Development Of Ethanol-induced Fatty Liver Research Articles

The inhibitory effect of ethanol on Sestrin3 in the pathogenesis of ethanol-induced liver injury.

Sestrins (Sesns) are a family of stress-sensitive genes that have been suggested to regulate lipid metabolism. Chronic ethanol feeding is known to cause lipid accumulation in hepatocytes. This study was designed to investigate the role of Sesn3 in the pathogenesis of alcohol-induced hepatic steatosis. We demonstrated that ethanol inhibited the expression of Sesn3 in VL-17A cells. Overexpression of Sesn3 ameliorated triglyceride accumulation; downregulation using short hairpin RNA significantly deteriorated triglyceride accumulation in these cells. The expression of Sesn3 was also reduced in mice fed with ethanol for 4 wk. Overexpression of Sesn3 prevented hepatic steatosis, whereas knockdown of Sesn3 worsened hepatic steatosis in ethanol-fed mice. Overexpression of Sesn3 significantly reduced the expression of genes encoding for lipid synthesis through AMPK pathway. Overexpression of Sesn3 augmented the effect of ethanol on phospho-p70 S6 kinase. The levels of hepatic light chain 3, a marker for autophagy, expression were significantly decreased in ethanol-fed mice after Sesn3 gene was knocked down. Our findings suggest that inhibitory effect of ethanol on Sesn3 may play an important role in the development of ethanol-induced fatty liver.

Hepatic lipid profiling of deer mice fed ethanol using 1H and 31P NMR spectroscopy: A dose-dependent subchronic study

Chronic alcohol abuse is a 2nd major cause of liver disease resulting in significant morbidity and mortality. Alcoholic liver disease (ALD) is characterized by a wide spectrum of pathologies starting from fat accumulation (steatosis) in early reversible stage to inflammation with or without fibrosis and cirrhosis in later irreversible stages. Previously, we reported significant steatosis in the livers of hepatic alcohol dehydrogenase (ADH)-deficient (ADH−) vs. hepatic ADH-normal (ADH+) deer mice fed 4% ethanol daily for 2months [Bhopale et al., 2006, Alcohol 39, 179–188]. However, ADH− deer mice fed 4% ethanol also showed a significant mortality. Therefore, a dose-dependent study was conducted to understand the mechanism and identify lipid(s) involved in the development of ethanol-induced fatty liver. ADH− and ADH+ deer mice fed 1, 2 or 3.5% ethanol daily for 2months and fatty infiltration in the livers were evaluated by histology and by measuring dry weights of extracted lipids. Lipid metabolomic changes in extracted lipids were determined by proton (1H) and 31phosphorus (31P) nuclear magnetic resonance (NMR) spectroscopy. The NMR data was analyzed by hierarchical clustering (HC) and principle component analysis (PCA) for pattern recognition. Extensive vacuolization by histology and significantly increased dry weights of total lipids found only in the livers of ADH− deer mice fed 3.5% ethanol vs. pair-fed controls suggest a dose-dependent formation of fatty liver in ADH− deer mouse model. Analysis of NMR data of ADH− deer mice fed 3.5% ethanol vs. pair-fed controls shows increases for total cholesterol, esterified cholesterol, fatty acid methyl esters (FAMEs), triacylglycerides and unsaturation, and decreases for free cholesterol, phospholipids and allylic and diallylic protons. Certain classes of neutral lipids (cholesterol esters, fatty acyl chain (COCH2) and FAMEs) were also mildly increased in ADH− deer mice fed 1 or 2% ethanol. Only small increases were observed for allylic and diallylic protons, FAMEs and unsaturations in ADH+ deer mice fed 3.5% ethanol vs. pair-fed controls. PCA of NMR data showed increased clustering by gradual separation of ethanol-fed ADH− deer mice groups from their respective pair-fed control groups and corresponding ethanol-fed ADH+ deer mice groups. Our data indicate that dose of ethanol and hepatic ADH deficiency are two key factors involved in initiation and progression of alcoholic fatty liver disease. Further studies on characterization of individual lipid entities and associated metabolic pathways altered in our deer mouse model after different durations of ethanol feeding could be important to delineate mechanism(s) and identify potential biomarker candidate(s) of early stage ALD.

