This project was supported by Grant Numbers HL96593 from NIH and D56HP20783 from HRSA/ HHS. Its contents are solely the responsibility of the authors and do not necessarily represent the official views of the NIH or HRSA / HHS.

注意：获取处理用于获得毛细管图像先前已公布的和使用静止数字照相机与对应图像采集和分析的计算机程序来完成11 21本实验利用静止图像进行分析，而不是视频，简化用于分析图像采集。以下描述了一种用于从图像中定量的毛细血管的新技术。 1.图像增强处理<ol><li>获得的数字图像与单色的数码相机。通过取已知长度的对象的图片，例如用相机尺子校准图像以已知尺寸的目的。此过程允许计算机程序，测量和加工后准确地计算毛细管。理想情况下最高的精度，应使用一个手提袋（科学制造的一块玻璃，用蚀刻成它的统治者）。测量在一个1平方毫米的像素数框使用的计算机程序。 注意：关键此协议的重复性和标准化很大程度上取决于1mm 2中框被计数的正确放置。 </li><li>使用对比度增强工具变暗毛细血管，减轻的背景下，这将最大限度地提高毛细血管的可视化。毛细管从背景的初始分化为在后续步骤中的图像的适当的裁切重要。通过调整使用的图像做这个要做到这一点，点击“最适合的直方图。”：捕捉，亮度，图像的直方图，最适合的。 </li><li>作物感兴趣区域（ROI）的毛细管计数为新图像的区域（选择，感兴趣区域）。使用1毫米2中，这是通过校准图像530的像素等于1毫米来确定。检查该裁剪出的图像放置毛细血管袢的顶点在图像的最顶端。 </li><li>拼合因此未来所有的图像调整将均匀地涂在图像图片。要做到这一点点击：进程选项卡，2D过滤器，压平，“光明”，对象宽度BG强度为“75”，适用。 </li><li>提高图像的对比度，因此毛细管最大限度可视化。要做到这一点点击：调整选项卡，显示屏，对比度提升到75。 </li><li>去斑的图片，点击进程选项卡，2D过滤器，去斑，内核尺寸7×7平滑毛细血管的边缘，适用。 </li><li>最终确定的图像对比度使毛细血管是黑色的，背景是白色的。执行此步骤通过调整直方图的“最适合”的模式。 注： 请参考图2代表结果的处理图像应该是什么样子以下步骤的一个例子。 </li></ol> 2.执行毛细管计数/定性测定毛细血管密度<ol><li>每个图像，手动选择使用“目标对象”featur一个明确定义的毛细管中的一个部分E要被识别为对象的程序计算。然后，选择的背景的一小部分，采用了“背景”的功能，作为参考，以需要由所述计数算法被忽略的区域。 注意：这些亮点的组合导致而忽视了背景噪音所有毛细管加以强调。定量（计数）协议使用的计算机的功能来区分基于颜色和形态图像的某些部分。 </li><li>使用计数功能能够即时计算所有毛细管与成像设备的形象。为了避免计算的背景噪声如毛细血管设置计算的对象到5个像素的最小直径。 </li><li>对于每个单独的，计数为3的平均 - 4个图像，以获得一个更可靠的评估。 注： 请参考图3。代表结果的形象应该是什么样子的计算过程中的一个例子。 </li></ol&gt; 3.创建和使用宏来自动图像处理注：为了节省时间，宏可以以一个或多个图像自动执行的过程特定顺序创建。这些序列可以定制，以使该图像修改更快。从本质上说，这些宏记住如何图像进行处理，并迅速和没有用户输入执行所有的步骤。 12毛细管图像执行计数需要20至30分钟之间的这种实验室用宏（2至3每个画面分钟），相对于每个图象约8分钟而不宏。因此，使用宏是比手动处理每个单独的图像3至5倍更有效的。 <ol><li>为了创建宏，选择“记录宏”，并在一个图像执行的步骤和过程所需的，如在步骤1和2如上所述。在此基础上的图像处理步骤进行以供将来参考进行命名宏。当USI纳克未来图片宏，只需单击“运行宏”，程序将自动应用所记录的增强功能需要的图像（S）。 注：本实验使用宏来执行的在几秒钟的方法第1节所有，但其中一个步骤。需要用户输入唯一的步骤是选择在哪里裁剪图像，步骤1.2。 </li></ol>

