PURPOSE: Mechanical stress on the ocular surface, such as from eye-rubbing, has been reported to lead to inflammation and various ocular conditions. We hypothesized that the mechanosensitive Piezo1 channel in the conjunctival epithelium contributes to the inflammatory response at the ocular surface after receiving mechanical stimuli. METHODS: Human conjunctival epithelial cells (HConjECs) were treated with Yoda1, a Piezo1-specific agonist, and various allergens to measure cytokine expression levels using qRT-PCR. Piezo1 activation-induced intracellular signaling pathways were also investigated by Western blot. Mechanical stretching experiments were conducted to simulate Piezo1 activation in HConjECs. Specificity of Piezo1 was confirmed by PIEZO1 knockdown and GsMTx4. In in vivo studies, using immunohistochemistry, rats were administered Yoda1 eye drops to examine the inflammatory response in the conjunctiva and Piezo1-induced signaling activation. RESULTS: HConjECs expressed functional Piezo1 channel which was the dominant mechanoreceptor among putative channels and whose activation significantly increased IL-6 and IL-8 expression through the p38 MAPK-CREB pathway. Piezo1-induced [Ca CONCLUSION: Mechanical stress-induced Piezo1 channel activation in conjunctival epithelial cells can cause ocular inflammation by upregulating pro-inflammatory cytokines via the p38 MAPK-CREB pathway and promoting neutrophil infiltration. These findings suggest that mechanical stimuli on ocular surface tissues are significant risk factors for ocular inflammation.