我们感谢圣保罗研究基金会 （FAPESP） 通过第 2012/02500-8 （RM）、2014/25486-6 （RF） 和 2013/07559-3 号文件提供的支持。资助者在研究设计、数据收集和分析、发表决定或手稿准备方面没有任何作用。

该协议由坎皮纳斯大学地方委员会批准（协议号56602516.2.0000.5404）。在测量之前，从患者或法定代表人那里获得了书面知情同意。我们监测了坎皮纳斯大学诊所医院收治的患者，诊断为缺血性中风或影响前循环的蛛网膜下腔出血。影响后循环的缺血性中风患者，由于颅内压升高而导致减压性颅骨切除术的患者和其他神经退行性疾病（痴呆，帕金森氏症或任何其他可能与皮质萎缩相关的疾病）的患者被排除在研究方案之外。
1. 将系统移至重症监护室之前的准备工作
<ol><li>将所有光纤连接到相关的激光器和检测器，并确保它们正确连接到光学探头（图1B）。</li>
	<li>检查光学探头是否用黑布覆盖，以避免激光照射在房间内。</li>
	<li>将系统电源开关转到"ON"位置。系统通电后，等待30秒，然后将DCS激光钥匙开关转到"ON"位置。FD-DOS激光器在系统通电时自动打开。</li>
	<li>在准备系统时，请征得参与者或法定代表人的同意。获得同意后，将推车带到患者房间。 注意:由于混合动力系统具有可持续使用长达 45 分钟的内置电池，因此在运输过程中无需关闭。</li>
</ol>2. DOS系统的校准和增益设置
<ol><li>到达ICU后，通过将钥匙切换到"OFF"位置来关闭DCS激光器。</li>
	<li>从标有"校准"的固体模型开始，按照以下步骤在FD-DOS软件（BOXY，ISS）上运行校准过程。<ol><li>在"文件"菜单上，通过单击"加载设置文件"选项，为正在使用的探头加载相应的设置文件。</li>
		<li>将探头放在模型的弯曲侧，确保与表面良好接触，然后单击FD-DOS软件中的"优化所有探测器" 按钮来优化PMT偏置电压。</li>
		<li>通过单击"计算波形校准"选项，运行多个源检测器分离的校准。光学道具的值。和"校准" 菜单中的多个距离"。</li>
		<li>从"文本星期一"菜单中打开"用户定义的计算"选项，以检查测量的光学属性是否与预先指定的值（写在实体模型中）匹配，以及拟合 R2 是否接近 1。</li>
	</ol></li>
	<li>重复上述步骤（步骤2.2.3除外）以测量标记为"检查" 的模型的光学特性，以确保校准充分。测量的光学特性应在 10% 以内与模型中指定的值匹配。 注意:确保每次移动探头时都关闭PMT（通过单击"所有探测器关闭"按钮），以避免由于环境光的直接照明而损坏PMT。</li>
	<li>如果校准不充分，请重新运行校准过程（步骤 2.2 和 2.3）。确保FD-DOS系统的良好校准对于FD-DOS测量的有效性至关重要。</li>
</ol>3. 参与者在床边的准备
<ol><li>使用消毒湿巾清洁探头和患者额头。</li>
	<li>将双面胶带放在探头上（图1B），确保胶带不与光纤尖端直接接触。</li>
	<li>在主题上放置激光安全谷歌。</li>
	<li>将探针放在感兴趣区域（ROI）上，并将松紧带缠绕在受试者的头部。虽然FD-DOS和DCS并非绝对必要，但建议用黑布或黑色绷带覆盖光学探头，以减少环境光引起的噪音。 注意:重要的是要确保弹性带既不太紧也不太松。如果带子太紧，可能会给患者带来明显的不适，如果带子太松，可能会导致数据质量差，因为双面带的强度不足以将探头固定到位。</li>
	<li>将探头正确固定在患者的额头上后，通过将钥匙切换到"ON"位置来打开DCS激光器。 注意:DCS系统使用3B类激光，这对眼睛暴露是危险的。只有当探头正确连接到患者的额头上时，才打开激光非常重要。</li>
</ol>4. 数据质量评估
<ol><li>在开始使用GUI获取数据之前，请在GUI的"设置" 选项卡中编写DCS源检测器分离。 注意:DCS系统不需要校准步骤，但实时分析需要正确输入源探测器分离（详见 补充材料 ）。</li>
	<li>按GUI中的"开始" 按钮启动采集软件，并在FD-DOS软件中检查DOS信号:<ol><li>单击FD-DOS软件中的"优化所有检测器" 按钮以优化PMT偏置电压。</li>
		<li>在"文本星期一"菜单中的"用户定义计算"选项中检查光学特性和DOS拟合的R2。R2系数应接近单位，根据经验，人类患者的吸收系数应在0.05和0.2cm-1范围内，而散射系数应在6和13cm-113范围内。</li>
	</ol></li>
	<li>在GUI的"相关曲线"选项卡中检查DCS信号。<ol><li>通过将开关转到"ON" 位置来打开DCS探测器。</li>
		<li>确保每个DCS探测器测量的光强度都足够。根据经验，需要超过 10 kHz。</li>
		<li>如果测量强度高于800 kHz，请使用中性密度滤光片减少光子计数，以免损坏探测器。对于较短（< 1 cm）的源检测器分离，这通常是一个问题。 注意:除了可能损坏DCS探测器外，高于800 kHz的光子计数也可能由于探测器中的非线性效应而带来误差。</li>
		<li>检查自相关曲线以确保良好的皮肤耦合（参见 代表性结果 和 图2）， 并在必要时重新定位光学探头。</li>
		<li>如果在上一步中需要重新定位探头，请重复步骤 4.2 和 4.3。这些步骤可能需要重复多次。 注意:每次移动探头时，必须关闭DCS和FD-DOS探测器。要关闭DCS探测器，请手动将开关移动到"关闭"位置。通过单击FD-DOS软件中的"所有探测器关闭"按钮，将关闭FD-DOS探测器。</li>
	</ol></li>
	<li>当探头和皮肤之间实现良好接触时，通过单击 GUI 中的"停止"按钮停止数据收集。然后，在"文件夹"文本框中设置实验和患者标识符，并在"文件名"文本框中写入ROI名称。</li>
	<li>按GUI中的"开始"按钮开始数据采集。</li>
	<li>在协议要求的情况下，在第一个ROI中收集数据。如有必要，将探头移动到其他ROI并重复测量。 注意:监测期可能因研究目标而异。</li>
</ol>5. 测量过程中对实验者的考虑
<ol><li>开始测量后，在GUI的"实验信息"选项卡中写下相关的患者信息（例如，损伤的类型和位置，正在服用的药物，年龄，性别等）。</li>
	<li>确保通过单击 GUI 上的"标记"按钮来标记在监控期间发生的任何相关事件。每次标记后，请确保在 GUI 的"实验信息"选项卡中编写事件描述。</li>
</ol>6. 停止数据收集
<ol><li>通过按 GUI 中的"停止"按钮停止数据收集。</li>
	<li>通过按下FD-DOS软件中表示为两个红色方块的停止数据采集和记录按钮来停止FD-DOS软件。</li>
	<li>通过将开关翻转到"关闭"位置来关闭DCS探测器，通过将钥匙转到"关闭" 位置来关闭DCS激光器。</li>
	<li>通过单击"所有探测器关闭"按钮关闭FD-DOS模块的PMT。</li>
	<li>从患者头部取下探头，然后从探头上取下双面胶带。然后，用消毒湿巾清洁探头。</li>
	<li>尽快重复测量每个固体模型的光学特性，以确保在整个监测过程中校准保持充分（参见步骤4.2.2）。 注意:理想情况下，校准步骤应在从患者头部取下光学探头后立即完成（步骤6.6）。但是，由于时间问题，在下一节中介绍的示例中，这是在存储设施中完成的。</li>
	<li>用消毒湿巾清洁系统及其附件。</li>
	<li>将推车推回储藏室。</li>
</ol>

