本研究得到了国家自然科学基金第82102314号（至ZSJ）和32170977（至HSL）和广东省自然科学基金（编号：2022A1515010438（至ZSJ）和2022A1515012306（至HSL）的支持。本研究由暨南大学附属第一医院临床前沿技术项目支持，编号：JNU1AF-CFTP-2022-a01206（至HSL）。本研究得到了广州市科技计划项目 第202201020018号（至HSL）、2023A04J1284（至ZSJ）和2023A03J1024（至HSL）的支持。

涉及动物的研究由暨南大学伦理委员会审查和批准。
1.动物麻醉和T10椎板切除术
<ol><li>使用 8 周龄的雌性年轻成年 C57/6J 小鼠进行本研究。通过腹膜内注射氯胺酮（100mg / kg）和地西泮（5mg / kg）麻醉小鼠。检查麻醉是否成功，表现为疼痛反射丧失。在眼睛上涂抹兽医软膏，以防止麻醉下干燥。</li>
	<li>用剃须刀剃掉老鼠背部的毛发，露出皮肤。用三轮碘磷和酒精交替消毒皮肤。</li>
	<li>用手术刀在背侧皮肤上做 2.5 厘米的内侧纵向切口，并使用镊子在 T9-T11 水平暴露脊柱。</li>
	<li>使用脊柱固定器双侧固定 T10 小关节。确保脊柱稳定固定。确保剥离椎旁肌肉，并使用微磨钻去除棘突和椎板，以暴露 T10 节段的脊髓。</li>
</ol>2. 使用 G smart SCI 系统对 T10 脊髓进行挫伤
<ol><li>打开开关，等待设备自动恢复到原始状态。将脊柱固定器放入 G smart SCI 系统中，并用螺钉固定。</li>
	<li>使用操作触摸屏（图2A），设置损伤参数，包括冲击速度（1 m / s），冲击深度（三组不同的小鼠为0.5 mm，0.8 mm和1.1 mm）和停留时间（500 ms）15。</li>
	<li>通过移动平台将激光测距仪对准暴露的脊髓中心。（图2B）</li>
	<li>单击触摸屏上的 Ready 按钮（图 2C）。冲击头将根据设置参数自动调整到特定高度。托架台自动将脊髓撞击部位移动到撞击头下方。</li>
	<li>手动按压冲击头以进一步确定冲击部位。单击 “开始 ”按钮，冲击头将根据设置的参数撞击脊髓。</li>
	<li>将小鼠从设备中取出并在体视显微镜（20x）下观察以确定脊髓损伤（图3）。为了确定模型开发是否成功，请观察局部充血、塌陷和脊髓膜破裂。</li>
	<li>使用 3-0 缝合线逐层缝合肌肉、筋膜和皮肤。将小鼠放入温暖的盒子中，等待它们恢复。</li>
</ol>3. 术后护理
<ol><li>手术后每天皮下注射美洛昔康（5mg/kg），持续 7 天。每8小时手动排空膀胱，直到膀胱功能恢复。</li>
	<li>手术后 14 天，移除缝合线。</li>
</ol>4. 脊髓损伤的检测效果
<ol><li>从术后第一天开始计算小鼠的BMS评分16,17。</li>
	<li>术后第30天，进行动物行为实验，包括走秀、足部断层和旋转16,17。走秀：创纪录的45厘米距离;最大运行持续时间 8 秒;相机增益：28.02;强度阈值 0.01。脚故障：为每只鼠标记录 60 步。旋转杆：速度 20 rpm。记录鼠标下落的时间，并记录为 120 秒以上 120 秒。</li>
	<li>术后第31天，腹膜内注射氯胺酮（100mg/kg）和地西泮（5mg/kg）麻醉小鼠，然后用4%PFA灌注对小鼠实施安乐死。小心地取出脊髓，并在受伤部位上方和下方截取 5 毫米以进行石蜡包埋。在小鼠脊髓损伤中心做一个5μm的切片，并进行苏木精和伊红染色17。</li>
	<li>对于统计分析，请使用商业软件。将数据表示为均值±均值标准误差 （SEM），并使用单因素方差分析进行比较;p < 0.05 被认为是显著的。</li>
</ol>

用于精确小鼠模型的脊髓损伤挫伤装置

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作者声明没有相互竞争的经济利益。

脊髓损伤可导致感觉和运动缺陷，从而导致严重的身体和精神损伤。在中国，不同省份的脊髓损伤发病率从14.6/100万到60.6不等18。脊髓损伤患病率的增加将给医疗保健系统带来更大的压力。目前，脊髓损伤的有效治疗选择有限，因为其病理机制和修复过程尚未完全了解19。有必要创建准确且可重复的脊髓损伤动物模型，以研究脊髓损伤的病理机制和修复过程。为...

