The Receptor for AGEs (RAGE) Regulates Microglial Infiltration and Lesion Development in a Murine Model of Choroidal Neovascularisation (CNV)

Abstract

Purpose: : Activation of pro-inflammatory pathology is linked to wet age-related macular degeneration (AMD) and CNV. RAGE is a well-characterised pattern-recognition receptor that upon ligand-binding evokes pro-inflammatory cytokine expression. This study has sought to establish if RAGE has a role in CNV by controlling microglia infiltration and associated angiogenesis in the sub-retinal space.

Methods: : C57Bl/6 wild-type (WT) and RAGE knockout mice (RAGE-/-) (n=8/group) were subjected to diode laser photocoagulation (2/3 burns, 50µm spot size, 0.10s duration, 250 mW), rupturing Bruch’s membrane to produce CNV. Retinal flatmounts were analysed for CNV (Isolectin B4) and infiltrating microglia expression markers F4/80, Iba-1and Isolectin B4. In a parallel in vitro study, microglia were exposed to the RAGE ligand S100B and signal transduction leading to cytokine expression assessed.

Results: : CNV lesion size reached maximal diameter at 2 weeks in WT mice. At this time, RAGE -/- mice exhibited significantly reduced CNV lesion size compared to WT controls (p<0.05). Three different populations of microglia (ramified/ inactive, divergent/activating and amoeboid/active) were identified within and proximal to CNV lesions. Active microglia were higher in number in CNV induced mice compared to respective controls, which predominantly possessed inactive microglia. RAGE-/- mice showed significantly less microglia and the activated phenotype less apparent when compared to WT controls. S100B-immunoreactivity was high in CNV lesions and microglia exposed to protein in vitro upregulated RAGE protein. S100B also evoked phosphorylation of the p44/42 pathway, a response blocked by using a RAGE neutralising antibody. S100B also caused cytokine release from microglia.

Conclusions: : This study demonstrates a key role for RAGE in CNV by regulation of infiltrating microglia and pro-inflammatory signalling. Blockade of RAGE could represent an important therapeutic option for outer retinal inflammatory pathology.

Keywords: choroid: neovascularization • receptors • microglia