Objective To investigate the protective effect and mechanism of astragaloside IV (AST4) on H2O2-induced oxidative stress injury and apoptosis of SY5Y cells. Methods Human SY5Y cells were cultured in vitro and induced by H2O2 to establish oxidative stress model, which was divided into PBS group, H2O2 group and AST4 group. Cell viability was determined by MTT assay. Cell apoptosis was detected by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling assay (TUNEL). The supernatant was used to determine the activity of malondialdehyde (MAD), superoxide dismutase (SOD) and glutathione (GSH) in each group. Immunofluorescence cytochemistry was used to detect the nuclear factor E2-related factor (Nrf-2) and cleaved caspase-3 (c-caspase-3). B-lymphoblastoma-2 (Bcl2), Bcl2-associated X protein (BAX), c-caspase-3, Nrf-2 in cells and nuclei and heme oxygenase-1 (HO-1) were determined by Western blot analysis. Results AST4 had a protective effect on viability of SY5Y cells under oxidative stress damage, reduced the content of MAD, and increased the content of GSH and SOD. AST4 increased Bcl2 and decreased BAX, thus Bc12/BAX ratio was significantly increased compared with that in H2O2 group. Meanwhile, AST4 inhibited the expression of c-caspase-3. AST4 promoted nuclear translocation of Nrf-2 and increased the expression of the downstream antioxidant protein HO-1. Conclusion AST4 can promote Nrf-2 nuclear translocation, increase HO-1 expression, regulate oxidation/antioxidant balance, improve antioxidant level, protect cells from oxidative damage and reduce apoptosis by activating Nrf-2/HO-1 signaling pathway.