作者声明了与本文的研究、作者身份和/或出版有关的以下潜在利益冲突:RC Mesquita 有一项待决专利申请和与这项工作相关的另外两项专利（美国专利 10，342，488 和 10，064，554）。目前没有作者从这些专利中获得版税或付款。

本文提出了一种混合光学系统，该系统可以在旁边提供神经危重症患者的脑血流、脑氧合和脑氧代谢的实时信息。弥漫光学技术的使用以前曾被认为是临床情况下非侵入性床旁监测的潜在标志物。之前的一项研究通过病例报告9重点关注神经ICU住院期间光学监测的临床方面和可行性。这项工作的重点是详细说明与漫反射光学实时监测相关的相关和创新方面。具体来说，本文提出了...

An erratum was issued for:&#160;<a href="https://www.jove.com/t/61608">Real-Time Monitoring of Neurocritical Patients with Diffuse Optical Spectroscopies</a>. The Authors section was updated from:
Rodrigo Menezes Forti1,2 
Marilise Katsurayama2,3 
Lenise Valler2,3 
Andr&#233;s Quiroga1,2 
Luiz Simioni1 
Julien Menko4 
Antonio L. E. Falc&#227;o3 
Li Min Li2,5 
Rickson C. Mesquita1,2 
1Institute of Physics, University of Campinas 
2Brazilian Institute of Neuroscience and Neurotechnology 
3Clinical Hospital, University of Campinas 
4Department of Emergency Medicine, Albert Einstein College of Medicine 
5School of Medical Sciences, University of Campinas
to:
Rodrigo Menezes Forti1,2 
Marilise Katsurayama2,3 
Giovani Grisotti Martins1 
Lenise Valler2,3 
Andr&#233;s Quiroga1,2 
Luiz Simioni1 
Julien Menko4 
Antonio L. E. Falc&#227;o3 
Li Min Li2,5 
Rickson C. Mesquita1,2 
1Institute of Physics, University of Campinas 
2Brazilian Institute of Neuroscience and Neurotechnology 
3Clinical Hospital, University of Campinas 
4Department of Emergency Medicine, Albert Einstein College of Medicine 
5School of Medical Sciences, University of Campinas

An erratum was issued for:&#160;<a href="https://www.jove.com/t/61608">Real-Time Monitoring of Neurocritical Patients with Diffuse Optical Spectroscopies</a>. The Authors section was updated.

