这项研究的经费来自国家癌症研究所 (R01CA166376 和 R01CA191489 D.X.) 和国防部 (W81XWH-16-1-0227 D.X. 阮) 的赠款。

所有实验都是按照耶鲁大学机构动物护理和使用委员会 (IACUC) 批准的协议进行的。
1. 试剂的设置/准备。
<ol><li>
博莱 霉 素 注意: 根据全球统一的化学品分类和标签制度, 博莱霉素被归类为 GHS08 的健康危害。<ol>
<li>用化学罩准备博莱霉素。并用重悬 15 U 入5毫升无菌磷酸盐缓冲盐水 (PBS)。</li>
		<li>整除100-200 µL 的溶液成玻璃瓶, 并冻结在-20 摄氏度, 以供将来使用。用泥沙的日期正确地标注管子。从该日期起6月内使用。 注意: 每个鼠标的应变对博莱霉素26有不同的敏感性。建议对不同剂量的博莱霉素进行小鼠菌株的检测。表 1列出了在代表性实验中使用的鼠株和博莱霉素的相应剂量。</li>
	</ol>
</li>
	<li>
小 鼠
	<ol>
<li>购买实验所需的小鼠, 并允许老鼠在注射前7天适应。在生物安全罩中进行输液。在博莱霉素输液前使用标准的机构程序, 将动物安置在 non-BSL2 兼容的房间里, BSL2 房间。在6-8 周的年龄注射老鼠。 警告: 在危险剂、生物安全等级 2 (BSL2) 和 IACUC 批准的机构指南下注射老鼠。 注: 为代表性实验而购买的鼠标列在材料表中。只有雄性老鼠被使用。</li>
	</ol>
</li>
	<li>
肺癌细胞系
	<ol>
<li>在各自的媒体中培养所需的细胞线。注射前, 进行短串联重复 DNA 分析或基因检测, 以确保正确的细胞线识别。为了促进体内和体外成像, 感染细胞系的慢病毒北疆表达胸苷激酶, 绿色荧光蛋白 (GFP) 和荧光素酶融合记者27, 并排序报告阳性细胞的荧光植入前的活化细胞分类。每6月支原体试验线。<ol>
<li>培养 H2030 人类癌细胞系的建议, 制造商使用罗斯威尔公园纪念研究所培养基 (RPMI) 与10% 胎牛血清 (血清), 青霉素-链霉素, 和两性霉素 B。</li>
			<li>培养368T1 小鼠细胞系 (从KrasG12D;p 53-/ 老鼠) 使用 Dulbecco 的改良鹰的培养基 (DMEM) 与10% 的血清, 青霉素-链霉素, 和两性霉素 B。</li>
			<li>PC9 人类癌细胞系的 RPMI, 采用10% 血清、青霉素-链霉素和两性霉素 B。</li>
		</ol>
</li>
	</ol>
</li>
</ol>2. 博莱霉素治疗
<ol><li>计划在肿瘤细胞植入前14天注射博莱霉素 intratracheally 小鼠。</li>
	<li>在注射前将博莱霉素在冰上解冻2小时。</li>
	<li>解冻后, 将博莱霉素稀释至所需的工作浓度 (0.02 U/50 µL 或 0.005 U/50 µL), 并将其放在冰上。 注: 博莱霉素浓度取决于实验用的小鼠应变 (表 1)。应进行小鼠博莱霉素滴定, 以确定最佳剂量。</li>
	<li>用1毫升注射器和27克针, 通过将氯胺酮和甲苯噻嗪稀释到最终浓度10毫克/毫升和1毫克/毫升, 通过注射腹腔氯胺酮/甲苯噻嗪溶液在100/10 毫克/千克上麻醉每只老鼠, 以制备麻醉药。分别。</li>
	<li>监测老鼠的呼吸, 并使用脚趾捏。确认正确的麻醉时, 鼠标不响应脚趾捏。在麻醉时, 应将兽医软膏涂抹于眼部, 以防干燥。</li>
	<li>放置1鼠标在一个插管平台上, 通过挂在它的前牙与它的背部对平台。</li>
	<li>用光纤照明灯照亮上胸部, 帮助可视化气管。张开鼠标的嘴, 用无菌的扁平钳轻轻地拉出舌头。</li>
	<li>使用静脉导管没有针头, 以避免阻塞呼吸。将导管放置在从气管开口处发出的白色光线上。</li>
	<li>将导管插入气管, 直到导管顶部到达前牙。通过在口腔内打开导管, 通过可视化白光照射, 确认导管在气管内的正确放置。</li>
	<li>使用吸管, 50 µL 的博莱霉素或车辆 (PBS) 直接进入导管, 以确保整个体积被吸入。在无菌条件下执行此步骤。</li>
	<li>如果导管正确插入, 鼠标将立即吸入导管的内容。等待几秒钟, 直到整个音量沿着导管移动。然后从气管取出导管, 在10% 漂白剂溶液中处理。 注: 每个步骤 2.7-2.11 的鼠标平均时间约为5分钟。</li>
	<li>如果鼠标不吸入液体, 仔细监测呼吸并调整导管位置。如果鼠标停止呼吸, 立即取出导管, 并允许鼠标在重新插入导管前正常恢复呼吸。</li>
	<li>将注入的鼠标放在 IACUC 上的已批准的加热垫上, 然后放在平坦的表面上进行恢复。整个过程将采取每只鼠标10分钟。在完全恢复之前, 不要将已经注射到其他动物公司的动物退回。</li>
	<li>将 "危险化学品使用" 卡放在笼子里24小时。</li>
</ol>3. 在插管后监测小鼠
<ol><li>在插管后立即监测小鼠呼吸窘迫, 然后每15分钟, 直到老鼠从麻醉中醒来。不要让老鼠无人看管, 直到他们恢复足够的意识来维持胸骨卧床。</li>
	<li>注射酮酮 (5 毫克/千克) 腹腔减轻疼痛。</li>
	<li>检查小鼠 24 h 后插管和每周3次的任何发病率的迹象。</li>
