Comparative effects of ethanol, n-propranol and isopropanol on lipid disposal by rat liver.

Besides ethanol, other aliphatic alcohols such as n-propanol and isopropanol induce a triacylglycerol (TAG) accumulation in the liver. To determine whether a common mechanism is responsible for the effects of these three alcohols on hepatic lipid metabolism, each was administered by gastric tube to female Wistar rats at the dose of 50 mmol/kg body wt. Whichever alcohol was administered, the hepatic triacylglycerol accumulation was found to be related to the duration of elevated blood alcohol concentration. After administration of n-propanol or isopropanol, the liver [14C]palmitate uptake was increased whereas hepatic palmitate oxidation to 14CO2 was impaired and palmitate esterification into TAG enhanced; these perturbations were however more discrete than after ethanol administration. In contrast to ethanol and n-propanol which, at the dose presently used, increase precursor incorporation into blood TAG, isopropanol inhibits this incorporation. Interference with the process of very low density lipoprotein (VLDL) synthesis and/or secretion, which appears only at a late stage of isopropanol intoxication, is probably responsible for the intensity and duration of the fatty liver observed after administration of this alcohol.     

Action of isopropanol on rat lipid metabolism: complementary studies on mechanisms implicated in hepatic accumulation of triacylglycerides]

Isopropanol administration (3 g/kg, p.o.) determines in the rat liver an inhibition of fatty acid oxidation, an enhancement of fatty acid esterification into triacylglycerols as well as an inhibition of lipoprotein secretion which appears to be related to alterations in hepatic phospholipids and which differentiates isopropanol from ethanol induced fatty liver. Disturbances in peripheral lipolysis following isopropanol administration are found only in mature rats and are not necessary to the fatty liver induction.     

In vivo ethanol elimination in man, monkey and rat: a lack of relationship between the ethanol metabolism and the hepatic activities of alcohol and aldehyde dehydrogenases

The in vivo ethanol elimination in human subjects, monkeys and rats was investigated after an oral ethanol dosage. After 0.4 g. ethanol/kg of body weight, ethanol elimination was much slower in human subjects than in monkeys. In order to detect a rise in monkey plasma ethanol concentrations as early as observed in human subjects, ethanol had to be administered at a dose of 3 g/kg body weight. Ethanol metabolism in rats was also much faster than in human subjects. However, human liver showed higher alcohol dehydrogenase activity and higher low Km aldehyde dehydrogenase activity than rat liver. Thus, our data suggest a lack of relationship between hepatic ethanol-metabolizing activities and the in vivo ethanol elimination rate.     

The collagenolytic effects of the traditional Chinese medicine preparation,Han-Dan-Gan-Le,contribute to reversal of chemical-induced liver fibrosis in rats

Han-Dan-Gan-Le (HDGL), a Chinese herb preparation composed of Stephaniat tetrandra, Salvia miltorrhiza, Radix paeoniae, Astragalus membranaceus, and Ginkgo biloba, has been used to treat human liver fibrosis. This study was designed to examine the therapeutic effect of HDGL on chemical-induced liver fibrosis in adult Wistar rats. Liver fibrosis was produced in rats by carbon tetrachloride (1.2 ml CCl(4)/kg, 2 times/week, after an initial dose of 5.0 ml CCl(4)/kg, sc), plus a diet of 20% fat, 0.05% cholesterol (continuous) and 30% alcohol in the drinking water ad libitum (every other day) for 8 weeks. HDGL (0.5 and 1.0 g/kg, ig, daily for 6 weeks) was administered to rats 72 hrs after the last dose of CCl(4) to examine its therapeutic effects on chemical-induced liver fibrosis. Upon pathological examination, the HDGL treatment had significantly reversed chemical-induced liver fibrosis and other hepatic lesions. Hepatic collagen accumulation induced by CCl(4) was markedly reduced by HDGL treatment, as evidenced by hepatic collagen content and by immunohistochemical analysis of type-I collagen in liver. HDGL appeared to stimulate the collagenolytic process in the liver, as a 30-50% increase in urinary excretion of hydroxyproline was observed with HDGL treatment as compared to rats only given CCl(4). In conclusion, HDGL can effectively reverse chemically induced liver fibrosis, and this appears to be due, at least in part, to the stimulation of hepatic collagenolysis, resulting in a resolution of hepatic fibrosis.     