Erratum: Real-Time Monitoring of Neurocritical Patients with Diffuse Optical Spectroscopies

导致危重神经系统患者预后不良的大多数并发症与脑血流动力学损伤引起的继发性损伤有关。因此，监测这些患者的脑生理可能直接影响发病率和死亡率1，2，3，4，5，6，7。然而，目前还没有成熟的临床工具用于在床边对神经危重患者的脑生理进行连续实时无创监测。在潜在的候选者中，漫反射光学技术最近被提出作为填补这一空白的有前途的工具8，9，10，11。通过测量来自头皮的漫射近红外光（~650-900 nm）的缓慢变化（即数十到数百毫秒），漫反射光谱（DOS）可以测量大脑中主要发色团的浓度，例如脑氧（HbO）和脱氧血红蛋白（HbR）12，13。此外，可以通过量化光强度的快速波动（即从几μs到几毫秒）来测量脑血流量（CBF）10，14，15，16，17。当结合时，DOS和DCS还可以提供大脑代谢率的估计值（CMRO2）18，19，20。
已经探索了DOS和DCS的组合，以在几种临床前和临床情况下监测患者。例如，漫反射光学已被证明可以为危重新生儿提供相关的临床信息 21，22，23，24，包括在心脏手术期间治疗心脏缺陷 23，25，26，27，28.此外，几位作者还探索了在不同脑血管介入治疗过程中使用漫反射光学来评估脑血流动力学，例如颈动脉内膜切除术29、30、31、中风溶栓治疗32、床头操作33、34、35、心肺复苏36 和其他 37，38，39.当连续血压监测也可用时，漫反射光学器件可用于监测健康和危重受试者的大脑自动调节11，40，41，42，以及评估脑循环的临界闭合压力43。几位作者已经根据不同的金标准CBF测量18验证了DCS的CBF测量结果，而使用漫反射光学器件测量的CMRO2已被证明是神经关键监测的有用参数8，18，23，24，28，43，44，45.此外，先前的研究已经验证了光学衍生的脑血流动力学参数用于长期监测神经危重症患者8，9，10，11，包括用于预测缺氧46，47，48和缺血事件8。
漫反射光学技术在纵向测量和临床干预期间提供有价值的实时信息的可靠性在很大程度上仍未得到解决。以前将独立DOS系统的使用与侵入性脑组织氧张力监测仪进行比较，并且DOS被认为没有足够的敏感性来取代侵入性监测仪。然而，除了使用相对较小的人群外，侵入性和非侵入性监测仪的直接比较可能会被误导，因为每种技术探测包含大脑脉管系统不同部分的不同体积。尽管这些研究最终得出结论，漫反射光学器件不能替代侵入性监视器，但在这两项研究中，DOS都达到了中等到良好的精度，这对于没有侵入性监视器的情况和/或地方可能足够了。
相对于其他方法，漫反射光学器件的主要优点是它能够使用便携式仪器在床边无创（连续）同时测量血流和组织血氧合。与经颅多普勒超声（TCD）相比，DCS还有一个额外的优势:它在组织水平上测量灌注，而TCD测量大脑底部大动脉的脑血流速度。在评估近端大动脉血流和软脑膜侧支均有助于灌注的狭窄闭塞性疾病时，这种区别可能尤为重要。与其他传统成像模式（如正电子发射断层扫描（PET）和磁共振成像（MRI））相比，光学技术也具有优势。除了同时提供CBF和HbO/HbR浓度的直接测量（单独使用MRI或PET无法实现）外，光学监测还提供了明显更好的时间分辨率，例如，允许评估动态脑自动调节40，41，42和评估动态演变的血流动力学变化。此外，与PET和MRI相比，漫射光学仪器价格低廉且便于携带，鉴于中低收入国家血管疾病的高负担，这是一个关键优势。
这里提出的协议是对重症监护病房（ICU）的患者进行实时床边神经监测的环境。该方案使用混合光学设备以及临床友好的图形用户界面（GUI）和定制的光学传感器来探测患者（图1）。用于展示该协议的混合系统结合了来自独立模块的两个漫反射光谱:商用频域（FD-）DOS模块和自制DCS模块（图1A）。FD-DOS 模块49，50 由 4 个光电倍增管 （PMT） 和 32 个以四种不同波长（690、704、750 和 850 nm）发射的激光二极管组成。DCS 模块由一个发射 785 nm 的长相干激光器、16 个作为探测器的单光子计数器和一个相关器板组成。FD-DOS模块的采样频率为10 Hz，DCS模块的最大采样频率为3 Hz。为了集成FD-DOS和DCS模块，在我们的控制软件中编程了一个微控制器，以便在每个模块之间自动切换。微控制器负责打开和关闭FD-DOS和DCS激光器，以及FD-DOS探测器，以允许对每个模块进行交错测量。总的来说，所提出的系统可以每0.5至5秒收集一个FD-DOS和DCS组合样本，具体取决于信噪比（SNR）要求（收集时间越长，SNR越好）。为了将光耦合到额头，我们开发了一种3D打印的光学探头，可以为每个患者定制（图1B），源检测器的间隔在0.8到4.0厘米之间变化。此处示例中使用的标准源检测器分离对于DCS为2.5 cm，对于FD-DOS为1.5、2.0、2.5和3.0 cm。
本研究中提出的协议的主要特征是开发了一个实时界面，既可以使用友好的GUI控制硬件，又可以在不同的时间窗口下实时显示主要的大脑生理参数（图1C）。在所提出的GUI中开发的实时分析管道速度快，计算光学参数所需的时间不到50 ms（有关更多详细信息，请参阅 补充材料 ）。GUI的灵感来自神经ICU现有的临床仪器，并在将系统转换为神经ICU期间，通过临床用户的广泛反馈进行了调整。因此，实时GUI可以促进常规医院工作人员（如神经重症监护医生和护士）采用光学系统。漫反射光学作为临床研究工具的广泛采用有可能增强其监测生理上有意义的数据的能力，并最终证明漫反射光学是实时无创监测神经关键患者的良好选择。

