1。测量抑制<ol><li>在抑制测量开始之前，给病人详细的说明（见1.8）以下步骤1.2-1.8笔记本电脑的屏幕上显示的视觉刺激。所有的患者应戴眼镜/隐形眼镜屈光矫正（）在整个过程中，适当调整工作距离。 </li><li>的护目镜（eMagin公司8700），应安装的病人（上方的眼镜/隐形眼镜）和适当的调整。确保屏幕坐眼球表面平行。该屏幕可以调整，考虑到每个病人的瞳距。连接和校准护目镜可以在这里找到： <a href="http://www.3dvisor.com/wpcontent/uploads/2010/08/UserGuide10.7.pdf" target="_blank">http://www.3dvisor.com/wpcontent/uploads/2010/08/UserGuide10.7.pdf</a> 。 </li><li>刺激包括两类人群移动的小圆点，人口居无定所g的一个共同的方向（在&#39;signal&#39;点），在随机的方向（“噪音”点）和其他移动。观察员的任务是报告的信号方向。操作任务难度不同的刺激信号对噪声点的相对比例。 </li><li>自定义控制这些测量的Matlab程序，利用3下降1的楼梯程序。有两个独立的测量阶段。第一提供了双眼的阈值，即，信号点的阈值数时，需要两个信号和噪声的点呈现在高对比度双眼。第二程序时实现相同的阈值的点的种群分开的眼睛所需测量的对比度不平衡。 </li><li>的护目镜通过使用一个Matrox DualHead2Go的，（蒙特利尔，加拿大）的移动设备的屏幕上显示不同的图像。 </li><li>要开始的压制评估，测量一个双眼门槛。在这里，相同的视觉刺激AR两个眼睛和需要准确地判断点移动的方向上的信号点的最小数目的E测定。 </li><li>要对齐的眼睛，每只眼睛对一个白色的屏幕，屏幕将显示一个黑色的半剖。使用键盘时，病人被要求弱视眼看到的交叉移动的一半，直到它的另一只眼睛看到的一半。重要的是，建议患者弱视眼看到的图像会变得模糊，但是，重要的是，它是取向。如果运动图像从视图中消失询问病人更频繁地闪烁，因为这可能会提高知名度的图像。可以提取精确眼部对齐（像素值），每次测量后，以确定此任务的可重复性。 </li><li>一旦半十字架对准病人的报告，就可以开始测量程序。每次测量大约需要三分钟就可以完成。五短梯测量是completed的两个部分的每一个测量。测量开始之前，应给予以下说明： </li><li>为观察员的说明： “在这个测试中，你会被要求作出判断的方向移动图像。” “你会看到一组白色/灰色灰色背景点。，每个演示后，我会问你决定点多移动到左边或右边使用键盘上的左，右箭头键在你的面前“。 “有两组的点，一些移动顺利的左侧或右侧（信号点），另一组出现在随机的方向移动。” “请尝试决定哪个方向（向左或向右）移动的信号点的随机点之间的混乱。” “当你继续做出决定，随机点的数量将会增加，使得它更难以看到的信号点，当它变得如此困难时，你可以不告诉的方向，请你只需要您的最佳猜测“。 </li><li>现在就可以开始第一次测量。一个很好的&#39;最后一分钟&#39;的指令是要提醒参与者定期闪烁（闪烁的好时机是当你按下按钮“）。 </li><li>现在做五次测量的双眼楼梯范例。眼对准反复进行，在每次测量之前浏览网页。 </li><li>从每一次测量得到的结果是阈值的信号点（总共100点）时，两个眼睛看到的相同的刺激，正确判断方向。该计划还报告了一系列的反应，作为一个标准的测量误差。 </li><li>进行下一个测量，对比度楼梯，从双目楼梯五个结果进行平均，所得到的阈值定义的信号点的数目提交弱视眼。余下的噪声的点的非弱视眼在不同的对比来评估抑制。 </li><li>对于这种测量弱视眼总是能看到高对比度的点而使用的楼梯过程变化的噪声的对比度的点示出的非弱视眼。楼梯开始在一个固定数量的信号点（从步骤1.11）得到被显示在100％时的对比度和噪声点的剩余数的弱视眼对侧眼显示在0％的对比度（ 即无噪音点是可见的在最小的抑制所导致）。正确识别的信号点方向的查询结果在增加的对比度噪声点示出根据3 1-楼梯算法对侧眼。在整个测量显示对侧眼的噪声点，对比不同的楼梯，直到任务的性能收敛于79％的正确。这表明，信号和噪声的点的两只眼睛之间的组合，以产生相同的任务的性能水平，双眼观看条件（步骤1.11）下观察。的对比度的比值弱视眼（100％）的对比显示对侧眼在阈值的噪声点的信号点是衡量的平衡点。提高对比度显示对侧眼上面这点的噪声点，会造成抑制的信号点执行任务的弱视眼及减值。 </li><li>五个测量的对比度不平衡，并计算平均的结果。结果通知水平的对比失衡，是要克服抑制，使弱视眼和对侧眼看到点同时考官。 </li></ol> 2。使用这种方法的培训<ol><li>可以使用以上步骤1.14中发现的对比度不平衡，作为起点的培训制度。培训涉及到视频游戏格式（“俄罗斯方块”）结合使用dichoptic图像的对比失衡。的视频游戏可以实现对无论是虚拟现实护目镜或iPod Touch设备上。 </li><li>的视频游戏俄罗斯方块涉及一系列的下降块装配在一起，以形成完整的行。的弱视眼看到完整的对比度块，对侧眼看到了低对比度图像。信息提交给每只眼睛必须同时被视为成功的发挥。 </li><li>随着时间的推移，作为训练政权继续降低的对比度不平衡通过增加对比度对侧眼，使之更难以视觉系统来克服更多类似的dichoptic的图像所造成的抑制。 </li><li>培训时间应为1-2小时，一天，应该继续下去，直到没有进一步观察对比失衡的改善（增加对侧眼的对比介绍）。在培训制度，应定期进行检查单眼和双眼功能的病人年龄及相关条件决定。单眼视力，立体视功能的测试抑制和标准的测试尤为重要报告进展情况。 </li></ol>

