Intracellular events in retinal glial cells exposed to ICG and BBG.
OBJECTIVE
To investigate the intracellular events in retinal glial cells exposed to indocyanine green (ICG) and brilliant blue G (BBG).


METHODS
The human Müller cell line MIO-M1 was exposed to a low dose (0.25 mg/mL) and a clinical dose (2.5 mg/mL) of ICG and a clinical dose (0.25 mg/mL) of BBG for 15 minutes, respectively. To quantify the proliferation and viability of the cells, [(3)H]-thymidine incorporation was measured and cell numbers were counted 24 hours after treatment. Cell morphology was evaluated using phase-contrast microscopy and transmission electron microscopy. The effects of ICG and BBG on phosphorylation of p38 MAPK and cleavage of caspase-9 and caspase-3 were examined by Western blot.


RESULTS
ICG and BBG significantly reduced [(3)H]-thymidine incorporation in MIO-M1 cells compared with the vehicle-treated controls (P < 0.01). Cell number significantly decreased after exposure to ICG at 2.5 or 0.25 mg/mL (P < 0.01) but did not decrease after exposure to BBG at 0.25 mg/mL. Transmission electron microscopy revealed apoptotic changes only in the ICG-treated cells. Prominent p38 MAPK phosphorylation was observed in the presence of ICG, even at the low concentration and within a short time exposure; however, no apparent enhancement was observed in the presence of 0.25 mg/mL BBG. Furthermore, ICG, but not BBG, induced the cleavage of caspase-9 and caspase-3, which was inhibited by an inhibitor of p38 MAPK.


CONCLUSIONS
ICG is toxic to retinal glial cells because it induces apoptosis, involving induction of the caspase cascade through p38 MAPK phosphorylation. In contrast, BBG does not cause apoptosis and thus could be a safer adjuvant during vitreoretinal surgery.
Kawahara
Shuhei
S
Department of Ophthalmology, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
Hata
Yasuaki
Y
Miura
Muneki
M
Kita
Takeshi
T
Sengoku
Akihito
A
Nakao
Shintaro
S
Mochizuki
Yasutaka
Y
Enaida
Hiroshi
H
Ueno
Akifumi
A
Hafezi-Moghadam
Ali
A
Ishibashi
Tatsuro
T
D017209:Apoptosis / Q000187:drug effects; D015153:Blotting, Western; D053148:Caspase 3 / Q000378:metabolism; D053453:Caspase 9 / Q000378:metabolism; D002452:Cell Count; D018929:Cell Culture Techniques; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D004789:Enzyme Activation; D006801:Humans; D007208:Indocyanine Green / Q000633:toxicity*; D046529:Microscopy, Electron, Transmission; D008858:Microscopy, Phase-Contrast; D009457:Neuroglia / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D010766:Phosphorylation; D012160:Retina / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D012394:Rosaniline Dyes / Q000633:toxicity*; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
D012394:Rosaniline Dyes; D048051:p38 Mitogen-Activated Protein Kinases; D053148:Caspase 3; D053453:Caspase 9; D007208:Indocyanine Green; C004692:coomassie Brilliant Blue