蒂姆·戴利和安德鲁·史密斯在提供早期的许多有用的计算讨论;蒂姆·戴利写了协议5.计算资源是由高性能计算集群在南加州大学提供了必要的程序rotate_translate_cylindrical。这项工作是由美国国立卫生研究院授予＃GM098536（MDD）的支持。

所有的过程和人员由南加州学院的大学动物护理和使用委员会（IACUC），协议＃11394批准。 1.解剖短肛和准备<ol><li>通过二氧化碳过度曝光安乐死一个性成熟的雄性小鼠，根据相关机构的动物护理和使用委员会（IACUC）提出的协议。 </li><li>躺在仰卧位的动物，并通过施加压力拖延阴茎大拇指外侧包皮开幕。 </li><li>一旦阴茎旷日持久，通过包皮尽可能延长组织。 </li><li>用剪刀，剪断阴茎体近端，其中阴茎骨所在的龟头。 </li><li>转移解剖阴茎1.7毫升管中，加入200μl自来水。使某些阴茎在液体完全浸没。 </li><li>孵育在水中的组织在约50℃下3-5天。 </li><li&gt;适当温育后，从阴茎骨除去周围组织，用解剖显微镜下用镊子。轻轻喷70％乙醇推下剩余的组织和清理骨头。 </li><li>放置解剖阴茎骨与盖打开一个新的离心管中。请假封顶开O / N为干骨。 </li></ol> 2.显微扫描<ol><li>按一个显微扫描圆柱支架插入花店泡沫砖创造花店泡沫的圆柱体。 </li><li>提取花店泡沫缸和切割片〜2-5厘米厚。 </li><li>干燥后推入的bacula花店泡沫，围绕单个片的外围扫描过程中，尽量减少干扰。骨骼的精确定位应该说允许在协议4个人标本的正确识别。 </li><li>轻轻将切片与嵌入式骨头进入显微持有人。 </li><li>获得显微扫描。在小鼠的bacula 14的情况下</sup90 kVp的，155μA为0.5mm铝过滤器，每180（360覆盖）750预测，曝光时间:&gt;，我们根据以下设置在南加州大学分子影像中心使用的uCT50扫描器（SCANCO医疗AG，Bruttisellen，瑞士） 500毫秒，为15.5mm的体素尺寸。 </li></ol> 3.显微加工:转换一个.DCM堆栈到单个名为.xyz文件注意:每个显微扫描产生一个堆栈.DCM的，或"DICOM"，即表示通过对象拍摄图像片段文件。所有下游计算几何需要平面名为.xyz文件，这是一个简单的包含四列的文本文件 - 在x，y和每个像素的z坐标的，并且所述像素的强度，范围从-5000（黑色）到5000 （白色）。超过3000像素的阈值通常工作以及定义骨骼的阈值。 <ol><li>安装Python（www.python.org）和Python模块命令，DICOM，PYLAB，SYS和numpy的。 </li><li>打开"01_process_dicom.py"{} Figshare用任何文本编辑器，在变量部分，改变路径，像素阈值和目录名是必要的。 </li><li>运行"蟒蛇01_process_dicom.py"。进展情况将被打印到屏幕上。在每个步骤3.2命名的目录，新文件由命名;例如，directory_name.PT3000.xyz，其中PT3000指示像素阈在步骤3.2表示。 </li></ol> 4.显微加工:细分出单独的标本名为.xyz文件<ol><li>与库RGL安装R（https://www.r-project.org/）。 </li><li>用任何文本编辑器打开文件02_segment_dicoms.r&#39;{} Figshare。根据变量部分，更改路径名称指向上述协议3中创建的名为.xyz文件。 </li><li>从内部R，运行命令"源（&#39;02_segment_dicoms.r&#39;）"（不包括双引号）。 </li><li>在协议3中创建的名为.xyz文件的三维图像出现后，输入numbe在整体名为.xyz文件标本河然后贴上标签并选择使用滚动和缩放功能，每个样本点。 注:在后台，独立名为.xyz文件将每个样品上进行。这些出现在一个新的目录，例如，XYZ_FILES_PT3000，其中PT3000表示使用的像素阈值。 </li></ol> 5."对齐"标本名为.xyz文件到通用坐标。 <ol><li>打开Python脚本"03_transform.py"{} Figshare，这需要额外的模块mattdean_modules.py {} Figshare，以及两个自立的应用程序:"rotate_translate_cylindrical"（https://github.com/timydaley/dean_cylindrical_tranform）和"qconvex"（www.qhull.org/html/qconvex.htm）受此脚本中使用。 </li><li>根据变量部分，确定全路径名mattdean_modules.py，rotate_path和qconvex_dir。此外，确定到包含个体.xy目录的完整路径步骤4中创建ž文件。 </li><li>运行03_transform.py，它创建每个试样的新文件的后缀.TRANSFORMED.xyz。 </li></ol> 6."切片"不结盟标本名为.xyz文件识别半地标。 <ol><li>打开并运行Python脚本"04_identify_landmarks.py"{} Figshare。在变量部分，确定全路径名包含.TRANSFORMED.xyz文件的目录。此脚本标识可以用来量化该结构的尺寸和形状802半标。 </li></ol>

