Kawahara Shuhei, Hata Yasuaki, Miura Muneki, Kita Takeshi, Sengoku Akihito, Nakao Shintaro, Mochizuki Yasutaka, Enaida Hiroshi, Ueno Akifumi, Hafezi-Moghadam Ali, Ishibashi Tatsuro
Department of Ophthalmology, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
Invest Ophthalmol Vis Sci. 2007 Oct;48(10):4426-32. doi: 10.1167/iovs.07-0358.
To investigate the intracellular events in retinal glial cells exposed to indocyanine green (ICG) and brilliant blue G (BBG).
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.
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.
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.
研究视网膜神经胶质细胞暴露于吲哚菁绿（ICG）和亮蓝G（BBG）后的细胞内事件。
将人 Müller 细胞系 MIO-M1 分别暴露于低剂量（0.25 mg/mL）和临床剂量（2.5 mg/mL）的 ICG 以及临床剂量（0.25 mg/mL）的 BBG 中 15 分钟。为了量化细胞的增殖和活力，在处理后 24 小时测量[³H] - 胸腺嘧啶核苷掺入量并计数细胞数量。使用相差显微镜和透射电子显微镜评估细胞形态。通过蛋白质印迹法检测 ICG 和 BBG 对 p38 MAPK 磷酸化以及半胱天冬酶 -9 和半胱天冬酶 -3 裂解的影响。
与载体处理的对照组相比，ICG 和 BBG 显著降低了 MIO-M1 细胞中的[³H] - 胸腺嘧啶核苷掺入量（P < 0.01）。暴露于 2.5 或 0.25 mg/mL 的 ICG 后细胞数量显著减少（P < 0.01），但暴露于 0.25 mg/mL 的 BBG 后细胞数量未减少。透射电子显微镜仅在 ICG 处理的细胞中显示出凋亡变化。即使在低浓度和短时间暴露的情况下，在 ICG 存在时也观察到明显的 p38 MAPK 磷酸化；然而，在 0.25 mg/mL 的 BBG 存在时未观察到明显增强。此外，ICG 而非 BBG 诱导了半胱天冬酶 -9 和半胱天冬酶 -3 的裂解，这被 p38 MAPK 的抑制剂所抑制。
ICG 对视网膜神经胶质细胞有毒性，因为它诱导细胞凋亡，包括通过 p38 MAPK 磷酸化诱导半胱天冬酶级联反应。相比之下，BBG 不会引起细胞凋亡，因此在玻璃体视网膜手术期间可能是一种更安全的辅助剂。
