Hervé Zylberberg

Soluble tumor necrosis factor receptors in chronic hepatitis C: a correlation with histological fibrosis and activity.

BACKGROUND/AIMS Tumor necrosis factor-alpha (TNF) is a mediator of inflammation and cellular immune response. Soluble TNF receptors (sTNFR) sTNF-R55 and sTNF-R75, which compete with cellular receptors for the binding of TNF, have been detected at high levels in infectious diseases including human immunodeficiency virus and HBV infection. In order to investigate the activation of the TNF system in HCV infection, we have analyzed the balance between TNF and sTNF-R in 60 HCV-infected subjects according to their clinical, biological, virological and histological characteristics. METHODS Serum TNF, sTNF-R55 and sTNF-R75 levels were determined by ELISA before any therapy and were compared to a control group of 60 healthy subjects and a group of 34 HBV-infected patients. RESULTS Mean TNF levels were 50.5+/-4.5 pg/ml in HCV patients, and undetectable (<5 pg/ml) in the control subjects. sTNF-R55 and sTNF-R75 levels were significantly higher in HCV-infected patients than in the controls: 2.88+/-0.14 ng/ml vs. 1.30+/-0.05, (p = 0.0001), and 9.54+/-0.58 ng/ml vs. 4.19+/-016, (p = 0.0001), respectively. sTNF-R55 and TNF-alpha levels in HCV patients were not significantly different from levels in HBV patients. sTNF-R75 levels were slightly lower than in HBV patients (9.54+/-0.58 vs. 11.4+/-0.79 ng/ml, p = 0.03). In contrast to other infectious diseases, there was no correlation between levels of sTNF-R and TNF. sTNF-R75 but not TNF levels were correlated with aminotransferases levels (p = 0.0001 and p = 0.0015 for aspartate and alanine aminotransferase, respectively), while sTNF-R55 levels were significantly correlated only with aspartate aminotransferase levels (p = 0.003). sTNF-R75 levels were significantly correlated with the Metavir activity index (p = 0.01), and sTNF-R55 and sTNF-R75 levels were significantly higher in patients with vs. without cirrhosis (3.22+/-0.21 vs. 2.54+/-0.17 ng/ml (p<0.02) and 11.6+/-0.86 vs. 7.5+/-0.53 ng/ml (p<0.001), respectively). sTNF-R55, sTNF-R75 and TNF levels were not correlated with viral load, genotype or response to interferon therapy. CONCLUSIONS Levels of soluble TNF receptors, and particularly sTNF-R75, are significantly correlated with the severity of the disease but not with virological parameters such as quantitative viremia and genotype. High TNF-R production could thus suggest that HCV-related liver disease involves immunological mechanisms, including activation of the TNF system.

Retrospective analysis of the impact of HIV infection and alcohol use on chronic hepatitis C in a large cohort of drug users

BACKGROUND/AIM This retrospective study aimed to better define the respective biological and pathological impact of human immunodeficiency virus infection and chronic alcohol consumption on the course of hepatitis C virus infection in intravenous drug users. METHODS Two hundred and ten consecutive anti-HCV positive intravenous drug users, among whom 60 were also anti-HIV positive, took part in the study at the University Hospital, Paris, France. RESULTS The activity of aspartate aminotransferase and gamma-glutamyl transpeptidase was significantly increased in serum from anti-HIV positive patients. The mean hepatitis activity index was significantly higher in anti-HIV positive patients (p<0.05), among whom there was also a higher proportion of patients with cirrhosis as compared to anti-HIV negative patients (30.0 vs 15.3%, p<0.0001). Excessive alcohol drinking (recorded in around 35% of the patients, whatever their HIV status), as compared to non-excessive drinking, was more often associated with cirrhosis in anti-HIV negative (24.5 vs 11.3%, p<0.05) than in anti-HIV positive patients (30.4 vs 29.7%, not significant). In a multivariate analysis, HIV infection (relative risk 2.2, confidence interval 1.1-4.5) and excessive alcohol drinking (relative risk 1.9, confidence interval 1.0-3.9) were the variables independently associated with the risk of cirrhosis. CONCLUSION Human immunodeficiency virus infection worsens the course of chronic hepatitis C in intravenous drug users. Excessive alcohol drinking also appears to be a crucial negative cofactor, and therefore alcohol withdrawal should be proposed as an integral part of the therapy.

