Abstract
Purpose :
We have previously shown that in the chick eye, choroidal retinoic acid synthesis is exclusively regulated by expression of the enzyme, retinaldehyde dehydrogenase 2 (RALDH2). In chicks and humans, RALDH2 is synthesized by a population of uncharacterized extravascular stromal cells. RALDH2+ cells markedly increase in number over 1 – 7 days of recovery from induced myopia resulting in increased choroidal retinoic acid synthesis and slowing of ocular growth. The aim of this study was to identify the RALDH2+ cell type(s) in the choroid and determine how these cells modulate retinoic acid concentrations during recovery.
Methods :
Eyes were isolated from control and treated chick eyes following 4- 7 days of unrestricted vision preceded by 10 days of form deprivation (recovery). Choroids were co-immunolabeleld with anti-chicken RALDH2 antibodies together with antibodies specific for macrophages (Ia antigen, KuL01, MHC-II, IgY), smooth muscle/myofibroblast proteins (αSMA, smoothelin, desmin, myocardin), hematopoietic cells (CD-45, TCRgd, CD5, GRL2), neurons (neuron-specific beta III tubulin, NOS, tyrosine hydroxylase), pericytes (NG2), fibroblasts [collagen type 1 (Col1), vimentin] and endothelial cells (PECAM-1). Additionally, a group of chicks received 5-bromo-2′-deoxyuridine (BRDU; 300mg/kg/day, i.p.) for 1-3 days prior to choroid isolation and proliferating RALDH2+ cells were identified by co-immunolabelling with anti-chick RALDH2 and anti-BRDU. Choroidal whole mounts were imaged using confocal microscopy.
Results :
Immunohistochemical analyses indicated that the vast majority of RALDH2+ co-expressed Col1. A minor population of RALDH2+ cells co-labeled with the Ia antigen, indicating similarities with thymic macrophages/dendritic cells, but were negative for the macrophage markers KuL01, MHC-II, and IgY. Some RALDH2+ cells also expressed α-smooth muscle actin (αSMA) but were negative for other muscle/myofibroblast proteins. RALDH2+ cells did not co-localize with any other antibodies tested. The percentage of RALDH2+/BRDU+ choroidal cells was ≈ 3X higher in 4 day recovering choroids as compared with controls suggesting that RALDH2 activity is partially controlled by proliferation of RALDH2+ cells.
Conclusions :
Our results indicate that choroidal retinoic acid synthesis is largely mediated by the proliferation of collagen 1a1+/RALDH2+ perivascular stromal cells.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.