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Hepatitis C virus (HCV)-induced iron overload has been shown to promote liver fibrosis, steatosis, and hepatocellular carcinoma. The zonal-restricted histological distribution of pathological iron deposits has hampered the attempt to perform large-scale in vivo molecular investigations on the comorbidity between iron and HCV. Diagnostic and prognostic markers are not yet available to assess iron overload-induced liver fibrogenesis and progression in HCV infections. Here, by means of Spike-in SILAC proteomic approach, we first unveiled a specific membrane protein expression signature of HCV cell cultures in the presence of iron overload. Computational analysis of proteomic dataset highlighted the hepatocytic vitronectin expression as the most promising specific biomarker for iron-associated fibrogenesis in HCV infections. Next, the robustness of our in vitro findings was challenged in human liver biopsies by immunohistochemistry and yielded two major results: (i) hepatocytic vitronectin expression is associated to liver fibrogenesis in HCV-infected patients with iron overload; (ii) hepatic vitronectin expression was found to discriminate also the transition between mild to moderate fibrosis in HCV-infected patients without iron overload., (© 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Background/aims: The life cycle of hepatitis C virus (HCV) is intimately linked to the lipid metabolism of the host. In particular, HCV exploits the metabolic machinery of the lipoproteins in several steps of its life cycle such as circulation in the bloodstream, cell attachment and entry, assembly and release of viral particles. However, the details of how HCV interacts with and influences the metabolism of the host lipoproteins are not well understood. A study was undertaken to investigate whether HCV directly affects the protein composition of host circulating lipoproteins., Methods: A proteomic analysis of circulating very low-, low- and high-density lipoproteins (VLDL, LDL and HDL), isolated from either in-treatment naïve HCV-infected patients or healthy donors (HD), was performed using two-dimensional gel electrophoresis and tandem mass spectrometry (MALDI-TOF/TOF). The results obtained were further investigated using in vitro models of HCV infection and replication., Results: A decreased level of apolipoprotein A-I (apoA-I) was found in the LDL fractions of HCV-infected patients. This result was confirmed by western blot and ELISA analysis. HCV cellular models (JFH1 HCV cell culture system (HCVcc) and HCV subgenomic replicons) showed that the decreased apoA-I/LDL association originates from hepatic biogenesis rather than lipoprotein catabolism occurring in the circulation, and is not due to a downregulation of the apoA-I protein concentration. The sole non-structural viral proteins were sufficient to impair the apoA-I/LDL association. Functional evidence was obtained for involvement of apoA-I in the viral life cycle such as RNA replication and virion production. The specific siRNA-mediated downregulation of apoA-I led to a reduction in both HCV RNA and viral particle levels in culture., Conclusions: This study shows that HCV induces lipoprotein structural modification and that its replication and production are linked to the host lipoprotein metabolism, suggesting apoA-I as a new possible target for antiviral therapy.

It has been suggested that hepatitis C virus (HCV) infected patients with type II mixed cryoglobulinemia have less extensive liver damage than patients without cryoglobulinemia. We retrospectively evaluated 35 patients with type II mixed cryoglobulinemia associated with HCV infection, seeking for factors associated with normal alanine aminotransferase (ALT) values. The presence of anti-GOR and of other autoantibodies, including the recently described anti-LAG-3.1, was specifically investigated. Fifty-four percent of patients had anti-GOR, 46% anti-LAG-3.1, 40% anti-smooth muscle, 17% anti-nuclear, and 11% anti-liver-kidney microsome 1 antibodies. Anti-GOR was significantly (p = 0.037) associated with anti-LAG-3.1 but not with other autoantibodies. Persistently abnormal ALT levels were observed in 54% of patients. By univariate analyses, abnormal ALT was significantly associated with anti-GOR positivity (p = 0.018) and younger age (p = 0.03). Multivariate regression analysis confirmed that these variables were independently associated with abnormal ALT. Our data suggest that the presence of autoimmune manifestations as well as unidentified age-related host factor(s) may protect from liver injury in HCV-associated cryoglobulinemia.

Hepatitis C virus (HCV) infection induces an acute and chronic liver inflammation through an immune-mediated pathway that may lead to cirrhosis and liver failure. Indeed, HCV-related hepatitis is characterized by a dramatic lymphocyte infiltrate into the liver which is mainly composed by HCV non-specific cells. Several data indicated that interferon (IFN)-gamma secretion by intrahepatic lymphocytes (IHL) may drive non-specific cell homing to the liver, inducing interferon inducible protein-10 (IP-10) production. An interesting hallmark of these IHL is the recruitment of lymphocytes associated with mechanisms of innate immunity, such as natural killer (NK), natural killer T (NKT) and gamma delta T lymphocytes. CD81 triggering on NK cell surface by the HCV envelope glycoprotein E2 was recently shown to inhibit NK cell function in the liver of HCV-infected persons, resulting in a possible mechanism contributing to the lack of virus clearance and to the establishment of chronic infection. In contrast, intrahepatic NKT cells restricted to CD1d molecules expressed on the hepatocyte surface may contribute to a large extent to liver damage. Finally, an increased frequency of T cells expressing the gamma delta T cell receptor (TCR) was observed in HCV-infected liver and recent observations indicate that intrahepatic gamma delta T cell activation could be directly induced by the HCV/E2 particle through CD81 triggering. These cells are not HCV specific, are able to kill target cells including primary hepatocytes and their ability to produce T helper (Th)1 cytokines is associated with a higher degree of liver disease. Together, CD1d/NKT and/or E2/CD81 interactions may play a major role in the establishment of HCV immunopathogenesis. In the absence of virus clearance, the chemokine-driven recruitment of lymphocytes with an innate cytotoxic behavior in the liver of HCV-infected patients may boost itself, leading to necroinflammatory and fibrotic liver disease.