Predictive factors for development of cirrhosis in parenterally acquired chronic hepatitis C: a comparison between immunocompetent and immunocompromised patients

BACKGROUND/AIMS The aim of this study was to evaluate the impact of the host immune status and of virological and environmental parameters on the development of cirrhosis during chronic hepatitis C virus infection. METHODS Liver histology (cirrhosis or not, Knodell score) was evaluated according to age, sex, route and age of contamination, alcohol consumption and immune status in a large series of 553 HBsAg-negative patients (whose duration of hepatitis C virus infection could be precisely evaluated) divided into three groups: group 1 consisted of 462 immunocompetent subjects (46.1% intravenous drug users, 53.9% transfused), infected for a mean of 12.5+/-6.7 years, including 16.6% of alcohol abusers (>80 g/day); groups 2a and 2b consisted of 91 immunocompromised patients, 52 human immunodeficiency virus-coinfected patients corresponding to group 2a and 39 kidney recipients undergoing immunosuppressive therapy for group 2b, having been infected by hepatitis C virus for a mean of 12.6+/-5.3 and 11.5+/-5.3 years, respectively. RESULTS Group 1: cirrhosis was present in 11.0% of group 1 patients and in 23.6% of immunocompetent patients with a duration of hepatitis C virus infection of 20 years or more. Forty-three percent of patients with cirrhosis and with hepatitis C virus infection for more than 20 years were alcohol abusers. The time taken to develop cirrhosis was 14+/-7 years in patients infected before the age of 40 years as compared to 8+/-5 years in those infected after 40 years (p<0.001). Groups 2a and 2b: cirrhosis was present in 19.8% of immunocompromised patients, a significantly higher rate than in immunocompetent patients (p<0.01). Alcohol abuse did not increase the risk of cirrhosis in this group. All patients but one were infected by hepatitis C virus before the age of 40 and the calculated time elapsed until the occurrence of cirrhosis was 12.4+/-5.5 years. In groups 1, 2a and 2b, there was no relation between histological severity, hepatitis C virus genotype and viral load. Four variables were independently associated with the occurrence of cirrhosis in the multivariate analysis: age over 40 at time of contamination (RR=9.3 in age range 40 to 59 and 91.2 in > or =60 years); long duration (> or =20 years) of hepatitis C virus infection (RR=15.4); alcohol consumption over 80 g/d (RR=2.9); and human immunodeficiency virus-coinfection (RR=2.6). CONCLUSIONS Our study on a large series of well-characterized patients provides an accurate evaluation of the risk of cirrhosis in parenterally-contaminated immunocompetent hepatitis C virus-infected patients, with an overall figure of 11%. It also demonstrates the impact of the host immune status on the risk of severe histological lesions during chronic hepatitis C virus infection. It finally establishes the importance of age at the time of viral infection in the occurrence of cirrhosis, as well as the importance of alcohol consumption. Thus, at least following parenteral infection, both host-related and environmental cofactors play a major role in the severity of the liver lesions associated with hepatitis C virus infection.

In vivo expression of a new hepatitis B virus protein encoded by a spliced RNA

Hepatitis B virus (HBV) is a small DNA virus with a compact genomic organization. All HBV proteins identified to date have been encoded by unspliced HBV RNAs. Spliced HBV RNAs have been described, but their functions are unknown. We show here that a singly spliced HBV RNA encodes a novel HBV protein in vivo. This HBV splice-generated protein (HBSP) corresponds to the fusion of a part of the viral polymerase and a new open reading frame that is created by the splicing event. In vivo, HBSP protein was found in HBV-infected liver samples, and anti-HBSP antibodies occurred in one-third of sera samples collected from chronic HBV carriers. In vitro, the ectopic expression of HBSP had no effect on viral DNA replication or transcription but induced cell apoptosis without a cell-cycle block. Overall, our results suggest that HBV has evolved a mechanism that directly modulates virus-cell interaction through RNA splicing.

