Vascular permeability, angiogenesis and the role of delta-like ligand 4

Cardiovascular disease, of which ischaemic diseases such as stroke and peripheral arterial disease make up a large proportion, are the leading cause of death worldwide. The endogenous response to ischaemia is to upregulate growth factors to stimulate the growth of new vessels and in some cases, form a collateral network. The concept of therapeutic stimulation has therefore become a priority area of cardiovascular research. The collateral network formed must consist of intact, stable, non-leaky vessels that can respond appropriately to stimuli. While the potent angiogenic driver, vascular endothelial growth factor (VEGF) appeared to be a promising target, trials in peripheral ischaemia patients have been disappointing. Upregulated by VEGF during angiogenesis, the Notch ligand Dll4 is a key regulator of vessel maturation and function. Widely known to regulate tip and stalk cell selection during sprouting angiogenesis, its role in the recruitment and growth of mural cells, regulation of permeability, compliance and signalling is poorly understood. Inhibition of Dll4 results in non-functional, poorly perfused vessels with reduced pericyte coverage suggesting a greater role for Dll4 in the formation of mature, intact, operative vessels. I therefore tested the hypothesis that the physiological characteristics of the neovasculature are regulated by Dll4 during physiological neovascularisation. A recombinant human sDll4 protein was used to induce Notch signalling in an endothelial cell monolayer and resulted in increased expression of VE-Cadherin and cell-cell contacts. Using the Landis-Michel microvascular permeability technique, Dll4 signalling was shown to decrease the permeability of rat mesenteric vessels and then subsequent experiments showed that this could be prevented by the proteins kinase A (PKA) inhibitor H89 dihydrochloride. In the rat mesenteric angiogenesis model, inducing Notch with an adenovirus (Ad.) encoding sDll4 (Ad.sDll4) resulted in a less angiogenic vasculature and when added into arteriolargenesis stimulating adenovirus combinations it led to a more endogenous-like vasculature and a switch from sprouting to branching. In a mouse hindlimb ischaemia model, Ad.sDll4 hindered blood flow recovery to the hind paw but led to increase capillary and arteriolar density. These results show that Dll4 plays a key role in regulating vascular permeability and that this is through a cAMP/PKA dependent pathway and involves VE-Cadherin expression. They also demonstrate a role for Dll4 in arteriolargenesis in both physiological and pathological settings and the possibility of using Dll4 as part of an ischaemic therapeutic strategy cannot yet be ruled out