title: Modeling the Dynamic AMD-Associated Chronic Oxidative Stress Changes in Human ESC and iPSC-Derived RPE Cells.
abstract: OBJECTIVE
  Here we use human embryonic stem cells (hESCs) and human-induced pluripotent stem cell (hiPSC)-derived retinal pigment epithelium (RPE) cells to model chronic oxidative stress in vitro. This model allows us to understand the evolution of chronic stress response in RPE in vivo, as well as to monitor microRNAs changes. Finally, we use this in vitro model to identify a partial agonist of NRF2 that is protective against reactive oxygen species (ROS)-induced cytotoxicity.
  
  
  METHODS
  The hESCs and hiPSCs were differentiated toward an RPE fate. Upon maturation, RPE cells were subjected to chronic oxidative stress using Paraquat (PQ). The cells were then analyzed using immunocytochemistry and quantitative RT-PCR to look for changes in gene expression and microRNA changes. Small molecules targeting NRF2 pathways were utilized to look for protection against oxidative stress-induced apoptosis.
  
  
  RESULTS
  We show that 160 μM PQ can be used to generate a model of chronic oxidative stress in RPE cells derived from hESCs and hiPSCs. Using this model, we characterize the NRF2 pathway effectors during the early and late stages of chronic oxidative stress and identify microRNAs changes during oxidative stress. We find that hsa-miR144 modulates NRF2 activity during ROS stress. Lastly, we found a small molecule modulator of NRF2 that plays a protective role against oxidative stress-induced RPE apoptosis.
  
  
  CONCLUSIONS
  In summary, pluripotent stem cell-derived retinal cells can be used to model retinal diseases in a dish. This can provide an unprecedented opportunity to understand the evolution of disease processes and allow us to identify novel therapeutics.
authors.affiliation: Buck Institute for Research on Aging, Novato, California, United States.
authors.forename: Thelma Y
authors.initials: TY
authors.lastname: Garcia
authors.affiliation: Buck Institute for Research on Aging, Novato, California, United States.
authors.forename: Mark
authors.initials: M
authors.lastname: Gutierrez
authors.affiliation: Buck Institute for Research on Aging, Novato, California, United States.
authors.forename: Joseph
authors.initials: J
authors.lastname: Reynolds
authors.affiliation: Buck Institute for Research on Aging, Novato, California, United States 2Department of Ophthalmology, University of Washington, Seattle, Washington, United States.
authors.forename: Deepak A
authors.initials: DA
authors.lastname: Lamba
mesh_terms: D000293:Adolescent; D017209:Apoptosis; D002454:Cell Differentiation; D002460:Cell Line; D002648:Child; D006801:Humans; D057026:Induced Pluripotent Stem Cells / Q000378:metabolism*; D008268:Macular Degeneration / Q000378:metabolism* / Q000473:pathology; D008297:Male; D018384:Oxidative Stress; D017382:Reactive Oxygen Species / Q000378:metabolism; D055213:Retinal Pigment Epithelium / Q000378:metabolism* / Q000473:pathology
chemical_list: D017382:Reactive Oxygen Species