Staging chronic hepatitis C in seven categories using fibrosis biomarker (FibroTest™) and transient elastography (FibroScan®).

BACKGROUND & AIMS: FibroTest™ (FT) and Transient Elastography (TE) have been validated as non-invasive markers of METAVIR fibrosis stages from F0 to F4 using biopsy, and as prognostic markers of liver related mortality in patients with chronic hepatitis C. The aim was to extend the validation of FT and TE as markers of critical steps defined by occurrence of cirrhosis without complications (F4.1), esophageal varices (F4.2), and severe complications (F4.3): primary liver cancer, variceal bleeding, or decompensation (ascites, encephalopathy, or jaundice). METHODS: The updated individual data of 3927 patients (1046 cirrhotics) without complications at baseline were pooled from three prospective cohorts called "EPIC", "Paris", and "Bordeaux" cohorts. RESULTS: At 5 years, among 501 patients without varices at baseline (F4.1) varices occurred in 19 patients [F4.2 incidence of 4.0% (95% CI 2.2-5.8)]. The predictive performance (AUROC) of FT was 0.77 (0.66-0.84; p<0.001). At 10 years severe complications occurred in 203 patients, [F4.3 incidence of 13.4% (9.6-17.1)], including primary liver cancer in 84 patients [6.4% (3.5-9.3)]. FT was predictive (Cox adjusted on treatment) of severe complications [AUROC 0.79 (76-82); p<0.0001], including primary liver cancer [AUROC 0.84 (80-87); p<0.0001]. Similarly TE was predictive of severe complications [AUROC 0.77 (72-81); p<0.0001], including primary liver cancer [AUROC 0.86 (81-90); p<0.0001]. CONCLUSIONS: FibroTest™ and TE increase were associated with the occurrence of all severe complications including hepatocellular carcinoma, hepatic insufficiency, and variceal bleeding. FibroTest™ increase was also associated with the occurrence of esophageal varices.

Chronic Hepatitis C treatment for genotype 2 or 3 in Brazil: cost effectiveness analysis of peginterferon plus ribavirin as first choice treatment

Brazilian Guidelines to HCV treatment (2007) recommended that the first choice treatment for patients with chronic hepatitis C (CHC) and genotype 2 or 3 is interferon alpha (IFN) plus ribavirin (RBV) for 24 weeks. The aim of this study is compare the cost and effectiveness to Hepatitis C treatment in patients with genotype 2 or 3 of peginterferon alpha (PEG) as the first choice of treatment within PEG for those that do not respond to IFN. The target population is CHC patients with genotype 2 or 3 in Brazil. The interventions are: PEG-SEC (first IFN plus RBV for 24 weeks, after, for non-responders and relapsers subsequently PEG plus RBV for 48 weeks); PEG-FIRST24 (PEG+RBV for 24 weeks). The type of the study is cost-effectiveness analysis. The data sources are: Effectiveness data from meta-analysis conducted on the Brazilian population. Treatment cost from Brazilian micro costing study is converted into USD (2010). The perspective is the Public Health System. The outcome measurements are Sustained Viral Response (SVR) and costs. PEG-FIRST24 (SVR: 87.8%, costs: USD 8,338.27) was more effective and more costly than PEG-SEC (SVR: 79.2%, costs: USD 5,852.99). The sensitivity analyses are: When SVR rates with IFN was less than 30% PEG-FIRST is dominant. On the other hand, when SVR with IFN was more then 75% PEG-SEC is dominant (SVR=88.2% and costs USD $ 3,753.00). PEG-SEC is also dominant when SVR to PEG24 weeks was less than 54%. In the Brazilian context, PEG-FIRST is more effective and more expensive than PEG-SEC. PEG-SEC could be dominant when rates of IFN therapy are higher than 75% or rates of PEG24 therapy are lower than 54%.

