Abstract
Free all-trans-retinal (atRAL) and retinal pigment epithelium (RPE) lipofuscin are both considered to play etiological roles in Stargardt disease and age-related macular degeneration. A2E and all-trans-retinal dimer (atRAL-dimer) are two well characterized bisretinoid constituents of RPE lipofuscin. In this study, we found that, after treatment of primary porcine RPE (pRPE) cells with atRAL, atRAL-dimer readily formed and accumulated in a concentration- and time-dependent manner, but A2E was barely detected. Cell-based assays revealed that atRAL, the precursor of atRAL-dimer, significantly altered the morphology of primary pRPE cells and decreased cell viability at a concentration of 80 μm regardless of light exposure. By contrast, atRAL-dimer was not cytotoxic and phototoxic to primary pRPE cells. Compared with atRAL and A2E, atRAL-dimer was more vulnerable to light, followed by the generation of its photocleaved products. Moreover, we observed the presence of atRAL-dimer in reaction mixtures of atRAL with porcine rod outer segments (ROS), RPE/choroid, or neural retina. Taken together, we here proposed an alternative metabolic/antidotal pathway of atRAL in the retina: atRAL that evades participation of the visual (retinoid) cycle undergoes a condensation reaction to yield atRAL-dimer in both ROS and RPE. Translocation of atRAL, all-trans N-retinylidene-phosphatidylethanolamine (NR-PE), atRAL-dimer, and photocleavage products of atRAL-dimer from ROS into RPE is accomplished by phagocytosing shed ROS on a daily basis. Without causing damage to RPE cells, light breaks up total atRAL-dimer within RPE cells to release low-molecular-weight photocleavage fragments. The latter, together with ROS-atRAL-dimer photocleavage products, may easily move across membranes and thereby be metabolically eliminated.
Highlights
Free all-trans-retinal and retinal pigment epithelium (RPE) lipofuscin are both considered to play etiological roles in Stargardt disease and age-related macular degeneration
Ultraperformance LC (UPLC)/atmospheric pressure chemical ionization (APCI)-MS analysis of the 430/290-nm adduct from primary porcine RPE (pRPE) cells cultured with 80 M atRAL for 6 h in positive ion mode disclosed an m/z peak at 551.5 (MHϩ) (Fig. 3A, c) that was compatible with that of atRAL-dimer
These results demonstrated that the 430/290-nm pigment of interest in the extract of atRAL-treated primary pRPE cells was atRAL-dimer
Summary
UPLC/APCI-MS analysis of the 430/290-nm adduct from primary pRPE cells cultured with 80 M atRAL for 6 h in positive ion mode disclosed an m/z peak at 551.5 (MHϩ) (Fig. 3A, c) that was compatible with that of atRAL-dimer (theoretically, m/z 551.4, MHϩ). The health of primary pRPE cells cultured with 0, 20, 40, and 80 M of atRAL-dimer for 6 h, 24 h, 3 days, and 5 days was examined in the presence of 220 or 10,000 lx of LED light illumination. In the case of atRAL, morphologic changes of primary pRPE cells and decrease of cell viability were not detected at 20 M, they were both observed at a higher concentration (80 M) with or without light illumination (Fig. 7, A–D).
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
