A Protective Role for Microglial Cells in TNF -/- Mice in Ischemic Retinopathy

Abstract

Purpose: : Neovascularisation occurs in response to tissue ischemia and growth factor stimulation. In ischemic retinopathies however, new vessels fail to recover the hypoxic tissue; instead, they infiltrate the transparent vitreous for reasons that are poorly defined. In a model of oxygen-induced retinopathy (OIR) TNFα and iNOS, upregulated in retinal Müller and microglial cells in response to tissue ischemia, are cytotoxic and inhibitory to vascular repair. Here we investigated the mechanism for this effect.

Methods: : Wild type C57WT and TNFα ^-/- mice were subject to OIR by exposure to 75% oxygen (postnatal days 7-12). Retinas were subsequently removed during the hypoxic phase of the model (P12-P13). Retinal cell death was determined by TUNEL staining and microglial cells quantified following Z series capture with a confocal microscope. In situ peroxynitrite and superoxide were measured using the fluorescent dyes DCF and DHE. iNOS, nitrotyrosine, arginase and VEGF were analysed by real-time PCR, western blotting or the conversion of radiolabeled arginine.

Results: : In the TNFα ^-/- animals there was a significant reduction in TUNEL positive apoptotic cells in the inner nuclear layer of the avascular retina compared to the WT group. This coincided with an increase in the number of microglial cells which were actively engaged in phagocytosing apopotic debris and displayed low ROS/RNS/NO production and high arginase activity.

Conclusions: : Collectively, our results demonstrate that enhanced vascular recovery in the absence of TNFα is associated with preservation of a microglial cell population, which display an anti-inflammatory phenotype during the early ischemic phase of OIR.

Keywords: hypoxia • nitric oxide • retinal neovascularization