Abstract: Objective To explore the mechanism by which berberine inhibits ferroptosis of mouse hippocampal neuronal cells (HT22). Methods Cultured HT22 cells were pretreated with 30 or 60 μmol/L berberine for 2 h before exposure to 0.5 μmol/L erastin for 8 h, and the cell proliferation, intracellular ferric iron level, changes in intracellular reactive oxygen species (ROS) and cell apoptosis were detected using CCK-8, Fe2 + fluorescent probe, fluorescent dye (DAPI) and fluorescent probe (H2DCFH-DA). RT-qPCR and Western blotting were used to detect the mRNA and protein expressions of Nrf2, HO-1 and GPX4 in the cells. We further tested the effects of treatments with 2 μmol/L ML385 (a Nrf2 inhibitor), 60 μmol/L berberine and erastin in the cells to explore the protective mechanism of berberine against erastin-induced ferroptosis in the neuronal cells. Results Treatment with 0.5 μmol/L erastin significantly lowered the viability of HT22 cells (P<0.05) and increased the production of ROS, cell apoptosis rate and ferric iron level (P<0.05). Pretreatment with 30 and 60 μmol/L berberine both significantly increased the vitality of erastin-exposed cells (P<0.05) and lowered the levels of intracellular ROS and ferric iron content (P<0.05). RT-qPCR and Western blotting showed that berberine obviously promoted the expressions of Nrf2, HO-1 and GPX4 in the cells (P<0.05), and treatment with ML385 significantly inhibited the Nrf2-HO-1/GPX4 pathway, increased intracellular ROS and ferric iron contents and mitigated the protective effect of berberine against erastin-induced ferroptosis (P<0.05). Conclusion Berberine can inhibit erastin-induced ferroptosis in HT22 cells possibly by activating the Nrf2-HO-1/GPX4 pathway.

Key words: berberine; erastin; HT22 cells; ferroptosis; neuroprotection; Nrf2-HO-1/GPX4