Exploring a new model to study developmental eye diseases

项目总结

PROJECT SUMMARY Defects in human eye development can result in a broad spectrum of ocular disorders, many of which lead to partial or complete vision loss. The human eye is comprised of many specialized tissues derived from different primordial cell lineages, and their formation is highly coordinated over time, making ocular development a very complex process. Human ocular cell culture systems commonly used for disease modeling fail to recapitulate earlier stages of this stepwise developmental process. Consequently, it remains a challenge to efficiently test how patient variants impact developing structures within the eye, thus contributing to the observed complex ocular phenotypes. To overcome this barrier, we aim to explore the recently presented cell culture model where human iPSCs are induced to form multiple ocular cell types organized within four identifiable, concentric zones - referred to as self-formed ectodermal autonomous multi-zone of ocular cells (SEAMs). Uniquely, each zone is comprised of different cell types found within the eye, including neuroretina, retinal pigmented epithelium, cornea and lens, and zone formation occurs progressively over time, mimicking the timing and cell-cell coordination observed during human eye development. The primary objective of this proposal is to delineate quantifiable morphological benchmarks and gene expression signatures associated with SEAM generation under normal conditions and then test this model by introducing disease-associated genetic variants in well-known ocular genes. Specifically, we aim: (1) To characterize SEAM morphology, variability and gene/protein expression during their formation; (2) To explore the effects of pathogenic human variants in key ocular genes on SEAM formation. In our approach we will measure changes in size, shape, and other physical characteristics and appearance of SEAMs at four timepoints along the differentiation process. Concurrent scRNA-seq analysis will define the identity of the cell types within the multi-zones and determine their equivalence to developing human ocular tissues. These analyses will be established first under normal conditions and then extended to assessing SEAMs derived from hiPSCs carrying pathogenic variants in well-known ocular genes (PAX6 and FOXE3). Completion of these studies will identify strengths and possible limitations of this innovative model system for studying human whole eye development. Thus, this study will likely provide a platform for testing a wide range of genetic variants identified in congenital ocular phenotypes including identification of the direct and secondary effects of those variants on coordinated development of various ocular tissues.