Effect of Diquafosol on Hyperosmotic Stress-Induced Tumor Necrosis Factor-α and Interleukin-6 Expression in Human Corneal Epithelial Cells
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Purpose: Diquafosol is a pharmaceutical drug used for dry eye treatment with a novel mechanism of action. It is a purinergic P2Y2 receptor agonist that promotes the secretion of tears and healing of corneal epithelialwounds. However, its inhibitory effect on hyperosmotic stress-induced inflammation in human corneal epithelialcells (HCECs) remains unclear. Methods: A hyperosmotic stress model was established by transferring HCECs from isosmotic (312 mOsm/kg tohyperosmotic medium (500 mOsm/kg). HCECs were incubated with 500 mOsm/kg hyperosmotic medium for30 minutes, and then treated with diquafosol (0.6–6 mg/mL) for 4 or 24 hours. Cells were then harvested andanalyzed by western blot, immunocytochemistry, and real-time polymerase chain reaction to evaluate the expressionof interleukin-6, tumor necrosis factor-alpha, and the phosphorylation status of nuclear factor-kappa B. Results: Diquafosol significantly decreased the mRNA and protein expression of hyperosmotic stress-inducedtumor necrosis factor-alpha and interleukin-6. These results were supported by immunofluorescence stainingand quantitative real-time polymerase chain reaction analysis. Furthermore, diquafosol inhibits nuclear factor-kappa B activation by suppressing the phosphorylation and degradation of the inhibitor of кB. Conclusions: This study shows that diquafosol inhibits nuclear factor-kappa B signaling and inflammatory factorsinduced by hyperosmotic stress in HCECs. This suggests that using diquafosol for the improvement of dryeye syndrome could be effective in the treatment of inflammation-related corneal and conjunctival diseases.
Abstract
Purpose: Diquafosol is a pharmaceutical drug used for dry eye treatment with a novel mechanism of action. It is a purinergic P2Y2 receptor agonist that promotes the secretion of tears and healing of corneal epithelialwounds. However, its inhibitory effect on hyperosmotic stress-induced inflammation in human corneal epithelialcells (HCECs) remains unclear. Methods: A hyperosmotic stress model was established by transferring HCECs from isosmotic (312 mOsm/kg tohyperosmotic medium (500 mOsm/kg). HCECs were incubated with 500 mOsm/kg hyperosmotic medium for30 minutes, and then treated with diquafosol (0.6–6 mg/mL) for 4 or 24 hours. Cells were then harvested andanalyzed by western blot, immunocytochemistry, and real-time polymerase chain reaction to evaluate the expressionof interleukin-6, tumor necrosis factor-alpha, and the phosphorylation status of nuclear factor-kappa B. Results: Diquafosol significantly decreased the mRNA and protein expression of hyperosmotic stress-inducedtumor necrosis factor-alpha and interleukin-6. These results were supported by immunofluorescence stainingand quantitative real-time polymerase chain reaction analysis. Furthermore, diquafosol inhibits nuclear factor-kappa B activation by suppressing the phosphorylation and degradation of the inhibitor of кB. Conclusions: This study shows that diquafosol inhibits nuclear factor-kappa B signaling and inflammatory factorsinduced by hyperosmotic stress in HCECs. This suggests that using diquafosol for the improvement of dryeye syndrome could be effective in the treatment of inflammation-related corneal and conjunctival diseases.
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