title: Role for intestinal CYP2E1 in alcohol-induced circadian gene-mediated intestinal hyperpermeability.
abstract: We have shown that alcohol increases Caco-2 intestinal epithelial cell monolayer permeability in vitro by inducing the expression of redox-sensitive circadian clock proteins CLOCK and PER2 and that these proteins are necessary for alcohol-induced hyperpermeability. We hypothesized that alcohol metabolism by intestinal Cytochrome P450 isoform 2E1 (CYP2E1) could alter circadian gene expression (Clock and Per2), resulting in alcohol-induced hyperpermeability. In vitro Caco-2 intestinal epithelial cells were exposed to alcohol, and CYP2E1 protein, activity, and mRNA were measured. CYP2E1 expression was knocked down via siRNA and alcohol-induced hyperpermeability, and CLOCK and PER2 protein expression were measured. Caco-2 cells were also treated with alcohol or H₂O₂ with or without N-acetylcysteine (NAC) anti-oxidant, and CLOCK and PER2 proteins were measured at 4 or 2 h. In vivo Cyp2e1 protein and mRNA were also measured in colon tissue from alcohol-fed mice. Alcohol increased CYP2E1 protein by 93% and enzyme activity by 69% in intestinal cells in vitro. Alcohol feeding also increased mouse colonic Cyp2e1 protein by 73%. mRNA levels of Cyp2e1 were not changed by alcohol in vitro or in mouse intestine. siRNA knockdown of CYP2E1 in Caco-2 cells prevented alcohol-induced hyperpermeability and induction of CLOCK and PER2 proteins. Alcohol-induced and H₂O₂-induced increases in intestinal cell CLOCK and PER2 were significantly inhibited by treatment with NAC. We concluded that our data support a novel role for intestinal CYP2E1 in alcohol-induced intestinal hyperpermeability via a mechanism involving CYP2E1-dependent induction of oxidative stress and upregulation of circadian clock proteins CLOCK and PER2.
authors: Department of Internal Medicine, Rush University Medical Center, Chicago, IL 60612, USA. christopher_b_forsyth@rush.edu
authors: Christopher B
authors: CB
authors: Forsyth
authors: Robin M
authors: RM
authors: Voigt
authors: Maliha
authors: M
authors: Shaikh
authors: Yueming
authors: Y
authors: Tang
authors: Arthur I
authors: AI
authors: Cederbaum
authors: Fred W
authors: FW
authors: Turek
authors: Ali
authors: A
authors: Keshavarzian
mesh_terms: D000818:Animals; D056926:CLOCK Proteins / Q000235:genetics / Q000378:metabolism*; D018938:Caco-2 Cells; D019392:Cytochrome P-450 CYP2E1 / Q000235:genetics / Q000378:metabolism*; D000431:Ethanol / Q000633:toxicity*; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D007413:Intestinal Mucosa / Q000378:metabolism; D007422:Intestines / Q000187:drug effects*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D056950:Period Circadian Proteins / Q000235:genetics / Q000378:metabolism*; D010539:Permeability; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism
chemical_list: D056950:Period Circadian Proteins; D012333:RNA, Messenger; D000431:Ethanol; D019392:Cytochrome P-450 CYP2E1; D056926:CLOCK Proteins
keywords: clock genes; cytochrome P450 isoform 2E1; ethanol; leaky gut; oxidative stress
references: World J Gastroenterol. 2010 Mar 21;16(11):1321-9
references: 20238398
references: J Pharmacol Exp Ther. 2006 Apr;317(1):44-52
references: 16373529
references: Gastroenterology. 1989 Mar;96(3):736-49
