The role of avB3 and VEGFR2 endocytic recycling in angiogenesis

Placental growth factor (PIGF) binds to VEGFR1 and is known to play a role in pathological angiogenesis, but its mechanism of action remains unclear. Endothelial cell migration in response to angiogenic stimuli requires coordination of adhesive function with VEGFR signalling, and I have studied the intracellular trafficking of integrins and VEGFRs in primary cultured human umbilical vein endothelial cells (HUVECs). VEGFR2 and alphavbeta3 integrin cycled rapidly between the plasma membrane and EEA1/Rab4-positive early endosomes, whereas VEGFR1 remained at the plasma membrane and was not subject to rounds of endo- exocytosis. Treatment of HUVECs with PIGF or VEGF-A promoted the rapid and coordinate mobilisation of both VEGFR2 and alphavbeta3 from these endosomes to the plasma membrane, via a mechanism that was developed on inactivation of GSK-3beta but did not require the activity of PKD1, nor the tyrosine kinase activity of VEGFRs. Furthermore, RNAi of Rab4a and pharmacological inhibition of alphav integrin signalling opposed PIGF-promoted endothelial cell vessel branching and cross-bridge formation in an organotypic tube formation assay.;Taken together these data show that PIGF can influence endothelial cell function by controlling the endocytic recycling of alphavbeta3 integrin. This recycling pathway is required to induce vessel branching and the formation of a complex endothelial cell vessel network. In addition, regulation of VEGFR2 recycling by VEGFR1 represents a novel mechanism for mediating receptor cross talk during angiogenesis. Therefore, I have identified a novel pathway by which PIGF/VEGFR1 is able to positively influence the angiogenic response.