April 2011
Volume 52, Issue 14
ARVO Annual Meeting Abstract  |   April 2011
Hydroquinone Changes Complement Factor H And Reactive Oxygen Species Levels In ARPE-19 Cells
Author Affiliations & Notes
  • Ashish U. Sapkal
    Gavin Herbert Eye Institute, UC, Irvine, California
  • Maria F. Estrago-Franco
    Ophthalmology, Clinica Dres Estrago, Corrientes, Argentina
  • Alammaprabhu J. Patil
    Ophthalmology, Royal Lancaster Infirmary,University Hospitals of Morecambe Bay NHS Trust, Lancaster, United Kingdom
  • Vishal R. Sharma
    Gavin Herbert Eye Institute, UC, Irvine, California
  • Claudio A. Ramirez
    Gavin Herbert Eye Institute, UC, Irvine, California
  • Baruch D. Kupperamann
    Gavin Herbert Eye Institute, UC, Irvine, California
  • Cristina M. Kenney
    Gavin Herbert Eye Institute, UC, Irvine, California
  • Footnotes
    Commercial Relationships  Ashish U. Sapkal, None; Maria F. Estrago-Franco, None; Alammaprabhu J. Patil, None; Vishal R. Sharma, None; Claudio A. Ramirez, None; Baruch D. Kupperamann, None; Cristina M. Kenney, None
  • Footnotes
    Support  Discovery Eye Foundation, Lincy Foundation, Polly and Michael Smith Foundation, Research to Prevent Blindness.
Investigative Ophthalmology & Visual Science April 2011, Vol.52, 2315. doi:
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      Ashish U. Sapkal, Maria F. Estrago-Franco, Alammaprabhu J. Patil, Vishal R. Sharma, Claudio A. Ramirez, Baruch D. Kupperamann, Cristina M. Kenney; Hydroquinone Changes Complement Factor H And Reactive Oxygen Species Levels In ARPE-19 Cells. Invest. Ophthalmol. Vis. Sci. 2011;52(14):2315.

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      © ARVO (1962-2015); The Authors (2016-present)

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Purpose: : To evaluate in vitro effects of Hydroquinone (HQ), a cigarette smoke component on Complement Factor H (CFH) levels, Reactive Oxygen Species (ROS) generation and mitochondrial membrane potential (ΔΨm) in human retinal pigment epithelial cells (ARPE-19). CFH inhibits the alternative complement pathway and protects cells against over activated complement components, which might contribute to drusen formation. High oxidative stress initiates apoptosis, a known mechanism of cell death in age-related macular degeneration (AMD).

Methods: : ARPE-19 cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM) containing 10% fetal bovine serum. Cells were treated with 0, 50, 100 or 200µM HQ dissolved in Dimethyl sulfoxide (DMSO) or DMSO alone for 24 hours. Western blot was performed on 10% SDS-polyacrylamide gels using goat polyclonal antibody to CFH. Bands obtained at 85kDa were analyzed by Image Point software and standardized to 100% untreated. ROS production was measured with a fluorescent H2DCFDA assay. ΔΨm was measured with the JC-1 assay.

Results: : CFH levels in 200 and 100µM HQ were lower at 57.09±0.57 (p<0.001) and 64.03±4.35 (p<0.01) respectively, compared to DMSO controls of 95.02±0.59. ROS levels in 200, 100 and 50µM HQ were elevated to 13190±620.0 (p<0.001), 11720±347.3 (p<0.001) and 9834±94.31 (p<0.01) respectively, compared to DMSO levels of 5856±167.3. The ΔΨm values were lowered in 200 and 100µM HQ cultures at 3.24±0.25 (p<0.001) and 3.82±0.19 (p<0.001) respectively, compared to 6.16±0.40 in DMSO cultures. 50µM HQ had ΔΨm values of 5.28±0.49 which was non significant compared to DMSO.

Conclusions: : ARPE-19 cells had decreased CFH levels and ΔΨm values along with increased ROS levels after HQ exposure. This provides biochemical evidence of a relationship between HQ a known component of smoking and inflammatory elements that could be part of the pathogenic mechanisms involved in AMD.

Keywords: age-related macular degeneration • retinal culture • retinal pigment epithelium 

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