Retinyl Ester Synthase Activities in Müller Cells from Rod- and Cone-dominated Retinas

Abstract

Purpose: : Recent reports have suggested cone visual pigments regenerate via a retinal biochemical pathway involving Müller cells. Research from our laboratory showed that chicken retinal membranes synthesized 11-cis retinyl ester from all-trans retinol in the presence of plamitoyl coenzyme A; demonstrating retinol isomerase and retinyl ester synthase (ARAT) activities in the retina. We hypothesize that this synthase activity is present in retinal Müller cells of both cone- and rod-dominated retinas.

Methods: : In this study, rat Müller cells (rMC-1), human Müller cells, primary chicken Müller cells, primary bovine Müller cells, and primary mouse Müller cells were cultured and grown until confluence at 37°C/5% CO₂. Comparisons were made between cells incubated with 11-cis retinol or all-trans retinol in culture.Under dark/red light conditions, cells were incubated with all-trans or 11-cis retinol + Bovine Serum Albumin (BSA) for 24h at 37°C + 5% CO₂. Cells were harvested and homogenates prepared. Reaction products were extracted and analyzed using high performance liquid chromatography (HPLC). Products were extracted and analyzed by HPLC. Immunocytochemistry was performed on each Müller cell type using GFAP for positive verification of Müller cells.

Results: : : Incubation of rMC1 cells with all-trans retinol yielded 60 pmol/mg of all-trans retinyl esters and incubation with 11-cis retinol yielded 559.6 pmol/mg 11-cis retinyl esters (n=7). Incubation of MIO-MI (human Müller cells) with all-trans retinol yielded 124 pmol/mg of all-trans retinyl esters and incubation with 11-cis retinol yielded 128.5 pmol/mg of 11-cis retinyl esters (n=7). Primary chicken Müller cells incubated with all-trans retinol yielded 47 pmol/mg all-trans retinyl ester and incubation with 11-cis retinol yielded 5.6 pmol/mg of 11-cis retinyl esters (n=7). Primary bovine Müller cells incubated with all-trans retinol yielded 30 pmol/mg of all-trans retinyl esters and incubation with 11-cis retinol yielded 12 pmol/mg of 11-cis retinyl esters (n=4). Primary mouse Müller cells incubated with all-trans retinol yielded 50 pmol/mg of all-trans retinol and incubation with 11-cis retinol yielded 8280 pmol/mg of 11-cis retinyl esters (n = 5).

Conclusions: : rMC-1, human, and primary cultures of both Bovine and Mouse Müller cells have both 11-cis and all-trans retinyl ester synthase activity.

Keywords: Muller cells • retina • enzymes/enzyme inhibitors