作者希望感谢马克罗杰斯概述感染程序和 l. Eoin 卡和布赖恩 Lopresti 的指导, 建立这些成像程序。这项工作的经费由比尔和梅琳达·盖茨基金会 ( J . L . F . 、成功率 ) 、国家卫生研究院、国立反应和传染性疾病研究所 R01 AI111871 ( 成功率 ) 、国家心肺和血液研究所 R01 HL106804 ( J.l.f), R01 HL110811。

这项工作中列出的所有方法都已获得匹兹堡大学动物保育和使用委员会的批准。所有程序都遵循了机构生物安全和辐射安全性要求。CT 扫描需要穿铅围裙和喉盖。生物安全3级 (BSL3) 的外衣和与非人灵长类一起工作的程序必须按照机构的指导方针来遵循。所有扫描都是在 BSL3 设施中进行的. 
 1. 动物感染程序 <ol> <li> 安静的动物, 氯胺酮 (10 毫克/千克, 肌肉注射) 或 telazol (5-8 毫克/千克, 肌肉) 如果动物对氯胺酮有不良反应. </li>
	<li> 使用喉镜, 可视化会厌和声带。麻醉声带通过喷洒 cetacaine 喷雾 1 s (不超过2秒). </li>
	<li> 使用喉镜, 引导一个支气管镜 (2.5 毫米外径) 进入气管 通过 直接可视化进入右尾肺叶. </li>
	<li> 准备一个注射器, 由大约 5-20 (取决于研究) 的菌落形成单位的 m 结核 在2毫升的无菌生理盐水和管理的解决方案通过支气管镜通道。准备一个单独的注射器, 由2毫升无菌生理盐水和管理生理盐水通过支气管镜通道, 其次是5毫升空气, 以确保完全沉积细菌 22 . </li>
	<li> 取出支气管镜并观察猴子直到完全清醒和警觉. </li>
</ol> 2。图像采集、直方图和重建过程 <ol> <li> 准备用于成像的动物。
	<ol> <li> 镇静动物, 氯胺酮 (10 毫克/千克, 肌肉注射) 或 telazol (5-8 毫克/千克, 肌肉) 如果动物对氯胺酮有不良反应. 
		注意: 动物需要在夜间禁食, 以减少在成像过程中呕吐的风险, 并保持葡萄糖 PET 扫描的一致性. </li>
		<li> 将静脉导管插入两条腿的隐静脉, 并用布带固定. </li>
		<li> 将大约5居里剂量的葡萄糖与无菌生理盐水稀释到一个塑料注射器的总容积5毫升. </li>
		<li> 使用剂量校验仪记录注射器中的预放射性水平, 记录时间, 并将注射器放置在注射器支架上. </li>
		<li> 慢慢地通过 IV 导管注入放射性剂量, 并跟随5毫升无菌生理盐水。记录注射时间。注射时间应协调约45分钟-1 小时前 PET 成像. </li>
		<li> 使用剂量校验仪记录注射器的后放射性水平并记录时间。在适当的废物容器中处理注射器. </li>
		<li> 使用喉镜, 形象化的会厌和声带和麻醉与 cetacaine 喷雾. </li>
		<li> 引导气管插管 (3.5 毫米-4.5 毫米, 视猴大小而定) 进入气管, 并在导管的插入端充气. </li>
		<li> 使用长薄的无菌纱布条, 将试管周围的小条包裹好, 用动物的每只狗刺穿条带, 然后在鼻口的桥周围用剩余的纱布绑一个结, 最后绕在后面 o头. </li>
		<li> 用人工泪液遮住眼睛, 防止在成像过程中干燥. </li>
	</ol> </li> <li> 执行 CT 和 PET 扫描。
	<ol> <li> 将动物放在扫描床上. </li>
		<li> 将插管管连接到具有以下设置的呼吸器: 呼吸速率 = 15, 峰值压力 = 15-17, Oxygen% = 40, 窥视 (正端呼气压力) = 3, 潮汐容积 = 60, T i (吸气时间) = 0.4, i: E (吸气呼气时间) 比 = 1:3. 4, T 高原 (在失效前吸气停顿) = 0.5, 峰值流量 = 9.0 (这些值可以根据动物特定的肺顺应性或实验性需求来调整). </li>
		<li> 通过呼吸机开始吸入麻醉 (2% 异氟醚) 并继续, 直到动物对物理刺激没有反应. </li>
		<li> 将动物置于俯卧位置, 并支持其头部和腿部. </li>
		<li> 将动物置于 CT 视中, 并进行预览扫描, 以确保完整扫描中包括整个肺容积范围. </li>
		<li> 用以下参数获取 CT 扫描 (螺旋扫描, 轴向视 = 250 mm, 电压 = 140 伏, 电流 = 2.0 毫安, 切片厚度 = 1.25 mm, 锐度 = 额外锐度), 同时进行呼吸机呼吸保持. 
		注: 造影剂 CT 是可选的。如果进行对比扫描, 在注射造影剂和图像采集之间需要延迟, 因为在心脏中汇集造影剂会干扰 PET 扫描中肺部空间的正确重建, 并在肺部产生伪影。在 ct 扫描上 </li> <li> 确保在扫描过程中将异氟醚浓度降低到 0.7-0.8%. </li>
		<li> 将动物置于宠物视. 
		注: 该系统到位的这项工作是一个内嵌系统的一个单独的 CT 和 PET 扫描仪。根据 CT 坐标, 手动计算 PET 定位坐标. </li>
		<li> 获取每个床位置的 600s PET 图像. 
		注: 焦点220系统的轴向视为7.6 厘米。这项工作是使用四床的位置, 是手工缝制在后期处理. </li>
		<li> 关闭异氟醚, 逐渐关闭呼吸器, 从呼吸机管袖中取出空气, 一旦动物恢复了咳嗽反应并正常呼吸, 就将其取出。取出静脉导管, 在注射部位保持压力直到血流停止. </li>
	</ol> </li> <li> 执行 PET 图像直方图和重建。
	<ol> <li> 使用以下参数执行 PET 图像直方图: 3D 直方图无平滑, 跨度: 3, 环差:47, 全球平均死校正. </li>
		<li> 使用以下参数执行 PET 图像重建 = OSEM3D (有序子集期望 Maximum-3 维度) 算法与 ct 衰减, 斜坡投影滤波器, 和散射校正产生一个284层的图像. </li>
	</ol> </li> <li> 联合注册 PET 和 CT 图像. </li>
	<li> 将合作注册的 PET 和 CT DICOM 图像导出到软件 ( 例如 , OsiriX). </li>
</ol> 3。识别和分析个别病变 <ol> <li> 在轴向方向上从 OsiriX 数据库中打开 pet 和 ct DICOM 图像 (CT 图像将与 pet 图像融合, 并会有一个单独的 pet 图像窗口). </li>
	<li> 将扫描 (或串行扫描) 设置为轴向方向. </li>
	<li> 在 ct 扫描时单击 (任意位置) 并更改 和 #34; 西城/WW 和 #34; 在顶部菜单栏中 和 #34; ct 和 #8211; 肺和 #34; 。
	<li> 滚动扫描以确定肺裂片的起始和结束位置。(识别肺裂隙)
	<ol> <li> 滚动整个扫描, 聚焦于肺部空间的小区域一次. </li>
		<li> 注意正常肺部呈深色, 解剖特征显示较轻 (取决于密度)。气管出现黑色, 而血管几乎是白色的. </li>
		<li> 沿轴向切片滚动时, 跟随船只和呼吸道移动. </li>
		<li> 裂缝可以在没有船只或空中航线的地区识别。(这些是肺部的区域, 只出现在黑暗中, 没有其他解剖结构.) </li>
	</ol> </li> <li> 使用熔融 PET/CT 识别病灶。
