Evidence for involvement of oxygen free radicals in bile acid toxicity to isolated rat hepatocytes

Abstract

The mechanisms by which hydrophobic bile acids are toxic to the liver are unknown. To determine whether the generation of free radicals is involved in the hepatotoxicity of bile acids, freshly isolated rat hepatocytes were incubated with individual bile acids (100 to 200 mumol/L) for 4 hr. Hepatocyte viability (trypan blue exclusion) declined to 40% to 50% in incubations with taurochenodeoxycholic acid and taurolithocholic acid, whereas taurocholic acid and tauroursodeoxycholic acid were not toxic. Lipid peroxidation was significantly associated with the loss of cell viability. Preincubation with different antioxidants-D-alpha-tocopheryl succinate, D-alpha-tocopherol, diphenyl-p-phenylenediamine, superoxide dismutase, catalase, superoxide dismutase + catalase, deferoxamine or apotransferrin-protected against the loss of viability and inhibited lipid peroxidation in cells incubated with 200 mumol/L taurolithocholic acid. alpha-Tocopheryl succinate added after 90 min of incubation with taurolithocholic acid ameliorated further hepatocyte toxicity and lipid peroxidation. Incubation of hepatocytes with 500 mumol/L of taurochenodeoxycholic acid or taurolithocholic acid under a low oxygen tension (9% O2) similarly caused lipid peroxidation and cell injury that was reversed by preincubation with D-alpha-tocopherol. These data suggest that oxygen free radicals may be involved in the pathogenesis of bile acid hepatotoxicity.

Deoxycholate induces DNA damage and apoptosis in human colon epithelial cells expressing either mutant or wild-type p53

“…Apparently, secondary bile acids stimulate the production of reactive oxygen species (ROS) and also reactive nitrogen species (RNS). In considering ROS, lipid peroxidation products were increased in rat hepatocytes after being exposed to bile acids [30], but this was inhibited by various antioxidants [31]. The bile acid-induced DNA damage in HT-29 cells mentioned above was inhibited when cells were pre-treated with the antioxidant, vitamin E, before exposure to the bile acids [17].…”

The NAD+ precursors, nicotinic acid and nicotinamide protect cells against apoptosis induced by a multiple stress inducer, deoxycholate

“…The cell pellets were resuspended in 100 mM mercury orange (MO) for 5 min on ice, then centrifuged for another 5 min at 1200 r.p.m. 20 The pellets were resuspended in ice-cold PBS supplemented with 1% FCS.…”

Effects of bile acids on biliary epithelial cell proliferation and portal fibroblast activation using rat liver slices

“…3 Accumulation of toxic bile acids results in hepatocellular damage, ultimately leading to liver fibrosis and cirrhosis. Bile acid cytotoxicity has been demonstrated on hepatocytes in vitro 4,5 and in vivo in rat models, 6,7 and they are known to induce hepatocyte apoptosis. 8 There is generally a correlation between bile acid hydrophobicity and cytotoxicity: the hydrophobic lithocholic and deoxycholic acids exert the most toxic effects, whereas the hydrophilic ursodeoxycholic and cholic acids have little toxicity.…”