Tumor necrosis factor-α-induced release of plasminogen activator inhibitor-1 from human umbilical vein endothelial cells: involvement of intracellular ceramide signaling event

Abstract

We have investigated the biochemical mechanism of tumor necrosis factor (TNF)-α-induced release of plasminogen activator inhibitor-1 (PAI-1) from human umbilical vein endothelial cells (HUVEC). Treatment of HUVEC with TNF-α for 3 h resulted in a 2.8-fold increase in the PAI-1 release compared with control. The increase in PAI-1 release was accompanied by a 133% increase in the intracellular acidic sphingomyelinase (SMase) activity. High-performance liquid chromatographic (HPLC) analysis revealed that the intracellular ceramide levels increased to 126% of the control ( P<0.05), but the contents of membranous ceramide remained unaltered. We have previously shown that a cell-permeable ceramide analog, N-acetylsphingosine (C 2-ceramide) enhances the PAI-1 release from HUVEC. Here, N-acetylsphinganine (C 2-dihydroceramide) was found to specifically suppress both C 2-ceramide- and TNF-α-induced increase in PAI-1 release from HUVEC without affecting the control PAI-1 release. Treatment of HUVEC with staphylococcal SMase that may mimic the activation of the membranous neutral SMase also increased the PAI-1 release. The increase in PAI-1 release by this mechanism was suppressed by a cyclooxygenase inhibitor, aspirin, whereas the inhibitor did not affect TNF-α-induced increase in PAI-1 release. Taken together, these findings suggest that TNF-α prominently utilizes the lysosomal acidic SMase-ceramide signaling pathway in the induction of PAI-1 release from HUVEC.