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在这个手稿中描述的技术形成的基础上通过BT和RFH，其中一个还举行了由JC和LT持有的两项专利。

抑制测量和治疗本文中描述的技术关键取决于操纵每只眼睛看到的图像的对比度。特别是，在出现双眼组合的相对两眼对比失衡， 即 “平衡点对比”，提供了6,11,17抑制措施。此外，通过反复使患者弱视和抑制，以“平衡”的视觉刺激，它是可以减少抑制潜在的提高单眼和双眼视觉功能12,15,16。这些技术代表了提前抑制测量和弱视治疗的新方法。目前可用的临床试验用于评估抑制如Worth四灯测试和横纹肌镜片Bagolini评估抑制是是否存在或不存在，和其他测试，如斯比萨酒吧量化的水平抑制由使用中性密度过滤器。我们的技术提供了更多的临床信息，允许进行精确量化的力量抑制。弱视，往往与抑制，一般被阻断非弱视眼或有辱人格的非弱视眼的图像，以鼓励使用弱视眼的18。尽管此方法有效地提高弱视眼的功能18，它不直接处理伴有弱视双眼赤字。我们的技术直接针对双眼视觉功能，我们已经发现，这种方法既可以提高单眼弱视12,15,16和双眼视功能的成年患者。测量和治疗方法的描述是适合成人和儿童的使用，虽然游戏在年轻的儿童人群中的使用可能需要适应。目前，我们正在为此目的开发更多的游戏。 我们的技术也有其局限性。用的斜视和强抑制有些患者发现它具有挑战性所示的图像的每个眼睛在抑制测量协议的开始对齐。虽然它有可能为这样的患者的图像17对齐，这可能是耗时的。治疗方面，重要的是一起工作，整个培训制度，有适当资格的临床医生，如果抑制降低，并考虑复视（复视）的风险。我们以前的工作对这种治疗方法一直专注于屈光参差或小角度斜视的患者，没有病人有复视，复视，但风险应该仔细考虑门诊的基础上，由合格的临床医生对病人治疗前的管理。 第一次设立技术是很重要的，以确保所有的硬件​​连接是安全的，所有相关的驱动程序在主机电脑上停止和更新。同样重要的是使用的电脑配备的图形卡是兼容的协议中描述的Matrox图形卡，并支持1600x600的屏幕分辨率（Matr​​ox驱动程序安装一次），以确保每个的护目镜屏幕具有鲜明的图像。我们发现，大多数遇到的问题时，设立的发生是由于不正确的屏幕分辨率设置，并没有最新的驱动程序进行必要的硬件系统。同样重要的是确保护目镜屏幕前具有相同的亮度测量抑制，以确保准确和稳定的测量。 所描述的技术测量的抑制和治疗方法的进一步发展提供了基础。特别是眼睛之间的不同对比的核心原则的基础上更吸引人的视频游戏的发展可以使这种方法更高效，更是一个到年轻患者ppealing。除了 ​​从它们的临床使用，这些技术也可以被使用在研究设置探索的神经机制抑制和抑制的眼睛之间的相互作用发挥作用，在正常的视觉系统13。

