RESULTS: By contrast to sustained responders, non-responders (n = 105) did not achieve improvement of histological scores after therapy. Their yearly fibrosis progression rate was similar before and during follow-up (0.18, 95%CI: 0.16-0.20, vs 0.26 (95%CI: 0.12-0.40) fibrosis units/year, NS), and was not different in controls (0.17, 95%CI: 0.06-0.27). The levels of PIIINP and hyaluronate (HA) remained unchanged during follow-up. Histological improvement was observed for the second biopsy in 25% of non-responders, but also in 23% of controls. This improvement was not correlated with decrease of ALT level, viral load, PIIINP, or HA. CONCLUSIONS: Our results suggest that IFNalpha therapy is unable to decrease PIIINP or HA levels and cannot improve the histological outcome in virologic non-responder patients. The histological improvement observed in a subset of patients may be linked to sample fluctuation or lack of reproducibility of histological scores.

[Hepatitis C virus culture systems]. / Les systèmes de culture du virus de l'hépatite C.

Hepatitis C virus pathogenesis and cycle are difficult to study because of the lack of culture system able to replicate efficiently the virus. Furthermore such a system will permit screen new antiviral drugs. Studies were realized to select cell culture system able to allow hepatitis C virus replication. Primary cell cultures and cell lines were used to performed HCV culture. Most of these works used lymphocyte and hepatocyte primary cultures or cell lines because of HCV tropism in these cells in vivo. Animals and arthropods cell lines were used as well for their capacity to bind and replicate HCV. The aim of this review is to present the different cell systems used to replicate HCV in culture and the results obtained.

Mosquito cells bind and replicate hepatitis C virus.

Several studies have demonstrated some hepatitis C virus (HCV) replication in lymphocyte and hepatocyte cell lines such as in African green monkey Vero cells. The aim of the present study was to select other cell lines able to bind and replicate HCV. Human hepatoma PLC/PRF/5 cells, human lymphoma Namalwa cells, Vero and mosquito AP61 cells were inoculated with HCV-positive plasma, washed six times and examined for the presence of the viral genome at different times post infection, using an RT-PCR method. Binding of HCV to cells was estimated by HCV RNA detection in cells 2 hr after inoculation and in the last wash of these cells. Successive virus passages in cells were carried out. All the cells studied were able to bind HCV but only AP61 and Vero cells provided evidence of replication and production of infectious virus: virus RNA was detected during 28 days post-infection in four successive virus passages. CD81 molecules, a putative HCV receptor, were detected by cytofluorometric analysis. Vero cells express CD81 molecules whereas these molecules were not detected on AP61 cells. It is suggested that other receptors are involved in HCV binding to Vero and AP61 cells.

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.

Better efficacy of a 12-month interferon alfa-2b retreatment in patients with chronic hepatitis C relapsing after a 6-month treatment: a multicenter, controlled, randomized trial. Le Groupe D'étude et De Traitement du Virus De L'hépatite C (Get.Vhc).

We studied the efficacy of three interferon alfa-2b (IFN-2b) regimens for the retreatment of patients with chronic hepatitis C (CHC) with prior complete response followed by relapse. Consecutive patients with CHC who had a complete biochemical response but relapse after a first course of 6 months of IFN with 3 million units (MU) given subcutaneously three times per week were enrolled in the study. Six to 24 months after the end of the first treatment, the patients were randomly assigned to receive IFN with either the same regimen (group 1), a regimen of 12 months with 3 MU (group 2), or a regimen of 6 months with 10 MU (group 3). Sustained biochemical response was defined as normal serum alanine transaminase (ALT) values during the follow-up and sustained virological response as a clearance of hepatitis C virus (HCV) RNA from the serum at the end of follow-up (6 months' posttreatment). Histological improvement was defined as a decrease of 1 point in Metavir score between the first liver biopsy and a biopsy performed at 6 months' postretreatment. Two hundred forty-seven patients were randomized: 75 to group 1, 91 to group 2, and 81 to group 3. In an intent-to-treat analysis, 12%, 36.3%, and 18.5% of patients had a sustained biochemical response after retreatment in groups 1, 2, and 3, respectively (P <.001); 13. 8%, 32.4%, and 17.2% of patients had a sustained virological response after retreatment in groups 1, 2, and 3, respectively (P <. 05). A low viral load and patients in group 2 were independently associated with a sustained biochemical response. A low Knodell score index before treatment, patients with a high level of ALT before retreatment, genotype 3, low viral load, and patients in group 2 were independently associated with sustained virological response. Younger age, a high level of ALT, a low level of gamma-glutamyl transferase before retreatment, low viral load, and patients in group 2 were independently associated with sustained biochemical and virological response. Among the 80 patients with repeated liver biopsies, 47.6% had improved histological activity scores; this improvement was associated with a sustained biochemical and virological response. In patients with CHC initially treated with 3 MU of IFN given subcutaneously three times per week over a 6-month period, and who subsequently developed a relapse after a biochemical response, retreatment with a regimen of 3 MU of IFN given three times per week for 12 months produced better biochemical and virological sustained response rates than regimens involving a higher dose or a shorter duration of retreatment. The biochemical and virological sustained response was associated with histological improvement.