	<li>记录程序的日期、动物身份、程序类型、麻醉类型和术后监测观察时间。</li>
	<li>监测小鼠的体重减轻, 呼吸窘迫, 行为异常, 身体状况评分 < 2 (椎柱明显/背骨盆骨分割) 双周后注射博莱霉素。</li>
	<li>弄死小鼠在呼吸窘迫或小鼠经历了15% 的身体质量损失由 CO2或氯胺酮/甲苯噻嗪后, 颈椎脱位。通过对呼吸活动进行触诊确认安乐死。</li>
</ol>4. 植入肺腺癌细胞系。
注: 在注射博莱霉素14天后进行细胞植入 (步骤 2.1)。
<ol><li>允许细胞生长在75厘米2瓶不受干扰的3天前的注射与相应的媒体, 如步骤1.3 所述。 注: 在注射的当天, 它们应该是80% 汇合的 (在每个烧瓶中对应于 2-3 x 106 , 这取决于细胞线)。</li>
	<li>用5毫升的 PBS 在室温下冲洗75厘米2烧瓶上生长的细胞。</li>
	<li>吸入 PBS 和添加1.5 毫升胰蛋白酶到细胞和孵化细胞在37°c, 直到细胞分离 (通常2-5 分钟)。 注: 我们建议每隔2分钟检查从塑料的细胞脱离, 通过简单的可视化烧瓶底部或在光显微镜下。不要超过5分钟的孵化与胰蛋白酶。</li>
	<li>加入4毫升含有10% 种血清的培养基 (与步骤4.1 相同的培养基) 中和胰蛋白酶。将细胞收集成15毫升管, 在室温下将其离心在 200 x g 处3分钟。</li>
	<li>在5毫升的 PBS 中吸入培养基并并用重悬细胞颗粒以洗涤细胞。在室温下将细胞以 200 x g 为3分钟, 在颗粒细胞中进行离心。重复这1次。</li>
	<li>吸入 pbs 和并用重悬1.5 毫升的 pbs 每瓶。</li>
	<li>将50µL 的细胞混合物与50µL 的台盼蓝混合, 用细胞计数器/Hemocytometer 室28计算细胞数。</li>
	<li>通过稀释 PBS 中的细胞来制备细胞悬浮液, 在每只老鼠的50µL 中注射 1 x 105细胞 (或其他指示量)。在注射前把细胞放在冰上。 注: 准备至少两次更多的细胞注射, 以避免耗尽细胞悬浮。</li>
	<li>用1毫升注射器和27克针, 通过将氯胺酮和甲苯噻嗪稀释到最终浓度10毫克/毫升和1毫克/毫升, 通过注射腹腔氯胺酮/甲苯噻嗪溶液在100/10 毫克/千克上麻醉每只老鼠, 以制备麻醉药。分别。</li>
	<li>监测老鼠的呼吸, 并使用脚趾捏。确认正确的麻醉时, 鼠标不响应脚趾捏。在麻醉时, 应将兽医软膏涂抹于眼部, 以防干燥。</li>
	<li>放置1鼠标在一个插管平台上, 通过挂在它的前牙与它的背部对平台。</li>
	<li>用光纤照明灯照亮上胸部, 帮助可视化气管。</li>
	<li>用 p200 吸管轻轻地并用重悬细胞, 以确保它们不形成团簇。张开鼠标的嘴, 用无菌的扁平钳轻轻地拉出舌头。</li>
	<li>使用静脉导管与针移除, 以避免阻塞呼吸。将导管放置在从气管开口处发出的白色光线上。</li>
	<li>将导管插入气管, 直到导管顶部到达前牙。通过在口腔内打开导管, 通过可视化白光照射, 确认导管在气管内的正确放置。</li>
	<li>使用吸管, 将50µL 的细胞直接放入导管中, 以确保整个体积被吸入。在无菌条件下执行此步骤。 注: 如果导管正确插入, 鼠标将立即吸入导管的内容。</li>
	<li>等待几秒钟, 直到整个音量沿着导管移动, 然后从气管中取出导管, 并在10% 漂白剂溶液中处理。</li>
	<li>如果鼠标不吸入液体, 仔细监测呼吸并调整导管位置。如果鼠标停止呼吸, 立即取出导管, 并允许鼠标在重新插入导管前正常恢复呼吸。 注: 步骤 4.11-4. 18 的每个鼠标的平均时间约为5分钟。</li>
	<li>将注入的鼠标放在 IACUC 上的已批准的加热垫上, 然后放在平坦的表面上进行恢复。 注: 整个过程将采取10分钟每鼠标。在完全恢复之前, 不要将已经注射到其他动物公司的动物退回。</li>
	<li>在成像前用电动剃须刀剃掉整个肋区的腹部和背的 B6129SF1/J 和其他小鼠的黑头发。</li>
	<li>2-3 分钟后细胞注射, 注射100µL 荧光素复古轨道 (15 毫克/毫升) 使用胰岛素针。在每次成像会话中交替使用用于注射的眼睛。</li>
	<li>在至少2分钟后, 在一个动物生物发光成像仪中放置多达5只老鼠, 在背卧床位置, 并获得一个侧面图片使用发光设置29。</li>
	<li>在控制面板下调整分辨率和灵敏度设置, 以测量给定细胞线的发光。对于大多数应用程序, 从3分钟 (或 "自动" 如果饱和) 开始, binning = 中等 (4), F/Stop=1, 视野 = D, 主题高度 = 1.50。 注: 如果注射成功, 在上胸区, 在一个肺或两个肺部检测到发光信号。</li>
	<li>翻转小鼠到胸骨卧床位置, 并获得一个背部图片如上所述。 注: 如果细胞注射未正确执行, 细胞可能会聚集在气管入口或颈部。</li>
	<li>注射酮酮5毫克/千克腹腔减轻疼痛。</li>
	<li>把老鼠搬回笼子里, 在笼子里放一张 "BSL-2 剂"。</li>
	<li>在插管后立即监测小鼠呼吸窘迫, 然后每隔15分钟, 直到老鼠从麻醉中醒来。不要让老鼠无人看管, 直到他们恢复足够的意识来维持胸骨卧床。</li>
