这项工作部分是由 jsp KAKENHI 授予号 JP17K11914 支持的。

这项研究得到了作者的机构审查委员会的批准, 所有患者都完成了书面同意表格。
1. 材料的制备
<ol><li>使用个人计算机, 计算机断层扫描 (CT) 数据的面部骨骼和腓骨, 转换软件, 如 InVesalius20和三维 (3 维) CAD 软件 (如搅拌机21)。 注: 为精确设计推荐1毫米 CT 数据的最大厚度。对于实际的手术模拟, 使用病人的 CT 资料。为了研究, 使用免费的人类3维数据22。</li>
	<li>使用3维打印机23, 螺丝, 金属板, 和一个小锯, 不仅检查设计, 而且真实的对象和结果。 注: 本研究是实验性的。金属板、螺丝和小锯子可用于模型手术。而不是金属板, 塑料固定板也可以打印的3维打印机, 连同手术指南。</li>
	<li>使用 InVesalius20将面部骨骼和腓骨的成像数据转换为3维数据 (. stl 格式)。 注: CT 数据基本上以二维 (2 维) 图片的形式记录下来。因此, 在使用3维数据之前, 必须将数据转换为3维数据。免费软件足以达到这个目的。本报告不解释如何将数据传输到3维文件中;其他地方可提供教学视频和指南。</li>
	<li>将每个. stl 文件导入搅拌器21。 注意: CAD 软件通常接受. stl 样式的3维格式。首先, 应在特定的 CAD 软件中通过导入来打开上颌和腓骨. stl 文件。</li>
</ol>2. 设计
<ol><li>骨缺损的修复与骨修复
	<ol>
<li>决定要切除的区域。 注意: 在这个实验性的模拟手术中, 上颌的任何部分都可以被设置为切除区。因为重建后的总上颌骨是困难的, 只有一小部分的上颌将是一个选择初学者。在临床环境中, otorhinolaryngologists 将根据癌变区域决定该区域。</li>
		<li>制作一架大型飞机, 并将其放在该区域的边框上以在对象模式下进行删除 (图 1a和1b)。通过放置第二个平面 (图 1b-1d) 然后继续这样做, 直到飞机环绕整个区域进行拆卸。在对象模式中统一这些平面。</li>
		<li>选择所有这些平面的顶点, 并通过在编辑模式中制作边和面 (图 1e) 将它们连接起来, 以环绕区域进行移除。 注意: 切除的飞机应该被复制和维护, 因为这些原始的飞机在固化切除术时使用和丢弃。在本研究中, 建议每个场合都复制每一个平面和固体, 以使其能够重用它们。</li>
		<li>使用编辑模式中的布尔修饰符从面部骨骼中减去切除固体。这导致一个剃光面部骨 (图 1f), 这是上颌缺损模型。</li>
	</ol></li>
	<li>放置腓骨骨
	<ol>
<li>将腓骨放入上颌缺损区 (图 2a)。放置小立方体在两点 (远端8厘米从腓头和5厘米近侧踝) 在腓骨作为标记 (紫色小立方体显示在图 2)。 注: 在临床情况下, 腓骨从腓头远端8厘米至离侧踝近5厘米之间可使用。通过这个标记, 我们可以很容易地理解可以使用的区域。</li>
		<li>将小多维数据集链接到作为对象模式中的父设置的腓骨。</li>
		<li>将小立方体作为标记, 在上颌病变的几个点, 重建是必要的。用这个标记, 必要的再生产点的能见度增加。</li>
		<li>如果腓骨是从中线放置的, 则在物体模式下, 将腓骨与肺泡骨的前缘吻合。</li>
		<li>使用上上颌截骨前平面作为第一个腓骨截骨平面 (图 2b)。</li>
		<li>放置一个新的截骨平面在适当的对象模式 (图 2c)。将这架新飞机连接到腓骨作为父母的设置。 注意: 通过将父母设置为腓骨, 即使腓骨被移到不同的地方, 这种新的截骨平面与腓骨之间的相对方向仍然保持不变。由这两个切割面包围的腓骨的面积成为第一个腓骨块。</li>
		<li>将腓骨和两个截骨平面复制为对象模式中的父设置。移动这个复制的腓骨, 它有第一块区域, 两端有两个切割平面, 到第二个需要重建的区域 (图 2e) 来规划第二个腓骨块。</li>
		<li>通过在对象模式中添加新平面, 放置第二个切削平面。 注: 第一和第二切割平面将成为第二个腓骨块的两端。如果需要第三个块, 则添加类似的过程。应保持相邻腓骨块之间适当长度的间隙。 注意: 第一和第二块之间的差距被认为是一个舒适的截骨的关键。如果这一差距很大, 截骨将是容易的, 因为广泛的工作空间, 但血管长度是有点浪费。相比之下, 如果差距很小, 截骨变得麻烦, 但第二个或第三个街区可以通过消除浪费的未使用的骨头。</li>
