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Semaphorin 3F Promotes Transendothelial Migration of Leukocytes in the Inflammatory Response After Survived Cardiac Arrest

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Abstract

Leukocyte transmigration through the blood vessel wall is a fundamental step of the inflammatory response and requires expression of adhesion molecule PECAM-1. Accumulating evidence implicates that semaphorin (Sema) 3F and its receptor neuropilin (NRP) 2 are central regulators in vascular biology. Herein, we assess the role of Sema3F in leukocyte migration in vitro and in vivo. To determine the impact of Sema3F on leukocyte recruitment in vivo, we used the thioglycollate-induced peritonitis model. After the induction of peritonitis, C57BL/6 mice were intraperitoneally (i.p.) injected daily with recombinant Sema3F or solvent for 3 days. Compared with solvent-treated controls, leukocyte count was increased in the peritoneal lavage of Sema3F-treated mice indicating that Sema3F promotes leukocyte extravasation into the peritoneal cavity. In line with this observation, stimulation of human endothelial cells with Sema3F enhanced the passage of peripheral blood mononuclear cells (PBMCs) through the endothelial monolayer in the transwell migration assays. Conversely, silencing of endothelial Sema3F by siRNA transfection dampened diapedesis of PBMCs through the endothelium in vitro. xMechanistically, Sema3F induced upregulation of adhesion molecule PECAM-1 in endothelial cells and in murine heart tissue shown by immunofluorescence and western blotting. The inhibition of PECAM-1 by blocking antibody HEC7 blunted Sema3F-induced leukocyte migration in transwell assays. SiRNA-based NRP2 knockdown reduced PECAM-1 expression and migration of PBMCs in Sema3F-treated endothelial cells, indicating that PECAM-1 expression and leukocyte migration in response to Sema3F depend on endothelial NRP2. To assess the regulation of Sema3F in human inflammatory disease, we collected serum samples of patients from day 0 to day 7 after survived out-of-hospital cardiac arrest (OHCA, n = 41). First, we demonstrated enhanced migration of PBMCs through endothelial cells exposed to the serum of patients after OHCA in comparison to the serum of patients with stable coronary artery disease or healthy volunteers. Remarkably, serum samples of OHCA patients contained significantly higher Sema3F protein levels compared with CAD patients (CAD, n = 37) and healthy volunteers (n = 11), suggesting a role of Sema3F in the pathophysiology of the inflammatory response after OHCA. Subgroup analysis revealed that elevated serum Sema3F levels after ROSC are associated with decreased survival, myocardial dysfunction, and prolonged vasopressor therapy, clinical findings that determine the outcome of post-resuscitation period after OHCA. The present study provides novel evidence that endothelial Sema3F controls leukocyte recruitment through a NRP2/PECAM-1-dependent mechanism. Sema3F serum concentrations are elevated following successful resuscitation suggesting that Sema3F might be involved in the inflammatory response after survived OHCA. Targeting the Sema3F/NRP2/PECAM-1 pathway could provide a novel approach to abolish overwhelming inflammation after resuscitation.

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Acknowledgements

We are indebted to Jessica Beckert for her outstanding technical assistance.

Funding

This work was supported by the Deutsche Forschungsgemeinschaft to T. H. (HE 7432/1-1) and to M. M. (973/8-1).

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Authors and Affiliations

  1. Department of Cardiology and Angiology I, Heart Center Freiburg University, Faculty of Medicine, University of Freiburg, Hugstetter Strasse 55, 79106, Freiburg im Breisgau, Germany

    Stephanie Reichert, Stefanie Scheid, Tina Roth, Marius Herkel, Diana Petrova, Alexandra Linden, Miki Weberbauer, Jennifer Esser, Philipp Diehl, Sebastian Grundmann, Christoph Bode, Martin Moser & Thomas Helbing

  2. Department of Emergency Medicine, University Hospital of Freiburg, Faculty of Medicine, University of Freiburg, Freiburg im Breisgau, Germany

    Hans-Jörg Busch & Katrin Fink

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  1. Stephanie Reichert
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Correspondence to Thomas Helbing.

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Reichert, S., Scheid, S., Roth, T. et al. Semaphorin 3F Promotes Transendothelial Migration of Leukocytes in the Inflammatory Response After Survived Cardiac Arrest. Inflammation 42, 1252–1264 (2019). https://doi.org/10.1007/s10753-019-00985-4

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