340A A A S L D A B S T R A C T S HEPATOLOGY O c t o b e r 1995

933 CHRONIC HEPATITIS C OF DRUG USERS : INFLUENCE OF HIV INFECTION

S. Pol, N. Tnnh Thi, V. Thiers, F. Jaffredo, F. Camot, B. Lamorthe, H. Zylberberg, P. Berthelot, C. Br@chot, B. Nalpas. Liver Unit, INSERM U-370, H6pital Necker and Hybddotest, Institut Pasteur, Pads, France.

To evaluate the influence of HIV infection on HCV-related chronic hepatitis, we compared biological (AST, ALT and GGT activities), histological (cirrhosis or not, Knodell score), virological (HCV genotypes) and therapeutical data in a large sodas of intravenous drug users (IVDU), according to the HIV status.

We studied 212 patients (162 males and 50 females, mean age 32 years, mean anciennety of drug addiction 11 years), including 60 HIV-positive patients (28.3%). Seventy eight patients (16 HIV-positive and 62 HIV-negative) were given Interferon therapy (3 MU thdca weekly subcutaneously for 6 months).

AST and GGT activities were significantly higher in HIV- positive than in HIV-negative subjects (p<0.001 and p<0.01, respectively) although ALT were similar. Histological scores were not statistically different (7.5+4.1 vs. 5.1+3.1) but cirrhosis was 3.5-fold frequent in HIV-positive (25% vs. 7%)(p<0.001). The HCV genotype distibution was comparable in both groups. Chronic alcohol consumption was equally observed in both groups and had no clear influence on liver histology.

Interferon therapy led to a pdmary reponse (normalization of transaminase activities dudng treatment) and a sustained response (normalization of aminotransferase at least 6 months after Interferon withdrawal) in 64.5 and 40.3% of HIV-negative and 18.8 and 0% of HIV-positive patients.

In summary, although poody modifying the clinic.o-biological features of HCV infected drug users, HIV infection cleady increases the sevedty of the liver disease and decreases the hopes of HCV eradication. These results underline the needs of eady diagnosis and treatment in drug users, especially those infected by HIV.

934 HEPATITIS C VIRUS DENSITY HETEROGENEITY AND VIRAL TIT_RE IN ACUTE AND CHRONIC INFECTION: A COMPARISON OF IMMUNODEFICIENT AND IMMUNOCOMPETENT PATIENTS. JP Watson. DJ Bevitt. GP Suiekett. GL Toms and MF Bassendine. Departments of Medicine, Virology and Immunology, University of Newcastle upon Tyne, Framlington Place, Newcastle upon Tyne, NE2 4HH, UK.

Heterogeneities in the buoyant densit~ of hepatitis C virus (HCV) have been reported in different groups of patmnts, and have been attributed to differential binding of fl-lipoproteins and IgG, and the presence of HCV nucleocapsids in circulation. It may be that HCV density heterogeneity correlates with the severity of liver disease, the HCV RNA titre, and the immunocompetence of the patient. Methods: Patient groups tested were as follows: sequential samples from 3 immunodeficient pattents (2 with common variable immunodeficiency, I with X-linked hypogammaglobulinaemia) who had been acutely infect~i with a batch of ]fitravenous immunoglobulin contaminated with HCV (genotype la) and progressed to chronic disease; sequential samples from 1 immunocompetent patient who presented with acute HCV (genotype 1, subtype not classified) and progressed to chronic disease; single samples froni 7 immunocompetent patients with chronic HCV (2 patients:genotype la, 3 patients:genotype lb, 1 patient:genotype 2, 1 patlent:genotype 3). Serum samples were analysed by differential flotation ultracontrifugation in NaCI solution (density 1.06~g/ml). The high and low density~HCV fractions were tested for the presence of RNA by RT-PCR. All serum samples were also quantitated for HCV RNA (Amplicor HCV Monitor kit, Roche Diagnosuc Systems). Results: 3/4 of the acutely infected patients presented with low density HCV (fl-lipoprotein associated). High density _HCV (IgG essociated/nucleocapsid) was detected later in the acute intections, and was present in all samples from PCR positive patients with chronic infeetton. The .patient with X-linked agammaglobulinaemia progressed from low denstty to high density RNA in the absence of an antibody resVonse. 2/3 lmmunodeficient patients and 2/8 immunocompetent patients had high HCV RNA titres (> 1 x 106 viral copies/ml). Serum HCV RNA titre did not relate to clinical course: for example the immunodeficient patient with the lowest titre of the three immunodeficient patients analysed did not respond to a-interferon and progressed to cirrhosis within 2 years of infection, whereas one of the other two patients has responded to or-interferon and is now PCR negative, and the other remains PCR positive but stable with normal liver function tests. This suggests the host immune response is important in the progression of HCV/elated disease.

935 DYNAMIC CHANGES OF FLOATING DENSITY IN CIRCUlaTING HEPATITIS C VIRAL PARTICLES ASSOCIATED WITH DISEASE ACTIVITY. K Hino, K F~] ii, M Oknzaki, M. Korenaga, C. Murakami, M. Oknda, K. Okita. Dept. of Interual Mad. Yamaguchi University, School of Medicine, Y~maguohi Japan.