O protocolo brasileiro de tratamento da Hepatite C (2007) recomendava como primeira escolha para pacientes com hepatite C crônica e portadores de genótipo 2 ou 3 o tratamento com interferona alfa (IFN) associada à ribavirina (RBV), por 24 semanas. O objetivo deste estudo é comparar o custo e a efetividade para pacientes com hepatite C crônica e portadores do genótipo 2 ou 3 o uso de peguinterferon (PEG) como primeiro escolha com o PEG como secunda escolha para aqueles que não responderam ao tratamento com IFN. A população alvo compreende pacientes com hepatite C crônica portadores de genótipo 2 ou 3 no Brasil. As intervenções são: PEG-SEC (IFN + RBV por 24 semanas, para os não respondedores e recidivantes tratamento subsequente com PEG + RBV por 48 semanas; PEG-FIRST24 (PEG + RBV por 24 semanas). O tipo de estudo envolvido é Análise de Custo Efetividade. Os dados de efetividade são provenientes de um metanálise de estudos brasileiros e os dados de custo do tratamento de um estudo de custo do contexto brasileiro. A perspectiva é o Sistema Público de Saúde. Os desfechos avaliados foram Resposta Viral Sustentada (RVS) e Custos. PEG-FIRST24 (RVS: 87,8%, costs: USD 8.338,27) foi mais efetivo e apresentou maior custo que PEG-SEC (RVS: 79,2%, custo USD 5.852,99). A análise de sensibilidade demonstrou que PEG-SEC é dominado por PEG-FIRST24 quando RVS com IFN for menor que 30%. Por outro lado, quando RVS com IFN for maior que 75% PEG-SEC é dominante (RVS=88.2% e custo USD $ 3.753,00). PEG-SEC é também dominante quando RVS para PEG24 for menor que 54%. Conclusão: No contexto brasileiro, PEG-FIRST é mais efetivo e mais custoso que PEG-SEC. PEG-SEC poderia ser dominante quando as taxas de RVS do tratamento com IFN forem superiores a 75% ou as taxas de PEG24 forem inferiores a 54%.

[Chronic liver diseases--new therapy. Are biopsies necessary?]. / Chronische Lebererkrankungen--neue Therapie. Benötigen wir die Biopsie?

Chronic liver disease can often reliably be assessed only by examination of biopsy material. In this article the possible indications for liver biopsy in viral hepatitis B and C, autoimmune liver disease, steatohepatitis and hereditary metabolic diseases are described. A biopsy may be useful in cases with unclear clinical or serological situations or with questionable chronicity and comorbidities. The assessment of biopsy material should be based on guideline-based classification systems. The value of biopsy diagnosis benefits from a close interdisciplinary clinical pathological cooperation.

Poly-ε -caprolactone microspheres containing interferon alpha as alternative formulations for the treatment of chronic hepatitis C

Interferon-alpha (IFN-alpha) is one of the main drugs used in the treatment of hepatitis C. Use of IFN-alpha has some limitations that result in poor treatment efficacy and low patient compliance. Therefore, the aim of this study was to develop poly-ε-caprolactone (PCL) microspheres containing IFN-alpha as an alternative for the treatment of chronic hepatitis C. Microspheres were prepared using the multiple emulsion followed by solvent evaporation technique. Particle size, surface morphology, drug content and encapsulation efficiency of the microspheres produced were evaluated. The stability of the formulation was assessed after 90 days at -20ºC. An in vitro release study was performed in PBS. In vitro cytotoxicity of the formulation was studied using hepatic cell line. The freeze-dried microspheres had mean particle size, IFN-alpha content, and encapsulation efficiency of 38.52 ± 4.64 µm, 15.52 ± 3.28% and 83.93 ± 5.76%, respectively. There were no significant changes during storage and the structural integrity of the protein was not compromised by the preparation technique. A total of 82% of the IFN-alpha was released after 28 days and the developed microspheres did not present cytotoxicity to the hepatic cell line. In vivo studies are currently underway to evaluate the biological activity of IFN-alpha encapsulated into microspheres.

O interferon alfa (IFN-alfa) é um dos principais fármacos utilizados no tratamento de hepatite C, mas o seu uso apresenta limitações que resultam em baixa eficácia do tratamento e não adesão do paciente. Diante disso, este estudo objetiva o desenvolvimento de microesferas de poli-ε-caprolactona (PCL) contendo IFN-alfa como alternativa ao tratamento de hepatite C crônica. As microesferas foram preparadas pelo método de emulsão múltipla seguido de evaporação do solvente e caracterizadas quanto ao diâmetro médio das partículas, morfologia da superfície, taxa e eficiência de encapsulamento. A estabilidade da formulação foi acompanhada durante 90 dias a -20 ºC. O estudo de liberação in vitro foi realizado em PBS. A citotoxicidade da formulação foi avaliada utilizando linhagem de células hepáticas. As microesferas liofilizadas apresentaram diâmetro médio, taxa de encapsulamento e eficiência de encapsulamento de 38,52 ± 4,64 µm, 15,52 ± 3,28% e 83,93 ± 5,76%, respectivamente. Não foram observadas alterações significativas durante o armazenamento e a integridade estrutural da proteína foi mantida após o preparo. Oitenta e dois por cento de IFN-alfa foram liberados em 28 dias e a formulação desenvolvida não apresentou toxicidade para as células testadas. Estudos in vivo estão em andamento para avaliar a atividade biológica do IFN-alfa encapsulado nas microesferas.

