Evaluation of a role of acetaldehyde in the mechanism of inhibition of p-nitroanisole O-demethylation in isolated hepatocytes by ethanol

The rate of p-nitroanisole O-demethylation is markedly inhibited by ethanol. To evaluate a role of acetaldehyde in the inhibition by ethanol, a comparison was made of the effects of ethanol and acetaldehyde on the metabolism of p-nitroanisole by isolated liver cells. No effect on the metabolism of p-nitroanisole was found at low concentrations of acetaldehyde (<0.5 mm), whereas inhibition occurred at high concentrations (1 mm). In fact, acetaldehyde was not any more inhibitory than crotonaldehyde, which is a poor substrate for the low-K_m mitochondrial aldehyde dehydrogenase. Cyanamide, an inhibitor of acetaldehyde oxidation, did not prevent the inhibition by ethanol. Crotonol, an alcohol that does not change the mitochondrial redox state, in contrast to ethanol, proved to be a more effective inhibitor of the metabolism of p-nitroanisole than ethanol. Greater sensitivity to crotonol was also found in isolated microsomes and may reflect hydrophobic effects by crotonol, relative to ethanol. These results suggest that although high levels of acetaldehyde can be inhibitory, physiological levels of acetaldehyde did not affect the metabolism of p-nitroanisole. It is unlikely that acetaldehyde itself plays a major role in the mechanism by which ethanol inhibits the metabolism of p-nitroanisole. The inhibition of p-nitroanisole O-demethylation by ethanol was prevented by pyruvate or fructose, but not by xylitol, sorbitol, or lactate. All these substrates by themselves stimulated metabolism of p-nitroanisole. Pyruvate and glyceraldehyde (which arises from the metabolism of fructose) can oxidize cytosolic NADH. These results suggest that the generation of cytosolic NADH from the oxidation of ethanol, the subsequent requirement for substrate shuttles to transfer NADH into the mitochondria, and redox inhibition of the citric acid cycle, interfere with the transport of NADPH out of the mitochondria, and consequently with drug metabolism.     

Effects of ethanol-derived acetaldehyde on the phosphorylation potential and on the intramitochondrial redox state in intact rat liver

The role of acetaldehyde (AcH) in the ethanol-induced shift toward reduction of the cytosolic and mitochondrial free NAD⁺/free NADH ratios and its effect on the phosphorylation potential was investigated in livers of fed, intact rats given ethanol (1 g/kg ip). Calcium cyanamide, an inhibitor of mitochondrial aldehyde dehydrogenase, was administered to block predominantly intramitochondrial NADH production from AcH oxidation. Compared with ethanol alone, cyanamide almost totally reversed the elevation of the β-OH-butyrate/acetoacetate ratio but only slightly reduced the lactate/ pyruvate ratio, which was calculated to be in near equilibrium with the hepatic ethanol/ AcH ratio after cyanamide. Ethanol or cyanamide alone had no effect on ATP, ADP, or P_i, but together they significantly decreased the $\text{ATP}\text{ADP}\text{· P}{}_{\mathrm{<mtext>i</mtext>}}$ ratio by increasing both ADP and P_i levels. No association between changes in the phosphorylation potential and the redox states was, however, observed. An ethanol-induced increase in AMP was abolished by cyanamide. The results demonstrate that the effect of ethanol on the mitochondrial redox state requires active AcH oxidation and suggest that moderate AcH accumulation likely to occur during alcohol-aversive drug treatment significantly lowers the cellular phosphorylation potential.     

Amidino-substituted aromatic heterocycles as probes of the specificity pocket of trypsin-like proteases.

