Abstract
The aim of the present study was to observe the effect of zhenbao pill on the motor function of acute spinal cord injury (ASCI) rats and the molecular mechanisms involving miR-146a-5p and G-protein-coupled receptor 17 (GPR17). ASCI rat model was established by modified Allen method, and then the rats were divided into three groups. SH-SY5Y cells were cultured overnight in hypoxia condition and transfected with miR-146a-5p mimic or miR-146a-5p inhibitor. The hind limb motor function of the rats was evaluated by Basso, Beattie, Bresnahan (BBB) scoring system. Quantitative real-time PCR (qRT-PCR) and Western blot were used to detect the expression of miR-146a-5p, GPR17, inducible nitric oxide synthase (iNOS), interleukin 1β (IL-1β), and tumor necrosis factor α (TNF-α). Neuronal apoptosis was measured using flow cytometry assay. Luciferase reporter assay was performed to determine the regulation of miR-146a-5p on GPR17. Zhenbao pill could enhance hind limb motor function and attenuate the inflammatory response caused by ASCI. Moreover, zhenbao pill increased the level of miR-146a-5p and decreased GPR17 expression in vivo and in vitro. Bioinformatics software predicted that GPR17 3′-UTR had a binding site with miR-146a-5p. Luciferase reporter assay showed that miR-146a-5p had a negative regulatory effect on GPR17 expression. Knockdown of miR-146a-5p could reverse the effect of zhenbao pill on the up-regulation of GPR17 induced by hypoxia, reversed the inhibitory effect of zhenbao pill on the cell apoptosis induced by hypoxia and the recovery of zhenbao pill on hind limb motor function in ASCI rats. Zhenbao pill could inhibit neuronal apoptosis by regulating miR-146a-5p/GPR17 expression, and then promoting the recovery of spinal cord function.
Highlights
Acute spinal cord injury (ASCI) is a high disability of the central nervous system injury caused by trauma
Normal rats have a BBB score of 21, while ASCI rats with completely paralyzed hind limbs have a BBB score of 0
Results showed that the inhibitory effect of zhenbao pill on neurones’ apoptosis was not obvious at 0.2 g/kg dosing conditions, but the inhibitory effect increased with increasing dosage (Figure 1A)
Summary
Acute spinal cord injury (ASCI) is a high disability of the central nervous system injury caused by trauma. ASCI is mainly caused by various accidents, sports, natural disasters, falls, and violence, and its incidence has been high. The incidence of ASCI in developed countries is 236–1009 per million people [1]. In China, annual incidence (6.7 cases per million people) is lower than the developed countries. It is of great significance to strengthen the study of spinal cord injury (SCI). SCI can be divided into primary injury and secondary injury [2]. Primary injury mainly refers to the destruction of the spinal cord structure caused by external forces. The study of ASCI treatment and rehabilitation is focussed on secondary injury [5]
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