6PPD induces apoptosis and autophagy in SH-SY5Y cells via ROS-mediated PI3K/AKT/mTOR pathway: In vitro and in silico approaches

N-(1,3-dimethylbutyl)-N’-phenyl-p-phenylenediamine (6PPD), an extensively employed antioxidant in rubber materials, is considered as an emerging contaminant. 6PPD was proven to have potential neurotoxicity, which poses risks to human health. However, the research on its neurotoxicity is still limited. This work explored the neurotoxicity of 6PPD to SH-SY5Y cells and in-depth mechanisms with a combination of in vitro and in silico approaches. Our results indicated that 6PPD could reduce cell viability and cause oxidative damage by increasing reactive oxygen species (ROS) accumulation and altering the levels of glutathione (GSH), catalase (CAT), superoxide dismutase (SOD), and malondialdehyde (MDA). 6PPD induced neuronal apoptosis of mitochondrial pathway and autophagy dysfunction, as characterized by the increased expressions of Cleaved caspase-3, Bax/Bcl-2, Beclin-1, LC3-II/I, and P62. 6PPD downregulated the expression of PI3K, p-AKT, and p-mTOR, while the PI3K inhibitor suppressed PI3K/AKT/mTOR pathway and promoted both apoptosis and autophagy, indicating that PI3K/AKT/mTOR pathway was involved in 6PPD-induced apoptosis and autophagy. The inhibition of this pathway was attributed to ROS accumulation in SH-SY5Y cells. Molecular docking analysis further revealed that 6PPD exhibits strong binding affinity to PI3K, AKT, and mTOR protein molecules, which could effectively interfere with downstream signaling pathways. These findings enrich the understanding of 6PPD-induced neurotoxicity and contribute to the evaluation of ecological risks associated with 6PPD.