Tritherapy for Human Immunodeficiency Virus Infection Does Not Modify Replication of Hepatitis C Virus in Coinfected Subjects

Triple antiretroviral therapy combining reverse transcriptase and protease inhibitors modifies the prognosis for human immunodeficiency virus (HIV) infection, with dramatic improvement in immune status. In an attempt to evaluate the impact of anti-HIV triple combination therapy on the course of hepatitis C virus (HCV)-related chronic hepatitis and on HCV replication, we studied the biological and virological characteristics of 22 HCV/HIV-coinfected patients who were given triple combination therapy. In comparison with baseline values, there was (1) a significant increase in the CD4 and CD8 cell counts and a decrease in the HIV RNA load and (2) no significant variation in aminotransferase activities or the HCV RNA load at 3, 6, or 9 months of tritherapy. Antiretroviral tritherapy seems to modify neither the biological activity of HCV-related chronic hepatitis nor the HCV load, despite immune restoration. Hepatic histopathologic analysis is warranted to assess the impact of immune restoration on liver lesions.

Hepatitis C virus-related fibrosing cholestatic hepatitis after renal transplantation

Fibrosing cholestatic hepatitis is a well-described syndrome in patients with immunodeficiency and chronic hepatitis B. It is clinically, biologically, and histologically characterized by rapidly progressive hepatic failure, a mildly elevated serum aminotransferase level, an extensive periportal fibrosis associated with intense cholestasis, mild inflammatory cellular infiltrate, no cirrhosis, and a high hepatocellular level expression of B viral antigens. This syndrome reflected a direct hepatocytopathic injury linked to high intrahepatic viral antigen expression. Because the syndrome of fibrosing cholestatic hepatitis has not been described in chronic hepatitis C, we report the first well-characterized case in a renal transplant patient with chronic hepatitis C and discuss the clinical and pathogenic implications of such a syndrome in this setting.

HBV genotypic resistance to lamivudine in kidney recipients and hemodialyzed patients.

BACKGROUND Lamivudine is a potent inhibitor of human immunodeficiency virus reverse transcriptase and hepatitis B virus (HBV) DNA polymerase. Its overall efficiency is clearly hampered by relapse at discontinuation and by risk of genotypic resistance. We describe herein the first cases of HBV resistance to lamivudine in kidney recipients and hemodialyzed patients. METHODS We analyzed 26 HBV-infected kidney recipients and five hemodialyzed patients treated with lamivudine who became serum HBV DNA-negative (by Digene test). The biological and virological follow-up identified breakthrough as defined by the reappearance of serum HBV DNA. In two cases of breakthrough, HBV DNA was amplified and sequenced through the polymerase domain, including the YMDD motif, before the beginning of treatment and at time of breakthrough to determine genotypic mutations. RESULTS Ten breakthroughs (reappearance of serum HBV DNA) were observed after a median follow-up of 11 months in eight kidney recipients and two hemodialyzed patients after a median duration of treatment of 16.5 (from 4 to 31) months of treatment. Previous HBe/anti-HBe seroconversion was not observed in the patients who escaped. In two kidney recipients, the comparison of HBV-DNA sequences before the treatment and after the breakthrough identified in one case a mutation of the highly conserved YMDD motif (YVDD), whereas in the second case, no genotypic mutation was observed in the sequenced region. CONCLUSION We report the first cases of HBV genotypic resistance to lamivudine in kidney recipients and hemodialysis patients. Genotypic resistance is observed after 4-31 months of therapy. The YMDD mutation does not account for all cases of virological escape.

Influence of HFE gene polymorphism on the progression and treatment of chronic hepatitis C

Summary.  We analysed liver histology findings in a large cohort of patients with chronic hepatitis C and in roughly half of them their response to interferon‐α‐based on iron parameters and HFE status. Histological activity and virological response to antiviral therapy (n = 146) were analysed in 273 immunocompetent and nonalcoholic patients with chronic hepatitis C, in terms of serum iron load, intrahepatic iron load (n = 110) and HFE mutations. Patients who were heterozygous for the C282Y and H63D mutations exhibited higher iron serum parameters than subjects without these mutations. The intrahepatic iron load was higher in H63D patients only. No association was observed between HFE mutations and histological activity. Increased iron parameters were associated with liver disease severity by univariate analysis only. Genotype 1 and ferritinaemia were associated with a poor response to antiviral therapy, whereas the H63D mutation emerged as a positive predictive factor for end of treatment and sustained antiviral response. Therefore, in chronic hepatitis C patients serum and intrahepatic iron levels were weakly correlated with histological activity, while HFE mutations were not. As for the response to interferon‐α, elevated ferritinaemia constituted a negative predictive factor whereas the H63D mutation was a positive one. The H63D mutation might form part of an immunogenetic profile influencing the response to interferon therapy.

