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Acta Ophthalmol. 2012 Jun;90(4):e255-63. doi: 10.1111/j.1755-3768.2011.02350.x. Epub 2012 Feb 7.

Regulation of the expression of interleukin-8 induced by 25-hydroxycholesterol in retinal pigment epithelium cells.

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  • 1Centre of Ophthalmology and Vision Sciences (COCV), IBILI, Faculty of Medicine, University of Coimbra, Coimbra, Portugal.

Abstract

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.

© 2012 The Authors. Acta Ophthalmologica © 2012 Acta Ophthalmologica Scandinavica Foundation.

PMID:
22313893
[PubMed - indexed for MEDLINE]
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