Objective:To investigate the effect of hypoxia-inducible factor 1α (HIF-1α)/Bcl-2 and adenovirus E1B19kDa-interacting protein 3 (BNIP3)-mediated mitophagy on neuronal apoptosis after traumatic brain injury (TBI).Methods:HT22 cells (mouse hippocampal neurons) were chosen; oxygen-glucose deprivation/re-oxygenation (OGD/R) was used to establish in vitro TBI models. Expressions of HIF-1α and BNIP3 were regulated by HIF-1α, BNIP3-specific small interfering RNA (siRNA) or plasmid vector transfection. Experiment 1 was performed to investigate the effect of BNIP3-mediated mitophagy on neuronal apoptosis after TBI; HT22 cells were divided into 4 groups ( n=3): siRNA control group (normal culture after negative siRNA transfection), siRNA+TBI group (OGD/R after negative siRNA transfection), BNIP3-siRNA group (normal culture after BNIP3-siRNA transfection), and BNIP3-siRNA+TBI group (OGD/R after BNIP3-siRNA transfection); the expressions of mitochondrial autophagy related proteins such as HIF-1α, BNIP3, LC3-II, and P62 were detected by Western blotting, mitochondrial autophagosomes were observed by transmission electron microscopy, apoptosis was detected by TUNEL, and lactate dehydrogenase (LDH) activity in cell supernatant was determined by LDH kit. Experiment 2 was performed to investigate the effect of HIF-1α on BNIP3-mediated mitophagy and neuronal apoptosis after TBI; HT22 cells were divided into 8 groups ( n=3): siRNA control group (normal culture after negative siRNA transfection), siRNA+TBI group (OGD/R after negative siRNA transfection), HIF-1α-siRNA group (normal culture after HIF-1α-siRNA transfection), HIF-1α-siRNA+TBI group (OGD/R after HIF-1α-siRNA transfection), empty plasmid group (normal culture after pcDNA3.1[+] transfection), HIF-1α overexpression group (normal culture after HIF-1α plasmid transfection), empty plasmid+TBI group (OGD/R after empty plasmid transfection), HIF-1α overexpression+TBI group (OGD/R after HIF-1α plasmid transfection); the expressions of HIF-1α, BNIP3, LC3-II, P62, TOMM20 and COX IV, apoptosis rate and LDH activity in neurons of each group were determined. Results:(1) In Experiment 1, compared with siRNA control group, siRNA+TBI group had significantly increased BNIP3 expression and LC3-II/I ratio, significantly decreased P62, TOMM20 and COX IV expressions, and statistically increased apoptosis and LDH activity ( P<0.05); compared with siRNA+TBI group, BNIP3-siRNA+TBI group had significantly decreased BNIP3 expression and LC3-II/I ratio, significantly increased P62, TOMM20, and COX IV expressions, and significantly increased apoptosis and LDH activity ( P<0.05); under projective electron microscope, siRNA+TBI group had increased autophagosomes compared with siRNA control group, while BNIP3-siRNA+TBI group had decreased autophagosomes compared with siRNA+TBI group. (2) In Experiment 2, compared with siRNA+TBI group, HIF-1α-siRNA+TBI group had significantly decreased expressions of HIF-1α and BNIP3, and LC3-II/I ratio, significantly increased expressions of P62, TOMM20 and COX IV, and significantly increased apoptosis and LDH activity ( P<0.05); compared with empty plasmid+TBI group, HIF-1α overexpression+TBI group had statistically higher expressions of HIF-1α and BNIP3, and LC3-II/I ratio, significantly decreased expressions of P62, TOMM20 and COX IV, and significantly decreased apoptosis and LDH activity ( P<0.05). Conclusion:HIF-1α can mitigate TBI-induced neuronal apoptosis via promoting BNIP3-mediated mitophagy.