	<li>检查小鼠 24 h 后插管和每周三次的任何发病率的迹象。</li>
	<li>记录在程序日期, 动物身份, 程序类型, 麻醉类型和术后监测观察和时间的日志。</li>
	<li>监测小鼠的体重减轻, 呼吸窘迫, 行为异常, 身体状况评分 < 2 (脊柱明显/背骨盆骨分割可见) 两周后接种的博莱霉素。 注: 肺肿瘤小鼠可能表现为呼吸刺激、体重减轻、恶病质和/或死亡。</li>
	<li>弄死小鼠在呼吸窘迫或小鼠经历了15% 的身体质量损失由 CO2或氯胺酮/甲苯噻嗪后, 颈部脱位, 如果收集组织。通过对呼吸活动进行触诊确认安乐死。</li>
</ol>5. 生物发光成像对肿瘤生长的监测
<ol><li>重复成像的老鼠在3天后植入和每周。</li>
	<li>麻醉小鼠注射氯胺酮/甲苯噻嗪 (如步骤 2.4-2.5 中所述) 或吸入异氟醚 (2%)。监测小鼠的呼吸, 并使用脚趾捏来确认正确的麻醉。在麻醉时, 应将兽医软膏涂抹于眼部, 以防干燥。</li>
	<li>在成像前用电动剃须刀剃掉整个肋区的腹部和背的 B6129SF1/J 和其他小鼠的黑头发。</li>
	<li>一旦老鼠被麻醉, 注射100µL 的荧光素轨道 (15 毫克/毫升) 使用胰岛素针。等待2分钟交替的眼睛用于注射每一个成像会话。</li>
	<li>将鼠标放在成像仪中, 并获得4.22-4 步24中所述的背侧和腹侧图像。</li>
	<li>在成像后将老鼠放回笼子里, 在他们的背部在平坦的表面恢复。 注: 整个手术过程每组5只老鼠需要5分钟。不要让老鼠无人看管, 直到他们恢复足够的意识来维持胸骨卧床。</li>
	<li>利用成像/生物发光分析软件对图像进行分析。<ol>
<li>选择适当的图像并将它们作为一个组加载到生物发光分析软件中。</li>
		<li>单击roi 工具 | 正方形以添加感兴趣的正方形区域 (roi)。将 ROI 放在上胸部区域, 覆盖整个肺部图像。对每个鼠标图像重复此操作。</li>
		<li>右键单击并选择 "复制所有 roi函数" 将相同的 roi 应用于组中的所有图像, 并将它们放置在鼠标的上胸部, 无论是腹侧视图还是背景色。</li>
		<li>在图像窗口的左上角, 选择光子函数。单击测量控制面板函数以测量 ROIs。将包含总通量 (光子/s) 的输出表复制并粘贴到一个可供选择的软件中, 以便进一步分析数据。</li>
		<li>将每个鼠标的每日 roi 值正常化为其在0天测量的 roi 值。</li>
	</ol>
</li>
</ol>6. 组织分离和加工。
<ol><li>麻醉小鼠注射氯胺酮/甲苯噻嗪 (如步骤 2.4-2.5 或吸入异氟醚 (2%) 所述。监测老鼠的呼吸, 并使用脚趾捏。确认正确的麻醉时, 鼠标不响应脚趾捏。在麻醉时, 应将兽医软膏涂抹于眼部, 以防干燥。</li>
	<li>用胰岛素针注射100µL 荧光素复古轨道 (15 毫克/毫升)。等待2分钟。</li>
	<li>图象老鼠跟随步骤 4.22-4. 24 从这个协议。</li>
	<li>注射到另一只眼睛从步骤6.2 另外100µL 的荧光素复古轨道使用胰岛素针之前, 安乐死。</li>
	<li>弄死小鼠腹腔注射氯胺酮/甲苯噻嗪 (见步骤2.4 的细节), 其次是颈椎脱位, 如果小鼠在深度麻醉下, 根据 IACUC 提供的指导方针。通过对呼吸活动进行触诊确认安乐死。</li>
	<li>使用无菌手术剪刀在胸骨下面做一个皮肤切口。用镊子和手术剪刀在隔膜上打开肌肉层, 通过在两侧两侧的肋骨笼上切开胸腔, 避免胸腔内动脉。</li>
	<li>灌注通过在心脏右心房做一个小切口, 并通过左心室注入5毫升 PBS。如果血液灌注得当, 肺组织的颜色会从红到浅粉色白色。</li>
	<li>用镊子抓住食道或心脏, 仔细地将肺部从胸腔中切除。轻轻拉组织, 并使用手术剪刀仔细削减连接组织避免穿刺的肺组织。在以后的一步中尽量保持气管的膨胀。</li>
	<li>通过浸没和旋转组织3倍的6井板填充3毫升的 PBS, 以清除多余的血液, 洗肺。</li>
	<li>将肺放在6井板的盖子上, 将含有组织的盖子放在发光成像仪中, 并获得发光图像30。 注: 我们建议从 "采集控制面板" "视图 = A" 和 "主题高度 = 0.75 厘米" 中选择以下设置, 然后 "自动曝光" 以避免获得饱和图像。</li>
	<li>消费和图像其他器官, 如步骤 6.9-6.10 中所做的, 其中转移已被发现的发光, 如脑, 肝, 肾上腺, 骨, 淋巴结, 肾脏或脾脏31。</li>
	<li>灌注1毫升4% 多聚甲醛的肺, 将针头插入鼠标的气管内。如果灌注得好, 肺部会膨胀。 警告: 4% 多聚甲醛是一种毒性、健康危害 GHS07 和 GHS08。</li>
	<li>将肺部转移到15毫升圆锥, 5 毫升4% 多聚甲醛。</li>
	<li>修复肺或其他组织通过孵化的组织与4% 多聚甲醛隔夜在4°c 在一个震动装置在 30-50 rpm。</li>
	<li>根据应用32, 在石蜡或最佳切削温度化合物中嵌入肺部 (或其他组织)。 注: 石蜡是保存肺结构和马尾松三色和苏木精-伊红染色的首选方法。</li>
</ol>

