Generation of a Novel Immortalized Human Corneal Epithelial Cell Line for Reliable In Vitro Ocular Toxicity Studies
SOT 65th Annual Meeting and ToxExpo 2026
San Diego, California, United States
March 24, 2026Background and Purpose
The corneal epithelium is a transparent cell layer that covers the cornea and allows light to enter the eye. It serves as a critical barrier against pathogens and environmental insults, protecting deeper ocular tissues. Due to its protective and regenerative functions, the corneal epithelium is widely used in studies of wound healing, drug development, permeation, and toxicity. While animal models are commonly employed in ocular toxicity testing, they often fail to accurately reflect human responses. Human primary corneal epithelial cells offer a more relevant alternative, but their short lifespan and donor-dependent variability limit their suitability in long-term, reproducible studies. To address these limitations, we developed an immortalized human corneal epithelial cell line that retains key functional characteristics of primary cells while enabling extended culture and consistent performance in ocular toxicity assays.
Methods
We established a clonal immortalized human corneal epithelial cell line by stably expressing human telomerase reverse transcriptase (hTERT) and a mutant cyclin-dependent kinase 4 (CDK4R24C) in normal primary human corneal epithelial cells. We assessed cell morphology, proliferation rate, expression of epithelial markers, migratory capacity, and performance in in vitro eye irritation assays.
Results
The resulting cell line, hTERT HCEC (ATCC® CRL-4067™), was cultured for over 125 population doublings without signs of replicative senescence. It exhibited an abnormal female human karyotype and maintained epithelial-like morphology. Both primary cells and hTERT HCEC expressed cytokeratin 14 (KRT14) and demonstrated migratory ability. In irritation assays, treatment with glycolic acid or benzalkonium chloride led to a dose-dependent reduction in cell viability, confirming the cell line’s responsiveness to toxic agents.
Conclusions
The hTERT HCEC cell line successfully overcomes the limitations of primary human corneal epithelial cells while preserving essential functional attributes. This model provides a robust and reproducible platform for in vitro eye irritation testing and broader ocular toxicology applications.
Download the poster to learn about the hTERT-immortalized human corneal epithelial cell line.
DownloadPresenter
Xiangshan Zhao, PhD
Senior Scientist, Cell Biology R&D, ATCC
Xiangshan Zhao, PhD, is a Senior Scientist at ATCC. He has extensive experience in cell line development and cell-based assay development. Dr. Zhao leads the Immortalized Primary Cells program developing innovative immortalized cell lines for research, drug screening, cell therapy, and toxicology studies.
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