P. Czygan

Analysis of bile acid glucuronides in urine: group separation on a lipophilic anion exchanger

A chromatographic separation of glucuronidated bile acids using the anion exchanger diethylaminohydroxypropyl Sephadex LH-20 (DEAP LH-20) is described. Group separation of non-sulfated, non-glucuronidated bile acids, bile acid glucuronides, bile acid monosulfates, and bile acid disulfates was obtained. The method allowed analysis of all these bile acid derivatives in the urine of 15 patients with cirrhosis of the liver and cholestasis. The patients excreted in mean 30.4 mumol/24 h non-sulfated, non-glucuronidated bile acids, 90.3 mumol bile acid monosulfates, and 10.2 mumol bile acid glucuronides. Glycine- or taurine-conjugated were 68% of the non-sulfated, non-glucuronidated bile acids, 96% of bile acid sulfates, and 81% of bile acid glucuronides.

Hepatic Secretion of Bilirubin and Biliary Lipids in Patients with Alcoholic Cirrhosis of the Liver

In patients with cirrhosis of the liver elevated bilirubin concentrations in the plasma could be the result of decreased bilirubin excretion or an overproduction of bilirubin with insufficient excretion of the increased amounts of bilirubin. Under steady state conditions with constant serum bilirubin concentrations bilirubin synthesis equals biliary and urinary bilirubin excretion. In the present study in 10 healthy volunteers and 11 patients with alcoholic cirrhosis of the liver and serum bilirubin concentrations of 7.0 +/- 1.9 mg/dl the biliary excretion of bilirubin was studied by the intestinal perfusion method and compared with the excretion of bile lipids. Biliary excretion of bilirubin in the cirrhotics was 38.7 +/- 8.8 mumol/h, the 10 healthy controls excreted 17.9 +/- 0.9 mumol/h bilirubin. Only minor amounts of bilirubin were excreted in urine. In 4 of the 11 cirrhotics 51Cr-red blood cell half-lives were studied revealing ongoing hemolysis. Bilirubin production calculated from red cell life span was identical to biliary excretion of bilirubin with an error less than 5%. The data indicate that in patients with alcoholic cirrhosis of the liver serum concentrations of bilirubin may be elevated due to overproduction of bilirubin and a concomitant decrease of the biliary transport capacity of bilirubin.

Ethanol and intestinal carcinogenesis in the rat.

The effect of chronic ethanol administration on 1,2-dimethylhydrazine induced rectal carcinogenesis was investigated in 32 paired male Sprague-Dawley rats fed a nutritionally adequate liquid diet containing 36% of total calories either as ethanol or isocaloric carbohydrates. Chronic ethanol ingestion increased the total number of rectal tumors significantly (17 vs. 6, p less than 0.02), whereas no cocarcinogenic effect of ethanol was observed in other parts of the intestine. Alcohol did not influence tumor size or histopathology. A 47% increase in the activity of mucosal alcohol dehydrogenase in the distal colorectum was found between chronically ethanol fed and pair fed controls (0.241 +/- 0.019 vs. 0.164 +/- 0.020 mumol mg protein-1 hr-1, p less than 0.01). This could in part explain the cocarcinogenic effect of alcohol in this tissue. The data give experimental support to the epidemiologic findings of an increased incidence of rectal cancer in the alcoholic.

Microsomal ethanol oxidation in the colonic mucosa of the rat

SummaryA microsomal ethanol oxidizing system (MEOS) is present in the colonic mucosa of the rat. This MEOS metabolizes ethanol to acetaldehyde at the physiological pH of 7.4. Alcoholdehydrogenase or catalase are not involved in the reaction. The Michaelis Menten constante of the reaction is 13.7±0.3 mM and the maximal velocity is 219±30 pmoles acetaldehyde/mg microsomal protein x min.Bacterial ethanol metabolism does not contribute to the acetaldehyde production in the colonic MEOS.Chronic ethanol consumption has no effect on colonic MEOS activity. In addition, chronic ethanol ingestion does not affect colonic microsomal NADPH-cytochrome-c-reductase nor benzo(a)pyrene hydroxylase activity.

Hormone-sensitive adenylate cyclase in human colonic mucosa.

Human colonic adenylate cyclase has been shown to be sensitive to vasoactive intestinal polypeptide (VIP) and prostaglandins of the E- and F-type. Maximal activation of enzyme activity averaged 200% for VIP and 300-350% for the E-prostaglandins. Both classes of hormones had an additive effect on enzyme activity indicating the existence of two distinct hormone-sensitive adenylate cyclases in human colonic mucosa.