<table border="1"><tbody><tr><td> 的设备名称 </td><td> 公司 </td><td></td><td></td></tr><tr><td></td><td></td><td></td><td> 的抑制的测量设备 </td></tr><tr><td>计算机配备了图形卡兼容使用Matrox决斗头去设备（见下文）和Psychtoolbox 19,20 </td><td>任何合适的制造商</td><td></td><td></td></tr><tr><td> Matlab软件兼容的版本与Psychtoolbox </td><td> Mathworks公司</td><td></td><td></td></tr><tr><td>为刺激产生的Psychtoolbox和定制软件</td><td> Psychtoolbox </td><td></td><td> Psychtoolbox可以下载从<a href="http://psychtoolbox.org" target="_blank">psychtoolbox.org</a>和额外的定制软件可从相应的作者可达的要求。 </td></tr><tr><td>的Matrox决斗头去</td><td>的Matrox </td><td></td><td></td></tr><tr><td> Z800 3D双亲HMD视频眼镜</td><td> eMagin公司</td><td></td><td></td></tr><tr><td></td><td></td><td></td><td> 抑制治疗设备 </td></tr><tr><td> iPod的触摸</td><td>苹果</td><td></td><td></td></tr><tr><td>屏幕覆盖，以便dichopic的iPod观看</td><td>空间视角</td><td></td><td></td></tr><tr><td>俄罗斯方块处理游戏的自定义软件</td><td></td><td></td><td>这个软件是使用的空间视角专有的SDK，充满细节已经公布以前15 </td></tr></tbody></table>

the measurement and treatment of suppression in amblyopia

弱视是一种发育障碍的视觉皮层，处于工作年龄人口的视觉功能障碍的主要原因之一。目前估计弱视的患病率约为1-3％，1-3，大多数情况下是单眼2。最经常引起弱视眼偏差（斜视），模糊引起的不平等的屈光不正（屈光参差），并在某些情况下，形觉剥夺性。 ，虽然弱视最初所造成的异常视觉输入处于起步阶段，一旦建立，的视觉赤字时，往往仍然采用手术和/或屈光矫正已经恢复了正常的视觉输入。这是因为弱视是视觉皮层的异常发展，而不是一个问题弱视眼4,5的结果。弱视的特点是单眼和双眼两种赤字6,7，其中包括视力受损和差或无小号tereopsis。弱视的视觉功能障碍往往是相关的投入弱视眼的双筒望远镜观看条件下8上具有较强的抑制。最近的研究工作表明，抑制发挥了核心作用，无论是单眼和双眼赤字与弱视9,10。 目前临床试验的抑制倾向，来验证是否存在或不存在的抑制，而不是给抑制的程度的定量测量。在这里，我们描述了一种技术具有结构紧凑，便携式设备11,12为测量弱视抑制的。该装置由一台笔记本电脑连接到一对虚拟现实护目镜。该技术的新颖之处在于我们带来了视觉刺激来衡量抑制的方式。的刺激被显示在高对比度的弱视眼是多种多样的，而所示的非弱视眼的刺激的对比度。患者进行SIMPLE信号/噪声任务，允许精确测量兴奋的双眼相互作用的强度。既不眼具有性能上的优势的对比度偏移是“平衡点”的措施，是抑制的直接量度。对照13,14和病人6,9,11人口，这项技术已被验证之心。 除了 ​​测量抑制这种技术还形成了一种新的治疗方式的基础上，随着时间的推移，以减少抑制和改善功能的成年患者双眼和单眼弱视12,15,16。这种新的治疗方法，可部署的护目镜系统，上述或一个专门修改后的iPod touch设备15。

