Identification and spatial distribution of novel molecular features of adult human corneal cell subtypes

Abstract

Purpose : The human cornea has been defined as our “external window” to the visual world that serves as a barrier against the outside environment and as the main refractive lens to focus light into the retina. The histological structure is defined by three layers of cellular elements (epithelium, stroma, endothelium) and two layers of extracellular membranes. Here, we used a combination of single-cell RNA sequencing and in-situ hybridization to characterize the transcriptomic features of different cells and their localization in the human cornea

Methods : Six adult human corneas from healthy donors were processed by sequential tissue digestion and cell sorting. Single-cell suspensions were profiled using 10x Genomics Chromium Single Cell 3′ (v2) Gene Expression workflow. Spatial distribution of cell markers was localized by RNA in situ hybridization (RNAscope) in human cornea cross-sections.

Results : Unsupervised clustering of individual cell transcriptomes based on overall gene expression similarity identified 16 transcriptionally distinct clusters within corneal cells, including stromal keratocytes, endothelium, several subtypes of corneal epithelium, and supportive cells in the limbal stem cell niche. Epithelial cells represent the most diverse cell type with eleven sub-clusters. By combining pseudotime bioinformatic analysis and RNAscope we mapped the epithelial cell fate trajectory and location including their initial generation in the limbal region, differentiation, and migration to superficial epithelial layers.

Conclusions : Our study reveals the single-cell map of the adult human cornea and expands the knowledge of the molecularly define cellular subsets of the cornea on a whole genome transcriptional level. This information can be applied to better understand normal corneal biology, serve as a reference to study corneal diseases, and provide potential insights into disease pathology and therapeutics.

This is a 2021 ARVO Annual Meeting abstract.