The beta-Hydroxybutyrate Receptor GPR109A Functions as an Anti-inflammatory Receptor In Retinal Pigment Epithelium (RPE)

Abstract

Purpose: : GPR109A is a G-protein coupled receptor for nicotinic acid. beta-Hydroxybutyrate is the physiologic ligand. We reported recently expression of the receptor in RPE where it localizes to the basolateral membrane. Other than its anti-lipolytic actions in adipocytes, GPR109A is most noted for is anti-inflammatory properties. We have described GPR109A expression in RPE however, the function of the receptor in this cell type is not known. The purpose of the present study was to investigate the functional role of GPR109A in RPE.

Methods: : Immunofluorescence, real-time quantitative PCR, ELISA and bioluminescence resonance energy transfer (BRET) assays were used to study the expression/function of GPR109A in RPE cells under normal and inflammatory conditions. Human RPE cells (ARPE-19) and primary RPE cells isolated from mouse retina (mRPE) were cultured in the presence of TNF-alpha (10 microgram/ml) to induce inflammation and in the presence or absence of the GPR109A ligands nicotinic acid (1 mM) or beta-hydroxybutyrate (5 mM). The expression/secretion of the pro-inflammatory cytokines MCP-1/Ccl2 and IL-6 was then analyzed.

Results: : qPCR and immunofluorescence analyses demonstrated an upregulation of MCP-1/Ccl2 and IL-6 mRNA and protein in response to TNF-alpha. ELISA analyses showed an associated increase in the secretion of these cytokines into the culture medium. Treatment of cells in the presence of the GPR109A ligands nicotinic acid or beta-hydroxybutyrate suppressed the expression and secretion of IL-6 and MCP-1/Ccl2. Additional mRPE cells were isolated from GPR109A knockout mouse eyes and treated identically. The suppression of IL-6 and MCP-1/Ccl2 expression/secretion was not observed in GPR109A-deficient RPE cells.

Conclusions: : Our data support an anti-inflammatory role for GPR109A in RPE. RPE plays a pivotal role in regulating immunity/inflammation in retina. Expression of a receptor capable of stimulating anti-inflammatory pathways in RPE may have tremendous implications in terms of modulating the retinal inflammatory response and the development of novel treatments for retinal diseases in which inflammation plays a major role.

Keywords: retinal pigment epithelium • inflammation • receptors