The authors thank Perkin Elmer Company for supporting this publication. We also thank Alana Ross, B.S. for proofreading this manuscript.

注：所有的动物研究应在批准的机构的协议和动物保健的准​​则来进行。 1 在体内的CLI成像英美烟草公司与18F-FDG 1.1背景影像： <ol><li>前18 F-FDG注射液，放置4裸鼠中已连接与氧异氟烷平衡5分钟，以诱导麻醉的感应腔室。然后把四个麻醉小鼠成成像装置。 </li><li>获得具有下列参数的背景图像：打开过滤器中，f = 1，仓= 8，FOV = D和曝光时间= 120秒，段温度= 37℃。 </li></ol> 1.2 英美烟草成像与18架F-FGD： <ol><li>注射麻醉小鼠280微居里的18 F-FDG在PBS静脉内的尾部静脉。注射后，返回小鼠配备了食物和水的笼子里。 注意：这是neces萨里市静脉注射18F-FDG实现英美烟草周围地区一个体面的对比。只有非常弱的对比可以看出与18 F-FDG的相同量如果腹膜内注射使用。 </li><li>在30，60取得CLI图象及后18 F-FDG注射使用相同的参数作为背景图像（步骤1.1.2）120分钟。对于每个成像会议，允许5分钟的麻醉再诱导。 </li><li>量化的信噪比（S / N），使用图像处理软件接口，以得出两个同等大小的椭圆形的ROI在肩胛间BAT和一个区域相邻的BAT（参照区域）（ 图1b）。 </li></ol> 1.3验证CLI的来源： <ol><li>麻醉四只老鼠，并与280微居里的18F-FDG静脉注射的小鼠。 </li><li>通过注射戊巴比妥钠（200毫克/公斤，腹腔注射）牺牲注射18 F-FDG后的小鼠60分钟。小心地取出滑雪N从肩胛间区。图像中的所有小鼠具有相同的参数如上述（1.1.2），在相同的时间（ 图2a）。 </li><li>小心地取出肩胛间白色脂肪组织（WAT）和BAT，然后像解剖小鼠相同的参数（ 图2b）。 </li><li> R（BAT）=（CLI中-CLI设定二）（BAT）/（CLI中-CLI设定二）（BAT-除去），其中，ROI：从CLI的图像，通过使用两个ROI下面的公式计算出从BAT的贡献a是肩胛间区和ROI b为基准区域（ 图2b）。 </li></ol> 2，英美烟草成像中的应用 2.1监测激活与东北<ol><li>分裸鼠分成两组（每组4个）。 </li><li>注1组腹腔注射去甲肾上腺素（NE）（50微升，10毫）。使用第二组作为非激活控制手机升。 30分钟后，麻醉两组用异氟烷进行5分钟，然后静脉注射18 F-FDG（220微居里）注射各小鼠。 注意：腹膜内注射的NE应在30分钟前18 F-FDG注射，以避免显着搅动的小鼠。 </li><li>图像使用相同的参数作为上述协议后18 F-FDG注射的小鼠60分钟。 </li></ol> 2.2监控激活在冷暴露<ol><li>测量人体温度的小鼠在冷室中以直肠温度计。温度应在约30℃。 </li><li>图像后18 F-FDG注射的小鼠60分钟，通过使用相同的摄像参数作为上述协议。使用小鼠被保持在室温（25℃）为对照，并且图象它们具有相同的。 </li><li>对于感冒暴露研究，将小鼠在冷室（4℃）4小时前18F-FDG注入和返回米冰给每个成像会议后冷室和麻醉恢复后。 </li><li>如上面的参数。 </li></ol>英美烟草公司在长期的麻醉监测2.3停用<ol><li>对于长麻（60 - 70分钟），氯胺酮/甲苯​​噻嗪腹腔注射小鼠，使他们在麻醉状态下进行60 - 在室温下70分钟。 </li><li>一旦将小鼠麻醉，注射18 F-FDG（220微居里）通过尾静脉静脉内给药。 </li><li>图像后18 F-FDG注射的小鼠60分钟，通过使用相同的参数，如上述的协议。 </li></ol> 3，混合光谱和多光谱切伦科夫荧光断层成像研究<ol><li>麻醉小鼠5分钟，然后注入300微居里18F-FDG静脉注射。 F = 1，箱= 16，采集：在18F-FDG注射从动物的背侧使用以下参数获得多光谱影像60分钟每个过滤器，发射器= 580，600，620，640，660和680 nm的时间为300秒。 </li><li>进行光谱分离与生活影像4.3.1软件，并选择两个组件（英美烟草公司和非特异性信号）和自动解混（ 图4）。 </li><li>据报道Kuo 等 28,43的方法进行三维重建。从结构光成像结合的切伦科夫发光光谱成漫射光的传播模型，应用Tikhonov正则化中的NNLS（非负的最小平方）的残差优化，并产生表面断层扫描（这是用于3D图像配准）（ 图5中 ）。 注意：对于多光谱CLI光谱分离和断层，至少需要5张不同的过滤器。 </li></ol>

