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Lia Judice M. Relvas, Christophe Bouffioux, Brice Marcet, Didier Communi, Maya Makhoul, Michael Horckmans, Daniel Blero, Catherine Bruyns, Laure Caspers, Jean-Marie Boeynaems, François Willermain; Extracellular Nucleotides and Interleukin-8 Production by ARPE Cells: Potential Role of Danger Signals in Blood–Retinal Barrier Activation. Invest. Ophthalmol. Vis. Sci. 2009;50(3):1241-1246. doi: 10.1167/iovs.08-1902.
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purpose. RPE cell activation is an important feature of autoimmune uveitis. This investigation focused on whether extracellular nucleotides could contribute to this activation, and the effects of ATPγS, UTP, and UDP on the production of IL-8 by RPE cells was studied in relation to their expression of functional P2Y receptors.
methods. ARPE-19 cells were cultured with ATPγS, UTP, UDP, and TNF. IL-8 gene transcription and protein production were measured by semiquantitative RT-PCR and ELISA. Western blot analysis and RT-PCR were used to investigate ERK 1/2 activation and P2Y expression. Changes in intracellular calcium and cAMP concentration were analyzed by spectrofluorometry and radioimmunoassay.
results. Stimulation of ARPE-19 cells with ATPγS, UTP, and UDP induced IL-8 gene transcription and protein secretion. TNFα induction of IL-8 secretion was also increased by ATPγS, UTP, and UDP. Nucleotide induction of IL-8 production was blocked by PD98059, and all nucleotides stimulated ERK 1/2 phosphorylation. P2Y2 and P2Y6 mRNAs were detected in ARPE-19 cells. All tested nucleotides induced a pulse of intracellular calcium.
conclusions. ATPγS, UTP, and UDP stimulate both basal and TNFα-induced IL-8 secretion in RPE cells through an ERK 1/2-dependent pathway. The results suggest that those effects are mediated by P2Y2 and P2Y6 receptors.
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