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Sappington RM, Calkins DJ: Contribution of TRPV1 to microglia-derived IL-6 and NFkappaB translocation with elevated hydrostatic pressure. Invest Ophthalmol Vis Sci. 2008 Jul;49(7):3004-17. Epub 2008 Mar 24.
PURPOSE: The authors investigated the contributions of the transient receptor potential vanilloid-1 receptor (TRPV1) and Ca (2+) to microglial IL-6 and nuclear factor kappa B (NFkappaB) translocation with elevated hydrostatic pressure. METHODS: The authors first examined IL-6 colocalization with the microglia marker Iba-1 in the DBA/2 mouse model of glaucoma to establish relevance. They isolated microglia from rat retina and maintained them at ambient or elevated (+70 mm Hg) hydrostatic pressure in vitro and used ELISA and immunocytochemistry to measure changes in the IL-6 concentration and NFkappaB translocation induced by the Ca (2+) chelator EGTA, the broad-spectrum Ca (2+) channel inhibitor ruthenium red, and the TRPV1 antagonist iodo-resiniferatoxin (I-RTX). They applied the Ca (2+) dye Fluo-4 AM to measure changes in intracellular Ca (2+) at elevated pressure induced by I-RTX and confirmed TRPV1 expression in microglia using PCR and immunocytochemistry. RESULTS: In DBA/2 retina, elevated intraocular pressure increased microglial IL-6 in the ganglion cell layer. Elevated hydrostatic pressure (24 hours) increased microglial IL-6 release, cytosolic NFkappaB, and NFkappaB translocation in vitro. These effects were reduced substantially by EGTA and ruthenium red. Antagonism of TRPV1 in microglia partially inhibited pressure-induced increases in IL-6 release and NFkappaB translocation. Brief elevated pressure (1 hour) induced a significant increase in microglial intracellular Ca (2+) that was partially attenuated by TRPV1 antagonism. CONCLUSIONS: Elevated pressure induces an influx of extracellular Ca (2+) in retinal microglia that precedes the activation of NFkappaB and the subsequent production and release of IL-6 and is at least partially dependent on the activation of TRPV1 and other ruthenium red-sensitive channels. |
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