AR、XH和PE得到了NIH赠款R01HL140398和吉尔森·朗根博基金会赠款的支持。DEMM 和 KK 得到了 NIH 赠款 R01HL136333 和 R01HL134880 （KYK） 以及赫利斯医学研究基金会的赠款的支持。DEMM还得到霍华德·休斯医学研究所（HHMI）吉利亚姆高级研究奖学金的支持。PE 还得到精密环境卫生科学培训 NIEHS T32 ES027801 研究金计划的支持。JC 和 MF 由表观遗传学和分子致癌学系的烟草研究基金以及 MD Anderson 的表观遗传学中心（学者奖 MF）提供支持。FK 和 YZ 由 NIH 赠款 R01 ES029442-01 和 R01 AI135803-01 以及 VA 功绩赠款 CX000104 提供支持。该项目得到了贝勒医学院细胞测量和细胞分类核心的支持，并得到了CPRIT核心设施支持奖（CPRIT-RP180672）、NIH（CA125123和RR024574）和乔尔·塞德斯特伦的资助。

参与实验和开发这项技术的所有动物都根据我们的动物使用协议，由机构动物护理和使用委员会 （IACUC） 和贝勒医学院和 MD 安德森机构批准，这些机构已获得实验室动物护理评估和认证协会 （AAALAC） 的认证。
1. 建造设备
<ol><li>用阀门系统组装空气压缩机。
	<ol>
<li>使用 1 / 8 英寸螺纹男性适配器配件将流计（两个 15 L / min 带 Y 条和 2 个电源起飞）连接到微型压力调节器。请确保在所有螺纹端使用螺纹密封胶带。</li>
		<li>使用以下方式将组装的压力调节器与流计连接到医用空气压缩机仪器:压缩机气口上的 1 / 8 英寸六角、 1 / 8 英寸螺纹耦合配件以及连接到压力调节器的 1 / 8 英寸螺纹男性适配器配件。</li>
		<li>在每个 （4） 流表上安装氧气旋转带刺接头。</li>
		<li>在空气压缩机的顶部空气出口上安装男性适配器（部分包含在医用空气压缩机仪器中）。</li>
	</ol></li>
	<li>组装曝光室（制作4个单元）
	<ol>
<li>将一根 3/4 英寸氯化聚氯乙烯 （CPVC） 管道切成 8 个 4 英寸段。</li>
		<li>将每个段插入一个 3/4 英寸 90° 肘部 CPVC 配件，并将肘部的配件侧连接到直径为 3/4 英寸的 CPVC 男性适配器上。应有八个 CPVC 段，每个部分连接到一个 CPVC 肘部安装和一个 CPVC 男性适配器。</li>
		<li>在距离对方最远的 8.5 L 密封容器（11.25 x 7.75 x 6 英寸）上钻两个孔（直径 1 1/4 英寸），盖子（见图 1 曝光室）。孔的定位是从上到下和从左到右。</li>
		<li>将之前组装的 CPVC 男性适配器的螺纹侧插入容器中的每个孔中。</li>
		<li>从容器内部，在另一侧（室内烟雾输入）上安装一个 3/4 英寸 CPVC 盖，在一侧附加 3/4 英寸 CPVC 滴灌女性适配器（室烟输出）。</li>
		<li>在室内烟雾输出的 CPVC 盖顶部以昆昆克斯（淋浴头）模式钻出五个 3 毫米孔。这将使香烟烟雾以更高的速度进入房间，并确保它均匀地分布在室内的所有方向。</li>
	</ol></li>
	<li>组装卷烟室（最多4个穆林暴露单位）
	<ol>
<li>采取一个单孔橡胶塞（制造商大小8.5），并在较宽的一侧插入一个1/4英寸的带刺Y连接器和一个直刺配件（8毫米开口）在较窄的一侧。香烟将在吸烟过程中（香烟基座）放置在这里。</li>
		<li>将 12 英寸长医用级乙烯基管的一端连接到连接到橡胶塞的 Y 连接器上的一个带刺连接器，将另一端连接到 1/4 英寸的配件上，并将此配件的对面插入一个单孔橡胶塞子（制造商尺寸 1）。</li>
		<li>在另一个橡胶塞子（制造商尺寸 8.5），插入一个 1/4 英寸直管连接器在塞子的较宽的一侧，并将配件的外端连接到 7 英尺医用级乙烯基管。</li>
		<li>将之前组装的两个橡胶塞件结构（步骤 1.3.1–1.3.3）与实验室玻璃排水管中的 8 英寸 x 1.75 英寸玻璃缸连接起来。</li>
	</ol></li>
	<li>阀门控制系统
	<ol>
<li>该系统由有节奏的电磁阀打开和关闭控制，这些阀门模拟吸入（吹）香烟烟雾和清洁空气。控制电磁阀的系统是商业设计的（见 材料表）。</li>
	</ol></li>
	<li>将所有组件组装在一起（见 图 1）
	<ol>
<li>使用 1 英寸紧固件将四个电磁阀安装到阀门控制系统的两侧。</li>
		<li>按照制造商的指示将电磁阀连接到阀门控制系统。</li>
