Quantitative in vitro assessment of phototoxicity by a fibroblast-neutral red assay

We have adapted the neutral red uptake assay for quantitative assessment of injury to fibroblast cultures by potential phototoxins. Tetracycline derivatives, quinolone derivatives, and chlorpromazine were used as model compounds for development of the assay. Human fibroblasts were incubated with potential phototoxins, the cell cultures irradiated with UV, and the capacity for neutral red uptake determined. Demeclocycline and doxycycline, two known photosensitizers, showed a 94% and 95% decrease of neutral red uptake, respectively, indicating photo-induced cytotoxicity. Minocycline, a non-photosensitizing tetracycline derivative, showed no decrease in uptake. Tetracycline, a weak phototoxin, showed minor (10%) decrease at equivalent concentrations (20 micrograms/ml). Microscopic observation of neutral red uptake and cell damage paralleled the spectrophotometric findings. Chlorpromazine, a non-tetracycline phototoxin, showed 91% decrease. An additional group of phototoxic drugs, quinolone antibacterials, were studied. Nalidixic acid, ofloxacin, ciprofloxacin, and norfloxacin all demonstrated phototoxicity, with nalidixic acid showing the greatest decrease in neutral red uptake. This methodology may provide a useful rapid method to quantify phototoxic potential of new drugs or suspected phototoxins.