	</ol></li>
	<li>腓骨切割导轨的设计
	<ol>
<li>仅可视化腓骨和切割平面, 以便在对象模式下设计腓骨切割指南 (图 3a)。</li>
		<li>在编辑模式下, 通过沿边缘滑动顶点 (图 3b-3d), 使每个切割平面较小, 仅占用腓骨切割部分的一半面积。 注: 切口导轨的拟合侧为腓骨的侧面。由于饲用血管位于内侧, 指南不是设计在内侧的方面。</li>
		<li>将两端的两个平面联合起来, 以在对象模式中构建一个实体 (图 4a-4e)。选择所有这些平面的顶点, 并通过在编辑模式中制作边和面来形成矩形实心来连接它们。</li>
		<li>使用布尔修饰符 (图 5a-5c) 从这个矩形实体中减去腓骨。 注: 这种减法的表面完全适合腓侧侧面。同样的程序在每一个必要的腓骨块重复。</li>
		<li>在对象模式中统一每个减法实体。</li>
		<li>在减去的固体附近放置一个立方体 (图 5d)。挤出工作面制造柱子 (图 5e-5g)。把这些柱子与减去的固体结合起来。这是腓骨切割指南 (图 5h-5j)。</li>
	</ol></li>
	<li>上颌截骨切口指南 注意: 为了切开上颌, 没有必要为每一个切割面设计指南, 因为只有有限的区域是用腓骨重建的。通常, 两个切割指南, 包括内侧肺泡和侧颧骨地区, 设计。<ol>
<li>准备上颌和颧骨的飞机, 这是原始剩余的表面后, 上颌骨切除。宽度为1厘米的边距足够 (图 6a)。</li>
		<li>挤出步骤2.4.1 中准备的面使平面变厚, 并使用凝固修饰符将其凝固在编辑模式下 (图 6b)。</li>
		<li>删除在步骤2.1 中决定在两端的切除平面上加厚的固体;这就是如何设计上颌切割导轨。 注意: 如果管接头表面是锯齿状的, 则较小的拟合区域就足够了。如果拟合曲面易于平整, 则需要大面积的区域以避免导轨的任何滑动。</li>
	</ol></li>
	<li>腓骨段固定指南 注: 将转移到上颌骨的腓骨段被认为是准确的大小和长度, 但转移的位置可以自由地偏离, 如果不使用固定指南。腓骨和每一个切割平面 (如步骤2.2 所做的) 在这个段中再次使用。<ol>
<li>在布尔修饰符中构造每个腓骨块, 方法是在 "编辑" 模式下, 取出腓骨与实心之间的相交区域 (图 7a和7b)。</li>
		<li>提取每个腓骨块表面的一半。</li>
		<li>在对象模式中联合所有这些曲面 (图 7c)。</li>
		<li>使用刀切 (图 8a) 在编辑模式下删除小面孔, 以确保金属板的贴合空间。</li>
		<li>在编辑模式下使用凝固修饰符 (图 8b-8e) 使曲面变厚。 注: 要稳定固定导轨, 避免翘曲, 必须要有最低的2–3毫米厚度。如果机翼是两端设计的, 它将帮助指导上颌骨不使用任何金属板。</li>
	</ol></li>
</ol>3. 3 维印刷模型手术和真实指南
注: 本报告的主要目的是说明设计手术指南的方法;如果不需要3维打印, 则不需要下面描述的过程。
<ol><li>导出. stl 格式的指南设计, 可以打印3维。</li>
	<li>打印所有指南和骨骼。 注: 在印刷中, 筏被认为是扰乱平滑表面印刷和导致锯齿状表面和较差的骨骼, 所以需要平滑的平面必须向上指出。</li>
	<li>
执行模型手术如下:
	<ol>
<li>与实际手术类似, 先将上颌切口引导至面部骨模型 (图 9a)。然后, 用锯子将脸部骨与切割平面一起切开。</li>
		<li>将腓骨切割指南附在腓骨骨模型上, 将其切成块 (图 9b)。将腓骨块连接至固定指南 (图 9c和9d)。</li>
		<li>使用螺钉和钢板固定本固定指南复杂的上颌缺损 (图 9e)。在固定指南未用螺钉和钢板固定的地方, 将腓骨段修复至上颌骨后, 取下固定指南。这完成了重建 (图 9f)。</li>
	</ol>
</li>
	<li>扫描3维重建的图像, 并用3维扫描仪24将其记录到. stl 格式中。</li>
	<li>比较后模型手术. stl 文件和 CAD 重建设计 (图 10) 使用比较软件25。 注: 通过对虚拟重构设计与制导重建模型的比较, 计算出实际精度。由于 CAD/凸轮精度是在下颌重建10的 2.5 mm 偏差内获得的, 因此该方法需要类似的精度。如果无法获得所需的精度, 请重做虚拟设计。</li>
</ol>

