Induction of functional neovascularization by combined VEGF and angiopoietin-1 gene transfer using AAV vectors

Abstract

Vectors based on the adeno-associated virus (AAV) deliver therapeutic genes to muscle and heart at high efficiency and maintain transgene expression for long periods of time. Here we report about the synergistic effect on blood vessel formation of AAV vectors expressing the 165 aa isoform of vascular endothelial growth factor (VEGF165), a powerful activator of endothelial cells, and of angiopoietin-1 (Ang-1), which is required for vessel maturation. High titer AAV-VEGF165 and AAV-Ang-1 vector preparations were injected either alone or in combination in the normoperfused tibialis anterior muscle of rats. Long term expression of VEGF165 determined massive cellular infiltration of the muscle tissues over time, with the formation of a large set of new vessels. Strikingly, some of the cells infiltrating the treated muscles were found positive for markers of activated endothelial precursors (VEGFR-2/KDR and Tie-2) and for c-kit, an antigen expressed by pluripotent bone marrow stem cells. Expression of VEGF165 eventually resulted in the formation of structured vessels surrounded by a layer of smooth muscle cells. Presence of these arteriolae correlated with significantly increased blood perfusion in the injected areas. Co-expression of VEGF165 with angiopoietin-1-which did not display angiogenic effect per se-remarkably reduced leakage of vessels produced by VEGF165 alone.

Additive effect of AAV-mediated angiopoietin-1 and VEGF expression on the therapy of infarcted heart

“…Although co-expression of VEGF and ang-1 in ischemic hindlimb and heart has been proven to induce leakage resistant vessels and resulted in some improvements in angiogenesis and therapy (11)(12)(13)(14)(15)(16)(17)(18)(19)(20)(21). Most of the experiments used plasmid as the vector, which only results in short-term gene expression.…”

“…We used AAV, a vector that can mediate long-lasting transgene expression in tissues (22), to deliver ang-1 and VEGF genes into ischemic mouse myocardium. AAV vector has only been used to mediate co-expression of VEGF and ang-1 in hindlimb (14,19). Although skeletal and cardiac muscles have many similarities, the results obtained from hindlimb may not apply to the heart directly.…”

The enigmatic role of angiopoietin-1 in tumor angiogenesis

“…Since Ang1 is not a mitogen, the increased vascular branching may arise from reinforcement of VEGF-induced angiogenesis. Indeed, Ang1 has been shown to synergise with VEGF to enhance angiogenesis in the rat aorta model [33] and increase vessel density in the corneal implant assay [34] and several other in vivo assays [35][36][37][38]. While overexpression of VEGF alone gives rise to increased vascular branching, the vessels induced by VEGF are leaky [32].…”

“…Tumor vessels in the Ang1-transfected breast [50], colon [69], hepatic colon tumor [88], and squamous cell [77] xenografts mentioned above all demonstrate significantly increased association of pericytes with vessels, suggesting that enforced maturation of blood vessels may functionally inhibit tumor angiogenesis. First of all, the ability of Ang1 to inhibit vascular permeability is thought to be due partly to the enhancement of cell-cell junctions [40], as well stabilization of blood vessels by the promotion of mural cell recruitment [19,34,35,38]; it is interesting to note that the absence of pericytes leads to defects in endothelial junction formation [16]. In addition, peripheral blood vessels in the Ang1-transfected MCF7 tumors were not dilated, in contrast to those in the vector control counterparts [50].…”

High efficient adenoviral-mediated VEGF and Ang-1 gene delivery into osteogenically differentiated human mesenchymal stem cells

“…In this context, it has been demonstrated that angiopoietin-1 (Ang-1) plays an important role for stabilization and maturation of blood vessels by inducing the recruitment of perivascular mural cells such as pericytes and smooth muscle cells (Thurston et al, 1999;Thurston et al, 2000;Gamble et al, 2000). In the context of therapeutic angiogenesis it was recently shown that a combined gene transfer of VEGF and Ang-1 leads to the formation of functionally stable blood vessels with no signs of plasma leakage (Arsic et al, 2003).…”