Experimental study of the mechanism of induction of conjunctival goblet cell hyperexpression using CHIR-99021 in vitro

A component of the tear film, mucin is produced by conjunctival goblet cells and is crucial to preserving the tear film's stability. Severe thermal burns, chemical burns, and severe ocular surface diseases can cause extensive damage to the conjunctiva, destroy the secretory function of goblet cells, and affect the stability of the tear film and integrity of the ocular surface. Currently, the expansion efficiency of goblet cells in vitro is low. In this study, we observed that rabbit conjunctival epithelial cells exhibited dense colony morphology after stimulation with the Wnt/fi-catenin signaling pathway activator CHIR-99021 and promoted the differentiation of conjunctival goblet cells and the expression of its specific marker Muc5ac, among which the best induction effect was observed after 72 h in vitro culture with 5 mu mol/L CHIR-99021. Under optimal culture conditions, CHIR-99021 increased the expression levels of the Wnt/fi-catenin signaling pathway factors Frzb, fi-catenin, SAM pointed domain containing ETS transcription factor, and glycogen synthase kinase-3fi and the levels of the Notch signaling pathway factors Notch1 and Kruppel-like factor 4 while decreasing the expression levels of Jagged-1 and Hes1. The expression level of ABCG2, a marker of epithelial stem cells, was raised to keep rabbit conjunctival epithelial cells from self -renewing. Our study showed that CHIR-99021 stimulation successfully activated the Wnt/fi-catenin signaling pathway and conjunctival goblet cell differentiation was stimulated, in which the Notch signaling pathway played a combined role. Those results provide a novel idea for the expansion of goblet cells in vitro.(c) 2023 Elsevier Inc. All rights reserved.