<ol>
	<li>Gronenschild, E. H., 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=Semi-automatic+assessment+of+skin+capillary+density:+proof+of+principle+and+validation.">Semi-automatic assessment of skin capillary density: proof of principle and validation.</a> Microvasc Res. 90, 192-198 (2013).</li><li>Allen, J., Howell, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microvascular+imaging:+techniques+and+opportunities+for+clinical+physiological+measurements.">Microvascular imaging: techniques and opportunities for clinical physiological measurements.</a> Physiol Meas. 35, R91-R141 (2014).</li><li>Boettcher, M., Gloe, T., de Wit, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Semiautomatic+quantification+of+angiogenesis.">Semiautomatic quantification of angiogenesis.</a> J Surg Res. 162, 132-139 (2010).</li><li>Wild, R., Ramakrishnan, S., Sedgewick, J., Griffioen, A. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Quantitative+assessment+of+angiogenesis+and+tumor+vessel+architecture+by+computer-assisted+digital+image+analysis:+effects+of+VEGF-toxin+conjugate+on+tumor+microvessel+density.">Quantitative assessment of angiogenesis and tumor vessel architecture by computer-assisted digital image analysis: effects of VEGF-toxin conjugate on tumor microvessel density.</a> Microvasc Res. 59, 368-376 (2000).</li><li>Tresadern, P. A., 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=Simulating+nailfold+capillaroscopy+sequences+to+evaluate+algorithms+for+blood+flow+estimation.">Simulating nailfold capillaroscopy sequences to evaluate algorithms for blood flow estimation.</a> Conf Proc IEEE Eng Med Biol Soc. , 2636-2639 (2013).</li><li>Anderson, M. E., 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=Computerized+nailfold+video+capillaroscopy--a+new+tool+for+assessment+of+Raynaud's+phenomenon.">Computerized nailfold video capillaroscopy--a new tool for assessment of Raynaud's phenomenon.</a> J Rheumatol. 32, 841-848 (2005).</li><li>Neubauer-Geryk, J., 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=Decreased+reactivity+of+skin+microcirculation+in+response+to+L-arginine+in+later-onset+type+1+diabetes.">Decreased reactivity of skin microcirculation in response to L-arginine in later-onset type 1 diabetes.</a> Diabetes Care. 36, 950-956 (2013).</li><li>Pazos-Moura, C. C., Moura, E. G., Bouskela, E., Torres-Filho, I. P., Breitenbach, M. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nailfold+capillaroscopy+in+diabetes+mellitus:+morphological+abnormalities+and+relationship+with+microangiopathy.">Nailfold capillaroscopy in diabetes mellitus: morphological abnormalities and relationship with microangiopathy.</a> Braz J Med Biol Res. 20, 777-780 (1987).</li><li>Antonios, T. F., Singer, D. R., Markandu, N. D., Mortimer, P. S., MacGregor, G. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Structural+skin+capillary+rarefaction+in+essential+hypertension.">Structural skin capillary rarefaction in essential hypertension.</a> Hypertension. 33, 998-1001 (1999).</li><li>Kaiser, S. E., Sanjuliani, A. F., Estato, V., Gomes, M. B., Tibirica, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Antihypertensive+treatment+improves+microvascular+rarefaction+and+reactivity+in+low-risk+hypertensive+individuals.">Antihypertensive treatment improves microvascular rarefaction and reactivity in low-risk hypertensive individuals.</a> Microcirculation. 20, 703-716 (2013).</li><li>Cheng, C., Diamond, J. J., Falkner, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Functional+capillary+rarefaction+in+mild+blood+pressure+elevation.">Functional capillary rarefaction in mild blood pressure elevation.</a> Clinical and Translational Science. 1, 75-79 (2008).</li><li>Debbabi, H., Bonnin, P., Levy, B. I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+blood+pressure+control+with+perindopril/indapamide+on+the+microcirculation+in+hypertensive+patients.">Effects of blood pressure control with perindopril/indapamide on the microcirculation in hypertensive patients.</a> Am J Hypertens. 23, 1136-1143 (2010).</li><li>He, F. J., Marciniak, M., Markandu, N. D., Antonios, T. F., MacGregor, G. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+modest+salt+reduction+on+skin+capillary+rarefaction+in+white,+black,+and+Asian+individuals+with+mild+hypertension.">Effect of modest salt reduction on skin capillary rarefaction in white, black, and Asian individuals with mild hypertension.</a> Hypertension. 56, 253-259 (2010).</li><li>Murray, A. K., 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=The+influence+of+measurement+location+on+reliability+of+quantitative+nailfold+videocapillaroscopy+in+patients+with+SSc.">The influence of measurement location on reliability of quantitative nailfold videocapillaroscopy in patients with SSc.</a> Rheumatology (Oxford). 51, 1323-1330 (2012).</li><li>Ingegnoli, F., 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=Feasibility+of+different+capillaroscopic+measures+for+identifying+nailfold+microvascular+alterations.">Feasibility of different capillaroscopic measures for identifying nailfold microvascular alterations.</a> Semin Arthritis Rheum. 38, 289-295 (2009).</li><li>Debbabi, H., 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=Increased+skin+capillary+density+in+treated+essential+hypertensive+patients.">Increased skin capillary density in treated essential hypertensive patients.</a> Am J Hypertens. 19, 477-483 (2006).</li><li>Serne, E. H., 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=Impaired+skin+capillary+recruitment+in+essential+hypertension+is+caused+by+both+functional+and+structural+capillary+rarefaction.">Impaired skin capillary recruitment in essential hypertension is caused by both functional and structural capillary rarefaction.</a> Hypertension. 38, 238-242 (2001).</li><li>Shore, A. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Capillaroscopy+and+the+measurement+of+capillary+pressure.">Capillaroscopy and the measurement of capillary pressure.</a> Br J Clin Pharmacol. 50, 501-513 (2000).</li><li>Rieder, M. J., O'Drobinak, D. M., Greene, A. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+computerized+method+for+determination+of+microvascular+density.">A computerized method for determination of microvascular density.</a> Microvasc Res. 49, 180-189 (1995).</li><li>Vermeulen, P. B., 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=Quantification+of+angiogenesis+in+solid+human+tumours:+an+international+consensus+on+the+methodology+and+criteria+of+evaluation.">Quantification of angiogenesis in solid human tumours: an international consensus on the methodology and criteria of evaluation.</a> Eur J Cancer. 32A, 2474-2484 (1996).</li><li>Cheng, C., Daskalakis, C., Falkner, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Non-invasive+Assessment+of+Microvascular+and+Endothelial+Function.">Non-invasive Assessment of Microvascular and Endothelial Function.</a> Journal of Visualized Experiments. , (2012).</li><li>Antonios, T. F., 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=Maximization+of+skin+capillaries+during+intravital+video-microscopy+in+essential+hypertension:+comparison+between+venous+congestion,+reactive+hyperaemia+and+core+heat+load+tests.">Maximization of skin capillaries during intravital video-microscopy in essential hypertension: comparison between venous congestion, reactive hyperaemia and core heat load tests.</a> Clin Sci (Lond). 97, 523-528 (1999).</li></ol>