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The authors declare that they have no competing financial interests.

在上述协议中的关键步骤是:1）解剖的bacula，2）收集显微图像，3）显微输出转换到XYZ的平面文件坐标，4）分割出各试样的点云，5）将每个试样标准化的坐标系，和6）限定的半标。这些步骤很容易被修改，以适应不同的对象。 这些方法可能可以被应用到实质上是"棒状"，或者至少不太弯曲的任何对象。对象曲线回自己成为"U形"不能被目前分析，因为切片（ 图1G）将来自对象的不同部分返回点。这样的对象可以通过切片之前计算矫直对象被容纳在将来。 我们已经提出了数学上定义半地标一般方法从缺乏小号形状奥利德地标。这些一般方法已被修改，以研究鲸盆腔及肋骨16，它有非常不同形状的演变。我们的定义标计算方法应适用于任何系列的xyz坐标。我们采用显微扫描这里，给小鼠的bacula 14的小尺寸。对于较大的骨头，如鲸骨盆和肋骨，我们采用了重构的骨头16的表面的激光扫描仪。以目视检查半的地标所有集合以验证该方法的质量是很重要的。我们的计算方法的主要优点是，它们正是量化的大小和形状的变化，并保持该物体的不同区域之间的对应关系。

形态测量的领域包括的方法来量化的大小和生物形式的形状，在科学调查1，2，3，4，5，6的一个基本步骤的分集。传统上，大小和形状的统计分析开始于识别上的生物结构的地标，然后测量线性距离，角度和比例，这可以在一个多变量的框架进行分析。里程碑式的基于几何形态特征是保留的地标空间位置，通过分析和可视化5保存从数据收集几何信息的方法。广义普鲁克分析（GPA）可以应用到消除位置，规模，和地标旋转变化，以产生是m标本之间的对准inimizes他们的平方差-剩下的就是形状相异7。 任何形态分析的一个重要概念是同源性，或一个可以可靠地识别代表该标本或结构之间对应生物学意义的和离散的特征的地标的想法。例如，人头骨有同源流程，椎间孔，缝合线，和管道，可以使形态学分析。不幸的是，对应的地标标识是在许多生物结构困难，特别是那些具有光滑表面或曲线8，9，10。 我们下面的办法使用计算几何这个问题。的一般工作流程，以产生可以被表示为点的云的物体的三维扫描，然后旋转和转换，该转换点云，使得所有specimens被定向一个共同的坐标系上。然后我们数学定义从物体的特定区域的半标。放在这些地区离散半地标生物任意11。开展GPA和随后的统计分析可以产生不良的文物8，12，因为随意放置地标可能不是生物同源。因此，我们允许这些半地标数学"滑动"。此过程减少结构之间的电势差。如别处认为这里使用的滑动算法是适当的量化缺乏类似解剖区域容易识别相应的地标3，6，8，10，11，12。这些方法各有其利mitations 13，但应该适应不同尺寸和形状的物体。 这里，我们说明这个方法是如何在近一个鼠标阴茎骨14的研究应用，在已经获得和哺乳动物进化15期间丢失多个独立倍阴茎骨。我们讨论解剖和制备的特定骨的，所述阴茎骨（协议1），生成显微图像（协议2），并且这些图像以一种格式，使所有下游计算几何变换（协议3和4）。完成这些步骤后，由〜100K XYZ坐标每个试样表示。然后，我们通过一系列的变革，有效地调整所有标本到一个共同的方向（协议5）走，然后定义从对准标本半地标（协议6）。协议1-4应该是相似的，无论被分析的对象。协议5和6号议定书是SPEcifically专为阴茎骨，但我们希望通过详细介绍这些步骤，研究者可以想像的修改，这将是对有关他们感兴趣的对象。例如，这些方法改进了用于研究鲸骨盆和肋骨16。

