title: Regulation of endothelial migration and proliferation by ephrin-A1.
abstract: Endothelial migration and proliferation are fundamental processes in angiogenesis and wound healing of injured or inflamed vessels. The present study aimed to investigate the regulation of the Eph/ephrin-system during endothelial proliferation and the impact of the ligand ephrin-A1 on proliferation and migration of human umbilical venous (HUVEC) and arterial endothelial cells (HUAEC). Endothelial cells that underwent contact inhibition showed a massive induction of ephrin-A1. In contrast, an injury to a confluent endothelial layer, associated with induction of migration and proliferation, showed reduced ephrin-A1 levels. In addition, reducing ephrin-A1 expression by siRNA led to increased proliferation, whereas the overexpression of ephrin-A1 led to decreased proliferative activity. Due to the fact that wound healing is a combination of proliferation and migration, migration was investigated in detail. First, classical wound-healing assays showed increased wound closure in both ephrin-A1 silenced and overexpressing cells. Live-cell imaging enlightened the underlying differences. Silencing of ephrin-A1 led to a faster but more disorientated migration. In contrast, ephrin-A1 overexpression did not influence velocity of the cells, but the migration was more directed in comparison to the controls. Additional analysis of EphA2-silenced cells showed similar results in terms of proliferation and migration compared to ephrin-A1 silenced cells. Detailed analysis of EphA2 phosphorylation on ligand-dependent phospho-site (Y588) and autonomous activation site (S897) revealed a distinct phosphorylation pattern. Furthermore, the endothelial cells ceased to migrate when they came in contact with an ephrin-A1 coated surface. Using a baculoviral-mediated expression system, ephrin-A1 silencing and overexpression was shown to modulate the formation of focal adhesions. This implicates that ephrin-A1 is involved in changes of the actin cytoskeleton which explains the alterations in migratory actions, at least in part. In conclusion, ephrin-A1 expression is regulated by cellular density and is itself a critical determinant of endothelial proliferation. According to current knowledge, ephrin-A1 seems to be remarkably involved in elementary processes of endothelial migration like cellular polarization, migratory direction and speed. These data support the notion that ephrin-A1 plays a pivotal role in basal mechanisms of re-endothelialization.
authors.affiliation: Department of Internal Medicine and Cardiology, TU Dresden, Germany.
authors.forename: Elisa
authors.initials: E
authors.lastname: Wiedemann
authors.affiliation: Department of Internal Medicine and Cardiology, TU Dresden, Germany.
authors.forename: Stefanie
authors.initials: S
authors.lastname: Jellinghaus
authors.affiliation: Department of Internal Medicine and Cardiology, TU Dresden, Germany.
authors.forename: Georg
authors.initials: G
authors.lastname: Ende
authors.affiliation: Department of Internal Medicine and Cardiology, TU Dresden, Germany.
authors.forename: Antje
authors.initials: A
authors.lastname: Augstein
authors.affiliation: Lightmicroscopy Facility, BioTec Dresden, Germany.
authors.forename: Ronny
authors.initials: R
authors.lastname: Sczech
authors.affiliation: Department of Clinical Pathobiochemistry, TU Dresden, Germany.
authors.forename: Ben
authors.initials: B
authors.lastname: Wielockx
authors.affiliation: Department of Internal Medicine, Division of Cardiology and Angiology, Magdeburg University, Germany.
authors.forename: Sönke
authors.initials: S
authors.lastname: Weinert
authors.affiliation: Department of Internal Medicine and Cardiology, TU Dresden, Germany.
authors.forename: Ruth H
authors.initials: RH
authors.lastname: Strasser
authors.affiliation: Department of Internal Medicine and Cardiology, TU Dresden, Germany. Electronic address: david.poitz@tu-dresden.de.
authors.forename: David M
authors.initials: DM
authors.lastname: Poitz
mesh_terms: D002452:Cell Count; D059447:Cell Cycle Checkpoints; D002465:Cell Movement; D049109:Cell Proliferation; D036382:Ephrin-A1 / Q000378:metabolism*; D022001:Focal Adhesions / Q000378:metabolism; D061307:Human Umbilical Vein Endothelial Cells / Q000166:cytology* / Q000378:metabolism*; D006801:Humans; D060888:Real-Time Polymerase Chain Reaction; D036104:Receptor, EphA2 / Q000378:metabolism
chemical_list: D036382:Ephrin-A1; D036104:Receptor, EphA2
keywords: Endothelial cells; In-stent-restenosis; Migration; Proliferation