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InVesalius Available from: <a target="_blank" href="https://www.cti.gov.br/en/invesalius">https://www.cti.gov.br/en/invesalius</a> (2018)</li><li>. Blender Available from: <a target="_blank" href="https://www.blender.org/">https://www.blender.org/</a> (2018)</li><li>. Free3D Available from: <a target="_blank" href="https://free3d.com/3d-models/human">https://free3d.com/3d-models/human</a> (2018)</li><li>. MakerBot Replicator+ Available from: <a target="_blank" href="https://www.makerbot.com/replicator/">https://www.makerbot.com/replicator/</a> (2018)</li><li>. Artec Eva Lite Available from: <a target="_blank" href="https://www.artec3d.com/portable-3d-scanners/artec-eva-lite">https://www.artec3d.com/portable-3d-scanners/artec-eva-lite</a> (2018)</li><li>Guerrero-de-Mier, A., Espinosa, M. 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E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Warping+deformation+of+desktop+3D+printed+parts+manufactured+by+open+source+fused+deposition+modeling+(FDM)+system.">Warping deformation of desktop 3D printed parts manufactured by open source fused deposition modeling (FDM) system.</a> International Journal of Mechanical and Mechatronics Engineering (IJMME) - International Journal of Engineering and Sciences (IJENS). 17 (4), 7-16 (2017).</li><li>Maschio, F., Pandya, M., Olszewski, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Experimental+validation+of+plastic+mandible+models+produced+by+a+"low-cost"+3-dimensional+fused+deposition+modeling+printer.">Experimental validation of plastic mandible models produced by a "low-cost" 3-dimensional fused deposition modeling printer.</a> Medical Science Monitor. 22, 943-957 (2016).</li><li>Rendon-Medina, M. A., Andrade-Delgado, L., Telich-Tarriba, J. E., Fuente-Del-Campo, A., Altamirano-Arcos, C. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dimensional+error+in+rapid+prototyping+with+open+source+software+and+low-cost+3D-printer.">Dimensional error in rapid prototyping with open source software and low-cost 3D-printer.</a> Plastic and Reconstructive Surgery - Global Open. 6 (1), 1646 (2018).</li><li>Nizam, A., Gopal, R. N., Naing, L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dimensional+accuracy+of+the+skull+models+produced+by+rapid+prototyping+technology+using+stereolithography+apparatus.">Dimensional accuracy of the skull models produced by rapid prototyping technology using stereolithography apparatus.</a> Archives of Orofacial Sciences. 1, 60-66 (2006).</li></ol>

作者没有什么要申报的。

CAD/CAM 重建被认为有助于达到准确的截骨长度, 宽度和角度的切割骨骼, 而使用切割指南4,5,6,7,8 ,9,10,11,12,13,14,</sup...

cad/cam 技术的使用最近在牙科和义齿工作中有所增加。继 cad/cam 的进化后, 腓骨瓣转移使用 cad/cam 现在用于下颌骨重建领域的肿瘤广泛切除恶性肿瘤1,2,3。一些西方国家的公司已经开始为下颌骨地区提供和销售 CAD/凸轮切割指南。下颌骨的 CAD/凸轮重建被认为在精确度4、5、6、7、8、9、10等方面有优势. ,11。然而, 一个缺点是这项技术在世界范围有限的地区是可利用的, 并且它是非常昂贵的12。因此, 上颌病变的 cad/cam 重建尚未流行。上颌重建的病例数低于下颌骨, 商业指南不常见。
由于在日本没有销售商用上颌 cad/凸轮导轨, 我们使用内部方法开发了 cad/cam 外科指南。cad/cam 指南的临床效果已报告13、14、15、16、17、18、19, 但没有报告如何设计它们。本报告的目的是用一种低成本的内部方法显示 cad/cam 设计方法。

