Exploring the structural properties, osteoarthritis-relieving effects and potential mechanisms of peptides from eggshell membrane byproduct

Food protein-derived bioactive peptides hold great potential to improve osteoarthritis (OA). Eggshell membrane is an abundant protein-rich byproduct of egg processing. Our previous study revealed that the eggshell membrane peptides (EMP) exhibited cytoprotection against oxidative stress. However, the structural properties, protective effects against OA in vivo and molecular mechanism of EMP have not been elucidated. This study aimed to explore these issues by combining molecular simulation and in vivo studies. The results showed that EMP comprised novel high-abundance peptides, predominantly VEDIRIRPIL, DKDGPFRL and VFFDMFNAL, enriched with antioxidant and anti-inflammatory amino acids at the N- and C- terminals. These peptides possessed high binding affinities with Keap1 at the Nrf2-binding site mainly by hydrogen bonds. In a papaininduced OA rat model, EMP significantly prevented OA cartilage destruction by regulating collagen II and MMP-3 expression, and inhibited inflammation and oxidative stress by decreasing the level of inflammatory mediators, NO free radical production, and increasing the antioxidant enzyme SOD activity. Corresponding with molecular docking, EMP activated the Keap1-Nrf2 pathway by regulating Keap1 and Nrf2 expression in OA cartilages, which may relate to the inhibition effect on the Keap1-Nrf2 interaction by EMP. This study provides novel insights into the structural basis, protective effects, and potential molecular mechanism of EMP against OA, as well as lays a theoretical basis for the value-added utilization of egg byproduct proteins in functional products to alleviate OA.