<ol>
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作者声明没有竞争的财政利益。

在肺癌和其他慢性疾病的肺36之间有明显的临床相似之处。尤其是特发性肺纤维化 (IPF) 患者对发展肺癌的偏爱增加, 而这个协会独立于吸烟史37,38。森林小组的特点是逐步破坏肺结构和受损的呼吸功能, 通过沉积 ECM39。同时, 在手术切除后, 早期的无细胞肺癌患者并发森林小组的结局较差40。多数 NSCLC ?...

肺癌是全球1癌症相关死亡的主要原因。肺癌患者最终从转移到远端器官, 主要是中枢神经系统, 肝脏, 肾上腺, 骨骼2,3,4。胸恶性肿瘤历来被归类为小细胞肺癌或非小细胞肺癌 (NSCLC)5。NSCLC 是最常见的恶性肿瘤, 可分为不同的组织学亚型, 包括肺腺癌 (LUAD) 和肺鳞状细胞癌 (LUSC)6。经手术切除的人原发性肺癌的基因组分析显示, 某一 histotype 内的肿瘤也能表达多种分子扰动, 进一步促进其临床进展和混淆患者预后。肺癌的显著异质性对合理的设计、临床前试验和有效的治疗策略的实施都是一个重大挑战。因此, 有必要扩大可听话的实验性肺癌模型的汇辑, 以研究这种疾病的不同细胞来源、分子亚型和分期。
使用动物模型的各种方法在体内研究肺癌, 每个都有各自的优缺点, 这取决于所关心的生物学问题。基因工程小鼠模型 (GEMMs) 可以针对特定祖细胞类型的特异基因改变, 导致肿瘤在免疫宿主7中取得进展。虽然非常强大和临床相关的, 潜伏期, 变异性和/或肺部肿瘤发病率与 GEMMs 可能会禁止某些定量测量和检测晚期转移在遥远的器官8。一个补充的方法是使用同种异体模型, 藉以肺癌细胞从老鼠肿瘤直接获得或首先被获取作为建立的细胞线在文化中被重新介绍入自体寄主。类似, 肺癌移植是由人体细胞系或病人衍生的肿瘤样本建立的。人细胞系移植或患者衍生的移植 (PDXs) 一般维持在免疫缺陷小鼠, 因此排除完全免疫监测9。尽管有这个缺点, 它们提供了一个途径来传播限制数量的人类 biospecimens 和研究人类癌细胞的基本的体内性质, 这编码为更复杂的基因组畸变比 GEMM 肿瘤。
同种异体移植和异种移植的一个有用的特性是, 他们可以使用传统的限制性细胞稀释方法, 用于量化10恶性细胞中肿瘤起始细胞 (抽搐) 的频率。在这些实验中, 在动物的侧面注入了一定数量的细胞, 并根据肿瘤的发生率来估计抽搐频率。然而皮下肿瘤可以是更低氧11并且可能不模型主要生理制约肺肿瘤微环境。气管内向小鼠肺内输送上皮干细胞或祖细胞是研究肺再生和气道干细胞生物学12的方法。然而, 这种技术的植入率可能相对较低, 除非肺部首先受到生理形式的伤害, 如病毒感染13,14。炎症基质细胞的支持和 (或) 肺基底膜的破坏, 可改善移植细胞在远端气道15的相关干细胞龛位的保留。纤维化诱导剂也可以预条件肺, 以增强植入诱导多能细胞16和间充质干细胞17。类似的气道损伤形式是否会影响植入率, 肿瘤的启动能力和肺癌细胞的产生还有待系统地评估。
本文通过对损伤小鼠肺的预处理, 描述了提高原位肺癌细胞植入效率的方法。LUAD 出现在远端呼吸道与这些癌症的一个重要的子集发展纤维间质基质18 , 往往与不良预后19。博莱霉素是一种天然非核糖体杂交肽-聚酮体, 已广泛用于诱导小鼠肺纤维化20。博莱霉素的气道灌注首先促进肺泡上皮的磨损和炎症细胞的招募, 包括巨噬细胞、中性粒细胞和单核细胞21。其次是在远端呼吸道组织重塑, 基底膜重组22,23和细胞外基质 (ECM) 沉积24。单个博莱霉素注射液的影响是瞬变的, 在大多数研究中30天后纤维化解决25。利用同种异体移植和异种移植模型, 我们对博莱霉素小鼠呼吸道进行预适应试验, 可以显著提高肺 LUAD 细胞的服用率。