Oral Keto Analogs of Branched-Chain Amino Acids in Hyperammonemia in Patients with Cirrhosis of the Liver

Previous uncontrolled studies indicated a positive effect of keto analogs of amino acids on plasma ammonia in patients with cirrhosis of the liver and on portal-systemic encephalopathy. In the present double-blind study the influence of keto analogs of the branched chain amino acids valine, leucine and isoleucine on plasma ammonia and encephalopathy was investigated in 12 patients with cirrhosis of the liver and surgical portal systemic shunts. In addition to the usual therapy with lactulose and protein restriction (40 g protein/day) all patients received 15.24 g keto analogs and placebo orally over 4 weeks in a crossover regimen. In contrast to uncontrolled studies, plasma ammonia, which was elevated in all patients before the beginning of the study, was not significantly changed. In addition plasma amino acids, electroencephalogram, number connection test, clinical state and laboratory tests were not influenced by the therapy with keto analogs.

Induction and activation of rat liver microsomal bile salt glucoronyltransferase

In vivo induction and in vitro activation of the recently described bile salt glucuronyltransferase were investigated in rat. A radioactive assay for the determination of glucuronyltransferase activity was used. 14C-Labeled bile salts served as substrates, and the glucuronides were separated by thin layer chromatography. Lithocholate glucuronyltransferase activity was determined in liver microsomes of phenobarbital- and 3-methylcholanthrene-treated rats and of untreated controls. Pretreatment with phenobarbital induced lithocholate glucuronyltransferase activity to 150.5% of controls. In contrast, 3-methylcholanthrene treatment decreased activity to 29.6% of controls. In vivo activation of lithocholate glucuronyltransferase by Triton X-100 was observed in controls and in the 3-methylcholanthrene group, but not in the phenobarbital group. Substrate activation of the enzyme by lithocholate was demonstrated in microsomes of untreated controls. Pretreatment with 3-methylcholanthrene, but not phenobarbital, increased the latency of lithocholate glucuronyltransferase. The results indicate that rat liver microsomal bile salt glucuronyltransferase activity is increased by in vivo induction with phenobarbital and by in vitro activation with detergents like Triton X-100. The induction of bile salt glucuronide formation by phenobarbital is most likely one of the factors contributing to the increased biliary and fecal excretion of bile salts in patients with cholestasis following phenobarbital therapy.

Stimulation of Thymidine Incorporation in Isolated Rat Intestinal Mucosal Cells by Feeding an Ethanol-Containing Liquid Diet

The effect of chronic ethanol administration on the incorporation of 3H-thymidine into DNA of isolated intestinal cells in the rat was investigated. Chronic ethanol consumption increased significantly DNA synthesis in intestinal mucosal cells. Whether this enhancement of DNA synthesis as a measure for cellular regeneration is due to the local effect of ethanol via tissue injury, to a direct stimulatory effect of ethanol on enzyme activities involved in DNA metabolism or to a change in the intestinal cell population with a shift to immature crypt cells by ethanol is not known.

Different effects of dietary chenodeoxycholic acid and ursodeoxycholic acid on colonic adenylate cyclase in the rat.

SummaryThe oral administration of dietary chenodeoxycholic acid (1%), but not of ursodeoxycholic acid (1%), to male Sprague Dawley rats results in a significant increase in the colonic adenylate cyclase activity without any influence on the colonic cyclic-AMP phosphodiesterase activity. No effect of chronic bile acid feeding on the response of colonic adenylate cyclase to prostaglandin E2 and vasoactive intestinal peptide is observed. These data emphasize a dependence of the cyclic-AMP adenylate cyclase activation on the chemical structure of the bile acid. This may be of pathophysiologic relevance with respect to the frequently observed diarrhea as a side effect of oral chenodeoxycholic, but not ursodeoxycholic acid therapy for cholesterol gallstone dissolution in man.

Colonic cyclic AMP metabolism following chronic ethanol consumption in the rat: Effect of hormonal secretagoques

The colonic cyclic AMP system is known to be involved in intestinal secretion and can be stimulated by a variety of gastrointestinal hormones including prostaglandins. We have investigated the effect of chronic ethanol ingestion on the activity of the key enzymes in cyclic AMP metabolism--adenylate cyclase and cyclic AMP phosphodiesterase--in the colonic mucosa of the rat. Chronic ethanol consumption by feeding a nutritionally adequate liquid diet enhanced basal colonic adenylate cyclase activity significantly by 168% (p less than 0.01), but had no effect on colonic low Km cyclic AMP phosphodiesterase activity. In addition, various hormonal secretagogues were used to stimulate colonic adenylate cyclase. Colonic adenylate cyclase exhibited a significantly greater sensitivity and efficacy to prostaglandins and vasoactive intestinal peptide after chronic ethanol ingestion. Since increased intestinal cyclic AMP production due to an increased activity of intestinal adenylate cyclase is known to promote intestinal secretion of water and electrolytes, the frequently observed diarrhea in alcoholics may be explained at least in part by an enhanced production of colonic cyclic AMP.