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Tomohito Sato, Yoko Karasawa, Naoko Kato, Masataka Ito, Masaru Takeuchi; Oxidative Stress Caused By Continuous Fluorescent Lamp Illumination And/Or Indocyanine Green On Cultured Human Retinal Pigment Epithelial Cells. Invest. Ophthalmol. Vis. Sci. 2012;53(14):4760.
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© ARVO (1962-2015); The Authors (2016-present)
Indocyanine green (ICG) is a photosensitive tricarbocyanine dye, and is generally used for retinal angiography. However, ICG is known to remain in retinal pigment epithelium (RPE)-Bruch's membrane complex after angiography. In this study, we investigated the cellular damage from continuous illumination of visible rays and/or ICG by assessment of modifier oxide via photo-induced oxidative stress.
Human retinal pigment epithelial (RPE; ARPE-19) cells maintained in DMEM/F12 supplemented with 10 % FBS were cultured to confluence in multi-well polystyrene plates. Then, culture medium was changed to a colorless medium (PBS supplemented with 1% FBS, 1 mg/mL glucose, 1 mg/mL CaCl2 and 1 mg/mL MgCl2). The cells were incubated with or without ICG (10 μg/mL) for 24 hours in the dark or under continuous illumination of fluorescent lamp (daylight, 6500 K, 2000 lux). After the incubation, oxidative stresses were evaluated by examining production of total reactive oxygen species (ROS; 2’, 7’-Dichlorodihydrofluorescein assay), protein carbonyl formation (immunoblotting of dinitrophenylhydrazine), lipid peroxidation (immunostaining of malondialdehyde, MDA) and mitochondrial membrane potential (fluorescence intensity of tetramethylrhodamine ester, TRME).
After 24-hour incubation, the levels of total ROS were higher in the cultures with ICG in dark (B), without ICG under illumination (C) or with ICG under illumination (D) than the cultures without ICG in dark (A; control). Molecular weights of protein carbonyls over 240 kDa were detected in C and D but not in A or B. Furthermore, the molecular weights of protein carbonyls in D were higher than in C. The values of lipid peroxidation were almost equal in A and B, higher in C than A (P<0.01), and the highest in D (vs. C; P<0.01). The values of mitochondrial membrane potential in B and C were almost equal to A, but lower in D (P<0.01).
Continuous illumination of visible rays produces ROS in cultured RPE cells. ICG enhanced the ROS production via photo-oxidative stress and damages RPE cells. Persistent ICG in RPE-Bruch's membrane complex after ICG administration could enhance production of total ROS and modifier oxide, and impede functions of mitochondria under the illumination.
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