The authors have no conflicts of interest.

甲襞capillaroscopy表明承诺作为一个在未来各种肿瘤，心血管疾病，风湿病和疾病临床上的应用程序非常有用的工具。图像采集过程是研究人员之间相当一致的，但目前有多种方法用于图像处理和分析。方法目前包括计算机及人工毛细血管计数。手动计数是有问题的，因为它们是费时，并受到用户的主观性和疲劳。当前的计算机为基础的方法需要用户输入的一个高层次，无论在图像增强处理和量化?...

微血管成像是一个快速增长的领域有许多潜在的临床应用价值。2例如 ，肿瘤学家使用微血管成像，以确定肿瘤血管生成的程度，产生关于肿瘤和洞察力的状态变成可能的治疗选择有价值的信息。3 4然而，甲襞capillaroscopy也许是微血管成像的最具成本效益和广泛适用的形式。研究人员正在利用视频甲襞capillaroscopy研究血流量和调查毛细血管形态。5 6视频和静态图像甲襞capillaroscopy的附属品照顾诊断和治疗雷诺氏现象和各种结缔组织病如系统性硬化症。2 甲襞capillaroscopy具有多种潜在的心血管应用程序。目前的研究使用甲襞capillaroscopy建议该糖尿病类型1和类型2的患者表现出异常毛细管形态的高患病率，但有不变的毛细管密度相比，非糖尿病个体7-8 Capillaroscopy也进行了研究实验中高血压。结构毛细管稀疏导致降低的毛细血管密度已被证明在高血压个体相比，非高血压的个体。9-10相对于这些老年高血压患者（平均年龄40以上）谁展示稀疏结构，最近的研究已经证明，年轻的高血压患者（平均年龄40岁）有没有结构疏松功能稀疏。11这表明，功能性稀薄之前发生，并且随着时间的推移可能会发展为结构疏松。 有趣的是，高血压患者与特定的降压药如培哚普利治疗/吲达帕胺显示规范人毛细血管密度及治疗后血管内皮功能，而与ACE（血管紧张素转化酶）抑制剂或利尿剂治疗维持，尽管可比控制血压低的毛细血管密度。12这表明，一些抗高血压药物可能会恢复正常毛细血管密度扭转毛细管稀疏由高血压引起的。此外，其他研究人员已经表明，在食盐摄入量的减少导致逆转高血压个体功能和结构的毛细管稀薄。13 尽管这种技术的各种潜在的临床应用中，很少有标准化技术定量毛细血管密度的图像。2迄今为止，研究人员已经发现，毛细血管密度的结果是从两帧内观察者和观察者间的角度仅当精确重现的同一地区被每次计数。1,14 15 </sup&gt;值得注意的是，以往的研究人员已经基本上完成毛细血管计数手动使用肉眼，9 16 17 18这是一个缓慢而主观的过程。 标准化，基于计算机算法毛细血管图像的定量理论提供更加高效和可重复的数据分析，用更少的主体性，促进capillaroscopy的临床应用。一些研究人员已经确实使用基于计算机的程序，以从甲襞capillaroscopic图片定量的数据。1,6 19 20然而，只有一个出版迄今为止描述了一种复杂的软件程序的可用的可靠性定量capillaroscopy数据，1，并将该软件是复杂的作为先前在上文所指出的要求来算完全相同的视野。在这里，我们提出了一个简单的，可靠的协议，使用一个标准化的算法允许定量毛细管使用多个视场。使用多个视野，不仅简化了程序，而且还允许正常生物变化的毛细管计数的评估。 这项研究的目的是描述一个基于可重复和高效的计算机算法，它规范了毛细管定量的过程。虽然这些方法都没有完全自动化它们需要非常少的用户输入，并且提供图片的快速，可靠的定量。