Regulation of ethanol metabolism in the rat

The purpose of these experiments was to examine the factors which regulate ethanol metabolism in vivo. Since the major pathway for ethanol removal requires flux through hepatic alcohol dehydrogenase, the activity of this enzyme was measured and found to be 2.9 mumol/(min X g liver). Ethanol disappearance was linear for over 120 min in vivo and the blood ethanol fell 0.1 mM/min; this is equivalent to removing 20 mumol ethanol/min and would require that flux through alcohol dehydrogenase be about 60% of its measured maximum velocity. To test whether ethanol metabolism was limited by the rate of removal of one of the end products (NADH) of alcohol dehydrogenase, fluoropyruvate was infused to reoxidize hepatic NADH and to prevent NADH generation via flux through pyruvate dehydrogenase. There was no change in the rate of ethanol clearance when fluoropyruvate was metabolized. Furthermore, enhancing endogenous hepatic NADH oxidation by increasing the rate of urea synthesis (converting ammonium bicarbonate to urea) did not augment the steady-state rate of ethanol oxidation. Hence, transport of cytoplasmic reducing power from NADH into the mitochondria was not rate limiting for ethanol oxidation. In contrast, ethanol oxidation at the earliest time periods could be augmented by increasing hepatic urea synthesis.     

Hepatic redox state: attenuation of the acute effects of ethanol induced by chronic ethanol consumption

The effects of an acute dose of a diet containing ethanol (3g/kg) on hepatic redox state was compared in rats fed ethanol for 25 days and in their littermates given isocaloric carbohydrate. In both groups, cytoplasmic and mitochondrial redox states of pyridine nucleotides shifted to a more reduced level, but the changes were much less extensive in rats chronically fed ethanol. This metabolic adaption may reflect the oxidation of ethanol by a pathway not involving alcohol dehydrogenase, such as the microsomal ethanol osidizing system, which increases in activity after chronic ethanol ingestion.     

Significant pathways of hepatic ethanol metabolism.

Rat liver microsomes oxidized ethanol two to three times faster than propanol when incubated with either an NADPH- or an H2O2-generating system. In addition, solubilized, purified microsomal subfractions were found to contain protein with an electrophoretic mobility identical to rat liver catalase on SDS polyacrylamide gels, suggesting that the separation of catalase from cytochrome P-450 and other microsomal components may not be feasible. These data support the postulate that catalase is responsible for NADPH-dependent microsomal ethanol oxidation. Direct read-out techniques for pyridine nucleotides, the catalase-H2O2 complex, and cytochrome P-450 were utilized to evaluate the specificity of inhibitors of alcohol dehydrogenase (4-methylpyrazole; 4 mM) and catalase (aminotriazole; 1.0 g/kg) qualitatively in perfused rat livers. 4-Methylpyrazole and aminotriazole are specific inhibitors for alcohol dehydrogenase and catalase, respectively, under these conditions. Neither inhibitor nor a combination of them altered the mixed function oxygen of p-nitroanisole to p-nitrophenol as observed by oxygen uptake and product formation. When ethanol utilization was measured over the concentration range 20-80 mM in perfused liver, a concentration dependence was observed. At low concentrations of ethanol, ethanol oxidation was almost totally abolished by 4-methylpyrazole; however, the contribution of 4-methylpyrazole-insensitive ethanol uptake increased as a function of ethanol concentration. At 80 mM ethanol, ethanol utilization was nearly 50% methylpyrazole-insensitive. This portion of ethanol oxidation, however, was abolished by aminotriazole. The data indicate that alcohol dehydrogenase and catalase-H2O2 are responsible for hepatic ethanol oxidation. At low ethanol concentrations (less than 20 mM), alcohol dehydrogenase is predominant; however, at higher ethanol concentrations (up to 80 mM), the contribution of catalase-H2O2 to overall ethanol utilization is significant. No evidence that the endoplasmic reticulum is involved in ethanol metabolism in the perfused liver emerged from these studies.     