Analysis of the PKR-eIF2alpha phosphorylation homology domain (PePHD) of hepatitis C virus genotype 1 in HIV-coinfected patients by ultra-deep pyrosequencing and its relationship to responses to pegylated interferon-ribavirin treatment.

Chronic coinfection with hepatitis C virus (HCV) and human immunodeficiency virus (HIV) is among the greatest challenges facing public health worldwide. In this population, the response to hepatitis C therapy by treatment with pegylated interferon plus ribavirin (PEG-IFN+RBV) is lower than in HCV-monoinfected patients, particularly in those infected by HCV genotype 1. A PKR/eIF-2α phosphorylation homology domain (PePHD) within the E2 protein has been found to interact with PKR and inhibit PKR in vitro, suggesting a possible mechanism for HCV to evade the antiviral effects of IFN. The aim of this work was to analyze the amino acid conservation in the HCV-E2-PePHD and quasispecies diversity among HCV-HIV-coinfected patients exhibiting sustained virological response, non-response, or partial response with viral relapse to PEG-IFN+RBV by ultra-deep pyrosequencing. For this purpose, HCV-E2-PePHD PCR products were generated and sequenced directly for four patients with a sustained response, seven patients with no virological response, and four patients with viral relapse before and after treatment with PEG-IFN+RBV. HCV-E2-PePHD amino acid sequences were obtained for isolates from serum collected before and during treatment (24 h, 4 weeks, and 12 weeks). Quasispecies analysis of the HCV-E2-PePHD and flanking genomic regions was performed using 454/Roche pyrosequencing, analyzing 39,364 sequence reads in total. The HCV-E2-PePHD sequence at the amino acid and nucleotide level was highly conserved among HCV genotype 1 strains, irrespective of the PEG-IFN+RBV response. This high degree of amino acid conservation and sporadic mutations in the HCV-E2-PePHD domain do not appear to be associated with treatment outcome. The HCV-E2-PePHD sequence before or during treatment cannot be used to predict reliably the outcome of treatment in patients coinfected with HCV genotype 1 and HIV.

Comparison of eight diagnostic algorithms for liver fibrosis in hepatitis C: new algorithms are more precise and entirely noninvasive.

UNLABELLED: The sequential algorithm for fibrosis evaluation (SAFE) and the Bordeaux algorithm (BA), which cross-check FibroTest with the aspartate aminotransferase-to-platelet ratio index (APRI) or FibroScan, are very accurate but provide only a binary diagnosis of significant fibrosis (SAFE or BA for Metavir F ≥ 2) or cirrhosis (SAFE or BA for F4). Therefore, in clinical practice, physicians have to apply the algorithm for F ≥ 2, and then, when needed, the algorithm for F4 ("successive algorithms"). We aimed to evaluate successive SAFE, successive BA, and a new, noninvasive, detailed classification of fibrosis. The study included 1785 patients with chronic hepatitis C, liver biopsy, blood fibrosis tests, and FibroScan (the latter in 729 patients). The most accurate synchronous combination of FibroScan with a blood test (FibroMeter) provided a new detailed (six classes) classification (FM+FS). Successive SAFE had a significantly (P < 10(-3) ) lower diagnostic accuracy (87.3%) than individual SAFE for F ≥ 2 (94.6%) or SAFE for F4 (89.5%), and required significantly more biopsies (70.8% versus 64.0% or 6.4%, respectively, P < 10(-3) ). Similarly, successive BA had significantly (P ≤ 10(-3) ) lower diagnostic accuracy (84.7%) than individual BA for F ≥ 2 (88.3%) or BA for F4 (94.2%), and required significantly more biopsies (49.8% versus 34.6% or 24.6%, respectively, P < 10(-3) ). The diagnostic accuracy of the FM+FS classification (86.7%) was not significantly different from those of successive SAFE or BA. However, this new classification required no biopsy. CONCLUSION: SAFE and BA for significant fibrosis or cirrhosis are very accurate. However, their successive use induces a significant decrease in diagnostic accuracy and a significant increase in required liver biopsy. A new fibrosis classification that synchronously combines two fibrosis tests was as accurate as successive SAFE or BA, while providing an entirely noninvasive (0% liver biopsy) and more precise (six versus two or three fibrosis classes) fibrosis diagnosis.

