Objective To demonstrate the characteristics and contribution of different status and components of rod outer segments (ROS) to the formation of autofluorescence(AF) of lipofuscin.
Methods Different ROS (bleached, unbleached and oxidized-ROS) and different components of ROS (liposome, retinoids and protein, etc.) were added to post confluent, stationary ARPE-19 cell for 7 days. The AF in different groups were taken by FACS and confocal. The characteristics of the different components (A2E, spectral lifetime, etc.) were demonstrated by the spectral analysis and HPLC.
Results Cells fed with HNE oxidized-ROS demonstrated the highest level of AF compared to other groups. Unbleached ROS contribute more than bleached ROS to the formation of AF. Among the different components, lipid instead of retinoids showed higher level of AF. There were small amount of bisretinoids, including A2E, in the RPE cells under the ROS conditions unlike cells exposed to all-trans retinal and other retinoids. In addition, even there is no A2E detected, the liposome still showed strong contribution to the autofluorescence. However, the spectral characteristics of these components have the similar nature but with different lifetime, which showed differences still exist.
Conclustion Ocular lipofuscin has a complex nature, including different fluorophores. In addition to the well-know A2E fluorophore, the oxidative-ROS and lipid may contain more important fluorophores in lipofuscin. The results give more information for the mechanism of formation of lipofuscin and provide potential treatment direction for the age-related macular degeneration and other lipofucin accumulation disease.