美国国立卫生研究院（NIH），约书亚Frase基金会，肌肉萎缩协会（MDA）。

1。设置实验<ol><li>负载动态肌肉控制（DMC）软件。 </li><li>上电生理钻机制造商的指示。 </li><li>运行的DMC软件： </li></ol>这是假设，硬件连接到计算机，运行DMC软件，以及系统校准。软件可能会不时修订，因此，说明当前的软件版本，我们使用下面有关。软件打开时，它已在运行。 用户可以先写在DMC的协议（例如，在50赫兹为1.5秒，以刺激产生等距破伤风）的协议，然后可以保存供日后使用。我们使用两种标准协议：（1） 等长收缩协议 （例如，抽搐或破伤风）;（2） 偏心拉伸损伤协议 。我们开始为抽搐等长收缩协议 。为了优化抽搐，监视器输出的画面是用来在抽搐产生的扭矩可以观察到的电极移动。取得最大的抽搐反应时，收集的数据返回到主屏幕选择继续， 选择开始和完成。 作为弹出窗口保存的文件都保存。保持在同一位置上的电极，破伤风（50赫兹），然后收集和保存。当完成所有等距收缩，偏心拉伸损伤的协议，可以加载和运行。因为有一个一系列收缩在本协议执行（10收缩），在主屏幕上运行一个按钮被选中来揭示运行的所有状态。现在，当开始被选中， 执行所有协议弹出窗口出现，询问到其中的文件要保存的根文件名 ​​和目录。然后， 开始议定书“系列测试和做的事是选择启动偏心协议。这些文件会自动保存到选定的目录。 2。定位于动物的生理钻机<ol><li>动物护理和使用委员会（ACUC）的指导方针，必须遵守所有涉及动物的程序。 </li><li>麻醉犬使用吸入麻醉（如异氟醚）按照批准的协议。检查眨眼反射，以确保该动物是深深麻醉。在整个过程中的CO 2监测，脉搏血氧饱和度和温度，应密切监测。我们通常使用为七氟醚吸入麻醉诱导异丙酚。 </li><li>剃周围头发的横向扼杀（膝盖），将刺激电极放置（即在2-3厘米，大粗隆远端腓骨头的侧面，肢体开始。 </li><li>狗背斜卧，股骨和胫骨，做出正确的角度定位，使盆腔的肢体，和胫骨的生理表上的位置是在桌面平行。无论是左边或右边的肢体可能进行测试。 </li><li>之前附加狗爪子力踏板，检查，以确保该部队踏板是明确的和安全的旋转。按照开机过程的生理钻机和激活踏板。注：力踏板应该是“硬”，当用户尝试以手动方式移动它。 </li><li>轻轻地，其余的狗的爪子在脚踏板;测量和记录狗的爪子的长度从旋转中心轴的横向数字结束。 </li><li>安全在脚下的踏板采用弹性坚持自我包装的爪子。 </li></ol> 3。等距抽搐扭矩（tibiotarsa​​l屈曲） <ol><li>经过安全检查，无菌屏蔽单极连接24轨刺激电极刺激隔离单元（SIU）。 </li><li>设置的刺激：连续脉冲输出，1赫兹（每秒1个脉冲） </li><li>软件设置：负荷等长收缩的协议 （主屏幕上选择Load协议） 。 运行一个按钮应该是可见的“ 开始 ”按钮的旁边。这确保该协议是只运行一次。 </li><li>戴手套的手，用酒精清洁皮肤，和触诊腓总神经近端外侧腓骨远端。一个标记的神经方面的手指，小心地插入皮肤下的阴极（黑色）电极和公正的外部至骨膜。电极应平行放置，深度约2cm的​​皮肤表面，足以防止电极移动时手被删除。 </li><li>阳极（红色）电极放置在皮肤下，并行和阴极电极略有远端。现在应该是内部腓总神经远端腓骨头的两个电极。 </li><li>激活的stimula猛龙在1赫兹。观察显示器上的抽搐反应。在电极位置的小的调整，以尽可能获得最大的抽动幅度。电极应完全去除，并取代在三至四年半厘米的增量平行于胫骨，找到最大的抽搐反应。这是一个过程的重要组成部分，必须十分小心发现的最大的响应。 </li><li>记录抽搐反应退出监视器输出的屏幕（ 选择继续），然后选择开始和完成收集数据。 作为弹出窗口保存所需的文件名 ​​保存文件。关闭的刺激。 </li></ol> 4。