It is assumed that intact HCV virions have a low d~msity close to tlmt of low density lipoproteins and nuoleosapsid proteins or HCV immtme complexes have a higher density. We therefore ez~ni- nod the change of flsating density in circulating HCVloarticles ansoding to the disease activity in chronic hepatitis u. ~ t ~ Serum samples were obtained at intervals of one year from 5 patiests whose ALT levels had remained normal for over [ years (group A); and at the time of high and lowALT levels from patients who showed remarkable fluctations in ALT levels(group B. Each sart~n sample was layered on an NaC1 solution with a density of 1.063 g/ml. Low density a~d high density particles were respect-

collected from top and bottom fractions after ultracentrifu-

~ tien at 139500Xg for 22h. 100/g from bottom fractions were incu- ted witl~ rabbit anti-human immuuggloblin and separated into Supernatan. ts and pellets after centri~gation at 680×g for 15m~q to tect HCV lmmuue osmplexes. HOV-RNAwas detected by l~r-POR and l~s levels in the top fractions were compared with those in the bottom fractious by serial end point dilution. HCV-RNA levefls inpellets were similarly comA0ared with those in supermatan~s, m additlon we e]mminod t~tal HCV-RNA levels using the branched DNA assay ard the quas_ ispecies in hyL0ervariable region~HV~) of HCV-I~NAby hetero-dfiplex method in a±± serum samples. ~uuizu High density particles were 10 to 100-fold more compared to low density ones in all patients of group A, regardless of sampling time, while titers of both particles became tOe same accordingly, as the elevation of ALT levels took place in 4 patients, and were constantly the same in one patient of group B. HCV immune compleXes were detected in all patients except for one of group B who showed the highest ALT level. HOV-RNA mas detected from superr~tants after immunopreoi- pitation in only onepatient of group A. However, it was detected and its levels were ~ne same or less than those from pellets at tl~ time of high levels of ALT in 5 pa t ients of group B. Total HCV-ENA laa~is in serum or quasispecies of HVRwere not significantly changed aecerding to sa~oling time in both groups. Comclmmi~ These results suggest that low density particles of HCV, indicating free virioss ; and high denmty particles except for immune complexes, assumed to be released nueleocapsid proteir~ from hepatocytes, increase according to the disease activity. By contrast, HCV circulates predomir~ntly as immune complexes, and dyr~mie changes of floating density were not observed in asymptomatio carriers.

936 THE "TAKE-OVER" PHENOMENON IN PATIENTS INFECTED WITH MULTIPLE HEPATITIS C VIRUS (HCV) GENOTYPES Qian KP, Natov SN 1, Pereira BJG 1. Lau JYN Section of Hepatobiliar S Diseases, University of Florida, Galnesville, FL, and New England Medical Center 1, Boston, MA.

Background Patients infected with HCV genotypes 1, 2 and 3 had similar liver biochemistry and viremia profile in both HCV+ blood donors or chronic HCV patients. However, the haman body is a complex biological system and subtle differences in replication efficacy between genotypes may not be revealed. Hveothesis In patients infected with multiple HCV genotypes, one genotype may prevail, either related to a difference in repfication efficacy, or to cruss-intederenre. Ai___m To study HCV genotypes in patients with serological evidence of previous exposure to multiple HCV genotypes. Method In this cross-sectional study, sera from 120 anti-HCV+ patients with chronic renal failure on hemodialysis (HD, a group known to be at risk for infection with multiple HCV genotypes) were screened for genotype-specific antibodies by competitive EIA based on NS4 region (Murex, Kent, UK). This competitive binding assay has been shown to be highly specific for genotype-specific antibody which can be taken as evidence for previous exposure to a particular HCV genotype. HCV genotypes in these samples were determined by restriction fragment length polymorphism (RFLP) of 5'UTR. This assay required a ratio 5-10:1 (i.e. 10-20% of viral population) before the minor genotypes can be detected. For those patients shown to have antibodies reactive to multiple genotypes, a line probe assay (LiPA-HCV, Innogenetics, Ghent, Belgium) was also used to detect the minor genotypes since this assay can detect a ratio 100:1 (i.e. 1% of the population). Results Of the 120 anti-HCV+ HD patients tested, 50 were positive for genotype-specific antibody (sensitivity 42%; compared to 70-80% in chronic HCV patients), consistent with the suggestion that these patients were less capable of mounting a good antibody response. 8/50 patients had antibody reactivity to more than one genotype (type 1 and 2, n=2; 1 and 3, n=5; 1, 2 and 3, n = 1). However, only genotype I was detected by RFLP in the same samples (la n = 6; lb n = 1; la + lb, n = 1). When LiPA was applied for a more sensitive detection, the same genotyping results as RFLP were obtained. PCR Analysis of the 120 samples 101/120 were PCR+ based on the 5'UTR and 9/101 had more than one major HCV type detected, further confirming that infection with multiple HCV genotypes was common in this population. Conclusion This preliminary data suggests that HCV genotype I may have survival advantage over genotypes 2 and 3. A longitudinal study is currently being conducted to further confirm this observation.