The apelin gene can promote vascular endothelial cell (VEC) proliferation, migration, and angiogenesis. However, the molecular mechanism for regulation of the apelin gene is still unknown. Real-time PCR and Western blotting analysis were employed to detect the effect of all-trans retinoic acid (ATRA) in up-regulating apelin expression in human umbilical vein endothelial cells (HUVECs). Furthermore, the in vivo study also indicated that ATRA could increase apelin expression in balloon-injured arteries of rats, which is consistent with the results from the cultured HUVECs. To ensure whether retinoic acid receptor (RAR) alpha (RAR alpha) could be induced by ATRA in regulating apelin, the expression of RAR alpha was tested with a siRNA method to knock down RAR alpha or adenovirus vector infection to overexpress RAR alpha. The results showed that ATRA could up-regulate apelin expression time- and dose-dependently in HUVECs. ATRA could induce a RAR alpha increase; however, the expression of RAR beta and RAR gamma were unchanged. The blocking of RAR alpha signaling reduced the response of apelin to ATRA when HUVECs were treated with RAR alpha antagonists (Ro 41-5253) or the use of siRNA against RAR alpha (si-RAR alpha) knockdown RAR alpha expression before using ATRA. In addition, induction of RAR alpha overexpression by infection with pAd-GFP-RAR alpha further increased the induction of apelin by ATRA. These results suggested that ATRA up-regulated apelin expression by promoting RAR alpha signaling.