Effect of iron overload on the severity of liver histologic alterations and on the response to interferon and ribavirin therapy of patients with hepatitis C infection

Brazilian Journal MedicalCand Biological Research (2006) 39: 79-83 Iron overload andofhepatitis ISSN 0100-879X Short Communication 79 Effect of iron overload on the severity of liver histologic alterations and on the response to interferon and ribavirin therapy of patients with hepatitis C infection R.M. Souza1,2, L.A.R. Freitas3, A.C. Lyra1,2, C.F. Moraes2, E.L. Braga1,2 and L.G.C. Lyra1,2 1Serviço de Gastro-Hepatologia, Hospital Prof. Edgard Santos, Universidade Federal da Bahia, Salvador, BA, Brasil 2Serviço de Gastro-Hepatologia, Hospital São Rafael, Salvador, BA, Brasil 3CPqGM, Fundação Oswaldo Cruz, Salvador, BA, Brasil Abstract Correspondence L.G.C. Lyra Av. Juracy Magalhães Jr., 1855/501 41940-060 Salvador, BA Brasil Fax: +55-71-276-2106 or 399-6266 E-mail: or Received September 28, 2004 Accepted September 9, 2005 The objective of the present study was to determine the presence of hepatic iron overload in patients with chronic HCV infection and to correlate it with histologic alterations, HCV genotype and response to therapy. Liver tissue samples from 95 patients with chronic hepatitis C were divided into two groups: group I, presence of iron overload in hepatic tissue (Perls’ staining) and group II, no iron overload. Hepatic iron overload was detected in 30 (31.6%) of 95 patients. Of the 69 patients tested by genotyping, 49 (71.01%) were genotype 1 and 20 (28.99%) genotype non-1. Iron overload was detected in 14 (28.6%) patients with genotype 1 and in 6 (30%) with genotype non-1 (P = 0.906). There was a significant difference in fibrosis stage between groups (P = 0.005). In group I (N = 30), one patient had stage F0/F1 of fibrosis, while in group II (N = 65), 22 (33.8%) patients had minimal or no fibrosis. Fibrosis stage F2/F3 was observed in 70% of group I patients compared to 46.2% of group II. Eighty-five patients were treated with a combination of interferon and ribavirin; 29 of them (34.1%) had a sustained virologic response and 8 (27.6%) of them had hepatic iron overload. Iron overload was detected in 18 (32.1%) of the 56 non-responders (P = 0.73). Hepatic iron overload was frequent among patients with chronic hepatitis C and was associated with a more severe stage of liver fibrosis. There was no association between iron overload and HCV genotype and response to interferon and ribavirin therapy. Elevated iron parameters and mild iron overload are common in the liver of patients with chronic hepatitis C. It has been suggested that ferritin and serum iron might be correlated with liver inflammation and serum markers of fibrogenesis (1,2). Bassett et Key words • Hepatitis C infection • Iron overload • Genotype • Inflammatory activity • Fibrosis • Antiviral therapy al. (3) showed that iron overload was associated with higher ALT levels and more severe histological findings in HCV-infected chimpanzees. Increased iron stores may stimulate hepatic fibrogenesis, by leading to oxygen free radical injury and/or by inducing the Braz J Med Biol Res 39(1) 2006 80 R.M. Souza et al. production of profibrogenic cytokines such as tumor growth factor ß (TGF-ß) (2). Of note, there appears to be a discrepancy between the frequency of altered iron parameters in serum and in liver tissue. Riggio et al. (4) found that 40% of patients had increased iron in serum compared to 10% in tissue. The presence of iron overload has been reported to possibly be involved in fibrosis progression and in the development of hepatocellular carcinoma, although the studies are controversial (5,6). Over the last few years there has also been much interest in the role of iron in the outcome of antiviral therapy in patients with chronic HCV infection (7,8). Several studies have demonstrated that iron overload is associated with lower response rates to interferon-α (IFN-α) monotherapy. Little is known about whether iron overload also has an impact on the response rate to combination therapy with interferon and ribavirin. The aims of the present study were to determine the presence of hepatic iron overload in patients with chronic HCV infection and to correlate it with histologic liver alterations, HCV genotype and the response to combination therapy with interferon and ribavirin. We evaluated patients who were antiHCV positive by commercial second- or third-generation ELISA and HCV-RNA positive by PCR (Amplicor, Roche, Indianapolis, IN, USA). Thirty-two patients had been referred to Hospital São Rafael and 63 to Hospital Universitário Prof. Edgard Santos, both in Salvador, BA, Brazil, during the period from 1997 to 2000. These 95 patients had participated in a previous protocol for the treatment of hepatitis C and fulfilled the following criteria: 18-65 years old, ALT level ≥1.5 times the upper normal limit, compensated liver disease, and negative serum markers for HIV and HBV. All subjects had an appropriate Braz J Med Biol Res 39(1) 2006 amount of liver tissue for analysis and none had been submitted to blood transfusion during the previous 30 days or had other clinical condition that might lead to iron overload. Eighty-five of 95 patients had completed therapy with interferon using one of the following two schedules: induction therapy with 3 MU IFN-α daily during the first 3 months, followed by treatment three times per week until 1 year was completed, or 3 MU IFN-α three times per week. In both schedules ribavirin was given orally in combination at the dose of 1 g/day. Patients were divided into two groups according to the presence (group I) or absence (group II) of iron overload in liver biopsies. Hepatic tissue fragments were fixed in 10% formaldehyde, embedded in paraffin and stained with a) hematoxylin and eosin, b) picrosirius red, c) Gomori’s silver impregnation, d) PAS with or without diastasis, and e) Perls’ staining (Prussian blue reaction). Iron overload was defined as the presence of any amount of iron detected by Perls’ staining. Necro-inflammatory activity and stage of liver fibrosis were determined using the METAVIR system (The French METAVIR Cooperative Study Group, 1994). The Hospital São Rafael Ethics Committee approved the study and the protocol conformed to the ethical guidelines of the 1975 Declaration of Helsinki. Continuous variables are reported as means ± SD. Data were analyzed statistically by the chi-square test or Fisher exact test to determine the differences between proportions, and by the Mann-Whitney test, with the level of significance set at P < 0.05. Twenty-four of the 95 patients were females (25.3%) and 71 were males (74.7%). Mean age was 45.8 ± 8.7 years (range 2762). In most cases the grade of iron overload determined by Perls’ staining was mild (1+) or mild to moderate (2+) according to Searle’s criteria. According to these data, patients 81 Iron overload and hepatitis C were divided into two groups: with or without iron detected in liver tissue. Group I contained 30 patients (31.6%) with iron overload in hepatic tissue and group II con (...truncated)