Endoscopy, extracorporeal shockwave lithotripsy (ESWL) and local lysis with alkaline solution of EDTA and bile salts in water were applied in combination in four patients with extra- and intrahepatic pigment stones as well as calcium bilirubinate covered concrements of the biliary tract. In the first patient (a man aged 80 years) a giant concrement of the bile duct was broken up after ESWL by three weeks of local chemical lysis and the fragments were removed by endoscopy. In the second case (man, aged 72), a nonextractable pigment stone was at first reduced in size by four-day local lysis and then removed endoscopically. Intrahepatic pigment stones were completely removed in the other two patients (boy of 12, man of 62) by local lysis only in 3 and 15 weeks, respectively. Even long-term use of the alkaline solution may not cause any serious side effects. Breaking up of stones after size reduction with ESWL of giant stones, size reduction of intact stones and contact lysis of intrahepatic stones are three important indications for chemical dissolution of biliary tract stones, respectively.

Blind liver biopsy is sufficient for patients with diffuse-parenchymatous liver diseases. The accuracy rate of guided, laparoscopic liver biopsy is not enhanced.

In 15 patients (13 women and two men) with cholesterol stones in the gall-bladder a special (Thistle) catheter was introduced into the gall-bladder under local anaesthesia by percutaneous transhepatic puncture. Methyl-tert-butyl ether, 2-15 ml, was injected via the catheter and removed again after 2 min. The number of stones per gall-bladder averaged 6.3 (1-20), size of stones 1.7 cm (0.5-2.8 cm), and duration of treatment 11.9 h (5-24 h). The stones dissolved in 13 patients (87%). In three patients stone débris remained: in one it was ultimately sucked out after reduction of the amount of débris with an EDTA-containing solution. The side effects of treatment--nausea and vomiting--were minor. In one patient there was a leak of bile from the gall-bladder after the procedure; a cholecystectomy was uneventfully performed. Another patient developed haemobilia which responded to conservative treatment. MTBE treatment has thus proved to be a successful and cheap method, low in side effects, in the treatment of patients with gall-stones.

Objective:To investigate whether enzyme release caused by chenodeoxycholic acid (CDCA) can be prevented by ursodeoxycholic acid (UDCA),and to study the effects of bile acids on bile secretion, glutamate dehydrogenase (GLDH) release and mitochondrial membrane structure. Methods: Totally 0.1-0.5 mmol/L bile acids were perfused into rat livers for 120 min. The bile duct was cannulated for collection of bile flow and GLDH was determined. Intact mitochondria were isolated and mitochondrial suspension was detected with electron paramagnetic resonance spectroscopy (EPR) for membrane mobility and polarity values. Results: Compared with control, CDCA at 0.1,0.3, 0.5 mmol/L decreased the bile flow by 12%, 77.25% and 78.98%, and enhanced GLDH release by 3, 9 and 21 times, respectively. It also increased the mobility of 4-maleimido-TEMPO spin label and the polarity of hydrophobic membrane interior. UDCA increased bile flow by 1.8 times at 0.3 mmol/L and 1.9 times at 0.5 mmol/L. It did not influence enzyme release and membrane structure. Prior infusion with UDCA (0.1 mmol/L) for 30 min followed by combination of UDCA and CDCA improved bile secretion, delayed enzyme release and partly prevented the membrane lesion caused by CDCA compared with CDCA alone. Conclusion:CDCA can damage mitochondrial membrane structure and result in liver dysfunction. UDCA improves secretion of bile and partly prevents liver mitochondrial lesion against CDCA. Low concentration of CDCA does not damage the liver function.

Large unilamellar vesicles were prepared from phosphatidylcholine (PC), sphingomyelin (SM), cholesterol (Chol) and cardiolipin (CL) by an extrusion technique (LUVETs). Diffusion of the more hydrophobic lithocholic acid (LCA) and the less hydrophobic chenodeoxycholic acid (CDCA) was investigated by using the pyranine fluorescence method. Membrane permeability was studied by measuring the inclusion of carboxyfluoresceine (CF) into the lipid vesicles, and membrane fluidity was determined with diphenylhexatriene (DPH) and trimethylammonium-diphenylhexatriene (TMA-DPH). All results indicate that, CDCA compared to LCA, exhibits a significantly better penetration into vesicles containing SM. LCA penetrates better into vesicles containing cholesterol. Small amounts of CL influenced the diffusional properties of CDCA more than those of LCA. Since Lamcharfi et al. (1997a) Euro. Biophys. 25, 285-291 have observed differences in the conformational forms of CDCA and LCA in solution, it is suggested that the diffusion rate of bile acids through (model-)membranes is not only dependent on hydrophobicity, but also on bile acid di-(poly-)meric associations and on membrane-lipid composition.