等距强直扭矩（tibiotarsa​​l屈曲） <ol><li>无需移动电极，改变刺激设置为1.5秒到50赫兹，使外部触发功能。这使得通过软件激活的刺激。 </li><li>软件设置：同样的等长收缩协议是用于收集到的抽搐，强直响应。假设刺激电极没有被感动了， 开始和完成选择产生破伤风。在等距的协议产生了破伤风，刺激是引发软件，以保持正确的时序。保存在保存为弹出窗口强直响应。 </li></ol> 5。偏心协议<ol><li>而不移动电极，更改的刺激设置：1秒（100V）的50赫兹，使外部触发功能。这使得通过软件激活的刺激。 </li><li>软件设置：负载的偏心牵拉伤的协议 ，命令伺服电机，旋转29度（30度是最高的）。选择运行所有 。选择“开始”;输入偏心系列文件的根文件名 ​​，他们是要保存到的文件夹。选择“ 开始”议定书“的系列测试 ，然后完成启动协议。 </li><li>在这个协议中的刺激是一套软件，以保持正确的时序触发。由于每个离心收缩的反应出现在屏幕上，它会自动保存。 </li><li> 4分钟的休息时间如下每十个离心收缩。 </li><li>共收集了30离心收缩（三套十个收缩;四分钟的休息时间分隔每套）。 </li></ol> 6。等距抽搐扭矩（tibiotarsa​​l扩展） <ol><li>重复上述抽搐协议，但放置电极垂直于皮肤和近端屈曲协议的原始电极放置。胫骨神经刺激gastrocnemious肌肉肚子尾端和远端课程内。配对刺激和参比电极放置在肌肉尾鳍深近端膝关节。近似​​的安置点是沿直线平分的膝关节（膝盖）形成90度角。由于胫神经腓总神经在于更深层次和尾鳍，电极应放置在几个方面引出的最佳抽搐潜力。这是一个过程的关键组成部分。注：tibiotarsa​​l抽搐和手足抽搐扭矩描会出现倒相tibiotarsa​​l屈曲描。 </li></ol> 7。等距强直扭矩（tibiotarsa​​l扩展） <ol><li>无需移动电极，改变刺激设置为1.5秒到50赫兹，使外部触发功能。这使得通过软件激活的刺激。 </li><li>软件设置：50赫兹为1.5秒，使外部触发功能。这使得通过软件激活的刺激。 </li><li>软件设置： 相同的等长收缩协议是用于破伤风的抽搐。假设刺激电极没有被感动了， 开始和完成选择产生破伤风。 </li><li>在等距的协议产生了破伤风，刺激是引发软件，以保持正确的时序。保存强直响应，在弹出的保存文件窗口。 </li></ol> 8。数据分析<ol><li>动态肌肉分析（DMA）方案（极光科学）是用来获取的扭矩和抽搐，强直和离心收缩收集的时间信息。 </li><li>运行DMA程序：软件可能会不时修订，因此，说明当前的软件版本，我们使用下面有关。当DMA软件打开它没有运行。激活该程序，选择左上角的白色箭头按钮。 DMA软件具有多项功能，但我们的目的S，只有一些相关的功能进行说明。 载入档案来分析： 重载最近的数据文件或新的数据文件的弹出窗口时出现的DMA程序启动。选择“ 新建”，在接下来的窗口中选择Labview的 ;在接下来的窗口中，选择相应的文件夹和文件，然后选择“ 打开”。选择删除过滤器删除此窗口的阴谋。在截止频率中输入滤波器的频率。最佳滤波器的频率将有具体的数据进行评估的基础上加以确定。作为一个例子，我们用我们等距反应和10赫兹的偏心响应，30赫兹。选择使用过滤后的数据按钮。在选择地块数据 ，选择筛选队 。使用底部的两个滑杆，扩大显示器和移动文件。 </li><li>抽搐分析（ 图1）：假设文件加载完成上述步骤，设置游标的位置：有3游标：（1）蓝色（2）黄色（3）白色。对于等距抽搐，破伤风或其他等距收缩，蓝色的地步，从静止扭矩扭矩上升，黄色的是放置扭矩返回休息扭矩（即放宽后阶段）;放置白色黄色光标的权利（不需要）。郁闷鼠标拖动选定的光标移动到其所需位置左边的按钮轻松移动光标。 