Effect of Alpha-melanocyte-stimulating Hormone on Interleukin 8 and Monocyte Chemotactic Protein 1 Expression in a Human Retinal Pigment Epithelial Cell Line

Purpose: To examine melanocortin receptor (from MC-1 to MC-5) mRNA and the effect of alpha-melanocyte-stimulating hormone (alpha-MSH) on interleukin 8 (IL-8) and monocyte chemotactic protein 1 (MCP-1) expression in a human retinal pigment epithelial cell line (ARPE-19) stimulated with IL-1beta or tumor necrosis factor alpha (TNF-alpha).

Methods: Expressions of MC-1 to MC-5 mRNA were examined by semiquantitative reverse transcription polymerase chain reaction (RT-PCR). alpha-MSH and IL-1beta or TNF-alpha were added to serum-free medium. IL-8 and MCP-1 mRNA were measured by real-time PCR. IL-8 and MCP-1 protein concentrations were measured using enzyme-linked immunosorbent assay. Nuclear factor kappaB (NF-kappaB) translocation was examined by immunofluorescent staining/microscopy.

Results: MC-1 to MC-5 receptor mRNA was expressed in unstimulated cells. IL-1beta stimulated IL-8 and MCP-1 mRNA at 6 h. TNF-alpha stimulated IL-8 and MCP-1 mRNA expression at 1.5 and 3 h. alpha-MSH (10(-14) to 10(-10)M) inhibited IL-8 and MCP-1 mRNA expression in the cells stimulated with IL-1beta or TNF-alpha. alpha-MSH inhibited IL-1beta or TNF-alpha-stimulated IL-8 and MCP-1 protein levels in the media. Immunofluorescent staining/microscopy of NF-kappaB in the nucleus was dense 30 min after stimulation with IL-1beta or TNF-alpha and was decreased by alpha-MSH.

Conclusions: ARPE-19 cells had MC-1 mRNA. alpha-MSH inhibited IL-8 and MCP-1 expression and protein secretion. Possibly, the effect on chemotactic factors may be via suppression of NF-kappaB translocation.