Supplementary Material for: Hydroxysafflor Yellow A Promotes Angiogenesis via the Angiopoietin 1/ Tie-2 Signaling Pathway

<i>Background:</i> The flowers of <i>Carthamus tinctorius</i> L. are widely used in traditional Chinese medicine to treat cerebrovascular and cardiovascular diseases. Hydroxysafflor yellow A (HSYA), the main constituent of <i>C. tinctorius</i> L. flowers, is known for its multiple biological activities. The present study investigated the effects of HSYA on angiogenesis in vitro and in a mouse hindlimb ischemia model. <i>Methods:</i>Using human umbilical vein endothelial cells (HUVEC) in vitro and a mouse hindlimb ischemia model in vivo, the angiogenic role of HSYA was evaluated. <i>Results:</i> HSYA significantly increased the capillary-like tube formation and migration of HUVEC. HSYA not only induced a rise in the expression of angiopoietin 1 and Tie-2 but it also increased phosphorylation of Tie-2, Akt, and extracellular signal-regulated kinase 1/2. Furthermore, an anti-Tie-2 neutralizing antibody significantly inhibited HSYA-induced HUVEC tube formation and migration. In vivo, the recovery of perfusion of ischemic hindlimb tissue after femoral artery interruption was significantly increased in HSYA-treated mice compared to vehicle controls. Consistent with these results, the arteriole and capillary densities in ischemic gastrocnemius muscles were significantly increased in HSYA-treated mice. <i>Conclusions:</i> These results indicate the potential utility of HSYA for the treatment of ischemic diseases.