title: Heme modulates intestinal epithelial cell activation: involvement of NADPHox-derived ROS signaling.
abstract: In many gut chronic inflammatory conditions, intestinal epithelium (IE) is deprived of the protection of the mucus secreted by IE-specialized cells. In these events, bleeding and subsequent lysis of erythrocytes are common. This may lead to the release of high amounts of heme in the intestinal lumen, which interacts with IE. Previous works from our group have shown that heme itself is a proinflammatory molecule, activating a number of phlogistic signaling events in a nicotinamide adenine dinucleotide phosphate oxidase (NADPHox)-dependent manner. In this study, we aim to evaluate the effects of heme upon a well-established nontransformed small intestine epithelial cell lineage (IEC 6). Our results show that free heme evokes intracellular reactive oxygen species (ROS) production by IEC 6 cells, which is inhibited both by pharmacological inhibition with diphenyleneiodonium (10 μM), a NADPHox inhibitor, and small interfering RNA-mediated suppression of NOX1, a constitutive NADPHox isoform present in intestinal epithelial cells. Focal adhesion kinase phosphorylation and actin cytoskeleton polymerization are also induced by heme in a NADPHox-dependent manner. Heme increases monolayer permeability and redistributes key modulators of cell-cell adhesion as zona occludens-1 and E-cadherin proteins via NADPHox signaling. Heme promotes IEC 6 cell migration and proliferation, phenomena also regulated by NADPHox-derived ROS. Heme, in NADPHox-activating concentrations, is able to induce mRNA expression of IL-6, a cytokine implicated in inflammatory and tumorigenic responses. These data indicate a prominent role for heme-derived signaling in the pathophysiology of intestinal mucosa dysfunction and address an important role of NADPHox activity on the pathogenesis of intestinal inflammatory conditions.
authors.affiliation: Laboratório de Farmacologia Celular e Molecular, Departamento de Biologia Celular, Instituto de Biologia Roberto Alcantara Gomes, Universidade do Estado do Rio de Janeiro, Rio de Janeiro, Brazil.
authors.forename: Pedro
authors.initials: P
authors.lastname: Barcellos-de-Souza
authors.forename: João Alfredo
authors.initials: JA
authors.lastname: Moraes
authors.forename: Julio Cesar Madureira
authors.initials: JC
authors.lastname: de-Freitas-Junior
authors.forename: José Andrés
authors.initials: JA
authors.lastname: Morgado-Díaz
authors.forename: Christina
authors.initials: C
authors.lastname: Barja-Fidalgo
authors.forename: Maria Augusta
authors.initials: MA
authors.lastname: Arruda
mesh_terms: D008841:Actin Cytoskeleton / Q000378:metabolism; D000818:Animals; D015820:Cadherins / Q000502:physiology; D002460:Cell Line; D002465:Cell Movement / Q000187:drug effects / Q000502:physiology; D049109:Cell Proliferation / Q000187:drug effects; D004386:Duodenum / Q000187:drug effects / Q000201:enzymology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004847:Epithelial Cells / Q000187:drug effects* / Q000201:enzymology; D051416:Focal Adhesion Protein-Tyrosine Kinases / Q000378:metabolism; D020868:Gene Silencing; D006418:Heme / Q000494:pharmacology*; D015850:Interleukin-6 / Q000096:biosynthesis; D007413:Intestinal Mucosa / Q000187:drug effects* / Q000201:enzymology; D009247:NADH, NADPH Oxidoreductases / Q000037:antagonists & inhibitors / Q000235:genetics; D000074624:NADPH Oxidase 1; D019255:NADPH Oxidases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D009861:Onium Compounds / Q000494:pharmacology; D010539:Permeability / Q000187:drug effects; D010766:Phosphorylation; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects* / Q000235:genetics; D062826:Zonula Occludens-1 Protein / Q000502:physiology
chemical_list: D015820:Cadherins; D004791:Enzyme Inhibitors; D015850:Interleukin-6; D009861:Onium Compounds; D017382:Reactive Oxygen Species; C569044:Tjp1 protein, rat; D062826:Zonula Occludens-1 Protein; D006418:Heme; C007517:diphenyleneiodonium; D009247:NADH, NADPH Oxidoreductases; D000074624:NADPH Oxidase 1; D019255:NADPH Oxidases; C000614712:NOX1 protein, rat; D051416:Focal Adhesion Protein-Tyrosine Kinases