大鼠骨骼肌急性损伤修复过程中电针联合跑台训练对神经肌肉接头相关蛋白nestin、Cdk5和ε-nAChR的影响研究

发布时间：2018-03-07 01:19

  本文选题：nestin　切入点：Cdk5　出处：《重庆医科大学》2017年硕士论文　论文类型：学位论文

【摘要】：目的:通过观察电针与跑台训练联合作用于大鼠急性骨骼肌损伤修复过程中对nestin、Cdk5和ε-n ACh R表达的影响,探讨电针联合跑台训练对于骨骼肌损伤及促进神经肌肉接头修复的可能机制。方法:将60只成年SD雄性大鼠,设置为正常组和干预组和自然恢复组,采用随机数字表法随机抽取6只作为正常组,剩余54只大鼠作为自然恢复组和干预组。通过自制打击器制备右下肢的腓肠肌急性钝挫伤模型。将随机抽取的48只建模成功大鼠按照随机数字表法分为2组(每组n=24):自然恢复组和干预组。干预组在造模成功48h后,按照跑台计划训练表干预,跑台结束后即刻选取阿是穴、足三里穴位进行电针干预,每日一次,连续干预5天后休息2天。每组分别于造模后7天、14天、21天、28天四个时间点分别随机选取6只大鼠进行检测。采用HE染色观察损伤局部组织形态变化;采用Western blot、免疫荧光和RT-PCR观察nestin、Cdk5以及ε-n ACh R蛋白及m RNA的表达水平。结果:1、HE染色显示:正常组肌纤维形态完整,肌细胞大小规则,排列整齐;自然恢复组、干预组与正常组相比,均出现不同程度的肌纤维断裂、变性,坏死;干预组与自然恢复组相比,肌细胞结构损伤后恢复更迅速,肌组织内的瘀血、出血灶和水肿减轻更快,新生肌细胞更多,整体肌肉组织形态恢复更好。2、免疫荧光显示:Nestin与Cdk5免疫荧光双标记染色:正常组中,nestin与Cdk5无共表达;在自然恢复组和干预组中,nestin与Cdk5荧光共表达逐渐降低,干预组中共表达低于自然恢复组。3、Western blot检测结果:自然恢复组和干预组中,nestin蛋白在7天,14天,21天表达量均高于正常组(P0.05),且干预组表达量高于自然恢复组(P0.05),28天各组相比无明显差异(P0.05);Cdk5蛋白在7天,14天,21天表达量均比正常组高(P0.05),干预组明显低于自然恢复组(P0.05),在28天各组无明显差异(P0.05);ε-n ACh R蛋白表达逐渐升高,各个时间点干预组表达量高于自然恢复组(P0.05),7天、14天、21天干预组和自然恢复组中表达量均低于正常组(P0.05),28天干预组表达量高于自然恢复组(P0.05),与正常组表达量无明显差异(P0.05)。4、RT-PCR检测结果:自然恢复组和干预组中,nestin m RNA在7天,14天均高于正常组(P0.05),且呈现递减趋势,在21天,28天干预组和自然恢复组中nestin m RNA表达量与正常组相比无统计学意义(P0.05);Cdk5 m RNA在7天,14天,21天干预组与自然恢复组表达量均比正常组高(P0.05),各时间点的干预组明显低于自然恢复组(P0.05),28天各组中Cdk5m RNA表达量与正常组相比无统计学意义(P0.05);ε-ACh R m RNA的表达在四个时间点逐渐升高,7天、14天、21天干预组和自然恢复组中表达量均低于正常组(P0.05),干预组表达量高于自然恢复组(P0.05),28天干预组表达量高于自然恢复组(P0.05),与正常组表达量无统计学差异(P0.05)。结论:大鼠骨骼肌急性损伤后,电针联合运动训练在一定程度上促进神经肌肉接头的修复,其作用机制可能是通过抑制nestin和Cdk5蛋白在肌细胞膜上的共表达,促进了ε-n ACh R与nestin的表达,从而促进神经肌接头的再生。
[Abstract]:Objective: To observe the effects of electroacupuncture combined with the treadmill training in repairing rat acute skeletal muscle injury of nestin, expression of Cdk5 and R ACh e -n, electroacupuncture combined with treadmill training on skeletal muscle injury and promote the mechanism of neuromuscular junction repair. Methods: 60 adult male SD rats that is set to normal group and intervention group and spontaneous recovery group were randomly selected 6 rats as normal group, the remaining 54 rats as the natural recovery group and intervention group. The acute gastrocnemius blunt contusion model was prepared to attack right lower limb system. 48 rats were randomly selected were randomly divided into 2 groups (n=24 each): natural recovery group and intervention group. The intervention group in rats after 48h, in accordance with the treadmill treadmill training schedule intervention plan, immediately after the selection of Ashi point, Zusanli acupoint for acupuncture The intervention, once a day, continuous intervention after 5 days of rest for 2 days. Each group after modeling for 7 days, 14 days, 21 days, 28 days four time points were randomly selected 6 rats were detected. HE staining was used to observe the morphological changes of local tissue injury; by Western blot, immunofluorescence and RT-PCR. Nestin, Cdk5 and the expression level of R protein and m ACh e -n RNA. Results: 1. HE staining: normal group muscle fiber morphology, muscle cell size rule, neatly arranged; natural recovery group, the intervention group compared with normal group, there were different degrees of muscle fiber fracture, degeneration, necrosis; intervention group and natural recovery group, injury of muscle cell structure recovered more quickly, muscle tissue congestion, hemorrhage and edema in neonatal myocytes faster, more whole muscle morphology recovery better.2, immunofluorescence showed that Nestin and Cdk5 double immunofluorescence staining: normal 缁勪腑,nestin涓嶤dk5鏃犲叡琛ㄨ揪;鍦ㄨ嚜鐒舵仮澶嶇粍鍜屽共棰勭粍涓，

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