执行分析：当光标定位，选择做肌肉的分析 ，然后显示出结果 。在肌肉的数据分析结果弹出窗口，显示一些变数，如最大的力量 ; 基线队 ; 半弛豫时间等数据在于游标1（蓝色）和2（黄之间的转矩响应，提出）。所有这些变量或特定变量可以被记录到一个数据文件，但是，这一过程不是本文所述。如果数据记录是不成立的，数据都必须由专人记录。选择关闭窗口 ，返回到主屏幕。选择退出计划关闭DMA，然后选择白色箭头开始分析一个新的文件。 </li><li>手足搐搦分析（ 图2）：上面所描述的抽搐的相同的步骤。 </li><li>偏心分析（ 图3）：偏心分析需要更多的步骤来设置和使用所有3个游标。销售如果偏心力矩的反应是负面的（例如，扭矩描出现向下的方向），它可以倒进入-1参考面积（平方厘米）箱（扭矩值不变），并选择不要正常化;。按钮反转指示数据已归（在这种情况下，倒）。输入截止频率所需的切割 （例如，10赫兹）， 并选择使用未经过滤的数据反向按钮来使用过滤后的数据，现在筛选的数据将被用于分析。位置等距扭矩开始上升，从休息扭矩的地步蓝色光标位置的地步偏心拉伸扭矩开始上升等距高原（见下图）黄色光标位置的白色光标的地方扭矩返回到休息扭矩。选择做肌肉的分析 ，然后显示结果 。在所分析的肌肉弹出窗口中的数据结果，您可以选择游标1（蓝色）和2（黄）之间的等距的扭矩数据数据，或游标2（黄色）和3（白色）之间的偏心力矩数据。所有这些变量或特定变量可以被记录到一个数据文件，但是，这一过程不是本文所述。如果数据记录是不成立的，数据都必须由专人记录。选择关闭窗口 ，返回到主屏幕。选择退出计划关闭DMA，然后选择白色箭头开始分析一个新的文件。 </li></ol> 9。代表性的成果<ol><li>抽搐扭矩：胫神经刺激（1赫兹）激活单tibiotarsal伸肌的肌肉抽搐（ 图1） 。请注意，Y轴表示“武力”;扭矩（力和距离的产品）是由用户计算。我们衡量的爪子的长度（从旋转轴外侧的数字结束）。这个长度的距离（力臂），用于确定扭矩（Nm）。 </li><li>等距强直扭矩（tibiotarsa​​l扩展名）：胫神经刺激tetanized tibiotarsa​​l伸肌（1.5秒50HZ脉冲列车）。我们用动态的肌肉控制计算机软件（DMC，震旦科学公司），控制刺激的时间，并捕捉扭矩反应（ 图 2 ）。 </li><li>偏心协议：在tibiotarsa​​l屈肌离心收缩进行。经皮腓总神经stimulATION激活一个初步的联合旋转等长收缩引起的激活屈肌肉（离心收缩;图 3）延长。 </li></ol><img src="/files/ftp_upload/2623/2623fig1.jpg" alt="图1" /> 图1：蓝色（＃1），黄色（＃2）和白色（＃3）游标的位置分析抽搐。一个代表的DMA软件屏幕截图显示在一个健康的13公斤的狗tibiotarsa​​l扩展抽搐。请注意，由软件进行后续分析，数据导出到电子表格，和动力输出（如图所示队（G）可转换为扭矩（Nm）扭矩计算力的产品和力臂的距离（这个距离=长度从旋转轴的测量横向爪子位）。 <img src="/files/ftp_upload/2623/2623fig2.jpg" alt="图2" /> 图2：蓝色（＃1），黄色（＃2）和白色（＃3）游标的位置来分析破伤风。 tibiotarsal扩展手足抽搐，同样的动物，如在图 1所示为代表的DMA软件拍摄的画面显示。 <img src="/files/ftp_upload/2623/2623fig3.jpg" alt="图3" /> 图3：蓝色（＃1），黄色（＃2）和白色（＃3）游标的位置分析偏心拉伸损伤收缩。一个代表的DMA软件屏幕截图所示为一个单一的偏心拉伸损伤收缩在一个健康的13公斤的狗。最初的高原（绿线），说明等距的收缩阶段。等距偏心（拉伸）所示的阶段，在绿线的快速高峰阶段之后。重复离心收缩通常在等距扭矩的稳步下降的结果。

