Nitric Oxide as a Second Messenger in Phagocytosis by Cultured Retinal Pigment Epithelial Cells

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.