		<li>将 10+32 （M） 螺纹直接器连接到电磁阀上的排气 （"EXH"） 连接，以及同一电磁阀的"IN"和"OUT"连接上的螺纹端口适配器。</li>
		<li>使用 7 英尺医用级乙烯基管通过"OUT"连接将连接到压缩机的流计连接到电磁阀。</li>
		<li>将 7 英尺乙烯基管与第 1.3.3 步的橡胶塞连接起来。在电磁阀上的"IN"连接器上。</li>
		<li>在烟室烟雾输入上插入卷烟室的小橡胶塞。</li>
		<li>将电磁阀连接到香烟基座上按步骤 1.3.1 组装的带刺 Y 连接器的第二个连接。</li>
	</ol></li>
</ol><img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/61793/61793fig01.jpg"> 图1:我们WBIS与CS接触的连接示意图。此图演示了所有组件的组装方式，以形成工作装置。该图仅显示机器能够操作的四个烟室中的一个组装吸烟室。 <a href="https://jove.unicartagenaproxy.elogim.com/files/ftp_upload/61793/61793fig01large.jpg" target="_blank">请单击此处查看此图的较大版本。</a>
2. 香烟烟雾暴露
注意:避免二手和三手接触香烟烟雾。香烟和暴露室应在 II 类 B2 层流生物安全柜中使用。在进行烟雾暴露实验时应佩戴适当的PPE（即口罩、手套、发网、礼服）。
<ol><li>
设置压力和气流
	<ol>
<li>一旦所有部件都按照 图 1的显示进行组装，请打开空气压缩机，等待安全警报自行关闭。</li>
		<li>通过打开压力调节器的旋钮，将空气压缩机的压力调整到 40–50 psi。</li>
		<li>使用流量计将空气压缩机的气流调整到 5 升。</li>
		<li>打开阀门控制器。</li>
		<li>通过按下 SET/LOCK 键，同时按住计时器第一位数字的 UP 键，将阀门控制器上的数字定时器调整为 PULSE-C（显示屏上显示为"Pu-c"）操作模式。然后，按上键，直到达到 Pu-c 模式。按重置键将显示的操作模式（即 Pu-C）设置为工作模式。</li>
		<li>按下 SET/LOCK 更改定时器 1（显示屏上显示为"T1"）。</li>
		<li>按上或向下键设置 T1 到 20 s。</li>
		<li>按下 SET/LOCK 更改定时器 2（显示屏上显示为"T2"）。</li>
		<li>按上或向下键设置T2到3 s。 注:步骤 2.1.5 到 2.1.9 是定制用于特定定时器（参见 材料表）。有关本产品其他用途的进一步说明，请参阅其相应的用户手册。</li>
	</ol>
</li>
	<li>
香烟烟雾处理 注意:该系统允许同时使用1–4个穆林暴露室。<ol>
<li>打开空气压缩机，等待安全警报自行关闭。</li>
		<li>打开阀门控制器。</li>
		<li>将 5 只小鼠转移到四个暴露室中，每个暴露室配有气密可拆卸盖，体积为 8.5 L。 将四个暴露室与小鼠放在 II 类 B2 层层流生物安全柜内。</li>
		<li>在层流生物安全柜内，点亮香烟，将香烟插入卷烟室内。使用市售香烟，其中含有15毫克/烟焦油和1.1毫克/西格尼古丁8相比，肯塔基州3RF4研究香烟（9.5毫克/西格焦油和0.73毫克/西格尼古丁）9。</li>
		<li>打开与当前使用的机舱对应的阀门控制器上的阀门。暴露分为两个阶段:（T1）清洁空气被泵入暴露室20s和（T2）气流导致香烟燃烧，从烟室抽到暴露室的烟雾3s。让香烟完全烧坏，直到它到达过滤器。<ol>
<li>调整定时器设置，在~4分钟的时间内平均执行约10次泡芙/香烟。请注意，定时器和系统很容易根据调查人员的研究需要定制，用于增强或降低CS给注方案。</li>
		</ol>
</li>
		<li>取下香烟过滤器，将烟头放入玻璃烧杯中，用水灭火并抑制气味。</li>
		<li>确保烟室再次关闭，无需香烟。让机器泵清洁空气10分钟。保持对接触CS的脊椎动物的持续监测至关重要。这种暴露方案针对每个暴露室中9周以上的5只雌性小鼠进行了优化。</li>
		<li>重复步骤 2.3.4 到 2.3.7 三次，每天总共 4 支香烟。只要研究人员需要进行实验，这个程序每周重复5天。</li>
		<li>将小鼠从暴露室中取出，带回相应的笼子里。</li>
		<li>关闭阀门控制器和空气压缩机。</li>
		<li>取出暴露和烟室，用水和肥皂清洗，以去除焦油残留物。</li>
		<li>让房间完全干燥，然后再使用它们。</li>
	</ol>
</li>
</ol>

