Fluorometholone inhibits corneal epithelial proliferation, migration via targeting Rho GTPases: RhoA, Rac1, and Cdc42

Abstract

Abnormal corneal epithelial hyperplasia is a common complication following refractive surgery. 0.1 % fluorometholone (FML) eye drops are commonly used for treatment. However, their efficacy varies among patients, potentially attributed to differences in the patient's microenvironment. The underlying reason remains incompletely understood. This study aimed to elucidate the molecular mechanisms of FML's action on corneal epithelial cells (CECs). The effects of FML on the cell viability, proliferation, cell cycle, and migration of human corneal epithelial cells (HCECs) were evaluated using MTS assay, EdU staining, flow cytometry, and scratch assay, respectively. Mouse corneal sections were immunofluorescently stained to assess cell proliferation. A corneal wound model, monitored by slit-lamp photography, was utilized to evaluate the impact of FML on wound healing. Gene expression alterations were detected via RNA sequencing. RT-qPCR and Western blot were employed to validate gene and protein expression in HCECs and mouse corneal epithelia. Proteomic analysis was conducted on tear samples from patients. FML treatment significantly inhibited CEC proliferation, migration, and wound healing. At the molecular level, FML treatment led to a remarkable downregulation of RhoA, Rac1, and Cdc42. Correspondingly, reductions in the downstream Erk and NF-κB signaling pathways were observed in both HCECs and mouse corneal epithelia. Moreover, these pathways were similarly downregulated in tear samples from clinical patients. In conclusion, FML inhibits CEC proliferation and migration by modulating the Rho GTPase signaling network, especially through RhoA/Rac1/Cdc42, thereby suppressing the Erk/NF-κB pathway.