The effect of pargyline on the uptake of acetaldehyde (in the presence of pyrazole) by isolated rat liver cells was studied after incubating the liver cells for 0, 10, 30, 45, and 60 min with 0.40, 1.30, and 2.6 mm pargyline. Without any incubation period, pargyline had no effect on acetaldehyde uptake. With increasing time of incubation, there was a progressive increase in the extent of inhibition of acetaldehyde uptake by pargyline. This suggests the possibility that pargyline is metabolized to the effective inhibitor or the incubation period allows pargyline to reach its site(s) of action. Pargyline was also a more effective inhibitor of the uptake of lower concentrations of acetaldehyde, e.g., 0.167 mm, than of higher concentrations (1.0 mm) of acetaldehyde, especially after short incubation periods or when pyrazole was omitted from the reaction medium. After a 20- to 30-min incubation period, pargyline inhibited the control rate of ethanol oxidation by the liver cells, as well as the accelerated rate of ethanol oxidation found in the presence of pyruvate or an uncoupling agent. Pargyline had no effect on hepatic oxygen consumption. During ethanol oxidation, a time-dependent release of acetaldehyde into the medium was observed. Pyruvate, by increasing the rate of ethanol oxidation, increased the output of acetaldehyde five- to tenfold. Pargyline increased the output of acetaldehyde two- to threefold, despite decreasing the rate of ethanol metabolism by the liver cells. These data indicate that pargyline inhibits the low K_m aldehyde dehydrogenase in intact rat liver cells and that this enzyme plays the major role in oxidizing the acetaldehyde which arises during the metabolism of ethanol. Although most of the acetaldehyde generated during the oxidation of ethanol is removed by the liver cells in an effective manner, changes in the activity of aldehyde dehydrogenase or the rate of acetaldehyde generation significantly alter the hepatic output of acetaldehyde.     

Whole-cell fluorescence for observing reductive metabolism from carbon compounds and hydrogen inAzotobacter vinelandii

The change in fluorescence of intactAzotobacter vinelandii was observed to study the oxidation and reduction of flavin and pyridine nucleotides resulting from carbon and hydrogen metabolism. Metronidazole, acetaldehyde, and oxygen each oxidized flavin. Flavin oxidized by metronidazole or acetaldehyde was reduced by addition of mannitol or ethanol, but not by acetate or hydrogen. The fluorescence induced by oxygen was transient. Mannitol, ethanol, acetate, acetaldehyde, and hydrogen shortened the duration of the oxygen-dependent fluorescence and supported respiration. The changes in redox state of pyridine nucleotides corresponded to the changes in flavin redox state. This indicates that the use of reducing equivalents from uptake hydrogenase is limited to the respiratory electron transport system inAzotobacter vinelandii.     

Inhibition of the oxidation of acetaldehyde and formaldehyde by hepatocytes and mitochondria by crotonaldehyde

Crotonaldehyde was oxidized by disrupted rat liver mitochondrial fractions or by intact mitochondria at rates that were only 10 to 15% that of acetaldehyde. Although a poor substrate for oxidation, crotonaldehyde is an effective inhibitor of the oxidation of acetaldehyde by mitochondrial aldehyde dehydrogenase, by intact mitochondria, and by isolated hepatocytes. Inhibition by crotonaldehyde was competitive with respect to acetaldehyde, and the Ki for crotonaldehyde was about 5 to 20 microM. Crotonaldehyde had no effect on the oxidation of glutamate or succinate. Very low levels of acetaldehyde were detected during the metabolism of ethanol. Crotonaldehyde increased the accumulation of acetaldehyde more than 10-fold, indicating that crotonaldehyde, besides inhibiting the oxidation of added acetaldehyde, also inhibited the oxidation of acetaldehyde generated by the metabolism of ethanol. Formaldehyde was a substrate for the low-Km mitochondrial aldehyde dehydrogenase, as well as for a cytosolic, glutathione-dependent formaldehyde dehydrogenase. Crotonaldehyde was a potent inhibitor of mitochondrial oxidation of formaldehyde, but had no effect on the activity of formaldehyde dehydrogenase. In hepatocytes, crotonaldehyde produced about 30 to 40% inhibition of formaldehyde oxidation, which was similar to the inhibition produced by cyanamide. This suggested that part of the formaldehyde oxidation occurred via the mitochondrial aldehyde dehydrogenase, and part via formaldehyde dehydrogenase. The fact that inhibition by crotonaldehyde is competitive may be of value since other commonly used inhibitors of aldehyde dehydrogenase are irreversible inhibitors of the enzyme.     

