PDGF-BB regulates splitting angiogenesis in skeletal muscle by limiting VEGF-induced endothelial proliferation

R Gianni-Barrera,M G Burger,M Bartolomeo,R Hlushchuk,D J Schaefer,P Valente,E Groppa,A Uccelli,A Banfi,V Djonov,M Heberer,L Gürke,A Certelli,B Vollmar,A Butschkau

doi:10.1007/s10456-018-9634-5

Abstract

VEGF induces normal or aberrant angiogenesis depending on its dose in the microenvironment around each producing cell in vivo. This transition depends on the balance between VEGF-induced endothelial stimulation and PDGF-BB-mediated pericyte recruitment, and co-expression of PDGF-BB normalizes aberrant angiogenesis despite high VEGF doses. We recently found that VEGF over-expression induces angiogenesis in skeletal muscle through an initial circumferential vascular enlargement followed by longitudinal splitting, rather than sprouting. Here we investigated the cellular mechanism by which PDGF-BB co-expression normalizes VEGF-induced aberrant angiogenesis. Monoclonal populations of transduced myoblasts, expressing similarly high levels of VEGF alone or with PDGF-BB, were implanted in mouse skeletal muscles. PDGF-BB co-expression did not promote sprouting and angiogenesis that occurred through vascular enlargement and splitting. However, enlargements were significantly smaller in diameter, due to a significant reduction in endothelial proliferation, and retained pericytes, which were otherwise lost with high VEGF alone. A time-course of histological analyses and repetitive intravital imaging showed that PDGF-BB co-expression anticipated the initiation of vascular enlargement and markedly accelerated the splitting process. Interestingly, quantification during in vivo imaging suggested that a global reduction in shear stress favored the initiation of transluminal pillar formation during VEGF-induced splitting angiogenesis. Quantification of target gene expression showed that VEGF-R2 signaling output was significantly reduced by PDGF-BB co-expression compared to VEGF alone. In conclusion, PDGF-BB co-expression prevents VEGF-induced aberrant angiogenesis by modulating VEGF-R2 signaling and endothelial proliferation, thereby limiting the degree of circumferential enlargement and enabling efficient completion of vascular splitting into normal capillary networks despite high VEGF doses.

Highlights

Coronary and peripheral artery diseases (CAD and PAD) remain the major causes of morbidity and mortality in Western countries, despite current medical and surgical options [1]
Platelet-Derived Growth Factor-BB (PDGF-BB) co-delivery caused vessel length density (VLD) to be already doubled compared to controls by 3 days (Fig. 3g; Ctrl = 10.0 ± 0.5 and VIPhigh = 23.3 ± 3.9 mm/ mm2; p < 0.05), reaching a sixfold increase by 7 days (Fig. 3g; Ctrl. = 11.1 ± 0.8 and VIPhigh = 65.1 ± 6.2 mm/mm2; p < 0.0001). These results indicate that PDGF-BB coexpression (a) limits the degree of vascular enlargement induced by high levels of VEGF; and (b) enables completion of vascular splitting leading to robust generation of normal capillary networks
Taking advantage of a highly controlled myoblast-based gene delivery platform, here we found that PDGF-BB normalizes aberrant angiogenesis by high levels of VEGF in the therapeutic target tissue of skeletal muscle by enabling efficient vascular splitting and without inducing any sprouting

Summary

Introduction

Coronary and peripheral artery diseases (CAD and PAD) remain the major causes of morbidity and mortality in Western countries, despite current medical and surgical options [1]. In order to investigate how VEGF gene delivery regulates vascular growth in the therapeutic target tissue of skeletal muscle, we have previously developed a highly controlled gene delivery platform, based on monoclonal populations of transduced myoblasts that ensure homogeneous expression of specific doses or combinations of angiogenic factors [6, 7] and we found that (1) VEGF induces either physiological microvascular networks or aberrant angioma-like structures depending on its dose localized in the microenvironment around each producing cell in vivo [7]; (2) at the doses required for functional ischemia relief [8], VEGF induces muscle angiogenesis essentially without sprouting and rather by vascular splitting [9]; and (3) the switch from normal to aberrant angiogenesis does not depend exclusively on VEGF dose, but rather on the balance between endothelial stimulation by VEGF and pericyte recruitment by Platelet-Derived Growth Factor-BB (PDGF-BB), such that PDGF-BB codelivery ensures normal and functional microvascular growth despite high or uncontrolled VEGF expression [6, 10]. VEGF-induced angiogenesis in skeletal muscle entailed two distinct phases, namely, an initial stage of circumferential vascular enlargement during the first 4 days, followed by longitudinal splitting that is complete within day 7 [9]

Methods

Results

Conclusion