Adenosine A2A receptor stimulation increases angiogenesis by down-regulating production of the antiangiogenic matrix protein thrombospondin 1

Topical adenosine A(2A) receptor agonists promote wound healing by, among other effects, increasing microvessel formation. Results of representational display analysis of human umbilical vein endothelial cells suggested that A(2A) receptor occupancy modulates expression of the antiangiogenic matrix protein thrombospondin 1 (TSP1). We therefore determined whether A(2A) receptor occupation stimulates angiogenesis by modulating TSP1 secretion. Human microvascular endothelial cells (HMVEC) were treated with medium alone, 2-p-[2- carboxyethyl] phenethyl-amino-5'-N-ethylcarboxamido-adenosine (CGS21680), or 2-[2-(4-chlorophenyl)ethoxy]adenosine (MRE0094), selective A(2A) receptor agonists. TSP1 protein secretion was down- regulated after treatment with the A 2A agonists CGS21680 or MRE0094 in a dose-dependent manner (EC50 = 6.65 nM and 0.23 mu M, respectively). The selective A(2A) receptor antagonist 4-{2-[7-amino-2-(2-furyl)[1,2,4] triazolo-[2,3a][ 1,3,5] triazin-5-ylamino]ethyl}phenol (ZM241385) but not the A1 and A(2B) receptor antagonists diphenylcyclopentylxanthine, enprofylline, and N-(4-acetylphenyl)-2-[4-(2,3,6,7-tetrahydro2,6- dioxo-1,3-dipropyl-1H-purin-8-yl)phenoxy]acetamide (MRS1706) completely abrogated the A(2A) receptor agonistmediated effect on TSP1. Vascular tube formation by HMVEC was increased by adenosine A 2A receptor agonists in a dosedependent fashion (EC50 = 6.65 mu M for both), and this effect was reversed by the A(2A) antagonist. Moreover, in the presence of antibodies to TSP1 and CD36, the receptor for TSP1, the adenosine A(2A) receptor agonists stimulated no increase in vascular tube formation. These results indicate that the angiogenic effects of adenosine A(2A) receptor activation are, at least in part, caused by the suppression of TSP1 secretion.