Effect of chronic ethanol or acetaldehyde on hepatic alcohol and aldehyde dehydrogenases, aminotransferases and glutamate dehydrogenase

Ethanol or acetaldehyde orally administered (15% and 2% respectively in drinking water) to male Wistar rats for three months induced alterations in the main liver enzymes responsible for ethanol metabolism, aspartate and alanine aminotransferases and NAD glutamate dehydrogenase. Ethanol produced a significant decrease in the activity of soluble alcohol dehydrogenase, while acetaldehyde induced alterations both in soluble and mitochondrial aldehyde dehydrogenases: soluble activity was significantly higher than in the control and ethanol-treated groups, and mitochondrial activity was significantly diminished. Both soluble aspartate and alanine aminotransferases showed pronounced increases by the chronic effect of acetaldehyde, while mitochondrial activities were practically unchanged by the effect of ethanol or acetaldehyde. Mitochondrial NAD glutamate dehydrogenase showed a rise in its activity both by the effect of chronic ethanol and acetaldehyde consumption. The level of metabolites assayed in liver extracts showed marked differences between ethanol and acetaldehyde treatment which indicates that ethanol produced a remarkable increase in glutamate, aspartate and free ammonia together with marked decrease in pyruvate and 2-oxoglutarate concentrations. Acetaldehyde consumption induced a significant decrease in 2-oxoglutarate and pyruvate concentrations. These observations suggest that ethanol has an important effect on the urea cycle enzymes, while the effect of acetaldehyde contributes to the impairment of the citric acid cycle.     

The metabolic activation of cyanamide to an inhibitor of aldehyde dehydrogenase is catalyzed by catalase

The inhibition of aldehyde dehydrogenase by cyanamide is dependent on an enzyme catalyzed conversion of the latter to an active metabolite. The following results suggest that catalase is the enzyme responsible for this bioactivation. The elevation of blood acetaldehyde elicited by cyanamide after ethanol administration to rats was attenuated more than 90 percent by pretreatment with the catalase inhibitor, 3-amino-1,2,4-triazole. This attenuation was dose dependent and was accompanied by a reduction in total hepatic catalase activity. Although hepatic catalase was also inhibited by cyanamide, a positive correlation between blood acetaldehyde and hepatic catalase activity was observed. In vitro, the activation inhibitor, 3-amino-1,2,4-triazole. This attenuation was dose dependent and was accompanied by a reduction in total hepatic catalase activity. Although hepatic catalase was also inhibited by cyanamide, a positive correlation between blood acetaldehyde and hepatic catalase activity was observed. In vitro, the activation of cyanamide was catalyzed by a) the rat liver mitochondrial subcellular fraction, b) the 50-65% ammonium sulfate mitochondrial fraction and c) purified bovine liver catalase. Cyanamide activation was inhibited by sodium azide. Since much of the hepatic catalase is localized in the peroxisomes and since peroxisomes and mitochondria cosediment, the cyanamide activating enzyme, catalase, is likely of peroxisomal and mitochondrial origin.     

THE EFFECTS OF INTOXICATING DOSES OF ETHANOL UPON INTERMEDIARY METABOLISM IN RAT BRAIN

Abstract— The effect of acute (8-min) and prolonged (13-h) exposures to high doses of ethanol upon the intermediary metabolites of rat brain has been studied, with the use of a new freezing technique which minimizes post-mortem changes. Injection of ethanol (80 mmol/kg body wt) produced general anaesthesia within 8 min after administration. At this time there were increases in the brain contents of glucose, glucose-6-phosphate and citrate; there was no change in arterial pCO₂. Rats under ethanol anaesthesia for 13 h showed increases in brain contents of glycogen, glucose and glucose 6-phosphate; and decreases in lactate, pyruvate, α-oxoglutarate and malate. Under similar experimental conditions, arterial pCO₂, increased from 37 to 51 Torr. The changes in levels of metabolites after injection of ethanol were similar to those after administration of many volatile anaesthetic agents or elevation of brain CO₂ by other means. Although brain levels of malate and α-oxoglutarate decreased after prolonged exposure to ethanol, the mitochondrial redox state was maintained. Accordingly, the levels of glutamate and aspartate fell in accordance with the law of mass action. The maintenance of the cytoplasmic and mitochondrial redox states in the brain during ethanol intoxication was in marked contrast to the effects on the liver. We suggest that the different effects observed in brain and liver result from the action of ethanol upon the nerve cell membrane in brain, whereas the primary target in liver is alcohol dehydrogenase.     

