Kai O. Lindros

Centrilobular expression of ethanol-inducible cytochrome P-450 (IIE1) in rat liver

Western blot analysis of digitonin eluates as well as immunohistochemical analysis revealed a 30-fold higher concentration of cytochrome P-450IIE1 in the centrilobular than in the periportal regions of the rat liver. Ethanol treatment caused a selective centrilobular induction of P-450IIE1, whereas phenobarbital induced P-450IIB1/2 in both liver lobule regions. The heterogeneous distribution pattern of P-450IIE1 was also observed in cells isolated from either region and correlated to the relative content of P-450IIE1 mRNA in the two cell types. The regiospecific expression and induction of P-450IIE1 may explain why several hepatotoxins, known to be metabolized by this isozyme, primarily damage the centrilobular region in the liver.

Liver aldehyde and alcohol dehydrogenase activities in rat strains genetically selected for their ethanol preference

Abstract Rat strains raised by genetical selection for either high (AA strain) or low (ANA strain) voluntary ethanol consumption were compared with respect to their hepatic alcohol and aldehyde dehydrogenase activities. Liver alcohol dehydrogenase activity was lower in both sexes in the AA strain compared with the ANA strain. The NAD-dependent aldehyde dehydrogenase activity was higher in the mitochondrial and microsomal fractions and lower in the soluble fraction in the AA strain than in the ANA strain. These differences were more pronounced in females than in males. The NAD-dependent utilization of acetaldehyde in liver homogenates was higher in the AA strain in both sexes, when the initial acetaldehydehyde level was 0·40 mM, but there was no difference at 0¢13 mM acetaldehyde. It is concluded that the higher activities in the AA strain are due mainly to those aldehyde dehydrogenases of mitochondrial and microsomal fractions, which have K m -values for aldehydes in the millimolar range. The higher alcohol dehydrogenase and lower aldehyde dehydrogenase activity in livers of rats of the ethanol-avoiding ANA strain may contribute to the previously found higher acetaldehyde levels in blood and liver of rats of this strain after ethanol administration.

Effects of long-term ethanol treatment on aldehyde and alcohol dehydrogenase activities in rat liver.

Abstract Administration of intoxicating doses of ethanol by gavage for 3 weeks caused weight loss and reduced hepatic aldehyde dehydrogenase activity in the soluble, mitochondrial and microsomal fractions. Rats receiving equivalent amounts of ethanol as a constituent of a liquid diet for 5 weeks gained weight and showed no changes in aldehyde dehydrogenase activity. Alcohol dehydrogenase activity was decreased in the rats treated by gavage and unchanged in those given ethanol in the diet, but in spite of this the rate of ethanol elimination was accelerated in both groups. In the livers of two strains of rats genetically selected for their difference in voluntary alcohol consumption, the mitochondrial and microsomal aldehyde dehydrogenase activities had previously been shown to be significantly higher in the alcohol-consuming (AA) than in the alcohol-avoiding (ANA) rats. Similar differences were now found after long-term intragastric ethanol administration, although in both strains the absolute levels of aldehyde dehydrogenase were reduced. Profound reduction of mitochondrial low- K m aldehyde dehydrogenase activity and high blood acetaldehyde were observed, especially in the ANA rats. This suggests a possible connection between the low activity of this enzyme and the increased acetaldehyde level.

Ethanol-induced hypoglycaemia in man: its suppression by the alcohol dehydrogenase inhibitor 4-methylpyrazole.

Abstract. Infusion of ethanol (0.6 g/kg body wt) caused marked hypoglycaemia in subjects fasted for 36 h. Previous administration of the alcohol dehydrogenase (ADH) inhibitor 4‐methylpyrazole (4‐MP, 7 mg/kg body wt i.v.) strongly suppressed the ethanol‐induced hypoglycaemia. The rate of ethanol elimination was 84.6 mg/kg per hour. 4‐MP at the dose used caused a 21% reduction of ethanol elimination, but had no significant effect on blood acetaldehyde levels. 4‐MP also significantly suppressed the ethanol‐induced elevation of blood lactate and almost completely prevented the increase in the 3‐hydroxy‐butyrate/acetoacetate ratio, but had only a slight effect on the lactate/pyruvate ratio of venous blood. The results demonstrate that the hypoglycaemia and lactacidaemia produced by the oxidation of alcohol can be prevented by a dose of 4‐MP that diminishes or prevents the ethanol‐induced shift in the NAD‐coupled redox state of the liver.

