Abstract
Purpose :
We previously showed that topical application of RCI001 had excellent anti-inflammatory and antioxidant effects in dry eye disease and ocular chemical burn model. In this study, we investigated the inhibiting effects of RCI001 on Rac1 and NLRP3 inflammasome in vitro and in vivo model.
Methods :
We confirmed Rac1 activity by RCI001 treat in RAW264.7 cells and Swiss 3T3 cells by GST (glutathione-S-Transferase) pull-down assay and G-protein activation assay kit. And the production and gene expression changes of IL-1β, IL-6 and TNF-α by RCI001 were quantified by ELISA and real-time PCR on RAW264.7 cell stimulated by LPS. In the mouse ocular alkali burn model, RCI001 was administered via eye drop (10mg/ml, twice daily) for 5 days. As a positive control, prednisolone acetate (PDE) ophthalmic suspension 1% was used. Corneal epithelial integrity (on Day 0, 3, 4 and 5) and histological changes were compared between RCI001 and PDE. Gene and Protein levels of Rac1, NLRP3, Caspase-1 and IL-1β were quantified by real-time PCR and western blotting in corneal tissues collected on day 3 and 5.
Results :
RCI001 inhibited Rac1 activity and various inflammatory cytokines dose-dependently in LPS-stimulated murine macrophage. Additionally, RCI001 restored corneal epithelial integrity and thickness faster than PDE in chemically injured cornea. Activation of Rac1 and NLRP3 inflammasome/ IL-1β axis was suppressed in RCI001 group compared with saline group especially in early phase of ocular alkali burn model.
Conclusions :
Topical RCI001 suppressed expressions of activated Rac1 and inflammatory cytokines in vitro and restored injured cornea quickly by inhibiting the activation of Rac1 and NLRP inflammasome/ IL-1β axis in vivo. We believe that RCI001 could be a promising therapeutic agent for ocular surface disease.
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.