Ten days after consuming the experimental diets, the mice were orally administered maltose dextrin solution (9 g of maltose dextrin/kg of body weight; CTRL
group) or ethanol solution (5 g of PKC412 purchase ethanol/kg of body weight; EtOH group) at zeitgeber time (ZT) 3 (9 am), and were sacrificed at ZT12, 18, 0, and 6. Serum and livers were collected at each time point. [Results] Serum ALT and AST levels were induced by alcohol at all time points, but with ALT showing a stronger oscillation, which was highest at ZT12 and lowest at ZT0. Serum triglyceride (TG) levels exhibited the highest induction by alcohol at ZT0, which declined to basal levels by ZT12. Interestingly, hepatic TG reached the highest levels in the EtOH group at ZT12, which was gradually decreased to the lowest levels by ZT6. Serum cholesterol levels did not show marked differences in CTRL and EtOH groups, whereas liver cholesterol content was constantly higher in the EtOH group with a moderate rhythm. Consistently, oil red O staining revealed the highest hepatic neutral lipid accumulation at ZT12 and lowest at ZT6 in the EtOH group. Gene expression analysis by qPCR uncovered a striking effect of alcohol on the alteration MLN0128 in vitro of rhythmic expression of transcription factors E2F1 and
Egr-1, nuclear receptors SHP and RORĪ³, bile acid synthesis enzyme Cyp7a1, lipid metabolic gene VLDLR, and the key clock gene NPAS2. [Conclusions] The effect of alcohol consumption by chronic and binge ethanol feeding in mice on the disruption of serum and hepatic lipid metabolism is strongly associated with alterations in the expression of key liver circadian clock genes. Disclosures: The following people have nothing to disclose: Hiroyuki Tsuchiya, Sangmin Lee, Yuxia Zhang, Rana Smalling, Li Wang FOXO3 is a multifunctional transcription factor that initiates several different transcriptional programs including oxidative stress resistance, cell proliferation, apoptosis, autophagy, and metabolism. The mechanisms that regulate
transcriptional specificity of FOXO3 are unknown. We have recently shown that ethanol and HCV infection each individually activate FOXO3 but they do so by different post-translational modifications. The AIM of this study was to determine the effects of ethanol on the transcriptional ASK1 specificity and post-translational modifications of FOXO3 and their consequences. METHODS: Huh7.5 cells were transfected with HA-tagged FOXO3, treated with 50 mM ethanol for 48 h and/or infected with HCV strain JFH1. ChiP assays were performed with anti-HA or FOXO3 antibodies. A phospho-specific S574-P_FOXO3 antibody was generated by Epitomics. RESULTS: Ethanol treatment increased mRNA for the apoptotic FOXO3 target protein Bim but not the antioxidant target protein SOD2. HCV-infection, which similarly stimulated FOXO3 reporter activity, had the opposite effect activating SOD2 but not Bim. We performed ChIP assays on Huh7.