Single cell RNA-seq of human cornea organoids identifies cell fates of a developing immature cornea

Abstract

The cornea is a protective and refractive barrier in the eye crucial for vision. Understanding the human cornea in health, disease and cell-based treatments can be greatly advanced with cornea organoids developed in culture from induced pluripotent stem cells. While a limited number of studies have investigated the single-cell transcriptomic composition of the human cornea, its organoids have not been examined similarly. Here we elucidated the transcriptomic cell fate map of 4-month-old human cornea organoids and human donor corneas. The organoids harbor cell clusters that resemble cells of the corneal epithelium, stroma and endothelium with sub-populations that capture signatures of early developmental states. Unlike the adult cornea where the largest cell population is stromal, the organoids contain large proportions of epithelial and endothelial-like cells. These corneal organoids offer a three-dimensional model to study corneal diseases and integrated responses of different cell types.

Single-cell transcriptomic analysis of corneal organoids during development

“…KRT15 was in limbal epithelium ( Figure 5 D), while KRT7 was expressed in the conjunctival epithelium ( Figure 5 E). Curiously, and similar to the findings of Maiti et al. (2022) , there was very low expression of KRT3 or KRT12 ( Figure 4 B), the definitive mature corneal epithelium markers ( Figure 5 F) ( Collin et al., 2021 ).…”

“…(2022) , there was very low expression of KRT3 or KRT12 ( Figure 4 B), the definitive mature corneal epithelium markers ( Figure 5 F) ( Collin et al., 2021 ). We did not observe any expression of myofibroblast markers MYH6, MYL3, or MYL4 ( Figure S4 ) at the 4-month time point as was observed by Maiti et al. (2022) in their organoids.…”

“…dataset in each of these bins, we observed that across organoid development, the proportion of cells mapping to the PCW 10–14 corneas decreased while the proportion of cells mapping to the adult cornea increased ( Figure 6 C). However, even at month 4, there were still <40% of cells mapping to adult cornea ( Figure 6 C), indicating that the month 4 organoid described in this as well as a previous study ( Maiti et al., 2022 ) does not best represent fully mature adult cornea, and that extending organoid growth beyond this time may be beneficial for modeling adult corneal diseases. A detailed fetal cornea and corneal organoid development comparison is shown in Figure S5 ( Eghrari et al., 2015b ).…”

“…Ocular, corneal, retinal, and lacrimal organoid cultures have been described using iPSCs or tissue biopsies with the immense potential for studying ocular disease development, testing therapeutics, and preparing transplantable tissue ( Foster et al., 2017 ; Susaimanickam et al., 2017 ; Fligor et al., 2018 ; Manafi et al., 2021 ). While there have been single-cell RNA-seq (scRNA-seq) studies characterizing retinal ( Fligor et al., 2018 ), lacrimal ( Hayashi et al., 2022 ), and more recently corneal organoids ( Maiti et al., 2022 ) and comparing them with the fetal and adult human cornea ( Collin et al., 2021 ), there have been no studies specifically analyzing corneal organoids during their differentiation and maturation. In this study, we classify the cell types of developing corneal organoids using scRNA-seq and compare them with fetal and adult human corneas to determine if the organoids recapitulate normal human corneal growth and development.…”

Analysis of gene expression dynamics and differential expression in viral infections using generalized linear models and quasi-likelihood methods

“… Wang et al (2022) explored patient-derived organoid cultures to study colorectal cancer (CRC) mechanisms, revealing how culture methods influence gene expression patterns. Maiti et al (2022) dissected human cornea organoids and donor corneas, elucidating early developmental states and their relevance for corneal diseases. Kathuria et al (2020) generated cerebral organoids from individuals with bipolar disorder, uncovering gene expression differences and synaptic impairments.…”