TY - JOUR T1 - Acetoacetate Induces CYP2E1 Protein and Suppresses CYP2E1 mRNA in Primary Cultured Rat Hepatocytes JF - Journal of Pharmacology and Experimental Therapeutics JO - J Pharmacol Exp Ther SP - 203 LP - 213 DO - 10.1124/jpet.105.084608 VL - 315 IS - 1 AU - Mohamed A. Abdelmegeed AU - Nicholas J. Carruthers AU - Kimberley J. Woodcroft AU - Sang K. Kim AU - Raymond F. Novak Y1 - 2005/10/01 UR - http://jpet.aspetjournals.org/content/315/1/203.abstract N2 - The ketone body acetoacetate (AA) in the absence of insulin or in the presence of diabetic insulin levels decreases CYP2E1 mRNA expression in a concentration- and time-dependent manner in primary cultured rat hepatocytes. AA activates p70 ribosomal S6 kinase (p70S6K) and protein kinase C (PKC) by ∼2- to 2.5-fold, respectively, following 6-h treatment. The AA-mediated activation of p70S6K, but not PKC, was abolished by inhibition of PI 3-K with LY294002 [2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one] or wortmannin, in agreement with p70S6K being downstream of phosphatidylinositol 3-kinase (PI 3-K). Inhibition of PI 3-K, mTOR with rapamycin, or PKC with bisindolylmaleimide ameliorated the AA-mediated down-regulation of CYP2E1 mRNA expression. Neither the mitogen-activated protein kinase kinase inhibitor PD98059 (2′-amino-3′-methoxyflavone) nor the p38 mitogen-activated protein kinase inhibitor SB203580 [4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)1H-imidazole] ameliorated the AA-mediated suppression of CYP2E1 mRNA expression. Heterogeneous nuclear RNA analysis revealed that AA suppressed CYP2E1 gene transcription by ∼50% and that inhibition of PI 3-K and PKC diminished this AA-mediated effect on transcription. CYP2E1 mRNA half-life slightly increased from ∼24 h in untreated hepatocytes to ∼32 h in AA-treated cells. Interestingly, AA increased CYP2E1 protein levels by ∼2- and 2.5-fold at 24 and 48 h, respectively. dl-β-Hydroxybutyrate was without effect. Polysomal distribution studies revealed that AA increased the proportion of RNA associated with the actively translated polysomal fractions versus the 40S to 60S untranslated fractions by ∼40%. CYP2E1 protein half-life increased from ∼8 h in untreated hepatocytes to ∼24 in AA-treated cells. These data show that AA decreases CYP2E1 mRNA expression through inhibition of gene transcription while simultaneously elevating CYP2E1 protein levels through increased translation and decreased protein degradation. The American Society for Pharmacology and Experimental Therapeutics ER -