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J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Damage+to+skeletal+muscle+from+eccentric+exercise.">Damage to skeletal muscle from eccentric exercise.</a> Exerc Sport Sci Rev. 33, 98-104 (2005).</li><li>Warren, G. L., Lowe, D. A., Armstrong, R. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Measurement+tools+used+in+the+study+of+eccentric+contraction-induced+injury.">Measurement tools used in the study of eccentric contraction-induced injury.</a> Sports Med. 27, 43-59 (1999).</li></ol>

生产这种视频文章极光科学，公司赞助

此协议是用来评估在大型动物体内的肌肉功能来分析，文件，并确定治疗干预的疗效，或文件疾病进展的生物特性。1-6动物定位，肢体的位置和电极放置的关键变量显着影响的结果。在初步研究中，平均肺泡浓度（MAC）的异氟醚并没有显着影响力测量（Schueler反渗透，科赫H，Kornegay JN，未公布的数据，1993年）的值。硬件和软件市售和行之有效的方法。 1,2,8,4,9-11,6,12建议用户获得和使用本议定书时，上面所述的软件和设备。 修改：当前伺服电机的限制，包括体质量约23公斤和爪的长度为20cm。注意，转矩限制〜8米的扭矩传感器（模型310LR）的N -，这里描述的。这个值可能会超出正常犬在6个月或以上。目前的协议可能修改包括更新软件和增加对传感器的扭矩范围。极光科学制造一个大型动物的肢体定位装置，在目前的协议中使用的立体框架，可用于取代。此方法已成功地被应用到其他的动物模型，包括非人类灵长类动物。

<table border="1"><tbody><tr><th>试剂名称</th><th>公司</th><th>目录编号</th><th>评论（可选） </th></tr><tr><td>刺激电极</td><td> TECA </td><td> 25mmx26G </td><td>一次性使用无菌单极电极针</td></tr><tr><td>刺激</td><td>草</td><td> S88X </td><td></td></tr><tr><td>刺激隔离病房</td><td>草</td><td>绍尔 - C </td><td></td></tr><tr><td>肌肉杠杆系统</td><td>极光科学</td><td> 310C - LR </td><td>踏板选项</td></tr><tr><td>信号接口</td><td>极光科学</td><td>系列604A </td><td></td></tr><tr><td>动态肌肉的控制及分析软件</td><td>极光科学</td><td>极光科学</td><td></td></tr><tr><td> PC /显示器</td><td>极光科学</td><td> Windows XP的计算机</td><td></td></tr><tr><td>肢位设备</td><td>极光科学</td><td>不适用</td><td>可替代立体框架</td></tr><tr><td>表</td><td> Northstar的海关，LLC </td><td>不适用</td><td>定制表</td></tr><tr><td>灵活的包装爪子附件踏板</td><td>安多弗</td><td> PetFlex（5cmx2m） </td><td>可以重复使用一个或两个时间</td></tr></tbody></table>

in vivo canine muscle function assay

我们描述了一种微创和可重复性的方法来衡量犬骨盆四肢肌力和反复离心收缩的肌肉反应。麻醉狗的骨盆四肢被固定在一个立体的框架对准直角股骨胫骨。胶粘剂包装词缀的爪子上安装测量扭矩伺服电机轴踏板。经皮神经刺激激活的爪子骨盆四肢肌肉或者推（延长）或拉（FLEX）对踏板生成等距扭矩。经皮胫神经刺激激活tibiotarsa​​l伸肌。 tibiotarsa​​l屈肌肌肉经皮腓总神经刺激诱导重复偏心（延长）收缩。偏心的协议包括由伺服电机施加的强制拉伸的初始等距收缩。旋转，有效地延长了肌肉，而它的合同， 例如，一个离心收缩。在刺激屈肌肌肉受到一个800毫秒的等距和200毫秒离心收缩。重复此过程每5秒。为了避免疲劳，休息4分钟以下，共30个执行收缩每10收缩。

Watch this Scientific Journal Video about In Vivo Canine Muscle Function Assay at JoVE.com

In Vivo Canine Muscle Function Assay

我们描述了一种微创，无痛的方法来衡量犬后肢的肌肉力量和肌肉反应反复离心收缩。

在犬体内的肌肉功能测定

We describe a minimally-invasive and reproducible method to measure canine pelvic limb muscle strength and muscle response to repeated eccentric 
...

Research

JoVE Journal

Medicine

In Vivo Canine Muscle Function Assay

16.7K 次观看.  Wake Forest University. 我们描述了一种微创，无痛的方法来衡量犬后肢的肌肉力量和肌肉反应反复离心收缩。

视频: In Vivo Canine Muscle Function Assay

An in vivo Assay to Test Blood Vessel Permeability

This method is based on the intravenous injection of Evans Blue in mice as the test animal model. Evans blue is a dye that binds albumin. Under physiologic conditions the endothelium is impermeable to albumin, so Evans blue bound albumin remains restricted within blood vessels. In pathologic conditions that promote increased vascular permeability endothelial cells partially lose their close contacts and the endothelium becomes permeable to small proteins such as albumin. This condition allows for extravasation of Evans Blue in tissues. A healthy endothelium prevents extravasation of the dye in the neighboring vascularized tissues. Organs with increased permeability will show significantly increased blue coloration compared to organs with intact endothelium. The level of vascular permeability can be assessed by simple visualization or by quantitative measurement of the dye incorporated per milligram of tissue of control versus experimental animal/tissue. Two powerful aspects of this assay are its simplicity and quantitative characteristics. Evans Blue dye can be extracted from tissues by incubating a specific amount of tissue in formamide. Evans Blue absorbance maximum is at 620 nm and absorbance minimum is at 740 nm. By using a standard curve for Evans Blue, optical density measurements can be converted into milligram dye captured per milligram of tissue. Statistical analysis should be used to assess significant differences in vascular permeability.