HIF-1α induction suppresses excessive lipid accumulation in alcoholic fatty liver in mice

Chronic alcohol intake stimulates hepatic oxygen consumption and subsequently causes liver hypoxia, leading to activation of hypoxia inducible factor-1 (HIF-1). Although HIF-1 plays a crucial role in the metabolic switch from aerobic to anaerobic metabolism in response to hypoxia, its roles in the regulation of lipid metabolism in alcoholic fatty liver remain unknown. Wild-type and hepatocyte-specific HIF-1α-null mice were subjected to a 6% ethanol-containing liquid diet for 4 weeks, and functional effects of loss of the HIF-1α gene on lipid metabolism were examined in the liver. Hepatocyte-specific HIF-1α-null mice developed severe hypertriglyceridemia with enhanced accumulation of lipids in the liver of mice exposed to a 6% ethanol-containing liquid diet for 4 weeks. Sterol regulatory element-binding protein 1c (SREBP-1c) and its downstream target acetyl-CoA carboxylase were greatly activated as the hepatic steatosis progressed, and these alterations were inversely correlated with the expression of the HIF-1-regulated gene DEC1. Overexpression of DEC1 in the mutant liver abrogated the detrimental effects of loss of HIF-1α gene on ethanol-induced fatty liver with reduced SREBP-1c expression. Conversely, co-administration of the HIF hydroxylase inhibitor dimethyloxalylglycine for the last 2 weeks improved markedly the ethanol-induced fatty liver in mice. The current results provide direct evidence for protective roles of HIF-1 induction in the development of ethanol-induced fatty liver via activation of the HIF-1-regulated transcriptional repressor DEC1.

Ethanol consumption impairs regulation of fatty acid metabolism by decreasing the activity of AMP-activated protein kinase in rat liver

The mechanisms by which ethanol consumption causes accumulation of hepatic triacylglycerols are complex. AMP-activated protein kinase (AMPK) plays a central role in the regulation of lipid metabolism. Therefore, in the present study we investigated whether AMPK may have a role in the development of ethanol-induced fatty liver. Hepatocytes isolated from rats fed with an ethanol-containing liquid diet showed higher rates of fatty acid and triacylglycerol syntheses, but a decreased rate of fatty acid oxidation, concomitant to a lower activity of carnitine palmitoyltransferase I. Hepatocytes from both ethanol-fed and pair-fed control rats were incubated with 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR), an AMPK activator in intact cells. In both hepatocyte preparations AICAR strongly inhibited the activity of acetyl-CoA carboxylase in parallel to fatty acid synthesis, but cells from ethanol-fed rats showed significantly lower sensitivity to inhibition by AICAR. Moreover, AICAR strongly decreased triacylglycerol synthesis and increased fatty acid oxidation in control hepatocytes, but these effects were markedly attenuated in hepatocytes from ethanol-fed rats. In parallel, AMPK in liver of ethanol-fed rats showed a decreased specific activity and a lower sensitivity to changes in the AMP/ATP ratio, compared to the enzyme of control rats. These effects are consistent with the impairment of AMPK-mediated regulation of fatty acid metabolism after ethanol consumption, that will facilitate triacylglycerol accumulation. Taken together, these findings suggest that a decreased AMPK activity may have an important role in the development of alcoholic fatty liver.

Differential Contributions of C3, C5, and Decay-Accelerating Factor to Ethanol-Induced Fatty Liver in Mice

The complement pathway is an important component of the innate and adaptive immune response. Here we tested the hypothesis that activation of complement is required for development of ethanol-induced fatty liver. Wild-type mice and mice lacking the third (C3) or fifth (C5) components of the complement activation pathway, as well as mice lacking decay-accelerating factor (CD55/DAF), a complement regulatory protein, were fed Lieber-DeCarli ethanol-containing diets for 6 weeks or pair-fed control diets. Ethanol feeding to wild-type mice increased C3a in plasma. Wild-type and C5-/- mice fed the ethanol diet developed hepatic steatosis characterized by microvesicular and macrovesicular lipid accumulation and increased triglyceride content. C3-/- mice did not develop steatosis, while CD55/DAF-/- mice accumulated even more hepatic triglyceride after ethanol feeding than wild-type mice. Levels of serum alanine aminotransferase and hepatic tumor necrosis factor alpha, indicators of hepatocyte injury and inflammation, respectively, were increased in wild-type and CD55/DAF-/- mice but not in C5-/- mice after ethanol feeding. In contrast to the protective effect of C3-/- against ethanol-induced steatosis, levels of both alanine aminotransferase and tumor necrosis factor alpha were increased in C3-/- mice after ethanol feeding. Here we have identified several elements of the complement system as important contributors to ethanol-induced fatty liver. C3 contributed primarily to the accumulation of triglyceride in the liver, whereas C5 was involved in inflammation and injury to hepatocytes. Further, the absence of CD55/DAF exacerbated these responses, suggesting that CD55/DAF serves as a barrier to ethanol-induced fatty liver.

