Complement-induced phospholipase A2 activation in experimental membranous nephropathy.
In the passive Heymann nephritis (PHN) model of membranous nephropathy, C5b-9 induces glomerular epithelial cell (GEC) injury and proteinuria, which is partially mediated by eicosanoids. By analogy, in cultured rat GEC, sublytic C5b-9 injures plasma membranes and releases arachidonic acid (AA) and eicosanoids, due to activation of phospholipase A2 (PLA2). This study addresses the mechanisms of PLA2 activation. PLA2 expression was assessed with the polymerase chain reaction or immunoblotting, and activity was determined using an in vitro assay or by measurement of free AA. Under basal conditions, GEC in culture expressed a relatively low level of cytosolic PLA2 (cPLA2) protein, while mRNAs of groups IB, IIA and V secretory PLA2s (sPLA2) were not detectable. Incubation of GEC with sublytic C5b-9 induced 1.5- to 2.0-fold increases in free [3H]AA at 40 minutes, and three and 24 hours. C5b-9 did not increase cPLA2 protein, and did not induce group IB, IIA or V sPLA2 mRNAs. Stable overexpression of cPLA2 in GEC amplified the C5b-9-induced increases in free [3H]AA, while analogous overexpression of group IIA sPLA2 had no effect. PLA2 activity was increased in glomeruli of rats with PHN, and this enhanced activity was characterized as cPLA2. There were no differences in cPLA2 protein expression between PHN and control glomeruli. Release of AA by C5b-9 in GEC in culture and in vivo is mediated by cPLA2, and the mechanism is consistent with post-translational regulation of cPLA2 activity. C5b-9 does not induce expression or stimulate activity of sPLA2 isoforms in GEC.