Purpose <p>The corneal epithelium is a biologically active tissue that helps maintain corneal homeostasis through continuous interaction with the underlying stroma. While oxidative stress–related DNA damage has been implicated in corneal disease, direct experimental evidence of genotoxic alterations in corneal epithelial cells in keratoconus remains limited. We evaluated genotoxicity in human corneal epithelial cells associated with keratoconus using the micronucleus (MN) assay.</p> Methods <p>This prospective study included 31 eyes with progressive keratoconus undergoing corneal cross-linking (CXL) and 34 healthy control eyes undergoing photorefractive keratectomy (PRK). Corneal epithelial samples were obtained intraoperatively during CXL and PRK. Genotoxicity was assessed with the MN assay by scoring 1000 epithelial cells per sample; MN frequency was expressed per mille (‰). Associations with smoking exposure and contact lens use were analyzed (p &lt; 0.05).</p> Results <p>MN frequency was higher in the keratoconus group than in controls (5.45 ± 1.12‰ vs. 0.50 ± 0.71‰; p &lt; 0.001). In keratoconus, MN frequency correlated positively with smoking exposure (r = 0.628; p = 0.038). In controls, MN frequency correlated with contact lens use duration (r = 0.523; p = 0.009). MN frequency was not associated with keratoconus staging parameters.</p> Conclusions <p>Corneal epithelial cells in keratoconus show increased genotoxicity, supporting increased epithelial genomic instability in keratoconus.</p>