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W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Three-dimensional+reconstruction+of+in+vivo+bioluminescent+sources+based+on+multispectral+imaging.">Three-dimensional reconstruction of in vivo bioluminescent sources based on multispectral imaging.</a> J. Biomed. Optics. 12, 024007 (2007).</li><li>Xu, X., 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=Exercise+ameliorates+high-fat+diet-induced+metabolic+and+vascular+dysfunction,+and+increases+adipocyte+progenitor+cell+population+in+brown+adipose+tissue.">Exercise ameliorates high-fat diet-induced metabolic and vascular dysfunction, and increases adipocyte progenitor cell population in brown adipose tissue.</a> Amer. J. Physiol. Regulatory, integrative and comparative physiology. 300, 1115-1125 (2011).</li></ol>

公布此视频文章是由Perkin Elmer公司的支持。

研究与英美烟草公司已经进行了几十年。此前，它已被认为具有人的成年期1中没有显著的生理意义。然而，最近的大规模临床PET成像与18 F-FDG和其它研究已经证实了BAT中仍存在的上胸部，颈部和成人2,3等位置。最近的研究强烈暗示BAT起着肥胖和糖尿病2,6中起重要作用。其它研究也表明，BAT中老化12的过程中，它的活性可通过锻炼10,44增强起着重要的作用。 无论是在人类临床研究和临床前研究，PET显像与18F-FDG是研究英美烟草公司最常用的方法。然而，对于临床前动物研究中，PET通常比光学成像昂贵得多。在这个协议中，我们证明了发光我跟18F-FDG可应用于小动物光学成像最佳可行技术。像其他的光学成像技术，组织穿透限制和对纵深目标的低敏感性的CLI 25,32的内在局限性。尽管如此，最近Spinelli 等。证明体面CLI信号可以观察到10 mm的组织深度用32 P 28,43，和托雷克等 。表明16毫米渗透可在患者的淋巴结后18F-FDG注入39，40来实现。相比于PET成像，CLI可以用相对低的成本的成像系统来执行。最近，托雷克等人证明了显著高灵敏度可以使用CLI来实现的18 F-FDG作为低量的患者39，节点积累（大约2 Cl）40。 体外试验表明，CLI信号从0.01次90Y是检测溶液中的25,32,33 </s起来&gt;。此外，CLI还具有高空间分辨率和高通量筛选的能力的潜力。此外，CLI是很容易学习和使用。 在实验过程中，我们发现BAT的18 F-FDG的CLI对比度强烈相关的注射方法。静脉注射18F-FDG可提供出色的对比度间BAT，而腹腔注射的18F-FDG相同金额仅表现在英美烟草地区弱对比度。 光谱分离，一个非常实用的技术来分离信号两套，目前已广泛应用于荧光成像。这是众所周知的18 F-FDG的摄取没有高度靶向特异性的，并且不同目标/组织中具有不同的光衰减特性。我们发现BAT的CLI光谱的峰是大约640纳米，并且该数据反映BAT的实际环境。三维重建这个公关otocol是非常可操作的，因为它可以与基于不同波长的光漫射特性的市售光学成像系统中进行。通过这种方法，我们就可以避免使用，需要多视角图像的三维重建一个专门的成像系统。 对于这两种光谱分离和三维重建，从每个图像更大最低600光子计数英美烟草公司是必需的。为此，大量的分级（分级= 16），小F停止中（f = 1）和一个长的采集时间（5分钟），需要针对每个图像。 综上所述，利用英美烟草公司的独特位置和形状与18F-FDG在英美烟草公司的显著高摄取，我们将演示如何英美烟草小动物可以通过CLI技术进行光学成像与18F-FDG。该方法可以可靠地用于成像的BAT和监测BAT激活。此外，我们还表明，光谱分离和3D RECOnstruction切实可行和三维体积定量可以为今后的研究。

