April 2010
Volume 51, Issue 13
Free
ARVO Annual Meeting Abstract  |   April 2010
Effects of Benzo(e)Pyrene, a Toxic Component of Cigarette Smoke, on Müller Cells (MIO-M1) in vitro
R. M. Piche Lopez; N. Gupta; C. A. Ramirez; S. Mansoor; A. G. Limb; B. D. Kuppermann; M. C. Kenney
Author Affiliations & Notes
  • R. M. Piche Lopez
    Ophthalmology, University of California, Irvine, Fountain valley, California
  • N. Gupta
    Ophthalmology, University of California, Irvine, Fountain Valley, California
  • C. A. Ramirez
    Ophthalmology, University of California, Irvine, Fountain Valley, California
  • S. Mansoor
    Department of Chemical and Biomolecular Engineering, Georgia Institute of Technology, Atlanta, Georgia
  • A. G. Limb
    Division of Ocular Biology and Therapeutics, UCL Institute of Ophthalmology, London, UK, United Kingdom
  • B. D. Kuppermann
    Ophthalmology, University of California, Irvine, Fountain Valley, California
  • M. C. Kenney
    Ophthalmology, University of California, Irvine, Fountain Valley, California
  • Footnotes
    Commercial Relationships  R.M. Piche Lopez, None; N. Gupta, None; C.A. Ramirez, None; S. Mansoor, None; A.G. Limb, None; B.D. Kuppermann, None; M.C. Kenney, None.
  • Footnotes
    Support  Discovery Eye Foundation, Guenther Foundation, Lincy Foundation, Iris and B. Gerald Cantor Foundation, Ko Family Foundation, Gilbert Foundation, Research to Prevent Blindness
Investigative Ophthalmology & Visual Science April 2010, Vol.51, 4108. doi:
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      R. M. Piche Lopez, N. Gupta, C. A. Ramirez, S. Mansoor, A. G. Limb, B. D. Kuppermann, M. C. Kenney; Effects of Benzo(e)Pyrene, a Toxic Component of Cigarette Smoke, on Müller Cells (MIO-M1) in vitro. Invest. Ophthalmol. Vis. Sci. 2010;51(13):4108.

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Abstract

Purpose: : To understand the cellular and molecular basis of the association between cigarette smoking and age-related macular degeneration (AMD), we examined the effect of Benzo(e)Pyrene (B(e)P), a toxic element in cigarette smoke, on a Müller cell line (MIO-M1).

Methods: : MIO-M1cells were treated for 24 hours with 200µM, 100µM, 50µM and 25µM of B(e)P. The cytotoxicity was measured by: Cell viability trypan blue dye-exclusion assay, reactive oxygen species/reactive nitrogen species (ROS/RNS) assay, mitochondrial membrane potential (ΔΨ[[Unsupported Character - М]]), caspase-3/7 activity, and DNA fragmentation assay

Results: : All concentrations of B(e)P decreased the cell viability of the MIO-M1 cells:200µM (75.13±1.52, P<0.001), 100µM (84.3±0.16,P<0.001), 50µM (87.18±0.69,P<0.05), and 25µM (89.25±1.47,P<0.05) compared to DMSO equivalent controls 86.9±0.48, 90.3±0.55, 90.8±0.27, 91.3±0.30 respectively. The increase in ROS/RNS production was 2835 ± 190.0, P<0.001 (200µM); 2704 ± 111.7, P<0.001 (100µM); 2339 ± 76.0, P<0.05 (50µM) and 2274 ± 67.6, P<0.05 (25µM), compared to with equivalent DMSO controls 1199±36.1, 1193±49.3, 1369±28.9, 1281±26.4 respectively. The ΔΨ[[Unsupported Character - &#1052;]] was significantly decreased after treatment with B(e)P at 200µM (3.25±0.14, P<0.001), 100µM (4.0±0.14, P<0.001), and 50µM (5.16±0.19, P<0.001),compared to with equivalent DMSO controls 6.64±0.14, 6.44±0.22, 6.78±0.14 and 6.81±0.15 respectively. Caspase-3/7 activity was increased for cells treated with 200µM (9804 ± 150.4, P<0.001), 100µM (8999 ± 146.5, P<0.001), 50µM (6864 ± 283.7, P<0.001) and 25µM (4592 ± 193.4, P<0.05), compared to DMSO controls 4113±80.5, 2619±232.3, 2559±211.9 and 2535±254.2 respectively. There was marked DNA fragmentation in cells exposed to B(e)P at 200 µM only.

Conclusions: : B(e)P, an important toxin of cigarette smoke, causes oxidative damage and apoptosis of MIO-M1 cells. Disruption of these important functions by B(e)P may contribute to the significant risk that smoking has for cells of AMD.

Keywords: drug toxicity/drug effects • retinal pigment epithelium • age-related macular degeneration 
© 2010, The Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Permission to republish any abstract or part of an abstract in any form must be obtained in writing from the ARVO Office prior to publication.
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