<table border="0" cellpadding="0" cellspacing="0" style="width:1292px;" width="1291">
	<tbody>
		<tr height="66">
			<td height="66" style="height:66px;width:523px;">Information Technology Center, Renato Archer, Campinas, Brazil</td>
			<td style="width:316px;">InVesalius</td>
			<td style="width:383px;">Free software https://www.cti.gov.br/en/invesalius</td>
			<td style="width:71px;"></td>
		</tr>
		<tr height="66">
			<td height="66" style="height:66px;">The Blender Foundation, Amsterdam, Netherlands</td>
			<td>Blender</td>
			<td style="width:383px;">Free software https://www.blender.org/</td>
			<td></td>
		</tr>
		<tr height="66">
			<td height="66" style="height:66px;">TurboSquid, Inc. 935 Gravier St., Suite 1600, New Orleans, LA.</td>
			<td>Free 3D skeletal data file</td>
			<td style="width:383px;">Free3D https://free3d.com/3d-models/human</td>
			<td></td>
		</tr>
		<tr height="66">
			<td height="66" style="height:66px;">MakerBot Industries, LLC One MetroTech Center, 21st Fl, Brooklyn, NY.</td>
			<td>MakerBot Replicator+</td>
			<td>https://www.makerbot.com/replicator/</td>
			<td></td>
		</tr>
		<tr height="66">
			<td height="66" style="height:66px;width:523px;">YouTube (Google, Inc.), 901 Cherry Ave. San Bruno, CA</td>
			<td>video sharing website.</td>
			<td>https://www.youtube.com/results?search_query=invesalius+dicom+to+stl</td>
			<td></td>
		</tr>
		<tr height="66">
			<td height="66" style="height:66px;">Artec 3D, 2, rue Jean Engling, Luxembourg</td>
			<td>Artec Eva Lite</td>
			<td>https://www.artec3d.com/portable-3d-scanners/artec-eva-lite</td>
			<td></td>
		</tr>
		<tr height="66">
			<td height="66" style="height:66px;">CloudCompare</td>
			<td>CloudCompare</td>
			<td>http://www.danielgm.net/cc/</td>
			<td></td>
		</tr>
	</tbody>
</table>

designing cad/cam surgical guides for maxillary reconstruction using an in-house approach

计算机辅助设计/计算机辅助制造 (cad/cam) 现正作为颌面外科的制备技术进行评估。由于这项技术成本高昂, 而且在世界上只有有限的地区可用, 我们开发了一种新的 CAD/CAM 外科指南, 采用内部方法。采用 CAD 软件, 确定上颌切除面积、截割面、腓骨切割面和角度。一旦确定了切除区域, 就用布尔修饰符提取必要的面孔。这些肤浅的面孔被团结了适合骨头的表面和加厚稳定固体。不仅在腓骨和上颌骨的切割指导, 而且转移骨段的位置安排是通过加厚表面的面孔定义的。CAD 设计被记录为. stl 文件和三维 (3 维) 打印为实际的外科指南。为了检查指南的准确性, 采用3维打印的面部和腓骨模型进行手术。这些方法可用于协助未提供商业指南的外科医生。

使用这里提出的程序, 首先确定切除区。使用 CAD 软件, 切除区域完全受面约束。这个区域从面部骨头被一个布尔操作减去。腓骨图像被放置在缺陷上, 腓骨切割面放置在适当的重建点。所有腓骨切开的面孔在父母设置连接了腓骨。这些面孔被做了更小和团结了做固体。腓骨被从这些固体中减去, 然后成为腓骨切割指南。面部骨的其余表面也加厚;这些成为上颌切割指南。腓?...

Watch this Scientific Journal Video about Designing CAD/CAM Surgical Guides for Maxillary Reconstruction Using an In-house Approach at JoVE.com

Designing CAD/CAM Surgical Guides for Maxillary Reconstruction Using an In-house Approach

介绍了计算机辅助设计/计算机辅助制造 (cad/cam) 外科指南的设计方法。切割平面分离, 联合, 加厚, 以方便地可视化必要的骨转移。这些设计可以三维打印和检查的准确性。

用内部方法设计上颌重建的 cad/cam 手术指南

Computer-aided design/computer-assisted manufacturing (CAD/CAM) is now being evaluated as a preparative technique for maxillofacial surgery. Because this ...

designing-cadcam-surgical-guides-for-maxillary-reconstruction-using

Research

JoVE Journal

Medicine

8.9K 次观看.  Kyoto Prefectural University of Medicine. 介绍了计算机辅助设计/计算机辅助制造 (cad/cam) 外科指南的设计方法。切割平面分离, 联合, 加厚, 以方便地可视化必要的骨转移。这些设计可以三维打印和检查的准确性。

视频: Designing CAD/CAM Surgical Guides for Maxillary Reconstruction Using an In-house Approach

A Novel Surgical Approach for Intratracheal Administration of Bioactive Agents in a Fetal Mouse Model