褐色脂肪组织（BAT）是用于产热的哺乳动物一种特殊的组织，并且其主要功能之一是通过消耗大量的化学/食品能量作为热量1保持整个身体的能量平衡。英美烟草公司的最独特的特征包括丰富的解偶联蛋白1（UCP-1）的表达，丰富的小油滴，在单个细胞中大量的线粒体，并显著血管的组织2-5。这些独特的功能强烈的代谢和能量消耗组织的重要作用相关联。英美烟草以前被认为是不再存在，并拥有成人1人无显著生理功能，但最近的PET / CT显像的调查清楚地表明，英美烟草公司还提出了在人类成人2,3,6-9。 （BMI）英美烟草质量和身体质量指数呈负相关关系，从一些研究确定，并重新分研究表明，体育锻炼可以增加英美烟草公司的质量。这些结果有力地表明，BAT的功能障碍是紧紧与肥胖症和糖尿病2,6,10,11的病状相关联。此外，越来越多的证据表明，BAT的功能密切相关的各种其他病症，例如神经变性疾病和癌症3,7,12,13。 BAT激活，进程，以增加产热，可以在各种条件如冷曝光，锻炼和药物治疗和基因操作1,14,15下才能实现。冷曝光和去甲肾上腺素的治疗是最常用的方法来激活BAT。冷，这可以通过多种机制来感测诸如在皮肤thermoreceptors，刺激交感神经和导致的去甲肾上腺素（NE）的释放，BAT。所释放的NE触发UCP-1来初始化产热以保持正常体温。在这种conditi上，葡萄糖的摄取也增加，以提供增加的代谢多种碳源中的BAT 1,16,17。 PET显像与18F-FDG已经确认标记葡萄糖的摄取人体研究6寒冷的条件下增加。 在光学成像方面，英美烟草公司是一个理想的目标。该间BAT对小鼠独特的地理位置，位于距较大的器官，如肝脏，心脏和胃。因此，从这些大器官信号干扰忽略不计（ 图1a）。与此同时，肩胛间BAT的浅位置，让更多的信号由检测照相机来捕获。此外，英美烟草公司是一家集中质量的器官，它地限制了光信号在某些领域。此外，英美烟草公司的独特的三角物理形状可以很容易地从其他组织（ 图1a）区别开来。 切伦科夫冷光成像（CLI），新​​发生的摩尔ecular成像技术18-26，线束从+和所产生的发光-衰变的放射性核素如18 F和131 I在该介质中。带电粒子（例如+和- ）极化分子，而行进的介质18-20，并且当偏振分子松弛回到平衡状态的发光/光被发射。所发射的荧光被称为切伦科夫发光（CL）。 CL的独特光谱特性包括在整个紫外（UV）的宽光谱和可见光谱18-20，并且其强度和波长（λ2）的平方之间的逆相关。所发射的光的UV和可见光范围内可用于不同的应用。切伦科夫发光的紫外部分已用于体内光活化笼萤光素21的，而在较长波长发射的光可以是用于体内光学成像18,27-31。 对于小动物研究中，CLI成像的光学成像系统的速度更快，比PET更高的成本效益。此外，CLI可以用于高通量筛选与具有高吞吐能力的成像系统。这种技术的优点和缺点在一些评论25,32,33进行了讨论。 CLI的3D断层已被广泛研究的几组28,34-37和CLI的内窥镜成像和术中成像的应用已经成功地证明了在小鼠和30,38。此外，斯皮内利和托雷克等人已经证明，CLI成像可应用于人类受试者，从而该技术还具有潜在的临床应用39，40。 在英美烟草在人类重新发现的过程中，PET图像清楚地表明，一个显著量18F-FDG在一定的条件下2,3,6英美烟草积累。此外，PET成像的小鼠也无疑表明，英美烟草公司可以与18F-FDG 41 42突出。在这份报告中，我们将演示如何切伦科夫冷光从18F-FDG发射可以使用光学成像系统被用于成像英美烟草的小动物。我们的方法提供了一种快速，廉价和方便的英美烟草成像小动物的方法。这种技术可以用作用于PET成像与18 F-FDG的替代方法，尤其是对于实验室无PET设施。

<table border="0" cellpadding="0" cellspacing="0" width="885">
	<tbody>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Name of Material/ Equipment</td>
			<td style="width:71px;">Company</td>
			<td style="width:119px;">Catalog Number</td>
			<td style="width:479px;">Comments/Description</td>
		</tr>
		<tr height="231">
			<td height="231" style="height:231px;width:216px;">Optical Imaging system</td>
			<td style="width:71px;">Perkin Elmer</td>
			<td style="width:119px;"></td>
			<td style="width:479px;">IVIS Spectrum is an optical imaging system that is equiped with very sensitive camera for Cerenkov Luminescence. Some instrumental information is listed below: CCD Sensor: Back thinned, back illuminated 
			CCD Size: 2.7cm x 2.7cm 
			Pixels: 2048 x 2048 
			Quantum Efficiency: 85% 
			Min detectable photons: 70 photons/s/sr/cm &#60; sup &#62; 2 &#60; /sup &#62; 
			Dark Current: &#60; 100 electrons/s/cm &#60; sup &#62; 2 &#60; /sup &#62; 
			Lens: f 0.95 50mm 
			CCD Cooling: Cooled to-90 degree.&#160;</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">18F-FDG</td>
			<td style="width:71px;">IBA Molecular</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">norepinephrine</td>
			<td style="width:71px;">Sigma</td>
			<td>N5785-250MG</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Ketamine/Xylazine</td>
			<td style="width:71px;">Sigma</td>
			<td>K113-10ML</td>
			<td></td>
		</tr>
	</tbody>
</table>

cerenkov luminescence imaging of interscapular brown adipose tissue

Brown adipose tissue (BAT), widely known as a &#8220;good fat&#8221; plays pivotal roles for thermogenesis in mammals. This special tissue is closely related to metabolism and energy expenditure, and its dysfunction is one important contributor for obesity and diabetes. Contrary to previous belief, recent PET/CT imaging studies indicated the BAT depots are still present in human adults. PET imaging clearly shows that BAT has considerably high uptake of 18F-FDG under certain conditions. In this video report, we demonstrate that Cerenkov luminescence imaging (CLI) with 18F-FDG can be used to optically image BAT in small animals. BAT activation is observed after intraperitoneal injection of norepinephrine (NE) and cold treatment, and depression of BAT is induced by long anesthesia. Using multiple-filter Cerenkov luminescence imaging, spectral unmixing and 3D imaging reconstruction are demonstrated. Our results suggest that CLI with 18F-FDG is a practical technique for imaging BAT in small animals, and this technique can be used as a cheap, fast, and alternative imaging tool for BAT research.

