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Kazuyuki Shimizu, Guey-Shuang Wu, Chand Sultana, Vijay K. Kalra, Narsing A. Rao; Stimulation of Macrophages by Retinal ProteinsProduction of Reactive Nitrogen and Oxygen Metabolites. Invest. Ophthalmol. Vis. Sci. 1999;40(13):3215-3223.
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purpose. In previous work, it has been shown that in experimental autoimmune
uveitis, the peroxynitrite-mediated protein nitration product
nitrotyrosine was localized in the degenerating photoreceptors.
Subsequently, phagocyte-generated inducible nitric oxide synthase
(iNOS) was also found to localize, primarily in the outer retina and to
a lesser extent in the anterior segments. This study was intended to
determine whether retinal soluble proteins such as S-antigen and
interphotoreceptor retinoid-binding protein (IRBP) play a role in the
induction of ·NO and superoxide by a macrophage cell line and by
rat and rabbit peritoneal macrophages.
methods. Cells from the murine macrophage cell line RAW 264.7 and rat and rabbit
peritoneal macrophages were incubated in the presence of retinal
soluble proteins. The nitrite level in the cultured supernatant was
evaluated for ·NO production using the Griess reaction.
Activation of nuclear transcription factor κB (NF-κB) was
determined by electrophoretic mobility shift assay. Superoxide
production was measured by superoxide dismutase-inhibitable reduction
of cytochrome C.
results. Both S-antigen and IRBP induced significant, dose-dependent nitrite
production in RAW 264.7 and rat peritoneal macrophages. Induction of
iNOS by retinal proteins was inhibited by the iNOS-specific inhibitor
aminoguanidine and the tyrosine kinase inhibitor genistein. This iNOS
induction was accompanied by the activation of NF-κB. S-antigen also
induced superoxide production in rabbit peritoneal macrophages, but not
in RAW 264.7.
conclusions. These results show that soluble retinal proteins significantly induce·
NO and superoxide production by macrophages. Increased
production of reactive oxygen species by macrophages in the presence of
these soluble retinal proteins in vivo may accelerate photoreceptor
degeneration in uveitis.
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