Comparison of accuracy of fibrosis degree classifications by liver biopsy and non-invasive tests in chronic hepatitis C.

BACKGROUND: Non-invasive tests have been constructed and evaluated mainly for binary diagnoses such as significant fibrosis. Recently, detailed fibrosis classifications for several non-invasive tests have been developed, but their accuracy has not been thoroughly evaluated in comparison to liver biopsy, especially in clinical practice and for Fibroscan. Therefore, the main aim of the present study was to evaluate the accuracy of detailed fibrosis classifications available for non-invasive tests and liver biopsy. The secondary aim was to validate these accuracies in independent populations. METHODS: Four HCV populations provided 2,068 patients with liver biopsy, four different pathologist skill-levels and non-invasive tests. Results were expressed as percentages of correctly classified patients. RESULTS: In population #1 including 205 patients and comparing liver biopsy (reference: consensus reading by two experts) and blood tests, Metavir fibrosis (FM) stage accuracy was 64.4% in local pathologists vs. 82.2% (p < 10-3) in single expert pathologist. Significant discrepancy (≥ 2FM vs reference histological result) rates were: Fibrotest: 17.2%, FibroMeter2G: 5.6%, local pathologists: 4.9%, FibroMeter3G: 0.5%, expert pathologist: 0% (p < 10-3). In population #2 including 1,056 patients and comparing blood tests, the discrepancy scores, taking into account the error magnitude, of detailed fibrosis classification were significantly different between FibroMeter2G (0.30 ± 0.55) and FibroMeter3G (0.14 ± 0.37, p < 10-3) or Fibrotest (0.84 ± 0.80, p < 10-3). In population #3 (and #4) including 458 (359) patients and comparing blood tests and Fibroscan, accuracies of detailed fibrosis classification were, respectively: Fibrotest: 42.5% (33.5%), Fibroscan: 64.9% (50.7%), FibroMeter2G: 68.7% (68.2%), FibroMeter3G: 77.1% (83.4%), p < 10-3 (p < 10-3). Significant discrepancy (≥ 2 FM) rates were, respectively: Fibrotest: 21.3% (22.2%), Fibroscan: 12.9% (12.3%), FibroMeter2G: 5.7% (6.0%), FibroMeter3G: 0.9% (0.9%), p < 10-3 (p < 10-3). CONCLUSIONS: The accuracy in detailed fibrosis classification of the best-performing blood test outperforms liver biopsy read by a local pathologist, i.e., in clinical practice; however, the classification precision is apparently lesser. This detailed classification accuracy is much lower than that of significant fibrosis with Fibroscan and even Fibrotest but higher with FibroMeter3G. FibroMeter classification accuracy was significantly higher than those of other non-invasive tests. Finally, for hepatitis C evaluation in clinical practice, fibrosis degree can be evaluated using an accurate blood test.

Editorial: staging liver fibrosis in hepatitis C: a challenge for this decade.

The limitations of and the invasive nature of liver biopsy has spurred extensive interest in the development of non-invasive tests to measure liver fibrosis in patients with chronic hepatitis C. Clinically applicable non-invasive tests, including radiological studies, elastography, and serum markers, all of which perform extremely well in excluding significant disease and diagnosing cirrhosis. FibroScan and acoustic radiation force impulse elastography are two elastography-based tests that show promise. In this new era of increased cure rates with newly Food and Drug Administration-approved drugs and the availability of multiple non-invasive tests of liver fibrosis, we anticipate a decreasing need for liver biopsies in the management of chronic hepatitis C.

Comparison of transient elastography and acoustic radiation force impulse for non-invasive staging of liver fibrosis in patients with chronic hepatitis C.