An in vivo Assay to Test Blood Vessel Permeability

We are presenting an in vivo assay to test blood vessel permeability. This assay is based on intravenous injection of a dye and subsequent visualization of its diffusion into interstitial spaces.

一个<em在体内</em分析测试血管通透性

This method is based on the intravenous injection of Evans Blue in mice as the test animal model. Evans blue is a dye that binds albumin. Under ...

科研

医学

In vivo 19F MRI for Cell Tracking

In vivo 19F MRI allows quantitative cell tracking without the use of ionizing radiation. It is a noninvasive technique that can be applied to humans. Here, we describe a general protocol for cell labeling, imaging, and image processing. The technique is applicable to various cell types and animal models, although here we focus on a typical mouse model for tracking murine immune cells. The most important issues for cell labeling are described, as these are relevant to all models. Similarly, key imaging parameters are listed, although the details will vary depending on the MRI system and the individual setup. Finally, we include an image processing protocol for quantification. Variations for this, and other parts of the protocol, are assessed in the Discussion section. Based on the detailed procedure described here, the user will need to adapt the protocol for each specific cell type, cell label, animal model, and imaging setup. Note that the protocol can also be adapted for human use, as long as clinical restrictions are met.

In vivo 19F MRI for Cell Tracking

We describe a general protocol for in vivo cell tracking using MRI in a mouse model with ex vivo labeled cells. A typical protocol for cell labeling, image acquisition processing and quantification is included.

在体内 19女性 MRI对细胞跟踪

In vivo 19F MRI allows quantitative cell tracking without the use of ionizing radiation. It is a noninvasive technique that can be applied to humans. ...

High-throughput Flow Cytometry Cell-based Assay to Detect Antibodies to N-Methyl-D-aspartate Receptor or Dopamine-2 Receptor in Human Serum

Over the recent years, antibodies against surface and conformational proteins involved in neurotransmission have been detected in autoimmune CNS diseases in children and adults. These antibodies have been used to guide diagnosis and treatment. Cell-based assays have improved the detection of antibodies in patient serum. They are based on the surface expression of brain antigens on eukaryotic cells, which are then incubated with diluted patient sera followed by fluorochrome-conjugated secondary antibodies. After washing, secondary antibody binding is then analyzed by flow cytometry. Our group has developed a high-throughput flow cytometry live cell-based assay to reliably detect antibodies against specific neurotransmitter receptors. This flow cytometry method is straight forward, quantitative, efficient, and the use of a high-throughput sampler system allows for large patient cohorts to be easily assayed in a short space of time. Additionally, this cell-based assay can be easily adapted to detect antibodies to many different antigenic targets, both from the central nervous system and periphery. Discovering additional novel antibody biomarkers will enable prompt and accurate diagnosis and improve treatment of immune-mediated disorders.

Over the recent years, live cell-based assays have been used successfully to detect antibodies against surface and conformational antigens. Here, we describe a method using high-throughput flow cytometry enabling the analysis of large cohorts of patients. Detection of novel antibodies will improve diagnosis and treatment of immune-mediated disorders.

高通量流式细胞仪检测细胞为基础的试验检测抗体，以N-甲基-D-天冬氨酸受体或多巴胺2受体在人血清

Over the recent years, antibodies against surface and conformational proteins involved in neurotransmission have been detected in autoimmune CNS diseases ...

Assessment of Right Ventricular Structure and Function in Mouse Model of Pulmonary Artery Constriction by Transthoracic Echocardiography