	<ol> <li> 滚动整个扫描, 同时聚焦于肺部的小面积. 
		注意: 集中在一个肺叶的时间, 以确定和计数病变. </li>
		<li> 在肺部发现葡萄糖-狂热的病灶。它们看起来像是热球体, 与肺部背景非常不同。小的, 冷的病灶将不太明显, 更难以辨认。它们将在扫描时显示为在滚动时不移动的稠密结构 (如容器). 
		注意: 小病变和血管看起来非常相似。区分两者的一个简单方法是将光标悬停在有问题的结构上, 然后向上和向下滚动一两个切片。如果结构停留在光标下方, 则结构是一个损伤。如果在向上或向下滚动切片时结构从光标处移开, 则最有可能是容器或气道. </li>
		<li> 用于标识目的, 请使用 和 #34; 箭头和 #34; 工具指向扫描中的每个病灶. </li>
		<li> 为了便于定位, 请使用 和 #34;P 点和 #34; 工具并单击病变部位, 使 ROI (感兴趣的区域) 直接位于肉芽肿的中心。此 ROI 中包含的信息将包括可在其中找到病灶的笛卡尔坐标 (XYZ 坐标). </li>
	</ol> </li> <li> 使用 #34; 长度和 #34; 和 #34; 椭圆形和 #34; 用于测量每个病灶大小 (mm) 和葡萄糖亲和力 (SUV) 的工具。
	<ol> <li> 若要测量病变的大小, 请删除 PET 信号, 以便只有 CT 可见. </li>
		<li> 选择 和 #34; 长度和 #34; 工具 </li>
		<li> 滚动直到包含病灶的最大部分的切片被识别 (病灶表面看起来是最大的切片). </li>
		<li> 在病变最长的长度上画一条线。此 ROI 中包含的信息将表示病灶直径的长度 (毫米). </li>
		<li> 要测量病变的葡萄糖亲和力, 首先点击 pet 扫描并打开 pet。转到 #34; #38; 查找和 #34; Osirix 菜单在屏幕顶部, 选择 和 #34; 在 "西城/ww" 下拉菜单中手动和 #34 设置。在对话框中, 将 0 输入到 和 #34; 从和 #34; 和 20 进入 和 #34; #34; 以将窗口限制为0到 20 SUV. </li>
		<li> 选择 和 #34; 椭圆和 #34; 来自 和 #34 的工具; 鼠标按钮功能和 #34; 工具下拉菜单. </li>
		<li> 在病灶上方滚动以评估病灶的最热部分。在病灶周围画一个椭圆形。 #34; 椭圆和 #34; 工具 ROI 信息包括区域内所有体的 suv 的描述性统计数据。在该区域中记录 最大 SUV . </li>
		<li> 每个 和 #34; 椭圆和 #34; ROI 只代表该特定轴向平面的 SUV 值和典型病变是球形的形状, 在几个切片上绘制椭圆形, 以确保该病变的实际最大 suv 捕获. 
		注意: 如果 pet/ct 扫描被人工重建, pet 和 ct 图像可能不会完全匹配。如果是这样的话, 所有的 SUV 分析和 roi 应进行 pet 扫描, 而不是融合 pet/CT 扫描。由于许多病变都小于 PET 探测器晶体的分辨率, 所有测量的单个病灶的 suv 都被输入一个恢复系数计算器电子表格, 对每个病灶执行部分音量校正 23 . </li> </ol> </li> </ol> 4。全肺葡萄糖亲和力测定法测定全肺炎症 <ol> <li> 在轴向方向上从 OsiriX 数据库中打开 pet 和 ct DICOM 图像 (ct 图像将与 pet 图像融合, 并会有一个单独的 pet 图像窗口). </li>
	<li> 在 CT 图像上执行肺容积的分割。
	<ol> <li> 在 CT 扫描的任意位置单击以确保它是活动窗口. </li>
		<li> 转到 ROI 下拉菜单并选择 和 #34; 增长区域 (2 维/3 维) 分割和 #8230; #34; . </li>
		<li> 为了捕获正常肺部的密度, 将下限阈值设置为-1024, 将上限阈值设为-200。这些都是 Hounsfield 单位的指示, 尽管 "分割" 框并没有这样命名. </li>
		<li> 一旦设置了下限和上限阈值, 请单击肺部内的任意位置。整个肺部应以绿色突出显示. </li>
		<li> 下一步, 单击 和 #34; 计算和 #34; 在 "分段参数" 对话框中。这将扩大生长区域从一个切片到整个肺部体积. </li>
	</ol> </li> <li> 移动和 #34; 增长区域和 #34; 从 CT 扫描到 PET 扫描的肺部。
	<ol> <li> 单击 CT 扫描名称左侧的小图标, 然后将图标拖动到 PET 扫描. </li>
		<li> 选择 和 #34; 复制 roi 和 #34; 。现在应该有一个覆盖的肺部在 PET 扫描. </li>
	</ol> </li> <li> 从 CT 扫描中删除 ROI (可选). 
	注意: 它有助于能够看到整个肺部没有 roi 的 CT 扫描, 以确保所有病理在肺部被抓获。为此, 请删除 roi。请确保 ct 窗口处于活动状态 (单击 ct 扫描) 选择 ROI 下拉菜单, 然后选择 和 #34;D elete 本系列中的所有 roi. 和 #34; </li> <li> 填充在 PET 扫描中显示为间隙的肺部的高密度区域. 
	注: 在许多情况下, PET 扫描的 ROI 有孔, 在 CT 扫描中肺组织密度大于-200 胡 (如果不发生此步骤可以跳过)。
	<ol> <li> 突出显示 ROI 下拉菜单并选择 和 #34; 画笔 roi 和 #34; #160; #8594; #34; 关闭. #34; 当出现对话框时, 将箭头滑动到 3, 以便对话框的顶部读取 和 #34; 结构元素半径: 3 和 #34; 并检查 和 #34; 应用于同名的所有 roi. #34; 注意: 当有很大一部分疾病 (如合并的密度比周围的肺部组织), 经常关闭刷子 roi将不足以填补整个肺部。如果是这种情况, 则必须手动填充间隙. </li>
		<li> 转到 #34; 鼠标按钮功能和 #34; 区域在顶部菜单上, 单击右侧的小箭头. </li>
		<li> 选择 和 #34; 画笔和 #34; 工具. </li>
		<li> 一旦选择了此工具, 请在 ROI 中手动绘制以填充孔. </li>
	</ol> </li> <li> 在 PET 扫描中隔离肺部 ROI。
	<ol> <li> 现在有一个代表整个肺部的宠物扫描, 删除肺部以外的所有像素. </li>
		<li> 突出显示 ROI 下拉菜单并选择 和 #34; 将像素值设置为和 #8230; #34; . </li>
		<li> 单击 外部 roi " 复选框, 并将 roi 以外的所有像素设置为 0. </li>
	</ol> </li> <li> 隔离和 #34; 热 #34; 病理。
	<ol> <li> 使用任何希望用作和 #34 的阈值; 热的 #34; 超过2.3 的 suv 被认为是和 #34; 热 #34; 基于文献价值的结核病损害诊断 24 . </li>
		<li> 选择 ROI 下拉菜单并选择 和 #34; 将像素值设置为和 #8230; #34; . </li>
		<li> 单击 内部 ROI 复选框。确保单击 "#34" 和 "#34"; 框, 使 0 和 2.3 之间的所有值都设置为 0. </li>