Effect of 4-Methylpyrazole on ethanol elimination rate and hepatic redox changes in alcoholics with adequate or inadequate nutrition and in nonalcoholic controls☆

Abstract Ethanol elimination rates produced by chronic alcohol consumption were higher in chronic alcoholics with adequate nutrition than in alcoholics with inadequate nutrition. 4-Methylpyrazole (4-MP, an inhibitor of alcohol dehydrogenase, ADH) at a dose of 7 mg/kg body weight inhibited ethanol elimination by 29% in nonalcoholic controls and by 35% in alcoholics with adequate nutrition, but only by 19% in alcoholics with inadequate nutrition. Galactose elimination rate (which is inhibited by NADH during ethanol oxidation) appeared to be a more sensitive indicator of hepatic redox changes than the lactate/pyruvate ratio of the peripheral venous blood. Ethanol-induced inhibition of galactose elimination was reduced in alcoholics as compared to nonalcoholic controls, and it was more reduced in alcoholics with poor nutrition than in alcoholics with adequate nutrition. Both in controls and in alcoholics, 4-MP reduced the inhibitory effect of ethanol on galactose elimination. In alcoholics with inadequate adequate nutrition, however, the further effect of 4-MP in this respect was negligible. The results indicate that the NADH reoxidation rate in chronic alcoholics is increased and support the existence, in chronic alcoholics with inadequate nutrition, of a non-ADH pathway for ethanol that does not produce reducing equivalents. In alcoholics with inadequate nutrition the proportional contribution of the ADH pathway to total ethanol elimination appears to be more decreased, resulting in reduction of the redox-mediated acute metabolic effects of alcohol.

Acinar distribution of glutathione-dependent detoxifying enzymes. Low glutathione peroxidase activity in perivenous hepatocytes.

The acinar distribution of glutathione S-transferase (GST), glutathione peroxidase (GPx), glutathione reductase (GR), and glucose-6-phosphate dehydrogenase (G-6-PDH) was examined by analyzing periportal (p.p.) and perivenous (p.v.) rat hepatocytes selectively isolated by the digitonin-collagenase perfusion. The cytosolic GST activity was higher in p.v. cells, but the microsomal GST and cytosolic GR were found to be evenly distributed in the acinus. In contrast, the activity of both the Se-dependent GPx and the microsomal (Se-independent) GPx, as well as G-6-PDH, was much lower in the p.v. than in the p.p. cells. The heterogeneous distribution of GST, GPx and G-6-PDH was confirmed by analyzing liver perfusion effluents collected after ante- or retrograde digitonin infusion. The relatively low activities of GPx and G-6-PDH in the p.v. cells could partly explain the susceptibility of this region to chemical injury.

Breath and blood acetaldehyde concentrations and their correlation during normal and calcium carbimide-modified ethanol oxidation in man

With the use of new and improved analytical techniques, concentrations of acetaldehyde in antecubital venous blood and breath of human volunteers were measured after (a) pretreatment of subjects with ethanol and the aldehyde dehydrogenase inhibitor, calcium carbimide and (b) treatment with ethanol only. Breath acetaldehyde concentrations were converted to equivalent pulmonary blood concentrations using an experimentally determined blood: breath partition ratio for acetaldehyde of 190. Under all experimental conditions, blood acetaldehyde concentrations calculated from breath analysis were seen to closely reflect those measured by direct blood analysis. Treatment of subjects with calcium carbimide resulted in elevated blood and breath acetaldehyde concentrations which were rapidly lowered by the intravenous infusion of 4-methyl pyrazole. Peak blood acetaldehyde concentrations ranged from 25 to 188 muM after calcium carbimide and ethanol treatment, but were only 6-11 muM after ethanol treatment alone (1.2g/kg).