β-Lapachone alleviates alcoholic fatty liver disease in rats

Alcohol-induced liver injury is the most common liver disease in which fatty acid metabolism is altered. It is thought that altered NAD⁺/NADH redox potential by alcohol in the liver causes fatty liver by inhibiting fatty acid oxidation and the activity of tricarboxylic acid cycle reactions. β-Lapachone (βL), a naturally occurring quinone, has been shown to stimulate fatty acid oxidation in an obese mouse model by activating adenosine monophosphate-activated protein kinase (AMPK). In this report, we clearly show that βL reduced alcohol-induced hepatic steatosis and induced fatty acid oxidizing capacity in ethanol-fed rats. βL treatment markedly decreased hepatic lipids while serum levels of lipids and lipoproteins were increased in rats fed ethanol-containing liquid diets with βL administration. Furthermore, inhibition of lipolysis, enhancement of lipid mobilization to mitochondria and upregulation of mitochondrial β-oxidation activity in the soleus muscle were observed in ethanol/βL-treated animals compared to the ethanol-fed rats. In addition, the activity of alcohol dehydrogenase, but not aldehyde dehydrogenase, was significantly increased in rats fed βL diets. βL-mediated modulation of NAD⁺/NADH ratio led to the activation of AMPK signaling in these animals. Conclusion: Our results suggest that improvement of fatty liver by βL administration is mediated by the upregulation of apoB100 synthesis and lipid mobilization from the liver as well as the direct involvement of βL on NAD⁺/NADH ratio changes, resulting in the activation of AMPK signaling and PPARα-mediated β-oxidation. Therefore, βL-mediated alteration of NAD⁺/NADH redox potential may be of potential therapeutic benefit in the clinical setting.     

Deuterium isotope effects in the oxidation of alcohols in vitro and in vivo

The oxidation of ethanol and isopropanol by liver alcohol dehydrogenase was studied in vitro and in vivo. Oxidation of ethanol by horse liver alcohol dehydrogenase was carried out in the presence of lactaldehyde and other aldehydes which reoxidized enzyme-bound NADH. Under these conditions the oxidation of ethanol was accelerated 7 to 22-fold, depending on the nature of the aldehyde. (An acceleration of ethanol oxidation by lactaldehyde was previously reported by Gupta and Robinson [(1966) Biochim, Biophys. Acta118, 431]. In the presence of lactaldehyde ping-pong kinetics were observed and a deuterium isotope effect on V of 4.2 was seen. In the absence of acceptor aldehyde no, or small, isotope effects (Baker, R. H. (1962) Biochemistry1, 41) are observed. Therefore, when dissociation of NADH is no longer rate limiting the hydrogen transfer step becomes largely rate determining. Oxidation of isopropanol shows an isotope effect on V of 2.5 in the absence of acceptor aldehyde. With mouse liver alcohol dehydrogenase results similar to those obtained with the horse liver enzyme were obtained.When ethanol metabolism was examined in vivo, in mice by measuring blood alcohol levels, no isotope effect was observed with ethanol-1-d₂. On the other hand, an isotope effect of 2.0 was observed when the metabolism of isopropanol and isopropanol-2-d₁ were compared. This isotope effect is very close to that observed in vitro with the mouse liver enzyme. The relative rate of metabolism of ethanol and isopropanol in vivo was similar to that observed in vitro with the mouse liver enzyme (ethanol:isopropanol, 2.1 in vivo:2.2 in vitro). It was concluded that in the metabolism of ethanol and isopropanol, alcohol dehydrogenase is partially rate determining. Administration to mice of lactaldehyde, as well as other aldehydes, ketones, or fructose, simultaneously with ethanol produced no increase in the rate of ethanol metabolism.     

