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Pawan Kumar Singh, Ajay Kumar, Ashok Kumar; TLR2 and ALX/FPR2 receptor “cross talk” is essential for Resolvin D1 (RvD1)-mediated inflammation resolution and protection in Staphylococcus aureus endophthalmitis.. Invest. Ophthalmol. Vis. Sci. 2016;57(12):No Pagination Specified. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
TLR2 has been shown to play a critical role in providing retinal innate defense against staphylococcal endophthalmitis. In this study, using a mouse model of S. aureus (SA) endophthalmitis, we demonstrate a novel function of TLR2 in regulating inflammation resolution actions of RvD1 via direct interaction with ALX/FPR2 receptor.
Endophthalmitis was induced in C57BL/6 (WT) and TLR2-/- mice via intravitreal injections of SA. The therapeutic effects of RvD1 were assessed by treating SA-infected eyes at 6h or 12h post bacterial infection. The role of ALX/FPR2 receptor was investigated using agonist (WKYMVM) and antagonists (WRW4/PBP10) respectively. The role of TLR2 was investigated using TLR2 agonist (Pam3) and TLR2-/- mice. Diseases progression was monitored via ophthalmoscopic, histological, and microbiological parameters. The expression of inflammatory mediators was assessed by qRT-PCR and/or ELISA. Flowcytometry was used to determine cellular infiltration. The receptor interaction studies were performed using Co-immunoprecipitation (Co-IP) followed by Western blotting.
Intravitreal administration of RvD1 significantly improved the disease outcome in the WT mice. RvD1-mediated protective mechanisms include reduced inflammatory mediators (IL-1β, IL-6, TNF-α, MIP2, and KC), bacterial burden, and diminished PMN infiltration. Interestingly, RvD1 treatment resulted in the induced expression of several anti-inflammatory molecules (IL-4, IL-10, and TGF-β1). RvD1 treatment also showed an increased population of alternatively activated anti-inflammatory macrophages (M2). The presence of ALX/FPR2 antagonist prior to RvD1 treatment abolished its protective effects. In contrast to WT mice, RvD1 treatment failed to induce protection in TLR2-/- mice and had no effect on either pro- or anti-inflammatory mediators. The Co-IP experiments revealed direct physical interaction of TLR2 and FPR2 receptors.
These results indicate, for the first time, that RvD1 promotes the resolution of inflammation and protects the eye from endophthalmitis and that both TLR2 and FPR2 are required for RvD1 to exert its actions. Furthermore, the failure of RvD1 to induce protection in TLR2-/- mice suggests the existence of a novel cross-talk between TLR2 and pro-resolving signaling pathways.
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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