<table>
	<tbody>
		<tr>
			<td class="xl65">Image-Pro Premier</td>
			<td class="xl65">Media Cybernetics, Inc</td>
			<td align="right" class="xl63">9.1</td>
			<td class="xl63">Image processing software</td>
		</tr>
	</tbody>
</table>

a reproducible computerized method for quantitation of capillary density using nailfold capillaroscopy

Capillaroscopy为直接可视化的微循环的非侵入性的，高效的，相对便宜和容易学习方法学。的capillaroscopy技术可以提供深入了解患者的微血管健康，导致各种潜在的有价值的皮肤病，眼科，风湿病和心血管临床应用。此外，肿瘤生长可能是依赖于血管生成，这可以通过测量肿瘤内微血管密度进行定量。然而，目前还几乎没有标准化的技术，只有一个出版迄今为止报告的当前可用的，复杂的基于计算机的算法的可靠性为定量capillaroscopy数据。1本文介绍了一种新的，更简单的，可靠的，标准化的毛细管计数算法对于定量甲襞capillaroscopy数据。一个简单的，可重复的电脑capillaroscopy算法，因为这将有利于更多广泛使用，研究人员和临床医生的技术。许多研究者目前正在分析capillaroscopy图像手工，促进成果的使用者的疲劳和主观性。本文介绍了一种新的，易于使用的自动除可重复的，半自动化计数算法的图像处理算法。这个算法使图像的分析中，同时降低主观性分钟;的训练时间仅有极少量（在我们的经验，不到1小时），需要学习的技术。

该图像处理程序的目标是从背景图像区分的毛细血管，这样他们可以被精确定量。这两个不完整的图像处理和过度的图像处理，损害程序的量化毛细血管能力。 正如图3中，不完整的图像处理使得毛细管难以从背景区别开来。该用户能够容易地辨别一个毛细管的边界，因为上述的计数方法取决于用户的准确强调几个毛细管能力是至关重要的。另一方面， 如在?...

Watch this Scientific Journal Video about A Reproducible Computerized Method for Quantitation of Capillary Density using Nailfold Capillaroscopy at JoVE.com

A Reproducible Computerized Method for Quantitation of Capillary Density using Nailfold Capillaroscopy

Capillaroscopy is a non-invasive, efficient, relatively inexpensive and easy-to-learn methodology for directly visualizing capillaries in the microcirculation. However, only one publication to date describes the reliability of a complex software program available for quantitating capillaroscopy data. Here, we present a simple, reliable protocol for quantitating capillaries using a standardized algorithm.

对于毛细血管密度的定量使用甲襞Capillaroscopy可再生的计算机化方法

Capillaroscopy is a non-invasive, efficient, relatively inexpensive and easy to learn methodology for directly visualizing the microcirculation. The ...

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8.9K 次观看.  Thomas Jefferson University. Capillaroscopy is a non-invasive, efficient, relatively inexpensive and easy-to-learn methodology for directly visualizing capillaries in the microcirculation. However, only one publication to date describes the reliability of a complex software program available for quantitating capillaroscopy data. Here, we present a simple, reliable protocol for quantitating capillaries using a standardized algorithm. 