The antiestrogen toremifene protects against alcoholic liver injury in female rats

BACKGROUND/AIMSnFemales are generally considered to be more susceptible to alcohol-induced liver injury than males. To elucidate whether gonadal hormones are involved, female rats were chronically treated with ethanol and with an antiestrogen.nnnMETHODSnEthanol was administered in a low-carbohydrate liquid diet. Estrogen action was blocked by daily intubation of toremifene, a non-hepatotoxic second generation estrogen receptor antagonist.nnnRESULTSnThe female rats consuming intoxicating amounts of ethanol diet for 6 weeks developed massive microvesicular/macrovesicular steatosis, frequent inflammatory foci and spotty necrosis. Serum alanine aminotransferase increased 7-fold. Toremifene treatment did not affect steatosis, but significantly reduced inflammation and necrosis. Ethanol increased the expression of CD14 and tumor necrosis factor- (TNF) alpha mRNA and also the production of TNF-alpha by isolated Kupffer cells, but toremifene had no significant counteracting effect. However, toremifene significantly alleviated both ethanol induction of the pro-oxidant enzyme CYP2E1 and ethanol reduction of the oxidant-protective enzyme Se-glutathione peroxidase.nnnCONCLUSIONSnThe partial protection by toremifene against ethanol-induced liver lesions suggests a pathogenic contribution of estrogens, possibly associated with an oxygen radical mediated mechanism.

Alcoholic liver disease in rats fed ethanol as part of oral or intragastric low-carbohydrate liquid diets

The intragastric administration of ethanol as part of a lowcarbohydrate diet results in alcohol hepatotoxicity. We aimed to investigate whether comparable liver injury can be achieved by oral diet intake. Male Sprague-Dawley rats were fed ethanol as part of low-carbohydrate diets for 36–42 days either intragastrically or orally. Liver pathology, blood ethanol concentration, serum alanine amino transferase (ALT), endotoxin level, hepatic CYP2E1 induction, and cytokine profiles were assessed. Both oral and intragastric low-carbohydrate ethanol diets resulted in marked steatosis with additional inflammation and necrosis accompanied by significantly increased serum ALT, high levels of CYP2E1 expression, and production of auto-antibodies against malondialdehyde and hydroxyethyl free radical protein adducts. However, cytokine profiles differed substantially between the groups, with significantly lower mRNA expression of the anti-inflammatory cytokine interleukin 4 observed in rats fed low-carbohydrate diets orally. Inflammation and necrosis were significantly greater in rats receiving low-carbohydrate alcohol diets intragastrically than orally. This was associated with a significant increase in liver tumor necrosis factor α and interleukin 1β gene expression in the intragastric model. Thus, oral low-carbohydrate diets produce more ethanol-induced liver pathology than oral high-carbohydrate diets, but hepatotoxicity is more severe when a low-carbohydrate diet plus ethanol is infused intragastrically and is accompanied by significant increases in levels of proinflammatory cytokines.

Acetaldehyde levels during ethanol oxidation: A diet-induced change and its relation to liver aldehyde dehydrogenases and redox states

Abstract 3 different diets that had previously been observed to cause large differences in blood acetaldehyde levels of rats administered ethanol were compared with respect to their influence on liver enzymes metabolizing alcohol, on ethanol elimination and on the ethanol-induced changes in the hepatic content of metabolites that reflect the cytosolic or the mitochondrial redox state of the nicotine-amide dinucleotide couple. The results demonstrate that an unknown dietary factor affects the activity of liver aldehyde dehydrogenase, especially that of the low-K m enzyme. It is suggested that these enzyme activity changes are reflected in the observed alterations in acetaldehyde levels, which in turn may be associated with the magnitude of the shift in the mitochondrial redox state during ethanol oxidation.