脊髓损伤通常会导致损伤节段以下的永久性脊髓功能障碍。它多是由物体撞击脊柱和脊柱过度伸展引起的，例如交通事故和跌倒1.由于脊髓损伤的有效治疗方案有限，使用动物模型阐明脊髓损伤的发病机制将有助于制定适当的治疗方法。由脊髓撞击引起的脊髓损伤挫伤模型导致开发了与大多数临床脊髓损伤病例具有相似病理的动物模型 2,3。因此，为脊髓损伤挫伤生成准确、可重复和方便的动物模型非常重要。
自 Allen 于 1911 年发明第一个脊髓损伤动物模型以来，用于建立脊髓损伤动物模型的仪器的发展取得了重大进展 4,5。根据损伤机制，脊髓损伤模型分为挫伤、压迫、牵引、脱位、横断或化学6。其中，利用外力移位和损伤脊髓的挫伤模型最接近大多数脊髓损伤患者的临床病因。因此，挫伤模型已被许多研究人员用于脊髓损伤研究 3,7。各种仪器用于开发脊髓损伤挫伤模型。纽约大学 （NYU） 多中心动物脊髓损伤研究 （MASCIS） 撞击器通过减重装置产生脊髓损伤挫伤8.经过几个更新版本后，MASCIS撞击器被广泛用于开发脊髓损伤挫伤动物模型9。然而，当MASCIS的冲击杆掉落并撞击脊髓时，可能会发生多处损伤，这会影响脊髓损伤模型的损伤程度。此外，实现机械精度以确保仪器的精度和制造模型的可重复性也具有挑战性。无限地平线撞击器通过控制施加在脊髓上的力而不是重跌落来引起挫伤10.它使用连接到传感器的计算机直接测量撞击器和脊髓之间的冲击力。当达到阈值时，冲击器立即缩回，从而避免重量反弹并提高精度10,11。然而，使用这种精细运动方式造成损伤会导致不一致的损伤和功能缺陷6.俄亥俄州立大学（OSU）设备通过电磁驱动器12,13以瞬态速率压缩脊髓的背面。该装置类似于无限地平线撞击器，因为它使用短距离按压来引起脊髓损伤。然而，它有各种局限性，因为零点的初始确定会由于脑脊液的存在而引起误差6,14。综上所述，可用于开发脊髓损伤挫伤动物模型的仪器很多，但它们都存在一些局限性，导致动物模型的准确性和可重复性不足。因此，为了更准确、更方便、更可重复地创建脊髓损伤小鼠挫伤模型，需要一种自动化、智能化的脊髓损伤撞击器。
我们提出了一种新型的脊髓损伤冲击器，广州暨南大学智能脊髓损伤系统（G smart SCI system; 图1），用于生成脊髓损伤挫伤模型。该设备使用激光测距仪作为定位装置，结合自动化移动平台，根据设定的打击参数（包括打击速度、打击深度和停留时间）自动进行打击。自动化操作减少了人为因素，提高了动物模型的准确性和可重复性。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>0.01M PBS (powder, pH7.2-7.4)</td>
			<td>Solarbio Life Sciences</td>
			<td>P1010</td>
			<td></td>
		</tr>
		<tr>
			<td>2,2,2-Tribromoethanol</td>
			<td>Macklin</td>
			<td>75-80-9</td>
			<td></td>
		</tr>
		<tr>
			<td>4% paraformaldehyde tissue fixative</td>
			<td>Biosharp life science</td>
			<td>BL539A</td>
			<td></td>
		</tr>
		<tr>
			<td>Biomicroscope</td>
			<td>Leica</td>
			<td>LCC50 HD</td>
			<td></td>
		</tr>
		<tr>
			<td>CatWalk&#160;</td>
			<td>Noldus Information Technology</td>
			<td>CatWalk XT 9.1</td>
			<td></td>
		</tr>
		<tr>
			<td>Cover glass</td>
			<td>CITOTEST Scientific</td>
			<td>10212432C</td>
			<td></td>
		</tr>
		<tr>
			<td>Embedding machine</td>
			<td>Changzhou Zhongwei Electronic Instrument</td>
			<td>BMJ-A</td>
			<td></td>
		</tr>
		<tr>
			<td>Ethanol absolute</td>
			<td>DAMAO</td>
			<td>64-17-5</td>
			<td></td>
		</tr>
		<tr>
			<td>FootFaultScan</td>
			<td>Clever Sys Inc.</td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>Glass slide</td>
			<td>CITOTEST Scientific</td>
			<td>80302-2104</td>
			<td></td>
		</tr>
		<tr>
			<td>Hematoxylin and Eosin Staining Kit</td>
			<td>Beyotime Biotechnology</td>
			<td>C0105S</td>
			<td></td>
		</tr>
		<tr>
			<td>micro-grinding drill&#160;</td>
			<td>FEIYUBIO</td>
			<td>19-7010</td>
			<td></td>
		</tr>
		<tr>
			<td>Mouse spinal fixator</td>
			<td>RWD Life Science</td>
			<td>68094</td>
			<td></td>
		</tr>
		<tr>
			<td>Paraffin microtome</td>
			<td>Thermo</td>
			<td>shandon finesse 325</td>
			<td></td>
		</tr>
		<tr>
			<td>RotaRod for Mice</td>
			<td>Ugo Basile</td>
			<td>47600</td>
			<td></td>
		</tr>
		<tr>
			<td>Stereomicroscope</td>
			<td>KUY NICE</td>
			<td>SZM-7045</td>
			<td></td>
		</tr>
		<tr>
			<td>Tert-Amyl alcohol</td>
			<td>Macklin</td>
			<td>75-85-4</td>
			<td></td>
		</tr>
		<tr>
			<td>Xylene</td>
			<td>China National Pharmaceutical</td>
			<td>#10023418</td>
			<td></td>
		</tr>
	</tbody>
</table>