<table>
	<tbody>
		<tr>
			<td>Bleomycin</td>
			<td>Sigma</td>
			<td>B5507-15UN</td>
			<td>CAUTION Health hazard GHS08</td>
		</tr>
		<tr>
			<td>Exel Catheter 24G</td>
			<td>Fisher</td>
			<td>1484121</td>
			<td>Remove needle. For intratracheal injection</td>
		</tr>
		<tr>
			<td>Ketamine (Ketaset inl 100 mg/mL C3N 10 mL)</td>
			<td>Butler Schein</td>
			<td>56344</td>
			<td>To anesthetize mice</td>
		</tr>
		<tr>
			<td>Xylazine</td>
			<td>Butler Schein</td>
			<td>33198</td>
			<td>To anesthetize mice</td>
		</tr>
		<tr>
			<td>Ketoprofen, 5,000 mg</td>
			<td>Cayman Chemical</td>
			<td>10006661</td>
			<td>Analgesic</td>
		</tr>
		<tr>
			<td>Puralube Veterinary Ophthalmic Ointment</td>
			<td>BUTLER ANIMAL HEALTH COMPANY LLC</td>
			<td>8897</td>
			<td>To prevent eye dryness while under anesthesia</td>
		</tr>
		<tr>
			<td>D-Luciferin powder</td>
			<td>Perkin Elmer Health Sciences Inc</td>
			<td>122799</td>
			<td>For luminescent imaging. Reconstitute powder with PBS for a working concentration of 15mg/mL. Protect from Light</td>
		</tr>
		<tr>
			<td>Rodent Intubation stand</td>
			<td>Braintree Scientific</td>
			<td>RIS-100</td>
			<td>Recommended stand for intratracheal injection</td>
		</tr>
		<tr>
			<td>MI-150 ILLUMINATOR 150W MI-150</td>
			<td>DOLAN-JENNER INDUSTRIES</td>
			<td>MI-150 / EEG2823M</td>
			<td>To illuminate and visualize trachea</td>
		</tr>
		<tr>
			<td>Graefe Forceps, 2.75 (7 cm) long serrat</td>
			<td>Roboz</td>
			<td>RS-5111</td>
			<td>For intratracheal injection</td>
		</tr>
		<tr>
			<td>Syringe Luer-Lok Sterile 5ml</td>
			<td>BD / Fisher</td>
			<td>309646</td>
			<td></td>
		</tr>
		<tr>
			<td>Satiny Smooth by Conair Dual Foil Wet/Dry Rechargeable Shaver</td>
			<td>Conair</td>
			<td>-</td>
			<td>To shave mice</td>
		</tr>
		<tr>
			<td>Bonn Scissors, 3.5&#34; straight 15 mm sharp/sharp sure cut blades</td>
			<td>Roboz</td>
			<td>RS-5840SC</td>
			<td></td>
		</tr>
		<tr>
			<td>15 mL conical tube</td>
			<td>BD / Fisher</td>
			<td>352097</td>
			<td></td>
		</tr>
		<tr>
			<td>1.5 mL centrifuge tubes</td>
			<td>USA SCIENTIFIC INC</td>
			<td>1615-5500</td>
			<td></td>
		</tr>
		<tr>
			<td>Vial Scintillation 7 mL Borosilicate Glass GPI</td>
			<td>Fisher</td>
			<td>701350</td>
			<td></td>
		</tr>
		<tr>
			<td>Filter pipette tips (200 &#956;L)</td>
			<td>USA SCIENTIFIC INC</td>
			<td>1120-8710</td>
			<td></td>
		</tr>
		<tr>
			<td>Phosphate Buffered Saline</td>
			<td>Life Technologies</td>
			<td>14190-144</td>
			<td></td>
		</tr>
		<tr>
			<td>0.25% Trypsin-EDTA</td>
			<td>Life Technologies</td>
			<td>25200-056</td>
			<td></td>
		</tr>
		<tr>
			<td>DMEM high glucose</td>
			<td>Life Technologies</td>
			<td>11965-092</td>
			<td></td>
		</tr>
		<tr>
			<td>RPMI Medium 1640</td>
			<td>Life Technologies</td>
			<td>11875-093</td>
			<td></td>
		</tr>
		<tr>
			<td>Fetal bovine serum USDA</td>
			<td>Life Technologies</td>
			<td>10437-028</td>
			<td></td>
		</tr>
		<tr>
			<td>Penicillin-Streptomycin</td>
			<td>Life Technologies</td>
			<td>15140-122</td>
			<td></td>
		</tr>
		<tr>
			<td>Amphotericin B</td>
			<td>Sigma</td>
			<td>A2942-20ML</td>
			<td></td>
		</tr>
		<tr>
			<td>Trypan Blue Stain 0.4%</td>
			<td>Life Technologies</td>
			<td>15250-061</td>
			<td></td>
		</tr>
		<tr>
			<td>Countess Automated Cell Counter</td>
			<td>Life Technologies</td>
			<td>AMQAX1000</td>
			<td></td>
		</tr>
		<tr>
			<td>Flask T/C 75cm sq canted neck, blue cap</td>
			<td>Fisher / Corning</td>
			<td>353135</td>
			<td></td>
		</tr>
		<tr>
			<td>IVIS Spectrum Xenogen Bioluminiscence</td>
			<td>Perkin Elmer Health Sciences Inc</td>
			<td>124262</td>
			<td>For in vivo bioluminescence imaging</td>
		</tr>
		<tr>
			<td>Living image software</td>
			<td>Perkin Elmer Health Sciences Inc</td>
			<td>128113</td>
			<td>For in vivo bioluminescence analysis</td>
		</tr>
		<tr>
			<td>XGI-8 Gas Anesthesia System</td>
			<td>Perkin Elmer Health Sciences Inc</td>
			<td>118918</td>
			<td>For Isoflurane anesthesia</td>
		</tr>
		<tr>
			<td>BD Ultra-Fine II Short Needle Insulin Syringe 1 cc. 31 G x 8 mm (5/16 in)</td>
			<td>BD / Fisher</td>
			<td>BD328418</td>
			<td>For retro-orbital luciferin injection</td>
		</tr>
		<tr>
			<td>Syringe 1ml</td>
			<td>BD / Fisher</td>
			<td>14-823-434</td>
			<td>For intraperitoneal injections</td>
		</tr>
		<tr>
			<td>26 G x 1/2 in. needle</td>
			<td>BD / Fisher</td>
			<td>305111</td>
			<td>For intraperitoneal injections</td>
		</tr>
		<tr>
			<td>4% Paraformaldehyde</td>
			<td>VWR</td>
			<td>43368-9M</td>
			<td>CAUTION Health hazard GHS07, GHS08. For fixing tissue</td>
		</tr>
		<tr>
			<td>Pipet-Lite Pipette, Unv. SL-200XLS+</td>
			<td>METTLER-TOLEDO INTERNATIONAL</td>
			<td>17014411</td>
			<td></td>
		</tr>
		<tr>
			<td>Mayer&#39;s Hematoxylin</td>
			<td>ELECTRON MICROSCOPY SCIENCES</td>
			<td>517-28-2</td>
			<td></td>
		</tr>
		<tr>
			<td>Eosin Y stain 0.25% (w/v) in 57%</td>
			<td>Fisher</td>
			<td>67-63-0</td>
			<td></td>
		</tr>
		<tr>
			<td>Masson Trichrome Stain Kit</td>
			<td>IMEB Inc</td>
			<td>K7228</td>
			<td>For masson trichrome stain to visualize collagen</td>
		</tr>
		<tr>
			<td>Superfrost plus glass slides</td>
			<td>Fisher</td>
			<td>1255015</td>
			<td></td>
		</tr>
		<tr>
			<td>6 well plate</td>
			<td>Corning</td>
			<td>C3516</td>
			<td></td>
		</tr>
		<tr>
			<td>Universal Mycoplasma Detection Kit</td>
			<td>ATCC</td>
			<td>30-1012K</td>
			<td></td>
		</tr>
		<tr>
			<td>OCT Embedding compound</td>
			<td>ELECTRON MICROSCOPY SCIENCES</td>
			<td>62550-12</td>
			<td>For embedding tissue for frozen sections</td>
		</tr>
		<tr>
			<td>Leica CM3050 S Research Cryostat</td>
			<td>Leica</td>
			<td>CM3050 S</td>
			<td>To section tissue for staining analysis</td>
		</tr>
		<tr>
			<td>Keyence All-in One Fluorescence Microscope</td>
			<td>Keyence</td>
			<td>BZ-X700</td>
			<td></td>
		</tr>
		<tr>
			<td>ImageJ</td>
			<td>US National Institutes of Health</td>
			<td>IJ1.46</td>
			<td>http://rsbweb.nih.gov/ij/ download.html</td>
		</tr>
		<tr>
			<td>Prism 7.0 for Mac OS X</td>
			<td>GraphPad Software, Inc.</td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>Athymic (Crl:NU(NCr)-Foxn1nu) mice</td>
			<td>Charles River</td>
			<td>NIH-553</td>
			<td></td>
		</tr>
		<tr>
			<td>NSG (NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ) mice</td>
			<td>Jackson Laboratories</td>
			<td>5557</td>
			<td></td>
		</tr>
		<tr>
			<td>B6129SF1/J mice</td>
			<td>Jackson Laboratories</td>
			<td>101043</td>
			<td></td>
		</tr>
		<tr>
			<td>NIH-H2030 cells</td>
			<td>ATCC</td>
			<td>CRL-5914</td>
			<td></td>
		</tr>
		<tr>
			<td>368T1</td>
			<td>generously provided by Monte Winslow (Standford University)</td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>PC9 cells</td>
			<td>Nguyen DX et al. Cell. 2009;138:51&#8211;62</td>
			<td>-</td>
			<td></td>
		</tr>
		<tr>
			<td>H2030 BrM3 cells</td>
			<td>Nguyen DX et al. Cell. 2009;138:51&#8211;62</td>
			<td>-</td>
			<td></td>
		</tr>
	</tbody>
</table>

pre-conditioning the airways of mice with bleomycin increases the efficiency of orthotopic lung cancer cell engraftment