在图1中 ，肩胛间BAT（ 图1a）被高亮显示在所有时间点（30，60，120分钟），BAT和参考区域之间的对比度，可容易地观察到（ 图1b）。值得注意的是，蝙蝠的图像的轮廓贴切地反映其外表，它有一个三角形的轮廓。 BAT和参考区域间的信号比分别为2.37，2.49，和2.53倍，在30，60，和注射（ 图1a）后120分钟。 从阶梯状切除的实验中，在肩胛间部位的信号的85％的源自BAT（ 图2）。然而，仍然靠近BAT，这可能源自于BAT残留组织和包括血管和肌肉等邻近组织的上边缘部分的残余信号。 据报道，BAT激活可以通过完成去甲肾上腺素（NE）的治疗和冷暴露3,6。首先，我们观察到BAT活化与网元中的同一组的小鼠有和没有NE的治疗下短（5分钟），异氟醚麻醉。该NE处理的条件（1.23倍），比没有NE处理（ 图3a）下的数据表明显著更高的CLI信号。 在人类研究中，PET显像表明，在冷暴露者有更高的18F-FDG摄取的英美烟草公司比在室温6。在这种小鼠的研究中，18架F-FDG摄取增加了39％，观察冷曝光（ 图3b）治疗的动物的最佳可行技术。 在小鼠中，英美烟草公司的活动可以由不同的麻醉的影响服法3,41。例如，18 F-FDG摄取BAT可以显麻醉一小时后下降用氯胺酮41。比较18F-FDG uptakE在小鼠下短（5分钟，用异氟烷）和长麻醉（70分钟用氯胺酮/甲苯噻嗪）的方案，减少了18 F-FDG的BAT摄取54％同一组观察用氯胺酮/甲苯噻嗪的组中（ 图3c）。 这是众所周知的血红素含有蛋白质（如血液中的血红蛋白和细胞色素c的释放）可导致显著的光吸收，并且因此预 ​​计在血管丰富的BAT 1将改变CLI的光谱，和频谱从英美烟草区域形状会与其他地区有所不同。可以想象，光谱分离技术，可以让我们分开的两个命令行谱。从图4a中 ，没有特定的区域被从组分＃1（UNMIX＃1），以及相应的光谱（蓝线在图4d）的未混合的图像突出显示非常相似的18 F在纯介质的发射光谱，其中，所述的CLI的IntensiTY是负相关的波长18,20的平方。该数据表明，UNMIX＃1表示的非特异性的CLI信号，这可能是从非常浅的深度，例如18 F-FDG蓄积在皮肤上。未混合＃2光谱（红色线）的峰值大约为640毫微米，表明该信号是从BAT。有趣的是，相反的CLI短于640纳米的强度的显著衰减，强度没有显着降低，观察，一旦它达到峰值（红线图4），这表明在较长的波长所发射的光的更好的组织渗透。 多光谱切伦科夫发光体层摄影术（msCLT）被用于3D重建。 msCLT先前已经报道了Kuo 等。28,43，和从一组与多个窄的带通滤波器（&gt; 5过滤器）获取的二维平面图像的构造。重建的三维图像coregistERED用从结构光产生表面成像，图像显示有关的CLI的信号的主要部分源自间BAT（ 图5）。值得注意的是，在冠状图像（ 图5a）清楚地概述了两个凸起BAT的，这非常类似于在图5e所示的BAT的三角形轮廓。 <img alt="图1" fo:content-width="5in" src="/files/ftp_upload/51790/51790fig1highres.jpg" width="500" /> 图1：间BAT（蓝色箭头）在小鼠（一）三角轮廓; （左）BAT覆盖有白色脂肪组织，和（右）BAT被暴露。在图30（b）的CLI鼠标图像和后18F-FDG（280微居里）静脉注射60分钟。图像清晰地勾勒英美烟草的轮廓。 （三）由INTE的CLI信号的定量分析rscapular英美烟草区和参考区域。从基准42再版。 <a href="https://www.jove.com/files/ftp_upload/51790/51790fig1highres.jpg" target="_blank">请点击这里查看该图的放大版本。</a> <img alt="图2" fo:content-width="5in" src="/files/ftp_upload/51790/51790fig2highres.jpg" width="500" /> 图2：（一）小鼠前（左）后（右）英美烟草去除代表性的图像。 （二）定量分析表明&gt; 85％的CLI源自BAT。从基准42再版。 <a href="https://www.jove.com/files/ftp_upload/51790/51790fig2highres.jpg" target="_blank">请点击这里查看该图的放大版本。</a> <img alt="图3" fo:content-width="5in" src="/files/ftp_upload/51790/51790fig3highres.jpg" width="500" /> 图3：（一）小鼠英美烟草公司的代表与CLI（左）和无下短异氟醚麻醉（右）东北刺激的图像。 （b）该CLI信号从（a）中的2组（n = 4每组）的定量分析。 （三）小鼠英美烟草公司的代表与CLI（左）和无（右）在短异氟醚麻醉寒冷刺激的图像。 （四）从（e）中所示的两组CLI的信号的定量分析（n = 3时- 4每个组）。 （五）后18 F-FDG注射下短的异氟醚麻醉（5分钟）代表的CLI BAT的图像，在60分钟（左）和氯胺酮/甲苯噻嗪麻醉的（70分钟）（右）。 （F）从（g）中所示的两组CLI的信号的定量分析（n = 4时为每个组）。从参考42。转载<a href="https://www.jove.com/files/ftp_upload/51790/51790fig3highres.jpg" target="_blank">认罪</a>SE点击这里查看该图的放大版本。 <img alt="图4" fo:content-width="5in" src="/files/ftp_upload/51790/51790fig4highres.jpg" width="500" /> 图4：混合光谱的非特异性的CLI和BAT CLI（一）UNMIX＃1表明，从18 F-FDG的非特异性的CLI信号被分布在整个身体（未混合的频谱为这个图像示于（d）（蓝线））。 （二）UNMIX＃2表明，大多数的CLI在肩胛间部位是从BAT（非混合光谱示于（d）（红色线））。 CLI的峰值大约为640纳米。 （ 三）UNMIX＃1和＃2合并后的图像。 （ 四）UNMIX＃1和＃2的CLI谱。从基准42再版。 <a href="https://www.jove.com/files/ftp_upload/51790/51790fig4highres.jpg" target="_blank">请点击这里查看该图的放大版本。</a> &lt;/ P&gt; <img alt="图5" fo:content-width="5in" src="/files/ftp_upload/51790/51790fig5highres.jpg" width="500" /> 图5：多光谱切伦科夫冷光成像 （A - D）的图像的三维重建。间BAT可以在日冕（一），矢状位（B）中可以看出，和横向（C）的意见，以及在3D图像（D）; （五）身体英美烟草形状如图（蓝色箭头），这与相关的重建图像。转载和引用42相适应。 <a href="https://www.jove.com/files/ftp_upload/51790/51790fig5highres.jpg" target="_blank">请点击这里查看该图的放大版本。</a>

