靶肌肉注射胶质细胞源性神经营养因子对面神经损伤修复的作用[*]
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西南医科大学附属医院耳鼻咽喉头颈外科

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泸州市科技局研究计划项目(2021-SYF-30)


Effects of target intramuscular injection of glial cell-derivedneurotrophic factor on the repair of crushed facial
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Department of Otorhinolaryngology Head and Neck Surgery,The Affiliated Hospital,Southwest Medical University

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    目的:通过靶肌肉注射胶质细胞源性神经营养因子(GDNF),观察其对面神经压榨损伤后大鼠功能恢复、神经形态学及GDNF在中枢面神经元中表达的影响,探讨经靶肌肉注射GDNF治疗周围性面瘫的可行性及作用机理。方法:SD大鼠随机分为假手术组(只暴露右侧面神经主干)、模型组(面神经主干压榨)、实验对照组(面神经主干压榨+颊肌注射生理盐水)和实验组(面神经主干压榨+颊肌注射GDNF),通过颊肌电生理、面瘫症状评分观察大鼠的神经功能恢复, Masson染色观察颊肌纤维形态学变化、甲苯胺蓝染色观察面神经形态学变化、Western Blot检测面神经元中GDNF的表达。结果:①大鼠面瘫症状评分:假手术组无面瘫表现,评分为0分,造模后各组大鼠均出现周围性面瘫表现,评分均为5分;术后28天,实验组大鼠面瘫症状已基本完全恢复,模型组及实验对照组较前有不同程度改善但未完全恢复,评分均大于3分;②颊肌电生理:各组造模后与假手术组相比,峰潜伏期有不同程度延长、最大振幅有不同程度下降;随时间的推移,实验组峰潜伏期较模型组及实验对照组明显缩短(P<0.05),最大振幅较其明显上升(P<0.05);③甲苯胺蓝染色:模型组、实验对照组、实验组术后均出现神经纤维形态不规则,外膜不连续不清晰,轴突数量减少。术后28天,实验组面神经形态与假手术无明显差异,较模型组及实验对照组有明显恢复;④Masson染色:各组造模后肌纤维数量减少,肌肉组织占比面积减少;实验组肌纤维形态较模型组及实验对照组恢复快,至术后28天,肌纤维形态接近正常(P<0.05);⑤Western Bolt检测:各组造模后中枢面神经元组织GDNF的蛋白表达下降,观察期内逐渐增强;与模型组及实验对照组相比,术后各时间点,实验组中枢面神经元组织GDNF的蛋白表达显著增强(P<0.01)。结论:GDNF可作为改善周围神经损伤的有效给药方式之一,促进神经纤维修复、靶肌肉功能的恢复及中枢神经系统对周围损伤神经的修复。

    Abstract:

    AIM: Glial cell-derived neurotrophic factor (GDNF) was injected into the target muscle to observe its effects on functional recovery, neuromorphology and GDNF expression in central facial neurons in rats after facial nerve crush injury, so as to investigate the feasibility and mechanism of action of GDNF injection via the target muscle in the treatment of peripheral facial paralysis. METHODS: SD rats were randomly divided into sham-operated group (right facial nerve trunk exposed only), model group (facial nerve trunk pressed), experimental control group (facial nerve trunk pressed + buccal muscle injected with saline) and experimental group (facial nerve trunk pressed + buccal muscle injected with GDNF). The neurological recovery of rats was observed by buccal muscle electrophysiology, animal facial palsy score, morphological changes of buccal muscle fibres by Masson staining, morphological changes of facial nerve by toluidine blue staining and GDNF expression in facial neurons by Western Blot. RESULTS: ① The rats in the experimental group had basically recovered from facial palsy 28 days after surgery, while the rats in the model and control groups had improved to different degrees but not fully recovered from facial palsy, with a score of more than 3; ② Buccal muscle electrophysiology: the peak latency was prolonged to different degrees and the maximum amplitude decreased to different degrees in each group compared with the sham-operated group; over time, the peak latency was significantly shorter (P<0.05) and the maximum amplitude increased (P<0.05) in the experimental group compared with the model group and the experimental control group; ③Toluidine blue staining: the model group, the experimental control group and the experimental group all showed irregular nerve fiber morphology, discontinuous and unclear outer membrane and reduced number of axons after surgery. 28 days after surgery, the facial nerve morphology of the experimental group was not significantly different from that of the sham operation, and there was significant recovery compared with the model group and the experimental control group; ④Masson staining: the number of muscle fibers decreased and the area occupied by muscle tissue decreased in each group; the morphology of muscle fibers in the experimental group recovered faster than that of the model group and the experimental control group, and by 28 days postoperatively, the morphology of muscle fibers was close to normal (P<0.05); ⑤Western Bolt assay: the protein expression of central facial neurons decreased in each group after modelling. The protein expression of GDNF in the central facial neuron tissues decreased in each group after modelling and gradually increased during the observation period; compared with the model group and the experimental control group, the protein expression of GDNF in the central facial neuron tissues was significantly enhanced in the experimental group at each time point after surgery (P<0.01). CONCLUSION: GDNF can be used as one of the effective modes of administration to improve peripheral nerve injury, promoting nerve fibre repair, recovery of target muscle function and repair of peripherally injured nerves by the central nervous system.

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  • 收稿日期:2023-01-06
  • 最后修改日期:2023-03-24
  • 录用日期:2023-03-30
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