Endothelial ephrinB2 induces pro-inflammatory activation of monocytes at atherosclerosis-prone sites Jennifer Braun, Sabrina Hoffmann, Anja Feldner, Markus Hecker, Thomas Korff Institute of Physiology and Pathophysiology, University of Heidelberg, Germany The arterial marker molecule ephrinB2 is expressed in endothelial and vascular smooth muscle cells and is crucial for the embryonic development of the vascular system. Although the mechanistic impact of this molecule on angiogenesis has been analyzed in detail, much less is known about its expression and function in adult arteries. To this end, we studied the distribution of ephrinB2 at different sites of murine arteries. Using immunohistochemistry we noted a strong up-regulation of ephrinB2 at atherosclerosis-prone sites of the aortic arch. Subsequent analysis of LDL receptor and ApoE-deficient mice revealed that ephrinB2 to some extend is also expressed by neointimal cells. Furthermore, monocytes up-regulate the receptor EphB2 upon their transdifferentiation into macrophages enabling the interaction with vascular ephrinB2. By applying atomic force microscopy, we demonstrated that monocytes physically interact with ephrinB2. In line with this obervation, adhesion of these cells to endothelial-intact mouse aortic segments abundantly expressing ephrinB2 was inhibited by pre-treatment with soluble ephrinB2. Consequently, the phopsphorylation state of the monocytic EphB2 receptor was analyzed, demonstrating an activation of this receptor upon interaction with ephrinB2. Subsequent functional studies revealed that endothelial ephrinB2 affects the transmigration of monocytes and facilitates their differentiation into a pro-inflammatory phenotype as evidenced by the secretion and up-regulation of the pro-atherosclerotic chemokines MCP-1 (monocytic chemoattractant protein -1) and interleukin-8 on the protein and RNA level, respectively. Taken together, these data establish ephrinB2 as a determinant of the atherosclerotic microenvironment which amplifies the pro-inflammatory differentiation of infiltrating monocytes.