15, 16 Recently, β-catenin MG-132 in vivo was shown to be the master regulator of hepatic metabolic zonation. 17, 18 We and others have previously reported that β-catenin regulates the expression of Cyp2E1, the loss of which makes β-catenin knockout (KO) mice resistant to acetaminophen-induced hepatotoxicity. 19-21 On the other hand, liver-specific loss of β-catenin leads to increased susceptibility to steatohepatitis in the methionine choline-deficient diet model of liver injury. 22 In addition to its metabolic role, β-catenin has also been implicated in the response to oxidative stress. 23 Because alcohol metabolism generates oxidative stress in the liver, we hypothesized that β-catenin may regulate the

coordinated response of the liver to alcohol-metabolism and the associated increase in oxidative stress. Thus, this study was Selleckchem CAL-101 undertaken to determine the effect of hepatocyte-specific loss of β-catenin on ethanol metabolism and alcohol-mediated liver injury in vivo in a murine model using the Lieber-DeCarli ethanol diet. ADH, alcohol dehydrogenase; ALDH, aldehyde dehydrogenase; ALT, alanine aminotransferase; AST, aspartate aminotransferase; Cyp2E1, cytochrome P450 2E1; EtOH, ethanol-fed; FOXO, forkhead box; kb, kilobase; KO, Knockout; LPO, lipid peroxidation; MDA, malondialdehyde; NAC, N-acetylcysteine; NAD, nicotinamide adenine dinucleotide; NADPH, nicotinamide adenine dinucleotide phosphate-reduced;

PCR, polymerase chain reaction; PF, pair-fed; SOD, superoxide dismutase;

TCA, tricarboxylic acid; TCF4, T-cell factor 4; WT, wild type. Liver-specific β-catenin KO mice were generated as previously described. 19 Female KO mice (Ctnnb1−/−, Cre+/−) and wild-type (WT) littermates (Ctnnb1loxp/loxp;Cre−/−; or Ctnnb1loxp/−,Cre−/−; or Ctnnb1loxp/−;Cre+/−) were between the ages of 8 and 12 weeks at the start of the experiments. All three WT genotypes were used in the experiments as controls and showed indistinguishable phenotype among them on both diets. Mice were maintained on 12-hour light-dark cycles and had free access to the diets. The high-fat Lieber-DeCarli liquid diet (5% final ethanol concentration) was used with a 6-day ramp-up period (2 days of control diet, 2 days of 1.8% ethanol, 2 days at 3.4% ethanol, and then 5% ethanol for 1, 6, or 22 days). The control group received an isocaloric maltodextrin-containing diet in a see more pair-fed (PF) fashion. For collection of blood for plasma ethanol and ammonia levels, mice were fed the high-fat Lieber-DeCarli liquid diet for 7 days (6 days of ramp-up followed by 1 day of 5% ethanol), and blood was collected at the end of the dark cycle at 7 a.m. The University of Pittsburgh Institutional Animal Care and Use Committee approved the study. Other reagents and methods are described in the Supporting Materials. During the ethanol ramp-up period, both genotypes had similar food intake, weight change, and exhibited normal behavior.