	</ol> </li> <li> 确保在 ROI 中只考虑病理学。
	<ol> <li> 请注意, 某些区域 (如肝脏) 的温度高于2.3。通过删除 roi 并创建另一个增长区域, 确保只捕获所需的区域。其他常见的组织, 在这个时候干扰包括心脏, 纵隔淋巴结, 椎骨, 肋骨. </li>
		<li> 突出显示 roi 下拉菜单并选择 和 #34;D elete 本系列中的所有 roi. #34; 下一步, 转到 roi 并选择 和 #34; 增长区域 (2 维/3 维) 分割和 #8230; #34; 。
		<li> 更改阈值以将阈值降低到 2.3, 将上限阈值转换为 100. </li>
		<li> 滚动整个 PET 窗口, 点击疾病病理学和点击 和 #34; 计算. #34; 对热疾病的每个区域重复。在 roi 菜单下, 确保使用 #34; 保存 roi 和 #34; 选项, 以节省整个肺投资回报率. </li>
	</ol> </li> <li> 将原始值导出到电子表格中。
	<ol> <li> 转到下拉菜单栏中的 2D 查看器 , 然后选择 和 #34; 还原系列. #34; </li> <li> 下一步, 转到 插件 下拉菜单栏. </li>
		<li> 选择 和 #34; ROI 工具和 #34; #160; #8594; #34; 导出 roi #34; 命名并保存导出的原始数据文件。请确保在该对话框的底部选择 #34; CSV 和 #34; . </li>
	</ol> </li> <li> 从原始数据中计算总的葡萄糖亲和力。此电子表格中的每一行表示扫描中的单个切片。感兴趣的列是 and #34; RoiTotal #34; <ol> <li> 以计算和 #34; 总葡萄糖亲和力, #34; 添加所有的 和 #34; RoiTotal 和 #34; 将切片的 组合在一起。计算列 F (RoiTotal) 的总和。这是总的葡萄糖亲和力测量. </li>
		<li> 如果 OsiriX 没有导出 ROI 插件, 请转到 插件 在下拉菜单中。选择 #34;P lugins 管理器和 #8230; #34; 单击对话框顶部的 & #34;D ownload 和 #8230; #34; 选项卡。从 和 #34; 可用插件和 #34; 下拉菜单中选择 #34; ExportROIs 和 #34; 。选择 #34;D ownload 和 #38; 安装. #34; </li> </ol> </li> </ol> 5。测定 #34 中葡萄糖摄取量的分析程序; 热 #34; 淋巴结肿大 <ol> <li> 在轴向方向 OsiriX 数据库中打开 pet 和 ct DICOM 图像 (CT 图像将与 pet 图像融合, 并会有一个单独的 pet 图像窗口). </li>
	<li> 确保在对 PET 图像执行手动 ROI 分析时, 图像强度窗口是一致的。
	<ol> <li> 单击 PET 图像窗口以确保它是活动窗口. </li>
		<li> 在 "OsiriX" 菜单中, 单击 " 和 #34; 妇女协会/ww 和 #34; 下拉菜单, 然后单击 和 #34; 手动和 #34; 。
		<li> 当下拉框出现在活动 PET 窗口中时, 请在 和 #34 中填充所需的最小强度值; 从和 #34; 字段和在 和 #34 中所需的最大强度值; to 和 #34; 字段 (如 到窗口PET 图像从0到 20 SUV, 类型0进入和 #34; 从和 #34; 字段和20进入和 #34; #34; 字段). </li>
		<li> 或者, 如果希望始终加载具有相同亮度值的图像, 请在顶部菜单突出显示 Osirix -#62; PET 和 #62; 然后根据 和 #34; 窗口级别和 #38; 宽度和 #34; 部分, 单击 使用固定级别 气泡并在 & #34 中插入所需的值; 从和 #34; 和 和 #34; #34; 字段. </li>
	</ol> </li> <li> 一旦确定了所需的淋巴结, 请在淋巴结边缘周围手工绘制一个 ROI。
	<ol> <li> 突出显示 PET/CT 融合图像, 以确保它是活动窗口. </li>
		<li> 由于对此分析使用彩色查找表非常有用, 要更改此设置, 请单击 和 #34; 查找和 #34; 主 OsiriX 工具栏中的下拉框, 并选择所需的查找表设置 (UCLA 首选). </li>
		<li> 若要在淋巴结周围绘制手动 ROI, 请单击 和 #34 的右侧的下拉菜单; 鼠标按钮功能和 #34; 在主工具栏中选择 和 #34; 关闭多边形和 #34; 。点击边缘的淋巴结根据宠物视窗查找表建立第一个点的 ROI. </li>
		<li> 单击淋巴结外边缘的另一点, 继续追踪, 直到淋巴结几乎封闭. </li>
		<li> 若要确定 roi 的最终点, 请双击以在 roi 中关闭. </li>
		<li> 在多个切片上重复此过程以确保确定淋巴结内的最大 SUV. </li>
		<li> 在单独的电子表格中记录所需的 SUV 数据. </li>
	</ol> </li> </ol> 6。葡萄糖浓度的测定标准值规范化的背景摄取量 注意: 为了保持多个成像时间点在葡萄糖摄取和动物体内代谢活性变化的一致性不同的时间, 所有的 PET 分析应归为肌肉, 并提出这样的。在这项工作中提出的所有定量 PET 数据表示为一个 SUVCMR (标准摄取值缸肌比). 
<ol> <li> 在轴向方向上从 OsiriX 数据库中打开 pet 和 ct DICOM 图像 (ct 图像将与 pet 图像融合, 并会有一个单独的 pet 图像窗口). </li>
	<li> 单击 Co 注册的 PET 和 CT 图像以确保它是活动窗口. </li>
	<li> 滚动图像, 直到到达包含主支气管 (底座) 的汇点的切片. </li>
	<li> 在背部肌肉上绘制 roi 以获取背景 SUV 值。
	<ol> <li> 在 OsiriX 主菜单中选择 & #34; 鼠标按钮选项和 #34; 的 "ROI" 下拉工具. </li>
		<li> 突出显示 和 #34; 椭圆和 #34; 作为 ROI 工具. </li>
		<li> 在脊柱后部和侧面的肌肉上绘制近似相同大小的 roi. </li>
		<li> 单击 Co 注册 pet/CT 扫描左侧的图标, 然后将图标拖动到 pet 窗口. </li>
		<li> 选择 和 #34; 复制 roi 和 #34; 。现在应该在 PET 扫描窗口中看到 roi. </li>
		<li> 在主菜单上, 选择 和 #34; 模式和 #34; 复选框并确保 和 #34; MIP 和 #8211; 最大强度投射和 #34; 在右侧的下拉菜单中立即选中. </li>
		<li> 确保 和 #34; 厚板和 #34; 滑动比例设置为 10 。这表明, 10 切片是结合在 PET 图像作为最大强度投影, 使 #34; 气缸和 #34; 兴趣量 (气缸肌肉比的来源). </li>
	</ol> </li> <li> 在电子表格中记录两个 roi 的平均 SUV 值. </li>
	<li> 平均两个值以获取背景葡萄糖摄取值。这是用于获得目标部位摄取量与基础代谢率之间的比值值的值. </li>
</ol>

