title: UV-A Irradiation Activates Nrf2-Regulated Antioxidant Defense and Induces p53/Caspase3-Dependent Apoptosis in Corneal Endothelial Cells.
abstract: OBJECTIVE
  To examine whether Nrf2-regulated antioxidant defense and p53 are activated in human corneal endothelial cells (CEnCs) by environmental levels of ultraviolet A (UV-A), a known stimulator of oxidative stress.
  
  
  METHODS
  Immortalized human CEnCs (HCEnCi) were exposed to UV-A fluences of 2.5, 5, 10, or 25 J/cm2, then allowed to recover for 3 to 24 hours. Control HCEnCi did not receive UV-A. Reactive oxygen species (ROS) were measured using H2DCFDA. Cell cytotoxicity was evaluated by lactate dehydrogenase (LDH) release. Levels of Nrf2, HO-1, NQO-1, p53, and caspase3 were detected by immunnoblotting or real-time PCR. Activated caspase3 was measured by immunoblotting and a fluorescence assay.
  
  
  RESULTS
  Exposure of HCEnCi to 5, 10, and 25 J/cm2 UV-A increased ROS levels compared with controls. Nrf2, HO-1, and NQO-1 mRNA increased 1.7- to 3.2-fold at 3 and 6 hours after irradiation with 2.5 and 5 J/cm2 UV-A. At 6 hours post irradiation, UV-A (5 J/cm2) enhanced nuclear Nrf2 translocation. At 24 hours post treatment, UV-A (5, 10, and 25 J/cm2) produced a 1.8- to 2.8-fold increase in phospho-p53 and a 2.6- to 6.0-fold increase in activated caspase3 compared with controls, resulting in 20% to 42% cell death.
  
  
  CONCLUSIONS
  Lower fluences of UV-A induce Nrf2-regulated antioxidant defense and higher fluences activate p53 and caspase3, indicating that even near-environmental levels of UV-A may affect normal CEnCs. This data suggest that UV-A may especially damage cells deficient in antioxidant defense, and thus may be involved in the etiology of Fuchs' endothelial corneal dystrophy (FECD).
authors.affiliation: Schepens Eye Research Institute Massachusetts Eye and Ear, Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts, United States.
authors.forename: Cailing
authors.initials: C
authors.lastname: Liu
authors.affiliation: Schepens Eye Research Institute Massachusetts Eye and Ear, Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts, United States.
authors.forename: Dijana
authors.initials: D
authors.lastname: Vojnovic
authors.affiliation: Wellman Center for Photomedicine, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts, United States.
authors.forename: Irene E
authors.initials: IE
authors.lastname: Kochevar
authors.affiliation: Schepens Eye Research Institute Massachusetts Eye and Ear, Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts, United States.
authors.forename: Ula V
authors.initials: UV
authors.lastname: Jurkunas
mesh_terms: D017209:Apoptosis / Q000502:physiology / Q000528:radiation effects*; D015153:Blotting, Western; D053148:Caspase 3 / Q000502:physiology*; D002460:Cell Line; D004307:Dose-Response Relationship, Radiation; D004728:Endothelium, Corneal / Q000378:metabolism / Q000502:physiology / Q000528:radiation effects*; D006801:Humans; D015151:Immunoblotting; D051267:NF-E2-Related Factor 2 / Q000502:physiology*; D017382:Reactive Oxygen Species / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction; D015398:Signal Transduction / Q000502:physiology; D016159:Tumor Suppressor Protein p53 / Q000502:physiology*; D014466:Ultraviolet Rays / Q000009:adverse effects*
chemical_list: D051267:NF-E2-Related Factor 2; C495635:NFE2L2 protein, human; D017382:Reactive Oxygen Species; D016159:Tumor Suppressor Protein p53; D053148:Caspase 3