Mechanism of microRNA-31-5p regulating protein kinase Cε pathway in protection of rabbit spinal cord ischemia-reperfusion injury by remote ischemic preconditioning

  Objective  To investigate the role and mechanism of microRNA-31-5p (miR-31-5p) in the protection of rabbit spinal cord ischemia-reperfusion injury (SCIRI) by remote ischemic preconditioning (RIPC).

  Methods  A total of 60 Japanese white rabbits were divided into sham group, ischemia/reperfusion (I/R) group, RIPC group, RIPC+miR-NC group and RIPC+miR-31-5p mimic group, with 12 rabbits in each group. SCIRI model was established by clamping the abdominal aorta for 30 minutes, and the miR-31-5p overexpression model was constructed by intrathecal injection of miR-31-5p mimic plasmid. Except for the sham group and the I/R group, RIPC was performed in other groups one hour before the abdominal aorta closed. After 48 hours of reperfusion, the animals were evaluated for hindlimb motor function score and blood-spinal cord barrier (BSCB) permeability; Hematoxylin-Eosin (HE) staining was used to observe the morphological changes of spinal cord tissue; TUNEL was used to detect neuronal apoptosis in spinal cord tissue; reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) was used to detect the mRNA expression of miR-31-5p, protein kinase Cε (PKCε) and brain-derived neurotrophic factor (BDNF) in spinal cord tissue; Western Blot was used to detect the expression of PKCε and BDNF protein in spinal cord tissue.

  Results  Compared with the sham group, the hindlimb motor function score of the I/R group was significantly reduced, and the Evans blue (EB) content and neuron apoptosis rate were significantly increased (P < 0.05), and the number of spinal cord neurons was reduced, vacuolation was obvious, and nucleus pyknosis was observed; compared with the I/R group, the hindlimb motor function score, PKCε and BDNF mRNA and protein expression of RIPC group were significantly increased, and the EB content, neuronal apoptosis rate and miR-31-5p expression were significantly decreased (P < 0.05), the normal neurons increased significantly, a few cells had vacuolar degeneration, and nucleoli were obvious; upregulation of miR-31-5p expression with miR-31-5p mimic significantly weakened the protective effect of RIPC on rabbit SCIRI(P < 0.05).

  Conclusion  The expression of miR-31-5p decreases after RIPC, and it can increase the expression of BDNF in the spinal cord and relieve SCIRI by activating the PKCε protein.