Beneficial effect of nitric oxide synthase inhibitor on hepatotoxicity induced by allyl alcohol

The effect of aminoguanidine (a selective inhibitor of inducible nitric oxide synthase) on allyl alcohol-induced liver injury was assessed by the measurement of serum ALT and AST activities and histopathological examination. When aminoguanidine (50-300 mg/kg, i.p.) was administered to mice 30 min before a toxic dose of allyl alcohol (75 microL/kg, i.p.), significant changes related to liver injury were observed. In the presence of aminoguanidine the level of ALT and AST enzymes were significantly decreased. All symptoms of liver necrosis produced by allyl alcohol toxicity almost completely disappeared when animals were pretreated with aminoguanidine at 300 mg/kg. Depletion of hepatic glutathione as a consequence of allyl alcohol metabolism was minimal in mice pretreated with aminoguanidine at 300 mg/kg. It was found that the inhibition of toxicity was not due to alteration in allyl alcohol metabolism since aminoguanidine did not effect alcohol dehydrogenase activity both in vivo and in vitro.     

In vivo inhibition of liver alcohol dehydrogenase by ethanol administration

The activity of liver alcohol dehydrogenase (LADH) from rats sacrificed two hours after the administration of ethanol 3, 4 or 5 g/kg intraperitoneally was significantly inhibited compared to the activity of LADH from control rats. LADH activity was inversely related to the dose of ethanol administered, to the concentration of ethanol in the liver, and to the concentration of ethanol in the blood. The clearance of blood ethanol in rats was dose-dependent and was inversely related to the dose administered. The half-life of ethanol elimination increased as the dose of ethanol increased. These results suggest that ethanol-induced inhibition of LADH can occur in vivo and that the level of activity of this enzyme determines the rate of oxidation of ethanol.     

Hepatic Deficiency of Augmenter of Liver Regeneration Exacerbates Alcohol-Induced Liver Injury and Promotes Fibrosis in Mice

Why only a subpopulation (about 15%) of humans develops liver cirrhosis due to alcohol is a critical as yet unanswered question. Liver-specific depletion of augmenter of liver regeneration (ALR) protein in mice causes robust steatosis and hepatocyte apoptosis by 2 weeks; these pathologies regress subsequently with return of ALR expression even at lower than control levels, but the mice develop modest steatohepatitis by 8 weeks. We aimed to investigate whether chronic alcohol ingestion promotes excessive hepatic fibrosis in these ALR-deficient mice. Liver-specific ALR-deficient and wild type (WT) female mice (8–10 weeks old) were placed on 4% alcohol-supplemented or isocaloric diet for 4 weeks. Liver sections were examined for histopathology, and parameters of steatosis and fibrosis were quantified. The mRNA expression of alcohol dehydrogenase-1, acetaldehyde dehydrogenase-1 and cytochrome P450-2E1 increased in WT mice but decreased in ALR-deficient mice upon alcohol ingestion. While alcohol induced steatosis and mild inflammation in WT mice, ALR-deficient mice showed minimal steatosis, strong hepatocellular injury and inflammation, prominent ductular proliferation, and robust fibrosis. Compared to the WT mice, alcohol feeding of ALR-deficient mice resulted in significantly greater increase in hepatic TNFα and TGFβ, and oxidative stress; there was also hepatic iron accumulation, robust lipid peroxidation and mitochondrial DNA damage. Importantly, similar to ALR-deficient mice, lower hepatic ALR levels in human alcoholic liver cirrhosis were associated with increased iron content, reduced expression of alcohol dehydrogenase and acetaldehyde dehydrogenase, and elevated fibrogenic markers. We conclude that ALR deficiency or anomaly can play a critical role in alcohol-induced hepatic fibrosis/cirrhosis, mechanisms of which may involve dysregulation of alcohol metabolism and iron homeostasis, mitochondrial damage and oxidative injury.     

