Progression of Fibrosis in Chronic Hepatitis C Treated with Interferon or Interferon and Ribavirin. Impact of Co-Factors: Alcohol, Steatosis, Iron Serum Markers

Fatima Serejo, Adilia Costa, Jose Velosa, Miguel C. Moura, Maria J. Lacerda

 

Material/ Methods Two hundred-seventeen with chronic hepatitis C were studied (164 treated with interferon 5 MU monotherapy and 53 with IFN 3 MU and ribavirin), 140 males ( 37.3 2.2years), 77 females (41.5 5.8 years), Genotype 1- 71%, Genotype 3a- 23%. BMI 25- 38%, 30%- 7.3%.Liver biopsy was performed before and 1 year after stopping therapy. Fibrosis score was defined according Knodell/Peter Scheuer and Steatosis graded 0- none, 1- mild focal, 2- mild diffuse (10-25% of hepatocytes), 3- moderate (25-50%), 4- severe (more 50%).Fibrosis progression per year (FPPY) defined by Poynard (Lancet 1997;349).

 

Results

Using the first liver biopsy, median FPPY was 0.37( min 0.00, max 4.0), estimating 11 years for the time to cirrhosis. Age at first biopsy more than 40 years and sporadic form were independent predictors of more severe fibrosis. Genotypes and viremia did not influence disease progression. After treatment, median FPPY was -0.000 (min -1.5, max 0.7), similar in mono and combination therapy). Type of response: sustained response (SR)- 90,Relapse (RR)- 44, Non response (NR)- 83. Predictive factors associated with response: younge age (p <0.005), short duration of infection (p <0.003), history of drug addict (p <0.03) but none was independent predictor of SR. Median FPPY in SR was -0.000 (min -1.5, max 0.00)and in NR and RR was -0.000 (min -1.0, max 0.67.

 

 

Conclusions

Age 40 years and sporadic form of infection are independent determinants of liver fibrosis in chronic hepatitis C. Antiviral therapy slows the rate of fibrosis progression, with regression in the group of sustained response (58% of the cases). The data suggest that BMI 25, iron overload and alcohol in low doses (40g/d) are important co-factors in reducing the antiviral response.

 

Antiviral Properties of Pegylated Versus Non-Pegylated Interferon Alpha-2b Against Hepatitis C in Cell Culture

Virendra Joshi, Karim Awad, Cesar Fermin, Srikanta Dash

 

Introduction

Interferon Alfa in combination with Ribavirin has been effective in the treatment of chronic hepatitis C, but the mechanism is unknown. We reported that Interferon Alfa inhibits hepatitis C virus replication by blocking internal ribosomal entry site (IRES) mediated translation.

 

Aim

To compare efficacy of pegylated versus non pegylated interferon alfa on translation of hepatitis C virus (HCV) using a chimeric clone between green fluorescent protein (GFP) and full length HCV in cell culture.

 

Methods

High level expression of HCV-GFP chimera was achieved in Huh 7 cells using a two step transfection procedure that involves first infection with a replication defective Adenovirus that that expresses T7 polymerase followed by transfection with HCV-GFP plasmid DNA. Huh 7 cells transfected with pDsRed2, in which translation of red fluorescence proteins is mediated by non-IRES mechanism, was used as a control. All cells were counterstained with the nuclear dye (DAPI) to verify each dish had equal number of cells. Antiviral effects of interferon on IRES and non-IRES dependent translation were then examined by looking at protein expression under a fluorescent microscope.

 

Results

Pegylated or non-pegylated Interferon Alfa specifically inhibited IRES-mediated translation of green fluorescent protein of HCV 1b strain in a concentration dependent manner. Both types of interferon did not block translation of red fluorescent protein mediated by non-IRES mechanism. The antiviral effects of Pegylated and Non-Pegylated interferon are comparable (mu/ml versus IU/ml).

 

Conclusions

Results of this study suggest that Pegylated or Non-Pegylated Interferon alfa block internal ribosomal entry site mediated translation of HCV. Addition of Polyethylene glycol to Interferon Alfa did not alter or enhance antiviral properties of Interferon in cell culture.