Prenatal pulmonary delivery of cells, genes or pharmacologic agents could provide the basis for new therapeutic strategies for a variety of genetic and acquired diseases. Apart from congenital or inherited abnormalities with the requirement for long-term expression of the delivered gene, several non-inherited perinatal conditions, where short-term gene expression or pharmacological intervention is sufficient to achieve therapeutic effects, are considered as potential future indications for this kind of approach. Candidate diseases for the application of short-term prenatal therapy could be the transient neonatal deficiency of surfactant protein B causing neonatal respiratory distress syndrome1,2 or hyperoxic injuries of the neonatal lung3. Candidate diseases for permanent therapeutic correction are Cystic Fibrosis (CF)4, genetic variants of surfactant deficiencies5 and &alpha;1-antitrypsin deficiency6.
Generally, an important advantage of prenatal gene therapy is the ability to start therapeutic intervention early in development, at or even prior to clinical manifestations in the patient, thus preventing irreparable damage to the individual. In addition, fetal organs have an increased cell proliferation rate as compared to adult organs, which could allow a more efficient gene or stem cell transfer into the fetus. Furthermore, in utero gene delivery is performed when the individual's immune system is not completely mature. Therefore, transplantation of heterologous cells or supplementation of a non-functional or absent protein with a correct version should not cause immune sensitization to the cell, vector or transgene product, which has recently been proven to be the case with both cellular and genetic therapies7.
In the present study, we investigated the potential to directly target the fetal trachea in a mouse model. This procedure is in use in larger animal models such as rabbits and sheep8, and even in a clinical setting9, but has to date not been performed before in a mouse model. When studying the potential of fetal gene therapy for genetic diseases such as CF, the mouse model is very useful as a first proof-of-concept because of the wide availability of different transgenic mouse strains, the well documented embryogenesis and fetal development, less stringent ethical regulations, short gestation and the large litter size.
Different access routes have been described to target the fetal rodent lung, including intra-amniotic injection10-12, (ultrasound-guided) intrapulmonary injection13,14 and intravenous administration into the yolk sac vessels15,16 or umbilical vein17. Our novel surgical procedure enables researchers to inject the agent of choice directly into the fetal mouse trachea which allows for a more efficient delivery to the airways than existing techniques18.

We developed a novel surgical approach for intratracheal administration of bioactive agents into the mouse fetus. The delivery route is more efficient in targeting the fetal mouse lungs than the commonly used intra-amniotic injection. This procedure has to date not been described in a mouse model.

一种新的手术方法气管内注射生物活性剂在胎鼠模型

Prenatal pulmonary delivery of cells, genes or pharmacologic agents could provide the basis for new therapeutic strategies for a variety of genetic and ...

科研

医学

Surgical Procedures for a Rat Model of Partial Orthotopic Liver Transplantation with Hepatic Arterial Reconstruction

Orthotopic liver transplantation (OLT) in rats using a whole or partial graft is an indispensable experimental model for transplantation research, such as studies on graft preservation and ischemia-reperfusion injury 1,2, immunological responses 3,4, hemodynamics 5,6, and small-for-size syndrome 7. The rat OLT is among the most difficult animal models in experimental surgery and demands advanced microsurgical skills that take a long time to learn. Consequently, the use of this model has been limited. Since the reliability and reproducibility of results are key components of the experiments in which such complex animal models are used, it is essential for surgeons who are involved in rat OLT to be trained in well-standardized and sophisticated procedures for this model.
While various techniques and modifications of OLT in rats have been reported 8 since the first model was described by Lee et al. 9 in 1973, the elimination of the hepatic arterial reconstruction 10 and the introduction of the cuff anastomosis technique by Kamada et al. 11 were a major advancement in this model, because they simplified the reconstruction procedures to a great degree. In the model by Kamada et al., the hepatic rearterialization was also eliminated. Since rats could survive without hepatic arterial flow after liver transplantation, there was considerable controversy over the value of hepatic arterialization. However, the physiological superiority of the arterialized model has been increasingly acknowledged, especially in terms of preserving the bile duct system 8,12 and the liver integrity 8,13,14.
In this article, we present detailed surgical procedures for a rat model of OLT with hepatic arterial reconstruction using a 50% partial graft after ex vivo liver resection. The reconstruction procedures for each vessel and the bile duct are performed by the following methods: a 7-0 polypropylene continuous suture for the supra- and infrahepatic vena cava; a cuff technique for the portal vein; and a stent technique for the hepatic artery and the bile duct.

Orthotopic liver transplantation in rats is an indispensable experimental model for biomedical research. Here we present our surgical procedures for orthotopic rat liver transplantation with hepatic arterial reconstruction using a 50% partial graft.