Nutritional Aspects of Alcoholic Liver Disease

Development of ethanol-induced fatty liver, alcoholic hepatitis, and cirrhosis has been attributed in part to nutritional deficiencies for many years. Special attention must be focused on treating alcohol-induced liver disease while providing replacement of deficient amino acids, vitamins, minerals, and other nutrients. Avoidance of alcohol intake is required to eliminate progressive liver disease in alcoholics. This is best achieved by using educational and social programs to convince patients and their caretakers of the great necessity to eliminate alcohol intake.

Metabolic Studies on the Development of Ethanol-induced Fatty Liver in KK-Ay Mice

Mechanisms involved in the development of the alcoholic fatty liver in KK-Ay mice were investigated. Incorporation studies using [14C]acetate and [3H]palmitate indicated that the half-life of hepatic triglycerides was doubled in the ethanol-ingesting mice, and utilization of the exogenous fat was significantly increases as compared with that of the control. No persistent alteration was recognized in hepatic oxidation of palmitate, as estimated by in vitro experiments using liver slices obtained from control and ethanol-drinking mice. Enzymic studies indicated that the activities of acetyl COA carboxylase, ATP citrate lyase, malic enzyme, and 6-phosphogluconate dehydrogenase were increased with ethanol drinking. The increment in hepatic triglycerides accumulated during ethanol ingestion was largely accounted for by palmitoleic, oleic, and linoleic acids. These findings demonstrated an augmentation in hepatic lipogenesis as well as an increased utilization of exogenous fats. Ethanol drinking did not cause any appreciable change in plasma triglyceride level and metabolism of adipose tissue. In summary of the present studies, accelerated lipogenesis and increased utilization of the dietary fats may be possible causal factors in the alcoholic fatty liver of KK-Ay mice.

Effects of Nutritional Factors on the Development of Ethanol-induced Fatty Liver in KK and KK-Ay Mice

KK and KK-Ay mice developed a steady and reproducible fatty liver when they were given free access to an ethanol solution as a drinking fluid for 10 to 20 days. The present studies were undertaken to elucidate effects of nutritional factors on liver fat contents of the mice given water or ethanol solution. In contrast to cornstarch, sucrose tended to increase the liver fat of control mice. A higher concentration of dietary casein lowered the liver fat of control mice, whereas the dietary concentration of cottonseed oil did not significantly affect the liver rat levels either in control or ethanol groups. Thus the standard basal diets favorable for the development of the alcoholic fatty liver have been established, for example, 10% cottonseed oil, 25 and 30% casein, 58.4 and 53.4% cornstarch for KK (12-15 weeds old) and KK-Ay (5-10 weeks old), respectively. Neither choline, myoinositol, nor any lipotropic agent tested prevented the development of the alcoholic fatty liver. Unlike in rats, orotic acid did not induce a fatty liver but rather alleviated the ethanol-induced fatty liver in these mice.

Inhibition of the Acute Ethanol-Induced Fatty Liver by Pyrazole

SummarySince pyrazole specifically inhibits liver alcohol dehydrogenase and impairs the metabolism of ethanol, studies were undertaken in rats to evaluate the influence of inhibition of ethanol metabolism on the development of an acute ethanol-induced fatty liver. In agreement with previous observations, 20 hr after ethanol administration a 5-fold increase in hepatic triglyceride concentration occurred. In marked contrast, the pyrazole- and ethanol-treated group showed complete inhibition of ethanol-induced hepatic triglyceride accumulation. The prevention of the ethanol-induced hepatic triglyceride accumulation occurred in the presence of enhanced blood ethanol concentrations in the pyrazole group. The carbon tetrachloride-induced fatty liver and plasma hypotrigly-ceridemia was not modified by pyrazole, denoting specificity in the pyrazole inhibition of ethanol-induced hepatic injury. These findings demonstrate that pyrazole can completely prevent the development of the ethanol-induced fatty liver in the...

An evaluation of plasma triglyceride formation as a factor in the development of the ethanol-induced fatty liver

The development of an acute fatty liver in rats following the admistration of ethanol was associated with a normal intravascular removal rate of albumin-bound palmitic acid-l-C 14. When compared to isocaloric glucose-treated control rats, the ethanol group manifested a normal rate of incorporation of palmitate-l-C 14 into hepatic triglycerides prior to, and during, the development of a fatty liver. The release of triglyceride from liver, as determined by plasma triglyceride radioactivity following palmitate-l-C 14, was also unaltered. The hypertriglyceredemic response to Triton was also normal in acute ethanol-treated rats. Significant depression of hepatic triglyceride metabolism was observed in the ethanol group and is considered to be a contributing factor to the development of the acute ethanol-induced fatty liver.