ABSTRACT: The repair of corneal damage is crucial for maintaining clear vision. When the corneal epithelium is damaged, epithelial cells at the corneal limbus quickly initiate a series of complex biological processes, including cell migration, extracellular matrix remodeling, and cell proliferation and differentiation, to promote repair and wound healing. However, comprehensive studies on the transcriptional heterogeneity of corneal limbal cell populations during migration, proliferation, and differentiation in the context of corneal epithelial injury repair are still lacking. In this study, we conducted a systematic analysis of five major cell types in the corneal limbus, including terminally differentiated cells of corneal epithelium (TDCE), basal cells (BC), transit-amplifying cells (TAC), limbal stem cells (LSC), and conjunctival cells (CC ), using high-throughput long-read single-cell RNA sequencing of corneal limbal tissues from Macaca fascicularis. The corneal epithelium was examined at three time points: before injury (0 days) and after injury (1 day and 3 days). The study focused on the migration, proliferation, and differentiation dynamics of these cells during different stages of corneal injury repair. Additionally, key regulatory genes and their RNA isoforms were investigated to uncover their roles in these processes. The results revealed that LSCs and BCs play pivotal roles in corneal epithelial healing, highlighting the regulatory functions of several key genes and their RNA isoforms in cell migration, proliferation, and differentiation. These include growth factors (e.g., IGF2), extracellular matrix components (e.g., FN1 and LAMC2), integrins (e.g., ITGB1 and ITGAV), and Keratin (e.g., KRT3, KRT12 and KRT6A). The study provides novel insights into the transcriptional landscape at the single-cell level during corneal epithelial damage repair, identifying important cell types and key regulatory genes with corresponding RNA isoforms.
