Grub polypeptide extracts protect against oxidative stress through the NRF2-ARE signaling pathway

Grub polypeptide extracts (GPEs) have antioxidant effects; however, their underlying molecular mechanisms are unknown. This study explored the antioxidant molecular mechanism of GPE via the nuclear factor-erythroid 2-related factor 2 (NRF2)-antioxidant response element (ARE) signaling pathway in C2C12 muscle satellite cells exposed to oxidative stress. The effects of GPE/or H2O2 on C2C12 were investigated by the MTT (3- (4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) viability assay and immunofluorescence and small interfering RNA (siRNA) analyses. The cell viability, cell damage, intracellular reactive oxygen species (ROS) levels, and NRF2 signaling pathways related to proteins were measured. GPE significantly increased the antioxidant capacity of cells, evident by increased cell viability and decreased lactate dehydrogenase leakage, DNA damage, malondialdehyde content, and ROS level. GPE also markedly increased mRNA expression levels and activities of antioxidant enzymes including superoxidase 1 and 2, catalase, and glutathione peroxidase. In addition, GPE increased the gene and protein expression of NRF2 and heme oxygenase 1 by promoting NRF2 translocation from the cytoplasm to the nucleus and activating NRF2-ARE signaling pathways. The antioxidant effects of GPE through these signaling pathways were further confirmed by NRF2-specific siRNA silencing. Thus, GPE enhances antioxidant capacity and alleviates oxidative damage of C2C12 cells via the NRF2-ARE signaling pathway.