Sandra Grässle1, Volker Huck1, Karin I Pappelbaum1, Christian Gorzelanny1, Camilo Aponte-Santamaría1, Carsten Baldauf1, Frauke Gräter1, Reinhard Schneppenheim1, Tobias Obser1, Stefan W Schneider2. 1. From the Experimental Dermatology Division, Department of Dermatology, Medical Faculty Mannheim, Heidelberg University, Mannheim, Germany (S.G., V.H., K.I.P., C.G., S.W.S.); Molecular Biomechanics Group, Heidelberg Institute for Theoretical Studies, Heidelberg, Germany (C.A.-S., F.G.); Theory Department, Fritz Haber Institute of the Max Planck Society, Berlin, Germany (C.B.); and Department of Pediatric Hematology and Oncology, University Medical Center, Hamburg-Eppendorf, Hamburg, Germany (R.S., T.O.). 2. From the Experimental Dermatology Division, Department of Dermatology, Medical Faculty Mannheim, Heidelberg University, Mannheim, Germany (S.G., V.H., K.I.P., C.G., S.W.S.); Molecular Biomechanics Group, Heidelberg Institute for Theoretical Studies, Heidelberg, Germany (C.A.-S., F.G.); Theory Department, Fritz Haber Institute of the Max Planck Society, Berlin, Germany (C.B.); and Department of Pediatric Hematology and Oncology, University Medical Center, Hamburg-Eppendorf, Hamburg, Germany (R.S., T.O.). Stefan.Schneider@medma.uni-heidelberg.de.
Abstract
OBJECTIVE: Inflammatory conditions provoke essential processes in the human vascular system. It leads to the formation of ultralarge von Willebrand factor (VWF) fibers, which are immobilized on the endothelial cell surface and transform to highly adhesive strings under shear conditions. Furthermore, leukocytes release a meshwork of DNA (neutrophil extracellular traps) during the process of the recently discovered cell death program NETosis. In the present study, we characterized the interaction between VWF and DNA and possible binding sites to underline the role of VWF in thrombosis and inflammation besides its function in platelet adhesion. APPROACH AND RESULTS: Both functionalized surfaces and intact cell layers of human umbilical vein endothelial cells were perfused with isolated, protein-free DNA or leukocytes from whole blood at distinct shear rates. DNA-VWF interaction was monitored using fluorescence microscopy, ELISA-based assays, molecular dynamics simulations, and electrostatic potential calculations. Isolated DNA, as well as DNA released by stimulated leukocytes, was able to bind to shear-activated, but not inactivated, VWF. However, DNA-VWF binding does not alter VWF degradation by a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13. Moreover, DNA-VWF interaction can be blocked using unfractionated and low-molecular-weight heparin, and DNA-VWF complexes attenuate platelet binding to VWF. These findings were supported using molecular dynamics simulations and electrostatic calculations of the A1- and A2-domains. CONCLUSIONS: Our findings suggest that VWF directly binds and immobilizes extracellular DNA released from leukocytes. Therefore, we hypothesize that VWF might act as a linker for leukocyte adhesion to endothelial cells, supporting leukocyte extravasation and inflammation.
OBJECTIVE: Inflammatory conditions provoke essential processes in the human vascular system. It leads to the formation of ultralarge von Willebrand factor (VWF) fibers, which are immobilized on the endothelial cell surface and transform to highly adhesive strings under shear conditions. Furthermore, leukocytes release a meshwork of DNA (neutrophil extracellular traps) during the process of the recently discovered cell death program NETosis. In the present study, we characterized the interaction between VWF and DNA and possible binding sites to underline the role of VWF in thrombosis and inflammation besides its function in platelet adhesion. APPROACH AND RESULTS: Both functionalized surfaces and intact cell layers of human umbilical vein endothelial cells were perfused with isolated, protein-free DNA or leukocytes from whole blood at distinct shear rates. DNA-VWF interaction was monitored using fluorescence microscopy, ELISA-based assays, molecular dynamics simulations, and electrostatic potential calculations. Isolated DNA, as well as DNA released by stimulated leukocytes, was able to bind to shear-activated, but not inactivated, VWF. However, DNA-VWF binding does not alter VWF degradation by a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13. Moreover, DNA-VWF interaction can be blocked using unfractionated and low-molecular-weight heparin, and DNA-VWF complexes attenuate platelet binding to VWF. These findings were supported using molecular dynamics simulations and electrostatic calculations of the A1- and A2-domains. CONCLUSIONS: Our findings suggest that VWF directly binds and immobilizes extracellular DNA released from leukocytes. Therefore, we hypothesize that VWF might act as a linker for leukocyte adhesion to endothelial cells, supporting leukocyte extravasation and inflammation.
Authors: Thejaswi Kalagara; Tracy Moutsis; Yi Yang; Karin I Pappelbaum; Anne Farken; Lucia Cladder-Micus; Sabine Vidal-Y-Sy; Axel John; Alexander T Bauer; Bruno M Moerschbacher; Stefan W Schneider; Christian Gorzelanny Journal: Blood Adv Date: 2018-09-25
Authors: Erin F Carlton; Walker M McHugh; Kelli McDonough; Julie Sturza; Karl Desch; Timothy T Cornell Journal: Am J Respir Crit Care Med Date: 2020-02-01 Impact factor: 21.405
Authors: Nicoletta Sorvillo; Daniella M Mizurini; Carmen Coxon; Kimberly Martinod; Ronak Tilvawala; Deya Cherpokova; Ari J Salinger; Robert J Seward; Caleb Staudinger; Eranthie Weerapana; Nathan I Shapiro; Catherine E Costello; Paul R Thompson; Denisa D Wagner Journal: Circ Res Date: 2019-06-28 Impact factor: 17.367