| Abstract |
Hepatitis C virus (HCV) is the cause of a chronic hepatitis that may frequently evolve into cirrhosis or hepatocellural carcinoma. In recent years many attempts of treating hepatitis C have been made with interferon, either alone or in combination with various drugs. Treatment response however is not uniform and factors determining a favourable outcome are still under investigation. The aim therefore of the present investigation was: A. To perform an epidemiological study of HCV genotypes in patients with chronic hepatitis C in Crete. Associations among viral genotypes in patients with chronic hepatitis C in Crete. Associations among viral genotypes and disease severity, histological classification, demographic characteristics of patients and viral load were also examined. B. To assess response of patients to various therapeutic regimes. Short and long term biochemical and virological response was assessed and an effort to identity factors influencing treatment response was done through univariate and multivariate analysis. C. Finally mutation in the NS-5A region of HCV genome were examined as possible factors determining the response to interferon treatment in patients infected with the 1b genotype of HCV. Results A. Genotype epidemiology: A1. By a home made nested RT-PCR the presence of HCV RNA was verified in 102 patients with liver biopsy proven chronic hepatitis. A2. Genotypic classification according to Simmonds was performed using reversed immunoblotting (INNO-Lipa, INNOFENETICS, Belgium). A3. Viral load was measured using the Amplicor Monitor System (Roche Diagnostics). A4. The Ishak classification was used for grading and staging of liver biopsies. · Age, sex, serum transaminases, alkaline phosphate, γ-GT, viral load , histological staging and grading were the variables used in univariate and multivariate analysis to look for correlations with genotypes. 64% of patients were infected with genotype 1. Genotype 2 was present in 13%, genotype 3 in 18% and genotype 4 in 5% of patients. Genotypes 5 and 6 were not detected. · Female sex and older age were independent variables related to genotype 1. · Younger age, lower viral loads and high staging were related to genotype 2. · Intravenous drug abuse and higher viral loads were related to genotype 3. · Lower staging and lower viral loads were related to genotype 4. However, only the relation between statistically sign cant mean age of genotype 1 patients was 55,8 years, significantly higher than non - 1 genotypes (p=0,007) B. Treatment response 6 treatment schedules were examined: Group 1 : 30 patients treated for 3 months with 3MU IFN three times a week Group 2 : 35 patients treated for 6 months with 5MU IFN three times a week Group 3 : 60 patients treated for 12 months with 3MU IFN three times a week plus 15mg/kgr ursodeoxycholate (UDCA) Group 4 : 35 patients treated for 6 months with 5MU IFN three times a week plus 500 mg D-Penicillamine. Group 5 : 36 patients treated for 6 months with 5MU IFN three times a week plus 150mg nimesulide Group 6 : 21 patients treated for 12 months with 5MU IFN three times a week plus 1000mg Ribavirin Serum enzymes, viral load and genotyping was performed in all these patients before and after treatment. A liver biopsy before treatment was grated and staged according to Ishak classification. o Short term biochemical response varied between 80% for the D-penicillamine group and 47% for group 1. o Short term virological response varied between 48% for the ribavirin group and 10% for group 1 o Long term biochemical response varied between 33% for the ribavirin group and 14% for groups 1 and 2. o Long term virological response varied between 19% for the ribavirin group and 0% for group 2 Long term biochemical and virological response were generally disappointing. Multivariate analysis showed no relation to treatment response of age, sex, viral load, genotypes or histological grading and staging. C. Sequencing of NS-5A region of 1b genotype: An amplification of the region 6741-7275 of the NS-5A area of 1b genotype was performed by RT-nested PCR in 28 patients infected with this genotype. The PCR product was isolated and purified and then a direct sequencing of nucleotides was done in an automatic sequencer. Results were correlated with response to IFN treatment. Conclusions can be summarized as follows: o Mutated viral clones were very rare in Crete o Wild type clones were different at nucleotide levels form Japanese clones and similar to other reported European clones. o Biochemical response of mutated viruses is favorable and prediction of response can be made from the number of amino acid mutations. By contrast, virological response cannot be predicted.
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Γονιδιακή ανάλυση και ταξινόμηση των ιών ηπατίτιδας C. Κλινικοεργαστηριακές συσχετίσεις
