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Nitric oxide as a second messenger in phagocytosis by cultured retinal pigment epithelial cells
J I Kogishi1, M Akimoto, M Mandai
1Department of Ophthalmology and Visual Science, Kyoto University Graduate School of Medicine, Kyoto, Japan.
Purpose:
To investigate a possible role of the nitric oxide (NO)-cGMP signal transduction system in phagocytosis of rod outer segments (ROS) by cultured retinal pigment epithelial (RPE) cells.
Methods:
Primary cultures of RPE cells from 10-day-old Brown Norway rats were used to study the phagocytosis of ROS by these cells. Phagocytosis of ROS was evaluated with or without an inhibitor of nitric oxide synthase (NOS), N(G)-nitro-L-arginine (L-NNA), and the reverse effects of L-NNA by L-arginine and 8-bromo-cGMP on phagocytosis were also studied. NO-associated cGMP production by RPE cells was monitored during phagocytosis using L-NNA. NOS activity was assayed in RPE cells and ROS to locate the source of NO.
Results:
Phagocytosis of ROS was inhibited by L-NNA but not by D-NNA. L-NNA inhibited the ingestion in a dose-dependent manner, but not the binding of ROS. The inhibition was reversed by L-arginine and also by an NO donor, SIN-1. RPE cells challenged with ROS showed increased cGMP activity, which was significantly reduced by L-NNA and again restored by an overdose of L-arginine. NOS activity was found in RPE cells but not in ROS.
Conclusions:
Our data show that cGMP plays a role in the ingestion phase of ROS phagocytosis by RPE cells via a cGMP second-messenger system.