Biochemical Pharmacology, Vol. 36, No. 1, pp, 51-57, 1987. Printed in Great Britain. oocl5-2952/8753.cKJ + 0.00 Pergamon Journals Ltd. zyxwvut THE ROLE OF ACROLEIN IN ALLYL ALCOHOL-INDUCED LIPID PEROXIDATION AND LIVER CELL DAMAGE IN MICE HARTMUT JAESCHKE, CHRISTIN KLEINWAECHTER and ALBRECHT WENDEL Physiologisch-Chemisches Institut der Universitaet Hoppe-Seyler-Str. 1, D-7400 Tuebingen, Federal Republic of Germany zyxwvutsrqponmlkjihgfedcbaZYXWVUTSR (Received 30 May 1985; accepted 7 July 1986) Abstract-Male NMRI mice were fed a sucrose diet for 48 hr in order to reduce the hepatic glutathione content and to level off its diurnal variation. After administration of ally1 alcohol (AA: 1.1 mmol/kg), hepatic glutathione (24.3 2 7.0 nmol GSH/mg protein) was almost totally lost within the first 15 min (<OSnmol GSH/mg protein). Subsequently, a massive lipid perioxidation was observed, i.e. the animals exhaled 414 * 186 nmol ethaue/kg/hr compared to 0.9 + 0.8 of controls, and the hepatic TBA- reactive compounds had increased from 55 * 16 pmol/mg protein in controls to 317 2 163 after 1 hr. Con~mitan~y, a 40-45% loss of the pol~nsaturated fatty acids (arac~donic and do~osahexaenoic acid) in the liver lipids was observed. About 80% of the cytosolic alcohol dehy~ogenase activity and about 50% of the microsomal P450-content were destroyed. In vivo-inhibition of alcohol dehydrogenase by pyrazole or induction of aldehyde dehydrogenase by phenobarbital abolished AA-induced liver damage as well as glutathione depletion and lipid peroxi- dation, while inhibition of aldehyde dehydrogenase by cyanamide made a subtoxic dose of AA (0.60 mmol/kg) highly toxic. These results strongly favour the importance of acrylic acid formation as an additional detoxification pathway. Enhanced hepatic levels of glutathione protected in vivo against the damaging effects of AA. Depletion of the liver glutathione content by phorone or diethylmaleate alone caused marginally enhanced lipid peroxidation (phorone) but no liver cell damage. Mono- oxygenase inhibitors (metyrapone, diethyldithiocarbamate, cu-naphthoflavone) or an inducer (benz(a)pyrene) did not affect AA-induced toxicity. The ferric iron chelator desferoxamine- methanesulfonate prevented AA-induced lipid peroxidation and liver cell damage in vivo. In vitro, acrolein alone failed to initiate lipid peroxidation in soy bean phospholipid liposomes or in mouse liver microsomes. Thus, acrolein not only impairs the glutathione defense system but also directly destroys cellular proteins and evokes lipid peroxidation by an indirect iron-depending mechanism. In a series of papers [l-5], we reported that acute intoxication of mice with certain drugs, such as par- acetamol, leads to lipid peroxidation and simul- taneously to liver damage. All compounds used were metabolized by the microsomal monooxygenase system, and we discussed reactive oxygen species, derived from the monooxygenase system during the metabolism of these compounds, as possible initiators of lipid peroxidation. In contrast to this type of compounds, ally1 alcohol (AA)* represents an established hepatotoxin [6] which is metabolized via the cytosolic enzyme alcohol dehydrogenase to acrolein [7]. Acrolein is considered as the toxic metabolite [8, 91 which may be respon- sible for the extensive periportal necrosis observed in the rodent liver after acute ally1 alcohol intoxi- cation [6, 91. Acrolein is of great toxicological importance, not only since AA and acrolein are important synthetic intermediates in chemical indus- tries [RI-123 but acrolein occurs also as a toxic metab- olite of the widely used anticancer drug cyclo- phosphamide [13], as a combustion product of petrol, * Abbreviations used: AA, ally1 alcohol; LPO, lipid per- oxidation; TBA, thiobarbitu~c acid-reactive material; SGOT, serum glutamic oxaloacetic trans~inase; SGPT, serum glutamic pyruvate transminase; ADH, alcohol dehydrogenase; AlDH, aldehyde dehydrogenase. coal, wood and plastics [14] and is a component of tobacco smoke [12]. Acrolein is a powerful elec- trophile which spontaneously reacts with sulphhydryl groups [ 151. Incubation of isolated hepatocytes with acrolein decreased cellular glutathione levels and caused cell death [14, 161. In this system cytosolic enzymes were released with a similar time course as TBA-reactive material was formed [14]. Therefore, peroxidative damage of lipids might be the cause of the cytotoxicity of acrolein. The aim of the present study was to investigate the special contribution of glutathione depletion, lipid peroxidation and protein denaturation to ally1 alcohol-induced liver cell damage as well as the role of different detoxification pathways in vim. Fur- thermore, mechanistic aspects of AA-induced lipid peroxidation were addressed. MATERIALSAND METHODS Chemicals. Ally1 alcohol (AA), pyrazole, phorone, diethylmaleate and a-naphthoflavone were purchased from Fluka AG (Buchs, Switzerland), reduced glutathione was obtained from Boehringer AG (Mannheim, F.R.G.), metyrapone and des- feroxaminemethanesulfonate (Desferal~) were obtained from Ciba Geigy AG (Basel, Switzerland) and cyanamide, diethyldithiocarbamate and 51