视频: A Reproducible Computerized Method for Quantitation of Capillary Density using Nailfold Capillaroscopy

Photothrombotic Ischemia: A Minimally Invasive and Reproducible Photochemical Cortical Lesion Model for Mouse Stroke Studies

The photothrombotic stroke model aims to induce an ischemic damage within a given cortical area by means of photo-activation of a previously injected light-sensitive dye. Following illumination, the dye is activated and produces singlet oxygen that damages components of endothelial cell membranes, with subsequent platelet aggregation and thrombi formation, which eventually determines the interruption of local blood flow. This approach, initially proposed by Rosenblum and El-Sabban in 1977, was later improved by Watson in 1985 in rat brain and set the basis of the current model. Also, the increased availability of transgenic mouse lines further contributed to raise the interest on the photothrombosis model. Briefly, a photosensitive dye (Rose Bengal) is injected intraperitoneally and enters the blood stream. When illuminated by a cold light source, the dye becomes activated and induces endothelial damage with platelet activation and thrombosis, resulting in local blood flow interruption. The light source can be applied on the intact skull with no need of craniotomy, which allows targeting of any cortical area of interest in a reproducible and non-invasive way. The mouse is then sutured and allowed to wake up. The evaluation of ischemic damage can be quickly accomplished by triphenyl-tetrazolium chloride or cresyl violet staining. This technique produces infarction of small size and well-delimited boundaries, which is highly advantageous for precise cell characterization or functional studies. Furthermore, it is particularly suitable for studying cellular and molecular responses underlying brain plasticity in transgenic mice.

Photothrombosis is a quick, minimally-invasive technique for inducing small and well-delimited infarction in areas of interest in highly reproducible manner. It is particularly suitable for studying cellular and molecular responses underlying brain plasticity in transgenic mice.

光化学缺血：一种微创和可重复性的光化学皮质病变模型小鼠中风研究

The photothrombotic stroke model aims to induce an ischemic damage within a given cortical area by means of photo-activation of a previously injected ...

科研

医学

Generation and 3-Dimensional Quantitation of Arterial Lesions in Mice Using Optical Projection Tomography

The generation and analysis of vascular lesions in appropriate animal models is a cornerstone of research into cardiovascular disease, generating important information on the pathogenesis of lesion formation and the action of novel therapies. Use of atherosclerosis-prone mice, surgical methods of lesion induction, and dietary modification has dramatically improved understanding of the mechanisms that contribute to disease development and the potential of new treatments.
Classically, analysis of lesions is performed ex vivo using 2-dimensional histological techniques. This article describes application of optical projection tomography (OPT) to 3-dimensional quantitation of arterial lesions. As this technique is non-destructive, it can be used as an adjunct to standard histological and immunohistochemical analyses.
Neointimal lesions were induced by wire-insertion or ligation of the mouse femoral artery whilst atherosclerotic lesions were generated by administration of an atherogenic diet to apoE-deficient mice.
Lesions were examined using OPT imaging of autofluorescent emission followed by complementary histological and immunohistochemical analysis. OPT clearly distinguished lesions from the underlying vascular wall. Lesion size was calculated in 2-dimensional sections using planimetry, enabling calculation of lesion volume and maximal cross-sectional area. Data generated using OPT were consistent with measurements obtained using histology, confirming the accuracy of the technique and its potential as a complement (rather than alternative) to traditional methods of analysis.
This work demonstrates the potential of OPT for imaging atherosclerotic and neointimal lesions. It provides a rapid, much needed ex vivo technique for the routine 3-dimensional quantification of vascular remodelling.

Ex vivo analysis of arterial lesions from animal models of cardiovascular disease classically relies on histological and immunohistochemical techniques. These provide 2-dimensional measurements in 3-dimensional lesions. This manuscript describes the generation of arterial lesions for quantitative analysis in 3-dimensions using optical projection tomography.

生成和动脉病变的小鼠的三维定量使用光学投影层析成像

The generation and analysis of vascular lesions in appropriate animal models is a cornerstone of research into cardiovascular disease, generating ...