的抑制水平，发现在所描述的方法是依赖于弱视，以前的治疗中，使用的屈光校正视力的根本原因。因为每个病人有一个非常独特的历史，它是很难界定“正常”值，抑制或使人与人之间的比较。通常情况下，我们期望那些视力较差，有一个更深层次的抑制9。在训练过程中，我们降低对比度眼睛之间的失衡，抑制的深度通常会降低，这提高了一系列的双眼和单眼功能。已被证明在抑制这种变化在几个患者人群（见参考12， 表3和图9中的参考文献15和图2图3和4中，在参考文献16）。重要的是要注意，详细的测量抑制日的技术有可能提高认识弱视综合征。 <img alt="图1" src="/files/ftp_upload/3927/3927fig1.jpg" /> 图1。设备用于测量抑制，其中包括1）笔记本电脑2）护目镜3）信号分配器。 <img alt="图2" src="/files/ftp_upload/3927/3927fig2.jpg" /> 图2。研究参与者的护目镜上正确安装。 <img alt="图3" fo:content-width="5in" fo:src="/files/ftp_upload/3927/3927fig3highres.jpg" src="/files/ftp_upload/3927/3927fig3.jpg" /> 图3概述的的随机点kinetogram刺激（步骤1.2/1.3）。顶部面板显示的对齐方式相半跨显示各眼单眼双目提示<html> cular对齐。中间面板中演示的信号噪声范式信号点呈现给弱视眼只和噪声点被提交至对侧眼，在双目条件下的信号和噪声相结合，得到的运动相干阈值，然后，利用在对比度不同阶段测量。底部面板中显示的对比度阈值，该过程使用的议案连贯性阈值的信号对噪声在一个固定水平，并显示不同的两只眼睛之间的对比度水平。这一点在两只眼睛看到一个平衡的双眼输入的对比度阈值或“平衡点”。 <img alt="图4" src="/files/ftp_upload/3927/3927fig4.jpg" /> 图4（1.7）对准屏幕上看到通过护目镜，水平和垂直方向键是用来校准目标，直到被视为整个横。 e_content的“Fo：保持together.within”页面=“总是”&gt; <img alt="图5" fo:content-width="5in" fo:src="/files/ftp_upload/3927/3927fig5highres.jpg" src="/files/ftp_upload/3927/3927fig5.jpg" /> 图5。训练协议的概述。 <img alt="图6" src="/files/ftp_upload/3927/3927fig6.jpg" /> 图6例临床及心理获得的数据从一个斜视性弱视，成年参与者完成了训练程序（步骤2.1至2.4）。 图6A表明视力的变化（测量四个星期的LogMAR单位的对数款式图训练弱视眼（AME）和对侧眼（FFE））。较小的的的logMAR视力值表示更好的视力。的累积数量小时的游戏的是， 图中所示的x轴的支架上。6B演示imba的对比度的变化枪超过四个星期的培训。 Y轴显示的对比度是可以容忍的，因此，较大的值表示抑制少对侧眼。在只有30％的对比度是可以容忍的，在训练开始前，对侧眼弱视眼的抑制。但是作为训练的进展，更多的对比可以指示抑制降低的耐受性， 图6C显示了在训练期间（测量与Randot音响测试）在立体视改善。在y-轴的零表示没有立体视觉和增加值表明改善立体声灵敏度（单位是立体声弧秒的阈值的倒数）。 <img alt="图7" src="/files/ftp_upload/3927/3927fig7.jpg" /> 图7。标准的临床试验来评估培训，包括Worth四点测试（左上）前后双眼视觉状态，Bagollini乐网络搜索引擎（右上），Randot立体视觉测试（左下）和TNO立体视觉检查（右下）。

Watch this Scientific Journal Video about The Measurement and Treatment of Suppression in Amblyopia at JoVE.com

The Measurement and Treatment of Suppression in Amblyopia

弱视是一种发育障碍，往往是伴随着强烈的抑制一只眼的视觉皮层。我们提出了一种新的技术测量两眼抑制和治疗，在弱视患者可以利用虚拟现实护目镜或便携式iPod Touch设备部署的。

测量和制止在弱视治疗

Amblyopia, a developmental disorder of the visual cortex, is one of the leading causes of visual dysfunction in the working age population. Current ...

the-measurement-and-treatment-of-suppression-in-amblyopia

Research

JoVE Journal

Medicine

49.5K 次观看.  University of Auckland. 弱视是一种发育障碍，往往是伴随着强烈的抑制一只眼的视觉皮层。我们提出了一种新的技术测量两眼抑制和治疗，在弱视患者可以利用虚拟现实护目镜或便携式iPod Touch设备部署的。