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作者没有什么可透露的。

在这里，我们提供构建小鼠WBIS到CS的仪器所需的信息。系统安装后，调查人员根据动物中尼古丁或可替宁的交付剂量校准系统至关重要。该仪器包含一个定时器和压力计，可用于调节香烟泡芙量、泡芙频率、合并烟雾暴露期以及动物在每支香烟之间获得的休息间隔。此外，每天服用的香烟的实际数量可能因焦油和尼古丁含量而异。最后，必须定期清洁接触香烟烟雾的任何成分，以确保动物的烟雾?...

吸烟仍然是美国可预防疾病的主要原因之一，尽管在过去50-60年中吸烟的成年人数量持续下降。众所周知，吸烟与肺和血液的多种疾病有关，包括慢性阻塞性肺病（COPD），这是一组疾病，包括肺气肿和慢性支气管炎2、3、4。根据美国疾病控制中心（CDC）的数据，2014年，慢性阻塞性肺病是美国第三大死因，超过1500万美国人患有这种疾病。
CS最近也与患克隆造血症（CH）6、7的较高风险有关，在这种情况下，单个造血干细胞不成比例地产生一个人的外周血液的很大一部分。这一发现表明吸烟和骨髓功能之间有潜在的联系。鉴于CS对健康的广泛和非常重要的影响，鉴于穆林疾病模型是生物医学研究进步的基石，开发高效且经济实惠的系统来模拟小鼠的CS是有益的。
在这里，我们提供一个分步指南，建立一个负担得起的系统，用于治疗和研究CS对肺肺气肿和骨髓平衡的体内影响。此设备的组装不需要用户具有专业知识，因此允许 DIY 组装。