automated impactor for contusive spinal cord injury model in mice

由车祸和跌倒等创伤引起的脊髓损伤 （SCI） 与永久性脊髓功能障碍有关。通过撞击脊髓来创建脊髓损伤的挫伤模型会导致与临床实践中大多数脊髓损伤相似的病理。准确、可重复和方便的脊髓损伤动物模型对于研究脊髓损伤至关重要。我们提出了一种新型的小鼠自动脊髓损伤挫伤装置，即广州暨南大学智能脊髓损伤系统，该系统可以生成具有准确性、可重复性和便利性的脊髓损伤挫伤模型。该系统 通过 激光距离传感器与自动化移动平台和高级软件相结合，精确地生成不同程度的脊髓损伤模型。我们使用该系统创建了三个级别的脊髓损伤小鼠模型，确定了它们的 Basso 小鼠量表 （BMS） 评分，并进行了行为和染色测定以证明其准确性和可重复性。我们展示了使用该设备开发损伤模型的每个步骤，形成了一个标准化的程序。该方法可产生可重复的脊髓损伤挫伤小鼠模型，并通过方便的处理程序减少人为操作因素。开发的动物模型对于研究脊髓损伤机制和相关治疗方法是可靠的。

Spinal cord injury usually results in permanent spinal cord dysfunction below the injured segment. It is mostly caused by objects striking the spine and hyperextension of the spine, such as traffic accidents and falls1. Due to the limited availability of effective treatment options for spinal cord injury, elucidation of the pathogenesis of spinal cord injuries using animal models will be informative for the development of appropriate treatment approaches. The contusion model of spinal cord injury caused by impact on the spinal cord results in the development of animal models with similar pathologies to most clinical spinal cord injury cases2,3. Therefore, it is important to produce accurate, reproducible, and convenient animal models for spinal cord injury contusion.
Since Allen&#39;s invention of the first animal model of spinal cord injury in 1911, there have been major advances in the development of instruments for establishing spinal cord injury animal models4,5. Based on injury mechanisms, spinal cord injury models are classified as contusion, compression, distraction, dislocation, transection, or chemical6. Among them, the contusion models, which use external forces to displace and injure the spinal cord, are closest to the clinical etiology of most spinal cord injury patients. Therefore, the contusion model has been used by many investigators in spinal cord injury studies3,7. Various instruments are used to develop spinal cord injury contusion models. The New York University (NYU)-multicenter animal spinal cord injury studies (MASCIS) impactor produces spinal cord injury contusions by weight-drop device8. After several updated versions, the MASCIS impactor is widely used to develop spinal cord injury contusion animal models9. However, when the impact rod of MASCIS falls and hits the spinal cord, multiple injuries may occur, which affects the degree of injury in spinal cord injury models. Moreover, achieving mechanical precision to ensure the accuracy of the instrument and the repeatability of the manufacturing model is also challenging. The infinite horizon impactors cause contusions by controlling the force applied to the spinal cord rather than heavy drops10. It uses a computer connected to a sensor to directly measure the impact force between the impactor and the spinal cord. When the threshold is reached, the impactor is immediately retracted, thereby avoiding weight rebound and improving accuracy10,11. However, the use of this fine motor modality to inflict damage can result in inconsistent damage and functional deficits6. The Ohio State University (OSU) device compresses the dorsal surface of the spinal cord at a transient rate by an electromagnetic driver12,13. This device is similar to the infinite horizon impactors, as it uses short-distance compressions to cause spinal cord injuries. However, it has various limitations in that initial determination of the zero point will cause errors due to the presence of the cerebrospinal fluid6,14. In summary, there are many instruments that can be used to develop spinal cord injury contusion animal models, but they all have some limitations that lead to insufficient accuracy and reproducibility of animal models. Therefore, in order to more accurately, conveniently and reproducibly create mouse contusion models of spinal cord injury, an automated and intelligent spinal cord injury impactor is needed.
We present a novel spinal cord injury impactor, Guangzhou Jinan University smart spinal cord injury system (G smart SCI system; Figure 1), for producing spinal cord injury contusion models. The device uses a laser rangefinder as a positioning device, combined with an automated mobile platform to automate strikes according to set strike parameters, including strike speed, strike depth and dwell time. Automated operation reduces human factors and improves the accuracy as well as reproducibility of animal models.