肺癌是一种致命的治疗顽固性疾病, 是生物异构的。为了了解和有效地治疗胸恶性肿瘤的完整临床谱, 需要额外的动物模型来重述不同的人肺癌亚型和分期。同种异体或异种移植模型是多种多样的, 可以利用小鼠或人类的恶性细胞,在体内癌变能力的量化。然而, 以前所描述的肺癌细胞植入的方法已经在非生理部位进行, 如小鼠的侧面, 由于细胞移植到肺部的效率低下。在本研究中, 我们描述了一种通过对小鼠肺纤维化诱发博莱霉素进行预调节来增强原位肺癌细胞植入的方法。作为一种概念验证实验, 我们将这种方法应用于从老鼠或人源获得的肺腺癌亚型的嫁接肿瘤细胞, 并将其转化为不同品系的小鼠。我们证明, 在肿瘤细胞注射前, 与博莱霉素的呼吸道损伤增加了肿瘤细胞从0-17% 到71-100% 的植入。显著地, 该方法通过不同模型和小鼠菌株增强肺肿瘤发生率和后续生长。此外, 嫁接肺癌细胞从肺部传播到相关的远端器官。因此, 我们提供了一个协议, 可用于建立和维持新的肺癌原位模型, 限制数量的细胞或 biospecimen, 并定量评估肺癌细胞在生理相关设置的癌变能力.

为了提高 LUAD 癌细胞植入到小鼠肺中的效率, 我们制定了一项协议, 首先使用博莱霉素的呼吸道, 其次是原位肿瘤细胞注射 (图 1)。我们证实, 即使在免疫裸小鼠, 博莱霉素诱导瞬变纤维化14天, 证明了气道结构的损失和增加胶原沉积 (图 2)。这些小鼠的毛纤维化在博莱霉素注射液后50天解决 (图 2; 右面板) 与以...

Watch this Scientific Journal Video about Pre-Conditioning the Airways of Mice with Bleomycin Increases the Efficiency of Orthotopic Lung Cancer Cell Engraftment at JoVE.com

Pre-Conditioning the Airways of Mice with Bleomycin Increases the Efficiency of Orthotopic Lung Cancer Cell Engraftment

本文介绍一种通过对气道损伤进行预适应来显著提高肺癌细胞原位植入的方法。这种方法也可以应用于研究肺微环境中的间质相互作用, 转移传播, 肺癌的共同发病, 并更有效地产生病人的移植。

博莱霉素对小鼠气管的预处理提高原位肺癌细胞植入的效率

Lung cancer is a deadly treatment refractory disease that is biologically heterogeneous. To understand and effectively treat the full clinical spectrum of ...

pre-conditioning-airways-mice-with-bleomycin-increases-efficiency

Universidad de Cartagena UDEC

Research

JoVE Journal

Medicine

13.5K 次观看.  Yale University School of Medicine. 本文介绍一种通过对气道损伤进行预适应来显著提高肺癌细胞原位植入的方法。这种方法也可以应用于研究肺微环境中的间质相互作用, 转移传播, 肺癌的共同发病, 并更有效地产生病人的移植。

视频: Pre-Conditioning the Airways of Mice with Bleomycin Increases the Efficiency of Orthotopic Lung Cancer Cell Engraftment

Bronchial Thermoplasty:  A Novel Therapeutic Approach to Severe Asthma

Bronchial thermoplasty is a non-drug procedure for severe persistent asthma that delivers thermal energy to the airway wall in a precisely controlled manner to reduce excessive airway smooth muscle. Reducing airway smooth muscle decreases the ability of the airways to constrict, thereby reducing the frequency of asthma attacks. Bronchial thermoplasty is delivered by the Alair System and is performed in three outpatient procedure visits, each scheduled approximately three weeks apart. The first procedure treats the airways of the right lower lobe, the second treats the airways of the left lower lobe and the third and final procedure treats the airways in both upper lobes. After all three procedures are performed the bronchial thermoplasty treatment is complete.
Bronchial thermoplasty is performed during bronchoscopy with the patient under moderate sedation. All accessible airways distal to the mainstem bronchi between 3 and 10 mm in diameter, with the exception of the right middle lobe, are treated under bronchoscopic visualization. Contiguous and non-overlapping activations of the device are used, moving from distal to proximal along the length of the airway, and systematically from airway to airway as described previously. Although conceptually straightforward, the actual execution of bronchial thermoplasty is quite intricate and procedural duration for the treatment of a single lobe is often substantially longer than encountered during routine bronchoscopy. As such, bronchial thermoplasty should be considered a complex interventional bronchoscopy and is intended for the experienced bronchoscopist. Optimal patient management is critical in any such complex and longer duration bronchoscopic procedure. This article discusses the importance of careful patient selection, patient preparation, patient management, procedure duration, postoperative care and follow-up to ensure that bronchial thermoplasty is performed safely.
Bronchial thermoplasty is expected to complement asthma maintenance medications by providing long-lasting asthma control and improving asthma-related quality of life of patients with severe asthma. In addition, bronchial thermoplasty has been demonstrated to reduce severe exacerbations (asthma attacks) emergency rooms visits for respiratory symptoms, and time lost from work, school and other daily activities due to asthma.

Bronchial thermoplasty is a non-drug procedure for severe persistent asthma that delivers thermal energy to the airway wall in a precisely controlled manner to reduce excessive airway smooth muscle. Reducing airway smooth muscle decreases the ability of the airways to constrict, thereby reducing the frequency of asthma attacks.

支气管Thermoplasty:严重哮喘的一种新的治疗方法

Bronchial thermoplasty is a non-drug procedure for severe persistent asthma that delivers thermal energy to the airway wall in a precisely controlled ...

科研

医学

An Orthotopic Model of Murine Bladder Cancer

In this straightforward procedure, bladder tumors are established in female C57 mice through the use of catheterization, local cauterization, and subsequent cell adhesion. After their bladders are transurethrally catheterized and drained, animals are again catheterized to permit insertion of a platinum wire into bladders without damaging the urethra or bladder. The catheters are made of Teflon to serve as an insulator for the wire, which will conduct electrical current into the bladder to create a burn injury. An electrocautery unit is used to deliver 2.5W to the exposed end of the wire, burning away extracellular layers and providing attachment sites for carcinoma cells that are delivered in suspension to the bladder through a subsequent catheterization. Cells remain in the bladder for 90 minutes, after which the catheters are removed and the bladders allowed to drain naturally. The development of tumor is monitored via ultrasound. Specific attention is paid to the catheterization technique in the accompanying video.