references: 2914637
references: Am J Physiol Regul Integr Comp Physiol. 2008 Dec;295(6):R2034-40
references: 18843092
references: J Biol Chem. 1995 Dec 15;270(50):29632-5
references: 8530344
references: Alcohol. 1993 Nov-Dec;10(6):459-64
references: 8123200
references: Am J Physiol Regul Integr Comp Physiol. 2009 Sep;297(3):R729-37
references: 19553498
references: Semin Liver Dis. 2004 Aug;24(3):217-32
references: 15349801
references: Science. 2012 Aug 17;337(6096):839-42
references: 22859819
references: J Hepatol. 2009 Mar;50(3):538-47
references: 19155080
references: Cell. 2008 Jul 25;134(2):329-40
references: 18662547
references: Subcell Biochem. 2013;67:131-43
references: 23400919
references: Am J Physiol Gastrointest Liver Physiol. 2010 Sep;299(3):G549-55
references: 20558764
references: Science. 2001 Jul 20;293(5529):510-4
references: 11441146
references: Arch Toxicol. 2009 Jun;83(6):519-48
references: 19448996
references: Alcohol. 2008 Aug;42(5):349-61
references: 18504085
references: Antioxid Redox Signal. 2013 Jul 10;19(2):192-208
references: 23198849
references: Nature. 2011 Jan 27;469(7331):498-503
references: 21270888
references: Lancet. 2009 Jun 27;373(9682):2223-33
references: 19560604
references: Annu Rev Biochem. 2002;71:307-31
references: 12045099
references: Drug Metab Dispos. 1985 Sep-Oct;13(5):548-52
references: 2865101
references: Sleep Med. 2009 Jun;10(6):597-603
references: 19403332
references: Rev Endocr Metab Disord. 2009 Dec;10(4):293-300
references: 19798581
references: Free Radic Biol Med. 2008 Mar 1;44(5):723-38
references: 18078827
references: Biochem Biophys Res Commun. 1970 Aug 24;40(4):858-65
references: 4395603
references: Am J Gastroenterol. 1999 Jan;94(1):200-7
references: 9934756
references: Gastroenterology. 2008 Dec;135(6):2019-29
references: 18848557
references: J Pharmacol Exp Ther. 2000 Sep;294(3):997-1008
references: 10945852
references: J Pharmacol Exp Ther. 1999 Dec;291(3):1075-85
references: 10565827
references: Alcohol Clin Exp Res. 2011 Jul;35(7):1305-14
references: 21463335
references: Nat Rev Neurosci. 2003 Aug;4(8):649-61
references: 12894240
references: Science. 2005 May 13;308(5724):1043-5
references: 15845877
references: Gastroenterology. 2000 Aug;119(2):536-49
references: 10930389
references: J Hepatol. 1987 Feb;4(1):8-14
references: 3571935
references: Am J Physiol Gastrointest Liver Physiol. 2010 Mar;298(3):G364-74
references: 20007851
references: Alcohol Clin Exp Res. 1998 Nov;22(8):1724-30
references: 9835287
references: Lancet. 1984 Jan 28;1(8370):179-82
references: 6141332
references: Biochem Pharmacol. 1996 Jan 12;51(1):61-9
references: 8534269
references: Am J Pathol. 2006 Apr;168(4):1148-54
references: 16565490
references: Free Radic Biol Med. 2012 Sep 15;53(6):1346-57
references: 22819980
references: Hepatology. 2010 Jan;51(1):307-28
references: 20034030
references: Nature. 2002 Aug 29;418(6901):935-41
references: 12198538
references: Hepatology. 1995 Jun;21(6):1610-7
references: 7768506
references: Cell. 1999 Jan 22;96(2):271-90
references: 9988221
references: Physiol Rev. 1997 Apr;77(2):517-44
references: 9114822
references: Alcohol Clin Exp Res. 2008 Apr;32(4):669-82
references: 18341647
references: Chronobiol Int. 2011 Apr;28(3):204-15
references: 21452916
