Cryoinjury, or injury caused by extremely low temperatures, can occur in corneal endothelial cells (CECs) and lead to visual impairment. However, the mechanism of cryoinjury in CECs is not clear. The Stk11‑p53 signaling pathway regulates the proliferation and division of cells. Activity of the Stk11‑p53 signaling pathway arrests the cell cycle at the G0/G1 phase and induces apoptosis. In this study, a mouse model of cryoinjury in CECs was used. Following injury, significant mouse CEC death and shedding were observed. In addition, the mRNA and protein levels of core factors from the Stk11‑p53 signaling pathway (Stk11, p21 and p53) were elevated and Caspase‑3 was activated following cryoinjury. In addition, chromatin immunoprecipitation revealed that Stk11 catalyzed p53 serine 15 phosphorylation, and the Stk11‑p53 complex bound to the p21 promoter and stimulated gene transcription. Thus, the results of the present study suggest that cryoinjury leads to the damage and apoptosis of mouse CECs by activation of the Stk11‑p53 signaling pathway, phosphorylation of p53 serine 15 and p21 gene transcription.