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Regulation of the expression of interleukin‐8 induced by 25‐hydroxycholesterol in retinal pigment epithelium cells
Author(s) -
Catarino Steve,
Bento Carla F.,
Brito Ana,
Murteira Eliana,
Fernandes Alexandre F.,
Pereira Paulo
Publication year - 2012
Publication title -
acta ophthalmologica
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.534
H-Index - 87
eISSN - 1755-3768
pISSN - 1755-375X
DOI - 10.1111/j.1755-3768.2011.02350.x
Subject(s) - secretion , retinal pigment epithelium , phosphorylation , microbiology and biotechnology , cytokine , chemistry , pi3k/akt/mtor pathway , transcription factor , signal transduction , kinase , mapk/erk pathway , biology , retinal , biochemistry , gene , immunology
. Purpose: This study aimed at elucidating the molecular mechanisms involved in the regulation of IL‐8 production by several oxysterols in retinal pigment epithelium (RPE) cells. Methods: A human cell line from RPE (ARPE‐19) was used to test the role of cholesterol and several oxysterols (25‐OH, 7‐KC and 7β‐OH) in the expression and secretion of IL‐8. Expression of IL‐8 was assessed by real‐time PCR, while IL‐8 secretion was evaluated by ELISA. PI3K‐, MEK1/2‐, ERK1/2‐ and NF‐κB‐specific inhibitors were used to assess the specific role of the several players on the regulation of IL‐8 production by oxysterols. A gene‐reporter assay for AP‐1 activity was also conducted to evaluate the putative role of this transcription factor on IL‐8 expression induced by oxysterols. Results: Here, we demonstrate that 25‐OH specifically increases transcription and secretion of the cytokine IL‐8 in ARPE‐19 cells. Indeed, treatment of ARPE‐19 with 25‐OH, but not with 7‐KC, 7β‐OH or cholesterol, induced the secretion of IL‐8 from cells. 25‐OH also induced the activation/phosphorylation of ERK1/2 through a mechanism dependent on MEK, ERK1/2 and PI3K kinase activity. Real‐time PCR and ELISA experiments demonstrated that 25‐OH increased transcription and secretion of IL‐8 through a mechanism that is dependent on ERK1/2 and PI3K activity. Furthermore, 25‐OH triggered the activation/phosphorylation of the AP‐1 component c‐Jun and, consistently, increased the transcriptional activity of AP‐1. Additionally, we also found that 25‐OH decreases the levels of IκB and increases the nuclear levels of NF‐κB p65 subunit and that inhibition of NF‐κB activity partially prevents the increased secretion of IL‐8 induced by 25‐OH. Conclusions: The results presented in this study suggest a role for 25‐OH in inducing IL‐8 production through pathways that are likely to involve AP‐1 and NF‐κB in ARPE‐19 cells. Our data may also provide new molecular targets for the treatment of AMD.