Effect of Rpe65 knockout on accumulation of lipofuscin fluorophores in theretinal pigment epithelium

Purpose. In all mammalian species examined to date the retinal pigment epit helium (RPE) has been found to accumulate autofluorescent lysosomal storage bodies (lipofuscin) during senescence. Substantial evidence indicates that retinoids in the RPE-retina complex play a major role in RPE lipofuscin fo rmation. Indeed, at least one RPE lipofuscin fluorophore is derived in part from vitamin A aldehyde. However, the precise mechanisms by which retinoid s modulate RPE lipofuscin accumulation have not been elucidated. In mice wi thout a functional Rpe65 gene, isomerization of all-trans- to 11-cis-retino l is blocked. Experiments were performed to determine whether this impairme nt of retinoid metabolism alters RPE lipofuscin accumulation. Methods. RPE lipofuscin fluorophore content was compared in 12- to 13-month -old Rpe65(+/+), Roe65(+/-), and Rpe65(-/-) mice. Lipofuscin fluorophore co ntent was determined using quantitative fluorometric measurements. RPE lipo fuscin content was also estimated with quantitative Ultrastructural techniq ues. Results. In the Rpe65(-/-) mice, RPE lipofuscin fluorophore accumulation wa s almost abolished. In addition, a significantly reduced accumulation of li pofuscin fluorophores was also observed in the RPe65(+/-) animals. The inab ility of the RPE of Rpe65(-/-) mice to supply 11-cis-retinal from the RPE t o the retinal photoreceptors was accompanied by a massive accumulation of l ipid droplets in the RPE that appeared to contain substantial amounts of re tinoids. Conclusions. These findings indicate that formation of RPE lipofuscin fluor ophores is almost completely dependent on a normal visual cycle. The absenc e of retinal (both all-trans and 11-cis) in Rpe65 knockout mice drastically reduced formation of lipofuscin fluorophores in these animals. Even an exc essive accumulation of retinyl fatty acid esters in the RPE of Rpe65 knocko ut mice did not contribute to lipofuscin accumulation.