Retinol/ethanol drug interaction during acute alcohol intoxication in mice involves inhibition of retinol metabolism to retinoic acid by alcohol dehydrogenase

Substantial evidence indicates that one consequence of alcohol intoxication is a reduction in retinoic acid (RA) levels. Studies on the mechanism have shown that chronic ethanol consumption induces P450 enzymes that increase RA degradation, thus accounting for much but not all of the observed decrease in RA. A reduction in RA synthesis may also be involved as ethanol competitively inhibits retinol oxidation catalyzed by alcohol dehydrogenase (ADH) in vitro. This may be important during acute ethanol intoxication and may contribute to adverse retinol/ethanol drug interactions. Here we have examined mice for the effect of either acute ethanol intoxication or Adh1 gene disruption on RA synthesis and degradation. RA produced following a dose of retinol (50 mg/kg) was reduced 87% by pretreatment with an intoxicating dose of ethanol (3.5 g/kg). RA produced in Adh1-null mutant mice following a 50-mg/kg dose of retinol was reduced 82% relative to wild-type mice, thus similar to wild-type mice pretreated with ethanol. Reduced RA production was associated with increased retinol levels in both ethanol-treated wild-type mice and Adh1-null mutant mice, indicating reduced clearance of the retinol dose. RA degradation following a dose of RA (10 mg/kg) was increased only 42% by ethanol pretreatment (3.5 g/kg) and only 26% in Adh1-null mutant mice relative to wild-type mice. These findings demonstrate that the reduced RA levels observed during acute retinol/ethanol drug interaction are due primarily to a decrease in ADH-catalyzed RA synthesis and secondarily to an increase in RA degradation.     

Interleukin-22 Protects against Acute Alcohol-Induced Hepatotoxicity in Mice

The protective effects of interleukin-22 (IL-22) on acute alcohol-induced liver injury were investigated. Mice were gavaged with 7 doses of alcohol (56% wt/vol, 15.2 mL/kg of body weight for each dose) over the 24 h, and IL-22 (0.5 mg/kg BW) was given to the mice by injection into the tail vein 1 h after alcohol administration. The results indicated that acute alcohol administration caused prominent hepatic microvesicular steatosis and an elevation of serum transaminase activities, induced a significant decrease in hepatic glutathione in conjunction with enhanced lipid peroxidation, and increased hepatocyte apoptosis as well as hepatic TNF-alpha production. IL-22 treatment attenuated these adverse changes induced by acute alcohol administration. The protective effects of IL-22 on alcohol-induced hepatotoxicity were due mainly to its anti-inflammatory, anti-oxidant, and anti-apoptotic features.     

Interleukin-22 protects against acute alcohol-induced hepatotoxicity in mice

The protective effects of interleukin-22 (IL-22) on acute alcohol-induced liver injury were investigated. Mice were gavaged with 7 doses of alcohol (56% wt/vol, 15.2 mL/kg of body weight for each dose) over the 24 h, and IL-22 (0.5 mg/kg BW) was given to the mice by injection into the tail vein 1 h after alcohol administration. The results indicated that acute alcohol administration caused prominent hepatic microvesicular steatosis and an elevation of serum transaminase activities, induced a significant decrease in hepatic glutathione in conjunction with enhanced lipid peroxidation, and increased hepatocyte apoptosis as well as hepatic TNF-alpha production. IL-22 treatment attenuated these adverse changes induced by acute alcohol administration. The protective effects of IL-22 on alcohol-induced hepatotoxicity were due mainly to its anti-inflammatory, anti-oxidant, and anti-apoptotic features.     