references: Gastroenterology. 2007 Oct;133(4):1250-60
references: 17919497
references: J Neurochem. 2004 Mar;88(6):1547-54
references: 15009656
references: Alcohol Clin Exp Res. 2007 Jun;31(6):1053-60
references: 17428293
references: Biochem Biophys Res Commun. 1994 Dec 15;205(2):1064-71
references: 7802633
references: J Biol Rhythms. 2009 Aug;24(4):304-12
references: 19625732
references: Int J Hepatol. 2012;2012:582790
references: 22028977
references: Methods Mol Biol. 2008;447:171-83
references: 18369919
references: PLoS One. 2009;4(3):e4882
references: 19287494
references: Gastroenterology. 1990 Oct;99(4):1044-53
references: 2203661
references: Free Radic Biol Med. 2009 Feb 15;46(4):480-91
references: 19063961
references: Am J Physiol. 1998 Dec;275(6):H2334-7
references: 9843836
references: Science. 2009 May 1;324(5927):654-7
references: 19286518
references: Semin Liver Dis. 1988 Feb;8(1):12-25
references: 3283941
references: Am J Physiol Gastrointest Liver Physiol. 2008 Apr;294(4):G892-8
references: 18239056
references: J Nephrol. 2010 Jan-Feb;23(1):102-10
references: 20091493
references: Alcohol Clin Exp Res. 1997 Dec;21(9):1599-605
references: 9438518
references: Alcohol Clin Exp Res. 2010 Jan;34(1):19-31
references: 19860811
references: Alcohol Clin Exp Res. 1999 Jun;23(6):991-1007
references: 10397283
references: Proc Soc Exp Biol Med. 2000 Sep;224(4):302-8
references: 10964266
references: Br J Clin Pharmacol. 2005 Jul;60(1):54-60
references: 15963094
references: Alcohol Clin Exp Res. 2005 Aug;29(8):1550-7
references: 16156052
references: Cell. 2002 May 3;109(3):307-20
references: 12015981
references: Am J Physiol Gastrointest Liver Physiol. 2004 Jun;286(6):G881-4
references: 15132946
references: Cell Cycle. 2009 Sep 1;8(17):2794-801
references: 19652538
references: Physiol Behav. 2005 Mar 31;84(4):537-42
references: 15811388
references: Genes Nutr. 2010 Jun;5(2):149-67
references: 19798529
references: Proc Natl Acad Sci U S A. 2007 Oct 2;104(40):15747-52
references: 17898172
references: Am J Physiol Gastrointest Liver Physiol. 2004 Sep;287(3):G510-7
references: 15331350
references: BMC Clin Pharmacol. 2005 Oct 27;5:4
references: 16253141
references: J Lipid Res. 2009 Sep;50(9):1800-13
references: 19387090
references: Alcohol Clin Exp Res. 1994 Oct;18(5):1280-5
references: 7847620
references: World J Gastroenterol. 2010 Mar 21;16(11):1304-13
references: 20238396
references: FASEB J. 1992 Jan 6;6(2):724-30
references: 1537462
references: Mol Cell Biochem. 1991 Mar 27;102(1):61-9
references: 2052000
references: Trends Cell Biol. 2010 Jan;20(1):36-44
references: 19926479
references: Free Radic Biol Med. 2012 Sep 1;53(5):1170-80
references: 22749809
references: Alcohol Clin Exp Res. 2011 Sep;35(9):1635-43
references: 21535025
references: Trends Biochem Sci. 2012 Nov;37(11):484-92
references: 22917814
references: Free Radic Biol Med. 2010 Nov 15;49(9):1406-16
references: 20692331
references: Nat Med. 2005 Jan;11(1):35-42
references: 15608650
references: Alcohol Clin Exp Res. 1996 Nov;20(8 Suppl):138A-146A
references: 8947253
references: Annu Rev Physiol. 2001;63:647-76
references: 11181971
references: Am J Physiol Gastrointest Liver Physiol. 2010 Feb;298(2):G143-50
references: 19926812
references: Curr Biol. 2012 Jul 24;22(14):R570-1
references: 22835791