<table><tbody><tr><td>Dissecting scissors</td><td>VWR</td><td>470106-338</td><td>Most sizes should work</td></tr><tr><td>Dissecting Forceps, Fine Tip, Curved</td><td>VWR</td><td>82027-406</td><td></td></tr><tr><td>1.7 mL microcentrifuge tube</td><td>VWR</td><td>87003-294</td><td></td></tr><tr><td>Absolute Ethanol</td><td>Fisher Scientific</td><td>CAS 64-17-5</td><td>To be diluted to 70% for dissections</td></tr><tr><td>Floral Foam</td><td>Wholesale Floral</td><td>6002-48-07</td><td></td></tr><tr><td>uCT50 scanner&amp;nbsp;</td><td>Scanco Medical AG, Bruttisellen, Switzerland</td><td></td><td></td></tr></tbody></table>

dissection, microct scanning and morphometric analyses of the baculum

现代形态测量提供了强大的方法来量化的大小和形状的变化。一个基本要求是，确定地标坐标列表;然而，这样的坐标必须代表整个标本同源结构。虽然许多生物对象包括容易识别标志，以满足同源性的假设，许多缺乏这种结构。一个可能的解决方案是在数学上横跨试样代表相同形态区域中的对象在地方半标。在这里，我们说明了一个新近开发的管道数学定义从鼠标阴茎骨（阴茎骨）半地标。我们的方法应该适用于广泛范围的对象。

