Inhibition, VegfR2 leads to Reduction, Angiogenesis

VEGF signals promote endothelial cell motility, filopodial extension and proliferation, and together with Notch signaling controls whether specific endothelial cells (ECs) become pioneering ‘EC-tip’ cells (non-proliferating) or trailing ‘EC-stalk’ cells (proliferating). VEGFR2 activation is the master switch that promotes motility and exploratory behaviors of leading EC-tip cells and a mitogenic effect on trailing EC-stalk cells [EIlken and Adams, 2010; Herbert and Stanier 2011; Blanco and Gerhardt, 2013]. An early step is EC-tip cell selection [Eilken and Adams, 2010]. Endothelial cells are normally suppressed in their tip cell behaviors by Notch-Delta signaling [Blanco and Gerhardt, 2013; Li et al. 2014]. This lateral inhibition is broken when VEGFR2 is activated by VEGF-A.  Delta-like 4 (Dll4), a membrane-bound ligand for Notch1 and Notch4, is selectively expressed in response to VEGF-A induction. This down-regulates VEGFR-2 expression in prospective EC-stalk cells but promotes VEGFR2 expression in EC-tip cells, enabling them to extend filopodial processes along VEGF-A rich paths thus orienting the angiogenic sprout [Williams et al. 2006]. VEGF-A rich corridors are established during in vivo development by local VEGFA gradients and the distribution of soluble VEGFR-1, a so-called ‘decoy receptor’ sequestered and released during enzymatic remodeling of ECM, both serving to channel sprouting progression along VEGFA-rich corridors [Roberts et al. 2004; Chappell et al. 2009 and 2016].

: Studies with pharmacological VEGFR2 inhibitors have shown their concentration dependent effect on angiogenic sprouting. For example, the VEGFR2 antagonist Vatalanib (PTK787) suppressed zebrafish ISV outgrowth in a concentration-dependent manner that was characterized quantitatively at 72 hours post-fertilization (hpf) and became evident at the 0.07 µM concentration level [Tal et al. 2014]. An even lower concentration of Vatalanib (0.01 µM) inhibited angiogenic sprouting dynamics in a 3D microsystem of human endothelial cells derived from induced pluripotent stem cells (iPSC-ECs) [Belair et al. 2016b]. The response-response relationship for Vatalnib in zebrafish was maintained for dysmorphogenesis at 120 hpf (0.22 µM) and adult survival curves at 10 days (0.70 µM) [Tal et al. 2014]. While Vatalanib inhibits both VEGFR-2 and PDGFRβ, it is most selective for VEGFR-2 [Wood et al. 2000].

Shirinifard et al. [2013] examined angiogenic sprouting dynamics in zebrafish embryos exposed to high concentrations of arsenic (As). This resulted in a suppressed but chaotic pattern of ISV outgrowth. Quantitative mathematical models inferred increased exploratory filopodial behaviors of EC-tip cells accounting for the loss of directional sensing of during ISV outgrowth [Shirinifard et al. 2013]. The chaotic versus ordered EC-tip cell dynamics may be mechanistically linked to key modulatory factors that regulate the cytoskeletal cycle and/or cell-ECM biomechanics. Molecular pathways such as the Aryl hydrocarbon receptor (AhR) and hypoxia-inducible factor-1 alpha (HIF-1α) that control genes in response to xenobiotic metabolism, hypoxia, and hypoglycemia have potential feedback roles. These pathways regulate genes in developmental angiogenesis. For example, functional inactivation of ARNT, the AhR nuclear translocator protein, results in critical embryonic vascular phenotypes in the yolk sac and branchial arches reminiscent of those observed in mouse embryos deficient in VEGF-signaling [Maltepe et al. 1997].

Domain of Applicability: The de novo assembly of endothelial cells into the primitive capillary network in an early embryo (vasculogenesis) or a tubular network in vitro (tubulogenesis) are both driven by VEGF-A signaling. A critical effect on developmental angiogenesis aligns with the Gene Ontology (GO) term ‘negative regulation of blood vessel morphogenesis’ (GO:0016525), defined as “Any process that stops, prevents, or reduces the frequency, rate or extent of angiogenesis”. Differences exist among the 110 genes mapped to this annotation in the Mouse Gene Ontology Browser (http://www.informatics.jax.org/vocab/gene_ontology/, last accessed November 30, 2021). Although the genetic signals and responses may differ between vasculogenesis and angiogenesis [Drake et al. 2007; Knudsen and Kleinstreuer, 2011], disruption of the former process ultimately leads to a reduction in the latter during development and so both are in the DoA for this KER.