Abstract
Purpose :
ELOVL2 (Elongation of Very Long Chain Fatty Acids-Like 2) encodes a transmembrane protein that plays critical roles in the biosynthesis of omega-3 docosahexaenoic acid (DHA) (22:6n-3) and very long-chain polyunsaturated fatty acids (VLC-PUFAs), which are highly enriched in photoreceptors and essential for maintaining healthy visual function. Adjacent retinal pigment epithelial (RPE) cells are responsible for daily phagocytosis and subsequent degradation of lipid-rich photoreceptor outer segment tips. The aim of this study was to investigate the functional role of ELOVL2 in RPE cells and to what extent alterations of DHA and VLC-PUFA synthesis contribute to RPE dysfunction.
Methods :
We used small interfering RNAs (siRNAs) directed against ELOVL2 in ARPE19 cells and performed immunohistochemistry and fluorescent microscopy studies. To investigate phagocytic function, cells were challenged with FITC-labelled rod outer segments (FITC-ROS) and stained with lysosomal marker LAMP1. The proportion of surface-bound and internalized ROS in control (n=9) and ELOVL2 knockdown (n=9) cells was quantified using ImageJ and normalized to the number of nuclei. Two-tailed Student’s t-test was used for statistical analysis.
Results :
ELOVL2 knockdown ARPE19 cells showed ~30% decrease in phagocytosed ROS (p<0.001) compared to control. In addition, lysosomal size and distribution differed between the two groups. In normal ARPE19 cells, lysosomes exhibited a perinuclear distribution, while in ELOVL2 knockdown ARPE19 cells, LAMP1 signal was localized to the cell periphery.
Conclusions :
Overall, the findings of this study support our hypothesis and suggest that ELOVL2 knockdown and subsequent decline in DHA and VLC-PUFA synthesis result in impaired phagocytic function of RPE cells. This study not only provides insight into the molecular mechanisms of RPE phagocytosis but also opportunities to explore therapeutic treatments for pathologic states such as age-related macular degeneration (AMD) using lipid supplementation.
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.