RESUMO
BACKGROUND:Tauroursodeoxycholic acid is a hydrophilic bile acid derivative that has neuroprotective effects in a variety of neurological disease models.However,there are few reports on the effects of tauroursodeoxycholic acid on spinal cord injury. OBJECTIVE:To investigate the effect of tauroursodeoxycholic acid on apoptosis of spinal cord neurons under hypoglycemic and hypoxic conditions,as well as the effect on recovery of motor function in mice after spinal cord injury. METHODS:(1)In vitro experiment:Primary spinal cord neurons were isolated from C57 BL/6 mouse embryos at 13.5 days of gestation.After 72 hours of culture,the cells were divided into three groups.In the normal group,cells were cultured in Neurobasal complete medium that was incubated in a CO2 incubator(5%CO2 + 95%air)for 24 hours.In the oxyglucose-deprived group,sugar-free Neurobasal medium was added and incubated in a triple-gas incubator(94%N2+5%CO2+1%O2)for 12 hours,and then the medium was replaced with Neurobasal complete medium and incubated in a CO2 incubator for 12 hours.In the experimental group,the treatment procedure was approximately the same as that in the oxyglucose-deprived group,except that taurodeoxycholic acid was added along with the sugar-free Neurobasal medium.TUNEL staining was used to detect apoptosis,cell counting kit-8 assay was applied to detect cell activity,and immunofluorescence staining was performed to detect cellular β-microtubule protein expression.(2)Animal experiment:Sixty C57 BL/6 mice were randomly divided into sham-operated group,spinal cord injury group and experimental group,with 20 mice in each group.Animal models of T9-T10 spinal cord injury were established using Allen's percussion method in the spinal cord injury group and the experimental group.Starting from the 1st day after modeling,taurodeoxycholic acid solution was given by gavage in the experimental group and normal saline was given by gavage in the sham-operated and spinal cord injury groups once a day for 14 consecutive days.Spinal cord tissue repair was assessed using behavioral and histological methods. RESULTS AND CONCLUSION:In vitro experiment:TUNEL staining,cell counting kit-8 and immunofluorescence staining showed that compared with the normal group,the number of apoptotic cells was higher(P＜0.01),while cell activity and β-microtubule protein expression were lower in the oxyglucose-deprived group(P＜0.01);compared with the oxyglucose-deprived group,the number of apoptotic cells was lower(P＜0.01),while cell activity and β-microtubule protein expression were higher in the experimental group(P＜0.01).Animal experiment:The Basso-Beattie-Bresnahan scores in the open field test and hind limb footprint experiments showed that the mice in the experimental group had better recovery of walking and motor functions than those in the spinal cord injury group.Hematoxylin-eosin staining showed that significant deformities and cavities were observed at the site of spinal cord injury and the number of nerve cells was significantly reduced in the spinal cord injury group.Compared with the spinal cord injury group,the experimental group showed a significant reduction in the area of spinal cord injury,less spinal cord deformity,fewer cavities,and an increase in the number of nerve cells.Immunofluorescence staining showed that the number of neuronal nucleus-labeled neuronal cells in the spinal cord injury group was less than that in the sham-operated group(P＜0.01),and the number of neuronal nucleus-labeled neuronal cells in the experimental group was higher than that in the spinal cord injury group(P＜0.01).To conclude,tauroursodeoxycholic acid could effectively reduce glucose/oxygen deprivation-induced apoptosis of spinal cord neurons and axonal loss,and promote the recovery of motor function in mice with spinal cord injury.