Computerized Dynamic Posturography for Postural Control Assessment in Patients with Intermittent Claudication

Computerized dynamic posturography with the EquiTest is an objective technique for measuring postural strategies under challenging static and dynamic conditions. As part of a diagnostic assessment, the early detection of postural deficits is important so that appropriate and targeted interventions can be prescribed. The Sensory Organization Test (SOT) on the EquiTest determines an individual&#39;s use of the sensory systems (somatosensory, visual, and vestibular) that are responsible for postural control. Somatosensory and visual input are altered by the calibrated sway-referenced support surface and visual surround, which move in the anterior-posterior direction in response to the individual&#39;s postural sway. This creates a conflicting sensory experience. The Motor Control Test (MCT) challenges postural control by creating unexpected postural disturbances in the form of backwards and forwards translations. The translations are graded in magnitude and the time to recover from the perturbation is computed.
Intermittent claudication, the most common symptom of peripheral arterial disease, is characterized by a cramping pain in the lower limbs and caused by muscle ischemia secondary to reduced blood flow to working muscles during physical exertion. Claudicants often display poor balance, making them susceptible to falls and activity avoidance. The Ankle Brachial Pressure Index (ABPI) is a noninvasive method for indicating the presence of peripheral arterial disease and intermittent claudication, a common symptom in the lower extremities. ABPI is measured as the highest systolic pressure from either the dorsalis pedis or posterior tibial artery divided by the highest brachial artery systolic pressure from either arm. This paper will focus on the use of computerized dynamic posturography in the assessment of balance in claudicants.

Individuals with intermittent claudication exhibit poor balance compared to healthy controls. Computerized dynamic posturography is an objective method for measuring an individual's postural responses to balance disturbances. This provides an objective reflection of the person&#x2019;s ability to respond to situations under which sensory stimuli are altered and unexpected perturbations occur.

Computerized dynamic posturography with the EquiTest is an objective technique for measuring postural strategies under challenging static and dynamic ...

Quantitation of Intra-peritoneal Ovarian Cancer Metastasis

Epithelial ovarian cancer (EOC) is the leading cause of death from gynecologic malignancy in the United States. Mortality is due to diagnosis of 75% of women with late stage disease, when metastasis is already present. EOC is characterized by diffuse and widely disseminated intra-peritoneal metastasis. Cells shed from the primary tumor anchor in the mesothelium that lines the peritoneal cavity as well as in the omentum, resulting in multi-focal metastasis, often in the presence of peritoneal ascites. Efforts in our laboratory are directed at a more detailed understanding of factors that regulate EOC metastatic success. However, quantifying metastatic tumor burden represents a significant technical challenge due to the large number, small size and broad distribution of lesions throughout the peritoneum. Herein we describe a method for analysis of EOC metastasis using cells labeled with red fluorescent protein (RFP) coupled with in vivo multispectral imaging. Following intra-peritoneal injection of RFP-labelled tumor cells, mice are imaged weekly until time of sacrifice. At this time, the peritoneal cavity is surgically exposed and organs are imaged in situ. Dissected organs are then placed on a labeled transparent template and imaged ex vivo. Removal of tissue auto-fluorescence during image processing using multispectral unmixing enables accurate quantitation of relative tumor burden. This method has utility in a variety of applications including therapeutic studies to evaluate compounds that may inhibit metastasis and thereby improve overall survival.

Ovarian cancer metastasis is characterized by numerous diffuse intra-peritoneal lesions, such that accurate visual quantitation of tumor burden is challenging. Herein we describe a method for in situ and ex vivo quantitation of metastatic tumor burden using red fluorescent protein (RFP)-labeled tumor cells and optical imaging.

腹膜内卵巢癌转移的定量

Epithelial ovarian cancer (EOC) is the leading cause of death from gynecologic malignancy in the United States.  Mortality is due to diagnosis of 75% of ...

Method of Direct Segmental Intra-hepatic Delivery Using a Rat Liver Hilar Clamp Model

Major hepatic surgery with inflow occlusion, and liver transplantation, necessitate a period of warm ischemia, and a period of reperfusion leading to ischemia/reperfusion (I/R) injury with myriad negative consequences. Potential I/R injury in marginal organs destined for liver transplantation contributes to the current donor shortage secondary to a decreased organ utilization rate. A significant need exists to explore hepatic I/R injury in order to mediate its impact on graft function in transplantation. Rat liver hilar clamp models are used to investigate the impact of different molecules on hepatic I/R injury. Depending on the model, these molecules have been delivered using inhalation, epidural infusion, intraperitoneal injection, intravenous administration or injection into the peripheral superior mesenteric vein. A rat liver hilar clamp model has been developed for use in studying the impact of pharmacologic molecules in ameliorating I/R injury. The described model for rat liver hilar clamp includes direct cannulation of the portal supply to the ischemic hepatic segment via a side branch of the portal vein, allowing for direct segmental hepatic delivery. Our approach is to induce ischemia in the left lateral and median lobes for 60 min, during which time the substance under study is infused. In this case, pegylated-superoxide dismutase (PEG-SOD), a free radical scavenger, is infused directly into the ischemic segment. This series of experiments demonstrates that infusion of PEG-SOD is protective against hepatic I/R injury. Advantages of this approach include direct injection of the molecule into the ischemic segment with consequent decrease in volume of distribution and reduction in systemic side effects.

