中国神经再生研究(英文版) ›› 2014, Vol. 9 ›› Issue (4): 413-419.doi: 10.4103/1673-5374.128248

• 原著:退行性病与再生 • 上一篇    下一篇

腓骨肌萎缩症2L型:揭示中国此病的转基因小鼠模型

  

  • 收稿日期:2014-01-20 出版日期:2014-02-25 发布日期:2014-02-25

A novel transgenic mouse model of Chinese Charcot-Marie-Tooth disease type 2L

Ruxu Zhang1, Fufeng Zhang2, Xiaobo Li1, Shunxiang Huang1, Xiaohong Zi1, Ting Liu1, Sanmei Liu1, Xuning Li1, Kun Xia3, Qian Pan3, Beisha Tang2, 3   

  1. 1 Department of Neurology, Third Xiangya Hospital, Central South University, Changsha, Hunan Province, China
    2 Department of Neurology, Xiangya Hospital, Central South University, Changsha, Hunan Province, China
    3 National Key Laboratory of Medical Genetics, Central South University, Changsha, Hunan Province, China
  • Received:2014-01-20 Online:2014-02-25 Published:2014-02-25
  • Contact: Ruxu Zhang, M.D., Department of Neurology, Third Xiangya Hospital, Central South University, Changsha 410013, Hunan Province, China, zhangruxu03@gmail.com.
  • Supported by:

    This project was funded by the National Natural Science Foundation of China, No. 81071001, 30900805.

摘要:

我们既往研究发现小热休克蛋白8的K141N突变与中国腓骨肌萎缩症2L型家系有关。为此,实验拟建立一个人小热休克蛋白8基因K141N突变的转基因小鼠模型,以确定该转基因小鼠模型是否可以用于腓骨肌萎缩症2L型的研究。将K141N突变的小热休克蛋白8的cDNA克隆到由人巨细胞病毒表达系统驱动的pCAGGS质粒中,转染受精卵中建立过表达K141NHSPB8的转基因小鼠。电镜观察显示,转基因小鼠坐骨神经髓纤维密度降低,且出现轴突水肿及空泡变性。PCR和Western bolt结果显示,141NHSPB8在转基因小鼠心脏和腓肠肌组织中整合并广泛表达。6个月时的行为评估显示K141NHSPB8转基因小鼠后肢肌肉力量下降,运动协调能力降低,但没有明显的感觉障碍。电生理分析表明,K141NHSPB8转基因小鼠6个月时坐骨神经的复合肌动作电位波幅显著下降,而运动神经传导速度未出现异常,表明其周围神经损伤主要发生在轴突。结果证实成功建立了K141NHSPB8转基因小鼠模型可以作为研究周围神经单基因遗传病腓骨肌萎缩症2L型发病机制的实验动物模型。

关键词: 神经再生, 周围神经损伤, 轴突损伤, 动物模型, 腓骨肌萎缩症2L型, 基因突变, 原核注射, 转基因模型, 小热休克蛋白B8, NSFC grant

Abstract:

We previously found that the K141N mutation in heat shock protein B8 (HSPB8) was responsible for Charcot-Marie-Tooth disease type 2L in a large Chinese family. The objective of the present study was to generate a transgenic mouse model bearing the K141N mutation in the human HSPB8 gene, and to determine whether this K141NHSPB8 transgenic mouse model would manifest the clinical phenotype of Charcot-Marie-Tooth disease type 2L, and consequently be suitable for use in studies of disease pathogenesis. Transgenic mice overexpressing K141NHSPB8 were generated using K141N mutant HSPB8 cDNA cloned into a pCAGGS plasmid driven by a human cytomegalovirus expression system. PCR and western blot analysis confirmed integration of the K141NHSPB8 gene and widespread expression in tissues of the transgenic mice. The K141NHSPB8 transgenic mice exhibited decreased muscle strength in the hind limbs and impaired motor coordination, but no obvious sensory disturbance at 6 months of age by behavioral assessment. Electrophysiological analysis showed that the compound motor action potential amplitude in the sciatic nerve was significantly decreased, but motor nerve conduction velocity remained normal at 6 months of age. Pathological analysis of the sciatic nerve showed reduced myelinated fiber density, notable axonal edema and vacuolar degeneration in K141NHSPB8 transgenic mice, suggesting axonal involvement in the peripheral nerve damage in these animals. These findings indicate that the K141NHSPB8 transgenic mouse successfully models Charcot-Marie-Tooth disease type 2L and can be used to study the pathogenesis of the disease.

Key words: nerve regeneration, peripheral nerve injury, axonal injury, animal models, Charcot-Marie-Tooth disease type 2L, gene mutation, pronuclear injection, transgenic model, small heat shock protein B8, NSFC grant, neural regeneration