Inflammatory cytokines and inhibitors in HIV infection : correlation between interleukin-1 receptor antagonist and weight loss

Objective:To determine serum levels of the interleukin-1 receptor antagonist (IL-1Ra), together with cytokines, other cytokine inhibitors and markers of immune activation in HIV-infected patients. Methods:Sixty-one HIV-patients were classified into Center for Disease Control and Prevention (CDC) groups A (n = 14), B (n = 14) and C (n = 33). Serum levels of IL-1Ra, IL-1β, IL-6, tumour necrosis factor (TNF)-α, TNF soluble receptors (TNF-sR) and IL-2sR were measured by enzyme-linked immunosorbent assay. CD4+ cell counts, p24 antigen, immunoglobulin (Ig) A, β2-microglobulin, triglycerides and neopterin were measured according to standard procedures. Weight variation was measured as the percentage of baseline weight lost or gained during the 3 months before sampling. Results:Serum levels of IL-1Ra were significantly elevated in HIV-infected patients, compared with control subjects (547 ± 104 and 133 ± 7 pg/ml), but did not vary significantly with the HIV disease stage, CD4+ cell count or p24 antigenaemia. IL-1Ra levels correlated with IL-1β(P < 0.005), IL-6 (P < 0.0001) and TNF-sR55 (P < 0.0001) levels, but not with those of TNF-α, TNF-sR75, IL-2sR, neopterin or IgA. IL-1Ra and IL-1Ra/IL-1β ratio were the only parameters significantly elevated (R = − 0.67, P < 0.0001) in the HIV-infected patients with marked weight loss (n = 12; mean of weight variation, −13.9 ± 2.1%) relative to the other patients, regardless of HIV disease stage and opportunistic infections. Conclusions:IL-1Ra levels are significantly elevated in HIV-infected patients, independently of immune deficiency. We propose that IL-1Ra accumulates in intense systemic inflammation, a state which does not seem to be reflected by the elevation of a single cytokine or the activation of a single cell system and which is correlated with marked weight loss.

Hepatocarcinoma-Intestine-Pancreas/Pancreatic Associated Protein (HIP/PAP) Is Expressed and Secreted by Proliferating Ductules as well as by Hepatocarcinoma and Cholangiocarcinoma Cells

Hepatocarcinoma-intestine-pancreas/pancreatic associated protein (HIP/PAP) gene was identified because of its increased expression in 25% of human hepatocellular carcinoma. HIP/PAP protein, a C-type lectin, binds laminin, acts as an adhesion molecule for hepatocytes, and has also been described as an acute phase secretory protein during acute pancreatitis in humans and rats. We investigated HIP/PAP protein expression in patients with various liver diseases associated with ductular reaction. At the same time, we analyzed patients with hepatocellular carcinoma and cholangiocarcinoma, and tested HIP/PAP protein levels in sera to establish the pattern of secretion. Our data show that HIP/PAP expression was not restricted to hepatocellular carcinoma, but was also detected in cholangiocarcinoma cells as well as in reactive non-malignant bile ductules. In contrast, HIP/PAP protein expression was undetectable in normal mature hepatocytes, but some ductular cells localized at the interface of portal tracts with parenchyma were HIP/PAP immunoreactive in normal liver. Finally, we present evidence that HIP/PAP serum levels were increased in 21/28 (75%) patients with hepatocellular carcinoma, and in 25/51 (49%) patients with nonmalignant cirrhosis. Altogether, these results suggest that HIP/PAP protein may be implicated in hepatocytic and cholangiolar differentiation and proliferation.