Proteome and Secretome Dynamics of Stem Cell-Derived Retinal Pigmented Epithelium in Response to Acute and Chronic ROS

Age-related macular degeneration (AMD) is the leading cause of blindness in developed countries, and is characterized by slow retinal degeneration linked to chronic oxidative stress in the retinal pigmented epithelium (RPE). The exact molecular mechanisms that lead to RPE death and dysfunction in response to chronic reactive oxygen species (ROS) are still unclear. In this work, human stem cell-derived RPE samples were treated with a low dose of paraquat (PQ) for 1 week or 3 weeks to induce chronic reactive oxygen species (ROS) stress. Cells were then harvested and both the intracellular and secreted RPE proteomes were quantified by mass spectrometry. Inside the RPE, chronic ROS caused concerted increase of glycolytic proteins but decreased mitochondrial proteins, as well as decreased extracellular matrix proteins and membrane proteins required for endocytosis. From the secreted proteins, we found that stressed RPE secrete over 1,000 detectable proteins, and the composition of the proteins secreted from RPE changes due to chronic ROS. Notably, secreted APOE is decreased 4-fold due to 3 weeks of chronic ROS stress, and urotensin-II, the strongest known vasoconstrictor, doubles. Further, secreted TGF-beta is increased, and its cognate signaler BMP1 decreased in the secretome. Together, these alterations of the RPE proteome and protein secretome paint a detailed molecular picture of the retinal stress response in space and time.