在协议6中产生的半地标XYZ坐标可以直接导入到任何基于具有里程碑意义的几何形态测量分析17。计算管道上面已经用于研究小鼠的bacula 14，以及鲸骨盆和肋骨16。上半的地标计算定义的更多细节，这里提出，以试图帮助研究人员可视可能被修改，以适应他们感兴趣的特定对象的步骤。该阴茎骨包含了被利用来自动化某些变换几个独特的功能。例如，经过计算切断骨成两半沿近侧 - 远侧轴，我们确定近端半部简单地通过比较点的总数（在近端有更多的）。只要存在着独特的功能，因为这样，我们的方法应该适应任何目的。此外，应当强调的是，我们凭经验确定特定的阈值，例如"10％近侧"，在我们的阴茎骨研究表现良好，但肯定需要重新评估其他对象。 在协议5开始，第一计算步骤是计算的凸包（最小集合，其中包含在样品中的所有其他点的点），以确定这两个点是从彼此相距最远。这两点（红球， 图1C）开始来定义运行近端-远端穿过骨一个新的z轴（红线， 图1C）。在阴茎骨的情况下，包含更多的点的点云的一半被定义为近端。 第二，整个点云被变换，使得近端点发生在将x，y，0,0,0 z坐标的和远端点t在x，Y，Z 0,0，+ z，其中+ z是依赖于骨的大小一些正值的坐标碟刹。在此步骤结束时，z轴穿过骨的长度。对于下面的程序，从最小到最大z坐标的长度将被称为Zlength。 第三，以校正与上面确定的近端和远端的点的确切位置，在10％的最近端和10％最远侧点被分别采样关联方差（ 图1D），它们各自的质心确定（红球， 图1E）和点云转化使得近侧质心是0,0,0，和远端形心是0,0，+ Z，与通过样品的中心通过一个新的z轴（红线， 图1E） 。 第四，点云围绕z轴通过首先采取SLIC旋转在结构的近端15到15.25％Zlength（蓝点， 图1E）点即分的这片在z方向是扁平的（ 即 z坐标被忽略），所采取的凸包，而最小外接矩形计算（包含所有其他点的最小矩形）。想象一下，这个连接最小外接矩形的两个短边的中点线。我们旋转点云直到这两个中点变得-x，0，+ z和+ X，0，+ Z，分别因而该行成为一个新的X轴。改造后的最大和最小的x值之间的距离被称为Xlength。一个新的文件是从specimen.xyz创建specimen.TRANSFORMED.xyz。 第五，内分1％z轴的Xlength被切出（蓝点， 图1F），和一个最近端和一个最远侧点从该中央SL识别冰和标记的远端和近端，分别。这些都是确定的前两个半地标。 第六，50均匀地间隔开的点片沿z轴（红点， 图1G）采样。每片是厚度为1％Zlength的。每个切片然后在z方向变平，和由7条垂直线（红线， 图1H）平分。每行的2％Xlength内的点被保持（红点， 图1H），再与最大值和点最小y坐标保持，投射到每个相应行，分别标有腹侧和背侧，。此外，标签包含片数和行数，例如P15_VENTRAL4是从第 15片的第 4垂直线取样腹侧点。重要的是，每点标记，例如，P15_VENTRAL4，发生一次且仅一次在所有试样，保留correspondence。除了各7行（14半标总数）的腹侧和背侧点，最大和最小的x值的点进行采样，并分别标有左，右，。左，右的Y和Z坐标采用了R中LOWESS功能对于阴茎骨平滑，共有16个半地标每片的定义（红球， 图1H）; 50片加近端和上面定义远端半地标，802半地标每个试样取样（绿球， 图1I）。从原来的显微扫描所有其它点都被丢弃。 应当指出的是，虽然腹侧/背和近端/远端极性数学确定的，所有检体比对用肉眼确认并根据需要手动调整。在我们的369的bacula的样品，大约10必须手动调整。 图 1: 计算工作流（协议4-6）的可视化表示。 （ 一 ）从02_segment_dicoms.r脚本（协议4），显示出明显的点云个别标本分配的屏幕截图。 （B）一种阴茎骨的放大图，由〜100K的xyz点云表示。的两个点（C）的鉴定从彼此（红球）相隔最远，用来定义运行近端-远端穿过骨（红线）一个新的z轴。 （D）的采样的最近侧10％，最远侧的10％点（红点）提供了在z轴的精确位置，以调整对轻度方差的装置。 （E） -基-至多10％和最远侧的10％（红色球）的质心被用来定义一个新的z轴（红线）。然后，分片之间1下降这种新的z轴（蓝点）5.00-15.25％是采取计算最小边界矩形。点云被旋转直到最小外接矩形的长边平行于一个新的X轴。 F），沿中线（蓝点运行点的切片）进行采样，而最近侧和最远侧点定义为一个半的地标。 G）50间隔均匀的点片进行采样（红点），用H）显示出一个这样的片。 16点（红圈）被定义为捕获每个切片的轮廓。 Ⅰ）当所有片重复，共802半标（绿球）定义的结构和在所有下游形态的应用程序被使用。 <a href="https://ecsource-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/55342/55342fig1large.jpg" target="_blank">请点击此处查看该图的放大版本。</a>

Watch this Scientific Journal Video about Dissection, MicroCT Scanning and Morphometric Analyses of the Baculum at JoVE.com