Alcohol dehydrogenase activity in the uremic rat

An increase in liver alcohol dehydrogenase activity was detected after 6 days of uremia in the rat, with a maximum increase occurring after 10 days. The increase was documented in the oxidative and reductive directions with ethanol and acetaldehyde as substrates, respectively; was not found in organs other than the liver and was not accompanied by increases in other cytosolic enzymes. The enzyme in the uremic animal did not differ in pH optimum for ethanol oxidation, K_m for ethanol and NAD⁺ or in electrophoretic mobility from the enzyme in the normal animal. Adrenalectomy prevented the increase. It is suggested that the increase in liver alcohol dehydrogenase requires intact adrenal glands and is most likely caused by the stress of surgery and the uremic state.     

Hypophysectomy does not alter the acinar zonation of gluconeogenesis or the mitochondrial redox state in rat liver.

The biochemical and functional heterogeneity of hepatocytes in different zones of the liver acinus may be related to the concentrations of hormones within the liver acinus. We examined the effects of hypophysectomy, which causes marked changes in plasma hormone levels and in activities of hepatic enzymes that are normally heterogeneously distributed, on the degree of metabolic zonation within the liver acinus. In hypophysectomized rats the activity of alanine aminotransferase was increased, but its normal zonation (predominance in the periportal zone) was preserved. The activity in cultured periportal and perivenous hepatocytes was increased by dexamethasone, but not by glucagon. Periportal hepatocytes from hypophysectomized rats expressed higher rates of gluconeogenesis in culture than did perivenous hepatocytes, irrespective of the absence or presence of dexamethasone, glucagon or insulin. Similar differences in rates of ketogenesis and in the mitochondrial redox state in response to glucagon were observed between periportal and perivenous hepatocytes from hypophysectomized rats as between cell populations from normal rats. Although hypophysectomy causes marked changes in hepatic enzyme activities, it does not alter the degree of zonation of alanine aminotransferase, gluconeogenesis or the mitochondrial redox state within the liver acinus.     

Ethanol metabolism in guinea pig: In vivo ethanol elimination,alcohol dehydrogenase distribution,and subcellular localization of acetaldehyde dehydrogenase in liver

Guinea pig ethanol metabolism as well as distribution and activities of ethanol metabolizing enzymes were studied. Alcohol dehydrogenase (ADH; EC 1.1.1.1) is almost exclusively present in liver except for minor activities in the cecum. All other organ tissues tested (skeletal muscle, heart, brain, stomach, and testes) contained only negligible enzyme activities. In fed livers, ADH could only be demonstrated in the cytosolic fraction (2.94 μmol/g liver/min at 38 °C) and its apparent K_m value of 0.42 mm for ethanol as substrate is similar to the average K_m of the human enzymes. Acetaldehyde dehydrogenase (ALDH; EC 1.2.1.3) of guinea pig liver was measured at low (0.05 mm) and high (10 mm) acetaldehyde concentrations and its subcellular localization was found to be mainly mitochondrial. The total acetaldehyde activity in liver amounts to 3.56 μmol/g/ min. Fed and fasted animals showed similar zero-order alcohol elimination rates after intraperitoneal injection of 1.7 or 3.0 g ethanol/kg body wt. The ethanol elimination rate of fed animals after 1.7 g ethanol/kg body wt (2.59 μmol/g liver/min) was inhibited by 80% after intraperitoneal injection of 4-methylpyrazole. Average ethanol elimination rates in vivo after 1.7 g/kg ethanol commanded only 88% of the totally available ADH activity in fed guinea pig livers. Catalase (EC 1.11.1.6), an enzyme previously implicated in ethanol metabolism, is of 3.4-fold higher activity in guinea pig (10,400 U/g liver) than in rat livers (3,100 U/g liver), but 98% inhibition by 3-amino-1,2,4-triazole did not significantly alter ethanol elimination rates. After ethanol injection, fed and fasted guinea pigs reacted with prolonged hyperglycemia.     

