Metabolism in Frog Retinal Pigment Epithelium of Docosahexaenoic and Arachidonic Acids Derived from Rod Outer Segment Membranes

Abstract

Frog photoreceptor cells shed about 10% of their rod outer segments (ROS) every 4th day. Packets of these membranes are phagocytized and digested by retinal pigment epithelial (RPE) cells. Large amounts of lipids must be processed daily, especially docosahexaenoic acid (22:6n-3, DHA), the major fatty acid of these membranes. To study the metabolism of ROS lipids in RPE cells, RPE-eyecups were incubated with [3H]DHA-, [3H]arachidonic acid (AA)-, or [2-3H]glycerol-labeled ROS membranes for 2 hr, followed by a chase for up to 8 hr. Lipid extracts of RPE cells and incubation media were resolved into classes and quantitated for radioactivity. In RPE cells, the relative proportion of DHA and AA label in triglycerides (TG) increased dramatically with incubation time, although the substrate ROS membranes did not contain labeled TG. Other RPE lipids showed prominent reductions or relative little change. The percentage of radioactivity in free fatty acids (FFA) was low (< 3%) in RPE cells. In the chase media, the majority (60-80%) of DHA and AA label was found in FFA, with little radioactivity in TG or phospholipids. When RPE cells were incubated with [3H]glycero-labeled ROS membranes which contained 22% of the label in TG, the most rapid reduction in relative radioactivity appeared in TG. We conclude that DHA and AA are released from phagocytized ROS membranes and are rapidly incorporated into RPE cellular lipids, primarily TG. This lipid class is very active metabolically, since TG derived from ROS are rapidly hydrolysed. Free DHA and AA of ROS origin are released from RPE cells. We suggest that RPE triglycerides are the source of FFA in the interphotoreceptor matrix.