Mechanisms of vascular stabilization by PDGF-BB

Peripheral artery disease is a chronic condition characterised by the narrowing of peripheral arteries due to atherosclerotic plaques and subsequent decrease in blood supply. The disease mainly affects the lower limbs and it manifests clinically as limb claudication in the early stages and can progress to a more severe form characterized by rest pain and tissue loss. Morbidity and mortality are high amongst individuals affected by the disease and although lifestyle changes and medical therapy have shown some efficiency, surgical and mini-invasive revascularization are still the mainstays of treatment. Novel strategies should be investigated in order to promote the formation of new collateral arteries (arteriogenesis) and/or capillaries (angiogenesis) and restore blood flow. Vascular Endothelial Growth Factor (VEGF) is the master regulator of angiogenesis, and its delivery in ischemic tissues has been associated with improved collateral artery development. However, its therapeutic potential is challenged by the need to control both the dose and duration of expression: sustained and uncontrolled levels cause the growth of angioma-like structures, whereas transient delivery shorter than about 4 weeks is insufficient for stabilization and persistence of induced vessels. We previously found that: 1) increasing VEGF doses impair vessel stabilization by inhibiting the endothelial Semaphorin3A/Neuropilin1+ monocytes (NEM)/TGF-b1 paracrine axis, and 2) PDGF-BB co-expression restores normal angiogenesis despite high VEGF levels. Here we investigated whether and how PDGF-BB accelerates the stabilization of VEGF dose-dependent angiogenesis. For this purpose, mouse muscles were implanted with monoclonal myoblast populations homogeneously producing low, medium or high VEGF levels, with or without PDGF-BB. VEGF signaling was abrogated after 2 and 3 weeks by systemic treatment with the receptor-body Aflibercept. As expected, increasing VEGF doses progressively impaired stabilization. PDGF-BB co-expression did not change the effects of low and medium VEGF, but it greatly accelerated vascular stabilization at high VEGF (80% at 3 weeks vs 0% with VEGF alone). Aberrant vascular structures were switched to normal pericyte-covered capillaries, similar to those induced by low VEGF alone. However, stabilization was significantly greater than with low VEGF (50%), suggesting pericyte restoration may not be the only mechanism. Moreover, PDGF-BB dose-dependently stimulated endothelial Sema3A expression in vitro and restored Sema3A production, NEM recruitment and TGF-b1 level in vivo despite high VEGF levels. These results suggest that PDGF-BB dose-dependently accelerates vascular stabilization independently of pericytes recruitment through Sema3A expression and the NEM/TGF-b1 axis. In conclusion, PDGF-BB co-delivery has the potential to overcome the limitations of VEGF gene delivery (dose and duration), providing a rational strategy to improve the clinical outcome of patients with limb ischemia

Extracellular matrix and growth factor engineering for controlled angiogenesis in regenerative medicine

Blood vessel growth plays a key role in regenerative medicine, both to restore blood supply to ischemic tissues and to ensure rapid vascularization of clinical-size tissue-engineered grafts. For example, vascular endothelial growth factor (VEGF) is the master regulator of physiological blood vessel growth and is one of the main molecular targets of therapeutic angiogenesis approaches. However, angiogenesis is a complex process and there is a need to develop rational therapeutic strategies based on a firm understanding of basic vascular biology principles, as evidenced by the disappointing results of initial clinical trials of angiogenic factor delivery. In particular, the spatial localization of angiogenic signals in the extracellular matrix (ECM) is crucial to ensure the proper assembly and maturation of new vascular structures. Here, we discuss the therapeutic implications of matrix interactions of angiogenic factors, with a special emphasis on VEGF, as well as provide an overview of current approaches, based on protein and biomaterial engineering that mimic the regulatory functions of ECM to optimize the signaling microenvironment of vascular growth factors

VEGF dose regulates vascular stabilization through Semaphorin3A and the Neuropilin-1+ monocyte/TGF-β1 paracrine axis

VEGF is widely investigated for therapeutic angiogenesis, but while short-term delivery is desirable for safety, it is insufficient for new vessel persistence, jeopardizing efficacy. Here, we investigated whether and how VEGF dose regulates nascent vessel stabilization, to identify novel therapeutic targets. Monoclonal populations of transduced myoblasts were used to homogeneously express specific VEGF doses in SCID mouse muscles. VEGF was abrogated after 10 and 17 days by Aflibercept treatment. Vascular stabilization was fastest with low VEGF, but delayed or prevented by higher doses, without affecting pericyte coverage. Rather, VEGF dose-dependently inhibited endothelial Semaphorin3A expression, thereby impairing recruitment of Neuropilin-1-expressing monocytes (NEM), TGF-β1 production and endothelial SMAD2/3 activation. TGF-β1 further initiated a feedback loop stimulating endothelial Semaphorin3A expression, thereby amplifying the stabilizing signals. Blocking experiments showed that NEM recruitment required endogenous Semaphorin3A and that TGF-β1 was necessary to start the Semaphorin3A/NEM axis. Conversely, Semaphorin3A treatment promoted NEM recruitment and vessel stabilization despite high VEGF doses or transient adenoviral delivery. Therefore, VEGF inhibits the endothelial Semaphorin3A/NEM/TGF-β1 paracrine axis and Semaphorin3A treatment accelerates stabilization of VEGF-induced angiogenesis