Activated coagulation factor X: A novel mitogenic stimulus for human mesangial cells

Intraglomerular activation of the coagulation cascade is a common feature o f mesangioproliferative glomerulonephritis. Besides thrombin, very little i s known about the cellular effects of other components of the coagulation s ystem. This study investigated the effect of activated factor X (FXa) on cu ltured human mesangial cells. This serine protease induced a significant an d dose-dependent increase in DNA synthesis. In addition to its mitogenic ef fect, FXa caused a striking upregulation of platelet-derived growth factor (PDGF) A and B chain gene expression. Next, the intracellular mitogenic sig naling pathways activated by FXa were investigated. FXa induced a rapid spi ke in cytosolic calcium concentration followed by a sustained plateau. This response was not influenced by the downregulation of thrombin receptors. I n addition, FXa stimulated a significant upregulation of different tyrosine -phosphorylated proteins. One of these phosphorylated cellular proteins was represented by the c-jun N-terminal kinase, a member of the mitogen-activa ted protein kinase family. To evaluate the role of FXa enzymatic activity a nd of PDGF autocrine secretion, FXa-induced DNA synthesis was studied in th e presence of leupeptin, a specific serine protease inhibitor, and neutrali zing anti-PDGF antibody. To investigate the role of tyrosine kinase (TK) ac tivation on FXa mitogenic effect, FXa-stimulated thymidine uptake was evalu ated in the presence of genistein and herbimycin A, two powerful and specif ic TK inhibitors. FXa-elicited DNA synthesis was also examined after protei n kinase C (PKC) downregulation by prolonged incubation with phorbol-12-myr istate-13-acetate to study the influence of the phospholipase C-PKC axis. T he proliferative effect of FXa required its proteolytic activity, and the a ctivation of TK was only partially dependent on PKC activation while it was PDGF independent. Finally, it was shown by reverse transcription-PCR that mesangial cells do not express the signaling splicing variant of the putati ve FXa receptor, effector protease receptor-1. In conclusion, the present s tudy demonstrated that FXa is a powerful mitogenic factor for human mesangi al cells, and it induces its cellular effect not through effector protease receptor-1, but most likely by binding a protease-activated receptor and ac tivating phospholipase C-PKC and TK signaling pathways.