title: Intracellular mechanisms involved in docosahexaenoic acid-induced increases in tight junction permeability in Caco-2 cell monolayers.
abstract: We recently showed that enrichment of Caco-2 cells with docosahexaenoic acid (DHA) increases lipid peroxidation and the formation of hydrogen peroxide and peroxynitrite, which disrupt the epithelial barrier function. Studies were designed to test whether the participation of phospholipase C (PLC)/Ca(2+)/protein kinase C (PKC), cyclooxygenase (COX), and 5-lipooxygenase pathways are involved in mediating the effects of DHA. Paracellular permeability was assessed from D-mannitol flux and transepithelial electrical resistance (TER) in differentiated Caco-2 cell monolayers incubated in control or DHA-enriched conditions (100 micromol/L). The effect of DHA was prevented by U73122 (PLC inhibitor), chelerytrine (PKC inhibitor), and 1-[5-iodonaphtalene-1-sulfonyl]-1H-hexahydro-1,4-diazepine hydrochloride (myosin light chain kinase inhibitor). In contrast, the effect of DHA was enhanced by A23187 (Ca(2+) ionophore) and BAPTA-AM (Ca(2+) chelator). Indomethacin (COX inhibitor) and AA961 (5-lipooxygenase inhibitor) also prevented the changes in D-mannitol flux induced by DHA, but no effect was detected for TER. Moreover, occludin and ZO-1 immunogold staining microscopy showed that the increase in paracellular permeability was accompanied by the redistribution of both tight junction proteins. We conclude that the disruption of epithelial barrier function by DHA is partly mediated by the PLC/Ca(2+)/PKC pathway and by the formation of eicosanoids.
authors.affiliation: Departament de Fisiologia, Facultat de Farmàcia, Universitat de Barcelona, Barcelona, Spain.
authors.forename: Sònia
authors.initials: S
authors.lastname: Roig-Pérez
authors.forename: Núria
authors.initials: N
authors.lastname: Cortadellas
authors.forename: Miquel
authors.initials: M
authors.lastname: Moretó
authors.forename: Ruth
authors.initials: R
authors.lastname: Ferrer
mesh_terms: D016227:Benzoquinones / Q000494:pharmacology; D018938:Caco-2 Cells; D002118:Calcium / Q000378:metabolism; D016861:Cyclooxygenase Inhibitors / Q000494:pharmacology; D004281:Docosahexaenoic Acids / Q000494:pharmacology*; D006801:Humans; D007213:Indomethacin / Q000494:pharmacology; D016859:Lipoxygenase Inhibitors / Q000494:pharmacology; D010539:Permeability; D011493:Protein Kinase C / Q000378:metabolism; D019108:Tight Junctions / Q000187:drug effects* / Q000378:metabolism*; D010738:Type C Phospholipases / Q000378:metabolism
chemical_list: D016227:Benzoquinones; D016861:Cyclooxygenase Inhibitors; D016859:Lipoxygenase Inhibitors; D004281:Docosahexaenoic Acids; C036837:2,3,5-trimethyl-6-(12-hydroxy-5,10-dodecadiynyl)-1,4-benzoquinone; D011493:Protein Kinase C; D010738:Type C Phospholipases; D002118:Calcium; D007213:Indomethacin