A novel soluble beta-glucan salecan protects against acute alcohol-induced hepatotoxicity in mice

This investigation was designed to determine the effect of a novel soluble beta-glucan salecan on acute alcohol-induced hepatic injury in mice. Mice were given salecan (15 or 30 mg/kg) or PBS for 4 d. Ethanol (6 g/kg) was administered orally 1 h after the last injection. The animals were sacrificed at 10 h after alcohol administration. Pretreatment with salecan significantly ameliorated the hepatic damage induced by ethanol, as evidenced by markedly reduced serum aminotransferase activities and hepatocyte steatosis. Salecan administration remarkably alleviated the formation of thiobarbituric acid-reactive substances and counteracted glutathione depletion. The mRNA level of peroxisome proliferator activated receptor alpha, a major gene responsible for fatty acid oxidation, was significantly increased after salecan pretreatment. The expression of diacylglycerol acyltransferase 1, an important gene responsible for triacylglycerol synthesis, was markedly decreased after salecan was administrated. These findings suggest that salecan might represent a novel protective strategy against alcoholic liver injury.     

Assessment of hepatoprotective potential of Radix Fici Hirtae on alcohol-induced liver injury in Kunming mice

ObjectiveThe objective of the present study was to investigate the hepatoprotective role of Radix Fici Hirtae on acute alcohol-induced liver injury in mice.MethodsThe component of Radix Fici Hirtae was extracted using petroleum ether, chloroform, ethyl acetate and n-butanol and divided into three dose groups of high, medium and low according to the clinical man's normal dose of the 50 g crude drug/d (0.83 g/kg body weight). Saline in concentration of 10 mg/mL, 5 mg/mL and 2.5 mg/mL and a dose of mouse lavage (0.2 mL/10 g mouse body weight) were added to the solution. Histopathlogical analysis of liver was performed. Finally, liver protection was validated by examining the effect of aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (AKP), and lactate dehydrogenase (LDH) on the hepatic function of mice in alcohol-induced liver injury model.ResultsExcept for group with saturated n-butyl alcohol, for the rest of the groups, pathological changes of hepatic lipid and inflammatory cells infiltration were alleviated and liver sinus was normal. As compared to model group, the concentrations of AST, ALT, AKP and LDH in chloroform groups and ethyl acetate groups were significantly decreased.ConclusionsExtracts of Radix Fici Hirtae are effective for the prevention of alcohol-induced hepatic damage in mice. The results revealed that extracts of Radix Fici Hirtae could be used as hepatoprotective agent.     

System of ethanol and acetaldehyde metabolism in the rat liver during the development of ethanol tolerance

Following daily intraperitoneal injections of ethanol at a dose of 3.5 g/kg rats developed tolerance to its hypnotic action, which was manifested in a drastic decrease of ethanol--induced sleep time and the number of animals sleeping more than 60 min. In the liver, this process was characterized by an elevated alcohol dehydrogenase activity and a decreased aldehyde dehydrogenase one, whereas that of MEOS remained unchanged.     

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.     

A Novel Soluble Beta-Glucan Salecan Protects against Acute Alcohol-Induced Hepatotoxicity in Mice

This investigation was designed to determine the effect of a novel soluble beta-glucan salecan on acute alcohol-induced hepatic injury in mice. Mice were given salecan (15 or 30 mg/kg) or PBS for 4 d. Ethanol (6 g/kg) was administered orally 1 h after the last injection. The animals were sacrificed at 10 h after alcohol administration. Pretreatment with salecan significantly ameliorated the hepatic damage induced by ethanol, as evidenced by markedly reduced serum aminotransferase activities and hepatocyte steatosis. Salecan administration remarkably alleviated the formation of thiobarbituric acid-reactive substances and counteracted glutathione depletion. The mRNA level of peroxisome proliferator activated receptor alpha, a major gene responsible for fatty acid oxidation, was significantly increased after salecan pretreatment. The expression of diacylglycerol acyltransferase 1, an important gene responsible for triacylglycerol synthesis, was markedly decreased after salecan was administrated. These findings suggest that salecan might represent a novel protective strategy against alcoholic liver injury.