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作者没有什么可透露的。

从 PET/CT 获得的数据可以作为替代测量的许多方面的m 结核感染, 将观测没有这样的技术。PET/CT 比 x 射线技术更灵敏, 它通常用于猕猴的研究。PET/CT 提供结构, 空间和功能的信息。上述分析有许多实际应用, 如监测疾病进展, 评估药物治疗的有效性, 并为重新激活6,10,11提供风险因素, 13。
<p class=...

结核分枝杆菌已经折磨了人类几千年, 造成的死亡率比当今世界上任何其他单一的传染性药物都要多。在 2015年, 有1050万报告的新的肺结核 (TB) 病例全球1 , 大多数病例来自印度、印度尼西亚、中国、尼日利亚、巴基斯坦和南非。估计将全球死亡人数从结核病的140万人在同一时期。这个值比100年前的死亡率低近25%。虽然药物敏感的结核病是可治疗的, 但疗程冗长需要多种药物, 依从性是一个关注的问题。多药耐药性 (MDR) 菌株的出现在2015年占新结核病病例的58万。m 结核耐多药菌株的成功治疗率估计为50% 左右。更令人担忧的是, 出现了广泛耐药 (XDR) 的m 结核菌株, 它对几乎所有可用的药物都有抗药性。因此, 在结核病研究领域需要新的技术, 提高诊断结核的能力, 提高对疾病过程的免疫学认识, 并允许对新的治疗方法和预防策略进行筛选, 包括抗生素方案和疫苗功效研究。
m. 结核是一种有氧的 acid-fast 芽孢杆菌, 其物理特征是其非常复杂的外细胞壁和缓慢的生长动力学。感染通常是通过吸入雾化液滴中的单个细菌而引起的, 它们是由有症状的、受感染的个体在咳嗽、打喷嚏或唱歌时排出的。在暴露的感染个体中, 只有 5-10% 的人发展为积极的临床结核病。其余90% 有不同程度的无症状感染, 范围从亚临床感染到无疾病, 所有这些都被归类为潜伏性结核病感染 (LTBI)2,3。在有这种无症状感染的人群中, 大约10% 将通过在其生命周期中重新激活所含的感染来发展活跃的结核病。如果有无症状感染的人染上 HIV 或接受免疫抑制剂治疗, 如 TNF 抑制剂4,5,6, 则重新激活的风险会显著增加。活动性结核病也表现为频谱, 大部分人患有肺结核, 影响肺部和胸淋巴结。然而, m. 结核可以感染任何器官, 因此感染也可以出现在外的参与部位。
m 结核感染的病理特征是宿主细胞的组织球形结构, 称为肉芽肿。巨噬细胞、T 细胞和 B 淋巴细胞是肉芽肿的主要成分, 有可变数量的中性粒细胞7。肉芽肿的中心经常坏死。因此, 肉芽肿功能作为一个免疫微环境杀死或控制杆菌, 防止扩散到其他部分的肺部。然而, m. 结核可以颠覆肉芽肿的致死, 并在这些结构中持续数十年。在新感染或重新激活 LTBI 后, 对积极结核病的发展进行持续和定期的监测是不切实际的, 具有科学挑战性和耗时。在人类和类似人类的动物模型中纵向研究这些过程的技术对于科学界进一步了解m. 结核感染和疾病的许多复杂性非常有用。
PET/CT 是一种非常有用的成像技术, 已被用来研究人类和动物模型的广泛疾病状态8。PET 是一种利用正电子发射的放射性化合物作为一个记者的功能技术。这些放射性同位素通常是功能化的代谢化合物, 如葡萄糖, 或目标组, 旨在绑定到一个受体的利益。由于 PET 同位素发出的辐射强度足以穿透组织, 因此极低的浓度可以用于研究受体靶向化合物中的饱和度, 且浓度低, 对新陈代谢没有影响。使用诸如 2-脱氧-2-(18F) 氟-d-葡萄糖 (葡萄糖) 等药剂的过程。CT 是一种三维 x 射线成像技术, 它使用不同程度的 x 射线衰减来识别人体内器官的物理特性9。当与宠物配对时, CT 被用作地图来确定特定的位置和结构, 以显示宠物示的摄取量。PET/CT 是一种功能强大的工具,在体内成像的人和动物模型感染的m 结核感染, 导致了许多重要的洞察力发病机制, 反应药物治疗, 疾病频谱等6 ,10,11,12。这项工作描述了特定的 PET/CT 分析方法, 在非人类灵长类动物模型中纵向使用参数, 如肉芽肿大小、个体病变中的葡萄糖摄取量、全肺和淋巴结葡萄糖亲和力, 以及检测外疾病6,10,11,12。
这篇手稿描述了非人类灵长类动物 (NHPs) 的成像分析方法, 特别是蟹猕猴, 用于纵向评估疾病进展和药物治疗感染后的m 结核.NHPs 是一个有价值的动物模型, 因为当接种了低剂量的m 结核艾德曼菌株, 动物显示了各种疾病的结果与〜50% 开发活动结核和其余的动物有无症状感染 (即控制感染, LTBI), 为人类所见的临床疾病谱提供最接近的模型3,13,14,15,16。LTBI 在猕猴中的重新激活是由引起人类重新激活的同一种药物引起的, 其中的例子包括人类免疫机能丧失病毒 (hiv、使用猿猴免疫缺陷病毒 (SIV) 作为猕猴版的艾滋病毒)、CD4 衰竭或肿瘤坏死因子 (TNF) 中和13,16。此外, 猕猴的病理学与人类所见的非常相似, 包括在肺部或其他器官中形成的有组织的肉芽肿17。因此, 该模型提供了重要的洞察力的基本主机-病原体相互作用的m 结核感染, 以及关于药物治疗方案和结核病疫苗的宝贵知识14,18,19,20,21。
