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Xin Chen, Wei Chi, Jin Li, Changjun Wang, Fang Bian, Stephen C Pflugfelder, Cintia S De Paiva, De-Quan Li; Imbalanced Activation between NLRP3 and NLRP6 by Ocular Innate Immunity in Experimental Dry Eye Mice. Invest. Ophthalmol. Vis. Sci. 2016;57(12):No Pagination Specified.
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© ARVO (1962-2015); The Authors (2016-present)
To explore the underlying mechanisms by which ocular mucosal innate immunity responds to desiccating stress by regulating NOD-like receptors (NLRs)-mediated inflammasomes using an experimental dry eye mouse model.
The experimental dry eye was induced by desiccating stress in C57BL/6 mice exposed to an air draft in <40% ambient humidity with subcutaneous injection of scopolamine hydrobromide for 5 days, with or without subconjunctival injection of a NLRP3 inhibitor glybenclamide. Reverse transcription and quantitative real time PCR (RT-qPCR), immunofluorescent staining, Western blotting, caspase activity assays and ELISA were performed to evaluate the inflammasome activation and regulation of inflammatory cytokines induced by desiccating stress.
The initial inflammasome priming was observed by upregulated mRNA expression of NLRP3 and pro-IL-1β with increased activity of deubiquitinase BRCC3, and the activation was evidenced by the increased protein production of NLRP3 and ASC with increased activation of caspase-1 from pro-caspase-1, in corneal and conjunctival epithelia of the dry eye mice, as evaluated by RT-qPCR, immunofluorescent staining, Western blot analysis, and caspase activity assays. The maturation, activation and release of IL-1β and IL-18 by activated caspase-1 were largely promoted. Glybenclamide inhibited NLRP3 activation, and also blocked the caspase-1 activation, which lead to suppressed activation and release of IL-1β and IL-18. Interestingly, NLRP6 was observed to be downregulated at both mRNA and protein levels in corneal and conjunctival epithelia of dry eye mice.
These findings uncovered a novel mechanism and signaling pathway of innate immunity by ocular surface mucosal epithelium in response to environmental desiccating stress, which activates NLRP3 mediated inflammasome but suppresses anti-inflammatory NLRP6. The imbalanced activity between NLRP3 and NLRP6 may contribute to casapase-1 dependent activation of inflammatory cytokines IL-1β and IL-18. These results provide new insight in the innate immunity that contributes to pathogenesis of dry eye.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
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