 

ACKNOWLEDMENTS: This work was possible due to partial support from the National Cancer Institute and Centralised Tulane Imaging Center (CTIC), Department of Pathology

 

Factors Associated with HCV-Stage III/IV Fibrosis: A Retrospective Study of a Large Cohort of US Patients

Ke-Qin Hu, Namgyal L. Kyulo, Victor W. Xia, Shirley X. Hu, Donald J. Hillebrand, Chang-Hong Yu

 

Background

Most previous studies on hepatitis C virus (HCV) related cirrhosis included a small cohort of patients and assessed limited variables. Thus, studies including a large cohort of patients with systematic analyses are required to further define factors associated with HCV-stage III/IV fibrosis.

 

Aims

The present study was aimed to determine the risk factors for and the clinical presentation of HCV-stage III/IV fibrosis in a large cohort of US patients.

 

Methods

The study subjects were collected consecutively from the Liver Clinics at Loma Linda University and the VA Medical Centers between July 1, 1999 and March 30, 2003. The inclusion criteria were positive HCV RNA PCR, liver biopsy-proven chronic hepatitis, negative HBsAg and anti-HIV, and absence of previous HCV treatment. The data were obtained through a retrospective chart review.

 

Results

Of the 460 patients, 331were males and 129 were females with mean age of 48.48.0 years, and 191 (41.7%) had stage III/IV (III = 25.3%, IV = 16.4 %). After adjusting for a history of alcohol use ≥30 g/day, age > 60 years at entry (p=0.03), estimated duration of HCV infection ≥ 25 years (p=0.04), being obese (body mass index ≥30 kg/m2, p=0.017) and a history of diabetes mellitus (DM, p=0.03) were independently associated with stage III/IV fibrosis. Univariate analysis revealed a significant association of 2 X upper limits of normal ALT and AST (i.e. > 80 U/L), AST/ALT ratio > 1, alpha fetoprotein (AFP) > 15 ng/ml, transferrin saturation > 50%, histological activity index (HAI) > 7, and grade II/III hepatic steatosis were significantly associated with stage III/IV fibrosis. After adjusting for elevated ALT, AST/ALT ratio, and transferrin saturation, HAI ≥7, and HCV genotype 1 infection, 2 X upper limit of normal AST (p=0.009), grade II/III hepatic steatosis (p=0.04), and serum AFP ≥ 15 ng/ml (p=0.04) were independently associated with stage III/IV fibrosis. In addition, significantly higher frequencies of hypoalbuminemia (< 3.5 g/L, 70.37%, p < 0.001), thrombocytopenia (< 130 X 109/L, 80.72%, p < 0.001), and splenomegaly (66.34%, p < 0.001) were seen in patients with stage III/IV fibrosis.

 

Conclusion

Elderly, longer duration of HCV infection, being obese, and a history of DM serve as independent risk factors for HCV-stage III/IV fibrosis. Clinically, elevated AST and AFP, grade II/III hepatic steatosis, hypoalbuminemia, thrombocytopenia, and splenomegaly are associated with HCV-stage III/IV fibrosis.

 

Proposal and Validation of the AST/ALT Ratio for Predicting the Presence of Significant Fibrosis in Patients with Chronic Hepatitis C

Edoardo Giannini, Luca Mastracci, Paola Ceppa, Roberto Testa

 

Background

The presence of significant fibrosis is an important diagnostic and prognostic histological hallmark in patients (pts) with chronic hepatitis C. The AST/ALT Ratio proved to be an easy and validated tool for non-invasively assessing cirrhosis in pts with chronic hepatitis C (CHC), although its diagnostic accuracy for significant fibrosis has never been assessed.

 

Aim

To assess and validate the diagnostic accuracy of the AST/ALT Ratio for the diagnosis of significant fibrosis in a cohort of CHC pts.

 

Patients and Methods

Liver biopsies of a cohort of 338 CHC pts were assessed by two pathologists (Ishak score). Nineteen biopsies were excluded because of non-adequate sample. Therefore, 319 pts made up the study group (232 m/87 f; mean age 4612 years). Data set was randomly split into two sub-sets (Identification and Validation sets). The AST/ALT Ratios of the pts were correlated to presence of significant fibrosis (score >2) and histological stage. ROC curves were used to identify and to confirm the accuracy (c-index) of the AST/ALT Ratio for the diagnosis of significant fibrosis. ANOVA was used to assess differences in AST/ALT Ratios among the various fibrosis stages.

