Tumor necrosis factor α increases antifibrinolytic activity of cultured human mesangial cells

Tumor necrosis factor α increases antifibrinolytic activity of cultured human mesangial cells. Tumor necrosis factorα (TNFα) is likely to exert a major influence in the pathogenesis of glomerulopathies. Besides its proinflammatory properties, TNFα interacts with cell growth and synthesis of components of the fibrinolytic system. In this study, we report the effects of recombinant human TNFα on the synthesis of tissue-type plasminogen activator (t-PA) and its inhibitor (PAI-1) by human mesangial cells in culture. We first demonstrate that TNFα binds specifically to a single class of high affinity receptors (Kd 5.10-11M; 1500 receptors/cell). TNFα has an antimitogenic effect on human mesangial cells since it decreased DNA synthesis, measured by 3H-thymidine incorporation, in a dose-dependent manner. Release of cytosolic LDH and incorporated 51Cr was not increased by 100 ng/ml TNFα as compared with control, indicating that this monokine is not cytotoxic for cultured human mesangial cells. Zymographic analysis and reverse fibrin autography disclosed a 120 kD t-PA-PAI-1 complex and a 50 kD free form of PAI-1 in the supernatants of both unstimulated and TNF-stimulated cells; PAI-1 was released in excess and free t-PA was not observed. TNFα (0 to 100 ng/ml) had no effect on t-PA synthesis, but enhanced PAI-1 release in a time- and dose-dependent manner (97% increase of PAI-1 synthesis after a 24 hour incubation). This effect was abolished by cycloheximide, suggesting that protein synthesis was required. Northern blot analysis showed that TNFα increased the steady-state PAI-1 mRNA levels in a time-dependent manner, with a maximal effect at two hours. Finally, in contrast to what was reported for rat mesangial cells, we failed to demonstrate a production of TNFα in human mesangial cells themselves by immunoradiometric assay, immunocytochemistry and Northern blot analysis using a specific TNFα cDNA probe. We conclude that TNFα has an antimitogenic activity on human mesangial cells in culture and enhances the synthesis of PAI-1. Whether TNFα plays a role in glomerular PAI-1 deposition and persistency of fibrin in vivo remains to be determined.