Relative Contributions of All-Trans and 11-Cis Retinal to Formation of Lipofuscin and A2E Accumulating in Mouse Retinal Pigment Epithelium

PURPOSE: Bis-retinoids are a major component of lipofuscin that accumulates in the retinal pigment epithelium (RPE) in aging and age-related macular degeneration (AMD). Although bis-retinoids are known to originate from retinaldehydes required for the light response of photoreceptor cells, the relative contributions of the chromophore, 11-cis retinal, and photoisomerization product, all-trans retinal, are unknown. In photoreceptor outer segments, all-trans retinal, but not 11-cis retinal, is reduced by retinol dehydrogenase 8 (RDH8). Using Rdh8(−/−) mice, we evaluated the contribution of increased all-trans retinal to the formation and stability of RPE lipofuscin. METHODS: Rdh8(−)(/)(−) mice were reared in cyclic-light or darkness for up to 6 months, with selected light-reared cohorts switched to dark-rearing for the final 1 to 8 weeks. The bis-retinoid A2E was measured from chloroform-methanol extracts of RPE-choroid using HPLC-UV/VIS spectroscopy. Lipofuscin fluorescence was measured from whole flattened eyecups (excitation, 488 nm; emission, 565–725 nm). RESULTS: Cyclic-light-reared Rdh8(−/−) mice accumulated A2E and RPE lipofuscin approximately 1.5 times and approximately 2 times faster, respectively, than dark-reared mice. Moving Rdh8(−/−) mice from cyclic-light to darkness resulted in A2E levels less than expected to have accumulated before the move. CONCLUSIONS: Our findings establish that elevated levels of all-trans retinal present in cyclic-light-reared Rdh8(−/−) mice, which remain low in wild-type mice, contribute only modestly to RPE lipofuscin formation and accumulation. Furthermore, decreases in A2E levels occurring after moving cyclic-light-reared Rdh8(−/−) mice to darkness are consistent with processing of A2E within the RPE and the existence of a mechanism that could be a therapeutic target for controlling A2E cytotoxicity.