部分原位肝移植大鼠模型的肝动脉重建的外科手术

Orthotopic liver transplantation (OLT) in rats using a whole or partial graft is an indispensable experimental model for transplantation research, such as ...

The Mouse Round-window Approach for Ototoxic Agent Delivery: A Rapid and Reliable Technique for Inducing Cochlear Cell Degeneration

Investigators have utilized a wide array of animal models and investigative techniques to study the mammalian auditory system. Much of the basic research involving the cochlea and its associated neural pathways entails exposure of model cochleae to a variety of ototoxic agents. This allows investigators to study the effects of targeted damage to cochlear structures, and in some cases, the self-repair or regeneration of those structures. Various techniques exist for delivery of ototoxic agents to the cochlea. When selecting a particular technique, investigators must consider a number of factors, including the induction of inadvertent systemic toxicity, the amount of cochlear damage produced by the surgical procedure itself, the type of lesion desired, animal survivability, and reproducibility/reliability of results. Currently established techniques include parenteral injection, intra-peritoneal injection, trans-tympanic injection, endolymphatic sac injection, and cochleostomy with perilymphatic perfusion. Each of these methods has been successfully utilized and is well described in the literature; yet, each has various shortcomings. Here, we present a technique for topical application of ototoxic agents directly to the round window niche. This technique is non-invasive to inner ear structures, produces rapid onset of reliably targeted lesions, avoids systemic toxicity, and allows for an intra-animal control (the contra-lateral ear). Results stemming from this approach have helped deeper understanding of auditory pathophysiology, cochlear cell degeneration, and regenerative capacity in response to an acute injury. Future investigations may use this method to conduct interventional studies involving gene therapy and stem cell transplantation to combat hearing loss.

Various methods exist for introducing ototoxic agents to the cochleae of animal models. Presented is a surgical protocol for delivery of ototoxic agents to the round window niche. The procedure is reliable, creates targeted intra-cochlear lesions, and avoids mechanical damage to the microarchitecture. Examination of cochlear self-repair/regeneration is possible.

老鼠圆窗途径耳毒性剂递送:一个快速和可靠的技术诱导耳蜗细胞变性

Investigators have utilized a wide array of animal models and investigative techniques to study the mammalian auditory system. Much of the basic research ...

Surgical Approach for Middle Cerebral Artery Occlusion and Reperfusion Induced Stroke in Mice

Stroke is a leading cause of death worldwide and continues to be one of the major causes of long-term adult disabilities. About 87% of strokes are ischemic in origin and occur in the territory of the middle cerebral artery (MCA). Currently the only Food and Drug Administration (FDA) approved drug for the treatment of this devastating disease is tissue plasminogen activator (tPA). However, tPA has a small therapeutic window for administration (3 - 6 hr), and is only effective in 4% of the patients who actually receive it. Current research focuses on understanding the pathophysiology of stroke in order to find potential therapeutic targets. Thus, reliable models are crucial, and the MCA occlusion (MCAo) model (also termed the intraluminal filament or suture model) is deemed to be the most clinically relevant surgical model of ischemic stroke, and is fairly non-invasive and easily reproducible. Typically the MCAo model is used with rodents, especially with mice due to all the genetic variations available for this species. Here we describe (and present in the video) how to successfully perform the MCAo model (with reperfusion) in mice to generate reliable and reproducible data.

In order to understand the pathophysiology of stroke, it is important to use reliable models. This paper will describe one of the most frequently used stroke models in mice, termed the middle cerebral artery occlusion (MCAo) model (also termed the intraluminal filament or suture model) with reperfusion.

对大脑中动脉闭塞再灌注损伤诱发中风小鼠的手术方法

Stroke is a leading cause of death worldwide and continues to be one of the major causes of long-term adult disabilities. About 87% of strokes are ...

Intravital Microscopy of Tumor-associated Vasculature Using Advanced Dorsal Skinfold Window Chambers on Transgenic Fluorescent Mice

Tumor and tumor vessel development, as well as tumor response to therapeutics, are highly dynamic biological processes. Histology provides static information and is often not sufficient for a correct interpretation. Intravital evaluation, in which a process is followed in time, provides extra and often unexpected information. With the creation of transgenic animals expressing cell-specific markers and live cell tracers, improvements to imaging equipment, and the development of several imaging chambers, intravital microscopy has become an important tool to better understand biological processes. This paper describes an experimental design for the investigation of tumor vessel development and of therapeutic effects in a spatial and temporal manner. Using this setup, the stage of vessel development, tip cell and lumen formation, blood flow, extravasation, an established vascular bed, and vascular destruction can be visualized and followed. Furthermore, therapeutic effects, intratumoral fate, and the localization of chemotherapeutic compounds can also be followed.