<table><tbody><tr><td>1 in fastener</td><td>Lowes</td><td>756990</td><td></td></tr><tr><td>1/4 in Barbed Y connector</td><td>VWR</td><td>89093-282</td><td></td></tr><tr><td>1/4 in straight tubing connector</td><td>VWR</td><td>62866-378</td><td></td></tr><tr><td>1/8 hex nipple</td><td>Lowes</td><td>877221</td><td></td></tr><tr><td>1/8 in threaded coupling fitting</td><td>Lowes</td><td>877208</td><td></td></tr><tr><td>1/8 in threaded male adapter nipple fitting</td><td>Lowes</td><td>877243</td><td></td></tr><tr><td>10/32 (M) threaded straight connector</td><td>Bimba</td><td>EB60</td><td></td></tr><tr><td>3/4 in 90-degree elbow CPVC fitting</td><td>Lowes</td><td>22643</td><td></td></tr><tr><td>3/4 in chlorinated polyvinyl chloride (CPVC) pipe</td><td>Lowes</td><td>23814</td><td></td></tr><tr><td>3/4 in CPVC cap</td><td>Lowes</td><td>23773</td><td></td></tr><tr><td>3/4 in CPVC Drip irrigation female adapter</td><td>Lowes</td><td>194629</td><td></td></tr><tr><td>3/4 in diameter CPVC male adapter</td><td>Lowes</td><td>23766</td><td></td></tr><tr><td>8.5 L airtight container with lid (11.25in x 7.75in x 6 in)</td><td>Komax</td><td>N/A</td><td>Listed as &quot;Komax Biokips Large Bread Box | (280-oz) Large Storage Container&quot;</td></tr><tr><td>Glass drain tube (1.75 in diameter x 8 in length)</td><td>KIMAX</td><td>6500</td><td></td></tr><tr><td>Isonic Solenoid Valves</td><td>Bimba</td><td>V2A02-AW1</td><td></td></tr><tr><td>Marlboro Red 100&#039;s</td><td>Marlboro</td><td>N/A</td><td></td></tr><tr><td>Oxygen swivel barbed connector</td><td>Global Medical Solutions</td><td>RES002</td><td></td></tr><tr><td>Panasonic Timer LT4H-W</td><td>Panasonic</td><td>LT4HW</td><td>Item was built-in the valve controller by Shepherd Controls &amp;amp; Associates</td></tr><tr><td>Pressure regulator</td><td>Allied Electronics and Automation</td><td>70600552</td><td>Also listed as &quot;Norgren R07-100-RGKA&quot;</td></tr><tr><td>Rubber stopper # 1 (one hole)</td><td>VWR</td><td>59581-163</td><td></td></tr><tr><td>Rubber stopper # 8.5 (one hole)</td><td>VWR</td><td>59581-389</td><td></td></tr><tr><td>Scireq inExpose system</td><td>Scireq and Emka Technologies</td><td>N/A</td><td>Commercial system used for comparison with our DIY WBIS</td></tr><tr><td>Straight barbed fitting (8mm opening)</td><td>VWR</td><td>10028-872</td><td></td></tr><tr><td>Thread Sealant tape</td><td>Lowes</td><td>1184243</td><td></td></tr><tr><td>Threaded port adaptor</td><td>Bimba</td><td>P1SA1</td><td></td></tr><tr><td>Timeter Aridyne 2000 Medical Air Compressor</td><td>MFI Medical</td><td>AHC-TE20</td><td></td></tr><tr><td>Timeter flowmeter</td><td>Allied Healthcare Products</td><td>15006-03YP2</td><td>Also listed as &quot;Puritan Air Meter&quot;</td></tr><tr><td>Valve Control system</td><td>Shepherd Controls and Associates</td><td>N/A</td><td>Company custom designed the valve control system for this model.</td></tr><tr><td>Vinyl pipes</td><td>Vitality Medical</td><td>RES3007</td><td></td></tr></tbody></table>

