Phototoxicity Of Indocyanine Green And Brilliant Blue G Under Continuous Fluorescent Lamp Illumination On Cultured Human Retinal Pigment Epithelial Cells

Abstract

Purpose: : Both indocyanine green (ICG) and brilliant blue G (BBG) are used for staining of internal limiting membrane. It has been reported about phototoxicity of ICG but not of BBG. The aim of this study was to compare phototoxicity of ICG and BBG under visible light irradiation on cultured human retinal pigment epithelial (RPE) cells.

Methods: : RPE cells (ARPE-19) were exposed to 0, 0.5 and 5.0 mg/mL ICG or 0, 0.05 and 0.5 mg/mL BBG according to clinical condition for 3 minutes and cultured in a colorless medium (PBS supplemented with 1% FBS, 1 mg/mL glucose, 1 mg/mL CaCl₂, 1 mg/mL MgCl₂) for 24 hours under fluorescent lamp illumination (daylight, 2000 Lux) or in the dark. After microscopical observation, cell viability was evaluated by measuring mitochondrial reductase activity, and the amount of cell death was evaluated by the assay of lactate dehydrogenase released into the culture supernatant from dead cells. Absorbance spectrum of the culture supernatant was measured.

Results: : Morphology of the cells exposed to 5.0 mg/mL ICG and cultured under the illumination changed from polygonal to spherical with enlargement of intercellular space. The cells maintained their original morphology in the other culture conditions. In the cultures with exposure to 5.0 mg/mL ICG and 0.5 mg/mL BBG, the amount of cell death was higher under the illumination than in the dark. In the culture under the illumination, cell viability was lower and the amount of cell death was higher in the culture exposed to 5.0 mg/mL ICG than 0.5 mg/mL BBG, although both values were almost the same when cultured in the dark. In the culture exposed to 5.0 mg/mL ICG, the absorbance at the peak spectral range of the supernatant was higher in the culture in the dark than under the illumination, while the spectra of the supernatant were almost same in the cultures exposed to 0.5 mg/mL BBG in the both culture conditions.

Conclusions: : Persistent BBG induced cytotoxicity on RPE cells in clinical condition in vitro under visible light irradiation as well as persistent ICG, however the degree of phototoxicity was lower. The mechanism of cell damage by BBG with the visible light irradiation might be different from ICG.

Keywords: retinal pigment epithelium • cell survival • clinical laboratory testing