This protocol describes the intravital imaging of transgenic mice expressing cell-specific fluorescent markers. Intravital imaging provides a non-invasive method for high-resolution observations in living animals using implantable windows through which the microscopic visualization of processes is possible. This method is especially useful to study longitudinal processes.

应用先进的背皮窗室对转基因荧光小鼠肿瘤相关血管的活体显微镜观察

Tumor and tumor vessel development, as well as tumor response to therapeutics, are highly dynamic biological processes. Histology provides static ...

Using In Vitro Live-cell Imaging to Explore Chemotherapeutics Delivered by Lipid-based Nanoparticles

Conventional imaging techniques can provide detailed information about cellular processes. However, this information is based on static images in an otherwise dynamic system, and successive phases are easily overlooked or misinterpreted. Live-cell imaging and time-lapse microscopy, in which living cells can be followed for hours or even days in a more or less continuous fashion, are therefore very informative. The protocol described here allows for the investigation of the fate of chemotherapeutic nanoparticles after the delivery of doxorubicin (dox) in living cells. Dox is an intercalating agent that must be released from its nanocarrier to become biologically active. In spite of its clinical registration for more than two decades, its uptake, breakdown, and drug release are still not fully understood. This article explores the hypothesis that lipid-based nanoparticles are taken up by the tumor cells and are slowly degraded. Released dox is then translocated to the nucleus. To prevent fixation artifacts, live-cell imaging and time-lapse microscopy, described in this experimental procedure, can be applied.

Using In Vitro Live-cell Imaging to Explore Chemotherapeutics Delivered by Lipid-based Nanoparticles

Live-cell imaging is a powerful tool to visualize dynamic processes. The examination of fixed cells provides only static pictures, which can lead to misinterpretation and confusion about the process. This work presents a method to study uptake, drug release, and intracellular localization of liposomal nanoparticles in living cells.

使用体外活体细胞成像技术研究脂纳米粒子化疗

Conventional imaging techniques can provide detailed information about cellular processes. However, this information is based on static images in an ...

A Rat Model of Orthotopic Liver Transplantation Using a Novel Magnetic Anastomosis Technique for Suprahepatic Vena Cava Reconstruction

The rat model of orthotopic liver transplantation (OLT) is essential for transplant research. It is a very sophisticated animal model and requires a steep learning curve. The introduction of the cuff technique for anastomosis of the portal vein (PV) and infrahepatic vena cava (IHVC) has significantly simplified the transplant procedure in rats. However, due to the short anterior wall of the recipients' suprahepatic vena cava (SHVC), the cuff technique is very difficult to use for the reconstruction of the SHVC. Most researchers in this field still use the hand-suture technique for SHVC reconstruction, which makes it the bottleneck step in rat orthotopic liver transplantation. The magnetic anastomosis technique (i.e., magnamosis) is a method of connecting two vessels using the attractive force between two magnets. Our recent study has shown that the magnetic anastomosis technique is superior to the hand-suture technique for SHVC reconstruction in rats. In this article, we show a step-by-step protocol for SHVC reconstruction in rats using the novel magnetic anastomosis technique. In this model, the reconstruction of the PV and IHVC was performed by the standard cuff technique, while the reconstruction of the bile duct (BD) was performed by a stent technique. The hepatic re-arterialization was not performed. The magnetic anastomosis technique made SHVC reconstruction much easier and significantly shortened the anphepatic phase. After a reasonable learning curve, even researchers without advanced microsurgical skills can produce reliable and reproducible results using this rat model of OLT.

The reconstruction of the suprahepatic vena cava (SHVC) remains a difficult step in rat orthotopic liver transplantation. In this article, we show a step-by-step protocol for SHVC reconstruction in rats using a novel magnetic anastomosis technique.

应用新型磁吻合技术 Suprahepatic 腔静脉重建大鼠原位肝移植模型的研究

The rat model of orthotopic liver transplantation (OLT) is essential for transplant research. It is a very sophisticated animal model and requires a steep ...