cigarette smoke exposure in mice using a whole-body inhalation system

据报道，2018年美国有近14%的成年人吸烟。香烟烟雾（CS）对肺部和心血管疾病的影响已得到广泛研究，然而，CS对血液和骨髓等其他组织和器官的影响仍然未完全确定。找到适当的系统来研究CS对啮齿动物的影响可能过于昂贵，需要购买市售系统。因此，我们着手构建一个经济实惠、可靠且多功能的系统，以研究CS在小鼠中的病理影响。这种全身吸入暴露系统 （WBIS） 设置通过交替接触 CS 和清洁空气来模拟香烟的呼吸和吹气。在这里，我们表明，这种自己动手（DIY）系统在接触了4个月的香烟烟雾后，在小鼠身上诱发气道炎症和肺气肿。使用这种仪器，CS的全身吸入（WBI）对造血干细胞和骨髓中的祖细胞（HSPCs）的影响也显示出来。

CS 暴露的主要特征之一是肺气肿，其特征是肺部气囊 （肺囊） 的损伤和破坏。因此，最初的实验侧重于DIY系统在反复接触CS后引起雌性小鼠肺部的肺部变化的能力。CS配药方案是根据我们先前的出版物选择的，我们利用此处描述的DIY系统用CS治疗小鼠，并研究肺气肿10、11、12、13、14、15、16的分子病理生理学。<sup class="...

Watch this Scientific Journal Video about Cigarette Smoke Exposure in Mice using a Whole-Body Inhalation System at JoVE.com

Cigarette Smoke Exposure in Mice using a Whole-Body Inhalation System

该协议展示了对香烟烟雾（CS）的病理生理影响的研究，内部内建立了全身吸入（WBI）暴露系统（WBIS）。该系统可以在受控的可重复条件下使动物接触CS，用于研究CS介于肺气肿和血细胞瘤的影响。

使用全身吸入系统在小鼠中暴露香烟烟雾

Close to 14% of adults in the United States were reported to smoke cigarettes in 2018. The effects of cigarette smoke (CS) on lungs and cardiovascular ...

cigarette-smoke-exposure-in-mice-using-a-whole-body-inhalation-system

Universidad de Cartagena UDEC

Research

JoVE Journal

Immunology and Infection

6.2K 次观看.  Baylor College of Medicine. 该协议展示了对香烟烟雾（CS）的病理生理影响的研究，内部内建立了全身吸入（WBI）暴露系统（WBIS）。该系统可以在受控的可重复条件下使动物接触CS，用于研究CS介于肺气肿和血细胞瘤的影响。

视频: Cigarette Smoke Exposure in Mice using a Whole-Body Inhalation System

Isolation of Mouse Respiratory Epithelial Cells and Exposure to Experimental Cigarette Smoke at Air Liquid Interface

Pulmonary epithelial cells can be isolated from the respiratory tract of mice and cultured at air-liquid interface (ALI) as a model of differentiated respiratory epithelium. A protocol is described for isolating and exposing these cells to mainstream cigarette smoke (CS), in order to study epithelial cell responses to CS exposure. The protocol consists of three parts: the isolation of airway epithelial cells from mouse trachea, the culturing of these cells at air-liquid interface (ALI) as fully differentiated epithelial cells, and the delivery of calibrated mainstream CS to these cells in culture. The ALI culture system allows the culture of respiratory epithelia under conditions that more closely resemble their physiological setting than ordinary liquid culture systems. The study of molecular and lung cellular responses to CS exposure is a critical component of understanding the impact of environmental air pollution on human health. Research findings in this area may ultimately contribute towards understanding the etiology of chronic obstructive pulmonary disease (COPD), and other tobacco-related diseases, which represent major global health problems.

Pulmonary epithelial cells can be isolated from the respiratory tract of mice and cultured at air-liquid interface as a model of differentiated respiratory epithelium. A protocol is described for isolating, culturing and exposing these cells to mainstream cigarette smoke, in order to study molecular responses to this environmental toxin.

