Complement-induced phospholipase A(2) activation in experimental membranous nephropathy

Background. In the passive Heymann nephritis (PHN) model of membranous neph ropathy. C5b-9 induces glomerular epithelial cell(GEC) injury and proteinur ia. 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 A(2) (PLA(2)). Th is study addresses the mechanisms of PLA(2) activation. Methods. PLA(2) expression was assessed with the polymerase chain reaction or immunoblotting, and activity was determined using an in vitro assay or b y measurement of free AA. Results. Under basal conditions. GEC in culture expressed a relatively low level of cytosolic PLA(2) (cPLA(2)) protein, while mRNAs of groups IB, IIA and V secretory PLA(2)s (sPLA(2)) were not detectable. Incubation of GEC wi th sublytic C5b-9 induced 1.5- to 2.0-fold increases in free [H-3]AA at 40 minutes, and three and 24 hours. C5b-9 did not increase cPLA(2) protein, an d did not induce group IB, IIA or V sPLA(2), mRNAs. Stable overexpression o f cPLA(2) in GEC amplified the C5b-9-induced increases in free [H-3]AA,whil e analogous overexpression of group IIA sPLA(2) had no effect. PLA(2) activ ity was increased in glomeruli of rats with PHN. and this enhanced activity was characterized as cPLA(2). There were no differences in cPLA(2) protein expression between PHN and control glomeruli. Conclusions. Release of AA by C5b-9 in GEC in culture and in vivo is mediat ed by cPLA(2) and the mechanism is consistent with post-translational regul ation of cPLA(2) activity. C5b-9 does not induce expression or stimulate ac tivity of sPLA(2) isoforms in GEC.