PET/CT 成像提供了跟踪单个肉芽肿的外观、分布和进展的能力.这项工作主要是作为一个探针, 作为葡萄糖模拟, 合并到新陈代谢活性宿主细胞, 如巨噬细胞, 嗜中性细胞和淋巴细胞8, 所有这些都在肉芽肿。因此, 葡萄糖是宿主炎症的代表。本文详细的分析程序使用 OsiriX, 一个广泛使用的 DICOM 查看器可供购买和使用。图象分析方法描述跟踪个体肉芽肿的形状、大小和新陈代谢活动 (通过葡萄糖吸收) 随着时间的推移, 并利用成像作为一个地图, 以确定特定的病变后, 动物尸检。此外, 还制定了一个单独的方法来量化在一个特定的阈值 (SUV ≥ 2.3) 的肺部葡萄糖摄取总和, 并使用这个值来评估控制和实验组之间的差异, 从疫苗的研究范围感染模型的试验。这些数据支持, 这一全面的措施, 在肺部摄取葡萄糖与细菌的负担, 从而提供有关疾病状况的信息。类似的分析可以进行的葡萄糖摄取胸淋巴结的研究进展的疾病, 以及。下面的协议描述了从动物感染通过图像分析的实验过程。

<table><tbody><tr><td>Ketamine</td><td>Henry Schein</td><td>23061</td><td>Henry Schein</td></tr><tr><td>Telazol</td><td>Zoetis</td><td>4866</td><td>Henry Schein</td></tr><tr><td>Cetacaine</td><td>Patterson Vet Generics</td><td>07-892-6862</td><td>Patterson</td></tr><tr><td>Sterile saline</td><td>Hospira</td><td>07-800-9721</td><td>Patterson</td></tr><tr><td>7H11 agar</td><td>BD</td><td>283810</td><td>BD Biosciences</td></tr><tr><td>IV catheter</td><td>Surflash</td><td>07-806-7659</td><td>Patterson</td></tr><tr><td>18F-FDG</td><td>Zevacor</td><td>N/A</td><td></td></tr><tr><td>Endotracheal tube</td><td>Jorgensen Labs Inc</td><td>07-887-0284</td><td>Patterson</td></tr><tr><td>Artificial tears</td><td>Patterson Vet Generics</td><td>07-888-1663</td><td>Patterson</td></tr><tr><td>Isoflurane</td><td>Zoetis</td><td>07-806-3204</td><td>Patterson</td></tr><tr><td>Neurologica Ceretom CT</td><td>Samsung Neurologica</td><td>N/A</td><td></td></tr><tr><td>Siemens Focus 220 microPET</td><td>Siemens Molecular Imaging Systems</td><td>N/A</td><td></td></tr><tr><td>Inveon Research Software</td><td>Siemens Molecular Imaging Systems</td><td>N/A</td><td></td></tr><tr><td>OsiriX</td><td>Pixmeo</td><td>N/A</td><td></td></tr></tbody></table>

analysis of 18fdg pet/ct imaging as a tool for studying mycobacterium tuberculosis infection and treatment in non-human primates

结核分枝杆菌仍然是当今世界头号传染性药物。随着抗生素耐药性菌株的出现, 需要新的临床相关方法来评估疾病的过程和筛选潜在的抗生素和疫苗治疗。正电子发射断层扫描/计算机断层扫描 (PET/CT) 已经建立, 作为一个有价值的工具, 研究许多疾病, 如癌症, 阿尔茨海默氏症, 炎症/感染。这里概述了一些策略, 已被用于评估蟹猕猴的 PET/CT 图像的感染 intrabronchially 低剂量的m 结核。通过对 pet 图像中病灶和淋巴结的 ct 和摄取量的葡萄糖 (葡萄糖) 的损伤大小进行评价, 这些方法表明 pet/ct 成像能预测活性与潜伏性疾病的未来发展, 并从潜在的感染状态重新激活的倾向。此外, 通过分析肺部炎症的总体水平, 这些方法确定了药物对m 结核的抗生素疗效, 这是目前临床上最相关的动物模型。这些图像分析方法是在阿森纳对抗这种疾病的一些最强大的工具, 他们不仅可以评估一些感染和药物治疗的特点, 但它们也可直接翻译成临床设置, 用于人类研究.

