ORIGINAL CONTRIBUTION CD40L contributes to angiotensin II-induced pro-thrombotic state, vascular inflammation, oxidative stress and endothelial dysfunction Michael Hausding • Kerstin Jurk • Steffen Daub • Swenja Kro ¨ller-Scho ¨n • Judith Stein • Melanie Schwenk • Matthias Oelze • Yuliya Mikhed • Jasmin Ghaemi Kerahrodi • Sabine Kossmann • Thomas Jansen • Eberhard Schulz • Philip Wenzel • Angelika B. Reske-Kunz • Christian Becker • Thomas Mu ¨ nzel • Stephan Grabbe • Andreas Daiber Received: 4 April 2013 / Revised: 14 August 2013 / Accepted: 6 September 2013 / Published online: 24 September 2013 Ó Springer-Verlag Berlin Heidelberg 2013 Abstract CD40 ligand (CD40L) is involved in the vas- cular infiltration of immune cells and pathogenesis of atherosclerosis. Additionally, T cell CD40L release causes platelet, dendritic cell and monocyte activation in throm- bosis. However, the role of CD40L in angiotensin II (ATII)-driven vascular dysfunction and hypertension remains incompletely understood. We tested the hypothesis that CD40L contributes to ATII-driven vascular inflam- mation by promoting platelet–leukocyte activation, vascu- lar infiltration of immune cells and by amplifying oxidative stress. C57BL/6 and CD40L -/- mice were infused with ATII (1 mg/kg/day for 7 days) using osmotic minipumps. Vascular function was recorded by isometric tension studies, and reactive oxygen species (ROS) were monitored in blood and heart by optical methods. Western blot, immunohistochemistry, FACS analysis and real-time RT- PCR were used to analyze immune cell distribution, pro- inflammatory cytokines, NAPDH oxidase subunits, T cell transcription factors and other genes of interest. ATII- treated CD40L -/- mice showed improved endothelial function, suppression of blood platelet–monocyte interac- tion (FACS), platelet thrombin generation (calibrated automated thrombography) and coagulation (bleeding time), as well as decreased oxidative stress in the aorta, heart and blood compared to wild-type mice. Moreover, ATII-treated CD40L -/- mice displayed decreased levels of T H 1 cytokines released by splenic CD4 ? T cells (ELISA) and lower expression levels of NOX-2, T-bet and P-selectin as well as diminished immune cell infiltration in aortic tissue compared to controls. Our results demonstrate that many ATII-induced effects on vascular dysfunction, such as vascular inflammation, oxidative stress and a pro- thrombotic state, are mediated at least in part via CD40L. Keywords Immune cell infiltration Á Vascular inflammation Á Phagocytic NADPH oxidase Á Platelet activation Á Endothelial function Á Bleeding time Introduction CD40 ligand (CD40L, CD154) is a member of the tumor necrosis factor (TNF) family preferentially expressed on the surface of activated CD4 ? T cells and platelets and to a variable degree by other immune cells, endothelial cells T. Mu ¨nzel, S. Grabbe and A. Daiber contributed equally and are joint senior authors. Electronic supplementary material The online version of this article (doi:10.1007/s00395-013-0386-5) contains supplementary material, which is available to authorized users. M. Hausding Á S. Daub Á S. Kro ¨ller-Scho ¨n Á M. Oelze Á Y. Mikhed Á J. G. Kerahrodi Á S. Kossmann Á T. Jansen Á E. Schulz Á P. Wenzel Á T. Mu ¨nzel Á A. Daiber (&) II. Medizinische Klinik, Department of Cardiology, Universita ¨tsmedizin der Johannes Gutenberg-Universita ¨t Mainz, Langenbeckstr. 1, 55131 Mainz, Germany e-mail: daiber@uni-mainz.de M. Hausding Á K. Jurk Á S. Kro ¨ller-Scho ¨n Á J. Stein Á M. Schwenk Á S. Kossmann Á P. Wenzel Á C. Becker Center for Thrombosis and Hemostasis (CTH), Medical Center of the Johannes Gutenberg University, Mainz, Germany J. Stein Á M. Schwenk Á A. B. Reske-Kunz Á C. Becker Á S. Grabbe (&) Department of Dermatology, Universita ¨tsmedizin der Johannes Gutenberg-Universita ¨t Mainz, Langenbeckstr. 1, 55131 Mainz, Germany e-mail: stephan.grabbe@unimedizin-mainz.de 123 Basic Res Cardiol (2013) 108:386 DOI 10.1007/s00395-013-0386-5