基于腺苷酸激活蛋白激酶-自噬信号通路探讨减重平板训练对脊髓损伤大鼠运动功能的影响OACSTPCD
Effect of Body Weight Supported Treadmill Training on Motor Function of Rats with Spinal Cord Injury Based on AMPK-Autophagy Signaling Pathway
目的 基于腺苷酸激活蛋白激酶(AMPK)-自噬信号通路探讨减重平板训练对脊髓损伤(SCI)大鼠运动功能的影响及其作用机制.方法 选择30只10周龄SPF级雌性SD大鼠,按照随机数字表法分为假手术组、模型组和运动组,每组10只.模型组和运动组均采用Allen's法制作T10不完全性脊髓损伤模型,假手术组仅进行假手术.3组均于SCI造模前3 d进行预适应运动训练,以适应跑台环境.运动组于造模后第8天进行减重平板训练,减重为大鼠体质量的20%~40%,跑台速度6 m/min,20 min/次,2次/d,持续训练3周;假手术组和模型组造模后均自由活动,不进行减重平板训练.分别在造模后第1、7、14、21、28天采用脊髓损伤行为学(BBB)评分法评估各组大鼠后肢运动功能;于干预结束后(SCI后第28天)采用步态分析处理系统评估大鼠的步态参数(足印面积、步幅长度、触地强度、摆动速度和运动速度);采用尼氏染色法观察脊髓神经元形态及数目;采用Western blot法检测脊髓组织p-AMPK、t-AMPK、LC3、p62蛋白表达水平.结果 ① BBB评分:与假手术组比较,模型组、运动组造模后第1、7、14、21、28天BBB评分明显更低,差异均具有统计学意义(P<0.05);与模型组同一时间点比较,运动组造模后14、21、28天BBB评分明显更高,差异均具有统计学意义(P<0.05);模型组、运动组内不同时间点BBB评分比较,差异均具有统计学意义(P<0.05).②步态参数:与假手术组比较,模型组足印面积、触地强度、摆动速度、步幅长度和运动速度均明显更低,运动组足印面积明显更低,差异均具有统计学意义(P<0.05);与模型组比较,运动组第28天足印面积、触地强度、摆动速度、步幅长度和运动速度均明显更高,差异均具有统计学意义(P<0.05).③ 脊髓前角神经元形态及数目:与假手术组比较,模型组脊髓前角神经元数目明显减少(P<0.05),可见细胞核固缩.与模型组比较,运动组脊髓前角可见较为清晰的尼氏体,细胞核固缩程度降低,脊髓前角神经元存活数目增加(P<0.05).④ p-AMPK、t-AMPK、LC3、p62蛋白表达水平:与假手术组比较,模型组LC3 Ⅱ/LC3 Ⅰ明显降低,p62明显升高,差异均具有统计学意义(P<0.05);与模型组比较,运动组LC3 Ⅱ/LC3 Ⅰ、p-AMPK/t-AMPK明显升高,p62明显降低,差异均具有统计学意义(P<0.05).结论 减重平板训练可改善SCI大鼠神经功能和后肢运动功能,这可能与激活脊髓AMPK-自噬信号通路有关.
Objective To explore the effect and mechanism of body weight supported treadmill training on motor function of rats with spinal cord injured(SCI)based on the adenosine monophosphate-activated protein kinase(AMPK)-autophagy signaling pathway.Methods A total of 30 10-week-old female SD rats were randomly divided into sham operation group,model group,and exercise group,with 10 cases in each group.Allen's method was used to establish T10 incomplete spinal cord injury models in the model group and the exercise group,while the sham operation group only underwent sham operation.All the three groups received preconditioning exercise training for 3 days before SCI modeling to adapt to the treadmill environment.From the 8th day after mod-eling,the exercise group received treadmill training with weight reduction of 20%-40%of the body mass,and the treadmill speed was set at 6 m a minute,20 minutes a time,twice a day for 3 weeks.The sham operation group and the model group were allowed to move freely without treadmill training after modeling.Basso-Beattie-Bresnahan(BBB)method was used to assess motor function of hind limbs of rats at the 1st,7th,14th,21st and 28th days after modeling.Gait analysis system was used to assess gait parameters(footprint area,stride length,touchdown intensity,swing speed and movement speed)after intervention(at the 28th day after SCI).Nissl staining was used to observe the morphology and count of spinal cord neurons.Western blot method was used to detect the pro-tein expression levels of p-AMPK,t-AMPK,LC3 and p62 in spinal cord tissue.Results(1)BBB score:compared with the sham operation group,the BBB score of the model group and the exercise group were significantly lower at the 1st,7th,14th,21st and 28th days after modeling(P<0.05).Compared with the model group at the same time,the BBB score of the exercise group was sig-nificantly higher at the 14th,21st and 28th days after modeling,and the differences were statistically significant(P<0.05).The BBB score in the model group and the exercise group were compared at different times,and the differences were statistically significant(P<0.05).(2)Gait parameters:compared with the sham operation group,the footprint area,touchdown intensity,swing speed,stride length and movement speed were significantly lower in the model group,and the footprint area was significantly lower of the exer-cise group(P<0.05);compared with the model group,the footprint area,touchdown intensity,swing speed,stride length and move-ment speed were significantly higher in the exercise group at the 28th day after modeling,and the differences were statistically sig-nificant(P<0.05).(3)Morphology and number of neurons in the anterior horn of the spinal cord:compared with the sham operation group,the number of neurons in the anterior horn of the spinal cord reduced significantly in the model group(P<0.05),and nuclear pyknosis was observed.Compared with the model group,Nissl bodies were more clearly observed in the exercise group,nuclear pyknosis reduced significantly,and the number of surviving neurons in the anterior horn of the spinal cord increased significantly(P<0.05).(4)Protein expression levels of p-AMPK,t-AMPK,LC3,and p62:compared with the sham operation group,LC3 Ⅱ/LC3 Ⅰdecreased significantly in the model group,and p62 increased significantly(P<0.05).Compared with the model group,LC3 Ⅱ/LC3 Ⅰ and p-AMPK/t-AMPK increased significantly in the exercise group,and p62 decreased significantly(P<0.05).Conclusion Body weight supported treadmill training can improve the neurological function and hindlimb motor function of rats with SCI,which may be related to the activation of the spinal AMPK-autophagy signaling pathway.
蔡吉利;翟晨元;王尊;伍琦;方露;王钰;王彤
南京医科大学康复医学院,江苏 南京 210029南京医科大学康复医学院,江苏 南京 210029||南京中医药大学针灸推拿学院·养生康复学院,江苏 南京 210023南华大学衡阳医学院附属第一医院,湖南 衡阳 421001南京医科大学康复医学院,江苏 南京 210029||南京医科大学第一附属医院,江苏 南京 210029南京医科大学第一附属医院,江苏 南京 210029
脊髓损伤减重平板训练腺苷酸激活蛋白激酶自噬运动功能神经功能
spinal cord injurybody weight supported treadmill trainingAMPKautophagymotor functionneurological func-tion
《康复学报》 2024 (004)
370-376 / 7
国家自然科学基金面上项目(82172541)
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