Lysosomes Are Sites of Fluoroquinolone Photosensitization in Human Skin Fibroblasts: A Microspectrofluorometric Approach*

Abstract

The fluoroquinolone antibiotics are widely used despite their strong phototoxicity under solar UV irradiation. Although they are known as good photodynamic photosensitizers, other factors than production of activated oxygen species may play a role in the effectiveness of the phototoxic effect. Subcellular localization is one of the important parameters that may determine this strength. Using microspectrofluorometry, it is shown that norfloxacin, ofloxacin, lomefloxacin, ciproflaxin and BAYy3118 are readily incorporated into lysosomes of HS68 human skin fibroblasts although weak staining of the whole cytoplasm also occurs especially with norfloxacin. Consistent with their photoinstability in solutions, the fluoroquinolones under study are readily photobleached by UVA in the HS68 fibroblasts. The BAYy3118 derivative that has the fastest bleaching rate also shows the strongest phototoxicity toward HS68 fibroblasts. Photosensitization with these fluoroquinolones induces lysosomal membrane damage as shown by the increased rate of leakage of the lysosomal probe lucifer yellow as compared to that observed with untreated cells.

Photophysical and Phototoxic Properties of the Antibacterial Fluoroquinolones Levofloxacin and Moxifloxacin

“…These data are in good agreement with the data reported by Wagai and Tawara [6] [7], who have demonstrated that the phototoxicity associated with quinolones in balb/c mice was initiated by the generation of ROS in the target tissue, which specifically involved hydroxyl radical, as well superoxide anion and hydrogen peroxide. Moreover, it has also been reported that the initial steps in the quinolone phototoxicity probably involve hydroxyl-radical attack on the cell membrane and mitochondria [32]. In this context, we have demonstrated a gradual loss of the mitochondrial potential (Dy m ) for LVX and OFX, suggesting that the mitochondria are readily damaged by photosensitization with these fluoroquinolones.…”

“…As recently demonstrated [19], lysosomes are major sites of localization of fluoroquinolones although other subcellular organelles can also be the target of fluoroquinolones. In fact, we also found that, upon fluoroquinolone addition, fluorescence in 3T3 cells was partially localized in mitochondria (data not shown), and co-localized with that of TMRM, a lipophilic cation commonly used for the assessment of the mitochondrial potential (Dy m ) [20] [21].…”

“…To investigate which cellular sites are involved in the phototoxicity induced by fluoroquinolones, we focused our attention on mitochondria. It has been previously shown that mitochondrial alterations are involved in cell death caused by many photosensitizers including fluoroquinolones [19].…”

Pyrrolo[3,2‐h]quinazolines as Photochemotherapeutic Agents

“…Lysosomal alterations can be involved in cell death caused by many photosensitizers such as certain fluoroquinolones, [20] acri- dine orange, [21] and some psoralen analogues. [7] To investigate the integrity of lysosomes after irradiation in the presence of 11 f and 11 g, flow cytometric analysis was performed with the fluorescent dye acridine orange (AO).…”

Effects of photodynamic therapy on the endocytic pathway

“…Santus’ group had earlier demonstrated that lysosomal photodamage mediated by fluoroquinolones could lead to impaired endocytosis 10,11. It was proposed that this result derived from release of lysosomal proteases into the cytoplasm, since the irreversible cysteine-protease inhibitor E64d prevented the impairment.…”