OBJECTIVES: Transient elastography (TE) is adequate for a diagnosis of cirrhosis, but its accuracy for milder stages of fibrosis is much less satisfactory. The objective of this study was to compare the performance and the discordance rate of acoustic radiation force impulse (ARFI) and TE with liver biopsy in a cohort of chronic hepatitis C (CHC) patients. METHODS: One hundred thirty-nine consecutive patients with CHC were enrolled in two tertiary centers, and evaluated for histological (Metavir score) and biochemical features. All patients underwent TE and ARFI. RESULTS: TE was unreliable in nine patients (6.5%), while in no cases (0%) were ARFI invalid measurements recorded (P=0.029). By area under receiver operating characteristic curve (AUROC), the best cutoff values for TE and ARFI for significant fibrosis (≥F2) were ≥6.5 kPa (AUROC: 0.78) and ≥1.3 m/s (AUROC: 0.86), respectively. For severe fibrosis (F3-F4), these cutoff values were 8.8 kPa (AUROC: 0.83) for TE and 1.7 m/s (AUROC: 0.94) for ARFI. For cirrhosis, TE had its best cutoff at ≥11 kPa (AUROC: 0.80) and ARFI at ≥2.0 m/s (AUROC: 0.89). By pairwise comparison of AUROC, ARFI was significantly more accurate than TE for a diagnosis of significant and severe fibrosis (P=0.024 and P=0.002, respectively), while this difference was only marginal for cirrhosis (P=0.09). By partial AUROC analysis, ARFI performance results significantly higher for all three stages of fibrosis. The average concordance rates of TE and ARFI vs. liver biopsy were 45.4 and 54.7%, respectively. By multivariate analysis, ARFI was not associated with alanine aminotransferase (ALT), body mass index, Metavir grade, and liver steatosis, while TE was significantly correlated with the ALT value (P=0.027). CONCLUSIONS: In a cohort of patients with CHC, ARFI imaging was more accurate than TE for the non-invasive staging of both significant and severe classes of liver fibrosis.

Evaluation of chronic HCV infection in transplanted livers using a modified histological activity index.

BACKGROUND: The majority of histopathological classifications of primary chronic viral hepatitis and recurrence of HCV infection in liver transplants is based on the histological activity index (HAI) introduced by Knodell et al in 1981; however, correlation between HAI and clinical/laboratory data is poor. Therefore, the aim of this study was to present a modification of HAI (mHAI) adapted to distinct features of graft infection, and to evaluate its usefulness in the description of disease activity. MATERIAL/METHODS: Inflammatory activity in 67 biopsies of HCV-infected grafted livers was semi-quantitatively assessed according to HAI based on Knodell's criteria and to mHAI proposed by the authors. Patients were divided into 4 groups according to level of clinical aggressiveness of HCV reinfection on the basis of laboratory data. Correlations between clinical aggressiveness and histological activity of the disease expressed as HAI or mHAI was estimated. RESULTS: Histological features of HCV reinfection of various activity were observed as early as in the second month after orthotopic liver transplantation. HAI and mHAI values were similar in 55.2% of cases, but in 38.8% HAI was lower than mHAI. Morphological and clinical features were found to be consistent in 32.8% and 49.3% of cases for HAI and mHAI evaluation, respectively. mHAI seems to correlate with clinical assessment of HCV recurrence in liver grafts significantly better than does the classical HAI. CONCLUSIONS: mHAI proposed in the present study appears to be more useful for evaluation of recurrence of HCV infection in post-transplant liver biopsies.

Noninvasive assessment of liver fibrosis with acoustic radiation force impulse imaging: increased liver and splenic stiffness in patients with liver fibrosis and cirrhosis.

PURPOSE: To evaluate acoustic radiation force impulse imaging (ARFI) of the liver and spleen as a new method for the noninvasive assessment of liver fibrosis (LF). MATERIALS AND METHODS: Three groups of 58 examinees were studied: (A) 20 healthy volunteers; (B) 18 patients with chronic viral hepatitis (CVH) B or C having liver fibrosis stages F 1 - 4 (assessed by liver biopsy; Ishak classification); and (C) 20 patients with liver cirrhosis (LC). All participants were examined using the Siemens ACUSON S 2000 Ultrasound Virtual Touch Tissue Quantification system. Ten measurements were performed on both liver lobes and three measurements on the spleen, and the obtained mean values (shear wave velocities [SWV] expressed in m/s) were compared between the groups. In 20 patients the splenic artery pulsatility index (SAPI) was also measured and correlated to the liver and splenic ARFI and histological stage of LF. RESULTS: Hepatic ARFI measurements demonstrated a significant correlation to LB results (Spearman's ρ = 0.766; ρ < 0.001) and SWV cut-off values of 1.3 (AUC 0.96) and 1.86 (AUC 0.99) could reliably differentiate between healthy (A) and non-cirrhotic CVH (B), as well as between non-cirrhotic CVH (B) and LC (C). Splenic SWV cut-off value of 2.73 (AUC 0.82) could differentiate between the patients with LC and non-cirrhotic CVH. A significant correlation was also observed between the SAPI and liver ARFI results (ρ = 0.56; p = 0.013). CONCLUSION: The hepatic and splenic SWV measured by ARFI increase with the LF stage, and the hepatic SWV correlate well with SAPI. This new technology enables simultaneous morphological, Doppler and elastometric examinations and might improve the accuracy of noninvasive liver fibrosis assessment.

