Nuclear factor erythroid 2-related factor 2 (NFE2L2) is a critical transcription factor in maintaining cellular redox balance and metabolism, which can induce an adaptive response against oxidative stress (OS). Tea polyphenols (TP) is a potent antioxidant that exerts protective effects on cells by scavenging free radicals and increasing the levels of NFE2L2 and phase II detoxifying enzymes. Thus, we used NFE2L2 short interfering siRNA (siRNA) to downregulate NFE2L2 expression in cultured bovine mammary epithelial cells (BMEC) and to investigate the cytoprotective effects of TP in BMEC exposed to hydrogen peroxide- (H₂O₂-) induced OS. The content of reactive oxygen species (ROS) and malondialdehyde (MDA), activities of superoxide dismutase (SOD), glutathione peroxidase (GPX), catalase (CAT), glutathione-S-transferase (GST) were determined to identify the oxidative damage of BMEC. All statistical tests were carried out using SAS 9.0. To establish an OS, isolated BMEC were exposed to increasing concentrations of H₂O₂ (0, 100, 200, 400, 600, 800, 1,000 μM) for 0, 2, 4, 6, 8, 12, and 24 h. Optimal doses of TP (0, 60, 80, 100 μg/mL) were evaluated by pretreatment of BMEC for 0, 2, 4, 6, 8, 12, and 24 h, followed by a H₂O₂ (600 µM) challenge for 6 h. BMEC were transfected with NFE2L2-siRNA, pretreated with 100 µg/mL TP for 12 h, followed by 600 μM H₂O₂ challenge for 6 h. The results revealed that treatment with 600 μM H₂O₂ for 6 h induced oxidative damage of BMEC, indicating this model could be used to establish an OS injury model. A concentration of TP of 100 μg/mL during 12 h incubation increased cell viability, protein and mRNA abundance of NFE2L2, and decreased intracellular ROS accumulation after H₂O₂ exposure. In addition, transfection with NFE2L2-siRNA confirmed that mRNA abundance of NFE2L2 and heme oxygenase-1 (HMOX-1) in response to TP was decreased, while ROS production was increased with H₂O₂ (600 μM). Overall, TP had beneficial effects on BMEC redox balance, TP can slow down cellular OS-related injury, decrease ROS production, and increase the mRNA abundance of NFE2L2 and HMOX-1 by activating the NFE2L2/HMOX1 pathway. TP may potentially serve as an antioxidant against OS in dairy cows.