一个快速的表型神经评分系统在ALS的SOD1-G93A小鼠模型评估疾病进展
Summary
This video protocol describes a sensitive, reliable, and quick method for evaluating the neuromuscular deficits in a transgenic mouse model of amyotrophic lateral sclerosis.
Abstract
The SOD1-G93A transgenic mouse is the most widely used animal model of amyotrophic lateral sclerosis (ALS). At ALS TDI we developed a phenotypic screening protocol, demonstrated in video herein, which reliably assesses the neuromuscular function of SOD1-G93A mice in a quick manner. This protocol encompasses a simple neurological scoring system (NeuroScore) designed to assess hindlimb function. NeuroScore is focused on hindlimb function because hindlimb deficits are the earliest reported neurological sign of disease in SOD1-G93A mice. The protocol developed by ALS TDI provides an unbiased assessment of onset of paresis (slight or partial paralysis), progression and severity of paralysis and it is sensitive enough to identify drug-induced changes in disease progression. In this report, the combination of a detailed manuscript with video minimizes scoring ambiguities and inter-experimenter variability thus allowing for the protocol to be adopted by other laboratories and enabling comparisons between studies taking place at different settings. We believe that this video protocol can serve as an excellent training tool for present and future ALS researchers.
Introduction
自从它在90年代中期开发的SOD1-G93A转基因小鼠模型已经肌萎缩侧索硬化症（ALS）1的最广泛使用的动物模型。这种转基因小鼠模型被基因工程化以过表达人铜/锌超氧化物歧化酶1（SOD1）基因携带有ALS相关甘氨酸至丙氨酸的突变在氨基酸93（G93A）的突变体形式。的G93A突变是许多突 变的SOD1基因，它们共同化妆大约五分之一的家族性ALS例2之一。
在SOD1-G93A模式已经成为了ALS药物开发研究的一个主力，因为除了要ALS其明显的遗传链接，这概括了许多ALS在人体内的病理特点，如运动神经元丧失，肌肉逐渐萎缩无力，与最终麻痹而死亡3。虽然，这是常有用的转基因动物模型的情况下，有一种内在的双向与SOD1-G93A小鼠相关ological变化。 Scott等。已经确定，需要在药物功效的研究设计，以克服由生物变异4中创建的噪声至少24垫料匹配性别平衡小鼠的队列。大量在这些研究中与侵袭性疾病进展和需要日常监测组合需要的动物的禁止使用的精心设计的，耗时的和潜在的压力的技术，用于测量神经肌肉强度和功能（例如，肌电图，握力，旋转棒等）。相反，它强调需要一个体内筛选工具，可靠地评估了一个快速的方式SOD1-G93A小鼠的神经肌肉功能。
在ALS TDI的协议被开发，允许SOD1-G93A小鼠的神经肌肉功能的每只小鼠不到30秒，平均可靠的评估。该协议encompa小规模企业后肢功能的使用是该报告在媒体和视频描述了一个简单的神经评分系统（NeuroScore）每天评估。 NeuroScore专注于后肢功能，因为后肢赤字是最早报道的疾病在SOD1-G93A小鼠5,6神经的迹象。此外，通过ALS TDI开发的协议提供了发病轻瘫（轻微或局部麻痹），进展和麻痹的严重程度的公正评估，这是不够敏感，以确定在疾病进展的药物引起的变化。
Protocol
Representative Results
Discussion
在这份报告中，我们描述了一个快速和简单的视频协议，如果应用得当，能够可靠地评估SOD1-G93A小鼠的疾病进展和确定药物引起的变化。尽管不同群体开发的表型的评分系统对SOD1-G93A小鼠8-10，他们往往不提供关于该过程足够的细节和不足进行复制。其结果，评分系统很少使用，开发他们的特定的组之外。在这份报告中，用视频详细手稿组合最大程度地减少评分含糊和跨实验者变异从而允?…
Disclosures
The authors have nothing to disclose.
Acknowledgements
We would like to acknowledge Beth Levine for critically reviewing the manuscript, Valerie Tassinari for developing the genotyping protocol, and Matt Ferola and Carlos Maya for their exceptional animal care.
Materials
| B6SJLTgN(SOD1G93A)1Gur strain | The Jackson Laboratory, Bar Harbor, ME | 002726 | Strain of mice used in this study |
| Breeders | Biomedical Research Models, Inc., Worcester, MA | Colony maintainance | |
| Scale | Navigator OHAUS, Parsippany, NJ | Model N14120 | Used to weigh mice |
| Nutra-GelH | Bio-Serv, Flemington, NJ | #S4798 | Wet food provided to sick mice |
| Irradiated Lab Animal Diet | Harlan, Indianapolis, IN | 2918-111914M | Chow diet |
| Lab-grade Sani-chips | Harlan Teklad, Indianapolis, IN | 7090 | Bedding |
| JMPH | SAS Institute, Inc., SAS Campus Drive, Cary, NC | v10.0.2 | Statistical software |
| Microsoft Excel | Microsoft, One Microsoft Way, Redmond, WA | Excel 2013 (v 15.0) | Spreadsheet software |
| GraphPad Prism 5 | GraphPad software Inc., La Jolla, CA | v5.4 | Graphing software |
| Mouse Huts | Bio-Serv, Flemington, NJ | K3272 | For environmental enrichment |
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