Complement-mediated injury reversibly disrupts glomerular epithelial cell actin microfilaments and focal adhesions

Background Foot process effacement and condensation of the glomerular epith elial cell (GEC) cytoskeleton are manifestations of passive Heymann nephrit is, a model of complement-mediated membranous nephropathy. Methods. To study the effects of complement on the actin cytoskeleton in th is model, we have used an in vitro system in which GECs are sublethally inj ured using a combination of complement-fixing anti-Fx1A IgG and human serum as a source of complement. We examined the effects of this injury on the o rganization of the cytoskeleton and focal contacts using immunohistology an d immunochemistry. Results. By immunofluorescence, sublethal complement mediated injury was ac companied by a loss of actin stress fibers and focal contacts but retention of matrix-associated integrins. Full recovery was seen after 18 hours. Wes tern blot analysis showed no change in the cellular content of the focal co ntact proteins. Inhibition of the calcium-dependent protease calpain did no t prevent injury. In addition, cycloheximide during recovery did not inhibi t the reassembly of stress fibers or focal contacts. Injury was associated with a reduction in tyrosine phosphorylation of paxillin and a currently un identified 200 kDa protein, but inhibition of tyrosine phosphatase activity with sodium vanadate did not prevent injury. Cellular adenosine triphospha te content was significantly reduced in injured cells. Conclusion. These results document reversible, complement dependent disrupt ion of actin microfilaments and focal contacts leading to the dissociation of the cytoskeleton from matrix-attached integrins. This may explain the al tered cell-matrix relationship accompanying podocyte effacement in membrano us nephropathy.