<table>
	<tbody>
		<tr>
			<td>3D Printer</td>
			<td>Sethi3D</td>
			<td>S2</td>
			<td>3D-printer used to print the customizable probes</td>
		</tr>
		<tr>
			<td>Arduino UNO</td>
			<td>Arduino</td>
			<td>UNO REV3</td>
			<td>Microcontroller responsible to interleave the DCS and FD-DOS measurements</td>
		</tr>
		<tr>
			<td>DCS Correlator</td>
			<td>Correlator.com</td>
			<td>Flex11-16ch</td>
			<td>Component of the DCS module</td>
		</tr>
		<tr>
			<td>DCS Dectectors IO Boards</td>
			<td>Excelitas Technology</td>
			<td>SPCM-AQ4C-IO</td>
			<td>Component of the DCS module</td>
		</tr>
		<tr>
			<td>DCS Detectors</td>
			<td>Excelitas Technology</td>
			<td>SPCM-AQ4C</td>
			<td>Component of the DCS module</td>
		</tr>
		<tr>
			<td>DCS Laser</td>
			<td>CrystaLaser</td>
			<td>DL785-120-SO</td>
			<td>Component of the DCS module</td>
		</tr>
		<tr>
			<td>DCS Power supply</td>
			<td>Artesyn</td>
			<td>UMP10T-S2A-S2A-S2A-S2A-IES-00-A</td>
			<td>Component of the DCS module (power supply for the DCS detecto; 2, 5 and 30V)</td>
		</tr>
		<tr>
			<td>FD-DOS fibers</td>
			<td>ISS</td>
			<td>Imagent supplies</td>
			<td>The fibers used for FD-DOS detection and illumination are provived by ISS</td>
		</tr>
		<tr>
			<td>Flexible 3D printer material</td>
			<td>Sethi3D</td>
			<td>NinjaFlex</td>
			<td>Material used to print the flexible customizable probes</td>
		</tr>
		<tr>
			<td>Imagent</td>
			<td>ISS</td>
			<td>Imagent</td>
			<td>FD-DOS module</td>
		</tr>
		<tr>
			<td>Laser safety googles</td>
			<td>Thorlabs</td>
			<td>LG9</td>
			<td></td>
		</tr>
		<tr>
			<td>Multi-mode fiber</td>
			<td>Thorlabs</td>
			<td>FT400EMT</td>
			<td>Multi-mode fiber used for DCS illumination</td>
		</tr>
		<tr>
			<td>Neutral density filter 1.0 OD</td>
			<td>Edmund Optics</td>
			<td>53-705</td>
			<td>Neutral density filter for the short source detector separations</td>
		</tr>
		<tr>
			<td>Single-mode optical fiber</td>
			<td>Thorlabs</td>
			<td>780HP</td>
			<td>Single-mode optical fiber used for the DCS detectors</td>
		</tr>
		<tr>
			<td>System battery</td>
			<td>SMS</td>
			<td>NET4</td>
			<td>System battery used for transportation</td>
		</tr>
	</tbody>
</table>

real-time monitoring of neurocritical patients with diffuse optical spectroscopies

神经生理学监测是神经危重症患者治疗的一个重要目标，因为它可以预防继发性损伤并直接影响发病率和死亡率。然而，目前缺乏合适的非侵入性实时技术来连续监测床边的脑生理。弥漫光学技术已被提出作为神经危重患者床旁测量脑血流量和脑氧合的潜在工具。以前已经探索过漫反射光谱法，以监测从新生儿监测到成人脑血管介入治疗等多种临床场景的患者。然而，该技术通过在床边提供实时信息来帮助临床医生的可行性在很大程度上仍未得到解决。在这里，我们报告了漫反射光学系统的翻译，用于在重症监护期间连续实时监测脑血流量，脑氧合和脑氧代谢。该仪器的实时功能可以实现基于患者特定脑生理学的治疗策略，而不是依赖于替代指标，例如动脉血压。通过使用相对便宜和便携式的仪器在不同时间尺度上提供有关脑循环的实时信息，这种方法在低预算医院、偏远地区和开放领域（例如国防和体育）的监测中可能特别有用。