Watch this Scientific Journal Video about Cerenkov Luminescence Imaging of Interscapular Brown Adipose Tissue at JoVE.com

Cerenkov Luminescence Imaging of Interscapular Brown Adipose Tissue

In this video report, we show the application of Cerenkov Luminescence Imaging (CLI) for interscapular brown adipose tissue in mice under activated and depressed conditions.

肩胛间褐色脂肪组织的切伦科夫发光成像

Brown adipose tissue (BAT), widely known as a &#8220;good fat&#8221; plays pivotal roles for thermogenesis in mammals. This special tissue is closely ...

cerenkov-luminescence-imaging-of-interscapular-brown-adipose-tissue

Research

JoVE Journal

Medicine

12.3K 次观看.  Massachusetts General Hospital/Harvard Medical School. In this video report, we show the application of Cerenkov Luminescence Imaging (CLI) for interscapular brown adipose tissue in mice under activated and depressed conditions.

视频: Cerenkov Luminescence Imaging of Interscapular Brown Adipose Tissue

Cerenkov Luminescence Imaging (CLI) for Cancer Therapy Monitoring

In molecular imaging, positron emission tomography (PET) and optical imaging (OI) are two of the most important and thus most widely used modalities1-3. PET is characterized by its excellent sensitivity and quantification ability while OI is notable for non-radiation, relative low cost, short scanning time, high throughput, and wide availability to basic researchers. However, both modalities have their shortcomings as well. PET suffers from poor spatial resolution and high cost, while OI is mostly limited to preclinical applications because of its limited tissue penetration along with prominent scattering optical signals through the thickness of living tissues.
 Recently a bridge between PET and OI has emerged with the discovery of Cerenkov Luminescence Imaging (CLI)4-6. CLI is a new imaging modality that harnesses Cerenkov Radiation (CR) to image radionuclides with OI instruments. Russian Nobel laureate Alekseyevich Cerenkov and his colleagues originally discovered CR in 1934. It is a form of electromagnetic radiation emitted when a charged particle travels at a superluminal speed in a dielectric medium7,8. The charged particle, whether positron or electron, perturbs the electromagnetic field of the medium by displacing the electrons in its atoms. After passing of the disruption photons are emitted as the displaced electrons return to the ground state. For instance, one 18F decay was estimated to produce an average of 3 photons in water5. 
 Since its emergence, CLI has been investigated for its use in a variety of preclinical applications including in vivo tumor imaging, reporter gene imaging, radiotracer development, multimodality imaging, among others4,5,9,10,11. The most important reason why CLI has enjoyed much success so far is that this new technology takes advantage of the low cost and wide availability of OI to image radionuclides, which used to be imaged only by more expensive and less available nuclear imaging modalities such as PET.
 Here, we present the method of using CLI to monitor cancer drug therapy. Our group has recently investigated this new application and validated its feasibility by a proof-of-concept study12. We demonstrated that CLI and PET exhibited excellent correlations across different tumor xenografts and imaging probes. This is consistent with the overarching principle of CR that CLI essentially visualizes the same radionuclides as PET. We selected Bevacizumab (Avastin; Genentech/Roche) as our therapeutic agent because it is a well-known angiogenesis inhibitor13,14. Maturation of this technology in the near future can be envisioned to have a significant impact on preclinical drug development, screening, as well as therapy monitoring of patients receiving treatments.

Cerenkov Luminescence Imaging CLI for Cancer Therapy Monitoring

Use of Cerenkov Luminescence Imaging (CLI) for monitoring preclinical cancer treatment is described here. This method takes advantage of Cerenkov Radiation (CR) and optical imaging (OI) to visualize radiolabeled probes and thus provides an alternative to PET in preclinical therapeutic monitoring and drug screening.