Alcohol induced hepatic fibrosis: role of acetaldehyde

Alcohol abuse is one of the major causes of liver fibrosis worldwide. Although the pathogenesis of liver fibrosis is a very complex phenomenon involving different molecular and biological mechanisms, several lines of evidence established that the first ethanol metabolite, acetaldehyde, plays a key role in the onset and maintenance of the fibrogenetic process. This review briefly summarizes the molecular mechanisms underlying acetaldehyde pro-fibrogenic effects. Liver fibrosis represents a general wound-healing response to a variety of insults. Although mortality due to alcohol abuse has been constantly decreasing in the past 20 years in Southern Europe and North America, in several Eastern-European countries and Great Britain Alcoholic Liver Disease (ALD) shows a sharply increasing trend [Bosetti, C., Levi, F., Lucchini, F., Zatonski, W.A., Negri, E., La, V.C., 2007. Worldwide mortality from cirrhosis: an update to 2002. J. Hepatol. 46, 827-839]. ALD has a complex pathogenesis, in which acetaldehyde (AcCHO), the major ethanol metabolite, plays a central role. Ethanol is mainly metabolized in the liver by two oxidative pathways. In the first one ethanol is oxidized to acetaldehyde by the cytoplasmic alcohol dehydrogenase enzyme (ADH), acetaldehyde is then oxidized to acetic acid by the mitochondrial acetaldehyde dehydrogenase (ALDH). The second pathway is inducible and involves the microsomal ethanol-oxidizing system (MEOS), in which the oxidation of ethanol to acetaldehyde and acetic acid also leads to generation of reactive oxygen species (ROS). Chronic ethanol consumption significantly inhibits mitochondrial ALDH activity while the rate of ethanol oxidation to acetaldehyde is even enhanced, resulting in a striking increase of tissue and plasma acetaldehyde levels [Lieber, C.S., 1997. Ethanol metabolism, cirrhosis and alcoholism. Clin. Chim. Acta 257, 59-84]. This review will focus on the molecular mechanisms by which acetaldehyde promote liver fibrosis.     

Stabilization of superoxide dismutase by acetyl-l-carnitine in human brain endothelium during alcohol exposure: novel protective approach

Oxidative damage of the endothelium disrupts the integrity of the blood-brain barrier (BBB). We have shown before that alcohol exposure increases the levels of reactive oxygen species (ROS; superoxide and hydroxyl radical) and nitric oxide (NO) in brain endothelial cells by activating NADPH oxidase and inducible nitric oxide synthase. We hypothesize that impairment of antioxidant systems, such as a reduction in catalase and superoxide dismutase (SOD) activity, by ethanol exposure may elevate the levels of ROS/NO in endothelium, resulting in BBB damage. This study examines whether stabilization of antioxidant enzyme activity results in suppression of ROS levels by anti-inflammatory agents. To address this idea, we determined the effects of ethanol on the kinetic profile of SOD and catalase activity and ROS/NO generation in primary human brain endothelial cells (hBECs). We observed an enhanced production of ROS and NO levels due to the metabolism of ethanol in hBECs. Similar increases were found after exposure of hBECs to acetaldehyde, the major metabolite of ethanol. Ethanol simultaneously augmented ROS generation and the activity of antioxidative enzymes. SOD activity was increased for a much longer period of time than catalase activity. A decline in SOD activity and protein levels preceded elevation of oxidant levels. SOD stabilization by the antioxidant and mitochondria-protecting agent acetyl-L-carnitine (ALC) and the anti-inflammatory agent rosiglitazone suppressed ROS levels, with a marginal increase in NO levels. Mitochondrial membrane protein damage and decreased membrane potential after ethanol exposure indicated mitochondrial injury. These changes were prevented by ALC. Our findings suggest the counteracting mechanisms of oxidants and antioxidants during alcohol-induced oxidative stress at the BBB. The presence of enzymatic stabilizers favors the ROS-neutralizing antioxidant redox of the BBB, suggesting an underlying protective mechanism of NO for brain vascular tone and vasodilation.     

