September 2016
Volume 57, Issue 12
Open Access
ARVO Annual Meeting Abstract  |   September 2016
Aging and antioxidants differentially modulate the effect of phagocytized lipofuscin granules on expression of antioxidant enzymes in ARPE-19 cells
Author Affiliations & Notes
  • Anna K Pilat
    Biophysics, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Krakow, Poland
  • Michal Sarna
    Biophysics, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Krakow, Poland
  • Dawid Wnuk
    Cell Biology, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Krakow, Poland
  • Magdalena Olchawa
    Biophysics, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Krakow, Poland
  • Tadeusz J Sarna
    Biophysics, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Krakow, Poland
  • Footnotes
    Commercial Relationships   Anna Pilat, None; Michal Sarna, None; Dawid Wnuk, None; Magdalena Olchawa, None; Tadeusz Sarna, None
  • Footnotes
    Support  Poland National Science Center (grant Maestro 2013/08/A/NZ1/00194). Faculty of Biochemistry, Biophysics and Biotechnology of Jagiellonian University is a partner of the Leading National Research Center (KNOW) supported by the Ministry of Science and Higher Education.
Investigative Ophthalmology & Visual Science September 2016, Vol.57, 6044. doi:
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      Anna K Pilat, Michal Sarna, Dawid Wnuk, Magdalena Olchawa, Tadeusz J Sarna; Aging and antioxidants differentially modulate the effect of phagocytized lipofuscin granules on expression of antioxidant enzymes in ARPE-19 cells. Invest. Ophthalmol. Vis. Sci. 2016;57(12):6044.

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

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Abstract

Purpose : Aged RPE cells are at elevated risk of oxidative stress due accumulation of lipofuscin (LF). It was postulated that chronic oxidative stress, mediated in the human RPE by LF, could contribute to the development of age-related macular degeneration (AMD).We have previously shown that LF under sub-lethal photic stress conditions photooxidizes cellular proteins in ARPE-19 cells. Here we ask whether LF-mediated oxidative stress modulates expression of antioxidant enzyme proteins such as heme oxygenase-1 (HO-1) and catalase in ARPE-19 cells.

Methods : Lipofuscin granules (LF), isolated from human RPEs from two age groups, were enriched with a combination of zeaxathin and alpha tocopherol (An). Control or antioxidant enriched lipofuscin granules (An-LF) were introduced to APRE-19 cells by phagocytosis. Control cells, An-treated cells or cells with LF and An-LF, irradiated with blue light for selected time intervals, were analyzed by Western blot for selected antioxidant enzymes. To monitor sub-lethal oxidative stress in ARPE-19 cells mediated by phagocytized human RPE lipofuscin atomic force microscopy (AFM) and laser–scanning confocal fluorescence microscopy were employed.

Results : Irradiation of LF containing cells with blue light induced HO-1 protein with the effect being light dose dependent. The extent of HO-1 expression by lipofuscin was higher for lipofuscin isolated from older donors (age: 50-59) compared to younger donors (age: 18-29). Enrichment of lipofuscin granules isolated from both age groups with zeaxathin and alpha tocopherol reduced HO-1protein level. AFM analysis revealed that after blue light-treatment, cells with lipofuscin granules, were significantly softer. Fluorescence analysis showed that lipofuscin-mediated photic stress to RPE cells induced significant reduction in the formation of actin stress fibers.

Conclusions : HO-1 upregulation may help to protect RPE cells form LF-mediated oxidative stress. Phototoxicity of lipofuscin granules in RPE can be modulated by combination of antioxidants. AFM and fluorescence image analysis could be employed as sensitive methods for detecting early sub-lethal photic changes in RPE cells mediated by LF.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.

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