小鼠呼吸道上皮细胞和实验香烟烟雾暴露在空气分离液界面

Pulmonary epithelial cells can be isolated from the respiratory tract of mice and cultured at air-liquid interface (ALI) as a model of differentiated ...

科研

医学

Impact Assessment of Repeated Exposure of Organotypic 3D Bronchial and Nasal Tissue Culture Models to Whole Cigarette Smoke

Cigarette smoke (CS) has a major impact on lung biology and may result in the development of lung diseases such as chronic obstructive pulmonary disease or lung cancer. To understand the underlying mechanisms of disease development, it would be important to examine the impact of CS exposure directly on lung tissues. However, this approach is difficult to implement in epidemiological studies because lung tissue sampling is complex and invasive. Alternatively, tissue culture models can facilitate the assessment of exposure impacts on the lung tissue. Submerged 2D cell cultures, such as normal human bronchial epithelial (NHBE) cell cultures, have traditionally been used for this purpose. However, they cannot be exposed directly to smoke in a similar manner to the in vivo exposure situation. Recently developed 3D tissue culture models better reflect the in vivo situation because they can be cultured at the air-liquid interface (ALI). Their basal sides are immersed in the culture medium; whereas, their apical sides are exposed to air. Moreover, organotypic tissue cultures that contain different type of cells, better represent the physiology of the tissue in vivo. In this work, the utilization of an in vitro exposure system to expose human organotypic bronchial and nasal tissue models to mainstream CS is demonstrated. Ciliary beating frequency and the activity of cytochrome P450s (CYP) 1A1/1B1 were measured to assess functional impacts of CS on the tissues. Furthermore, to examine CS-induced alterations at the molecular level, gene expression profiles were generated from the tissues following exposure. A slight increase in CYP1A1/1B1 activity was observed in CS-exposed tissues compared with air-exposed tissues. A network-and transcriptomics-based systems biology approach was sufficiently robust to demonstrate CS-induced alterations of xenobiotic metabolism that were similar to those observed in the bronchial and nasal epithelial cells obtained from smokers.

The aim of this protocol is to expose human organotypic 3D bronchial and nasal tissue models to mainstream cigarette smoke (CS) at the air-liquid interface. The impact of CS on the tissues is then investigated using a cytochrome P450 activity assay, a cilia beating measurement, and a systems biology approach.

反复接触器官的3D支气管及鼻部组织文化模型对整个卷烟烟雾的影响评估

Cigarette smoke (CS) has a major impact on lung biology and may result in the development of lung diseases such as chronic obstructive pulmonary disease ...

生物工程

Generation of Electronic Cigarette Aerosol by a Third-Generation Machine-Vaping Device: Application to Toxicological Studies

Electronic-cigarette (e-cig) devices use heat to produce an inhalable aerosol from a liquid (e-liquid) composed mainly of humectants, nicotine, and flavoring chemicals. The aerosol produced includes fine and ultrafine particles, and potentially nicotine and aldehydes, which can be harmful to human health. E-cig users inhale these aerosols and, with the third-generation of e-cig devices, control design features (resistance and voltage) in addition to the choice of e-liquids, and the puffing profile. These are key factors that can significantly impact the toxicity of the inhaled aerosols. E-cig research, however, is challenging and complex mostly due to the absence of standardized assessments and to the numerous varieties of e-cig models and brands, as well as e-liquid flavors and solvents that are available on the market. These considerations highlight the urgent need to harmonize e-cig research protocols, starting with e-cig aerosol generation and characterization techniques. The current study focuses on this challenge by describing a detailed step-by-step e-cig aerosol generation technique with specific experimental parameters that are thought to be realistic and representative of real-life exposure scenarios. The methodology is divided into four sections: preparation, exposure, post-exposure analysis, plus cleaning and maintenance of the device. Representative results from using two types of e-liquid and various voltages are presented in terms of mass concentration, particle size distribution, chemical composition and cotinine levels in mice. These data demonstrate the versatility of the e-cig exposure system used, aside from its value for toxicological studies, as it allows for a broad range of computer-controlled exposure scenarios, including automated representative vaping topography profiles.