视频: The Measurement and Treatment of Suppression in Amblyopia

Epidural Intracranial Pressure Measurement in Rats Using a Fiber-optic Pressure Transducer

Elevated intracranial pressure (ICP) is a significant problem in several forms of ischemic brain injury including stroke, traumatic brain injury and cardiac arrest. This elevation may result in further neurological injury, in the form of transtentorial herniation1,2,3,4, midbrain compression, neurological deficit or increased cerebral infarct2,4. Current therapies are often inadequate to control elevated ICP in the clinical setting5,6,7 . Thus there is a need for accurate methods of ICP measurement in animal models to further our understanding of the basic mechanisms and to develop new treatments for elevated ICP.
In both the clinical and experimental setting ICP cannot be estimated without direct measurement. Several methods of ICP catheter insertion currently exist. Of these the intraventricular catheter has become the clinical 'gold standard' of ICP measurement in humans8. This method involves the partial removal of skull and the instrumentation of the catheter through brain tissue. Consequently, intraventricular catheters have an infection rate of 6-11%9. For this reason, subdural and epidural cannulations have become the preferred methods in animal models of ischemic injury. 
Various ICP measurement techniques have been adapted for animal models, and of these, fluid-filled telemetry catheters10 and solid state catheters are the most frequently used11,12,13,14,15. The fluid-filled systems are prone to developing air bubbles in the line, resulting in false ICP readings. Solid state probes avoid this problem (Figure 1). An additional problem is fitting catheters under the skull or into the ventricles without causing any brain injury that might alter the experimental outcomes. Therefore, we have developed a method that places an ICP catheter contiguous with the epidural space, but avoids the need to insert it between skull and brain. 
An optic fibre pressure catheter (420LP, SAMBA Sensors, Sweden) was used to measure ICP at the epidural location because the location of the pressure sensor (at the very tip of the catheter) was found to produce a high fidelity ICP signal in this model. There are other manufacturers of similar optic fibre technologies13 that may be used with our methodology. Alternative solid state catheters, which have the pressure sensor located at the side of the catheter tip, would not be appropriate for this model as the signal would be dampened by the presence of the monitoring screw. 
Here, we present a relatively simple and accurate method to measure ICP. This method can be used across a wide range of ICP related animal models.

A novel technique to record the pressures within the skull is described. The minimally invasive method uses a fibre-optic pressure sensing system to accurately measure intracranial pressure (ICP) in anaesthetized rats without causing significant brain trauma. The technique may be used in a wide range of experimental models.

硬膜外颅内压测量中使用的光纤压力传感器的大鼠

Elevated intracranial pressure (ICP) is a significant problem in several forms of ischemic brain injury including stroke, traumatic brain injury and ...

科研

医学

In vivo Measurement of the Mouse Pulmonary Endothelial Surface Layer

The endothelial glycocalyx is a layer of proteoglycans and associated glycosaminoglycans lining the vascular lumen. In vivo, the glycocalyx is highly hydrated, forming a substantial endothelial surface layer (ESL) that contributes to the maintenance of endothelial function. As the endothelial glycocalyx is often aberrant in vitro and is lost during standard tissue fixation techniques, study of the ESL requires use of intravital microscopy. To best approximate the complex physiology of the alveolar microvasculature, pulmonary intravital imaging is ideally performed on a freely-moving lung. These preparations, however, typically suffer from extensive motion artifact. We demonstrate how closed-chest intravital microscopy of a freely-moving mouse lung can be used to measure glycocalyx integrity via ESL exclusion of fluorescently-labeled high molecular weight dextrans from the endothelial surface. This non-recovery surgical technique, which requires simultaneous brightfield and fluorescent imaging of the mouse lung, allows for longitudinal observation of the subpleural microvasculature without evidence of inducing confounding lung injury.

In vivo Measurement of the Mouse Pulmonary Endothelial Surface Layer

The endothelial glycocalyx/endothelial surface layer is ideally studied using intravital microscopy. Intravital microscopy is technically challenging in a moving organ such as the lung. We demonstrate how simultaneous brightfield and fluorescent microscopy may be used to estimate endothelial surface layer thickness in a freely-moving in vivo mouse lung.

在体测量小鼠肺血管内皮表面层

The endothelial glycocalyx is a layer of proteoglycans and associated glycosaminoglycans lining the vascular lumen. In vivo, the glycocalyx is highly ...