Short-term vitamin A supplementation at therapeutic doses induces a pro-oxidative state in the hepatic environment and facilitates calcium-ion-induced oxidative stress in rat liver mitochondria independently from permeability transition pore formation

There is a growing body of evidence showing that vitamin A induces toxic effects in several experimental models and in human beings. In the present work, we have investigated the effects of short-term vitamin A supplementation on the adult rat liver redox status. We have found that vitamin A at therapeutic doses induces a hepatic oxidative insult. Furthermore, we have observed increased antioxidant enzyme activity in the liver of vitamin-A-treated rats. Additionally, some mitochondrial dysfunction was found since superoxide anion production was increased in vitamin-A-treated rat liver submitochondrial particles, which may be the result of impaired mitochondrial electron transfer chain activity, as assessed here. We have also isolated rat liver mitochondria and challenged it with 75 μM CaCl₂, a non-oxidant agent that is able to induce mitochondrial oxidative stress indirectly. We have found that mitochondria isolated from vitamin-A-treated rat liver are more sensitive to CaCl₂ than control mitochondria regarding the redox status. Importantly, vitamin A seems to alter mitochondrial redox status independently of the participation of the mitochondrial permeability transition pore, which is activated by Ca²⁺ ions since cyclosporin A did not prevent the oxidative insult elicited by Ca²⁺ addition. Overall, we show here that mitochondria are a target of vitamin-A-associated toxicity also in vivo.     

Subcellular localization of the F1 and F2 isozymes of horse liver aldehyde dehydrogenase

The subcellular distribution and relative amounts of the two isozymes, F1 and F2, of aldehyde dehydrogenase (EC 1.2.1.3) which were recently purified to homogeneity from horse liver (Eckfeldt, J., et al. (1976) J. Biol. Chem.251, 236–240) have been investigated. A fresh horse liver homogenate was fractionated on DEAE-cellulose. The results indicate that approximately 60% of the total pH 7.0 acetaldehyde dehydrogenase activity is due to the F1 isozyme and 40% is due to the F2 isozyme. Several horse livers were then fractionated into subcellular components using a differential centrifugation method. Based on the disulfiram (Antabuse) inhibition and the aldehyde concentration dependence of the enzymatic activity, it appears that the disulfiram-sensitive F1 isozyme (K_m acetaldehyde ? 70 μm) is primarily cytosolic and the disulfiram-insensitive F2 isozyme (K_{m acetaldehyde} ? 0.2 μm) is primarily mitochondrial. Fluorescence studies showed that the acetaldehyde dehydrogenase of the intact mitochondria could utilize only the endogenous pyridine nucleotide pool and not externally added NAD. Also, the ethanol dehydrogenase activity was found to be nearly 10 times the total acetaldehyde dehydrogenase activity when assaying a horse liver homogenate at pH 7.0 and with saturating substrates. The significant differences between this work and the results reported in rat liver are discussed with respect to the physiological importance of the cytosolic and mitochondrial aldehyde dehydrogenase during the ethanol oxidation in vivo.     

The effect of methylene blue on the hepatocellular redox state and liver lipid content during chronic ethanol feeding in the rat.

Feeding of ethanol in a liquid diet to male Wistar rats caused decreases in the hepatic cytosolic and mitochondrial [NAD+]/[NADH] ratios. This redox-state change was attenuated after 16 days of feeding ethanol as 36% of the total energy intake. Supplementation of the ethanol-containing liquid diet with Methylene Blue largely prevented the ethanol-induced redox state changes, but did not significantly decrease the severity of the hepatic lipid accumulation that resulted from ethanol ingestion. Methylene Blue did not affect body-weight gain, ethanol intake or serum ethanol concentrations in ethanol-fed rats, nor did the compound influence the hepatic redox state or liver lipid content of appropriate pair-fed control animals. These findings suggest that the altered hepatic redox state that results from ethanol oxidation is not primarily responsible for the production of fatty liver after long-term ethanol feeding in the rat.     

Intralobular distribution of rat liver aldehyde dehydrogenase and alcohol dehydrogenase

1. The activity of liver microsomal high Km-ALDH and mitochondrial low Km-ALDH, which may be primarily responsible for the oxidation of acetaldehyde after ethanol administration was found to be predominantly distributed in the centrilobular area. 2. The activities of other ALDH isozymes in mitochondrial and soluble fractions were evenly distributed in periportal and perivenous regions. 3. The activity of ADH which is involved in production of acetaldehyde was predominantly located in the periportal area. 4. From these results it seems unlikely that a concentration of acetaldehyde after ethanol ingestion is higher in perivenous hepatocytes than in periportal ones. Additional data would be needed to understand fully the mechanism by which ethanol induces predominantly centrilobular liver injury.     