Dissection, MicroCT Scanning and Morphometric Analyses of the Baculum

许多生物结构缺乏容易定义的地标，从而难以应用现代形态的方法。在这里，我们举例说明的方法来研究的小鼠阴茎骨（在阴茎骨），包括夹层和显微扫描，随后计算方法来定义用于量化的尺寸和形状的变化的半标。

在中短肛解剖，显微扫描和分析形态

Modern morphometrics provides powerful methods to quantify size and shape variation. A basic requirement is a list of coordinates that define landmarks; ...

dissection-microct-scanning-and-morphometric-analyses-of-the-baculum

Nanyang Technological University

Research

JoVE Journal

Bioengineering

7.5K 次观看.  University of Southern California (USC). 许多生物结构缺乏容易定义的地标，从而难以应用现代形态的方法。在这里，我们举例说明的方法来研究的小鼠阴茎骨（在阴茎骨），包括夹层和显微扫描，随后计算方法来定义用于量化的尺寸和形状的变化的半标。 

视频: Dissection, MicroCT Scanning and Morphometric Analyses of the Baculum

Measuring Bone Remodeling and Recreating the Tumor-Bone Microenvironment Using Calvaria Co-culture and Histomorphometry

Bone is a connective tissue constituted of osteoblasts, osteocytes, and osteoclasts and a mineralized extracellular matrix, which gives it its strength and flexibility and allows it to fulfill its functions. Bone is continuously exposed to a variety of stimuli, which in pathological conditions can deregulate bone remodeling. To study bone biology and diseases and evaluate potential therapeutic agents, it has been necessary to develop in vitro and in vivo models.
This manuscript describes the dissection process and culture conditions of calvarias isolated from neonatal mice to study bone formation and the bone tumor microenvironment. In contrast to in vitro and in vivo models, this ex vivo model allows preservation of the three-dimensional environment of the tissue as well as the cellular diversity of the bone while culturing under defined conditions to simulate the desired microenvironment. Therefore, it is possible to investigate bone remodeling and its mechanisms, as well as the interactions with other cell types, such as the interactions between cancer cells and bone.
The assays reported here use calvarias from 5-7 day old BALB/C mice. The hemi-calvarias obtained are cultured in the presence of insulin, breast cancer cells (MDA-MB-231), or conditioned medium from breast cancer cell cultures. After analysis, it was established that insulin induced new bone formation, while cancer cells and their conditioned medium induced bone resorption. The calvarial model has been successfully used in basic and applied research to study bone development and cancer-induced bone diseases. Overall, it is an excellent option for an easy, informative, and low-cost assay.

The ex vivo culture of bone explants can be a valuable tool for the study of bone physiology and the potential evaluation of drugs in bone remodeling and bone diseases. The presented protocol describes the preparation and culture of calvarias isolated from newborn mice skulls, as well as its applications.

利用卡尔瓦里亚共同培养和教条法测量骨重塑和再造肿瘤-骨微环境

Bone is a connective tissue constituted of osteoblasts, osteocytes, and osteoclasts and a mineralized extracellular matrix, which gives it its strength ...

科研

癌症研究

Establishment of a Segmental Femoral Critical-size Defect Model in Mice Stabilized by Plate Osteosynthesis