Characteristics of patients with dual infection by hepatitis B and C viruses.

BACKGROUND/AIMS: The purpose of this study was to compare the epidemiological, biochemical, virological and histological characteristics of patients with chronic hepatitis B and C with those of patients suffering from chronic hepatitis C alone. METHODS: Twenty-three patients with chronic hepatitis C, who were anti-HCV positive and HBs antigen positive, were studied and subdivided into two groups according to the presence or absence of HBV DNA replication. They were compared to 69 age- and sex-matched patients with chronic hepatitis who were anti-HCV positive and HBs antigen negative. All patients were HCV RNA positive by PCR, anti-HIV negative and anti-HDV negative. HBV DNA and HCV RNA were detected in serum by means of a branched DNA assay and PCR. The HCV serotypes were determined by the Chiron Riba HCV serotyping SIA technique. The histological characteristics included the Knodell score. RESULTS: Epidemiological, biochemical and virological parameters were not different between the two groups. Only the prevalence of cirrhosis was greater in chronic hepatitis B and C patients than in patients with chronic hepatitis C alone (p = 0.01). Among chronic hepatitis B and C patients, HCV RNA level was significantly lower in HBV DNA positive than in HBV DNA negative patients (p = 0.01). Indeed, histological lesions were more severe in HBV DNA positive than in HBV DNA negative patients, including prevalence of cirrhosis (p = 0.01), Knodell score (p = 0.05) and, among the latter, piecemeal necrosis (p = 0.01) and fibrosis (p = 0.05). The characteristics of patients with dual infection did not differ according to the mode of contamination and duration of HBV disease, except for a shorter duration in patients contaminated by drug abuse than in other patients. CONCLUSIONS: These results suggest that HBV DNA replication inhibits HCV RNA replication in patients with chronic active hepatitis B and C but increases the severity of histological lesions.

Tenoxicam, a non-steroid anti-inflammatory drug, is unable to increase the response rate in patients with chronic hepatitis C treated by alpha interferon.

The purpose of this study is to compare a combination of interferon (IFN)-alpha2a (Roferon) + Tenoxicam with IFN-alpha2a alone in the treatment of chronic hepatitis C. This prospective, randomized double-blind study included 149 patients, all of whom were diagnosed with active chronic hepatitis C but non-cirrhotic (ALT > or = 1.5 upper limit of normal, anti-hepatitis C virus (HCV) positive by enzyme-linked immunosorbant assay2 and RIBA3). The patients were randomized in two groups, as follows: G1 (n = 76): IFNalpha2a 3 million units times per week during 6 months + placebo; and G2 (n = 73): IFNalpha2a 3 million units three times per week + Tenoxicam (20 mg/day) during 6 months. Alanine aminotransferase (ALT) and HCV RNA were determined before and at months 6 and 12 of treatment. 2'5' oligoadenylate synthetase activity (2'5' AS) was dosed in mononuclear cells before and at 3-month treatment intervals in 28 patients. Liver biopsy was performed before and 6 months after the end of therapy. Parameters were similar before therapy for both groups. Biochemical and virological responses were similar for both groups at month 6 (49.3% vs. 42.9% and 43.3% vs. 38.3%, respectively) and month 12 (28.3% vs. 23.8% and 17.2% vs. 17.5%, respectively). HCV RNA level significantly decreased in both groups at month 6, with no difference whatever the therapy; however, the HCV RNA level returned to initial values at month 12 and was the only significant prognostic factor of a sustained response. No peak of 2'5' AS activity was observed during treatment in patients with dual therapy. A histological improvement was also noted in both groups without difference, regardless of therapy. The percentage of adverse events was identical for both groups. Paracetamol intake, assessed in 80 patients, was 49.1 g per 6 months in the G1 group and 22.5 g per 6 months in the G2 group (not significant). In conclusion, the non-steroid anti-inflammatory drug, Tenoxicam, does not increase IFNalpha efficacy in the treatment of chronic hepatitis C. This combination is well tolerated and partially lowers Paracetamol intake, but not preexisting alpha-IFN adverse events.

