The absence of Nogo-B reduces the transmigration of neutrophils and monocytes across endothelial cell monolayers. (A) Western blot analysis for Nogo-B and the adhesion molecules VE-cadherin, PECAM-1, and ICAM-1, on HDMECs transfected for 48 hours with siRNA Nogo-B and control, and stimulated with TNF-α (10 ng/mL) for 24 hours. (B) FACS analysis for E-selectin, ICAM-1, VCAM, and PECAM-1 on HDMECs transfected for 48 hours with siRNA Nogo-B and control. Transfected cells were stimulated with TNF-α (10 ng/mL) for 4 or 24 hours, and surface expression of E-selectin or ICAM-1, VCAM, and PECAM-1 expression was analyzed by FACS. TNF-α–stimulated siRNA Nogo-B or control treated HDMEC monolayers were used in flow-chamber experiments, as described in “Methods.” The number of rolling neutrophils per square millimeter (C), the average rolling velocity of neutrophils (D), and firmly adherent neutrophils (E) were not different. However, knock-down of Nogo-B in HDMECs reduced the transmigration of (F) neutrophils and (G) monocytes (THP-1) compared with control, siRNA-treated HDMECs.