Effect of pituitary adenylate cyclase-activating polypeptide (PACAP) on IL-6, IL-8, and MCP-1 expression in human retinal pigment epithelial cell line

Abstract

PURPOSE: To examine pituitary adenylate cyclase-activating polypeptide (PACAP) receptors (PAC1, VPAC1, and VPAC2) mRNA and the effect of PACAP on interleukin-6 (IL-6), interleukin-8 (IL-8), and monocyte chemotactic protein-1 (MCP-1) expression in human retinal pigment epithelial cell line (ARPE-19) stimulated with interleukin-1beta (IL-1beta). METHODS: Expression of PACAP receptor mRNA was examined by reverse transcription polymerase chain reaction (RT-PCR). PACAP and IL-1beta were added to serum-free medium. IL-6, IL-8, and MCP-1 mRNA were measured by real-time PCR. IL-6, IL-8, and MCP-1 protein concentrations were measured using enzyme-linked immunosorbent assay. Nuclear factor kappaB (NF-kappaB) translocation was examined by immunofluorescence. RESULTS: PAC1 and VCAP1 receptors mRNA were expressed in unstimulated cells. VCAP2 mRNA was expressed in cells stimulated with IL-1beta. IL-1beta stimulated IL-6, IL-8, and MCP-1 mRNA expression and protein levels. PACAP (10(- 7) to 10(- 6) M) inhibited IL-1beta -stimulated IL-6, IL-8, and MCP-1 mRNA and protein levels. Immunofluorescence of NF-kappaB in the nucleus was dense 30 min after stimulation with IL-1beta, and it was decreased by PACAP. CONCLUSIONS: ARPE-19 cells had PACAP receptors mRNA. PACAP inhibited IL-6, IL-8, and MCP-1 expression and protein secretion. Possibly, the effect on cytokines may be via suppression of NF-kappaB translocation.

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