Abstract
Purpose :
While the retinal pigment epithelium (RPE) has been regarded as the primary site of pathology for age-related macular degeneration (AMD), the molecular mechanism responsible for initiating the disease has not been fully elucidated. To address this gap in knowledge, we identified changes in the proteome of RPE from donors diagnosed with early AMD compared with healthy age-matched controls.
Methods :
Donor eyes were obtained from the Lions Gift of Sight (St. Paul, MN). Evaluation of the presence and severity of AMD was done using criteria for eyebank eyes (RPE pigment changes, drusen size and location) established by the Minnesota Grading System. RPE harvested from donors with early AMD (N = 45) and healthy age-matched controls (N =32) were fractionated to isolate organelles. Proteins were analyzed by Ultra-high-resolution (UHR) Orbitrap mass spectrometry and quantified using UHR-IonStar, a label-free quantitative proteomics platform. Annotation of protein function and enrichment analysis were performed using Metascape. Upstream analysis was performed using Ingenuity Pathway Analysis (IPA).
Results :
A total of 5,941 proteins were quantified, with 132 significantly altered (90 up-regulated and 42 down-regulated) in RPE from donors with early AMD. The top two proteins highly upregulated in early AMD RPE were Kallistatin and Cathepsin S, which were 7.3- and 5.7- fold increased, respectively, compared with controls. The most significantly downregulated proteins in early AMD RPE were PARP16, ORML2/ORML3, decreased to 24% and 56%, respectively, compared with controls. Enrichment analysis indicated significant upregulation of protein translation, lipid metabolism, and mitochondrial translation, organization, and metabolism. Downregulated pathways include small GTPase-mediated signal transduction and negative regulation of sphingolipid biosynthesis. IPA analysis identified predicted upstream regulators associated with protein translation (LARP1, MLXIPL and MYC), ATP metabolic processes (ESRRG, RORC), and fatty acid biosynthesis (NCOA2).
Conclusions :
During the transition to early AMD, the RPE exhibit adaptive changes in protein content, which is reflected by the observed increase in protein translation. These changes can counteract detrimental conditions of early AMD by increasing lipid biosynthesis and mitochondrial metabolism.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.