Abstract

A novel antioxidant peptide AFYRW was previously isolated and identified from tartary buckwheat albumin in our laboratory. Human immortalized keratinocytes (HaCaT) are the main epidermal cells, and play a critical role in maintaining skin barrier function and mediating inflammatory and immune responses. This study explored the protective effect of AFYRW against oxidative damage in HaCaT cells and clarified the underlying molecular mechanism related to the SIRT1 pathway. HaCaT cells were divided into control, H₂O₂ model, and low/high-dose AFYRW intervention groups. A series of cytological and molecular biological assays were performed to detect cell viability, ROS levels, migration, apoptosis, and the expression of pathway-related proteins and genes. The results showed that AFYRW had no cytotoxicity, and dose-dependently reversed H₂O₂-induced cell viability decline, ROS overproduction, migration inhibition, inflammation and apoptosis. Mechanistically, AFYRW restored intracellular NAD⁺ content and enhanced SIRT1 activity, thereby reducing the acetylation levels of NF-κB p65 (Lys310) and p53 (Lys381). Furthermore, the SIRT1-specific inhibitor EX527 abolished the above protective effects of AFYRW. In conclusion, AFYRW alleviates H₂O₂-induced oxidative damage in HaCaT cells via activating SIRT1-mediated deacetylation, providing a theoretical basis for its application in skin-protective functional foods and cosmetics.