The use of tissue-engineered bone constructs is an appealing strategy to overcome drawbacks of autografts for the treatment of massive bone defects. As a model organism, the mouse has already been widely used in bone-related research. Large diaphyseal bone defect models in mice, however, are sparse and often use bone fixation which fills the bone marrow cavity and does not provide optimal mechanical stability. The objectives of the current study were to develop a critical-size, segmental, femoral defect in nude mice. A 3.5-mm mid-diaphyseal femoral ostectomy (approximately 25% of the femur length) was performed using a dedicated jig, and was stabilized with an anterior located locking plate and 4 locking screws. The bone defect was subsequently either left empty or filled with a bone substitute (syngenic bone graft or coralline scaffold). Bone healing was monitored noninvasively using radiography and in vivo micro-computed-tomography and was subsequently assessed by ex vivo micro-computed-tomography and undecalcified histology after animal sacrifice, 10 weeks postoperatively. The recovery of all mice was excellent, a full-weight-bearing was observed within one day following the surgical procedure. Furthermore, stable bone fixation and consistent fixation of the implanted materials were achieved in all animals tested throughout the study. When the bone defects were left empty, non-union was consistently obtained. In contrast, when the bone defects were filled with syngenic bone grafts, bone union was always observed. When the bone defects were filled with coralline scaffolds, newly-formed bone was observed in the interface between bone resection edges and the scaffold, as well as within a short distance within the scaffold. 
The present model describes a reproducible critical-size femoral defect stabilized by plate osteosynthesis with low morbidity in mice. The new load-bearing segmental bone defect model could be useful for studying the underlying mechanisms in bone regeneration pertinent to orthopaedic applications.

Although mouse models are invaluable tools for bone tissue engineering, models of long bone defects are sparse. This need motivated development of the present protocol which uses a locking plate with four screws and a dedicated jig to perform and stabilize a reproducible, femoral, critical-size defect with low morbidity.

在小鼠节段性股临界尺寸缺陷模型的建立接骨板稳定

The use of tissue-engineered bone constructs is an appealing strategy to overcome drawbacks of autografts for the treatment of massive bone defects. As a ...

医学

Rigid Embedding of Fixed and Stained, Whole, Millimeter-Scale Specimens for Section-free 3D Histology by Micro-Computed Tomography

For over a hundred years, the histological study of tissues has been the gold standard for medical diagnosis because histology allows all cell types in every tissue to be identified and characterized. Our laboratory is actively working to make technological advances in X-ray micro-computed tomography (micro-CT) that will bring the diagnostic power of histology to the study of full tissue volumes at cellular resolution (i.e., an X-ray Histo-tomography modality). Toward this end, we have made targeted improvements to the sample preparation pipeline. One key optimization, and the focus of the present work, is a straightforward method for rigid embedding of fixed and stained millimeter-scale samples. Many of the published methods for sample immobilization and correlative micro-CT imaging rely on placing the samples in paraffin wax, agarose, or liquids such as alcohol. Our approach extends this work with custom procedures and the design of a 3-dimensional printable apparatus to embed the samples in an acrylic resin directly into polyimide tubing, which is relatively transparent to X-rays. Herein, sample preparation procedures are described for the samples from 0.5 to 10 mm in diameter, which would be suitable for whole zebrafish larvae and juveniles, or other animals and tissue samples of similar dimensions. As proof of concept, we have embedded the specimens from Danio, Drosophila, Daphnia, and a mouse embryo; representative images from 3-dimensional scans for three of these samples are shown. Importantly, our methodology leads to multiple benefits including rigid immobilization, long-term preservation of laboriously-created resources, and the ability to re-interrogate samples.

We developed protocols and designed a custom apparatus to enable embedding of millimeter-scale specimens. We present sample preparation procedures with an emphasis on embedding in acrylic resin and polyimide tubing to achieve rigid immobilization and long-term storage of specimens for the interrogation of tissue architecture and cell morphology by micro-CT.

用微计算机断层扫描法对无节段3D 组织学进行固定染色、全尺寸、毫米度试样的刚性嵌入

For over a hundred years, the histological study of tissues has been the gold standard for medical diagnosis because histology allows all cell types in ...

