414A AASLD ABSTRACTS HEPATOLOGY October 1995

1 2 2 9 BILE ALCOHOLS UNDERGO AN ENTEROHEPATIC CIRCULATION IN AN ELASMOBRANCH, LITTLE SKATE, RAIA ~ R. Woessner*. J. Bover ~ . G. Kurz +. G Fricker s, Mount Desert Island Biological Laboratory, Salsbury Cove, ME; *Sandoz Pharma Ltd., Basle, e l l ; *Yale Medical School, New Haven, CT; +Institute for Org. Chem. and Biochem., Freiburg, FRG; *Institute for Pharm. Technol. and Biopharm., Heidelberg, FRG

The sulfated bile alcohol scynmol sulfate, 3a,7ot,12~24~,26,27- bexahydmxy-5B-cholestane-26(27)-sulfate is the major bile salt in bile of the elasmobranch little skate. To investigate the hepatic uptake and secretion of bile alcohols in skate liver [SH]-3ot,7~12ct-trihydroxy-5B-cholestane (C'T) was used as a model compound. Uptake of CT into freshly isolated hepatocytes occurred by both saturable and passive mechanisms. Transport of CT could not be inhibited by tanmcholate, whereas taumcholate transport was inhibited in a noncompetitive manner by CT, suggesting that bile alcohols were transported and delivered by a system being separate from the tanmcholate transporting system. To examine biliary secretion, [3H]-CT was injected into the portal vein of isolated perfused skate livers. Bile was collected over 7 hours because of the very slow bile flow of only 40-50 gl/h. Ten percent of a given, dose of radioactivity was secreted into bile as polar metabolites of CT during the time interval, whereas only nonmetabotized CT remained in the livers at the end of the peffusion. The slow secretion rate and the high recovery in the liver tissue suggest that metabolism may be the rate limiting step in the hepatic elimination of CT. The major metabolites secreted into bile were identified by mass spectrometry and chromatographic analysis as scymnol and scymnol sulfate. When the metabolites were administered to isolated perfused livers, more than 25% could be recovered in bile after 7 hours. To study the entembepatic circulation of bile alcohols, CT or the metabolites were administered imo the duodenum of free swimming skates and bile was collected through indwelling canulas over a 4-day period. More than 90% of radioactivity could be recovered from bile, indicating that there was a nighly effective absorptive mechanism in the GI-tract, as well as specific hepatic uptake and biliary secretion. In summary this is the first direct demonstration of an entemhepatic circulation of bile idcnhols in fish liver.

1230 P H E N O T Y P I C C H A R A C T E R I Z A T I O N O F H U M A N BILIARY E P I T H E L I A L C E L L S (BEC) IN S1TU AND IN V I T R O L Fabris .*M Strazzabosco.-E Mart inez-Anso.-J Medina:~J Prieto.M Auth .A Strain.J Neuberger & R Joplin; Liver Unit, University Hospital, Birmingham UK;*Universi ty of Padova, I taly.-Universi ty of Navarra, Spain

In cirrhosis, intra-hepatic BEC populations may be altered both by damage to bile ducts and an increased proport ion o f proliferative ductular cells. The a i m of this study was to identify the site o f origin of BEC which are isolated by immune-separation f rom cirrhotic human liver and used for in vitro studies of disease pathogenesis. M e t h o d s BOth BEC and tissue sections were obtained f rom the same normal (2) and cirrhotic (6) liver. Markers used were: H E A l 2 5 , cytokeratin 19 (CK19) , anion exchanger-2 (AE-2), hepatocyte growth factor receptor (HGFr), CFTR (R-domain specific) and epithelial membrane antigen (EMA). In histological sections, the distribution of antigens was studied by confocal microscopy after sequential double immupofluorescence staining. Light ankt electron microscopy together with "H-thymidine incorporation were used to determine the phenotype of isolated BEC and Ceils which proliferate in response to HGF. Results Bile ducts in tissue sections were categorised according to size: MD - marginal ducts (no discernible lumen), ILD - interlobular ducts ( < 1001xm), S D - septal ducts ( > 100~tm). H E A l 2 5 showed strongest staining in all compartments of the biliary tree, while CK19, AE2 and EMA stained ILD and SD but not MD. CFTR stained BEC of SD and ILD less intensely than MD. HGFr positivity appeared to be predominantly localised in ILD and SD. Confocal analysis of sections showed a clear apical localization for AE2 and CFTR, while HGFr and H E A l 2 5 were mainly bas01ateral. More than 90% of freshly isolated cells and those grown in the presence o f H G F were positive for all markers except CFTR. No difference between cells isolated from normal and cirrhotic liver was detected. Conclusions BEC lining the MD demonstrated some phenotypic differences when compared with the ILD and SD. Analysis o f BEC in vitro indicated that isolated cells and BEC cultured with HGF are derived from ILD and SD but not MD suggesting that the BEC preparation procedure may select against the purification of MD.

