Sitagliptin inhibits neuroinflammatory processes in retinal microglia

Abstract

Purpose: The breakdown of blood-retinal barrier (BRB) is a hallmark of diabetic retinopathy, and it has been correlated with increased nitric oxide (NO) production, mainly via inducible nitric oxide synthase (iNOS). Reactive microglia, which produce high amounts of NO, have been implicated in the pathogenesis of diabetic retinopathy. Sitagliptin is an inhibitor of dipeptidyl-peptidase-IV (DPP-IV) used for the treatment of type 2 diabetes. However, sitagliptin is also able to prevent the BRB breakdown in type 1 diabetic animals, likely by exerting anti-inflammatory effects. In this study, we investigated the potential anti-inflammatory effects of sitagliptin in retinal microglial cells.

Methods: We used primary retinal cell cultures and retinal organotypic cultures. Both cultures were exposed for 24h to lipopolysaccharide (LPS) to trigger an inflammatory response. iNOS immunoreactivity (iNOS-IR) in retinal microglia and microglia morphology (area, perimeter, Feret’s diameter and circularity) were evaluated by immunofluorescence. Images were acquired by confocal microscopy at the level of the ganglion cell layer. NO production was evaluated in primary cultures media by Griess reaction method.

Results: In primary retinal cell cultures, exposure to LPS increased iNOS-IR in microglial cells (1011±117% of control; p=0.001) and sitagliptin inhibited this increase (368.1±30% of control; p=0.01). LPS also triggered and increase in NO production (5.76±0.56 µM; p=0.001), which was inhibited by sitagliptin (3.80±0.3 µM; p=0.05). In retinal organotypic cultures, LPS also increased iNOS-IR in microglial cells (2960±289.3% of control; p=0.01) and this effect was inhibited by sitagliptin (1535±137.9% of control; p=0.01). Moreover, LPS induced alterations in microglia in several morphological parameters (for example, circularity index significantly increased from 0.20±0.01 µm in control conditions to 0.33±0.1 µm with LPS; p=0.01). Sitagliptin prevented the alterations triggered by LPS in microglia morphology.

Conclusions: These results indicate that sitagliptin has anti-inflammatory effects by controlling the reactivity of retinal microglial cells, as evidenced by its capacity to inhibit microglia morphological changes as well as the upregulation in iNOS-IR in microglia and NO production triggered by LPS.<br /> FCT (PTDC/NEU-OSD/1113/2012 and PEst-C/SAU/UI3282/2013), Portugal, and COMPETE-FEDER, and AIBILI.