Role of Rho-GTPases in complement-mediated glomerular epithelial cell injury

Abstract

Visceral glomerular epithelial cells (GEC) are essential for maintenance of normal glomerular permselectivity. The actin cytoskeleton is a key determinant of GEC morphology and function. In the rat passive Heymann nephritis (PHN) model of membranous nephropathy, complement C5b-9 induces nonlytic GEC injury associated with morphological changes of GEC and proteinuria. The current study addresses the role of Rho family of small GTPases in complement-mediated GEC injury. When cultured rat GEC were stimulated with complement C5b-9 for 18 h, RhoA activity increased, whereas Rac1/Cdc42 activities decreased, compared with control cells. Similar changes in Rho-GTPase activities were observed in glomeruli from rats with PHN. The amount of active p190RhoGAP, a negative upstream regulator of RhoA, was decreased in complement-stimulated GEC, potentially contributing to increased RhoA activity. To address the functional effects of Rho-GTPases, GEC were transfected with constitutively active (CA) or dominant negative (DN) Rho-GTPase mutants. GEC transfected with CA-RhoA showed a smaller and round contour and prominent cortical F-actin. In contrast, GEC transfected with CA-Rac1 demonstrated morphological changes that resembled process formation. In addition, expression of CA-RhoA attenuated complement-mediated cytotoxicity, whereas cytotoxicity was augmented by DN-RhoA. Thus exposure of GEC to complement alters the balance of RhoA, Rac1, and Cdc42 activities. The activity of Rac1 may contribute to process formation, while activation of RhoA (e.g., in the setting of complement attack), with or without blunting of Rac1 activity, may have an opposite effect, i.e., contribute to foot process effacement. Activation of RhoA increases the resistance of GEC to complement-mediated injury.