癌症治疗监测的切伦科夫发光成像（CLI）

In  molecular imaging, positron emission tomography (PET) and optical imaging (OI)  are two of the most important and thus most widely used modalities1-3. ...

科研

医学

Human Brown Adipose Tissue Depots Automatically Segmented by Positron Emission Tomography/Computed Tomography and Registered Magnetic Resonance Images

Reliably differentiating brown adipose tissue (BAT) from other tissues using a non-invasive imaging method is an important step toward studying BAT in humans. Detecting BAT is typically confirmed by the uptake of the injected radioactive tracer 18F-Fluorodeoxyglucose (18F-FDG) into adipose tissue depots, as measured by positron emission tomography/computed tomography (PET-CT) scans after exposing the subject to cold stimulus. Fat-water separated magnetic resonance imaging (MRI) has the ability to distinguish BAT without the use of a radioactive tracer. To date, MRI of BAT in adult humans has not been co-registered with cold-activated PET-CT. Therefore, this protocol uses 18F-FDG PET-CT scans to automatically generate a BAT mask, which is then applied to co-registered MRI scans of the same subject. This approach enables measurement of quantitative MRI properties of BAT without manual segmentation. BAT masks are created from two PET-CT scans: after exposure for 2 hr to either thermoneutral (TN) (24 &#176;C) or cold-activated (CA) (17 &#176;C) conditions. The TN and CA PET-CT scans are registered, and the PET standardized uptake and CT Hounsfield values are used to create a mask containing only BAT. CA and TN MRI scans are also acquired on the same subject and registered to the PET-CT scans in order to establish quantitative MRI properties within the automatically defined BAT mask. An advantage of this approach is that the segmentation is completely automated and is based on widely accepted methods for identification of activated BAT (PET-CT). The quantitative MRI properties of BAT established using this protocol can serve as the basis for an MRI-only BAT examination that avoids the radiation associated with PET-CT.

The method presented here uses 18F-Fluorodeoxyglucose (18F-FDG) positron emission tomography/computed tomography (PET-CT) and fat-water separated magnetic resonance imaging (MRI), each scanned following 2 hr exposure to thermoneutral (24 &#176;C) and cold conditions (17 &#176;C) in order to map brown adipose tissue (BAT) in adult human subjects.

人类的褐色脂肪组织的仓库的正电子发射断层扫描/计算机断层扫描自动分段及注册磁共振图像

Reliably differentiating brown adipose tissue (BAT) from other tissues using a non-invasive imaging method is an important step toward studying BAT in ...

Murine Isolated Heart Model of Myocardial Stunning Associated with Cardioplegic Arrest

The following protocol is of use to evaluate impaired cardiac function or myocardial stunning following moderate ischemic insults. The technique is useful for modeling ischemic injury associated with numerous clinically relevant phenomenon including cardiac surgery with cardioplegic arrest and cardiopulmonary bypass, off-pump CABG, transplant, angina, brief ischemia, etc. The protocol presents a general method to model hypothermic hyperkalemic cardioplegic arrest and reperfusion in rodent hearts focusing on measurement of myocardial contractile function. In brief, a mouse heart is perfused in langendorff mode, instrumented with an intraventricular balloon, and baseline cardiac functional parameters are recorded. Following stabilization, the heart is then subject to brief infusion of a cardioprotective hypothermic cardioplegia solution to initiate diastolic arrest. Cardioplegia is delivered intermittently over 2 hr. The heart is then reperfused and warmed to normothermic temperatures and recovery of myocardial function is monitored. Use of this protocol results in reliable depressed cardiac contractile function free from gross myocardial tissue damage in rodents.

The goal of this protocol to assess myocardial stunning following ischemic cardioplegic arrest in rodents.

心肌顿抑相关的小鼠离体心脏模型心脏停搏

The following protocol is of use to evaluate impaired cardiac function or myocardial stunning following moderate ischemic insults. The technique is useful ...

Encapsulation Thermogenic Preadipocytes for Transplantation into Adipose Tissue Depots

Cell encapsulation was developed to entrap viable cells within semi-permeable membranes. The engrafted encapsulated cells can exchange low molecular weight metabolites in tissues of the treated host to achieve long-term survival. The semipermeable membrane allows engrafted encapsulated cells to avoid rejection by the immune system. The encapsulation procedure was designed to enable a controlled release of bioactive compounds, such as insulin, other hormones, and cytokines. Here we describe a method for encapsulation of catabolic cells, which consume lipids for heat production and energy dissipation (thermogenesis) in the intra-abdominal adipose tissue of obese mice. Encapsulation of thermogenic catabolic cells may be potentially applicable to the prevention and treatment of obesity and type 2 diabetes. Another potential application of catabolic cells may include detoxification from alcohols or other toxic metabolites and environmental pollutants.

Here, we present a protocol for encapsulation of catabolic cells, which consume lipids for heat production in intra-abdominal adipose tissue and increase energy dissipation in obese mice.

封装产热前脂肪细胞移植到脂肪组织仓库

Cell encapsulation was developed to entrap viable cells within semi-permeable membranes. The engrafted encapsulated cells can exchange low molecular ...

