Disruption of the blood-spinal cord barrier (BSCB) results in secondary injury and apoptosis of neurons, leading to permanent neurological dysfunction after spinal cord injury. In this study, we evaluate the role of miRNA-125a-5p in the BSCB under hypoxia. The miRNA-125a-5p mimics group showed lower horseradish peroxidase (HRP) permeability and endothelial cell death rates compared with the transfection control group. By contrast, the miRNA-125a-5p inhibitor group demonstrated higher HRP permeability and endothelial cell death rates than the transfection control group. The expressions of ZO-1, occludin, VE-cadherin and their mRNA significantly increased in miRNA-125a-5p-overexpressing cells. By contrast, a remarkable reduction in ZO-1, occludin, and VE-cadherin expression and their mRNA were observed in miRNA-125a-5p-inhibited cells. MiRNA-125a-5p appears to reduce the permeability of the BSCB by up regulating the expression of ZO-1, occludin, and VE-cadherin and their mRNA, and against hypoxia-induced apoptosis of spinal cord microvascular endothelial cells. Taken together, our results clearly indicate that miRNA-125a-5p plays an important role in protecting the functions of the BSCB under hypoxia.