理想情况下，使用DCS模块获得的归一化自相关曲线在零延迟时间外推（使用单模光纤14时）应约为1.5，并且在较长的延迟时间下，曲线应衰减至1。曲线应该是平滑的，对于较长的源-检测器分离，它应该具有更快的衰减。 图2A显示了良好自相关的示例。 图2B 显示了不良自相关曲线的示例;在本例中，无法区分不同源检测器分离的曲线。 ...

Watch this Scientific Journal Video about Real-Time Monitoring of Neurocritical Patients with Diffuse Optical Spectroscopies at JoVE.com

Real-Time Monitoring of Neurocritical Patients with Diffuse Optical Spectroscopies

这里介绍的是一种方案，用于使用漫反射光学器件实时和在床边实时无创监测神经危重患者的脑血流动力学。具体而言，该协议使用混合漫反射光学系统来检测和显示有关脑氧合，脑血流和脑代谢的实时信息。

使用漫反射光谱实时监测神经危重症患者

Neurophysiological monitoring is an important goal in the treatment of neurocritical patients, as it may prevent secondary damage and directly impact ...

real-time-monitoring-neurocritical-patients-with-diffuse-optical

Research

JoVE Journal

Medicine

2.0K 次观看.  University of Campinas. 这里介绍的是一种方案，用于使用漫反射光学器件实时和在床边实时无创监测神经危重患者的脑血流动力学。具体而言，该协议使用混合漫反射光学系统来检测和显示有关脑氧合，脑血流和脑代谢的实时信息。

视频: Real-Time Monitoring of Neurocritical Patients with Diffuse Optical Spectroscopies

Movement Retraining using Real-time Feedback of Performance

Any modification of movement - especially movement patterns that have been honed over a number of years - requires re-organization of the neuromuscular patterns responsible for governing the movement performance. This motor learning can be enhanced through a number of methods that are utilized in research and clinical settings alike. In general, verbal feedback of performance in real-time or knowledge of results following movement is commonly used clinically as a preliminary means of instilling motor learning. Depending on patient preference and learning style, visual feedback (e.g. through use of a mirror or different types of video) or proprioceptive guidance utilizing therapist touch, are used to supplement verbal instructions from the therapist. Indeed, a combination of these forms of feedback is commonplace in the clinical setting to facilitate motor learning and optimize outcomes.
Laboratory-based, quantitative motion analysis has been a mainstay in research settings to provide accurate and objective analysis of a variety of movements in healthy and injured populations. While the actual mechanisms of capturing the movements may differ, all current motion analysis systems rely on the ability to track the movement of body segments and joints and to use established equations of motion to quantify key movement patterns. Due to limitations in acquisition and processing speed, analysis and description of the movements has traditionally occurred offline after completion of a given testing session.
This paper will highlight a new supplement to standard motion analysis techniques that relies on the near instantaneous assessment and quantification of movement patterns and the display of specific movement characteristics to the patient during a movement analysis session. As a result, this novel technique can provide a new method of feedback delivery that has advantages over currently used feedback methods.

Retraining abnormal movement patterns following injury or disease is a key component of physical rehabilitation. Recent advances in technology have permitted accurate assessment of movement during a variety of tasks, with near instantaneous quantification of results. This provides new opportunities for modification of faulty movement patterns in real time.

的运动再培训采用实时反馈的性能

Any modification of movement - especially movement patterns that have been honed over a number of years - requires re-organization of the neuromuscular ...

科研

医学

Cerebrospinal Fluid MicroRNA Profiling Using Quantitative Real Time PCR

MicroRNAs (miRNAs) constitute a potent layer of gene regulation by guiding RISC to target sites located on mRNAs and, consequently, by modulating their translational repression. Changes in miRNA expression have been shown to be involved in the development of all major complex diseases. Furthermore, recent findings showed that miRNAs can be secreted to the extracellular environment and enter the bloodstream and other body fluids where they can circulate with high stability. The function of such circulating miRNAs remains largely elusive, but systematic high throughput approaches, such as miRNA profiling arrays, have lead to the identification of miRNA signatures in several pathological conditions, including neurodegenerative disorders and several types of cancers. In this context, the identification of miRNA expression profile in the cerebrospinal fluid, as reported in our recent study, makes miRNAs attractive candidates for biomarker analysis.
There are several tools available for profiling microRNAs, such as microarrays, quantitative real-time PCR (qPCR), and deep sequencing. Here, we describe a sensitive method to profile microRNAs in cerebrospinal fluids by quantitative real-time PCR. We used the Exiqon microRNA ready-to-use PCR human panels I and II V2.R, which allows detection of 742 unique human microRNAs. We performed the arrays in triplicate runs and we processed and analyzed data using the GenEx Professional 5 software.
Using this protocol, we have successfully profiled microRNAs in various types of cell lines and primary cells, CSF, plasma, and formalin-fixed paraffin-embedded tissues.

