A1 adenosine receptor activation promotes angiogenesis and release of VEGF from monocytes

Adenosine is a proangiogenic purine nucleoside released from ischemic and hypoxic tissues. Of the 4 adenosine receptor (AR) subtypes (A(1), A(2A), A(2B), and A(3)), the A(2) and A(3) have been previously linked to the modulation of angiogenesis. We used the chicken chorioallantoic membrane (CAM) model to determine whether A1 AR activation affects angiogenesis. We cloned and pharmacologically characterized chicken AR subtypes to evaluate the selectivity of various agonists and antagonists. Application of the A(1) AR-selective agonist N-6-cyclopentyladenosine ( CPA; 100 nmol/L) to the CAM resulted in a 40% increase in blood vessel number (P < 0.01), which was blocked by the A(1) AR-selective antagonist C-8-( N- methylisopropyl)-amino-N-6-(5'-endohydroxy)-endonorbornan-2-yl-9-methyladenine (WRC-0571; 1 mu mol/ L). Selective A(2A) AR agonists did not stimulate angiogenesis in the CAM. In an ex vivo rat aortic ring model of angiogenesis that includes cocultured endothelial cells, fibroblasts, and smooth muscle cells, 50 nmol/L CPA did not directly stimulate capillary formation; however, medium from human mononuclear cells pretreated with CPA, but not vehicle, increased capillary formation by 48% (P < 0.05). This effect was blocked by WRC-0571 (1.5 mu mol/L) or anti-VEGF antibody (1 mu g/mL). CPA (5 nmol/L) stimulated a 1.7-fold increase in VEGF release from the mononuclear cells. This is the first study to show that A(1) AR activation induces angiogenesis. Stimulation of A(2) ARs on endothelial cells results in proliferation and tube formation, and A(2) and A(3) ARs on inflammatory cells modulate release of angiogenic factors. We conclude that adenosine promotes a coordinated angiogenic response through its interactions with multiple receptors on multiple cell types.