67 CHARACTERIZATION OF M9 BY WESTERNBLOT AND RELEVANCE OF ANTI-M9 AS A MARKER ANTIBODY FOR EARLY PRIMARY BILIARY CIRRHOSIS (PBC)

R.Klein,P.A.Ber~ Department of Internal Medicine,University of TObingen,West Germany

Recent ly an organ s p e c i f i c antzbody agaznst l i v e r m i tochondr ia (an t i -M9) was descr ibed in pts w i th PBC as w e l l as zn hea l thy r e l a t i v e s and normal c o n t r o l s . A n t i - M 9 were de tec ted by ELISA using t r y p s i n i z e d submi tochondr ia l p a r t i c l e s (SMP) from r a t l i v e r . Fu r the r studzes weceper- formed to f i n d out whether the s p e c i f i c i t y o f ant i -M9 zn PBC pts d i f f e r e d from t h a t in healthy persons and whether ant i -M9 can be considered as a t r ue marker f o r PBC as r e s e n t l y pos tu l a t ed .

The M9 an t igen was f u r t h e r p u r i f i e d by ion exchange chromatography using the lO0000g su- pe rna tan t from r a t l i v e r SMP,and the d i f f e r e n t peaks ob ta ined by t h i s procedure were t es ted by ELISA using s tandard ized a n t z m i t o c h o n d r i a l a n t i b o d i e s as w e l l as ant i -Mg.The f r a c t i o n conta inzng p r e f e r e n t i a l l y M9 was a lso t es ted by Westernblot .Two a n t i g e n i c de te rminants a t a mo lecu la r we ight o f 98 and 69kD were d i s t i n g u i s h e d by a l l an t i -M9 pos /an t i -M2 neg sera (n=lO) as w e l l as by ant i -M9 and ant i -M2 pos sera (n=lO),and by 14 o f 17 anti-MR pos sera from hea l thy r e l a t i v e s . I n con t ras t , none o f the ant i -M9 neg but ant i -M2 pos PBC sera (n=15) and the c o n t r o l s (n=15) reac ted in the Westernb lo t wzth t h i s M9 f r a c t i o n . Re t rospec t i ve a n a l y s i s o f pts w i t h e a r l y and l a t e PBC revea led t h a t the 98 and 69kD s p e c i f i c r e a c t i o n could be detec ted p r e f e r e n t i a l l y i n sera from pts w i t h e a r l y PBC (54% o f 104 pts w i th PBC stage I / I I vs 27% o f 74 pts w i t h PBC stage I I I / I V ) . S e r i a l t e s t i n g o f sera from on ly ant i -M9 pos pts w i th h i s t o l o g i c a l l y proven PBC showed t h a t a l l the sera recogn ized the same a n t i g e n i c determinants over an obse rva t i on per iod o f 6-12 years . We conclude that these two antigenic determinants of M9 are specif ical ly related to PBC (especially to early and anti-M2 neg PBC) and also to relatives of PBC pts including husbands. The detected epitopes could,therefore,be indzcative for the postulated infectious aetiology of PBC.

68 PERMEABILITIES TO SUCROSE AND FERROCYANATE AND MAXIMAL BILIARY PRESSURE ARE INCREASED IN CIRRHOTIC RATS.

S. Kr~henbUhl, J. Reichen Department of Clinical Pharmacology, University of Berne, CH-3010 Berne, Switzerland.

Hypercholeresls, presumably of ductular origln, has been described in patients with l iver cirrhosis. L i t t l e is known, however, aSout the characteristics of canalicular bi le formation in cirrhosis of the l iver . We therefore measured bi le flow, b i l i a ry pressure and permeability (as b11e to plasma ratio P) to sucrose (s) and ferrocyanate ( f ) under basal condltions and during TC-infusion at 0.4 and 0.8 umoles/min/lO0 g b.wt. in c i r - rhotic (CIR, induced by phenobarbital/CCl4 n=7) and control (CTR n=6) male SD rats. Cir- rhosis was present in al l animals; serum bile acid levels were higher in CIR (4.2±SD 2.6 vs 1.1_+0.6 ~mol/L; p<O.05). Maxlmal secretory pressure (MSP) did not d i f fe r during basal conditions but were higher during TC infusion in CIR (14.7±2.6 vs 10.6±2.4 mmHg; p<O.05). Basal bi le flow and bi le salt excretion in CIR was comparable to that of CTR. The osmotic choleretic capacity of TC (calculated as the slope of bi le flow vs TC output) was higher in CIR (I0.2±I.8 vs 7.3+ 1.2 ul/umol; p<O.Ol). Permeability to both, sucrose and ferro- cyanate, was higher in ~R than in CTR, averaging 0.27+0.16 vs 0.098+0.042 (p<O.O01) and 0.079_+0.049 vs 0.037+_0.018 (p<O.O01), respectively. -The ratio of-Ps/Pf, a measure of charge-selectivity, was not altered, however. We conclude from our data that the f i l t e r - ing surface for penetration of s and f is higher in c i r rhot ic rats whereas the electr ical barrier is unaltered.Cirrhotic rats produce more bi le for a given load of bi le salt there- by developping a higher MSP during TC infusion. Bile flow is not impaired in cirrhosis. To the contrary, since we have shown cirrhot ic l ivers to contain a smaller fractional volume of hepatocytes (Hepatology 6: 1123), bi le formation per hepatocyte must be higher in c irrhot ic rats. Thus, part of hypercholeresis in cirrhosis could be of canalicular rather than ductal origin.

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