Photodamage to human RPE cells by A2-E, a retinoid component of lipofuscin

Florian Schütt, Sallyanne Davies, Jürgen Kopitz, Frank G. Holz, Michael E. Boulton

Research output: Contribution to journalArticle

244 Scopus citations


PURPOSE. A fluorescent component of lipofuscin, A2-E (N-retinylidene-N- retinylethanol-amine) has been shown to impair lysosomal function and to increase the intralysosomal pH of human retinal pigment epithelial (RPE) cells. In addition to its lysosomotropic properties A2-E is known to be photoreactive. The purpose of this study was to determine the phototoxic potential of A2-E on RPE cells. METHODS. A2-E (synthesized by coupling all- trans-retinaldehyde to ethanolamine) was complexed to low-density lipoprotein (LDL) to allow for specific loading of the lysosomal compartment. Human RPE cell cultures were loaded with the A2-E-LDL complex four times within 2 weeks. A2-E accumulation was confirmed by fluorescence microscopy and flow cytometry analysis. Acridine orange staining allowed assessment of lysosomal integrity and intralysosomal pH. The phototoxic properties of A2-E were determined by exposing A2-E-free and A2-E-fed RPE cell cultures to short wavelength visible light (400-500 nm) and assessing cell viability and lysosomal integrity. RESULTS. Fluorescence microscopy and flow cytometry analysis demonstrated that the intralysosomal accumulation of A2-E in cultured RPE cells increased with the number of feedings. Acridine orange staining confirmed that the A2-E was located in the lysosomal compartment and induced an elevation of intralysosomal pH. Exposure of A2-E-fed cells to light resulted in a significant loss of cell viability by 72 hours, which was not observed in either RPE cells maintained in the dark or A2-E-free cultures exposed to light. Toxicity was associated with a loss of lysosomal integrity. CONCLUSIONS. A2-E is detrimental to RPE cell function by a variety of mechanisms: inhibition of lysosomal degradative capacity, loss of membrane integrity, and phototoxicity. Such mechanisms could contribute to retinal aging as well as retinal diseases associated with excessive lipofuscin accumulation for example, age-related macular degeneration and Stargardt's disease.

Original languageEnglish (US)
Pages (from-to)2303-2308
Number of pages6
JournalInvestigative Ophthalmology and Visual Science
Issue number8
StatePublished - Jul 24 2000
Externally publishedYes

ASJC Scopus subject areas

  • Ophthalmology
  • Sensory Systems
  • Cellular and Molecular Neuroscience

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    Schütt, F., Davies, S., Kopitz, J., Holz, F. G., & Boulton, M. E. (2000). Photodamage to human RPE cells by A2-E, a retinoid component of lipofuscin. Investigative Ophthalmology and Visual Science, 41(8), 2303-2308.