Evaluation of ocular surface damage and dry eye status in chronic hepatitis C at different stages of hepatic fibrosis.

PURPOSE: The purpose of this study was to explore changes in ocular surface and tear function parameters in chronic hepatitis C at different stages of hepatic fibrosis. METHODS: Fifty-four patients with biopsy-proven chronic hepatitis C and 54 age- and sex-matched healthy control subjects without systemic hepatitis C infection were examined with the Ocular Surface Disease Index questionnaire, Schirmer with and without anesthesia, tear film breakup time, and scoring of ocular surface Lissamine green staining using modified Oxford and van Bijsterveld scoring systems and corneal fluorescein staining. RESULTS: Patients with chronic hepatitis C scored significantly worse than the control subjects on all parameters: modified Oxford scores of Lissamine green staining (5.5/3.0; P <0.001), Oxford and van Bijsterveld scores (4.0/2.0; P <0.001), and corneal fluorescein staining (1.5/0.0; P = 0.001). The chronic hepatitis C group also had higher Ocular Surface Disease Index scores than the control subjects (22.3/13.7; P = 0.001). Schirmer with and without anesthesia and tear film breakup time scores were found to be lower in patients with chronic hepatitis C (P <0.001). Moreover, patients with advanced stages of hepatic fibrosis (stages 4-6) had significantly lower values of tear film breakup time and worse Ocular Surface Disease Index scores and ocular surface vital dye staining than those with initial stages of hepatic fibrosis (stages 0-3). CONCLUSION: Patients with chronic hepatitis C, especially those with advanced stages of hepatic fibrosis, were more likely to exhibit severe ocular surface damage and signs of dry eye.

Effective use of FibroTest to generate decision trees in hepatitis C.

AIM: To assess the usefulness of FibroTest to forecast scores by constructing decision trees in patients with chronic hepatitis C. METHODS: We used the C4.5 classification algorithm to construct decision trees with data from 261 patients with chronic hepatitis C without a liver biopsy. The FibroTest attributes of age, gender, bilirubin, apolipoprotein, haptoglobin, alpha2 macroglobulin, and gamma-glutamyl transpeptidase were used as predictors, and the FibroTest score as the target. For testing, a 10-fold cross validation was used. RESULTS: The overall classification error was 14.9% (accuracy 85.1%). FibroTest's cases with true scores of F0 and F4 were classified with very high accuracy (18/20 for F0, 9/9 for F0-1 and 92/96 for F4) and the largest confusion centered on F3. The algorithm produced a set of compound rules out of the ten classification trees and was used to classify the 261 patients. The rules for the classification of patients in F0 and F4 were effective in more than 75% of the cases in which they were tested. CONCLUSION: The recognition of clinical subgroups should help to enhance our ability to assess differences in fibrosis scores in clinical studies and improve our understanding of fibrosis progression.

[Current problems of hepatitis]. / Aktuelle Probleme der Hepatitis.

New findings have been made in recent years on the various forms of the hepatitis virus in terms of disease course, its etiopathogenetic link with comorbidities and the definition of new forms in Central Europe. Epstein-Barr virus (EBV)- and cytomegalovirus (CMV)-induced hepatitis may occur in the so-called sero-negative group of hepatitis and direct demonstration of the viral genome in paraffin liver tissues is required to confirm the diagnosis. Since diagnosis of autoimmune hepatitis in daily practice may be difficult, a scoring system with simplified criteria has recently been developed.

State of the iron metabolism in patients with chronic hepatitis C type C does not influence antiviral treatment with interferon and ribavirin.

BACKGROUND/AIMS: Comparison of the iron status in patients who responded and did not respond to combination treatment with interferon alpha and ribavirin in chronic hepatitis C. METHODOLOGY: The study group comprised of 61 patients with chronic hepatitis C (genotype 1) treated with alpha 2b interferon and ribavirin. The iron metabolism was evaluated based on serum iron level, total iron binding capacity, transferrin saturation, serum ferritin concentration and hepatic iron concentration. In the evaluation of antiviral treatment efficacy biochemical and virological responses were taken into account. RESULTS: End of treatment response was observed in 38 patients (62%). Significant differences in iron parameters were not observed between responders and non-responders. Also, sustained viral response, 6 months after treatment completion, was reached in 32 patients (52.5%). Iron metabolism parameters did not differ significantly in the group of sustained responders versus non- responders. Finally, ALT normalization was observed in 42 patients (68.9%). Again, no significant differences in iron status were observed between patients with and without biochemical response excluding significantly higher serum ferritin concentration in non-responders. CONCLUSIONS: Results of this study show that iron status does not significantly influence the efficacy of treatment with interferon and ribavirin in patients with chronic hepatitis C.

