Anti-oxidative and anti-apoptosis effects of egg white peptide, Trp-Asn-Trp-Ala-Asp, against H2O2-induced oxidative stress in human embryonic kidney 293 cells

Peptides derived from egg white protein are well-known for their abundant species and biological properties. The aim of this study was to investigate the anti-oxidative and anti-apoptosis effects of (Trp-Asn-Trp- Ala-Asp) WNWAD, a pentapeptide derived from egg white ovomucin pepsin hydrolysates, against the oxidative stress induced by H2O2 in HEK-293 cells. Oxygen radical absorbance capacity (ORAC) results showed that WNWAD possessed extraordinary oxygen radical absorption capacity (with an ORAC value = 2.91 mu M TE mu M-1) in vitro. Then, at the cellular level, MTS assay results displayed that WNWAD dose-dependently inhibited H2O2-induced cellular oxidative stress in HEK-293 cells and fully recovered the oxidative damage induced by 400 mu M H2O2 at a concentration of 1 mu M. The 2', 7'-dichlorofluorescein diacetate (DCFH-DA) fluorescent probe assay further proved that WNWAD inhibited cellular oxidative stress by reducing intracellular ROS accumulation in intact HEK-293 cells. In addition, scanning electron microscopy (SEM) images showed that the morphology of HEK-293 cells under H2O2 treatment displayed a cell apoptosis phenotype, while WNWAD pretreatment prevented the development of this phenotype. Furthermore, Western blot results indicated that WNWAD up-regulated the level of the anti-apoptotic protein Bcl-2 and down-regulated the levels of apoptosis executor proteins, cleaved caspase-3 and cleaved PRAP, in H2O2-induced oxidative stress in HEK-293 cells. All the above results suggested that WNWAD protected HEK-293 cells against H2O2-induced oxidative stress by inhibiting intracellular ROS accumulation and blocking the ROS activated mitochondria-mediated cell apoptosis pathway.