Slow-release Drug Delivery through Elvax 40W to the Rat Retina: Implications for the Treatment of Chronic Conditions

Diseases of the retina are difficult to treat as the retina lies deep within the eye. Invasive methods of drug delivery are often needed to treat these diseases. Chronic retinal diseases such as retinal oedema or neovascularization usually require multiple intraocular injections to effectively treat the condition. However, the risks associated with these injections increase with repeated delivery of the drug. Therefore, alternative delivery methods need to be established in order to minimize the risks of reinjection. Several other investigations have developed methods to deliver drugs over extended time, through materials capable of releasing chemicals slowly into the eye. In this investigation, we outline the use of Elvax 40W, a copolymer resin, to act as a vehicle for drug delivery to the adult rat retina. The resin is made and loaded with the drug. The drug-resin complex is then implanted into the vitreous cavity, where it will slowly release the drug over time. This method was tested using 2-amino-4-phosphonobutyrate (APB), a glutamate analogue that blocks the light response of the retina. It was demonstrated that the APB was slowly released from the resin, and was able to block the retinal response by 7 days after implantation. This indicates that slow-release drug delivery using this copolymer resin is effective for treating the retina, and could be used therapeutically with further testing.

This paper details how Elvax 40W can be used as a slow-release method for drug delivery to the adult rat retina. The protocol for preparing, loading, and delivering the drug-resin complex to the eye is described.

通过40W的Elvax缓释给药的大鼠视网膜：对慢性疾病的治疗

Diseases of the retina are difficult to treat as the retina lies deep within the eye. Invasive methods of drug delivery are often needed to treat these ...

Measurement of the Pressure-volume Curve in Mouse Lungs

In recent decades the mouse has become the primary animal model of a variety of lung diseases. In models of emphysema or fibrosis, the essential phenotypic changes are best assessed by measurement of the changes in lung elasticity. To best understand specific mechanisms underlying such pathologies in mice, it is essential to make functional measurements that can reflect the developing pathology. Although there are many ways to measure elasticity, the classical method is that of the total lung pressure-volume (PV) curve done over the whole range of lung volumes. This measurement has been made on adult lungs from nearly all mammalian species dating back almost 100 years, and such PV curves also played a major role in the discovery and understanding of the function of pulmonary surfactant in fetal lung development. Unfortunately, such total PV curves have not been widely reported in the mouse, despite the fact that they can provide useful information on the macroscopic effects of structural changes in the lung. Although partial PV curves measuring just the changes in lung volume are sometimes reported, without a measure of absolute volume, the nonlinear nature of the total PV curve makes these partial ones very difficult to interpret. In the present study, we describe a standardized way to measure the total PV curve. We have then tested the ability of these curves to detect changes in mouse lung structure in two common lung pathologies, emphysema and fibrosis. Results showed significant changes in several variables consistent with expected structural changes with these pathologies. This measurement of the lung PV curve in mice thus provides a straightforward means to monitor the progression of the pathophysiologic changes over time and the potential effect of therapeutic procedures.

Here we present a protocol to simply and reliably measure the lung pressure-volume curve in mice, showing that it is sufficiently sensitive to detect phenotypic parenchymal changes in two common lung pathologies, pulmonary fibrosis and emphysema. This metric provides a means to quantify the lung&#8217;s structural changes with developing pathology.

在小鼠肺压力容积曲线的测量

In recent decades the mouse has become the primary animal model of a variety of lung diseases. In models of emphysema or fibrosis, the essential ...

Studying Pancreatic Cancer Stem Cell Characteristics for Developing New Treatment Strategies

