Extracellular sphingomyelinase activity impairs TNF-α-induced endothelial cell death via ADAM17 activation and TNF receptor 1 shedding

Abstract

// Anselm Sommer 1 , Marie Düppe 1 , Lena Baumecker 1 , Felix Kordowski 1 , Joscha Büch 1 , Johaiber Fuchslocher Chico 2 , Jürgen Fritsch 2 , Stefan Schütze 2 , Dieter Adam 2 , Maria Sperrhacke 1 , Sucharit Bhakdi 1 and Karina Reiss 1 1 Department of Dermatology, University of Kiel, 24105 Kiel, Germany 2 Institute of Immunology, University of Kiel, 24105 Kiel, Germany Correspondence to: Karina Reiss, email: [e-mail] Keywords: sphingomyelinase, ADAM17, apoptosis, TNF-α, TNFR1 Received: February 28, 2017     Accepted: July 11, 2017     Published: August 07, 2017 ABSTRACT ADAM17, a prominent member of the “Disintegrin and Metalloproteinase” (ADAM) family, is an important regulator of endothelial cell proliferation and cell survival. The protease controls vital cellular functions through cleavage of growth factors, cytokines and their receptors including transforming growth factor-alpha (TGF-α), tumor necrosis factor-alpha (TNF-α) and TNF-α receptor 1 (TNFR1). TNF-α is the major inducer of endothelial cell death in cardiovascular diseases. The latter are also characterized by elevated plasma and tissue levels of extracellular sphingomyelinase (SMase). Whether the SMase affects ADAM activity and thus endothelial cell function has not been addressed to date. Here, we analyzed the effect of SMase on ADAM17-mediated shedding in COS7 cells and in human umbilical vein endothelial cells (HUVECs). Exposure to SMase significantly increased ADAM17-mediated release of alkaline-phosphatase (AP)-tagged TGF-α in COS7 cells and shedding of endogenously expressed TNFR1 in HUVECs. We previously presented evidence that surface exposure of phosphatidylserine (PS) is pivotal for ADAM17 to exert sheddase function. We found that SMase treatment led to PS externalization in both cell types. Transient non-apoptotic PS exposure is often mediated by Ca 2+ -dependent phospholipid scramblases. Accordingly, the Ca 2+ -chelator EGTA markedly reduced the breakdown of phospholipid asymmetry and shedding of TGF-α and TNFR1. Moreover, sheddase activity was significantly diminished in the presence of the competing PS-headgroup OPLS. SMase-stimulated TNFR1 shedding strikingly diminished TNF-α-induced signalling cascades and endothelial cell death. Taken together, our data suggest that SMase activity might act as protective factor for endothelial cells in cardiovascular diseases.