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Janet R. Sparrow, Koji Nakanishi, Craig A. Parish; The Lipofuscin Fluorophore A2E Mediates Blue Light–Induced Damage to Retinal Pigmented Epithelial Cells. Invest. Ophthalmol. Vis. Sci. 2000;41(7):1981-1989.
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purpose. To determine whether the lipofuscin fluorophore A2E participates in
blue light–induced damage to retinal pigmented epithelial (RPE) cells.
methods. Human RPE cells (ARPE-19) accumulated A2E from 10, 50, and 100 μM
concentrations in media, the levels of internalized A2E ranging from
less than 5 to 64 ng/105 cells, as assayed by quantitative
high-performance liquid chromatography (HPLC). Restricted zones (0.5-mm
diameter spots) of confluent cultures were subsequently exposed to
480 ± 20-nm (blue) or 545 ± 1-nm (green) light for 15 to 60
seconds. Phototoxicity was quantified at various periods after exposure
by fluorescence staining of the nuclei of membrane-compromised cells,
by TdT-dUTP terminal nick-end labeling (TUNEL) of apoptotic cells and
by Annexin V labeling for phosphatidylserine exposure.
results. Nonviable cells were located in blue light–exposed zones of
A2E-containing RPE cells, whereas cells situated outside the
illuminated areas remained viable. As shown by fluorescence labeling of
the nuclei of membrane-damaged cells and by the presence of
TUNEL-positive cells, the numbers of nonviable cells increased with
exposure duration and as a function of the concentration of A2E used to
load the cells before illumination. The numbers of blue light–induced
TUNEL-positive cells also increased in advance of the increase in
labeling of membrane-compromised cells, a finding that, together with
Annexin V labeling, indicates an apoptotic form of cell death.
Conversely, blue light–exposed RPE cells that did not contain A2E
remained viable. In addition, illumination with green light resulted in
the appearance of substantially fewer nonviable cells.
conclusions. These studies implicate A2E as an initiator of blue light–induced
apoptosis of RPE cells.
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