Pancreatic ductal adenocarcinoma (PDAC) contains a subset of exclusively tumorigenic cancer stem cells (CSCs) which have been shown to drive tumor initiation, metastasis and resistance to radio- and chemotherapy. Here we describe a specific methodology for culturing primary human pancreatic CSCs as tumor spheres in anchorage-independent conditions. Cells are grown in serum-free, non-adherent conditions in order to enrich for CSCs while their more differentiated progenies do not survive and proliferate during the initial phase following seeding of single cells. This assay can be used to estimate the percentage of CSCs present in a population of tumor cells. Both size (which can range from 35 to 250 micrometers) and number of tumor spheres formed represents CSC activity harbored in either bulk populations of cultured cancer cells or freshly harvested and digested tumors 1,2. Using this assay, we recently found that metformin selectively ablates pancreatic CSCs; a finding that was subsequently further corroborated by demonstrating diminished expression of pluripotency-associated genes/surface markers and reduced in vivo tumorigenicity of metformin-treated cells. As the final step for preclinical development we treated mice bearing established tumors with metformin and found significantly prolonged survival. Clinical studies testing the use of metformin in patients with PDAC are currently underway (e.g., NCT01210911, NCT01167738, and NCT01488552). Mechanistically, we found that metformin induces a fatal energy crisis in CSCs by enhancing reactive oxygen species (ROS) production and reducing mitochondrial transmembrane potential. In contrast, non-CSCs were not eliminated by metformin treatment, but rather underwent reversible cell cycle arrest. Therefore, our study serves as a successful example for the potential of in vitro sphere formation as a screening tool to identify compounds that potentially target CSCs, but this technique will require further in vitro and in vivo validation to eliminate false discoveries.

Pancreatic cancer stem cells (CSCs) can be expanded in vitro using the anchorage-independent sphere culture technique, which represents a powerful tool to study CSC biology and can serve as the first step to develop novel CSC-targeting therapies. Here the methodology for expanding, analyzing and targeting of pancreatic CSCs is provided.

研究胰腺癌干细胞特性开发新的治疗策略

Pancreatic ductal adenocarcinoma (PDAC) contains a subset of exclusively tumorigenic cancer stem cells (CSCs) which have been shown to drive tumor ...

Network Analysis of Foramen Ovale Electrode Recordings in Drug-resistant Temporal Lobe Epilepsy Patients

Approximately 30% of epilepsy patients are refractory to antiepileptic drugs. In these cases, surgery is the only alternative to eliminate/control seizures. However, a significant minority of patients continues to exhibit post-operative seizures, even in those cases in which the suspected source of seizures has been correctly localized and resected. The protocol presented here combines a clinical procedure routinely employed during the pre-operative evaluation of temporal lobe epilepsy (TLE) patients with a novel technique for network analysis. The method allows for the evaluation of the temporal evolution of mesial network parameters. The bilateral insertion of foramen ovale electrodes (FOE) into the ambient cistern simultaneously records electrocortical activity at several mesial areas in the temporal lobe. Furthermore, network methodology applied to the recorded time series tracks the temporal evolution of the mesial networks both interictally and during the seizures. In this way, the presented protocol offers a unique way to visualize and quantify measures that considers the relationships between several mesial areas instead of a single area.

This protocol describes a procedure to track the evolution of mesial network measures in temporal lobe epilepsy (TLE) patients. It is based on the combination of intracranial recordings with a novel numerical technique for data analysis. Specifically, we present a protocol for network analyses of foramen ovale recordings.

在耐药性颞叶癫痫患者卵圆孔电极记录的网络分析

Approximately 30% of epilepsy patients are refractory to antiepileptic drugs. In these cases, surgery is the only alternative to eliminate/control ...

Preparation Steps for Measurement of Reactivity in Mouse Retinal Arterioles Ex Vivo

Vascular insufficiency and alterations in normal retinal perfusion are among the major factors for the pathogenesis of various sight-threatening ocular diseases, such as diabetic retinopathy, hypertensive retinopathy, and possibly glaucoma. Therefore, retinal microvascular preparations are pivotal tools for physiological and pharmacological studies to delineate the underlying pathophysiological mechanisms and to design therapies for the diseases. Despite the wide use of mouse models in ophthalmic research, studies on retinal vascular reactivity are scarce in this species. A major reason for this discrepancy is the challenging isolation procedures owing to the small size of these retinal blood vessels, which is ~ &#8804; 30 &#181;m in luminal diameter. To circumvent the problem of direct isolation of these retinal microvessels for functional studies, we established an isolation and preparation technique that enables ex vivo studies of mouse retinal vasoactivity under near-physiological conditions. Although the present experimental preparations will specifically refer to the mouse retinal arterioles, this methodology can readily be employed to microvessels from rats.

Preparation Steps for Measurement of Reactivity in Mouse Retinal Arterioles Ex Vivo

Many vision-threatening ocular diseases are associated with dysfunctional retinal microvessels. Therefore, the measurement of retinal arteriole responses is important to investigate the underlying pathophysiological mechanisms. This article describes a detailed protocol for mouse retinal arteriole isolation and preparation to assess the effects of vasoactive substances on vascular diameter.