Emerging clinical data support the notion that RV dysfunction is critical to the pathogenesis of cardiovascular disease and heart failure1-3. Moreover, the RV is significantly affected in pulmonary diseases such as pulmonary artery hypertension (PAH). In addition, the RV is remarkably sensitive to cardiac pathologies, including left ventricular (LV) dysfunction, valvular disease or RV infarction4. To understand the role of RV in the pathogenesis of cardiac diseases, a reliable and noninvasive method to access the RV structurally and functionally is essential.
A noninvasive trans-thoracic echocardiography (TTE) based methodology was established and validated for monitoring dynamic changes in RV structure and function in adult mice. To impose RV stress, we employed a surgical model of pulmonary artery constriction (PAC) and measured the RV response over a 7-day period using a high-frequency ultrasound microimaging system. Sham operated mice were used as controls. Images were acquired in lightly anesthetized mice at baseline (before surgery), day 0 (immediately post-surgery), day 3, and day 7 (post-surgery). Data was analyzed offline using software.
Several acoustic windows (B, M, and Color Doppler modes), which can be consistently obtained in mice, allowed for reliable and reproducible measurement of RV structure (including RV wall thickness, end-diastolic&#160;and end-systolic dimensions), and function (fractional area change, fractional shortening, PA peak velocity, and peak pressure gradient) in normal mice and following PAC.
Using this method, the pressure-gradient resulting from PAC was accurately measured in real-time using Color Doppler mode and was comparable to direct pressure measurements performed with a Millar high-fidelity microtip catheter. Taken together, these data demonstrate that RV measurements obtained from various complimentary views using echocardiography are reliable, reproducible and can provide insights regarding RV structure and function. This method will enable a better understanding of the role of RV cardiac dysfunction.

Right ventricle (RV) dysfunction is critical to the pathogenesis of cardiovascular disease, yet limited methodologies are available for its evaluation. Recent advances in ultrasound imaging provide a noninvasive and accurate option for longitudinal RV study. Herein, we detail a step-by-step echocardiographic method using a murine model of RV pressure overload.

右心室结构和功能的肺动脉缩窄经胸超声小鼠模型的评估

Emerging clinical data support the notion that RV dysfunction is critical to the pathogenesis of cardiovascular disease and heart failure1-3. Moreover, ...

Early Detection of Drug-Induced Renal Hemodynamic Dysfunction Using Sonographic Technology in Rats

The kidney normally functions to maintain hemodynamic homeostasis and is a major site of damage caused by drug toxicity. Drug-induced nephrotoxicity is estimated to contribute to 19- 25% of all clinical cases of acute kidney injury (AKI) in critically ill patients. AKI detection has historically relied on metrics such as serum creatinine (sCr) or blood urea nitrogen (BUN) which are demonstrably inadequate in full assessment of nephrotoxicity in the early phase of renal dysfunction. Currently, there is no robust diagnostic method to accurately detect hemodynamic alteration in the early phase of AKI while such alterations might actually precede the rise in serum biomarker levels. Such early detection can help clinicians make an accurate diagnosis and help in in decision making for therapeutic strategy. Rats were treated with Cisplatin to induce AKI. Nephrotoxicity was assessed for six days using high-frequency sonography, sCr measurement and upon histopathology of kidney. Hemodynamic evaluation using 2D and Color-Doppler images were used to serially study nephrotoxicity in rats, using the sonography. Our data showed successful drug-induced kidney injury in adult rats by histological examination. Color-Doppler based sonographic assessment of AKI indicated that resistive-index (RI) and pulsatile-index (PI) were increased in the treatment group; and peak-systolic velocity (mm/s), end-diastolic velocity (mm/s) and velocity-time integral (VTI, mm) were decreased in renal arteries in the same group. Importantly, these hemodynamic changes evaluated by sonography preceded the rise of sCr levels. Sonography-based indices such as RI or PI can thus be useful predictive markers of declining renal function in rodents. From our sonography-based observations in the kidneys of rats that underwent AKI, we showed that these noninvasive hemodynamic measurements may consider as an accurate, sensitive and robust method in detecting early stage kidney dysfunction. This study also underscores the importance of ethical issues associated with animal use in research.

Early stage hemodynamic dysfunction is critical to the development of kidney disease. Yet, detection methodologies are limited. Recent advances in sonography provide a noninvasive, accurate option for early detection of kidney injury. This study outlines a step-by-step, sonographic methodology for detecting kidney dysfunction using a drug-induced nephrotoxicity rat model.

大鼠使用超声技术药物引起的肾血流动力学障碍的早期检测

The kidney normally functions to maintain hemodynamic homeostasis and is a major site of damage caused by drug toxicity. Drug-induced nephrotoxicity is ...