Development of JFH1-based cell culture systems for hepatitis C virus genotype 4a and evidence for cross-genotype neutralization.

Efficient in vitro systems to study the life cycle of hepatitis C virus (HCV) were recently developed for JFH1 (genotype 2a), which has unique replication capacity in Huh7 cells. We developed 4a/JFH1 intergenotypic recombinants containing the structural genes (Core, E1, and E2), p7, and all or part of NS2 of the 4a prototype strain ED43 that, after transfection of Huh7.5 cells with RNA transcripts, produced infectious viruses. Compared with the J6/JFH control virus, production of viruses was delayed. However, efficient spread of infection and high HCV RNA and infectivity titers were obtained in serial passages. Sequence analysis of recovered viruses and subsequent reverse genetic studies revealed a vital dependence on one or two NS2 mutations, depending on the 4a/2a junction. Infectivity of ED43/JFH1 viruses was CD81 dependent. The genotype 4 cell culture systems permit functional analyses as well as drug and vaccine research on an increasingly important genotype in the Middle East, Africa, and Europe. We also developed genotype 1a intergenotypic recombinants from H77C with vital mutations in NS3. Using H77C/JFH1 and ED43/JFH1 viruses, we demonstrated high homologous neutralizing antibody titers in 1a and 4a patient sera, respectively. Furthermore, availability of JFH1 viruses with envelope proteins of the six major HCV genotypes permitted cross-neutralization studies; 1a and 4a serum cross-neutralized 1a, 4a, 5a, and 6a but not 2a and 3a viruses. Thus, the JFH1 intergenotypic recombinants will be of importance for future studies of HCV neutralization and accelerate the development of passive and active immunoprophylaxis.

Liver biopsy assessment in chronic viral hepatitis: a personal, practical approach.

The terminology for assessment of chronic viral hepatitis in liver biopsy specimens has become confusing with the proliferation of grading and staging schemes that have paralleled the rise of the hepatitis C epidemic and the importance of mixed viral infections. This review represents a personal approach to the interpretation of these biopsy specimens, aiming at clarifying and simplifying the important points for the general pathologist confronted by these diagnostic dilemmas. The most commonly used schemes-Ishak modification of the Knodell 'hepatic activity index', Scheuer, Metavir, Batts-Ludwig classifications-are presented with evaluation of their pros and cons. Which scheme is selected is less important than the consistent use of a single scheme and the clear naming of that scheme in pathology reports. The importance and clinical implications of identifying severe necroinflammatory activity in the form of 'confluent necrosis' is discussed. Pathologists must also be clear about assessing concomitant diseases, in particular, alcoholic or non-alcoholic fatty liver disease, and be aware that grading/staging schemes for chronic hepatitis do not apply to mixed disease conditions. Other important features to be evaluated in all chronic hepatitis biopsy specimens include iron (which may represent hereditary hemochromatosis or secondary uptake) and neoplasia-associated changes, namely large cell change and small cell change; these findings and their clinical import are updated and reviewed. Sample approaches to composing useful diagnostic reports are also presented.

Correlation of amino acid variations within nonstructural 4B protein with initial viral kinetics during interferon-alpha-based therapy in HCV-1b-infected patients.

Chronic hepatitis C is a major cause of liver cirrhosis leading to chronic liver failure and hepatocellular carcinoma. Different hepatitis C virus (HCV) proteins have been associated with resistance to interferon-alpha-based therapy. However, the exact mechanisms of virus-mediated interferon resistance are not completely understood. The importance of amino acid (aa) variations within the HCV nonstructural (NS)4B protein for replication efficiency and viral decline during the therapy is unknown. We investigated pretreatment sera from 42 patients with known outcome to interferon-based therapy. The complete NS4B gene was amplified and sequenced. Mutational analyses of predicted conformational, functional, structural and phylogenetic properties of the deduced aa sequences were performed. The complete NS4B protein was highly conserved with a median frequency of 0.015 +/- 0.009 aa exchanges (median +/- SD, 4.00 +/- 2.31). Especially within the predicted transmembranous domains of the NS4B protein, the mean number of aa variations was low (median frequency, 0.013 +/- 0.013). Neither the number of aa variations nor specific aa exchanges were correlated with HCV RNA serum concentration at baseline. A rapid initial HCV RNA decline of >/=1.5 log(10) IU/mL at week 2 of interferon-based therapy was associated with a higher frequency of nonconservative aa exchanges within the complete NS4B protein in comparison with patients with a nonrapid HCV RNA decline (median frequency, 0.011 +/- 0.005 vs 0.004 +/- 0.003, P = 0.006). Overall, the aa sequence of the NS4B protein was highly conserved, indicating an important role for replication in vivo. Amino acid variations with relevant changes of physicochemical properties may influence replication efficiency, associated with a rapid early virological response.

