Abstract
Purpose: :
Human retinal pigment epithelial (RPE) cells, the sub-RPE space, Bruch's membrane and choriocapillaris undergo prominent aging changes not seen in most other mammals. In elderly eyes, age-related deposits may accumulate on both sides of the RPE basement membrane. The origin of the age-related material in Bruch's membrane is thought to include the RPE. Human RPE cells synthesize an extraneous splice isoform of the retinal G protein-coupled receptor (RGR), and an epitope of this exon-skipping protein variant (RGR-d) is released from the RPE into Bruch's membrane in young and old persons. The purpose of this work is to investigate the presence of extracellular RGR-d in drusen and age-related deposits in Bruch's membrane. We investigated the different types of drusen in which the RGR-d epitope is present and analyzed what types of organelle or structure, if any, are associated with RGR-d at the ultrastructural level.
Methods: :
RGR-d was analyzed in human donor eyes by immunohistochemical localization at the light and electron microscopic levels. Pre-embed immunohistochemical localization at the electron microscopic level was performed by peroxidase-based detection of electron dense diaminobenzidine reaction product.
Results: :
We have shown that extracellular RGR-d is present in most types of drusen, including hard, soft, large confluent and early-stage drusen. We have demonstrated the presence of the RGR-d epitope in vesicular deposits (diameter ~100 nm) and membranous debris in the inner collagenous zone and intercapillary regions at the electron microscopic level.
Conclusions: :
These results confirm that particular extracellular deposits in Bruch's membrane indeed originate from the RPE. RGR-d, an RPE-specific marker, is a constituent not only of various types of drusen, but of basal linear deposits consisting of a diffuse layer of vesicular and planar material external to the RPE basement membrane.
Keywords: age-related macular degeneration • drusen • proteins encoded by disease genes