Ultrasound Based Assessment of Coronary Artery Flow and Coronary Flow Reserve Using the Pressure Overload Model in Mice

Transthoracic Doppler echocardiography (TTDE) is a clinically useful, noninvasive tool for studying coronary artery flow velocity and coronary flow reserve (CFR) in humans. Reduced CFR is accompanied by marked intramyocardial and pericoronary fibrosis and is used as an indication of the severity of dysfunction. This study explores, step-by-step, the real-time changes measured in the coronary flow velocity, CFR and systolic to diastolic peak velocity (S/D) ratio in the setting of an aortic banding model in mice. By using a Doppler transthoracic imaging technique that yields reproducible and reliable data, the method assesses changes in flow in the septal coronary artery (SCA), for a period of over two weeks in mice, that previously either underwent aortic banding or thoracotomy. 
During imaging, hyperemia in all mice was induced by isoflurane, an anesthetic that increased coronary flow velocity when compared with resting flow. All images were acquired by a single imager. Two ratios, (1) CFR, the ratio between hyperemic and baseline flow velocities, and (2) systolic (S) to diastolic (D) flow were determined, using a proprietary software and by two independent observers. Importantly, the observed changes in coronary flow preceded LV dysfunction as evidenced by normal LV mass and fractional shortening (FS). 
The method was benchmarked against the current gold standard of coronary assessment, histopathology. The latter technique showed clear pathologic changes in the coronary artery in the form of peri-coronary fibrosis that correlated to the flow changes as assessed by echocardiography. 
The study underscores the value of using a non-invasive technique to monitor coronary circulation in mouse hearts. The method minimizes redundant use of research animals and demonstrates that advanced ultrasound-based indices, such as CFR and S/D ratios, can serve as viable diagnostic tools in a variety of investigational protocols including drug studies and the study of genetically modified strains.

Coronary flow reserve (CFR) is useful for assessment of myocardial oxygen demand and evaluation of cardiovascular risk. This study establishes a step-by-step transthoracic Doppler echocardiographic (TTDE) method for longitudinal monitoring of the changes in CFR, as measured from coronary artery in mice, under the experimental pressure overload of aortic banding.

冠脉流量和冠状动脉血流储备用小鼠压力超负荷基于模型的超声评估

Transthoracic Doppler echocardiography (TTDE) is a clinically useful, noninvasive tool for studying coronary artery flow velocity and coronary flow ...

Depletion of Mouse Cells from Human Tumor Xenografts Significantly Improves Downstream Analysis of Target Cells

The use of in vitro cell line models for cancer research has been a useful tool. However, it has been shown that these models fail to reliably mimic patient tumors in different assays1. Human tumor xenografts represent the gold standard with respect to tumor biology, drug discovery, and metastasis research 2-4. Tumor xenografts can be derived from different types of material like tumor cell lines, tumor tissue from primary patient tumors4 or serially transplanted tumors. When propagated in vivo, xenografted tissue is infiltrated and vascularized by cells of mouse origin. Multiple factors such as the tumor entity, the origin of xenografted material, growth rate and region of transplantation influence the composition and the amount of mouse cells present in tumor xenografts. However, even when these factors are kept constant, the degree of mouse cell contamination is highly variable.
Contaminating mouse cells significantly impair downstream analyses of human tumor xenografts. As mouse fibroblasts show high plating efficacies and proliferation rates, they tend to overgrow cultures of human tumor cells, especially slowly proliferating subpopulations. Mouse cell derived DNA, mRNA, and protein components can bias downstream gene expression analysis, next-generation sequencing, as well as proteome analysis 5. To overcome these limitations, we have developed a fast and easy method to isolate untouched human tumor cells from xenografted tumor tissue. This procedure is based on the comprehensive depletion of cells of mouse origin by combining automated tissue dissociation with the benchtop tissue dissociator and magnetic cell sorting. Here, we demonstrate that human target cells can be can be obtained with purities higher than 96% within less than 20 min independent of the tumor type.

Human tumor xenografts are vascularized and infiltrated by cells of mouse origin during the growth phase in vivo. To circumvent the bias caused by these contaminating cells during downstream analysis, we developed a method allowing for comprehensive depletion of all mouse cells by magnetic cell sorting.

从人类肿瘤异种移植的小鼠细胞的消耗显着改善靶细胞下游分析

The use of in vitro cell line models for cancer research has been a useful tool. However, it has been shown that these models fail to reliably mimic ...

Phosphorus-31 Magnetic Resonance Spectroscopy: A Tool for Measuring In Vivo Mitochondrial Oxidative Phosphorylation Capacity in Human Skeletal Muscle