生物工程

Biomechanical Testing of Murine Tendons

Tendon disorders are common, affect people of all ages, and are often debilitating. Standard treatments, such as anti-inflammatory drugs, rehabilitation, and surgical repair, often fail. In order to define tendon function and demonstrate efficacy of new treatments, the mechanical properties of tendons from animal models must be accurately determined. Murine animal models are now widely used to study tendon disorders and evaluate novel treatments for tendinopathies; however, determining the mechanical properties of mouse tendons has been challenging. In this study, a new system was developed for tendon mechanical testing that includes 3D-printed fixtures that exactly match the anatomies of the humerus and calcaneus to mechanically test supraspinatus tendons and Achilles tendons, respectively. These fixtures were developed using 3D reconstructions of native bone anatomy, solid modeling, and additive manufacturing. The new approach eliminated artifactual gripping failures (e.g., failure at the growth plate failure rather than in the tendon), decreased overall testing time, and increased reproducibility. Furthermore, this new method is readily adaptable for testing other murine tendons and tendons from other animals.

The protocol describes efficient and reproducible tensile biomechanical testing methods for murine tendons through the use of custom-fit 3D printed fixtures.

鼠头的生物力学试验

Tendon disorders are common, affect people of all ages, and are often debilitating. Standard treatments, such as anti-inflammatory drugs, rehabilitation, ...

Analysis of Craniomaxillofacial Malformations in Mice Using Three-dimensional Microcomputed Tomography

To model craniofacial malformations caused by vitamin A deficiency (VAD), we expressed a dominant-negative retinoid receptor mutation in osteoblasts to specifically inhibit RAR transcriptional activity in mice. This approach allowed us to investigate the effects of VAD on cranial hypomineralization, mandibular deformity, and clavicular hypoplasia in clinical cases. In this study, microcomputed tomography (microCT) scanning of the craniomaxillofacial region of mice represented a valuable tool for studying the growth and development of this animal model. The manual estimation of images is both time-consuming and inaccurate. Hence, here, we present a straightforward, efficient, and accurate approach for segmenting and quantifying the microCT images of each craniomaxillofacial bone. MicroCT software was used to slice the mandible, frontal bone, parietal bone, nasal bone, premaxilla, maxilla, interparietal bone, and occipital bone of mice and measure their corresponding lengths and widths. This segmentation method can be applied to study growth and development in developmental biology, biomedicine, and other related sciences and allows researchers to analyze the effects of genetic mutations on individual craniofacial bones.

Here, we describe in depth a method, which is based on microcomputed tomography, to segment and measure 3D models of craniomaxillofacial bones in mice, for better assessment of craniomaxillofacial bone development in mice than what is possible with current methods.

使用三维显微计算机断层扫描分析小鼠颅颌面畸形

To model craniofacial malformations caused by vitamin A deficiency (VAD), we expressed a dominant-negative retinoid receptor mutation in osteoblasts to ...

发育生物学

骨折易感性机械测试的原则和方案

Practical Considerations for the Design, Execution, and Interpretation of Studies Involving Whole-Bone Bending Tests of Rodent Bones

Skeletal fragility leading to fracture is an American public health crisis resulting in 1.5 million fractures each year and $18 billion in direct care costs. The ability to understand the mechanisms underlying bone disease and the response to treatment is not only desired, but critical. Mechanical testing of bone serves as a valuable technique for understanding and quantifying a bone&#39;s susceptibility to fracture. While this method appears simple to perform, inappropriate and inaccurate conclusions may be reached if governing assumptions and key steps are disregarded by the user. This has been observed across disciplines as studies continue to be published with misuse of methods and incorrect interpretation of results. This protocol will serve as a primer for the principles associated with mechanical testing along with the application of these techniques-from considerations of sample size through tissue harvesting and storage, to data analysis and interpretation. With this in hand, valuable information regarding a bone&#39;s susceptibility to fracture may be obtained, furthering understanding for both academic research and clinical solutions.

作者聚焦：提高全骨弯曲测试的准确性和可重复性 

Mechanical testing of rodent bones is a valuable method for extracting information regarding a bone's susceptibility to fracture. Lacking proper practical understanding, the results may be overinterpreted or lack validity. This protocol will serve as a guide to ensure mechanical tests are performed accurately to provide valid and functional data.

设计、执行和解释涉及啮齿动物骨骼全骨弯曲试验的研究的实际考虑因素

Skeletal fragility leading to fracture is an American public health crisis resulting in 1.5 million fractures each year and $18 billion in direct care ...