 

Results

Identification subset (n=162) and Validation subset (n=157) were no statistically different. In particular, prevalence of significant fibrosis was 43% in both sub-sets. In the Identification subset, an AST/ALT Ratio cut-off of 0.65 had 80% SS, 70% SP, and c-index of 0.802 for the diagnosis of significant fibrosis. In the Validation subset this cut-off had 70% SS, 71% SP, and a c-index of 0.769. Prevalence of significant fibrosis and cirrhosis in the whole cohort of 319 pts were 43% and 24%, respectively. Patients with increasing fibrosis score showed progressively increasing AST/ALT Ratios (score 0=0.53, score 1=0.56, score 2=0.58, score 3=0.60, score 4=0.75, score 5=0.79, score 6=1.18, p<0.001). AST/ALT ratio was significantly correlated to fibrosis score (n=319, rs=0.550, p<0.001).

 

Conclusions

In this study we have shown that in CHC pts the AST/ALT Ratio is able to identify noninvasively the presence of an important histological feature such as significant fibrosis, and were able to validate the results we had obtained. Furthermore, we demonstrated that this simple means is correlated to histological staging. Indeed, as fibrosis scores increased the AST/ALT Ratios of the pts increased as well. The use of the AST/ALT Ratio can be proposed as an alternative tool for non-invasively stage CHC.

 

A Comparative Study on Natural History of Chronic Hepatitis C in Patients with Chronic Renal Failure (CRF) vs. Normal Renal Function (NRF)

Ke-Qin Hu, Steve Lee, Namgyal L. Kyulo, Victor W. Xia, Donald J. Hillebrand, Shirley X. Hu, Chang-Hong Yu

 

Background

Chronic hepatitis C (CHC) occurs in 3-36% of patients with CRF. Although the natural history of CHC has been studied in patients with normal renal function (NRF), it remains to be defined in those with chronic renal failure (CRF).

 

Aims

This comparative study was aimed to determine the clinical presentation of CHC and factors associated with stage III/IV fibrosis in patients with CHC and CRF.

 

Methods

The study subjects were collected consecutively from the Liver Clinic at Loma Linda Univ. Med. Ctr. between July 1, 1999 and March 30, 2003. Group A and B included patients with CRF and NRF, respectively. The inclusion criteria for both groups included patients with positive HCV RNA PCR, liver biopsy-proven CHC, negative HBsAg and anti-HIV, and absence of previous HCV treatment. Both groups were matched in age, gender, history of alcohol use, and estimated duration of HCV infection. The data were obtained through a retrospective chart review.

 

Results

Groups A and B consisted of 91 and 159 patients, respectively. The frequencies of elevated ALT (13.4% vs. 68.8%, p<0.0001), AST (10.8% vs. 64.2%, p<0.0001), mean body mass index (BMI, 26.3 vs. 28.9, p<0.0001), hepatic steatosis (31.1% vs. 58.2%, p<0.0001), and stage III/IV fibrosis (25.0% vs. 42.8%, p=0.0053) were significantly lower, but the frequency of diabetes mellitus (DM, 50.0% vs. 13.9%, p<0.0001) was significantly higher in patients with CRF than in those with NRF. After adjusting for age > 50 years, a history of alcohol use (> 30g/day), BMI > 25 kg/m2, HCV RNA > 1 x 106 copies/ml, steatosis, and serum ferritin > 300 m g/L, a history of DM (p<0.0001), positive hepatic iron staining (p=0.002), AST/ALT ratio > 1 (p=0.005), and a lower frequency of stage III/IV fibrosis (p=0.048) were independently associated with CHC and CRF. In addition, mutivariate analysis indicated that after adjusting for AST > 40 IU/L, AST/ALT ratio > 1, hypoalbuminemia (<3.5g/dL), thrombocytopenia (< 130 X 109/L), and splenomegaly, a history of DM (p=0.047) and hepatic steatosis (p=0.011) were independently associated with stage III/IV fibrosis in patients with CHC and CRF, but not in those with CHC and NRF.

 

Conclusions

Compared to patients with CHC and NRF, those with CHC and CRF commonly present with normal ALT and AST, a history of DM, and lower frequencies of being overweight, hepatic steatosis, and stage III/IV fibrosis. In patients with HCV and CRF, a history of DM and hepatic steatosis are independently associated with stage III/IV fibrosis.

