小剂量超短波对大鼠脊髓损伤后早期A1型星形胶质细胞的影响

Effects of low dose ultrashort wave on early A1 astrocytes after spinal cord injury

目的:研究超短波治疗对脊髓损伤(spinal cord injury,SCI)后早期A1型星形胶质细胞分化的影响。方法:将45只SD大鼠随机分成3组:假手术组,仅暴露T10脊髓并不打击,其他操作同其他实验组;对照组,暴露胸10(T10)脊髓后给予Allen打击制备SCI模型,并不给予任何治疗;干预组,给予Allen法打击致SCI,并在损伤后24h开始给予小剂量超短波干预,每日1次,每周5次,每次7min,直至取材前。大鼠的运动功能用BBB评分及大鼠脊髓损伤联合行为评价方法(combined behavioral evaluation of spinal cord injury,CBS)评分进行评定,SCI大鼠取材后分别对组织进行纵向切片行免疫荧光染色,检测C3/GFAP、IBA-1/P65在组织内的表达部位和表达量。结果:①BBB评分显示,SCI后大鼠运动功能逐渐改善,7d时治疗组BBB评分相比对照组明显改善(P<0.05);SCI后5d、7d时,干预组CBS评分低于对照组,这说明干预组大鼠的功能恢复强于对照组(P<0.05,P<0.01);②SCI后1d、3d、7d小胶质数量逐渐增多,SCI后3d、7d,干预组IBA-1数量明显低于对照组(P<0.01)。③SCI后1d、3d、7d随着时间的推移,表达P65蛋白的细胞逐渐增多。SCI后3d,与对照组相比,干预组SCI大鼠的受损脊髓组织IBA-1/P65共染阳性细胞数量显著降低(P<0.01)。此外,干预组P65阳性细胞入核数明显低于对照组(P<0.01)。④SCI后A1型促炎性星形胶质细胞逐渐出现,相比SCI后1d、7d,SCI后3d大鼠受损脊髓当中的A1型星型胶质细胞(C3/GFAP阳性细胞之比)达峰值(P<0.01)。SCI后3d、7d时,干预组A1型星形胶质细胞数量均明显低于对照组(P<0.01)。结论:SCI后早期小剂量USW治疗可以抑制损伤周围小胶质细胞数量及炎症因子释放,进而抑制A1型星形胶质细胞的形成,促进运动功能恢复,这一现象与NF-κB通路相关。

Objective:To investigate the effect of ultrashort wave therapy on the differentiation of astrocytes with spinal cord injury(SCI)in the early stage.Method:Totally 45 SD rats were randomly divided into three groups:Sham group,only exposed the T10 spinal cord and did not strike;Control group,after exposing chest 10(T10)spinal cord,spinal cord contusion was inflicted using Allen's method without any treatment;Intervention group,spinal cord contusion was inflicted using Allen's method,a low dose ultra-short wave intervention was given 24h after injury,7min per time,once a day,5 times a week until sacrificed.The motor function of rats was evaluated by Basso Beattie Bresnahan score(BBB score)and combined behavioral evaluation of spinal cord injury(CBS).After sacrificed,spinal cord was cut longitudinally on a frozen biopsy machine for immunofluorescence staining to detect the expression sites and levels of C3/GFAP and IBA-1/P65 in the tissues.Result:BBB score showed that motor function of rats was gradually improved after SCI,and BBB score in the treatment group was significantly improved compared with the control group on day 7(P<0.05).At 5d and 7d after SCI,the CBS score of the intervention group was lower than that of the control group,indicating that the functional recovery of the rats in the intervention group was stronger than that in the control group(P<0.05,P<0.01).The quantity of microglia gradually increased on 1d,3d and 7d after SCI,and the quantity of IBA-1 in the intervention group was significantly lower than that in the control group(P<0.01).As time passed after SCI,the number of cells expressing P65 protein increased gradually.The number of IBA-1/P65 costaining positive cells in SCI rats in the USW intervention group was significantly lower than that in the control group(P<0.01)in rats of 3d after SCI.At the same time,the number of P65 positive cells in the intervention group was significantly lower than that in the control group(P<0.01).After SCI,A1 astrocytes reached the peak on day 3(P<0.01)and decreased some on day 7.The number of A1 astrocytes in the intervention group were significantly lower than that in the control group(P<0.01)on day 3 and day 7.Conclusion:Early low-dose USW therapy after SCI can inhibit the number of microglia and the release of inflammatory factors,thus inhibiting the formation of A1 astrocytes and promoting the recovery of motor function,which is related to the NF-B pathway.