个体病变的鉴别与分析
个别肉芽肿可以可视化的数量, 大小和葡萄糖摄取定性地了解感染过程的一般范围 (图 1)。使用这些图像, 计数肉芽肿随着时间的推移是一个定量的疾病传播的尺度。图 2描述了一组10只动物在一段时间内的个别肉芽肿计数。在这10动物中, 有三个被开发的...

Watch this Scientific Journal Video about Analysis of 18FDG PET/CT Imaging as a Tool for Studying Mycobacterium tuberculosis Infection and Treatment in Non-human Primates at JoVE.com

Analysis of 18FDG PET/CT Imaging as a Tool for Studying Mycobacterium tuberculosis Infection and Treatment in Non-human Primates

在这里, 我们提出了一个协议来描述的分析, 18f-葡萄糖 PET/CT 成像的非人灵长类已经感染了m 结核研究疾病的过程, 药物治疗, 和疾病的重新激活。

分析18葡萄糖 PET/CT 成像作为研究结核分枝杆菌感染和治疗非人类灵长类动物的工具

Mycobacterium tuberculosis remains the number one infectious agent in the world today. With the emergence of antibiotic resistant strains, new clinically ...

analysis-18fdg-petct-imaging-as-tool-for-studying-mycobacterium

Universidad de Cartagena UDEC

Research

JoVE Journal

Immunology and Infection

Analysis of 18FDG PET/CT Imaging as a Tool for Studying Mycobacterium tuberculosis Infection and Treatment in Non-human Primates

18.7K 次观看.  University of Pittsburgh School of Medicine. 在这里, 我们提出了一个协议来描述的分析, 18f-葡萄糖 PET/CT 成像的非人灵长类已经感染了m 结核研究疾病的过程, 药物治疗, 和疾病的重新激活。

视频: Analysis of 18FDG PET/CT Imaging as a Tool for Studying Mycobacterium tuberculosis Infection and Treatment in Non-human Primates

Utilizing 18F-FDG PET/CT Imaging and Quantitative Histology to Measure Dynamic Changes in the Glucose Metabolism in Mouse Models of Lung Cancer

A hallmark of advanced tumors is a switch to aerobic glycolysis that is readily measured by [18F]-2-fluoro-2-deoxy-D-glucose positron emission tomography (18F-FDG PET) imaging. Co-mutations in the KRAS proto-oncogene and the LKB1 tumor suppressor gene are frequent events in lung cancer that drive hypermetabolic, glycolytic tumor growth. A critical pathway regulating the growth and metabolism of these tumors is the mechanistic target of the rapamycin (mTOR) pathway, which can be effectively targeted using selective catalytic mTOR kinase inhibitors. The mTOR inhibitor MLN0128 suppresses glycolysis in mice bearing tumors with Kras and Lkb1 co-mutations, referred to as KL mice. The therapy response in KL mice is first measured by 18F-FDG PET and computed tomography (CT) imaging before and after the delivery of MLN0128. By utilizing 18F-FDG PET/CT, researchers are able to measure dynamic changes in the glucose metabolism in genetically engineered mouse models (GEMMs) of lung cancer following a therapeutic intervention with targeted therapies. This is followed by ex vivo autoradiography and a quantitative immunohistochemical (qIHC) analysis using morphometric software. The use of qIHC enables the detection and quantification of distinct changes in the biomarker profiles following treatment as well as the characterization of distinct tumor pathologies. The coupling of PET imaging to quantitative histology is an effective strategy to identify metabolic and therapeutic responses in vivo in mouse models of disease.

Utilizing 18F-FDG PET/CT Imaging and Quantitative Histology to Measure Dynamic Changes in the Glucose Metabolism in Mouse Models of Lung Cancer

In this protocol, we describe how to utilize [18F]-2-fluoro-2-deoxy-D-glucose positron emission tomography and computed tomography (18F-FDG PET/CT) imaging to measure the tumor metabolic response to the targeted therapy MLN0128 in a Kras/Lkb1 mutant mouse model of lung cancer and coupled imaging with high resolution ex vivo autoradiography and quantitative histology.

利用18f-葡萄糖 PET/CT 成像和定量组织学测量肺癌小鼠模型葡萄糖代谢的动态变化

A hallmark of advanced tumors is a switch to aerobic glycolysis that is readily measured by [18F]-2-fluoro-2-deoxy-D-glucose positron emission tomography ...

科研

癌症研究

Multimodal Bioluminescent and Positronic-emission Tomography/Computational Tomography Imaging of Multiple Myeloma Bone Marrow Xenografts in NOG Mice

Multiple myeloma (MM) tumors engraft in the bone marrow (BM) and their survival and progression are dependent upon complex molecular and cellular interactions that exist within this microenvironment. Yet the BM microenvironment cannot be easily replicated in vitro, which potentially limits the physiologic relevance of many in vitro and ex vivo experimental models. These issues can be overcome by utilizing a xenograft model in which luciferase (LUC)-transfected 8226 MM cells will specifically engraft in the mouse skeleton. When these mice are given the appropriate substrate, D-luciferin, the effects of therapy on tumor growth and survival can be analyzed by measuring changes in the bioluminescent images (BLI) produced by the tumors in vivo. This BLI data combined with positronic-emission tomography/computational tomography (PET/CT) analysis using the metabolic marker 2-deoxy-2-(18F)fluoro-D-glucose (18F-FDG) is used to monitor changes in tumor metabolism over time. These imaging platforms allow for multiple noninvasive measurements within the tumor/BM microenvironment.

