EphB4-Fc induces reverse signaling in osteoclast precursors but not in MSCs. (A) Osteoclast precursors (pOC) and MSCs from donors and MM patients were left untreated or stimulated with clustered EphB4-Fc (4 μg/mL) for 20 minutes and then subjected to Western blot. Note that ephrinB2 was expressed in pOC and MSCs (∼ 50-kDa band size) and that EphB4-Fc induced phosphorylation of ephrinB2 in pOC but not in donor or MM patient MSCs. (B) Human pOCs were treated with Fc or EphB4-Fc for 2 days and then subjected to quantitative RT-PCR for NFATc1 (ephrinB2 target). Note down-regulation of NFATc1 by EphB4-Fc. (C-D) Human pOCs were treated with Fc or EphB4-Fc for 7 to 10 days and then stained for TRAP. Note reduced number of TRAP-expressing multinucleated osteoclasts (red arrows) in EphB4-Fc–treated cultures.