Vertebrate embryos as tools for anti-angiogenic drug screening and function

Abstract

The development of new angiogenic inhibitors highlights a need for robust screening assays that adequately capture the complexity of vessel formation, and allow for the quantitative evaluation of the teratogenicity of new anti-angiogenic agents. This review discusses the use of screening assays in vertebrate embryos, specifically focusing upon chicken and zebrafish embryos, for the detection of anti-angiogenic agents.

Keywords: Angiogenesis; Chicken; Mouse; Non-human primates; Rabbit; Rat; Thalidomide; Zebrafish.

Fig. 1.

Vasculogenesis and angiogenesis. Endothelial cell precursors (angioblasts) are derived from mesodermal stem cells. Angioblasts are promoted by vascular endothelial growth factor (VEGF) and its co-receptor neuropilin-1, to migrate, differentiate, and assemble into endothelial cords. The primitive vascular plexus forms from merging of the endothelial cords with the endocardial tubes (vasculogenesis). The expansion and remodeling (angiogenesis) leads to stabilized vessels in a vascular network. This is mediated by VEGF, as well as other factors (such as Sonic hedgehog and Notch signaling). Vessels are stabilized by mural cells, a process mediated by platelet derived growth factor B (PDGF) acting upon its receptor (PDG FR). Adapted from [160] and [161].

Fig. 2.

Molecular mechanisms controlling angiogenesis.

Fig. 3.

Angiogenesis and angiogenesis inhibitors. (A) Angiogenesis is promoted by tumour cells and neighbouring stromal cells. Endothelial cells are activated by pro-angiogenic factors released by these cells and they begin to migrate. They then proliferate and stabilize, while the angiogenic factors continue to promote the process in a feedback loop. Vessels penetrate the environment of the cancerous cells allowing the tumour to grow. (B) At the molecular level, angiogenesis inhibitors reduce the inappropriate blood vessel growth by targeting VEGF receptors directly (sunitinib, sorafenib, axitinib), a downstream molecular target (sorafenib, vandetanib, pazopanib) or by acting within the tumour cells themselves to inhibit the production of pro-angiogenic factors such as VEGF (everolimus) [54,162]

Fig. 4.

Angiogenesis in the zebrafish and developing chicken embryo as a tool for assessing drug action. (A) Normal vascular development at 24 h post fertilization and (B) reduced vascular development in an anti-angiogenic drug (sunitinib) treated transgenic fli1:EGFP embryo. Angiogenesis is quantified by measuring intersomitic vessel (ISV) number and outgrowth. Selected ISVs are indicated by white arrow heads. The anterior (head) and posterior (tail) regions are indicated. The yolk sac (YS) is indicated. (C) A chicken embryo with normal eye (e), limb bud (1) and spinal (s) development (D) Chicken embryo following treatment for 24 h with an anti-angiogenic drug (sunitinib). Note hemorrhagic tissues (black asterisk) and bent spine of the embryo (black arrowhead). The chorioallantoic membrane (CAM) and yolk sac membrane (YSM) are indicated. Images are from previously unpublished work of Beedie, Figg and Vargesson from a published study [54].