Here we use bioluminescent, X-ray, and positron-emission tomography/computed tomography imaging to study how inhibiting mTOR activity impacts bone marrow-engrafted myeloma tumors in a xenograft model. This allows for physiologically relevant, non-invasive, and multimodal analyses of the anti-myeloma effect of therapies targeting bone marrow-engrafted myeloma tumors in vivo.

nog 小鼠多发性骨髓瘤异种移植的多模态生物发光和正电子发射成形断层扫描成像

Multiple myeloma (MM) tumors engraft in the bone marrow (BM) and their survival and progression are dependent upon complex molecular and cellular ...

Hybrid &#181;CT-FMT imaging and image analysis

Fluorescence-mediated tomography (FMT) enables longitudinal and quantitative determination of the fluorescence distribution in vivo and can be used to assess the biodistribution of novel probes and to assess disease progression using established molecular probes or reporter genes. The combination with an anatomical modality, e.g., micro computed tomography (&#181;CT), is beneficial for image analysis and for fluorescence reconstruction. We describe a protocol for multimodal &#181;CT-FMT imaging including the image processing steps necessary to extract quantitative measurements. After preparing the mice and performing the imaging, the multimodal data sets are registered. Subsequently, an improved fluorescence reconstruction is performed, which takes into account the shape of the mouse. For quantitative analysis, organ segmentations are generated based on the anatomical data using our interactive segmentation tool. Finally, the biodistribution curves are generated using a batch-processing feature. We show the applicability of the method by assessing the biodistribution of a well-known probe that binds to bones and joints.

Hybrid  &#181;CT-FMT imaging and image analysis

We describe a protocol for hybrid imaging, combining fluorescence-mediated tomography (FMT) with micro computed tomography (&#181;CT). After fusion and reconstruction, we perform interactive organ segmentation to extract quantitative measurements of the fluorescence distribution.

混合μCT-FMT成像和图像分析

Fluorescence-mediated tomography (FMT) enables longitudinal and quantitative determination of the fluorescence distribution in vivo and can be used to ...

生物工程

A 3D Human Lung Tissue Model for Functional Studies on Mycobacterium tuberculosis Infection

Tuberculosis (TB) still holds a major threat to the health of people worldwide, and there is a need for cost-efficient but reliable models to help us understand the disease mechanisms and advance the discoveries of new treatment options. In vitro cell cultures of monolayers or co-cultures lack the three-dimensional (3D) environment and tissue responses. Herein, we describe an innovative in vitro model of a human lung tissue, which holds promise to be an effective tool for studying the complex events that occur during infection with Mycobacterium tuberculosis (M. tuberculosis). The 3D tissue model consists of tissue-specific epithelial cells and fibroblasts, which are cultured in a matrix of collagen on top of a porous membrane. Upon air exposure, the epithelial cells stratify and secrete mucus at the apical side. By introducing human primary macrophages infected with M. tuberculosis to the tissue model, we have shown that immune cells migrate into the infected-tissue and form early stages of TB granuloma. These structures recapitulate the distinct feature of human TB, the granuloma, which is fundamentally different or not commonly observed in widely used experimental animal models. This organotypic culture method enables the 3D visualization and robust quantitative analysis that provides pivotal information on spatial and temporal features of host cell-pathogen interactions. Taken together, the lung tissue model provides a physiologically relevant tissue micro-environment for studies on TB. Thus, the lung tissue model has potential implications for both basic mechanistic and applied studies. Importantly, the model allows addition or manipulation of individual cell types, which thereby widens its use for modelling a variety of infectious diseases that affect the lungs.

A 3D Human Lung Tissue Model for Functional Studies on Mycobacterium tuberculosis Infection

Human tuberculosis infection is a complex process, which is difficult to model in vitro. Here we describe a novel 3D human lung tissue model that recapitulates the dynamics that occur during infection, including the migration of immune cells and early granuloma formation in a physiological environment.

关于功能研究的三维人体肺组织模型<em&gt;结核分枝杆菌</em&gt;感染

Tuberculosis (TB) still holds a major threat to the health of people worldwide, and there is a need for cost-efficient but reliable models to help us ...

免疫与感染

<meta charset="utf8"></head><body>使用 BCAM 和 SaaS 工具对小鼠进行 PET 成像和生物分布分析

Preclinical Positron Emission Tomography with Body Conforming Animal Molds for Cloud-Based Automated Image Analysis in Mice

Positron Emission Tomography (PET) is a molecular imaging modality that can be used to investigate a multitude of pharmacological questions, such as biomarker modulation, receptor occupancy, and biodistribution of compounds of interest. In biodistribution studies, experimental subjects are often longitudinally imaged after receiving the test article. The images are then analyzed to derive the compound&#39;s distribution profile in various organs at different timepoints. This constitutes a crucial step in drug development to understand the distribution and potentially binding profile of an investigative compound. Standard/manual methods of PET imaging-based biodistribution analyses, however, are labor-intensive and time-consuming and are often associated with high inter-operator variability. Further, it is challenging to keep the animals&#39; positions consistent across different timepoints. To address these shortcomings, a series of mouse Body Conforming Animal Molds (BCAMs) were used to enable rigid and consistent positioning of animals during PET/CT imaging acquisition. Further, a Software-as-a-Service (SaaS) platform consisting of a cloud-based Organ Probability Map (OPM) and an artificial intelligence-powered segmentation tool were employed to enable reliable and automated quantitation of in vivo PET imaging data. The workflow presented here includes (1) prepping mice for imaging with the BCAMs, including the proper implantation of subcutaneous tumors to be compatible with the molds, (2) acquiring PET/CT images with BCAMs using the G8 scanner, and 3) performing automated organ segmentation and biodistribution analysis using the cloud-based SaaS. [18F]FDG was used as an exemplar tracer here, but other biomarkers and/or radio-labeled compounds can be readily adapted into the workflow. This procedure can be executed accurately and effectively with minimal training, and the automated PET data analysis yielded satisfactory results consistent with the manual method.

<meta charset="utf8"></head><body>作者聚焦:使用人体贴合模具和自动分析实现小鼠体内 PET 成像的标准化

This protocol describes the procedure for performing in vivo PET imaging on mice using Body Conforming Animal Molds (BCAMs) in the G8 PET/CT scanner. Technical details on mouse preparation, including proper tumor implantation, optimal positioning, BCAM-assisted PET/CT image acquisition, and data analysis, are provided.