小鼠视网膜微动脉的反应性测定的制备步骤前体

Vascular insufficiency and alterations in normal retinal perfusion are among the major factors for the pathogenesis of various sight-threatening ocular ...

Complete and Partial Aortic Occlusion for the Treatment of Hemorrhagic Shock in Swine

Hemorrhage remains the leading cause of preventable deaths in trauma. Endovascular management of non-compressible torso hemorrhage has been at the forefront of trauma care in recent years. Since complete aortic occlusion presents serious concerns, the concept of partial aortic occlusion has gained a growing attention. Here, we present a large animal model of hemorrhagic shock to investigate the effects of a novel partial aortic balloon occlusion catheter and compare it with a catheter that works on the principles of complete aortic occlusion. Swine are anesthetized and instrumented in order to conduct controlled fixed-volume hemorrhage, and hemodynamic and physiological parameters are monitored. Following hemorrhage, aortic balloon occlusion catheters are inserted and inflated in the supraceliac aorta for 60 min, during which the animals receive whole-blood resuscitation as 20% of the total blood volume (TBV). Following balloon deflation, the animals are monitored in a critical care setting for 4 h, during which they receive fluid resuscitation and vasopressors as needed. The partial aortic balloon occlusion demonstrated improved distal mean arterial pressures (MAPs) during the balloon inflation, decreased markers of ischemia, and decreased fluid resuscitation and vasopressor use. As swine physiology and homeostatic responses following hemorrhage have been well-documented and are like those in humans, a swine hemorrhagic shock model can be used to test various treatment strategies. In addition to treating hemorrhage, aortic balloon occlusion catheters have become popular for their role in cardiac arrest, cardiac and vascular surgery, and other high-risk elective surgical procedures.

Here, we present a protocol demonstrating a hemorrhagic shock model in swine that uses aortic occlusion as a bridge to definitive care in trauma. This model has application in testing a wide range of surgical and pharmacological therapeutic strategies.

完全性和部分主动脉闭塞治疗猪失血性休克

Hemorrhage remains the leading cause of preventable deaths in trauma. Endovascular management of non-compressible torso hemorrhage has been at the ...

The Use of Mixed Reality in Custom-Made Revision Hip Arthroplasty: A First Case Report

The technology of 3D printing and visualization of anatomical structures is rapidly growing in various fields of medicine. A custom-made implant and mixed reality were used to perform complex revision hip arthroplasty in January 2019. The use of mixed reality allowed for a very good visualization of the structures and resulted in precise implant fixation. According to the authors' knowledge, this is the first described case report of the combined use of these two innovations. The diagnosis preceding the qualification for the procedure was the loosening of the left hip's acetabular component. Mixed reality headset and holograms prepared by engineers were used during the surgery. The operation was successful, and it was followed by early verticalization and patient rehabilitation. The team sees opportunities for technology development in joint arthroplasty, trauma, and orthopedic oncology.

A complex revision hip arthroplasty was performed using a custom-made implant and mixed reality technology. According to the authors&#39; knowledge, this is the first report of such procedure described in the literature.

混合现实在定制翻修髋关节置换术中的应用：第一份病例报告

The technology of 3D printing and visualization of anatomical structures is rapidly growing in various fields of medicine. A custom-made implant and mixed ...

Surgical Treatment of an Endolymphatic Sac Tumor

Endolymphatic sac tumors (ELST) are low-grade papillary adenocarcinoma originating from the endolymphatic sac. Usually slow-growing, with local aggressiveness and a low risk of distant metastases, ELST can be sporadic but also frequently associated with von Hippel Lindau disease. The current treatment of ELST is primarily surgical resection. A 55-year-old woman accessed our otologic tertiary level referral center for a sudden worsening of hearing loss in her left ear and vertigo. A magnetic resonance (MRI) and computer tomography scan study subsequently showed a mass in the petrous bone; hence, the presence of an ELST was hypothesized. After embolization of the mass, the patient underwent surgical removal of the lesion. The resection of the mass was done through a translabirinthine approach, with an uneventful procedure. No residual disease remained after surgery. After 24 months of radiologic follow up with MRI, there are no signs of recurrence disease. This paper reports the management of this sporadic ELST, as well as the follow up results, providing clinicians this protocol for the handling of such a challenging otologic skull base surgery and rare disease.

Here, we discuss clinical and radiological features of endolymphatic sac tumors and report the methodology and results of a surgical removal in a case in care at our otologic referral center.