BACKGROUND:In some patients with primary biliary cirrhosis, ursodeoxycholic acid causes full biochemical normalisation of laboratory data; in others, indexes improve but do not become normal.AIMS:To characterise complete and incomplete responders.METHODS:Seventy patients with primary biliary cirrhosis were treated with ursodeoxycholic acid 10-15 mg/kg/day and followed up for 6-13 years.RESULTS:In 23 patients (33%) with mainly stage I or II disease, cholestasis indexes and aminotransferases normalised within 1-5 years, except for antimitochondrial antibodies. Histological findings improved. Indexes were not normalised in 47 patients (67%) although the improvement of their biochemical functions parallelled the trend in the first group. In these incomplete responders histological findings improved to a lesser extent. The only difference between the two groups before treatment was higher levels of alkaline phosphatase and gamma glutamyl transpeptidase in the incomplete responders. At onset of treatment the discriminant value separating responders from incomplete responders was 660 U/l for alkaline phosphatase and 131 U/l for gamma glutamyl transpeptidase. One year later it was 239 and 27 U/l (overall predictive value for responders 92%, for incomplete responders 81%). There were no differences between the two groups concerning immune status, antimitochondrial antibody subtypes, liver histology, or any other data. HLA-B39, DRB1*08, DQB1*04 dominated in both groups.CONCLUSIONS:In patients with mainly early stages of primary biliary cirrhosis, higher values of alkaline phosphatase and gamma glutamyl transpeptidase are the only biochemical indexes which allow discrimination between patients who will completely or incompletely respond to ursodeoxycholic acid treatment.

Background & Aims: The Dubin-Johnson syndrome is characterized by conjugated hyperbilirubinemia and by impaired secretion of anionic conjugates from hepatocytes into bile. Absence of the multidrug-resistance protein 2 (MRP2; symbol ABCC2), an adenosine triphosphate-dependent conjugate export pump, from the hepatocyte canalicular membrane is the molecular basis of this syndrome. The aim of this study was the elucidation of all exon-intron boundaries of the MRP2 gene as a prerequisite for the analysis of mutations in patients with Dubin-Johnson syndrome. Methods: Exon-intron boundaries of MRP2 were determined, and the amplified exons were screened for mutations. Immunofluorescence microscopy served to localize the MRP2 protein in human liver. Results: The human MRP2 gene is similar to 45 kilobases long; it contains 32 exons and a high proportion of class 0 introns, In 2 patients with Dubin-Johnson syndrome, we detected a nonsense mutation at codon 1066 and a 6-nucleotide deletion mutation affecting codons 1392-1394, The MRP2 protein was absent from the canalicular membrane of both patients, Conclusions: The mutations detected so far show that Various mutations in the MRP2 gene can lead to the Dubin-Johnson syndrome. The exon-intron boundaries established in this article will facilitate the analysis of additional mutations in the MRP2 gene.

Intact mitochondria were incubated with and without calcium in solutions of chenodeoxycholate, ursodeoxycholate, or their conjugates. Glutamate dehydrogenase, protein and phospholipid release were measured. Alterations in membrane and organelle structure were investigated by electron paramagnetic resonance spectroscopy. Chenodeoxycholate enhanced enzyme liberation, solubilized protein and phospholipid, and increased protein spin label mobility and the polarity of the hydrophobic membrane interior, whereas ursodeoxycholate and its conjugates did not damage mitochondria. Preincubation with ursodeoxycholate or its conjugate tauroursodeoxycholate for 20 min partially prevented damage by chenodeoxycholate. Extended preincubation even with 1 mM ursodeoxycholate could no longer prevent structural damage. Calcium (from 0.01 mM upward) augmented the damaging effect of chenodeoxycholate (0.15–0.5 mM). The combined action of 0.01 mM calcium and 0.15 mM chenodeoxycholate was reversed by ursodeoxycholate only, not by its conjugates tauroursodeoxycholate and glycoursodeoxycholate. In conclusion, ursodeoxycholate partially prevents chenodeoxycholate-induced glutamate dehydrogenase release from liver cell mitochondria by membrane stabilization. This holds for shorter times and at concentrations below 0.5 mM only, indicating that the different constitution of protein-rich mitochondrial membranes does not allow optimal stabilization such as has been seen in phospholipid- and cholesterol-rich hepatocyte cell membranes, investigated previously.

