The role of ursodeoxycholic acid in bile acid-mediated kidney fragment toxicity.

Elevated levels of bile acids are thought to play an important role in the renal failure of patients with obstructive jaundice undergoing surgery. In contrast, ursodeoxycholic acid (UDA) is widely used to improve cholestasis and has been proposed as protective bile acids and antioxidant. The present study employs kidney fragments to determine the role of reactive oxygen species (ROS) in the mechanism of toxicity of hydrophobic bile acids and to determine the nephroprotectant properties of UDA against the hydrophobic bile acids. The hydrophobic bile acids chenodeoxycholic (200 microM) and deoxycholic acid (200 microM) significantly (P<0.05) increased lactate dehydrogenase leakage (LDH) from glomerular fragments from 2.7+/-0.4 to 5.03+/-0.23 and 4.66+/-0.37 (micromol NADH consumed/min/mg protein) for chenodeoxycholic and deoxycholic acid respectively. Preincubating the fragments with UDA (500 microM) did not prevent the leakage of LDH caused by the bile acids. The level of lipid peroxidation was not increased in fragments exposed to either ursodeoxycholic (0-500 microM), lithocholic (0-100 microM), chenodeoxycholic (0-500 microM) or deoxycholic acid (0-500 microM). Furthermore UDA (500 microM) did not prevent the increase in lipid peroxidation caused by tert-butyl hydroperoxide (0-1000 microM) in the fragments. These results suggest that hydrophobic bile acids do not cause lipid peroxidation in kidney fragments and that UDA is neither capable of preventing the loss of membrane integrity induced by hydrophobic bile acids or acting as an antioxidant in kidney fragments.

The role of reactive oxygen species in adriamycin and menadione-induced glomerular toxicity.

Redox cycling leading to oxidative stress has been proposed as the mechanism by which adriamycin induces glomerular toxicity in rats. The present study compares the extent of the oxidative stress and cytotoxicity induced by adriamycin to menadione (a model redox cycling quinone) in freshly isolated rat glomeruli. Adriamycin and menadione (25 microM) decreased de novo protein synthesis (measured by 3H-proline incorporation into acid-precipitable glomerular protein) by 50 and 85%, respectively, in 2 h. By contrast, menadione at 25 microM reduce glomerular membrane integrity (as assessed by lactate dehydrogenase leakage), adriamycin reduced membrane integrity at 500 microM adriamycin. Reactive oxygen species (ROS) were measured by the oxidation of dihydrodichlorofluorescein. Menadione (25 microM) and adriamycin (25 microM) increased ROS formation to 260 and 156% of controls after 30 min incubation, respectively. Oxidative stress was assessed by measuring the intracellular level of reduced glutathione (GSH) and the decrease of the NADPH/NADP- ratio which stimulates the pentose phosphate pathway (PPP): (a) menadione (25-100 microM) reduced glomerular GSH to 10-20% of controls, adriamycin (25-100 microM) had no effect; (b) menadione (10 microM) increased PPP activity 6-fold, while adriamycin (125 microM) had only a 2-fold effect. Although adriamycin and menadione generate extensive ROS and decrease protein synthesis, there was no correlation between the extent of oxidative stress and cytotoxicity in glomeruli exposed to adriamycin. These results suggest that oxidative stress may not be the primary mechanisms by which adriamycin induces selective glomerular toxicity.

The effects of Bile Acids on Freshly Isolated Rat Glomeruli and Proximal Tubular Fragments.

The role of bile acids in post-surgical acute renal failure in jaundiced patients is obscure. In this study the effects of 11 bile acids were assessed on freshly isolated rat glomeruli and proximal tubular fragments using de novo protein synthesis and lactate dehydrogenase (LDH) leakage as markers of cytotoxicity. Lithocholic acid inhibited protein synthesis from 5mum, chenodeoxycholic and deoxycholic acid from 50mum (P<0.05). The concentration of hydrophobic bile acids that inhibited protein synthesis by 50% (IC(50)) was 10mum, 75mum and 80mum for lithocholic, chenodeoxycholic and deoxycholic acids, respectively. The glycine and taurine conjugates of these bile acids had no significant effect on de novo protein synthesis up to 200mum. Lithocholic acid (50mum), chenodeoxycholic (200mum) and deoxycholic acids (200mum) caused a significant increase (P<0.05) in LDH leakage. Lithocholic acid also directly inhibited LDH activity above 50mum (P<0.05), whereas chenodeoxycholic acid and deoxycholic acid had no effect on LDH below 500mum, at which concentration they caused a slight increase in activity. The cytotoxic bile acids had no effect on the level of reactive oxygen species in kidney fragments. Hydrophobic bile acids inhibit protein synthesis and increase membrane permeability. Hydrophobic bile acids also directly alter LDH activity. Kidney cells are susceptible to the hydrophobic bile acids at concentration significantly below their critical micellar concentration. These results suggest that both glomeruli and tubules are highly sensitive to hydrophobic bile acids.