Assessment of Human Adipose Tissue Microvascular Function Using Videomicroscopy

While obesity is closely linked to the development of metabolic and cardiovascular disease, little is known about mechanisms that govern these processes. It is hypothesized that pro-atherogenic mediators released from fat tissues particularly in association with central/visceral adiposity may promote pathogenic vascular changes locally and systemically, and the notion that cardiovascular disease may be the consequence of adipose tissue dysfunction continues to evolve. Here, we describe a unique method of videomicroscopy that involves analysis of vasodilator and vasoconstrictor responses of intact small human arterioles removed from the adipose depot of living human subjects. Videomicroscopy is used to examine functional properties of isolated microvessels in response to pharmacological or physiological stimuli using a pressured system that mimics in vivo conditions. The technique is a useful approach to gain understanding of the pathophysiology and molecular mechanisms that contribute to vascular dysfunction locally within the adipose tissue milieu. Moreover, abnormalities in the adipose tissue microvasculature have also been linked with systemic diseases. We applied this technique to examine depot-specific vascular responses in obese subjects. We assessed endothelium-dependent vasodilation to both increased flow and acetylcholine in adipose arterioles (50 - 350 &#181;m internal diameter, 2 - 3 mm in length) isolated from two different adipose depots during bariatric surgery from the same individual. We demonstrated that arterioles from visceral fat exhibit impaired endothelium-dependent vasodilation compared to vessels isolated from the subcutaneous depot. The findings suggest that the visceral microenvironment is associated with vascular endothelial dysfunction which may be relevant to clinical observation linking increased visceral adiposity to systemic disease mechanisms. The videomicroscopy technique can be used to examine vascular phenotypes from different fat depots as well as compare findings across individuals with different degrees of obesity and metabolic dysfunction. The method can also be used to examine vascular responses longitudinally in response to clinical interventions.

Videomicroscopy systems are used to examine functional properties of isolated adipose tissue arterioles in response to physiological and pharmacological stimuli. This technique can be used to examine microvascular phenotypes in different adipose tissue domains in obese humans.

应用 Videomicroscopy 评价人脂肪组织微血管功能

While obesity is closely linked to the development of metabolic and cardiovascular disease, little is known about mechanisms that govern these processes. ...

Characterization of Immune Cells in Human Adipose Tissue by Using Flow Cytometry

Infiltration of immune cells in the subcutaneous and visceral adipose tissue (AT) deposits leads to a low-grade inflammation contributing to the development of obesity-associated complications such as type 2 diabetes. To quantitatively and qualitatively investigate the immune cell subsets in human AT deposits, we have developed a flow cytometry approach. The stromal vascular fraction (SVF), containing the immune cells, is isolated from subcutaneous and visceral AT biopsies by collagenase digestion. Adipocytes are removed after centrifugation. The SVF cells are stained for multiple membrane-bound markers selected to differentiate between immune cell subsets and analyzed using flow cytometry. As a result of this approach, pro- and anti-inflammatory macrophage subsets, dendritic cells (DCs), B-cells, CD4+ and CD8+ T-cells, and NK cells can be detected and quantified. This method gives detailed information about immune cells in AT and the amount of each specific subset. Since there are numerous fluorescent antibodies available, our flow cytometry approach can be adjusted to measure various other cellular and intracellular markers of interest.

This article describes a method to analyze immune cell content of adipose tissue by isolation of immune cells from adipose tissue and subsequent analysis using flow cytometry.

应用流式细胞仪对人体脂肪组织免疫细胞的表征

Infiltration of immune cells in the subcutaneous and visceral adipose tissue (AT) deposits leads to a low-grade inflammation contributing to the ...

Clinical Protocol of Producing Adipose Tissue-Derived Stromal Vascular Fraction for Potential Cartilage Regeneration

Osteoarthritis (OA) is one of the most common debilitating disorders.&#160;Recently, numerous attempts have been made to improve the functions of the knees by using different forms of mesenchymal stem cells (MSCs). In Korea, bone marrow concentrates and cord blood-derived stem cells have been approved by the Korean Food and Drug Administration (KFDA) for cartilage regeneration. In addition, an adipose tissue-derived stromal vascular fraction (SVF) has been allowed by the KFDA for joint injections in human patients. Autologous adipose tissue-derived SVF contains extracellular matrix (ECM) in addition to mesenchymal stem cells. ECM excretes various cytokines that, along with hyaluronic acid (HA) and platelet-rich plasma (PRP) activated by calcium chloride, may help MSCs to regenerate cartilage and improve knee functions. In this article, we presented a protocol to improve knee functions by regenerating cartilage-like tissue in human patients with OA. The result of the protocol was first reported in 2011 followed by a few additional publications. The protocol involves liposuction to obtain autologous lipoaspirates that are mixed with collagenase. This lipoaspirates-collagenase mixture is then cut and homogenized to remove large fibrous tissue that may clog up the needle during the injection. Afterwards, the mixture is incubated to obtain adipose tissue-derived SVF. The resulting adipose tissue-derived SVF, containing both adipose tissue-derived MSCs and remnants of ECM, is injected into knees of patients, combined with HA and calcium chloride activated PRP. Included are three cases of patients who were treated with our protocol resulting in improvement of knee pain, swelling, and range of motion along with MRI evidence of hyaline cartilage-like tissue.