Background & Aims: Ursodeoxycholic acid (UDCA) is used for treatment of primary biliary cirrhosis. Previous studies showed that, compared with UDCA mono-therapy, bile salts plus prednisolone had no further effect on laboratory data but improved liver histology. Thirty percent of these patients had prednisolone-related side effects. Budesonide is a glucocorticoid with a high receptor affinity and a high first-pass metabolism. In this study we investigated whether budesonide and UDCA are superior to UDCA mono-therapy. Methods: A 2-year prospective, controlled double-blind trial was performed. Twenty patients (mainly with early-stage disease) were treated with UDCA at a dose of 10-15 mg/kg daily in addition to 3 mg budesonide 3 times daily (group A), and 19 patients (1 dropped out for personal reasons) were treated with UDCA plus placebo (group B). Liver biopsy specimens were taken before, after 12 months, and at the end of study. Glucose tolerance tests, serum cortisol levels, and adrenocorticotropin-stimulated cortisol secretion were assessed at regular intervals. Bone mass density was measured by dual-energy photon absorptiometry. Results: Compared with pretreatment values, liver enzyme and immunoglobulin M and G levels decreased significantly in both groups. Improvement in group A was significantly more pronounced (P < 0.05) than in group B. Titers of antimitochondrial antibodies did not change. In group A, the point score of liver histology improved by 30.3%; in group B, it deteriorated by 3.5% (P < 0.001). Changes in bone mineral density after 2 years were -1.747% in group A and -0.983% in group B (P = 0.43). Budesonide had little influence on the hypothalamic-pituitary-adrenal axis. One patient in group A had budesonide-related side effects; in 3 patients in group B, complications of liver disease developed. Conclusions: Combination therapy with UDCA and budesonide is superior to UDCA and placebo.

Background: PBC without antimitochondrial antibodies (AMA) is called autoimmune cholangitis.PBC without or with AMA plus antinuclear antibodies (ANA) or smooth muscle antibodies (SMA) and the histological features of PBC and chronic autoimmune hepatitis is called overlap syndrome.Patients and Methods: Because it has been shown that patients with AMA-positive OS respond differently to UDCA therapy from patients without OS, we investigated whether AMA-positive OS is different from PBC with respect to biochemical, serological and morphological criteria.Results: From a collective of 103 PBC-patients the data of 70 patients have been evaluated.45 (64%) had an AMA-positive overlap syndrome, 25 (36%) a PBC.There were no statistically significant differences between the two groups concerning stages of the disease, histological activity, AMA and AMA-subtypes, IgM, IgG, inflammation-indicating enzymes (GLDH, AST, ALT), cholestasis enzymes and the course of the disease.15/45 (33%) of the patients with OS and 7/25 (28%) with PBC responded rapidly to medical therapy, 30 patients (67%) and 18 (72%) responded but slowly.Conclusions: AMA-positive overlap syndrome is not different from primary biliary cirrhosis with respect to biochemical, serological and morphological data.Although there were no differences between patients with OS and PBC concerning response to medical therapy, this needs to be confirmed in a larger study.

For the medical treatment of cholestasis plant alkaloids, phenobarbital and S-adenosyl-L-methionine (SAMe) have been used. Although the mode of action of these substances is understood in part, the treatment of patients was unsuccessful. In contrast, ursodeoxycholic acid, a physiologically occurring bile acid in man, was successful. The daily dosage of ursodeoxycholic acid is 10-15 mg/kg bodyweight. Best results have been obtained in primary biliary cirrhosis where symptoms improved markedly, the cholestasis-indicating enzymes and immunoglobulin M decreased significantly. After long-term therapy even liver histology improved. Recently it has been shown that ursodeoxycholic acid prolongs the interval to liver transplantation. Ursodeoxycholic acid has to be taken lifelong, because interruption of therapy, even after long periods of continued treatment will induce a rebound of cholestasis. Ursodeoxycholic acid therapy is without side effects. In patients with primary sclerosing cholangitis ursodeoxycholic acid prolongs life expectancy only in combination with endoscopic bile duct dilatation.