Time-resolved microspectrofluorimetry and fluorescence lifetime imaging of hypericin in human retinal pigment epithelial cells.
Hypericin is the active ingredient of the off-the-shelf antidepressant St. John's Wort. It is an effective phototoxic agent and its systemic administration at therapeutic doses could induce particular damage in the eye due to continuous light exposure. Hypercin is strongly fluorescent and its fluorescence properties can be monitored to investigate noninvasively its localization and interactions. To this aim, time-resolved microspectrofluorimetry and fluorescence lifetime imaging were used to assess the spectral and temporal properties as well as the spatial distribution of the fluorescence emitted by retinal pigment epithelium (RPE) cells treated with Hyp at concentrations in the micromolar range (0.5-10 microM). In the presence of hypericin, the emission peaks at 600-605 nm and the fluorescence decay is best fitted with three lifetimes (5.5-7 ns, 1.9-2.5 ns and <0.8 ns). Spectral and temporal differences were observed between high (> or =5 microM) and low hypericin concentrations. In particular, upon increasing concentration, the emission spectrum of the slow component broadens and its lifetime shortens. The latter change is observed also when high concentrations are reached locally, due to more efficient localization within the cell.
Taroni
Paola
P
INFM-Department of Physics and IFN-CNR, Politecnico di Milano, Piazza Leonardo da Vinci 32, 20133 Milan, Italy. paola.taroni@fisi.polimi.it
Valentini
Gianluca
G
Comelli
Daniela
D
D'Andrea
Cosimo
C
Cubeddu
Rinaldo
R
Hu
Dan-Ning
DN
Roberts
Joan E
JE
D000873:Anthracenes; D002478:Cells, Cultured; D004847:Epithelial Cells / Q000187:drug effects* / Q000528:radiation effects; D005453:Fluorescence; D006801:Humans; D015591:Microspectrophotometry / Q000379:methods; D010569:Perylene / Q000031:analogs & derivatives* / Q000032:analysis / Q000494:pharmacology; D010857:Pigment Epithelium of Eye / Q000166:cytology / Q000187:drug effects* / Q000528:radiation effects; D011838:Radiation-Sensitizing Agents / Q000494:pharmacology*; D013050:Spectrometry, Fluorescence / Q000379:methods; D013997:Time Factors
D000873:Anthracenes; D011838:Radiation-Sensitizing Agents; D010569:Perylene; C004965:hypericin