1 2 3 1 S e r o p r e v a l e n c e o f hepatitis A (HA) i n f e c t i o n : r e l a t i o n s h i p w i t h soc io -

e c o n o m i c c o n d i t i o n s . D.Facchinetti. A. Saleri, R Negrini, F. Donato, A. Lanzini. Departments of Medicine 1, Hygiene and Biochemistry, Spedali Civili.Brescia. Italy.

HA is declining in Italy, and we aimed to study the relationship between this changing pattern of infection with changes in socio-economie status. We studied 415 residents in a recently industrialized rural area, selected

according to age among consecutive persons attending general practitioners' clinics. Selected subjects filled in a questionnaire on their present and childhood (age o f 8 years) socio-economic conditions, and gave a blood sample for HA antibodies (Ab) detection. Mean seroprevalence of HAV-Ab was 86%. Age related prevalence is shown in the figure.

100 - 80 :

(*/,) so : 4 0 , 2 0 .

o l "18 22 26 30 34 38 42 46 so 154 58 52 6 6 70 Age(years)

Figure 1

Analysis of risk factors was limited to 128 subjects younger than 35 years, because prevalence was 100% in older age groups. HAV semprevalence was associated with the present number of adults sharing the house (13<0.01). Other present and Childhood indicators of socio-economic stares, including domestic overcrowding, housing tenure, shared toilet and absence of a hot water tap, and father's and subject's education were not risk factors for HAV seroprevalence. We conclude that changes in HAV Ab seroprovalence are independent from socio-economic factors. Better hygiene and sanitation are probably responsible for the reduced circulation of HAV.

1232 ENLARGED OF LYMPH NODES (LN) WITHIN THE HEPATO-DUODENAL LIGAMENT (HDL) AS INDICATOR OF LIVER DAMAGE IN PATIENTS ANTI-HCV PosITIVE. K__FAINBOIM. N. MENDEZ. E. ALVAREZ. T. SCHRODER, M. SAWICKI, E. GANCEDO. Hospital F. J.Mulliz, Buenos Aires. Argentina.

The presence of Anti-HCV antibodies (Abs) usually implies HCV infection and liver damage (LD). Even in this situation,fluctuation or even normality of ALT is well recognized,therefore this is not a good indicator of LD.On the other hand the lack of histological lesion may represent resolved infection or a "carrier" stataEalarged LN within the HDL have been found both in primary biliary ¢irrosis end acute viral hepatitis.In this study we investigated the association of the presence of LN in the HDL with Anti-HCV Abs,biofhemieal alterations and the grading of histological activity index (HAl). 125 patients with confirmed Anti-HCV (LIA- test),HIV and HBV neg,were ultrasound studied before the liver biopsy was performed.According to the slze,the LN were scored as 0:LN less of 9 mmJ:with a size between 10-15 mm and lI:bigger than 15 ram.On a previous study on 170 normal blood dunnors the size of the LN was never higher than 9 mm.Results:Messurable visualization of the HDL was technically sound in 118 out of the 125 patients studied.

Lymph nodes in HDL n (%) u 0 ! _~ I+~H

Normal (15) 15 (100) 0 (0) 0 (0) 0 (0) Minimal (28) 12 (43) 12 (43) 4 (14) 16(57) Mild (48) 8 (17) 17 (35). 23 (48) 40(83) Moderate (27) 3 (11) S (18,5) 19 (70) 24(89)

Enlarged LN in HDL in patients with normal ALT

Chronic hepatitis n: 13 10/13 (77%) Normalhistology n: 15 0/15 (0%) Conclusions:The presence of LN in HDL of patients Anti-HCV positive was observed in 78% of patients showing different degrees of LD.It was also detected an association between the size o f the LN end the HA/scot .This results indicate that the presence of LN is associated with LD independently of ALT modifications.The presence of this LN could represent an interesting model for further studies on the inmune mechanisms associated with liver damage.