AP-2β IN THE CRANIAL NEURAL CREST IS REQUIRED FOR SUB-PROGENITOR DIFFERENTIATION AND LIMBAL EPITHELIAL STEM CELL SURVIVAL

Abstract

The developing cornea partially derives from cranial neural crest cells (NCCs). These cells form the periocular mesenchyme (POM), giving rise to the corneal endothelium and stroma. The transcription factor, activating protein 2β (AP-2β) shows strong expression in the POM and when conditionally deleted in the NCCs of mice (the AP-2β NCC KO model), the endothelium fails to form, the stroma is neovascularized, and the epithelium does not stratify. Further, the corneal epithelium of the AP-2β NCC KO mice was devoid of the corneal epithelial marker, K12, and instead, the limbal epithelial marker, K15 was expressed. These findings suggested AP-2β in the NCC not only plays an important role in stromal and endothelial development, but also has a non-cell autonomous influence on the epithelium, a surface ectoderm-derived (SE) tissue. As the stroma displayed neovascularization, and the epithelium lost its characteristic marker, the AP-2β NCC KO cornea potentially exhibited ‘conjunctivalization,’ caused by limbal epithelial stem cell (LESCs) loss. This thesis investigated the presence LESCs by identifying slow-cycling, label retaining cells in the corneal epithelium with BrdU. Here, a significant reduction in BrdU+ cells was found KO mice. The conjunctival marker, K13, was also found across the corneal epithelium of the KO mice. These findings suggest the loss of LESCs and conjunctivalization of the AP-2β NCC KO cornea. The lineage of the corneal stroma and endothelium was also traced using a novel NCC-Confetti transgenic mouse line. Two sub-NCC progenitor populations were detected in the developing and adult wild-type cornea, both contributing to the endothelium and stroma. In contrast, in the absence of AP-2β in the NCC, one of the populations was lost by postnatal day 15. Overall, these findings indicate that two sub-NCC progenitor populations contribute to the corneal endothelium and stroma and that AP-2β is required for differentiation of the lost NCC progenitor population, leading to abnormalities in the endothelium, stroma, and potentially the epithelium through a non-cell autonomous process(es).