Skeletal muscle mitochondrial oxidative phosphorylation (OXPHOS) capacity, which is critically important in health and disease, can be measured in vivo and noninvasively in humans via phosphorus-31 magnetic resonance spectroscopy (31PMRS). However, the approach has not been widely adopted in translational and clinical research, with variations in methodology and limited guidance from the literature. Increased optimization, standardization, and dissemination of methods for in vivo 31PMRS would facilitate the development of targeted therapies to improve OXPHOS capacity and could ultimately favorably impact cardiovascular health. 31PMRS produces a noninvasive, in vivo measure of OXPHOS capacity in human skeletal muscle, as opposed to alternative measures obtained from explanted and potentially altered mitochondria via muscle biopsy. It relies upon only modest additional instrumentation beyond what is already in place on magnetic resonance scanners available for clinical and translational research at most institutions. In this work, we outline a method to measure in vivo skeletal muscle OXPHOS. The technique is demonstrated using a 1.5 Tesla whole-body MR scanner equipped with the suitable hardware and software for 31PMRS, and we explain a simple and robust protocol for in-magnet resistive exercise to rapidly fatigue the quadriceps muscle. Reproducibility and feasibility are demonstrated in volunteers as well as subjects over a wide range of functional capacities.

Phosphorus-31 Magnetic Resonance Spectroscopy: A Tool for Measuring In Vivo Mitochondrial Oxidative Phosphorylation Capacity in Human Skeletal Muscle

This work demonstrates the feasibility of an in vivo phosphorus-31 magnetic resonance spectroscopy (31PMRS) technique to quantify mitochondrial oxidative phosphorylation (OXPHOS) capacity in human skeletal muscle.

磷31磁共振波谱分析：一种测量工具<em&gt;在体内</em&gt;线粒体氧化磷酸化能力人骨骼肌

Skeletal muscle mitochondrial oxidative phosphorylation (OXPHOS) capacity, which is critically important in health and disease, can be measured in vivo ...

In Vivo Evaluation of the Mechanical and Viscoelastic Properties of the Rat Tongue

The tongue is a highly innervated and vascularized muscle hydrostat on the floor of the mouth of most vertebrates. Its primary functions include supporting mastication and deglutition, as well as taste-sensing and phonetics. Accordingly, the strength and volume of the tongue can impact the ability of vertebrates to accomplish basic activities such as feeding, communicating, and breathing. Human patients with sleep apnea have enlarged tongues, characterized by reduced muscle tone and increased intramuscular fat that can be visualized and quantified by magnetic resonance imaging (MRI). The abilities to measure force generation and viscoelastic properties of the tongue constitute important tools for obtaining functional information to correlate with imaging data. Here, we present techniques for measuring tongue force production in anesthetized Zucker rats via electrical stimulation of the hypoglossal nerves and for determining the viscoelastic properties of the tongue by applying passive Lissajous force/deformation curves.

In Vivo Evaluation of the Mechanical and Viscoelastic Properties of the Rat Tongue

We describe a surgical procedure in an anesthetized rat model for determining the muscle tone and viscoelastic properties of the tongue. The procedure involves specific stimulation of the hypoglossal nerves and application of passive Lissajous force/deformation curves to the muscle.

大鼠舌的机械和粘弹性的体内评估

The tongue is a highly innervated and vascularized muscle hydrostat on the floor of the mouth of most vertebrates. Its primary functions include ...

Skeletal Muscle Neurovascular Coupling, Oxidative Capacity, and Microvascular Function with 'One Stop Shop' Near-infrared Spectroscopy

Exercise represents a major hemodynamic stress that demands a highly coordinated neurovascular response in order to match oxygen delivery to metabolic demand. Reactive hyperemia (in response to a brief period of tissue ischemia) is an independent predictor of cardiovascular events and provides important insight into vascular health and vasodilatory capacity. Skeletal muscle oxidative capacity is equally important in health and disease, as it determines the energy supply for myocellular processes. Here, we describe a simple, non-invasive approach using near-infrared spectroscopy to assess each of these major clinical endpoints (reactive hyperemia, neurovascular coupling, and muscle oxidative capacity) during a single clinic or laboratory visit. Unlike Doppler ultrasound, magnetic resonance images/spectroscopy, or invasive catheter-based flow measurements or muscle biopsies, our approach is less operator-dependent, low-cost, and completely non-invasive. Representative data from our lab taken together with summary data from previously published literature illustrate the utility of each of these end-points. Once this technique is mastered, application to clinical populations will provide important mechanistic insight into exercise intolerance and cardiovascular dysfunction.

Skeletal Muscle Neurovascular Coupling, Oxidative Capacity, and Microvascular Function with &#039;One Stop Shop&#039; Near-infrared Spectroscopy

Here, we describe a simple, non-invasive approach using near-infrared spectroscopy to assess reactive hyperemia, neurovascular coupling and skeletal muscle oxidative capacity in a single clinic or laboratory visit.

"一站式" 近红外光谱对骨骼肌神经血管的耦合、氧化能力及微血管功能的作用

Exercise represents a major hemodynamic stress that demands a highly coordinated neurovascular response in order to match oxygen delivery to metabolic ...