作者聚焦：深入了解脊髓损伤研究的创新

用于小鼠挫伤脊髓损伤模型的自动撞击器

My research scope includes the pathogenesis and the treatment methods of spinal cord injury. Our recent research has found that various biomaterials can promote the repair of spinal cord injury in mice through antioxidant stress. The development of biomaterial sensors, technologies, and the gene modification technology, as well as the optimization of spinal cord injury models are advancing our research field.Due to compress and incompletely understood mechanism of spinal cord injury, it is currently difficult to cure spinal cord injury using a single intervention. In the future, multiple intervention methods may be needed for research. We found that mental immediate compress can participate in the treatment and the repair of spinal cord injury in animals through antioxidant stress.

如上所述，对 24 只雌性小鼠（8 周龄）进行椎板切除术。假手术组（n=6）小鼠未发生脊髓损伤，其余小鼠，包括0.5 mm组（n=6）、0.8 mm组（n=6）和1.1 mm组（n=6）均受到不同深度的脊髓撞击。定期记录 BMS 评分，直至术后 1 个月（图 4）。不同组小鼠术后BMS评分差异有统计学意义。1个月后，0.5mm组小鼠术后评分为4-6分，恢复到与假手术组相似的水平。0.8 mm 和 1.1 mm 组的小鼠术后评...

Watch this Scientific Journal Video about Automated Impactor for Contusive Spinal Cord Injury Model in Mice at JoVE.com

这里介绍的是一种新型的小鼠自动脊髓损伤挫伤装置，可以准确生成不同程度的脊髓损伤挫伤模型。

Spinal cord injury (SCI) due to traumatic injuries such as car accidents and falls is associated with permanent spinal cord dysfunction. Creation of ...

我的研究范围包括脊髓损伤的发病机制和治疗方法。我们最近的研究发现，各种生物材料可以通过抗氧化应激促进小鼠脊髓损伤的修复。生物材料传感器、技术和基因修饰技术的发展，以及脊髓损伤模型的优化正在推进我们的研究领域。由于脊髓损伤的压迫和机制不完全了解，目前很难使用单一干预措施来治愈脊髓损伤。未来，研究可能需要多种干预方法。我们发现精神即刻敷可以通过抗氧化应激参与动物脊髓损伤的治疗和修复。

automated-impactor-for-contusive-spinal-cord-injury-model-in-mice

Research

JoVE Journal

Neuroscience

Automated Impactor for Contusive Spinal Cord Injury Model in Mice

1.2K 次观看.  Jinan University. 我的研究范围包括脊髓损伤的发病机制和治疗方法。我们最近的研究发现，各种生物材料可以通过抗氧化应激促进小鼠脊髓损伤的修复。生物材料传感器、技术和基因修饰技术的发展，以及脊髓损伤模型的优化正在推进我们的研究领域。由于脊髓损伤的压迫和机制不完全了解，目前很难使用单一干预措施来治愈脊髓损伤。未来，研究可能需要多种干预方法。我们发现精?...