Modified Single-Loop Reconstruction for Pancreaticoduodenectomy

Modified single-loop reconstruction in pancreaticoduodenectomy separates pancreatic secretion from bile. It is performed in cases of high-risk pancreatic remnants to reduce the severity of postoperative pancreatic fistulas and moreover the overall postoperative morbidity. This reconstruction technique is characterized by an extra-long jejunal loop for the construction of the pancreaticojejunostomy and hepaticojejunostomy. The longer distance between these anastomoses and an additional jejuno-jejunostomy between the afferent and efferent limb of the hepaticojejunostomy separate the fluids and prevent backflow of bile towards the pancreaticojejunostomy. Thus, the secretions cannot activate each other and aggravate an existing anastomotic leakage. We observed a reduced rate of severe postoperative pancreatic fistulas after modified single-loop reconstruction compared to conventional single loop reconstruction. The technique is easy to perform, safe, and less time-consuming than a traditional double-loop reconstruction.

The technique of modified single-loop reconstruction following pancreaticoduodenectomy separates pancreatic secretion from bile, thus reducing the severity of POPF without prolonging the duration of surgery.

胰腺切除术的改性单循环重建

Modified single-loop reconstruction in pancreaticoduodenectomy separates pancreatic secretion from bile. It is performed in cases of high-risk pancreatic ...

Orthotopic Rat Kidney Transplantation: A Novel and Simplified Surgical Approach

Kidney transplantation offers increased survival rates and improved quality of life for patients with end-stage renal disease, as compared to any type of renal replacement therapy. Over the past few decades, the rat kidney transplantation model has been used to study the immunological phenomena of rejection and tolerance. This model has become an indispensable tool to test new immunomodulatory pharmaceuticals and regimens prior to proceeding with expensive preclinical large animal studies.
This protocol provides a detailed overview of how to reliably perform orthotopic kidney transplantation in rats. This protocol includes three distinctive steps that increase the probability of success: perfusion of the donor kidney by flushing through the portal vein and the use of a cuff system to anastomose the renal veins and ureters, thereby decreasing cold and warm ischemia times. Using this technique, we have achieved survival rates beyond 6 months with normal serum creatinine in animals with syngeneic or tolerant kidney transplants. Depending on the aim of the study, this model can be modified by pre- or posttransplant treatments to study the acute, chronic, cellular, or antibody-mediated rejection. It is a reproducible, reliable, and cost-effective animal model to study different aspects of kidney transplantation.

The purpose of this manuscript and protocol is to explain and demonstrate in detail the surgical procedure of orthotopic kidney transplantation in rats. This method is simplified to achieve the correct perfusion of the donor kidney and shorten the reperfusion time by using the venous and ureteral cuff anastomosis technique.

大鼠原位肾移植: 一种新的简化的手术方法

Kidney transplantation offers increased survival rates and improved quality of life for patients with end-stage renal disease, as compared to any type of ...

Whole-Mount Immunofluorescence Staining, Confocal Imaging and 3D Reconstruction of the Sinoatrial and Atrioventricular Node in the Mouse

The electrical signal physiologically generated by pacemaker cells in the sinoatrial node (SAN) is conducted through the conduction system, which includes the atrioventricular node (AVN), to allow excitation and contraction of the whole heart. Any dysfunction of either SAN or AVN results in arrhythmias, indicating their fundamental role in electrophysiology and arrhythmogenesis. Mouse models are widely used in arrhythmia research, but the specific investigation of SAN and AVN remains challenging.
The SAN is located at the junction of the crista terminalis with the superior vena cava and AVN is located at the apex of the triangle of Koch, formed by the orifice of the coronary sinus, the tricuspid annulus, and the tendon of Todaro. However, due to the small size, visualization by conventional histology remains challenging and it does not allow the study of SAN and AVN within their 3D environment.
Here we describe a whole-mount immunofluorescence approach that allows the local visualization of labelled mouse SAN and AVN. Whole-mount immunofluorescence staining is intended for smaller sections of tissue without the need for manual sectioning. To this purpose, the mouse heart is dissected, with unwanted tissue removed, followed by fixation, permeabilization and blocking. Cells of the conduction system within SAN and AVN are then stained with an anti-HCN4 antibody. Confocal laser scanning microscopy and image processing allow differentiation between nodal cells and working cardiomyocytes, and to clearly localize SAN and AVN. Furthermore, additional antibodies can be combined to label other cell types as well, such as nerve fibers.
Compared to conventional immunohistology, whole-mount immunofluorescence staining preserves the anatomical integrity of the cardiac conduction system, thus allowing the investigation of AVN; especially so into their anatomy and interactions with the surrounding working myocardium and non-myocyte cells.

We provide a step-by-step protocol for whole-mount immunofluorescence staining of the sinoatrial node (SAN) and atrioventricular node (AVN) in murine hearts.

小鼠窦房和房室结的全卡口免疫荧光染色、共聚焦成像和 3D 重建

The electrical signal physiologically generated by pacemaker cells in the sinoatrial node (SAN) is conducted through the conduction system, which includes ...