HIV and hepatis C virus (HCV) coinfection is frequently associated with rapid progression of HCV-related disease, resulting in a higher risk of cirrhosis. Data suggest that natural T cells expressing the Vdelta1 T cell receptor rearrangement are recruited in the liver of chronically HCV-infected patients and are increased in the peripheral blood of HIV-infected persons. We studied gammadelta T cell distribution in the peripheral blood and liver of HCV-infected and HIV/HCV-coinfected patients in the presence and absence of antiretroviral therapy. We observed that Vdelta1+ T cells releasing helper T cell type 1 cytokines are compartmentalized not only in the liver of HCV+ patients, but also of HIV/HCV-coinfected persons. HIV/HCV patients showed an increased frequency of both peripheral and intrahepatic Vdelta1 natural T lymphocytes, resulting in a higher degree of hepatic inflammation when compared with patients with other liver diseases. Finally, highly active antiretroviral therapy (HAART) was unable to restore Vdelta1T cell circulation to normal levels in chronically HIV-infected persons. We conclude that gammadelta T lymphocytes released from tissue to the bloodstream circulation under the influence of chronic HIV infection may contribute to intrahepatic Vdelta1 compartmentalization and progression of liver disease, independently of HAART.

We have prospectively studied patients with type II cryoglobulinemia since 1985 to assess the efficacy of treatment with interferon-alpha at cumulative doses ranging from 234 to 849 MU. In the present study we retrospectively evaluated in this cohort parameters associated with complete response to therapy in 31 consecutive patients with type II cryoglobulinemia associated with hepatitis C virus (HCV) infection. Prevalence of complete response of cryoglobulinemia (disappearance of symptoms and signs of vasculitis and decrease of cryocrit below 10% of the initial value) was 62%, with a median response duration of 33 months and a range of 3 to 100 months. Three patients were putatively cured, as they remained in complete remission for more than 5 years off therapy. Eighteen patients (58%) had liver disease evidenced by histopathology and/or raised transaminase levels. Prevalence of normalization of transaminase levels was 100%, with a median response duration of 36 months. Relapse of hypertransaminasemia occurred in 100% and 8% of patients receiving less than or greater than 621 MU, respectively. By logistic regression analysis, the only pretherapy parameter that associated significantly (P = .0393) with complete response of cryoglobulinemia was the solitary anti-C22 (HCV core) antibody pattern, which was observed in 29% of patients. Association with older age and low cryocrit approached statistical significance (P = . 06), while no significant correlations were found with serum IgM levels, duration of disease, HCV genotype, NS5a gene mutations, liver histology, HLA-DR phenotype, or WA cross-idiotype. Complete responses were also associated, on univariate statistical analysis, with low pretherapy HCV viremia. Responses were accompanied by decrease of viremia, of anti-HCV antibody levels and cryocrit. The usefulness of a high dose regimen is underscored by the higher rates of sustained responses of cryoglobulinemia and transaminase levels compared with previous studies.

The association between an acquired form of hyper-IgM syndrome and a chronic hepatitis C virus (HCV) infection in a 71-year-old female patient is described. Both diseases were diagnosed at the age of 58 years. She was started on intramuscular and then intravenous immunoglobulin replacement therapy. HCV RNA was detected in 1992. The patient remained in well-balanced clinical condition until 1994, when total and specific anti-HCV IgM levels increased and the patient developed an IgM kappa monoclonal gammopathy. Adherent cells and B cells were HCV RNA positive, while T cells were HCV RNA negative. Anti-IgM reactivity was specifically directed to the core antigen of the HCV. The patient we describe showed a picture of a late-onset form of hypogammaglobulinemia with a progressive increase in IgM antibodies, possibly due to the concomitant HCV infection. It is possible that the immunodeficiency might also result from the HCV infection, with formation of specific antibodies belonging to the IgM class, and that the worsening of the clinical condition may be directly related to the persistent viral infection.

Membranoproliferative glomerulonephritis (MPGN) is associated with hepatitis C virus infection predominantly in patients with mixed cryoglobulinemia. Viral-like particles reported in cryoglobulins and in glomerular deposits may be artifacts; in situ identification of viral genome or antigens is required to establish validity of such observations. Although the precise role for hepatitis C virus in the pathogenesis of MPGN remains to be determined, recent evidence suggests that chronic infection with hepatitis C virus may stimulate the production of the monoclonal rheumatoid factor in type II cryoglobulins that are deposited in the glomerular lesions. Interferon-alpha now appears to be the drug of choice in treating MPGN associated with hepatitis C virus infection. The association of hepatitis B virus infection with MPGN has not been convincingly established nor has its role in the pathogenesis of MPGN been demonstrated.