 

High Prevalence of Novel Genotypes in Vietnamese Patients with Chronic Hepatitis C

Mindie H. Nguyen, Huy Trinh, Ruel T. Garcia, Jing Ning, Emmet B. Keeffe

 

Purpose

According to the World Health Organization, the prevalence of infection with hepatitis C virus (HCV) in Vietnamese is 6.1%. Novel genotypes such as genotype 6, 7, 8, and 9 have been reported in Vietnamese and other Southeast Asians according to small published studies. Response to antiviral therapy is highly dependent on HCV genotypes; therefore, knowledge of viral genotype distribution and correct identification of these genotypes are important in the care of Vietnamese patients, who may represent a significant proportion of the population in many geographic areas of the United States.

 

Methods

To examine the prevalence of the various HCV genotypes among Vietnamese, we conducted a cross-sectional study of all new Vietnamese patients who underwent testing for HCV genotypes at an urban community-based gastroenterology medical practice between 2000 and 2002. All HCV genotype testings were performed at Nichols Institute with the Hepatitis C Viral Genotype Core Sequencing Assay (San Juan Capistrano, California). This assay correctly identifies all HCV genotypes including genotype 7, 8, and 9. The commonly used LiPA assay, which is based on variations in the 5 prime unstranslated region of the HCV genome, may misclassify genotype 7, 8, and 9 as genotype 1b (Dev AT, Hepatology, 2002).

 

Results

We identified 308 consecutive Vietnamese patients who underwent HCV genotype testings: 204 (66%) were male and the mean age of all patients was 49 (SD=11). The distribution of HCV genotypes were as follows:

 

 

Conclusions

The most common genotype is genotype 1. Genotype 6,7,8 & 9 make up the second most common group. Approximately one-third of HCV-infected Vietnamese Americans had the novel genotypes (genotype 6, 7, 8, and 9). Current literature suggests that patients with novel genotypes may have more favorable response to antiviral therapy than those with genotype 1 (Dev AT, Hepatology, 2002 and Hui CK, J Inf Dis, 2003). Thus, accurate genotyping with core sequencing assay is essential in Vietnamese patients with hepatitis C who undergo treatment evaluation.

 

Fibrosis Progression Is Dependent upon Hepatic Inflammation in Patients with Chronic HCV

Joon Mo Myung, Iliana Bouneva, Jennifer Salvatori, Melissa J. Contos, Mitchell L. Shiffman, Richard K. Sterling, R. Todd Stravitz, Velimir A. Luketic, Andrea Ferreira-Gonzalez, A. S. Mills, Arun J. Sanyal

 

Introduction

Many patients (pts) with chronic HCV develop progressive fibrosis while others remain stable for 5-10 yrs or longer. Liver biopsy (LBX) is utilized to stage fibrosis. However, the role of hepatic inflammation (INFL) and the effect of this on fibrosis remains undefined.

 

Methods

Patients with chronic HCV who were either non-responders (NR) to prior IFN/RBV or who deferred treatment (tx) were enrolled in a prospective study to monitor fibrosis progression. All pts underwent baseline LBX (prior to IFN tx if administered), were followed prospectively and had repeat LBX after a mean of 6 years (>5 yrs after stopping IFN). LBX were scored according to Knodell HAI without knowledge of clinical or previous histologic data. HCV RNA was determined by Amplicor and genotype by InnoLippa.

 

Results

Summary of the 168 patients:

 

Fibrosis increases stepwise with inflammation and was directly related to an increase in inflammation.

 

Progression was NOT related to:

1)     Serum HCV RNA

2)     Serum ALT

3)     Genotype

4)     Degree of inflammation or piecemeal necrosis

 

Conclusions

Fibrosis progression in NR with chronic HCV is directly related to INFL, specifically PMN, but not serum HCV RNA or demographic factors. This is unaffected by a single course of IFN in those with non-response. These data suggest that NR with mild INFL are at low risk for fibrosis progression and unlikely to benefit from maintenance IFN therapy regardless of histologic stage. Pts with significant INFL are at higher risk to progress and may benefit from agents which reduce INFL. These data underscore the importance of LBX in the management of chronic HCV