von Willebrand factor directly interacts with DNA from neutrophil extracellular traps.
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.


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.
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.).
Sandra
S
Grässle
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.).
Volker
V
Huck
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.).
Karin I
KI
Pappelbaum
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.).
Christian
C
Gorzelanny
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.).
Camilo
C
Aponte-Santamaría
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.).
Carsten
C
Baldauf
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.).
Frauke
F
Gräter
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.).
Reinhard
R
Schneppenheim
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.).
Tobias
T
Obser
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.
Stefan W
SW
Schneider
D051722:ADAM Proteins / Q000378:metabolism; D000071120:ADAMTS13 Protein; D001665:Binding Sites; D002448:Cell Adhesion / Q000187:drug effects; D002478:Cells, Cultured; D004247:DNA / Q000378:metabolism*; D006495:Heparin, Low-Molecular-Weight / Q000494:pharmacology; D061307:Human Umbilical Vein Endothelial Cells / Q000378:metabolism; D006801:Humans; D008958:Models, Molecular; D009504:Neutrophils / Q000187:drug effects / Q000378:metabolism*; D010973:Platelet Adhesiveness; D019038:Platelet Glycoprotein GPIb-IX Complex / Q000378:metabolism; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D059748:Proteolysis; D012039:Regional Blood Flow; D013314:Stress, Mechanical; D013997:Time Factors; D014841:von Willebrand Factor / Q000737:chemistry / Q000378:metabolism*
D006495:Heparin, Low-Molecular-Weight; D019038:Platelet Glycoprotein GPIb-IX Complex; C094990:adhesion receptor; D014841:von Willebrand Factor; D004247:DNA; D051722:ADAM Proteins; D000071120:ADAMTS13 Protein; C099604:ADAMTS13 protein, human
endothelium; heparin; inflammation; leukocytes; von Willebrand factor