A unique rat liver hilar clamp model was developed for studying the impact of pharmacologic molecules in ameliorating ischemia-reperfusion injury. This model includes direct cannulation of the portal supply to the ischemic liver segment via a branch of the portal vein, allowing for direct hepatic delivery.

直接节段性肝内交付使用大鼠肝门部夹具模型方法

Major hepatic surgery with inflow occlusion, and liver transplantation, necessitate a period of warm ischemia, and a period of reperfusion leading to ...

Reproducible Arterial Denudation Injury by Infrarenal Abdominal Aortic Clamping in a Murine Model

Percutaneous vascular interventions uniformly result in arterial denudation injuries that subsequently lead to thrombosis and restenosis. These complications can be attributed to impairments in re-endothelialization within the wound margins. Yet, the cellular and molecular mechanisms of re-endothelialization remain to be defined. While several animal models to study re-endothelialization after arterial denudation are available, few are performed in the mouse because of surgical limitations. This undermines the opportunity to exploit transgenic mouse lines and investigate the contribution of specific genes to the process of re-endothelialization. Here, we present a step-by-step protocol for creating a highly reproducible murine model of arterial denudation injury in the infrarenal abdominal aorta using external vascular clamping. Immunocytochemical staining of injured aortas for fibrinogen and &#946;-catenin demonstrate the exposure of a pro-thrombotic surface and the border of intact endothelium, respectively. The method presented here has the advantages of speed, excellent overall survival rate, and relative technical ease, creating a uniquely practical tool for imposing arterial denudation injury in transgenic mouse models. Using this method, investigators may elucidate the mechanisms of re-endothelialization under normal or pathological conditions.

Understanding the cellular and molecular mechanisms of re-endothelialization following arterial denudation injury is of paramount importance in preventing thrombosis and restenosis of arteries. Here we describe a protocol for reproducible arterial denudation injury of the infrarenal abdominal aorta. The procedure was developed to investigate the underlying mechanisms that regulate endothelial regeneration using mouse models.

重复性的动脉剥脱损伤腹主动脉阻断在小鼠模型

Percutaneous vascular interventions uniformly result in arterial denudation injuries that subsequently lead to thrombosis and restenosis. These ...

A Method for Quantifying Upper Limb Performance in Daily Life Using Accelerometers

A key reason for referral to rehabilitation services after stroke and other neurological conditions is to improve one's ability to function in daily life. It has become important to measure a person's activities in daily life, and not just measure their capacity for activity in the structured environment of a clinic or laboratory. A wearable sensor that is now enabling measurement of daily movement is the accelerometer. Accelerometers are commercially-available devices resembling large wrist watches that can be worn throughout the day. Data from accelerometers can quantify how the limbs are engaged to perform activities in peoples' homes and communities. This report describes a methodology to collect accelerometry data and turn it into clinically-relevant information. First, data are collected by having the participant wear two accelerometers (one on each wrist) for 24 h or longer. The accelerometry data are then downloaded and processed to produce four different variables that describe key aspects of upper limb activity in daily life: hours of use, use ratio, magnitude ratio, and the bilateral magnitude. Density plots can be constructed that visually represent the data from the 24 h wearing period. The variables and their resultant density plots are highly consistent in neurologically-intact, community-dwelling adults. This striking consistency makes them a useful tool for determining if upper limb daily performance is different from normal. This methodology is appropriate for research studies investigating upper limb dysfunction and interventions designed to improve upper limb performance in daily life in people with stroke and other patient populations. Because of its relative simplicity, it may not be long before it is also incorporated in clinical neurorehabilitation practice.

This protocol describes a method to quantify upper limb performance in daily life using wrist-worn accelerometers.

一种使用加速度计量化上肢表现在日常生活的方法

A key reason for referral to rehabilitation services after stroke and other neurological conditions is to improve one's ability to function in daily life. ...

Ablation of Ischemic Ventricular Tachycardia Using a Multipolar Catheter and 3-dimensional Mapping System for High-density Electro-anatomical Reconstruction

Ventricular tachycardia (VT) in patients with ischemic cardiomyopathy mainly results from endocardial scars after myocardial infarction; those scars represent zones of slow conduction that allow the occurrence and maintenance of reentrant circuits. Catheter ablation enables substrate modification of those low voltage areas and thus can help to alter the scar tissue in such a way that arrhythmias cannot appear anymore. Hospitalizations of concerned patients decrease, quality of life and outcome rise. Consequently, VT ablation represents a growing field in electrophysiology, especially for patients with endocardial scars in ischemic heart disease after myocardial infarction. However, ablation of ventricular tachycardia remains one of the most challenging procedures in the electrophysiology lab. Precise scar definition and localization of abnormal potentials are critical for ablation success. The following manuscript describes the use of a multipolar mapping catheter and 3-dimensional (3D) mapping system to create a high density electro-anatomical map of the left ventricle including a precise scar representation as well as mapping of fractionated and late potentials in order to allow a highly accurate substrate modification.

