title: Visible light may directly induce nuclear DNA damage triggering the death pathway in RGC-5 cells.
abstract: OBJECTIVE
  Visible light has been previously demonstrated to induce retinal ganglion cell (RGC)-5 cell death through the mitochondrial pathway. The present study was designed to determine whether visible light might also directly trigger the death pathway by damaging nuclear DNA.
  
  
  METHODS
  RGC-5 cells were exposed to various intensities and durations of visible light exposure. Cell viability and death were monitored with the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and propidium iodide staining. Nuclear DNA damage caused by light was determined with the plasmid assay, genome DNA assay, and in situ terminal deoxynucleotidyl transferase dUTP nick end labeling. The subsequent activation of nuclear enzyme poly(ADP-ribose) polymerase-1 (PARP-1) was measured with western blot, and PARP-1's role in the death pathway was assessed by using specific inhibitors. Poly (ADP-ribose) glycohydrolase and apoptosis-inducing factor (AIF) inhibitors were used to show their influence on light-induced cell death. Calcium influx was examined with the fura-2 assay and calcium channel blocker.
  
  
  RESULTS
  We found that visible light induced RGC-5 cell death in a time- and intensity-dependent manner. After the light intensity was increased to 2,600 lx, activation of the death pathway in RGC-5 cells was clearly observed by detecting double-strand DNA breaks and nuclear DNA damage in vitro. Nuclear enzyme PARP-1 was promptly activated after exposure to 2,600 lx of light for 2 days, and specific inhibitors of PARP-1 had significant neuroprotective effects. The poly(ADP-ribose) glycohydrolase inhibitor tannic acid and AIF inhibitor N-phenylmaleimide partially protected RGC-5 cells from light injury. A massive calcium influx was detected after 2 days of light exposure, and a calcium channel blocker partially protected cells against light injury.
  
  
  CONCLUSIONS
  These results suggest that visible light exposure may directly cause nuclear DNA damage, which consequently activates PARP-1. In addition, RGC-5 cells damaged by 2,600 lx of light exposure can be used as an appropriate cell death model for screening neuroprotective drugs, since this treatment induced remarkable cell death within 2 days. Moreover, these results show that 2,600 lx of light exposure provides a more apparent activation of the death pathway than 1,000 lx of light exposure, which was used in a previous study.
authors.affiliation: Department of Ophthalmology, Second Hospital of JiLin University, ChangChun, China. liguangyu_box@yahoo.com.cn
authors.forename: Guang-Yu
authors.initials: GY
authors.lastname: Li
authors.forename: Bin
authors.initials: B
authors.lastname: Fan
authors.forename: Tong-Hui
authors.initials: TH
authors.lastname: Ma
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D051033:Apoptosis Inducing Factor / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D053903:DNA Breaks, Double-Stranded / Q000187:drug effects / Q000528:radiation effects*; D004307:Dose-Response Relationship, Radiation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005456:Fluorescent Dyes; D016257:Fura-2; D015870:Gene Expression / Q000187:drug effects / Q000528:radiation effects*; D006026:Glycoside Hydrolases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D020287:In Situ Nick-End Labeling; D008027:Light; D008301:Maleimides / Q000494:pharmacology; D010957:Plasmids; D000071137:Poly (ADP-Ribose) Polymerase-1; D000067856:Poly(ADP-ribose) Polymerase Inhibitors; D011065:Poly(ADP-ribose) Polymerases / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D017382:Reactive Oxygen Species / Q000037:antagonists & inhibitors / Q000378:metabolism; D012165:Retinal Ganglion Cells / Q000166:cytology / Q000187:drug effects / Q000201:enzymology / Q000528:radiation effects*; D015398:Signal Transduction / Q000187:drug effects / Q000528:radiation effects*; D013634:Tannins / Q000494:pharmacology
chemical_list: C494831:Aifm1 protein, rat; D051033:Apoptosis Inducing Factor; D004791:Enzyme Inhibitors; D005456:Fluorescent Dyes; D008301:Maleimides; D000067856:Poly(ADP-ribose) Polymerase Inhibitors; D017382:Reactive Oxygen Species; D013634:Tannins; C028525:N-phenylmaleimide; C499322:Parp1 protein, rat; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases; D006026:Glycoside Hydrolases; D002118:Calcium; D016257:Fura-2