Bladder tumors are established in female mice in a minimally invasive fashion through catheterization, local cauterization, and subsequent adhesion of carcinoma cells to the burn sites.

一个小鼠膀胱癌原位模型

In this straightforward procedure, bladder tumors are established in female C57 mice through the use of catheterization, local cauterization, and ...

Multi-photon Imaging of Tumor Cell Invasion in an Orthotopic Mouse Model of Oral Squamous Cell Carcinoma

Loco-regional invasion of head and neck cancer is linked to metastatic risk and presents a difficult challenge in designing and implementing patient management strategies. Orthotopic mouse models of oral cancer have been developed to facilitate the study of factors that impact invasion and serve as model system for evaluating anti-tumor therapeutics. In these systems, visualization of disseminated tumor cells within oral cavity tissues has typically been conducted by either conventional histology or with in vivo bioluminescent methods. A primary drawback of these techniques is the inherent inability to accurately visualize and quantify early tumor cell invasion arising from the primary site in three dimensions. Here we describe a protocol that combines an established model for squamous cell carcinoma of the tongue (SCOT) with two-photon imaging to allow multi-vectorial visualization of lingual tumor spread. The OSC-19 head and neck tumor cell line was stably engineered to express the F-actin binding peptide LifeAct fused to the mCherry fluorescent protein (LifeAct-mCherry). Fox1nu/nu mice injected with these cells reliably form tumors that allow the tongue to be visualized by ex-vivo application of two-photon microscopy. This technique allows for the orthotopic visualization of the tumor mass and locally invading cells in excised tongues without disruption of the regional tumor microenvironment. In addition, this system allows for the quantification of tumor cell invasion by calculating distances that invaded cells move from the primary tumor site. Overall this procedure provides an enhanced model system for analyzing factors that contribute to SCOT invasion and therapeutic treatments tailored to prevent local invasion and distant metastatic spread. This method also has the potential to be ultimately combined with other imaging modalities in an in vivo setting.

A comprehensive overview of the techniques involved in generating a mouse model of oral cancer and quantitative monitoring of tumor invasion within the tongue through multi-photon microscopy of labeled cells is presented. This system can serve as a useful platform for the molecular assessment and drug efficacy of anti-invasive compounds.

多光子成像在口腔鳞状细胞癌原位小鼠模型的肿瘤细胞的侵袭

Loco-regional invasion of head and neck cancer is linked to metastatic risk and presents a difficult challenge in designing and implementing patient ...

An Orthotopic Bladder Tumor Model and the Evaluation of Intravesical saRNA Treatment

We present a novel method for treating bladder cancer with intravesically delivered small activating RNA (saRNA) in an orthotopic xenograft mouse bladder tumor model. The mouse model is established by urethral catheterization under inhaled general anesthetic. Chemical burn is then introduced to the bladder mucosa using intravesical silver nitrate solution to disrupt the bladder glycosaminoglycan layer and allows cells to attach. Following several washes with sterile water, human bladder cancer KU-7-luc2-GFP cells are instilled through the catheter into the bladder to dwell for 2 hours. Subsequent growth of bladder tumors is confirmed and monitored by in vivo bladder ultrasound and bioluminescent imaging. The tumors are then treated intravesically with saRNA formulated in lipid nanoparticles (LNPs). Tumor growth is monitored with ultrasound and bioluminescence. All steps of this procedure are demonstrated in the accompanying video.

Establishing an orthotopic bladder tumor model to evaluate antitumor effects of intravesically delivered saRNA and monitoring tumor growth by ultrasound and bioluminescent imaging.

原位膀胱肿瘤模型和评价治疗膀胱萨纳

We present a novel method for treating bladder cancer with intravesically delivered small activating RNA (saRNA) in an orthotopic xenograft mouse bladder ...

Orthotopic Aortic Transplantation in Mice for the Study of Vascular Disease

Vascular procedures involving anastomoses in the mouse are generally thought to be difficult and highly dependent on the skill of the individual surgeon. This is largely true, but there are a number of important principles that can reduce the difficulty of these procedures and enhance reproducibility. Orthotopic aortic transplantation is an excellent procedure in which to learn these principles because it involves only two end-to-end anastomoses, but requires good suturing technique and handling of the vessels for consistent success. This procedure begins with the procurement of a length of abdominal aorta from a donor animal, followed by division of the native aorta in the recipient. The procured aorta is then placed between the divided ends of the recipient aorta and sutured into place using end-to-end anastomoses. To accomplish this objective successfully requires a high degree of concentration, good tools, a steady hand, and an appreciation of how easily the vasculature of a mouse can be damaged, resulting in thrombosis. Learning these important principles is what occupies most of the beginner's time when learning microsurgery in small rodents. Throughout this protocol, we refer to these important points. This model can be used to study vascular disease in a variety of different experimental systems1-8. In the context shown here, it is most often used for the study of post-transplant vascular disease, a common long-term complication of solid organ transplantation in which intimal hyperplasia occurs within the allograft. The primary advantage of the model is that it facilitates quantitative morphometric analyses and the transplanted vessel lies contiguous to the endogenous vessel, which can serve as an additional control9. The technique shown here is most often used for mice weighing 18-25 grams. We have accumulated most of our experience using the C57BL/6J, BALB/cJ, and C3H/HeJ strains.

We describe a technique in which a section of the abdominal aorta from a mouse is transplanted orthotopically to just below the renal arteries in an allogeneic or syngeneic recipient. This technique can be useful in studies in which transplantation of large arteries of uniform size is deemed advantageous.

原位主动脉瓣移植在小鼠血管疾病的研究

Vascular procedures involving anastomoses in the mouse are generally thought to be difficult and highly dependent on the skill of the individual surgeon. ...

Bioluminescent Orthotopic Model of Pancreatic Cancer Progression

Pancreatic cancer has an extremely poor five-year survival rate of 4-6%. New therapeutic options are critically needed and depend on improved understanding of pancreatic cancer biology. To better understand the interaction of cancer cells with the pancreatic microenvironment, we demonstrate an orthotopic model of pancreatic cancer that permits non-invasive monitoring of cancer progression. Luciferase-tagged pancreatic cancer cells are resuspended in Matrigel and delivered into the pancreatic tail during laparotomy. Matrigel solidifies at body temperature to prevent leakage of cancer cells during injection. Primary tumor growth and metastasis to distant organs are monitored following injection of the luciferase substrate luciferin, using in vivo imaging of bioluminescence emission from the cancer cells. In vivo imaging also may be used to track primary tumor recurrence after resection. This orthotopic model is suited to both syngeneic and xenograft models and may be used in pre-clinical trials to investigate the impact of novel anti-cancer therapeutics on the growth of the primary pancreatic tumor and metastasis.

