Effect And Mechanism Of Resolvin D1 In Inflammatory Responses In Mice Corneas Treated With Lipopolysaccharide

ObjectiveTo observe the expression of cytokines and chemokines in a murine model of lipopolysaccharide (LPS)-induced corneal inflammation, to investigate the effect of resolvin D1 (RvD1), which is an anti-inflammatory and proresolving lipid mediator biosynthesized from the omega-3-polyunsaturated fatty acid docosahexaenoic acid (DHA) on corneal inflammation, to find possible mechanism and signaling pathway may be involved in its role in corneal inflammation.MethodThere are two parts in this research. In the first part, the corneal epithelium was scraped and LPS was applied to established the murine model of corneal inflammation. Corneal haze and re-epithelialization were measured by stereomicroscopy. Nrutrophil infiltrated were determind by HE staining and immunohistochemistry, and gene expression of corneal IL-1β,IL-6, macrophage chemoattractant protein (MCP)-1 and macrophage inflammatory protein (MIP)-2 was assessed with real-time polymerase chain reaction (PCR) before and 1,3,5days after stimulated by LPS. In the second part, ninety C57BL/6 mice were randomly divided into 3 groups. The does of RvD1 for three group were 0 ng/d (control group),50 ng/d (RvD1-50 group) and 500 ng/d (RvD1-500 group). We examined the effect of intervention RvD1 in vivo. Corneal haze and re-epithelialization were measured in 1,3,5days. Corneal inflammatory infiltration were observed by HE staining and immunohistochemistry. The expression levels of IL-1β, IL-6, MCP-1 and MIP-2 were determined by real-time PCR and enzyme-linked immunosorbent assay (ELISA). The expression of endogenous inhibitory factor of NF-κB signaling pathway, IκBα was evaluated by western blot.Result1. The corneal haze was detected by LPS stimulation. Immunohistochemical analyses indicated that more neutrophils were present in corneas.2. Enhanced mRNA expression of proinflammatory cytokines and chemokines, such as IL-1βp, IL-6, MCP-1 and MIP-2 was observed in 1 day after LPS stimulation, and significantly decreased in 5 days.3. Topical administration of RvD1 resulted in a significant reduction in the severity and incidence of corneal haze, and increased the re-epithelialization rate in the RvDl-treated animals compared to their untreated counterparts. Infiltration of fewer neutrophils into the cornea were observed by HE staining and immunohistochemistry with RvD1 treatment.4. Treatment with RvD1 diminished the production of IL-1β, IL-6, MCP-1 and MIP-2 in the corneas. Down-regulation of these proinflammatory cytokines and chemokines due to RvD1 was proportional to its concertration.5. Importantly, treatment with RvD1 increased the production of the IκBa.ConclusionIn the pathological damage process of corneal tissue, corneal haze, stromal edema and neutrophil infiltration were detected in a murine model of LPS-induced corneal inflammation. IL-1β, IL-6, MCP-1 and MIP-2 may play an important role that promote leukocyte infiltration, which promoting corneal inflammation. RvD1 inhibited the expression of IL-1β, IL-6, MCP-1 and MIP-2. RvD1 inhibited the synthesis of proinflammatory cytokines and chemokines by corneal fibroblasts in association with suppression of NF-κB signaling pathway. RvD1 may therefore prove effective for the treatment of corneal inflammatory diseases.