Rat liver alkaline phosphatases. Evidence hepatocyte and portal triad enzymes differ.

Using biochemical and electron microscopic histochemical techniques, we studied membrane-bound alkaline phosphatase activities of rat hepatocytes and portal triads. Activity in portal triads was localized to capillaries surrounding bile ducts (peribiliary plexus) and arterioles. Despite the reputation of alkaline phosphatase as a "biliary enzyme," activity was not observed in bile ducts. Livers were separated into hepatocyte and portal triad fractions with collagenase. Enzyme from hepatocytes migrated faster during electrophoresis and eluted later during anion-exchange chromatography than that from portal triads. Thus, hepatocyte enzyme is more negatively charged (and also possibly smaller) than portal triad enzyme. Twelve hours after bile duct obstruction, new activity appeared on lateral and sinusoidal membranes of hepatocytes; appearance of portal triads did not change with obstruction. Electrophoretic mobilities of the two forms were not altered by obstruction. We conclude that two distinct liver alkaline phosphatases exist, one in hepatocytes, the other in portal triad blood vessels.

Perineal Crohn's disease complicated by pyogenic liver abscess during metronidazole therapy.

A patient with ileal and perineal Crohn's disease developed a pyogenic liver abscess caused by Streptococcus milleri. The perineal process had been treated with high-dose metronidazole for the preceding 6 mo, and culture of a perianal abscess 2 mo before admission revealed heavy growth of Streptococcus milleri (which was misidentified as an anaerobic Peptostreptococcus). This case illustrates the difficulty in correctly identifying Streptococcus milleri, the most frequent isolate from pyogenic liver abscesses. Furthermore, the events suggest that metronidazole therapy can allow overgrowth of this organism in perineal abscesses which, in turn, may predispose to the development of purulent foci such as liver abscesses. This effect on the bacterial ecology of the perineum should be considered when opting for metronidazole therapy of perineal Crohn's disease.

Induction of hepatic alkaline phosphatase by colchicine: role of cholestasis and bile acids.

We studied the induction of hepatic alkaline phosphatase by colchicine, a drug which impairs bile flow. By analogy to bile duct obstruction, where retained bile acids induce alkaline phosphatase, we hypothesized that the mechanism of induction involves the drug's cholestatic property and subsequent intrahepatic retention of bile acids. After administering colchicine to bile-fistula rats, we monitored bile flow. In colchicine-treated rats which did not develop cholestasis, we varied the transhepatic flux of taurocholate 15-fold over the physiologic range in order to study the role of intrahepatic bile acids. Induction of alkaline phosphatase was observed when cholestasis was absent and the bile acid flux was 5% of normal. The magnitude of enzyme induction was not augmented by increasing the transhepatic flux of taurocholate. The results demonstrate that colchicine induces hepatic alkaline phosphatase by a mechanism which operates independently of cholestasis and is not modified by the level of bile acid in the liver.

Similarities and differences between models of extrahepatic biliary obstruction and complete biliary retention without obstruction in the rat.

Biliary obstruction in the rat causes known biochemical and morphological abnormalities. These abnormalities might be caused simply by retention of biliary constituents or might require other factors such as high biliary pressure, interruption of the normal flux of fluid, ions, and other biliary constituents through the hepatocyte and/or changes in intracellular concentrations, binding, and distribution of biliary constituents. We studied a choledochocaval (CC) fistula rat model characterized by complete biliary retention without bile stasis. CC fistula animals were compared with biliary obstructed rats over a 46-hr period. Bile flow and bile acid excretion rate reached high levels by hour 5 and remained high in CC fistula rats. Urinary bile acid excretion, shrinkage of the rapidly exchangeable bile acid pool, and serum bile acid and bilirubin concentrations were equivalent in the two models. Serum cholesterol concentration was higher and serum alkaline phosphatase and 5'-nucleotidase activities lower in obstructed rats. The percentage of beta-muricholate rose and the percentage of cholate decreased in bile acid from obstructed rats. This was associated with increased microsomal 6 beta-hydroxylase activity. Bile acid composition and microsomal 6 beta-hydroxylase activity changed little in CC fistula rats. Morphological examination revealed greater damage in obstructed rat livers. These data indicate that factors other than simple retention of biliary constituents contribute to the abnormalities observed in extrahepatic biliary obstruction.

Bile acid-dependent secretion of alkaline phosphatase in rat bile.

The role of bile acids in the biliary secretion of alkaline phosphatase was studied. Rats with external bile fistulae were drained for 4 hr. After 2 hr, bile acid secretion fell progressively. Alkaline phosphatase secretion also decreased progressively during the period of drainage, suggesting that secretion of the two components was related. Each rat was then given an i.v. infusion of the taurine conjugate of either cholate, chenodeoxycholate, or ursodeoxycholate. Alkaline phosphatase secretion increased in a dose-dependent manner as bile acid secretion was varied over and beyond the physiologic range. Each bile acid affected alkaline phosphatase secretion differently: given at 0.5 mumoles per min per 100 gm, tauroursodeoxycholate caused a 3-fold, taurocholate a 14-fold, and taurochenodeoxycholate a 75-fold increase in enzyme secretion. To determine if these findings might represent elution of canalicular enzyme by bile acids, isolated liver surface membranes were incubated with the bile acids. Like the findings in vivo, taurochenodeoxycholate was strongest and tauroursodeoxycholate weakest in removing alkaline phosphatase from the membrane. Differential centrifugation of liver surface membranes after exposure to bile acids and ultracentrifugation of bile showed that more than half of the enzyme released by the action of bile acids did not sediment at 100,000 g and, thus, could be considered soluble. These results document bile acid-dependent secretion of alkaline phosphatase in rat bile and suggest that the process involves solubilization of both membrane fragments and free enzyme from membranes lining the biliary space.