Improved understanding of pancreatic cancer biology is critically needed to enable the development of better therapeutic options to treat pancreatic cancer. To address this need, we demonstrate an orthotopic model of pancreatic cancer that permits non-invasive monitoring of cancer progression using in vivo bioluminescence imaging.

生物荧光原位模型胰腺癌进展

Pancreatic cancer has an extremely poor five-year survival rate of 4-6%.  New therapeutic options are critically needed and depend on improved ...

Noninvasive Intratracheal Intubation to Study the Pathology and Physiology of Mouse Lung

The use of a model that mimics the condition of lung diseases in humans is critical for studying the pathophysiology and/or etiology of a particular disease and for developing therapeutic intervention. With the increasing availability of knockout and transgenic derivatives, together with a vast amount of genetic information, mice provide one of the best models to study the molecular mechanisms underlying the pathology and physiology of lung diseases. Inhalation, intranasal instillation, intratracheal instillation, and intratracheal intubation are the most widely used techniques by a number of investigators to administer materials of interest to mouse lungs. There are pros and cons for each technique depending on the goals of a study. Here a noninvasive intratracheal intubation method that can directly deliver exogenous materials to mouse lungs is presented. This technique was applied to administer bleomycin to mouse lungs as a model to study pulmonary fibrosis.

The use of a model that mimics the condition of lung diseases in humans is critical for studying the pathophysiology and/or etiology of a particular disease and for developing therapeutic intervention. Here a noninvasive intratracheal intubation method that can directly deliver exogenous materials to mouse lungs is presented.

无创性气管内插管，以研究小鼠肺的病理生理

The use of a model that mimics the condition of lung diseases in humans is critical for studying the pathophysiology and/or etiology of a particular ...

A Murine Model of Myocardial Ischemia-reperfusion Injury through Ligation of the Left Anterior Descending Artery

Acute or chronic myocardial infarction (MI) are cardiovascular events resulting in high morbidity and mortality. Establishing the pathological mechanisms at work during MI and developing effective therapeutic approaches requires methodology to reproducibly simulate the clinical incidence and reflect the pathophysiological changes associated with MI. Here, we describe a surgical method to induce MI in mouse models that can be used for short-term ischemia-reperfusion (I/R) injury as well as permanent ligation. The major advantage of this method is to facilitate location of the left anterior descending artery (LAD) to allow for accurate ligation of this artery to induce ischemia in the left ventricle of the mouse heart. Accurate positioning of the ligature on the LAD increases reproducibility of infarct size and thus produces more reliable results. Greater precision in placement of the ligature will improve the standard surgical approaches to simulate MI in mice, thus reducing the number of experimental animals necessary for statistically relevant studies and improving our understanding of the mechanisms producing cardiac dysfunction following MI. This mouse model of MI is also useful for the preclinical testing of treatments targeting myocardial damage following MI.

We introduce a surgical method to induce experimental ischemia/reperfusion (I/R) injury to simulate myocardial infarction (MI) in mouse models that allows for more clarity in positioning of the ligation on the left anterior descending artery (LAD) to increase the reproducibility of MI experiments in mice.

心肌缺血再灌注损伤的通过左前降支结扎小鼠模型

Acute or chronic myocardial infarction (MI) are cardiovascular events resulting in high morbidity and mortality. Establishing the pathological mechanisms ...

Orthotopic Injection of Breast Cancer Cells into the Mammary Fat Pad of Mice to Study Tumor Growth.

Breast cancer growth can be studied in mice using a plethora of models. Genetic manipulation of breast cancer cells may provide insights into the functions of proteins involved in oncogenic progression or help to discover new tumor suppressors. In addition, injecting cancer cells into mice with different genotypes might provide a better understanding of the importance of the stromal compartment. Many models may be useful to investigate certain aspects of disease progression but do not recapitulate the entire cancerous process. In contrast, breast cancer cells engraftment to the mammary fat pad of mice better recapitulates the location of the disease and presence of the proper stromal compartment and therefore better mimics human cancerous disease. In this article, we describe how to implant breast cancer cells into mice orthotopically and explain how to collect tissues to analyse the tumor milieu and metastasis to distant organs. Using this model, many aspects (growth, angiogenesis, and metastasis) of cancer can be investigated simply by providing a proper environment for tumor cells to grow.

Cancer is a complex disease that is influenced by the tissue surrounding the tumor as well as local pro- and anti-inflammatory mediators. Therefore, orthotropic injection models, rather than subcutaneous models may be useful to study cancer progression in a manner that better mimics human pathology.

注射原位乳腺癌细胞到小鼠乳腺脂肪垫来研究肿瘤的生长。

Breast cancer growth can be studied in mice using a plethora of models. Genetic manipulation of breast cancer cells may provide insights into the ...

Non-invasive Assessment of the Efficacy of New Therapeutics for Intestinal Pathologies Using Serial Endoscopic Imaging of Live Mice

Animal models of inflammatory bowel disease (IBD) and colorectal cancer (CRC) have provided significant insight into the cell intrinsic and extrinsic mechanisms that contribute to the onset and progression of intestinal diseases. The identification of new molecules that promote these pathologies has led to a flurry of activity focused on the development of potential new therapies to inhibit their function. As a result, various pre-clinical mouse models with an intact immune system and stromal microenvironment are now heavily used. Here we describe three experimental protocols to test the efficacy of new therapeutics in pre-clinical models of (1) acute mucosal damage, (2) chronic colitis and/or colitis-associated colon cancer, and (3) sporadic colorectal cancer. We also outline procedures for serial endoscopic examination that can be used to document the therapeutic response of an individual tumor and to monitor the health of individual mice. These protocols provide complementary experimental platforms to test the effectiveness of therapeutic compounds shown to be well tolerated by mice.

We describe methods for longitudinal monitoring of the efficacy of therapeutics for the treatment of colonic pathologies in mice using a rigid endoscope. This protocol can be readily used for the characterization of the therapeutic response of an individual tumor in live mice and also for monitoring potential disease relapse.

新的治疗肠道病变的疗效，使用活体小鼠的串行内镜成像的非侵入性评估

Animal models of inflammatory bowel disease (IBD) and colorectal cancer (CRC) have provided significant insight into the cell intrinsic and extrinsic ...