We describe a protocol of real time PCR to profile microRNAs in the cerebrospinal fluid (CSF). With the exception of RNA extraction protocols, the procedure can be extended to RNA extracted from other body fluids, cultured cells, or tissue specimens.

脑脊液的microRNA分析采用实时定量PCR技术

MicroRNAs (miRNAs) constitute a potent layer of gene regulation by guiding RISC to target sites located on mRNAs and, consequently, by modulating their ...

Real-time Cytotoxicity Assays in Human Whole Blood

A live cell-based whole blood cytotoxicity assay (WCA) that allows access to temporal information of the overall cell cytotoxicity is developed with high-throughput cell positioning technology. The targeted tumor cell populations are first preprogrammed to immobilization into an array format, and labeled with green fluorescent cytosolic dyes. Following the cell array formation, antibody drugs are added in combination with human whole blood. Propidium iodide (PI) is then added to assess cell death. The cell array is analyzed with an automatic imaging system. While cytosolic dye labels the targeted tumor cell populations, PI labels the dead tumor cell populations. Thus, the percentage of target cancer cell killing can be quantified by calculating the number of surviving targeted cells to the number of dead targeted cells. With this method, researchers are able to access time-dependent and dose-dependent cell cytotoxicity information. Remarkably, no hazardous radiochemicals are used. The WCA presented here has been tested with lymphoma, leukemia, and solid tumor cell lines. Therefore, WCA allows researchers to assess drug efficacy in a highly relevant ex&#160;vivo condition.

The whole blood cytotoxicity assay (WCA) is a cytotoxicity assay developed by incorporating high-throughput cell positioning technology with fluorescence microscopy and automated image processing. Here, we describe how lymphoma cells treated with an anti-CD20 antibody can be analyzed real-time in human whole blood to provide quantitative cellular cytotoxicity analysis.

实时的细胞毒性分析在人全血

A live cell-based whole blood cytotoxicity assay (WCA) that allows access to temporal information of the overall cell cytotoxicity is developed with ...

Diffuse Optical Spectroscopy for the Quantitative Assessment of Acute Ionizing Radiation Induced Skin Toxicity Using a Mouse Model

Acute skin toxicities from ionizing radiation (IR) are a common side effect from therapeutic courses of external beam radiation therapy (RT) and negatively impact patient quality of life and long term survival. Advances in the understanding of the biological pathways associated with normal tissue toxicities have allowed for the development of interventional drugs, however, current response studies are limited by a lack of quantitative metrics for assessing the severity of skin reactions. Here we present a diffuse optical spectroscopic (DOS) approach that provides quantitative optical biomarkers of skin response to radiation. We describe the instrumentation design of the DOS system as well as the inversion algorithm for extracting the optical parameters. Finally, to demonstrate clinical utility, we present representative data from a pre-clinical mouse model of radiation induced erythema and compare the results with a commonly employed visual scoring. The described DOS method offers an objective, high through-put evaluation of skin toxicity via functional response that is translatable to the clinical setting.

We present a diffuse optical spectroscopic (DOS) approach that provides quantitative optical biomarkers of skin response to radiation. We describe DOS instrumentation design, optical parameters extraction algorithms and the animal handling procedures required to yield representative data from a pre-clinical mouse model of radiation induced erythema.

弥漫性光学光谱治疗急性电离辐射引起的皮肤毒性的定量评价使用小鼠模型

Acute skin toxicities from ionizing radiation (IR) are a common side effect from therapeutic courses of external beam radiation therapy (RT) and ...

Uncontrolled Hemorrhagic Shock Modeled via Liver Laceration in Mice with Real Time Hemodynamic Monitoring

Uncontrolled hemorrhage is an important cause of preventable deaths among trauma patients. We have developed a murine model of uncontrolled hemorrhage via a liver laceration that results in consistent blood loss, hemodynamic alterations, and survival.
Mice undergo a standardized resection of the left-middle lobe of the liver. They are allowed to bleed without mechanical intervention. Hemostatic agents can be administered as pre-treatment or rescue therapy depending on the interest of the investigator. During the time of hemorrhage, real-time hemodynamic monitoring via a left femoral arterial line is performed. Mice are then sacrificed, blood loss is quantified, blood is collected for further analysis, and organs are harvested for analysis of injury. Experimental design is described to allow for simultaneous testing of multiple animals.
Liver hemorrhage as a model of uncontrolled hemorrhage exists in the literature, primarily in rat and porcine models. Some of these models utilize hemodynamic monitoring or quantify blood loss but lack consistency. The present model incorporates quantification of blood loss, real-time hemodynamic monitoring in a murine model that offers the advantage of using transgenic lines and a high-throughput mechanism to further investigate the pathophysiologic mechanisms in uncontrolled hemorrhage.

