title: The oxysterol 27-hydroxycholesterol increases β-amyloid and oxidative stress in retinal pigment epithelial cells.
abstract: BACKGROUND
  Alzheimer's disease (AD) and age-related macular degeneration (AMD) share several pathological features including β-amyloid (Aβ) peptide accumulation, oxidative damage, and cell death. The causes of AD and AMD are not known but several studies suggest disturbances in cholesterol metabolism as a culprit of these diseases. We have recently shown that the cholesterol oxidation metabolite 27-hydroxycholesterol (27-OHC) causes AD-like pathology in human neuroblastoma SH-SY5Y cells and in organotypic hippocampal slices. However, the extent to which and the mechanisms by which 27-OHC may also cause pathological hallmarks related to AMD are ill-defined. In this study, the effects of 27-OHC on AMD-related pathology were determined in ARPE-19 cells. These cells have structural and functional properties relevant to retinal pigmented epithelial cells, a target in the course of AMD.
  
  
  METHODS
  ARPE-19 cells were treated with 0, 10 or 25 μM 27-OHC for 24 hours. Levels of Aβ peptide, mitochondrial and endoplasmic reticulum (ER) stress markers, Ca2+ homeostasis, glutathione depletion, reactive oxygen species (ROS) generation, inflammation and cell death were assessed using ELISA, Western blot, immunocytochemistry, and specific assays.
  
  
  RESULTS
  27-OHC dose-dependently increased Aβ peptide production, increased levels of ER stress specific markers caspase 12 and gadd153 (also called CHOP), reduced mitochondrial membrane potential, triggered Ca2+ dyshomeostasis, increased levels of the nuclear factor κB (NFκB) and heme-oxygenase 1 (HO-1), two proteins activated by oxidative stress. Additionally, 27-OHC caused glutathione depletion, ROS generation, inflammation and apoptotic-mediated cell death.
  
  
  CONCLUSIONS
  The cholesterol metabolite 27-OHC is toxic to RPE cells. The deleterious effects of this oxysterol ranged from Aβ accumulation to oxidative cell damage. Our results suggest that high levels of 27-OHC may represent a common pathogenic factor for both AMD and AD.
authors.affiliation: Department of Pharmacology, University of North Dakota School of Medicine and Health Sciences, 501 North Columbia Road, Grand Forks, North Dakota 58202, USA. oghribi@medicine.nodak.edu
authors.forename: Bhanu
authors.initials: B
authors.lastname: Dasari
authors.forename: Jaya R P
authors.initials: JR
authors.lastname: Prasanthi
authors.forename: Gurdeep
authors.initials: G
authors.lastname: Marwarha
authors.forename: Brij B
authors.initials: BB
authors.lastname: Singh
authors.forename: Othman
authors.initials: O
authors.lastname: Ghribi
mesh_terms: D000544:Alzheimer Disease / Q000378:metabolism; D016229:Amyloid beta-Peptides / Q000378:metabolism; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D002784:Cholesterol / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D006888:Hydroxycholesterols / Q000494:pharmacology*; D007150:Immunohistochemistry; D008268:Macular Degeneration / Q000378:metabolism* / Q000473:pathology; D018613:Microscopy, Confocal; D018384:Oxidative Stress / Q000187:drug effects*; D017382:Reactive Oxygen Species / Q000378:metabolism; D055213:Retinal Pigment Epithelium / Q000378:metabolism* / Q000473:pathology
chemical_list: D016229:Amyloid beta-Peptides; D006888:Hydroxycholesterols; D017382:Reactive Oxygen Species; C076996:27-hydroxycholesterol; D002784:Cholesterol; D002118:Calcium