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Zhiyuan Yu, Stephen C Pflugfelder, Jehan Alam, Cintia S De Paiva; Activation of NFkB and NLRP3 Inflammasome in the corneal epithelium following short-term desiccating stress. Invest. Ophthalmol. Vis. Sci. 2021;62(8):1299.
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To determine if 24-hour exposure to desiccating stress activates NFkB and NLRP3 inflammasome pathways in the mouse cornea epithelium.
6-to-8-week-old C57BL/6J mice were housed under normal humidity (nonstressed) or subjected to desiccating stress from a drafty, low humidity environment combined with SC scopolamine QID for 1 day to suppress tear production (DS1). Eyes were embedded and sectioned for immunofluorescent staining or the corneal epithelium was scraped for immunoassay, caspase-1 assay, western blot or real time PCR. A TransAM kit was used to detect phospho-NFkB p65 in the nuclear fraction. Western blot and real-time PCR were performed to detect NFkB and NLRP3 pathway proteins and gene transcripts, respectively.
There was significantly increased nuclear p-p-65 protein and gene expression of the NFkB inducible cytokine IL-12 (1.9-fold) and its receptors (IL-12Rb1 and IL-12Rb2, 3 and 1.9-fold, respectively) in DS1 compared to nonstressed control. NLRP3 and Caspase 1 proteins (6.4 and 3.1-fold, respectively), RNA transcripts (1.4 and 1.9 fold, respectively), as well caspase 1 enzyme activity (1.5-fold, P=0.006) increased in DS1. IL-18, the inflammasome inducible cytokine protein increased 3.1-fold, while RNA expression of its receptor IL-18Rap increased 1.5-fold (P=0.001). There was no change in IL-1b expression. Immunostaining for NLRP3, Caspase 1 and IL-18 increased in the cornea epithelium at DS1 compared to nonstressed.
These findings indicate that two key innate inflammatory pathways (NFkB and NLRP3 inflammasome) are activated following short term desiccating stress. Cytokines stimulated by these pathways, IL-12 and IL-18, are required for full expression of the dry eye associated cytokine IFN-γ by T and natural killer cells.
This is a 2021 ARVO Annual Meeting abstract.
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