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Microfluorometry using fluorescein diacetate reflects the integrity of the plasma membrane in UVA‐irradiated cultured skin fibroblasts
Author(s) -
Skoog MarjaLeena,
Öllinger Karin,
Skogh Marcus
Publication year - 1997
Publication title -
photodermatology, photoimmunology and photomedicine
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.736
H-Index - 60
eISSN - 1600-0781
pISSN - 0905-4383
DOI - 10.1111/j.1600-0781.1997.tb00106.x
Subject(s) - chemistry , lipid peroxidation , trypan blue , fluorescein , membrane , biophysics , photosensitizer , rose bengal , glutathione , intracellular , biochemistry , microbiology and biotechnology , antioxidant , cell , biology , fluorescence , photochemistry , enzyme , physics , organic chemistry , quantum mechanics
The damaging effect of long‐wave ultraviolet radiation (UVA) on the plasma membranes of cultured human skin fibroblasts was evaluated by cytofluorometry performed after vital staining with fluorescein diacetate. The damage was associated with lipid peroxidation, as shown by accumulation of malondialdehyde and 4‐hydroxyalkenals; such accumulation was induced by a UVA dose of only 8 J/cm 2 . Pretreatment with the effective membrane peroxidation inhibitor α‐tocopherol (added in the form of α‐tocopherol succinate) or the singlet oxygen quencher β‐carotene protected the cells from membrane damage. Further, depletion of intracellular glutathione by exposure of the cells to buthionine sulfoximine, an inhibitor of γ‐glutamylcysteine synthetase aggravated the membrane‐damaging effects. The results confirm the photodynamic effects of UVA, which presupposes the excitation of endogenous photosensitizer(s) and the production of reactive oxygen species. The present results indicate that this method of detection of alterations in plasma membrane stability may be highly suitable for studying various photobiological phenomena and for use as a model for testing substances that could protect skin from UVA damage. The trypan blue exclusion test proved to be too insensitive to detect these changes.