title: Effect of miR-23 on oxidant-induced injury in human retinal pigment epithelial cells.
abstract: OBJECTIVE
  Micro(mi)RNAs negatively regulate a wide variety of genes through degradation or posttranslational inhibition of their target genes. The purpose of this study was to investigate the role of miR-23a in modulating RPE cell survival and gene expression in response to oxidative damage.
  
  
  METHODS
  The expression level of miR-23a was measured in macular retinal pigment epithelial (RPE) cells of donor eyes with aged-related macular degeneration (AMD) and age-matched normal eyes by using qRT-PCR. Cultured human ARPE-19 cells were transfected with miR-23a mimic or inhibitor. Cell viability was assessed by the MTT assay. Apoptosis was determined by incubating cells with hydrogen peroxide (H(2)O(2)) or t-butylhydroperoxide (tBH). Caspase-3 activity and DNA fragmentation were measured by enzyme-linked immunosorbent assays. The protein relevant to apoptosis, such as Fas expression level, was analyzed by Western blot analysis.
  
  
  RESULTS
  miR-23a expression was significantly downregulated in macular RPE cells from AMD eyes. H(2)O(2)-induced ARPE-19 cell death and apoptosis were increased by an miR-23a inhibitor and decreased by an miR-23a mimic. Computational analysis found a putative target site of miR-23a in the 3'UTR of Fas mRNA, which was verified by a luciferase reporter assay. Forced overexpression of miR-23a decreased H(2)O(2) or tBH-induced Fas upregulation, and this effect was blocked by downregulation of miR-23a.
  
  
  CONCLUSIONS
  The protection of RPE cells against oxidative damage is afforded by miR-23a through regulation of Fas, which may be a novel therapeutic target in retinal degenerative diseases.
authors.affiliation: Departments of Ophthalmology and Visual Sciences, University of Texas Medical Branch, Galveston, Texas 77555, USA.
authors.forename: Haijiang
authors.initials: H
authors.lastname: Lin
authors.forename: Jinqiao
authors.initials: J
authors.lastname: Qian
authors.forename: Alexander C
authors.initials: AC
authors.lastname: Castillo
authors.forename: Bo
authors.initials: B
authors.lastname: Long
authors.forename: Kyle T
authors.initials: KT
authors.lastname: Keyes
authors.forename: Guanglin
authors.initials: G
authors.lastname: Chen
authors.forename: Yumei
authors.initials: Y
authors.lastname: Ye
mesh_terms: D000368:Aged; D000369:Aged, 80 and over; D017209:Apoptosis / Q000187:drug effects; D051033:Apoptosis Inducing Factor / Q000378:metabolism; D015152:Blotting, Northern; D015153:Blotting, Western; D053148:Caspase 3 / Q000378:metabolism; D002470:Cell Survival; D002478:Cells, Cultured; D045304:Cytochromes c / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D005786:Gene Expression Regulation / Q000502:physiology*; D006801:Humans; D006861:Hydrogen Peroxide / Q000633:toxicity; D008268:Macular Degeneration / Q000235:genetics* / Q000378:metabolism; D035683:MicroRNAs / Q000235:genetics*; D008875:Middle Aged; D008928:Mitochondria / Q000201:enzymology; D016877:Oxidants / Q000633:toxicity*; D018384:Oxidative Stress; D055213:Retinal Pigment Epithelium / Q000378:metabolism* / Q000473:pathology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014019:Tissue Donors; D014162:Transfection; D019014:fas Receptor / Q000378:metabolism; D020122:tert-Butylhydroperoxide / Q000633:toxicity
chemical_list: C494829:AIFM1 protein, human; D051033:Apoptosis Inducing Factor; C552430:MIRN23a microRNA, human; D035683:MicroRNAs; D016877:Oxidants; D019014:fas Receptor; D045304:Cytochromes c; D020122:tert-Butylhydroperoxide; D006861:Hydrogen Peroxide; D053148:Caspase 3