Cell culture and treatment

The neuronal cell line SH-SY5Y (Pricella, China) was cultured in MEM (Gibco, USA). Primary neuronal cells were obtained as previously described (Geng et al. 2019). The ventral mesencephalon was dissected from E12.5 C57BL/6 embryos in cold PBS, and DMEM containing 0.25% trypsin was added to obtain a single cell suspension after adding DMEM/F12 (Gibco. USA) supplemented with 10% FBS. The cells were seeded on 96‐well plates coated with poly-D-lysine and cultured in B27-supplemented neurobasal medium (Gibco, USA).

When the cell growth density was approximately 80%, oe-BDNF, si-STAT3, and si-NC (GenePharma, China) were transfected using a Lipofectamine 2000 kit (Solarbio, China) into neuronal cells. The cells were cultured at 37 °C in a 5% CO₂ cell incubator. After 24 h, the transfection efficiency was detected. Cells were treated with 1 mmol/L MPP⁺ for 24 h to establish a PD cell model. The NC group was cultured in DMEM substrate without drugs, and the experimental groups were cultured in DMEM substrate containing 5 mmol/L 3-MA (3-Methyladenine, PI3K inhibitor), 2 mmol/L Stattic (STAT3 inhibitor) or 5 mmol/L Recilisib (PI3K/Akt/mTOR agonist).