Purchase this article with an account.
Philip Gage, Amanda Zacharias, Chen Kuang; The homeodomain transcription factor PITX2 is required to establish correct cell lineages and angiogenic privilege in the developing cornea. Invest. Ophthalmol. Vis. Sci. 2013;54(15):4719.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
Vision depends critically on normal corneal development, including correct specification of the epithelium (from surface ectoderm), stroma and endothelium (from mesenchyme), and establishment of an avascular environment (angiogenic privilege). Phenotypes of humans and mice with heterozygous mutations have suggested PITX2 as an important mediator of these processes but anterior segment morphogenesis in global and tissue-specific Pitx2 knockout mice is blocked prior to initiation of corneal development. Therefore, we utilized our conditional Pitx2 allele together with a tamoxifen-inducible Cre recombinase to generate temporal knockout animals (Pitx2-tko) and assess gene function in the developing cornea.
Appropriate strains were crossed to generate control and conditional mutant embryos. Timed pregnant females were injected with a single dose of tamoxifen early in corneal development to temporally ablate Pitx2 in the mutant embryos. Immunohistochemistry or RNA in situ hybridization was employed to assess gene expression.
Ablation of Pitx2 at e11.5 rescues the early block to anterior segment development but significantly disrupts formation of all three corneal layers. Gene expression is altered in all three layers of Pitx2-tko corneas and the surface ectoderm and mesenchyme appear to adopt a conjunctival fate based on ectopic expression on specific marker expression, suggesting that PITX2 is required cell-autonomously for specification of the stroma and endothelium and non-cell autonomously for specification of the epithelium. The mutant corneal mesenchyme is highly vascularized, indicating that PITX2 is required to establish angiogenic privilege. Dkk2 expression is not maintained in mutant eyes and there is a corresponding increase in canonical Wnt signaling activity. Vegfa and Pdgfa expression are also increased.
Collectively, these results suggest that PITX2-mediated suppression of canonical Wnt signaling may be an important mechanism required to correctly specify cell lineages and establish angiogenic privilege during corneal development. We are testing this hypothesis further by activating canonical Wnt signaling activity in the corneal ectoderm and mesenchyme independently of mutations in Pitx2.
This PDF is available to Subscribers Only