Angiogenesis does not only depend on endothelial cell invasion and proliferation: it also requires pericyte coverage of vascular sprouts for vessel stabilization1, 2. These processes are coordinated by vascular endothelial growth factor (VEGF) and platelet-derived growth factor (PDGF) through their cognate receptors on endothelial cells and vascular smooth muscle cells (VSMCs), respectively3, 4. PDGF induces neovascularization by priming VSMCs/pericytes to release pro-angiogenic mediators5, 6, 7. Although VEGF directly stimulates endothelial cell proliferation and migration, its role in pericyte biology is less clear. Here we define a role for VEGF as an inhibitor of neovascularization on the basis of its capacity to disrupt VSMC function. Specifically, under conditions of PDGF-mediated angiogenesis, VEGF ablates pericyte coverage of nascent vascular sprouts, leading to vessel destabilization. At the molecular level, VEGF-mediated activation of VEGF-R2 suppresses PDGF-Rbeta signalling in VSMCs through the assembly of a previously undescribed receptor complex consisting of PDGF-Rbeta and VEGF-R2. Inhibition of VEGF-R2 not only prevents assembly of this receptor complex but also restores angiogenesis in tissues exposed to both VEGF and PDGF. Finally, genetic deletion of tumour cell VEGF disrupts PDGF-Rbeta/VEGF-R2 complex formation and increases tumour vessel maturation. These findings underscore the importance of VSMCs/pericytes in neovascularization8, 9 and reveal a dichotomous role for VEGF and VEGF-R2 signalling as both a promoter of endothelial cell function and a negative regulator of VSMCs and vessel maturation.

Nature advance online publication 9 November 2008 | doi:10.1038/nature07424; Received 11 March 2008; Accepted 10 September 2008; Published online 9 November 2008

Infantile hemangiomas are localized and rapidly growing regions of disorganized angiogenesis. We show that expression of vascular endothelial growth factor receptor-1 (VEGFR1) in hemangioma endothelial cells (hemECs) and hemangioma tissue is markedly reduced compared to controls. Low VEGFR1 expression in hemECs results in VEGF-dependent activation of VEGFR2 and downstream signaling pathways. In hemECs, transcription of the gene encoding VEGFR1 (FLT1) is dependent on nuclear factor of activated T cells (NFAT). Low VEGFR1 expression in hemECs is caused by reduced activity of a pathway involving beta1 integrin, the integrin-like receptor tumor endothelial marker-8 (TEM8), VEGFR2 and NFAT. In a subset of individuals with hemangioma, we found missense mutations in the genes encoding VEGFR2 (KDR) and TEM8 (ANTXR1). These mutations result in increased interactions among VEGFR2, TEM8 and beta1 integrin proteins and in inhibition of integrin activity. Normalization of the constitutive VEGFR2 signaling in hemECs with soluble VEGFR1 or antibodies that neutralize VEGF or stimulate beta1 integrin suggests that local administration of these or similar agents may be effective in hemangioma treatment.

Nature Medicine
Published online: 19 October 2008 | doi:10.1038/nm.1877

Serum response factor (SRF) is a transcription factor that controls the expression of cytoskeletal proteins and immediate early genes in different cell types. Here, we found that SRF expression is restricted to endothelial cells (ECs) of small vessels such as capillaries in the mouse embryo. EC-specific Srf deletion led to aneurysms and hemorrhages from 11.5 days of mouse development (E11.5) and lethality at E14.5. Mutant embryos presented a reduced capillary density and defects in EC migration, with fewer numbers of filopodia in tip cells and ECs showing defects in actin polymerization and intercellular junctions. We show that SRF is essential for the expression of VE-cadherin and β-actin in ECs both in vivo and in vitro. Moreover, knockdown of SRF in ECs impaired VEGF- and FGF-induced in vitro angiogenesis. Taken together, our results demonstrate that SRF plays an important role in sprouting angiogenesis and small vessel integrity in the mouse embryo.

Developmental Cell, Vol 15, 448-461, 16 September 2008

Purhonen et al. (1) have refuted the data published in >50 reports (2, 3), neglecting to quote key articles or utilize relevant models, and have drawn unsubstantiated conclusions about the contribution of endothelial progenitor cells (EPCs) to tumor angiogenesis that are not supported by their nonquantitative data and superficially executed experiments. Their study (1) is flawed in experimental design and data interpretation. For example, they do not cite their own publication demonstrating the existence of VEGFR2+ EPCs (4) and neglect mentioning clinical validation (5, 6) and acknowledging mouse genetic models (2, 3), which provide convincing evidence for functional incorporation of EPCs into neovessels. Every figure lacks stereoconfocal-microscopic quantification of vessels that are presented as poorly defined longitudinal–linear streaks. Plasma VEGF-A levels were not measured in vivo in mice treated with VEGF-A, questioning their low level of VEGFR2+ EPC detection (3). Indeed, their FACS analysis is inaccurate because of (i) unconvincing CD31/VE-cadherin/VEGFR2 expression detected on MS-1 endothelium used as positive control and (ii) failure to show long-term marrow engraftment of donor-derived hematopoietic and authentic VEGFR2+LacZ+ colony-forming EPCs. APCmin mice develop only obstructive adenomas, rather than adenocarcinomas; therefore, it is an inappropriate model to study EPC incorporation, as Spring et al. (7) (not quoted) demonstrate that EPCs do not contribute to adenomas but contribute only to carcinomas/metastatic tumors. In the parabiotic model, wild-type EPCs compete with GPF+ EPCs, which underestimates EPC recruitment. Finally, study of 6-month-old VEGF-A-loaded Matrigel plugs in mice is impossible because Matrigel plugs are degraded within 2 months, particularly when VEGF-A by itself does not induce neoangiogenesis. No quantification of patent vessels in Matrigel plugs was provided. This article fails to disprove the established role of EPCs in supporting neoangiogenesis in certain tumors (3, 5) and metastatic transition (2)

PNAS 2008 105:E54; published ahead of print August 20, 2008, doi:10.1073/pnas.0804876105