Uncontrolled hemorrhage, an important cause of mortality among trauma patients, can be modeled using a standard liver laceration in a murine model. This model results in consistent blood loss, survival, and allows for testing hemostatic agents. This article provides the step-by-step process to perform this valuable model.

通过实时血液动力学监测在小鼠中通过肝脏浸润建立的不受控制的出血性休克

Uncontrolled hemorrhage is an important cause of preventable deaths among trauma patients. We have developed a murine model of uncontrolled hemorrhage via ...

Integration of Brain Tissue Saturation Monitoring in Cardiopulmonary Exercise Testing in Patients with Heart Failure

Cerebral hypo-oxygenation during rest or exercise negatively impacts the exercise capacity of patients with heart failure with reduced ejection fraction (HF). However, in clinical cardiopulmonary exercise testing (CPET), cerebral hemodynamics is not assessed. NIRS is used to measure cerebral tissue oxygen saturation (SctO2) in the frontal lobe. This method is reliable and valid and has been utilized in several studies. SctO2 is lower during both rest and peak exercise in patients with HF than in healthy controls (66.3 &#177; 13.3% and 63.4 &#177; 13.8% vs. 73.1 &#177; 2.8% and 72 &#177; 3.2%). SctO2 at rest is significantly linearly correlated with peak VO2 (r = 0.602), oxygen uptake efficiency slope (r = 0.501), and brain natriuretic peptide (r = -0.492), all of which are recognized prognostic and disease severity markers, indicating its potential prognostic value. SctO2 is determined mainly by end-tidal CO2 pressure, mean arterial pressure, and hemoglobin in the HF population. This article demonstrates a protocol that integrates SctO2 using NIRS into incremental CPET on a calibrated bicycle ergometer.

This protocol integrated near-infrared spectroscopy into conventional cardiopulmonary exercise testing to identify the involvement of the cerebral hemodynamic response in exercise intolerance in patients with heart failure.

心力衰竭患者心肺运动测试中脑组织饱和监测的集成

Cerebral hypo-oxygenation during rest or exercise negatively impacts the exercise capacity of patients with heart failure with reduced ejection fraction ...

Digital Home-Monitoring of Patients after Kidney Transplantation: The MACCS Platform

The MACCS (Medical Assistant for Chronic Care Service) platform enables secure sharing of key medical information between patients after kidney transplantation and physicians. Patients provide information such as vital signs, well-being, and medication intake via smartphone apps. The information is transferred directly into a database and electronic health record at the kidney transplant center, which is used for routine patient care and research. Physicians can send an updated medication plan and laboratory data directly to the patient app via this secure platform. Other features of the app are medical messages and video consultations. Consequently, the patient is better-informed, and self-management is facilitated. In addition, the transplant center and the patient's local nephrologist automatically exchange notes, medical reports, laboratory values, and medication data via the platform. A telemedicine team reviews all incoming data on a dashboard and takes action, if necessary. Tools to identify patients at risk for complications are under development. The platform exchanges data via a standardized secure interface (Health Level 7 (HL7), Fast Healthcare Interoperability Resources (FHIR)). The standardized data exchange based on HL7 FHIR guarantees interoperability with other eHealth solutions and allows rapid scalability to other chronic diseases. The underlying data protection concept is in concordance with the latest European General Data Protection Regulation. Enrollment started in February 2020, and 131 kidney transplant recipients are actively participating as of July 2020. Two large German health insurance companies are currently funding the telemedicine services of the project. The deployment for other chronic kidney diseases and solid organ transplant recipients is planned. In conclusion, the platform is designed to enable home monitoring and automatic data exchange, empower patients, reduce hospitalizations, and improve adherence, and outcomes after kidney transplantation.

The MACCS platform&#160;is a comprehensive telemedicine concept aiming at better outcomes after kidney transplantation by sharing key medical information between patients and physicians. A telemedicine team reviews incoming data to detect potential complications and to improve adherence in kidney transplant recipients to achieve better long-term outcomes.

数字家庭-肾移植后患者的监测:MACCS平台

The MACCS (Medical Assistant for Chronic Care Service) platform enables secure sharing of key medical information between patients after kidney ...

Real-Time Electrocardiogram Monitoring During Treadmill Training in Mice

Regular physical exercise is a major contributor to cardiovascular health, influencing various metabolic as well as electrophysiological processes. However, in certain cardiac diseases such as inherited arrhythmia syndromes, e.g., arrhythmogenic cardiomyopathy (ACM) or myocarditis, physical exercise may have negative effects on the heart leading to a proarrhythmogenic substrate production. Currently, the underlying molecular mechanisms of exercise-related proarrhythmogenic remodeling are largely unknown, thus it remains unclear which frequency, duration, and intensity of exercise can be considered safe in the context of disease(s).
The proposed method allows to study proarrhythmic/antiarrhythmic effects of physical exercise by combining treadmill training with real-time monitoring of the ECG. Implantable telemetry devices are used to continuously record the ECG of freely moving mice over a period of up to 3 months both at rest and during treadmill training. Data acquisition software with its analysis modules is used to analyze basic ECG parameters such as heart rate, P wave duration, PR interval, QRS interval, or QT duration at rest, during and after training. Furthermore, heart rate variability (HRV) parameters and occurrence of arrhythmias are evaluated. In brief, this manuscript describes a step-by-step approach to experimentally explore exercise induced effects on cardiac electrophysiology, including potential proarrhythmogenic remodeling in mouse models.