EBV-B cells as antigen presenting cells in characterization of the self/donor context of allorecognition by T lymphocytes.

Alloreactivity is caused by T cell recognition of foreign histocompatibility antigens according to two models: (i) indirect recognition, in which processed allogeneic antigens are presented by self-major histocompatibility complexes like any other foreign antigen, and (ii) direct recognition, where the foreign MHC itself is recognized breaking the T cell recognition rule of self-restriction. This paper uses these two cases of alloantigen presentation as illustrative examples to investigate (i) the capacity of Epstein-Barr virus-transformed B cells (EBV-B cells) to process alloantigens, and (ii) in vitro assays with EBV-B cell lysate as a source of alloantigen, in order to characterize alloreactive T cell populations. A microculture system was established using donor EBV-B cell lysate as a source of the allogeneic antigen and donor or recipient EBV-B cells as antigen presenting cells to investigate whether alloantigen is recognized by effector T cells from the recipient. T lymphocytes produced after expansion in the presence of interleukin-2 from four samples of liver biopsies (three patients) and four samples of bronchoalveolar lavages (four patients) were used as effector cells. Upon human leucocyte antigen class II typing, these expressed the patient phenotype. When the T lymphocytes were from liver grafts, the recognition involved donor antigens presented by donor EBV-B cells (direct recognition). On the other hand, when the T lymphocytes were cultured from lung grafts, they mainly recognized antigens of donor EBV-B cell lysates in a self-restricted context (indirect recognition). These data suggest that EBV-B cells can provide allogeneic determinants recognized by T cells in donor or self-contexts, i.e. through either direct or indirect recognition.

[Value of high-dose interferon-alpha in chronic viral hepatitis C patients non-responder to a 1st treatment. Pilot study prospective and randomized trial]. / Intérêt de l'interféron alpha à forte dose chez les malades atteints d'une hépatite virale chronique C non répondeurs à un premier traitement. Etude pilote prospective et randomisée.

OBJECTIVES: The aim of this study was to evaluate the efficacy of two different doses of alpha interferon (IFN) for retreatment in chronic hepatitis C patients who were non responders to initial treatment by IFN at a dose of 3 MIU TIW for 6 months. METHODS: This open, pilot, prospective, randomized and bicentric study included patients with biopsy-proven chronic hepatitis C. Non response was defined as serum ALT levels > 2 upper limit of normal for the entire first treatment period, HCV RNA positivity by PCR at the end of the first treatment period, and the persistence of histologically-proven chronic active hepatitis after the first treatment period. Patients were randomized into two groups: group I received IFN alpha 2b 10 MIU TIW for 2 months, then 6 MIU TIW for 4 months, group 2 received IFN alpha 2b 6 MIU TIW for 6 months. RESULTS: Twenty three patients (17 male, 6 female, mean age: 38.7 +/- 9.1 years) were included: 14 were randomized in group 1 and 9 in group 2. Both groups were similar for the main clinical, biochemical, and histological variables. At the end of retreatment, 2 patients (14.2%) had biochemical and virological response in group 1 and 4 in group 2 (44.4%) (non significant). Only one biochemical and virological sustained response was observed in group 2 (11.1%) (non significant). There was no difference between the groups for complete and sustained response. An overall statistical significant improvement of Knodell score was observed (7.8 +/- 3.8 vs 9.6 +/- 3.2, P < 0.02) in the 18 patients who had a second biopsy 6 months after the end of therapy, while the Knodell score did not change at the end of the first treatment period. This improvement was statistically significant in group 2 (5.4 +/- 3.0 vs 9.2 +/- 9.5 before treatment, P < 0.02) and concerned intralobular necrosis (P < 0.05). The Metavir index did not change. The number of side-effects was similar in both groups. CONCLUSIONS: These results suggest that histological improvement may be obtained after IFN retreatment in some patients who are non-responders to the first treatment, despite an absence of biochemical and/or virological response.