The A-kinase anchoring protein (AKAP) 12 negatively regulates angiogenesis in astrocytes and tumour cells by reducing VEGF release and controlling cell cycle progression. However, the role of AKAP12 in the endothelium, a primary target for VEGF, is unknown. We therefore studied the role of AKAP12 in the angiogenic response of endothelial cells to VEGF.

In primary human endothelial cells transfected with a control siRNA, 72 hours stimulation with VEGF induced significant proliferation, increasing total cell numbers approximately 2-fold compared to non-stimulated cells (P<0.05). In cells treated with siRNA against AKAP12, no significant VEGF-induced proliferation was observed. AKAP12 deficiency also impaired adhesion of cells to fibronectin and migration in response to VEGF while enhancing FAK phosphorylation on Tyr576/577. In a modified spheroid assay, AKAP12 siRNA severely impeded (by 52%, P < 0.001) the sprouting of endothelial cells in response to VEGF. Similarly, in collagen-embedded aortic rings from AKAP12-deficient mice, the formation of PECAM-1-positive tubes was reduced compared to wild-type aortae both under basal conditions and after VEGF stimulation. Moreover in AKAP12^-/- mice, we observed an impaired recovery of blood flow in the ischemic hindlimb. Three weeks after complete excision of the iliac artery, perfusion of the paw was completely restored in wildtype animals, whereas in AKAP12^-/- mice, recovery of perfusion levelled off at approximately 50% of the non-ischemic limb.

Taken together, our data indicate that endothelial AKAP12 expression is essential for VEGF-induced angiogenesis in endothelial cells regulating proliferation, migration and sprouting and focal adhesion turnover. Functionally, loss of AKAP12 results in severely impaired restoration of blood flow in ischemic tissue.