RELEASE OF ARACHIDONIC-ACID BY COMPLEMENT C5B-9 COMPLEX IN GLOMERULAR EPITHELIAL-CELLS

In experimental membranous nephropathy, C5b-9 induces noncytolytic glomerular epithelial cell (GEC) injury and proteinuria, which in some models is partially mediated by metabolites of arachidonic acid. In cultured GEC, sublytic C5b-9 increases cytosolic Ca2+ concentration ([Ca2+]i), activates phospholipase C (PLC), and releases arachidonic acid and eicosanoids. This study examined mechanisms of arachidonic acid production by C5b-9. In GEC labeled with [H-3]arachidonate C5b-9 increased free [H-3]arachidonic acid and 1,2-[H-3]arachidonoyl-diacylglycerol (DAG), an endogenous activator of protein kinase C (PKC). Elevated [Ca2+]i was not sufficient to account for increased free arachidonic acid. Moreover, in GEC that had been depleted of PKC by preincubation for 18 h with 2-mu-M phorbol myristate acetate, the C5b-9-induced arachidonate release was inhibited by > 75%. Reacylation of phospholipids was not decreased by C5b-9. Homogenates of GEC that had been stimulated with C5b-9 released more [C-14]arachidonate from exogenously added 2-[C-14]arachidonoyl-phosphatidyl-ethanolamine or 2-[C-14]arachidonoyl-phosphatidylcholine than homogenates of unstimulated cells (assayed at a Ca2+ concentration of 2 mM). These experiments demonstrate directly that C5b-9 increased phospholipase A2 (PLA2) activity. PLA2 appeared to be stimulated as a result of PKC activation (probably secondary to increased DAG) in association with elevated [Ca2+]i. The C5b-9-induced activation of PLA2 may lead to release of eicosanoids, which may contribute toward impaired glomerular capillary wall permselectivity in experimental membranous nephropathy.