Liver gene expression signature of mild fibrosis in patients with chronic hepatitis C.

BACKGROUND & AIMS: The molecular mechanisms of hepatocellular carcinoma have been studied, but little is known of the changes in liver gene expression during the different stages of chronic hepatitis C virus (HCV) infection, in particular the transition from mild to moderate fibrosis. METHODS: We used real-time quantitative RT PCR to study the messenger RNA expression of 240 selected genes in 2 pools of liver specimens according to the stages of fibrosis (Metavir score; mild fibrosis = F1 and septal fibrosis = F2). Genes whose expression differed between pools (F2 vs F1) by at least 2-fold were selected. In addition, the expression level of these selected genes then was assessed in each of the 62 individual samples (F4, n = 6; F3, n = 17; F2, n = 21; vs F1, n = 18). RESULTS: The 22 genes that were up-regulated in the 21 F2 samples relative to the 18 F1 samples mainly encoded genes involved in cytoskeleton (KRT 19 and SCG 10), growth factors/cytokines (CXCL6, interleukin 8 [IL8], IL1A, IL2, and CXCL10), or growth factor receptors (CCR2, CXCR3, and CXCR4), or were involved in extracellular matrix production (COL1A1, CHI3L, and SPP1), in extracellular matrix remodeling (TIMP1, MMP7, and MMP9), and in cell junction (ITGA2 and CLDN 4). When hierarchically clustering the F2 and F1 samples according to the expression of the 11 most discriminatory genes (KRT 19, COL1A1, STMN2, CXCL6, CCR2, TIMP1, IL8, IL1A, ITGA2, CLDN 4, and IL2), the patient population was categorized into 2 subgroups: F1 and F2. Specifically, 15 of 18 F1 (83%) and 19 of 21 F2 (90%) were classified correctly (P < 10(-5)). We also studied the messenger RNA expression of these 240 selected genes in normal liver in comparison with F1. Genes dysregulated in the transition from normal liver to F1 mainly were interferon-inducible genes, and therefore were very different from those dysregulated in the transition from F1 to F2. CONCLUSIONS: Genes involved in extracellular matrix turnover and immune response are implicated in the transition from mild to moderate fibrosis. Eleven of the genes could form the basis for the gene expression signature of mild versus moderate fibrosis in patients with chronic hepatitis C.

Hepatitis crónica por virus C: factores asociados a la severidad del daño histológico / Factors associated with the severity of liver damage in chronic hepatitis C

Background: Twenty percent of patients with chronic hepatitis C evolve to cirrhosis in 10 to 20 years. The degree of steatosis and hepatic iron stores in liver biopsy increase the risk. Age, high body mass index, diabetes mellitus and alcohol consumption are factors associated to the severity of liver damage. Aim: To study the association of steatosis and increased iron stores in the liver biopsy and age, overweight, alcohol consumption and diabetes with the severity of liver damage in patients with hepatitis C virus infection. Patients and methods: Retrospective study of 84 liver biopsies of patients with chronic infection with hepatitis C virus were studied. The pathological appearance was classified as stage I when chronic hepatitis with mild activity without fibrosis was observed; as stage II when moderate chronic hepatitis with mild fibrosis was observed and as stage III when there was a moderate chronic hepatitis with fibrosis or cirrhosis. The amount of steatosis and iron deposition in the biopsy were also assessed. Results: Forty one percent of patients were in stage I, 32% in stage II and 27% in stage III. Patients in stage I were younger than those in stages II and III (40.7 and 52.2 years respectively, p <0,001). No association between the severity of liver damage and the degree of steatosis, hemosiderosis, body mass index or alcohol intake, was observed. The frecuency of diabetes mellitus increased along with pathological staging (3, 15 and 30% in stages I, II and III, respectively, p <0,05). Conclusions: This study confirms that severity of chronic hepatitis C is associated with age and the presence of diabetes mellitus.