PlexinD1 is required for proper patterning of the periocular vascular network and for the establishment of corneal avascularity during avian ocular development

Abstract

The anterior eye is comprised of an avascular cornea surrounded by a dense periocular vascular network and therefore serves as an excellent model for angiogenesis. Although signaling through PlexinD1 underlies various vascular patterning events during embryonic development, its role during the formation of the periocular vascular network is yet to be determined. Our recent study showed that PlexinD1 mRNA is expressed by periocular angioblasts and blood vessels during ocular vasculogenesis in patterns that suggest its involvement with Sema3 ligands that are concurrently expressed in the anterior eye. In this study, we used in vivo knockdown experiments to determine the role of PlexinD1 during vascular patterning in the anterior eye of the developing avian embryos. Knockdown of PlexinD1 in the anterior eye caused mispatterning of the vascular network in the presumptive iris, which was accompanied by lose of vascular integrity and profuse hemorrhaging in the anterior chamber. We also observed ectopic vascularization of the cornea in PlexinD1 knockdown eyes, which coincided with the formation of the limbal vasculature in controls. Finally we show that Sema3E and Sema3C transcripts are expressed in ocular tissue that is devoid of vasculature. These results indicate that PlexinD1 plays a critical role during vascular patterning in the iris and limbus, and is essential for the establishment of corneal avascularity during development. We conclude that PlexinD1 is involved in vascular response to antiangiogenic Sema3 signaling that guides the formation of the iris and limbal blood vessels by inhibiting VEGF signaling.

Vascular Sema3E-Plexin-D1 Signaling Reactivation Promotes Post-stroke Recovery through VEGF Downregulation in Mice

“…Previously, the absence of this signaling pathway was thought to unlock the normal limits on vessel growth, thereby increasing the size and branching of vascular networks. This has been observed in mouse peripheral and avian periocular vasculature; vessels devoid of Sema3E or Plexin-D1 grow densely across their boundaries [13,64]. However, this mechanism is not observed in the retinal vasculature, where Sema3e or Plxnd1 knockout results in sparse and underdeveloped network formation.…”

Knockdown of CXCL14 disrupts neurovascular patterning during ocular development

“…Since expression of CXCL14 in the embryonic cornea (Fig. 1A–C) coincides with the period of sustained corneal avascularity during the formation of the dense vascular network in the adjacent limbus region (Kwiatkowski et al, 2013; 2016), we investigated whether it is involved in preventing corneal vascularization. First, we performed bead implantation experiments to test whether addition of recombinant CXCL14 would inhibit induced vascularization of the cornea during development.…”

“…Formation of the dense limbal vascular network is underway by this time (Kwiatkowski et al, 2013). The pericorneal nerves and vasculature both express Nrp1 and they are repelled from the presumptive cornea by Semaphorins secreted by the cornea and adjacent lens (Lwigale and Bronner-Fraser, 2007; McKenna et al, 2012; 2014; Kwiatkowski et al, 2016). The cornea remains un-innervated and avascular until about E10 when only sensory nerves are permitted to project into its periphery.…”

“…Our findings are also consistent with a previous study showing that the efficacy of the VEGF inhibitor (Sorafenib) used in cancer antiangiogenic therapy depends on the induction of CXCL14 (Rivera et al, 2015). Interestingly, neovascularization of the CXCL14 knockdown corneas occurred later in development, suggesting that CXCL14 is dispensable as an antiangiogenic factor during early corneal development, which is regulated by Semaphorins (Kwiatkowski et al, 2013; 2016; McKenna et al, 2015). We propose that CXCL14 sustains antiangiogenic signaling to maintain corneal avascularity as the repulsive Semaphorin signals weaken to permit innervation.…”

“…This process is influenced by VEGF and possibly, other angiogenic factors expressed by the ocular tissues (Kwiatkowski et al, 2013). Similar to neural guidance, vasculogenesis in the anterior eye is regulated by Semaphorin signaling through Neuropilin-1 and PlexinD receptors that are expressed by the angioblasts and nascent vasculature (Kwiatkowski et al, 2013; McKenna et al, 2014; and Kwiatkowski et al, 2016). …”