Abstract
Abstract: :
Purpose: In studies of a2E, a major fluorophore of RPE lipofuscin, we have addressed the ability of light to modulate the formation of the A2E precursor, A2-PE in photoreceptor outer segments. We also examined for the formation of photoisomers with cis olefins at positions other than the C13-14 double bond. Furthermore, in A2E-laden RPE we have investigated blue light induced oxidation of A2E and blue light induced cellular damage. Methods: Outer segments (ROS) isolated from rats exposed to bright light for 6 hours were analyzed by normal phase HPLC. Chloroform/methanol extracts of RPE from human donor eyes were analyzed by reverse phase HPLC as were light exposed samples of HPLC purified A2E. To study blue light irradiated A2E, A2E-laden RPE were exposed to 430 nm light and examined by mass spectrometry. DNA damage was assessed by Comet analysis. Results: HPLC analysis of extracts of ROS isolated from illuminated rats revealed a peak that co-migrated with authentic A2-PE and which was not present when rats were maintained under cyclic lighting. LC-MS analysis of HPLC purified A2E exposed to light and extracts of human RPE revealed that some minor components of RPE lipofuscin having retention times similar to A2E are cis double-bond photoisomers of A2E other than iso-A2E. Analysis of FAB-MS revealed that blue light irradiation of A2E-laden RPE leads to the polyepoxidation of A2E while Comet analysis indicated that A2E photo-products induce DNA damage. Conclusion: Light is an essential factor in A2E formation, A2E isomerization and A2E polyepoxidation with the latter leading to cellular damage.
Keywords: 308 age-related macular degeneration • 567 retinal pigment epithelium