Effect of alcohol intoxication and aldehyde dehydrogenase inhibitors on lipid peroxidation in rat liver

A single intraperitoneal administration of ethanol (3.5 g/kg) to rats induced a marked increase in lipid peroxidation and a decrease of antioxidative activity in the liver after 1 h when assessed by chemi-luminescence in liver homogenates. The pretreatment with aldehyde dehydrogenase inhibitor, disulfiram (200 mg/kg 24 hr before ethanol), caused a 10-fold elevation of the blood acetaldehyde levels, with no effect on the hepatic lipid peroxidation compared to control. Cyanamide (50 mg/kg, 2 h before the ethanol) increased approximately 100-fold the acetaldehyde levels, however, the changes in lipid peroxidation were not significantly different from that produced by ethanol alone. The present results suggest, that the metabolism of acetaldehyde and not acetaldehyde itself is responsible for the in vivo activation of lipid peroxidation during acute alcohol intoxication. Disulfiram prevents the ethanol-induced lipid peroxidation in the rat liver.     

Dynamic Adaptation of Liver Mitochondria to Chronic Alcohol Feeding in Mice: BIOGENESIS,REMODELING, AND FUNCTIONAL ALTERATIONS*

Liver mitochondria undergo dynamic alterations following chronic alcohol feeding to mice. Intragastric alcohol feeding to mice resulted in 1) increased state III respiration (109% compared with control) in isolated liver mitochondria, probably due to increased levels of complexes I, IV, and V being incorporated into the respiratory chain; 2) increased mitochondrial NAD⁺ and NADH levels (∼2-fold), with no change in the redox status; 3) alteration in mitochondrial morphology, with increased numbers of elongated mitochondria; and 4) enhanced mitochondrial biogenesis in the liver, which corresponded with an up-regulation of PGC-1α (peroxisome proliferator-activated receptor γ coactivator-1α). Oral alcohol feeding to mice, which is associated with less liver injury and steatosis, slightly enhanced respiration in isolated liver mitochondria (30.8% compared with control), lower than the striking increase caused by intragastric alcohol feeding. Mitochondrial respiration increased with both oral and intragastric alcohol feeding despite extensive N-acetylation of mitochondrial proteins. The alcohol-induced mitochondrial alterations are probably an adaptive response to enhance alcohol metabolism in the liver. Isolated liver mitochondria from alcohol-treated mice had a greater rate of acetaldehyde metabolism and respiration when treated with acetaldehyde than control. Aldehyde dehydrogenase-2 levels were unaltered in response to alcohol, suggesting that the greater acetaldehyde metabolism by isolated mitochondria from alcohol-treated mice was due to increased mitochondrial respiration that regenerated NAD⁺, the rate-limiting substrate in alcohol/acetaldehyde metabolism. Overall, our work suggests that mitochondrial plasticity in the liver may be an important adaptive response to the metabolic stress caused by alcohol intake and could potentially play a role in many other vital functions performed by the liver.     

Activities of hepatic enzymes related to ethanol oxidation and effects of ethanol on hepatic metabolites in rats with chronic liver injury.

To study the effects of ethanol on liver chronically injured by CCl4, activities of hepatic enzymes related to ethanol oxidation, influences of ethanol on hepatic metabolites, and blood ethanol disappearance were observed. (1) Activities of alcohol dehydrogenase, low- and high-Km aldehyde dehydrogenase, microsomal ethanol-oxidizing system and drug-metabolizing enzyme were remarkably decreased in the injured liver. (2) Increases in lactate/pyruvate and beta-hydroxybutyrate/acetacetate ratios were shown in control liver 2 h after ethanol ingestion. Similar but less pronounced effects of ethanol on the 'redox state' were also seen in rats with chronic liver injury. (3) Delay in ethanol disappearance was not observed until 12 h after ethanol ingestion. The ethanol-induced changes in the redox state in the injured liver were similar to those in controls. Higher ethanol concentrations in blood from rats with chronic liver injury could be related to potentiate the injured liver.