A Morphometric and Cellular Analysis Method for the Murine Mandibular Condyle

The temporomandibular joint (TMJ) has the capacity to adapt to external stimuli, and loading changes can affect the position of condyles, as well as the structural and cellular components of the mandibular condylar cartilage (MCC). This manuscript describes methods for analyzing these changes and a method for altering the loading of the TMJ in mice (i.e., compressive static TMJ loading). The structural evaluation illustrated here is a simple morphometric approach that uses the Digimizer software and is performed in radiographs of small bones. In addition, the analysis of cellular changes leading to alterations in collagen expression, bone remodeling, cell division, and proteoglycan distribution in the MCC is described. The quantification of these changes in histological sections - by counting the positive fluorescent pixels using image software and measuring the distance mapping and stained area with Digimizer - is also demonstrated. The methods shown here are not limited to the murine TMJ, but could be used on additional bones of small experimental animals and in other regions of endochondral ossification.

This manuscript presents methods for analyzing morphometric and cellular changes within the mandibular condyle of rodents.

小鼠下颌骨髁突的形态学及细胞分析方法

The temporomandibular joint (TMJ) has the capacity to adapt to external stimuli, and loading changes can affect the position of condyles, as well as the ...

生物学

HE 染色和 micro-CT 评估小鼠骨小梁微结构

Trabecular Bone Microarchitecture Evaluation in an Osteoporosis Mouse Model

Bone microstructure refers to the arrangement and quality of bone tissue at the microscopic level. Understanding the bone microstructure of the skeleton is crucial for gaining insight into the pathophysiology of osteoporosis and improving its treatment. However, handling bone samples can be complex due to their hard and dense properties. Secondly, specialized software makes image processing and analysis difficult. In this protocol, we present a cost-effective and easy-to-use solution for trabecular bone microstructure analysis. Detailed steps and precautions are provided. Micro-CT is a non-destructive three-dimensional (3D) imaging technique that provides high-resolution images of trabecular bone structure. It allows for the objective and quantitative evaluation of bone quality, which is why it is widely regarded as the gold standard method for bone quality assessment. However, histomorphometry remains indispensable as it offers crucial cellular-level parameters, bridging the gap between two-dimensional (2D) and 3D assessments of bone specimens. As for the histologic techniques, we chose to decalcify the bone tissue and then perform traditional paraffin embedding. In summary, combining these two methods can provide more comprehensive and accurate information on bone microstructure.

作者聚焦:一种经济有效的小鼠骨质疏松症模型中骨微结构定量评估的方法 

This protocol presents an economical and efficient method for quantitative evaluation of bone microarchitecture in a mouse model of osteoporosis by combining Hematoxylin-Eosin (HE) staining and micro-computed tomography (Micro-CT) techniques.

骨质疏松症小鼠模型中的骨小梁微观结构评估

Bone microstructure refers to the arrangement and quality of bone tissue at the microscopic level. Understanding the bone microstructure of the skeleton ...

在中短肛解剖，显微扫描和分析形态

摘要

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此视频中的章节

利用卡尔瓦里亚共同培养和教条法测量骨重塑和再造肿瘤-骨微环境

在小鼠节段性股临界尺寸缺陷模型的建立接骨板稳定

用微计算机断层扫描法对无节段3D 组织学进行固定染色、全尺寸、毫米度试样的刚性嵌入

鼠头的生物力学试验

使用三维显微计算机断层扫描分析小鼠颅颌面畸形

设计、执行和解释涉及啮齿动物骨骼全骨弯曲试验的研究的实际考虑因素

设计、执行和解释涉及啮齿动物骨骼全骨弯曲试验的研究的实际考虑因素

设计、执行和解释涉及啮齿动物骨骼全骨弯曲试验的研究的实际考虑因素

小鼠下颌骨髁突的形态学及细胞分析方法

骨质疏松症小鼠模型中的骨小梁微观结构评估