Here, we present a protocol to produce an adipose tissue-derived stromal vascular fraction and its application to improve knee functions by regenerating cartilage-like tissue in human patients with osteoarthritis.

制备脂肪组织基质血管部分用于潜在软骨再生的临床协议

Osteoarthritis (OA) is one of the most common debilitating disorders.&#160;Recently, numerous attempts have been made to improve the functions of the ...

Whole Body and Regional Quantification of Active Human Brown Adipose Tissue Using 18F-FDG PET/CT

In endothermic animals, brown adipose tissue (BAT) is activated to produce heat for defending body temperature in response to cold. BAT&#39;s ability to expend energy has made it a potential target for novel therapies to ameliorate obesity and associated metabolic disorders in humans. Though this tissue has been well studied in small animals, BAT&#39;s thermogenic capacity in humans remains largely unknown due to the difficulties of measuring its volume, activity, and distribution. Identifying and quantifying active human BAT is commonly performed using 18F-Fluorodeoxyglucose (18F-FDG) positron emission tomography and computed tomography (PET/CT) scans following cold-exposure or pharmacological activation. Here we describe a detailed image-analysis approach to quantify total-body human BAT from 18F-FDG PET/CT scans using an open-source software. We demonstrate the drawing of user-specified regions of interest to identify metabolically active adipose tissue while avoiding common non-BAT tissues, to measure BAT volume and activity, and to further characterize its anatomical distribution. Although this rigorous approach is time-consuming, we believe it will ultimately provide a foundation to develop future automated BAT quantification algorithms.

Whole Body and Regional Quantification of Active Human Brown Adipose Tissue Using 18F-FDG PET/CT

Using free, open-source software, we have developed an analytical approach to quantify total and regional brown adipose tissue (BAT) volume and metabolic activity of BAT using 18F-FDG PET/CT.

18F-fdg Petct 对活性人棕色脂肪组织的全身和区域定量

In endothermic animals, brown adipose tissue (BAT) is activated to produce heat for defending body temperature in response to cold. BAT&#39;s ability to ...

Inguinal Subcutaneous White Adipose Tissue (ISWAT) Transplantation Model of Murine Islets

Pancreatic islet transplantation is a well-established therapeutic treatment for type 1 diabetes. The kidney capsule is the most commonly used site for islet transplantation in rodent models. However, the tight kidney capsule limits the transplantation of sufficient islets in large animals and humans. The inguinal subcutaneous white adipose tissue (ISWAT), a new subcutaneous space, was found to be a potentially valuable site for islet transplantation. This site has better blood supply than other subcutaneous spaces. Moreover, the ISWAT accommodates a larger islet mass than the kidney capsule, and transplantation into it is simple. This manuscript describes the procedure of mouse islet isolation and transplantation in the ISWAT site of syngeneic diabetic mouse recipients. Using this protocol, murine pancreatic islets were isolated by standard collagenase digestion and a basement membrane matrix hydrogel was used for fixing the purified islets in the ISWAT site. The blood glucose levels of the recipient mice were monitored for more than 100 days. Islet grafts were retrieved at day 100 after transplantation for histological analysis. The protocol for islet transplantation in the ISWAT site described in this manuscript is simple and effective.

Inguinal Subcutaneous White Adipose Tissue ISWAT Transplantation Model of Murine Islets

In this protocol, a method of murine islet isolation and transplantation into the inguinal subcutaneous white adipose tissue is described. Isolated syngeneic murine islets are transplanted into a murine recipient using a basement membrane hydrogel. The blood glucose level of the recipients is monitored, and histology analysis of the islet grafts is performed.

毛里岛皮下白脂肪组织（ISWAT）移植模型

Pancreatic islet transplantation is a well-established therapeutic treatment for type 1 diabetes. The kidney capsule is the most commonly used site for ...

Fat-Covered Islet Transplantation Using Epididymal White Adipose Tissue

Islet transplantation is a cellular replacement therapy for severe diabetes mellitus. The intraperitoneal cavity is typically the transplant site for this procedure. However, intraperitoneal islet transplantation has some limitations, including poor transplant efficacy, difficult graft detection ability, and a lack of graftectomy capability for post-transplant analysis. In this paper, "fat-covered islet transplantation", an intraperitoneal islet transplantation method that utilizes epididymal white adipose tissue, is used to assess the therapeutic effects of bioengineered islets. The simplicity of the method lies in the seeding of islets onto epididymal white adipose tissue and using the tissue to cover the islets. While this method can be categorized as an intraperitoneal islet transplantation technique, it shares characteristics with intra-adipose tissue islet transplantation. The fat-covered islet transplantation method demonstrates more robust therapeutic effects than intra-adipose tissue islet transplantation, however, including the improvement of blood glucose and plasma insulin levels and the potential for graft removal. We recommend the adoption of this method for assessing the mechanisms of islet engraftment into white adipose tissue and the therapeutic effects of bioengineered islets.

This fat-covered islet transplantation method is suitable for the detection of engrafted islets in the intraperitoneal cavity. Notably, it does not require the use of biobinding agents or suturing.

使用附睾白色脂肪组织进行脂肪覆盖的胰岛移植

Islet transplantation is a cellular replacement therapy for severe diabetes mellitus. The intraperitoneal cavity is typically the transplant site for this ...