Intracellular Localisation of Hypericin in Human Glioblastoma and Carcinoma Cell Lines

Abstract

Hypericin, a natural polycyclic quinone extracted from Hypericum perforatum, has been recently shown to be a powerful sensitiser for photodynamic therapy (PDT). However, its intracellular localisation remains unclear and contradictory. In the present work we compared the intracellular localisation of hypericin in three cultured cell lines (adenocarcinoma cells WiDr, carcinoma cells NHIK 3025 and glioblastoma cells D54Mg) with the distribution of fluorescent probes specific to lysosomes (LysoTracker Blue DND-22), mitochondria (MitoTracker Green FM) and endoplasmic reticulum (ERTracker Blue-White DPX). It was shown that the hypericin staining pattern was different compared to the intracellular distribution of mitochondria or lysosomes. Hypericin was concentrated in the perinucleolar cytoplasmic area mainly on one side of the nucleus--the region rich in endoplasmic reticulum and Golgi. Sometimes nuclear envelope was also stained. Plasma membrane was not stained but the dye was often accumulated in the intercellular space between the tightly contacting WiDr cells in colonies. Hypericin concentrations of 10 microM or less were not toxic for WiDr cells in the dark. Orange light (lambda max approximately 600 nm; 6 mW/cm2) killed the cells stained with 1 microM hypericin with LD50 approximately 1 J/cm2.

Photodynamic therapy of malignant glioma with hypericin: Comprehensive in vitro study in human glioblastoma cell lines

“…Illumination of HYtreated glioblastoma cells (2 h/2.5 μM) at 595 nm with light doses as low as 0.15-0.20 J/cm 2 resulted in a decrease in cell survival to 50% (ID 50 -values); after exposure to 0.4 J/cm 2 cell survival was reduced to about 10% as compared to nonilluminated controls. These results are basically in line with results obtained for other cell lines (11,13,16). In the case of AY-27 cells (transitional cell carcinoma), incubated with HY (0.05-10 μM/2 h) and illuminated (595 nm) of under quite similar experimental conditions as used in the present study, an incubation concentration of about 2.5 μM was required to reduced cell survival to 50% (IC 50 -value) when cells were illuminated with 0.45 J/cm 2 (12).…”

“…by receptor-mediated endocytosis or pinocytosis, rather than by (passive) diffusion. In contrast, no noticeable difference in the HY staining pattern in WiDr cells (human colon carcinoma cell line) was found when comparing fluorescence images after 1 h incubation at 4˚C or at room temperature by Uzdensky et al (16). Consequently, they concluded that HY penetrates cells mostly by diffusion.…”

“…Since contradictory results concerning the cellular uptake mechanism of HY have been published (16,38), this issue was addressed in some more detail. No increase in cellular HY accumulation was observed in U373 MG glioblastoma cells after 2-h incubation at 4˚C (Fig.…”

“…Consequently, phototoxicity of HY was diminished when specific quenchers of ROS were applied (8) or under hypoxic conditions (9). HY-mediated photodynamic inactivation of malignant cell lines of different origin (9)(10)(11)(12)(13)(14)(15), including glioblastoma cells (16,17), has been demonstrated during the last decade.…”

Photochemically stimulated drug delivery increases the cytotoxicity and specificity of EGF–saporin

“…, Saczko et al 2007), Golgi apparatus (Rodal et al 1998, Fabris et al 2001, endoplasmic reticulum (Rodal et al 1998, Uzdensky et al 2001, endosomes and lysosomes (Roberts & Berns 1989, Berg et al 1990b) and the microtubuli (Berg et al 1990a). Clinically relevant PSs for cancer treatment do usually not localize to the nucleus because of their negative charge (Evensen & Moan 1982).…”