使用身体顺应动物模具的临床前正电子发射断层扫描用于小鼠基于云的自动图像分析

Positron Emission Tomography (PET) is a molecular imaging modality that can be used to investigate a multitude of pharmacological questions, such as ...

Imaging Mycobacterium tuberculosis in Mice with Reporter Enzyme Fluorescence

Reporter enzyme fluorescence (REF) utilizes substrates that are specific for enzymes present in target organisms of interest for imaging or detection by fluorescence or bioluminescence. We utilize BlaC, an enzyme expressed constitutively by all M. tuberculosis strains. REF allows rapid quantification of bacteria in lungs of infected mice. The same group of mice can be imaged at many time points, greatly reducing costs, enumerating bacteria more quickly, allowing novel observations in host-pathogen interactions, and increasing statistical power, since more animals per group are readily maintained. REF is extremely sensitive due to the catalytic nature of the BlaC enzymatic reporter and specific due to the custom flourescence resonance energy transfer (FRET) or fluorogenic substrates used. REF does not require recombinant strains, ensuring normal host-pathogen interactions. We describe the imaging of M. tuberculosis infection using a FRET substrate with maximal emission at 800 nm. The wavelength of the substrate allows sensitive deep tissue imaging in mammals. We will outline aerosol infection of mice with M. tuberculosis, anesthesia of mice, administration of the REF substrate, and optical imaging. This method has been successfully applied to evaluating host-pathogen interactions and efficacy of antibiotics targeting M. tuberculosis.

Imaging Mycobacterium tuberculosis in Mice with Reporter Enzyme Fluorescence

We describe the optical imaging of mice infected with Mycobacterium tuberculosis (M. tuberculosis) using reporter enzyme fluorescence (REF). This protocol facilitates the sensitive and specific detection of M. tuberculosis in pre-clinical animal models for pathogenesis, therapeutics and vaccine research.

报告酶荧光法检测小鼠结核分枝杆菌的影像学

Reporter enzyme fluorescence (REF) utilizes substrates that are specific for enzymes present in target organisms of interest for imaging or detection by ...

Determining Glucose Metabolism Kinetics Using 18F-FDG Micro-PET/CT

This paper describes the use of 18F-FDG and micro-PET/CT imaging to determine in vivo glucose metabolism kinetics in mice (and is transferable to rats). Impaired uptake and metabolism of glucose in multiple organ systems due to insulin resistance is a hallmark of type 2 diabetes. The ability of this technique to extract an image-derived input function from the vena cava using an iterative deconvolution method eliminates the requirement of the collection of arterial blood samples. Fitting of tissue and vena cava time activity curves to a two-tissue, three compartment model permits the estimation of kinetic micro-parameters related to the 18F-FDG uptake from the plasma to the intracellular space, the rate of transport from intracellular space to plasma and the rate of 18F-FDG phosphorylation. This methodology allows for multiple measures of glucose uptake and metabolism kinetics in the context of longitudinal studies and also provides insights into the efficacy of therapeutic interventions.

Determining Glucose Metabolism Kinetics Using 18F-FDG Micro-PET/CT

This study describes a protocol that uses 18F-FDG and positron emission tomography/computed tomography (PET/CT) imaging, together with kinetic modelling, to quantify the in vivo, real-time uptake of 18F-FDG into tissues.

确定葡萄糖代谢动力学研究<sup&gt; 18</sup&gt; F-FDG微PET / CT

This paper describes the use of 18F-FDG and micro-PET/CT imaging to determine in vivo glucose metabolism kinetics in mice (and is transferable to rats). ...

医学

Functional Imaging of Brown Fat in Mice with 18F-FDG micro-PET/CT

Brown adipose tissue (BAT) differs from white adipose tissue (WAT) by its discrete location and a brown-red color due to rich vascularization and high density of mitochondria. BAT plays a major role in energy expenditure and non-shivering thermogenesis in newborn mammals as well as the adults 1. BAT-mediated thermogenesis is highly regulated by the sympathetic nervous system, predominantly via &beta; adrenergic receptor 2, 3. Recent studies have shown that BAT activities in human adults are negatively correlated with body mass index (BMI) and other diabetic parameters 4-6. BAT has thus been proposed as a potential target for anti-obesity/anti-diabetes therapy focusing on modulation of energy balance 6-8. While several cold challenge-based positron emission tomography (PET) methods are established for detecting human BAT 9-13, there is essentially no standardized protocol for imaging and quantification of BAT in small animal models such as mice. Here we describe a robust PET/CT imaging method for functional assessment of BAT in mice. Briefly, adult C57BL/6J mice were cold treated under fasting conditions for a duration of 4 hours before they received one dose of 18F-Fluorodeoxyglucose (FDG). The mice were remained in the cold for one additional hour post FDG injection, and then scanned with a small animal-dedicated micro-PET/CT system. The acquired PET images were co-registered with the CT images for anatomical references and analyzed for FDG uptake in the interscapular BAT area to present BAT activity. This standardized cold-treatment and imaging protocol has been validated through testing BAT activities during pharmacological interventions, for example, the suppressed BAT activation by the treatment of &beta;-adrenoceptor antagonist propranolol 14, 15, or the enhanced BAT activation by &beta;3 agonist BRL37344 16. The method described here can be applied to screen for drugs/compounds that modulate BAT activity, or to identify genes/pathways that are involved in BAT development and regulation in various preclinical and basic studies.

Functional Imaging of Brown Fat in Mice with 18F-FDG micro-PET/CT

A method of functional imaging of mouse brown adipose tissue (BAT) is described in which cold-stimulated uptake of 18F-Fluorodeoxyglucose (FDG) in BAT is non-invasively assessed with a standardized micro-PET/CT protocol. This method is robust and sensitive to detect differences in BAT activities in mouse models.

功能成像在小鼠体内的棕色脂肪与FDG micro-PET/CT

Brown adipose tissue (BAT) differs from white adipose  tissue (WAT) by its discrete location and a brown-red color due to rich  vascularization and high ...

分析¹⁸葡萄糖 PET/CT 成像作为研究结核分枝杆菌感染和治疗非人类灵长类动物的工具

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使用身体顺应动物模具的临床前正电子发射断层扫描用于小鼠基于云的自动图像分析

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功能成像在小鼠体内的棕色脂肪与FDG micro-PET/CT