Gas6 regulates EC activation in vitro and in vivo. (A,B) Stimulation of ECs with 100 ng/mL TNF-α revealed a role for Gas6 in the activation of endothelial cells in vitro. TNF-α stimulated the production of VCAM-1 (A) and ICAM-1 (B) by WT ECs but not by Gas6^−/− ECs. Data represent means plus or minus SEM (*P < .05 vs stimulated WT cells, N = 9). (C-J) In normal heart tissue (baseline), no (VCAM-1) or limited (ICAM-1) expression of the adhesion molecules VCAM-1 (C,D) and ICAM-1 (G,H) can be detected in the blood vessels of either WT or Gas6^−/− mice. At 24 hours after intraperitoneal injection of a sublethal dose of TNF-α, up-regulated expression of VCAM-1 (E) and ICAM-1 (I) can be seen in the endothelial cells in WT hearts but is absent (VCAM-1) or reduced (ICAM-1) in Gas6^−/− hearts (F-J). Bar represents 50 μm in all panels. (K) Knockdown of Gas6 or Axl in HUVECs by transfection of cells with 25 nM siRNA for, respectively, Gas6 or Axl, reduced the expression of hICAM-1 in response to TNF-α (10 ng/mL). In contrast, knockdown of Tyro3, using 100 nM of Tyro3 siRNA, failed to reduce the expression of hICAM-1 on HUVECs in response to TNF-α (10 ng/mL). Data represent the percentage of the control response (*P < .05 vs control siRNA, N = 3).