BNIP3-mediated mitophagy alleviates traumatic brain injury by inhibiting neuronal apoptosis and oxidative stress

Traumatic brain injury (TBI) significantly influences neurological outcomes, and its pathogenesis is primarily attributed to neuronal apoptosis and oxidative stress. However, the precise role of Bcl2/adenovirus E1B protein-interacting protein 3 (BNIP3)-mediated mitophagy in this process remains poorly understood. Therefore, this study was designed to elucidate the function of BNIP3-dependent mitophagy in neuronal fate following TBI. Furthermore, an in vitro TBI model was established by promoting oxygen-glucose deprivation/reperfusion in HT22 neuronal cells. The BNIP3 expression in TBI model cells was systematically manipulated via siRNA-mediated knockdown and plasmid-based overexpression. Moreover, mitophagy activity was assessed by monitoring key mitochondria-associated proteins, including mitochondrial adaptor protein P62, microtubule-associated protein 1 light chain 3B (LC3B), the translocase of outer mitochondrial membrane 20 (TOMM20), and cytochrome c oxidase IV (COX IV). Immunofluorescence microscopy and transmission electron microscopy were carried out to analyze mitochondrial-lysosomal colocalization and autophagosome formation, respectively. Similarly, neuronal apoptosis was quantified via TUNEL assay, while oxidative stress was evaluated by measuring superoxide dismutase (SOD) activity and malondialdehyde (MDA) levels to elucidate redox imbalance. The results indicated that TBI substantially upregulated BNIP3 expression and enhanced mitophagy in HT22 cells. Furthermore, it increased neuronal apoptosis and MDA levels, but reduced SOD activity. BNIP3 knockdown significantly attenuated TBI-induced mitophagy while exacerbating apoptotic cell death and oxidative stress. Whereas BNIP3 overexpression elevated mitophagy and conferred robust neuroprotection against TBI-induced neuronal apoptosis and oxidative damage. In conclusion, these in vitro findings indicated that BNIP3-mediated mitophagy is a crucial endogenous neuroprotective mechanism in neurons, suggesting its potential in promoting neuronal survival following TBI, which warrants further in vivo validation.