With the following protocol, we provide an approach to Ventricular Tachycardia (VT) ablation using high density mapping with a multipolar catheter and 3D mapping system enhancing the success of the procedure.

采用多极导管三维测图系统进行缺血性室性心动过速的高密度电解剖重建

Ventricular tachycardia (VT) in patients with ischemic cardiomyopathy mainly results from endocardial scars after myocardial infarction; those scars ...

A Computerized Functional Skills Assessment and Training Program Targeting Technology Based Everyday Functional Skills

Today, many functional skills are technology-based, so development of a technology-based training program has broad importance. Here we present a computerized functional skills training program that was paired in half of the participants with a commercially available cognitive training (CCT) program.
Non-impaired older individuals (NC) aged 60+ (n=45) and similarly aged individuals with mild cognitive impairment (MCI; n=50) were randomized to receive 12 weeks of twice-weekly computerized functional skills training (CFST) or 12 weeks of twice-weekly sessions split between CCT and CFST. Skills trained were use of an ATM; internet banking; ticket kiosk; telephone and internet prescription refill; medication management; and internet shopping. As with previous functional capacity assessments, we focus on completion time for each simulation.
51 participants completed the training program, either by mastering all 6 tasks (34) or completing 12 weeks of training. 44 more participants completed 4 or more training sessions so they were also analyzed for improvement up to their last training session. Completion time for all 6 tests significantly improved from the baseline assessment to the final training session in both groups of participants (all p&#60;0.001 with an average improvement in task completion time of 45%). Further, there was no differential improvement in MCI and NC in the 6 tests from baseline to end of training (all t&#60;1.66, all p&#62;0.12). Finally, combined CCT plus CFST did not differ from CSFT alone on any of the percent-change score measures (all t&#60;1.64, all p&#62;0.11).
Both NC and MCI groups evidenced substantial improvements in performance. CCT supplementation led to similar functional gains with half as many training sessions. The NC participants proceeded through the training fairly rapidly even without CCT supplementation; MCI participants required more training but learned equivalently. These findings suggest that even in cases with memory impairments, functional skills can be efficiently learned with training.

This training protocol uses computerized training to teach technology-related everyday functional skills. These skills include financial skills, travel and transit, as well as medication management.

以基于技术的日常功能技能为目标的计算机化功能技能评估和培训计划

Today, many functional skills are technology-based, so development of a technology-based training program has broad importance. Here we present a ...

A Reproducible Intensive Care Unit-Oriented Endotoxin Model in Rats

Sepsis and septic shock remain the leading cause of death in intensive care units. Despite significant improvements in sepsis management, mortality still ranges between 20 and 30%. Novel treatment approaches in order to reduce sepsis-related multiorgan failure and death are urgently needed. Robust animal models allow for one or multiple treatment approaches as well as for testing their effect on physiological and molecular parameters. In this article, a simple animal model is presented.
First, general anesthesia is induced in animals either with the use of volatile or by intraperitoneal anesthesia. After placement of an intravenous catheter (tail vein), tracheostomy, and insertion of an intraarterial catheter (tail artery), mechanical ventilation is started. Baseline values of mean arterial blood pressure, arterial blood oxygen saturation, and heart rate are recorded.
The injection of lipopolysaccharides (1 milligram/kilogram body weight) dissolved in phosphate-buffered saline induces a strong and reproducible inflammatory response via the toll-like receptor 4. Fluid corrections as well as the application of norepinephrine are performed based on well-established protocols.
The animal model presented in this article is easy to learn and strongly oriented towards clinical sepsis treatment in an intensive care unit with sedation, mechanical ventilation, continuous blood pressure monitoring, and repetitive blood sampling. Also, the model is reliable, allowing for reproducible data with a limited number of animals in accordance with the 3R (reduce, replace, refine) principles of animal research. While animal experiments in sepsis research cannot easily replaced, repetitive measurements allow for a reduction of animals and keeping septic animals anesthetized diminishes suffering.

Here, we present a reproducible intensive care unit-oriented endotoxin model in rats.

一种可重复的重症监护病房导向内毒素模型在大鼠中的应用

Sepsis and septic shock remain the leading cause of death in intensive care units. Despite significant improvements in sepsis management, mortality still ...