Hypervitaminosis A unmasked by acute viral hepatitis.

A case of acute hypervitaminosis A complicating viral hepatitis is reported. Twenty days after presenting with hepatitis B, a 42-yr-old vegetarian developed acute hypervitaminosis A in the absence of recent, massive exposure to the vitamin. Findings included headache, confusion, skin desquamation, and hypercalcemia. Prior to developing hepatitis, he had ingested supplemental vitamin A without recognized ill effect. Liver and serum vitamin A without recognized ill effect. Liver and serum vitamin A levels were both elevated; the liver biopsy showed abundant, lipid-filled Ito cells and perisinusoidal fibrosis. This case demonstrates that patients with excessive hepatic stores of vitamin A may develop hypervitaminosis A during acute, intercurrent liver disease. Levels of retinol binding protein are reduced in hepatitis. This phenomenon may account for the findings in this case, since vitamin A is more toxic when not specifically bound to retinol binding protein. The size of the population at risk for this complication of hepatitis in unknown, but presumably it is growing with the widespread use of supplemental vitamin A.

Nature of bile acid maximum secretory rate in the rat.

We studied the determinants of maximum bile acid secretory rate (SRm) in the rat. The choledochocaval fistula rat model manifested a bile acid secretory rate far in excess of the SRm previously reported for taurocholate in this species. We studied the ability of various bile acid solutions to maintain the high secretion rate in this model. Whole-rat bile, but not taurocholate in 2% albumin nor rat bile with bile acid content over 90% taurocholate, maintained secretion rate. We concluded that the mixture of bile acids in rat bile was the most important determinant of the high secretion rate and that the high rate was not due to a peculiarity of the model itself nor to the infusion of biliary lipids together with bile acids. Conventional determination of the SRm in the bile fistula rat confirmed this impression, with the least toxic bile acids manifesting the highest SRm. During infusion of taurocholate beyond the SRm, bile flow and bile acid secretion rate fell. This was accompanied only by scattered focal necrosis of single liver cells or of small aggregates of cells and not by any diffuse subcellular morphological change. We believe the maximum bile acid secretory rate is determined by toxicity of a specific bile acid for the secretory mechanism rather than by a limitation in transport receptor number as is usual with substances manifesting classical transport maxima. The high SRm of the 7 beta-hydroxy bile acid, ursodeoxycholic acid, is probably related to its very low toxicity. The high SRm in the choledochocaval fistula rat is probably related to the presence of 7 beta-hydroxy muricholic acids in the bile of this species.

Bile acids modify alkaline phosphatase induction and bile secretion pressure after bile duct obstruction in the rat.

Bile acids induce synthesis of alkaline phosphatase by cultured hepatocytes. To test whether bile acids account for this enzyme's elevation during cholestasis, we developed an experimental model in which the content of the bile acid pool is controlled at the beginning of cholestasis. After depleting the bile acid pool by external biliary drainage, we obstructed bile flow in four groups of rats and then replenished the pool in three groups with taurocholate, taurochenodeoxycholate, or tauroursodeoxycholate, respectively, and replaced no bile acid in the fourth. Hepatic bile acid concentrations were elevated in all obstructed groups; however, the levels were higher in the groups that received bile acid replacement. Cholate was not metabolized, but both chenodeoxycholate and ursodeoxycholate were transformed to beta-muricholate. Although hepatic alkaline phosphatase activity rose in all obstructed animals, the levels achieved were higher in each group treated with bile acids than in the group that was not. Increases in plasma activities of alkaline phosphatase and 5'-nucleotidase occurred only after treatment with cholate or chenodeoxycholate. Bile secretion pressure was higher after ursodeoxycholate or no bile acid treatment but was lower after cholate or chenodeoxycholate replacement. Thus, all bile acids tested induced hepatic alkaline phosphatase to some degree, but only certain ones--those which reduced bile secretion pressure--released the enzyme into plasma. We conclude that, in the rat, the hepatic response to acute cholestasis is influenced by the composition of the intrahepatic bile acid pool and that various bile acids have significantly different effects on this response.

Nitrofurantoin: another cause of drug-induced chronic active hepatitis? A report of a patient with HLA-B8 antigen.

Nitrofurantoin, an agent used extensively in chronic urinary tract infections, is rarely incriminated as a hepatotoxin. In this 27 year old women chronic active hepatitis first developed during exposures to nitrofurantoin, and she later suffered exacerbations. Earlier reported cases are reviewed which describe a spectrum of hepatotoxic reactions. This patient has HLA-B8, an antigen associated with autoimmune forms of chronic active hepatitis. It is postulated that the antigen may be associated with liver injury initiated by drugs such as nitrofurantoin.