Electrocardiogram (ECG) is the key variable to understanding cardiac electrophysiology. Physical exercise has beneficial effects but may also be harmful in the context of cardiovascular diseases. This manuscript provides a method of recording real-time ECG during exercise, which can serve to investigate its effects on cardiac electrophysiology in mice.

小鼠跑步机训练期间的实时心电图监测

Regular physical exercise is a major contributor to cardiovascular health, influencing various metabolic as well as electrophysiological processes. ...

Real-Time Monitoring and Modulation of Blood Pressure in a Rabbit Model of Ischemic Stroke

Control of blood pressure, in terms of both absolute values and its variability, affects outcomes in ischemic stroke patients. However, it remains challenging to identify the mechanisms that lead to poor outcomes or evaluate measures by which these effects can be mitigated because of the prohibitive limitations inherent to human data. In such cases, animal models can be utilized to conduct rigorous and reproducible evaluations of diseases. Here we report refinement of a previously described model of ischemic stroke in rabbits that is augmented with continuous blood pressure recording to assess the impacts of modulation on blood pressure. Under general anesthesia, femoral arteries are exposed through surgical cutdowns to place arterial sheaths bilaterally. Under fluoroscopic visualization and roadmap guidance, a microcatheter is advanced into an artery of the posterior circulation of the brain. An angiogram is performed by injecting the contralateral vertebral artery to confirm occlusion of the target artery. With the occlusive catheter remaining in position for a fixed duration, blood pressure is continuously recorded to allow for tight titration of blood pressure manipulations, whether through mechanical or pharmacological means. At the completion of the occlusion interval, the microcatheter is removed, and the animal is maintained under general anesthesia for a prescribed length of reperfusion. For acute studies, the animal is then euthanized and decapitated. The brain is harvested and processed to measure the infarct volume under light microscopy and further assessed with various histopathological stains or spatial transcriptomic analysis. This protocol provides a reproducible model that can be utilized for more thorough preclinical studies on the effects of blood pressure parameters during ischemic stroke. It also facilitates effective preclinical evaluation of novel neuroprotective interventions that might improve care for ischemic stroke patients.

Continuous arterial blood pressure recording allows the investigation of impacts of various hemodynamic parameters. This report demonstrates the application of continuous arterial blood pressure monitoring in a large animal model of ischemic stroke for determination of stroke pathophysiology, impact of different hemodynamic factors, and the assessment of novel treatment approaches.

缺血性脑卒中兔模型中血压的实时监测和调节

Control of blood pressure, in terms of both absolute values and its variability, affects outcomes in ischemic stroke patients. However, it remains ...

Non-Invasive Monitoring of Microvascular Oxygenation and Reactive Hyperemia using Hybrid, Near-Infrared Diffuse Optical Spectroscopy for Critical Care

The detection of levels of impairment in microvascular oxygen consumption and reactive hyperemia is vital in critical care. However, there are no practical means for a robust and quantitative evaluation. This paper describes a protocol to evaluate these impairments using a hybrid near-infrared diffuse optical device. The device contains modules for near-infrared time-resolved and diffuse correlation spectroscopies and pulse-oximetry. These modules allow the non-invasive, continuous, and real-time measurement of the absolute, microvascular blood/tissue oxygen saturation (StO2) and the blood flow index (BFI) along with the peripheral arterial oxygen saturation (SpO2). This device uses an integrated, computer-controlled tourniquet system to execute a standardized protocol with optical data acquisition from the brachioradialis muscle. The standardized vascular occlusion test (VOT) takes care of the variations in the occlusion duration and pressure reported in the literature, while the automation minimizes inter-operator differences. The protocol we describe focuses on a 3-min occlusion period but the details described in this paper can readily be adapted to other durations and cuff pressures, as well as other muscles. The inclusion of an extended baseline and post-occlusion recovery period measurement allows the quantification of the baseline values for all the parameters and the blood/tissue deoxygenation rate that corresponds to the metabolic rate of oxygen consumption. Once the cuff is released, we characterize the tissue reoxygenation rate, magnitude, and duration of the hyperemic response in BFI and StO2. These latter parameters correspond to the quantification of the reactive hyperemia, which provides information about the endothelial function. Furthermore, the above-mentioned measurements of the absolute concentration of oxygenated and deoxygenated hemoglobin, BFI, the derived metabolic rate of oxygen consumption, StO2, and SpO2 provide a yet-to-be-explored rich data set that can exhibit disease severity, personalized therapeutics, and management interventions.

We describe a protocol to non-invasively and continuously measure absolute microvascular blood flow index and blood oxygen saturation using a multi-modal device based on near-infrared diffuse optics. We then evaluate the metabolic rate of oxygen consumption and reactive hyperemia utilizing a vascular occlusion test.