Ischemia/reperfusion injury in the monolayers of human intestinal epithelial cell line caco-2 and its recovery by antioxidants.
We previously established a in vitro system for assessing early ischemia/reperfusion injury using monolayers of human intestinal epithelial cell line Caco-2, in which lipid peroxidation caused by tertiary-butylhydroperoxide (t-BuOOH), a lipid peroxidation inducer, acts as a trigger of the injury. By now, we have shown that superoxide anion participates in the opening of tight junctions (TJ) induced by reoxygenation following the induction of lipid peroxidation by t-BuOOH at a low concentration. The present objectives are to elucidate the dysfunction of P-glycoprotein (P-gp) in addition to the opening of TJ by t-BuOOH at a high concentration condition using rhodamine123 (Rho123) as a P-gp substrate and cyclosporine A (CyA) as a P-gp inhibitor. Also, we compared the inhibition effect of lutein and other compounds such as biliverdin as a radical scavenger on the opening of TJ and the dysfunction of P-gp. t-BuOOH at a high concentration increased the permeability of Rho123 in the apical to basal direction and decreased basal to apical direction when compared with control conditions. t-BuOOH at a high concentration showed no significant difference between directional transport of Rho123 and no inhibition was observed in the permeability of both directions by CyA. The staining intensity of Western blot was decreased by t-BuOOH at a high concentration. Although lutein and the other compounds had recovery effects on the opening of TJ and P-gp dysfunction induced by t-BuOOH, lutein is more advantageous than other compounds since it has effective effects at the lower concentration. In conclusion, the barrier dysfunction such as the inhibition of P-gp in addition to the opening of TJ was induced by t-BuOOH at a high concentration condition. The above two barrier dysfunctions was ameliorated by antioxidant such as lutein and biliverdin.
Nagira
Mayuko
M
Department of Drug Absorption and Pharmacokinetics, School of Pharmacy, Tokyo University of Pharmacy and Life Science, Japan. nagira@ps.toyaku.ac.jp
Tomita
Mikio
M
Mizuno
Satoshi
S
Kumata
Miyuki
M
Ayabe
Toshiyuki
T
Hayashi
Masahiro
M
D014013:1,2-Dihydroxybenzene-3,5-Disulfonic Acid Disodium Salt / Q000494:pharmacology; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000032:analysis / Q000378:metabolism; D000975:Antioxidants / Q000737:chemistry / Q000494:pharmacology*; D001664:Biliverdine / Q000494:pharmacology; D015153:Blotting, Western; D018938:Caco-2 Cells; D002463:Cell Membrane Permeability / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D004847:Epithelial Cells / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D005456:Fluorescent Dyes / Q000378:metabolism / Q000494:pharmacology; D006801:Humans; D007202:Indicators and Reagents / Q000494:pharmacology; D007413:Intestinal Mucosa / Q000378:metabolism; D007422:Intestines / Q000187:drug effects* / Q000473:pathology; D015227:Lipid Peroxidation / Q000187:drug effects; D014975:Lutein / Q000494:pharmacology; D015427:Reperfusion Injury; D020112:Rhodamine 123 / Q000378:metabolism / Q000493:pharmacokinetics; D014810:Vitamin E / Q000494:pharmacology; D024341:Xanthophylls / Q000494:pharmacology; D020122:tert-Butylhydroperoxide / Q000494:pharmacology
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000975:Antioxidants; D005456:Fluorescent Dyes; D007202:Indicators and Reagents; D024341:Xanthophylls; D014810:Vitamin E; D020112:Rhodamine 123; D014013:1,2-Dihydroxybenzene-3,5-Disulfonic Acid Disodium Salt; C005948:astaxanthine; D020122:tert-Butylhydroperoxide; D001664:Biliverdine; D014975:Lutein