title: Effects of Benzo(e)Pyrene, a toxic component of cigarette smoke, on human retinal pigment epithelial cells in vitro.
abstract: OBJECTIVE
  To better understand the cellular and molecular basis for the epidemiologic association between cigarette smoke and age-related macular degeneration (AMD), the authors examined the effects of Benzo(e)Pyrene (B(e)P), a toxic element in cigarette smoke, on human retinal pigment epithelial cells (ARPE-19).
  
  
  METHODS
  ARPE-19 cells were cultured in Dulbecco modified Eagle medium containing 10% fetal bovine serum. Cells were treated for 24 hours with 1000 microM, 400 microM, 200 microM, and 100 microM B(e)P. Cell viability was determined by a trypan blue dye-exclusion assay. Activities of caspase-3/7, caspase-8, caspase-9, and caspase-12 were measured by a fluorescence image scanner, and DNA laddering was evaluated by electrophoresis on 3% agarose gel.
  
  
  RESULTS
  The mean percentage of cell viabilities of ARPE-19 cells was decreased in a dose-dependent manner after exposure to B(e)P at the higher concentrations of 1000 microM (20.0 +/- 0.4; P < 0.001), 400 microM (35.6 +/- 6.4; P < 0.001), and 200 microM (58.7 +/- 2.3; P < 0.001) but not at 100 microM (95.9 +/- 0.7; P > 0.05) compared with the equivalent dimethyl sulfoxide (DMSO)-treated control cultures. There were significant increases in caspase-3/7, -8, -9, and -12 activities compared with the DMSO-treated controls (P < 0.001). DNA laddering revealed bands at 200-bp intervals.
  
  
  CONCLUSIONS
  These results show that B(e)P is a toxicant to human retinal pigment epithelial cells in vitro. It causes cell death and induces apoptosis by the involvement of multiple caspase pathways.
authors.affiliation: Department of Ophthalmology, School of Medicine, University of California, Irvine, California 92868, USA.
authors.forename: Ashish
authors.initials: A
authors.lastname: Sharma
authors.forename: Aneesh
authors.initials: A
authors.lastname: Neekhra
authors.forename: Ana L
authors.initials: AL
authors.lastname: Gramajo
authors.forename: Jayaprakash
authors.initials: J
authors.lastname: Patil
authors.forename: Marilyn
authors.initials: M
authors.lastname: Chwa
authors.forename: Baruch D
authors.initials: BD
authors.lastname: Kuppermann
authors.forename: M Cristina
authors.initials: MC
authors.lastname: Kenney
mesh_terms: D017209:Apoptosis; D001580:Benzopyrenes / Q000009:adverse effects*; D020169:Caspases / Q000378:metabolism; D002470:Cell Survival; D002478:Cells, Cultured; D004247:DNA / Q000187:drug effects; D053938:DNA Fragmentation; D004587:Electrophoresis, Agar Gel; D006801:Humans; D008268:Macular Degeneration / Q000209:etiology / Q000235:genetics / Q000473:pathology; D010857:Pigment Epithelium of Eye / Q000187:drug effects* / Q000473:pathology; D012307:Risk Factors; D014028:Tobacco Smoke Pollution / Q000032:analysis*
chemical_list: D001580:Benzopyrenes; D014028:Tobacco Smoke Pollution; C026487:benzo(e)pyrene; D004247:DNA; D020169:Caspases