Investigation of Indocyanine Green Toxicity in vitro: Effects of Illumination and Concentration

Abstract

Purpose:: To investigate the pathomechanism of indocyanine green (ICG) cytotoxicity in vitro, lens epithelial cells (LEC), hyalocytes, and retinal pigmentepithelial cells (RPE) were incubated with different concentrations of ICG in the dark and irradiated with different light intensities.

Methods:: LECs, hyalocytes, and RPE cells were prepared from human and porcine eyes and cultured to confluence in 12-well plates or in 96-well microtiter plates. The cells were exposed to ICG dissolved in glucose 5% at 0.1 %, 1%, and 2% concentrations for 1 min, rinsed with fresh medium and than illuminated with a light intensity commonly used in vitreoretinal surgery for 3 min (1mW/cm², 0.18 J/cm²) or with a ten-fold increased light intensity for 3 min and 15 min. Following exposure to ICG cells were cultivated for 4 days before testing viability with the mitochondrial dehydrogenase assay (MTT-test) and vitality with the trypan blue exclusion test. Experiments were repeated in the dark.

Results:: ICG was not toxic to human and porcine LECs, hyalocytes and porcine RPE cells with light intensities commonly used in vitreoretinal surgery. Human RPE cells showed reduced vitality after ICG > 0.1% concentration and 3 min illumination, but the toxic effect of ICG on human cells in vitro may be aggravated by extended post-mortem times (<12 hours) and aged donors. Ten-fold increased light intensity and prolonged illumination lead to decreased viability and vitality of all cell types even after the use of 0.1% ICG-concentration. ICG was not toxic in the dark at any concentration.

Conclusions:: The results suggest that the cytotoxic effect of ICG is mainly caused by light. Possible underlying mechanisms include a photodynamic effect of ICG itself and the generation of cytotoxic degradation products of ICG by light. Although the present techniques for intraocular application of ICG seem to be within the safety margins reduction of light exposure during and after the use of ICG may help to further reduce toxic effects.

Keywords: drug toxicity/drug effects • retinal culture • vitreoretinal surgery