Electronic cigarette (e-cig) users are increasing worldwide. Little, however, is known about the health effects induced by inhaled e-cig aerosols. This article describes an e-cig aerosol generation technique suitable for animal exposures and subsequent toxicological studies. Such protocols are required to establish experimentally reproducible and standardized e-cig exposure systems.

第三代机械 Vaping 装置生成电子烟气溶胶: 毒理学研究的应用

Electronic-cigarette (e-cig) devices use heat to produce an inhalable aerosol from a liquid (e-liquid) composed mainly of humectants, nicotine, and ...

化学

Automated Measurement of Pulmonary Emphysema and Small Airway Remodeling in Cigarette Smoke-exposed Mice

COPD is projected to be the third most common cause of mortality world-wide by 2020(1). Animal models of COPD are used to identify molecules that contribute to the disease process and to test the efficacy of novel therapies for COPD. Researchers use a number of models of COPD employing different species including rodents, guinea-pigs, rabbits, and dogs(2). However, the most widely-used model is that in which mice are exposed to cigarette smoke. Mice are an especially useful species in which to model COPD because their genome can readily be manipulated to generate animals that are either deficient in, or over-express individual proteins. Studies of gene-targeted mice that have been exposed to cigarette smoke have provided valuable information about the contributions of individual molecules to different lung pathologies in COPD(3-5). Most studies have focused on pathways involved in emphysema development which contributes to the airflow obstruction that is characteristic of COPD. However, small airway fibrosis also contributes significantly to airflow obstruction in human COPD patients(6), but much less is known about the pathogenesis of this lesion in smoke-exposed animals. To address this knowledge gap, this protocol quantifies both emphysema development and small airway fibrosis in smoke-exposed mice. This protocol exposes mice to CS using a whole-body exposure technique, then measures respiratory mechanics in the mice, inflates the lungs of mice to a standard pressure, and fixes the lungs in formalin. The researcher then stains the lung sections with either Gill&#8217;s stain to measure the mean alveolar chord length (as a readout of emphysema severity) or Masson&#8217;s trichrome stain to measure deposition of extracellular matrix (ECM) proteins around small airways (as a readout of small airway fibrosis). Studies of the effects of molecular pathways on both of these lung pathologies will lead to a better understanding of the pathogenesis of COPD.

The goal of this protocol is to provide automated methods to quantify chronic lung pathologies in a murine model of COPD. The protocol includes exposing mice to cigarette smoke (CS), measuring pulmonary function, inflating the lungs, and using morphometry methods to measure emphysema and small airway remodeling in mice.

肺气肿，小气道重塑的香烟烟雾暴露小鼠自动测量

COPD is projected to be the third most common cause of mortality world-wide by 2020(1).  Animal models of COPD are used to identify molecules that ...

An Improved Method for Rapid Intubation of the Trachea in Mice

Despite some anatomical and physiological differences, mouse models continue to be an essential tool for studying human lung disease. Bleomycin toxicity is a commonly used model to study both acute lung injury and fibrosis, and multiple methods have been developed for administering bleomycin (and other toxic agents) into the lungs. However, many of these approaches, such as transtracheal instillation, have inherent drawbacks, including the need for strong anesthetics and survival surgery. This paper reports a quick, reproducible method of intratracheal intubation that involves mild inhaled anesthesia, visualization of the trachea, and the use of a surrogate spirometer to confirm exposure. As a proof of concept, 8-12 week old C57BL/6 mice were administered either 2.0 U/kg of bleomycin or an equivalent volume of PBS, and both damage and fibrotic endpoints were measured post-exposure. This procedure allows researchers to treat a large cohort of mice in a relatively short period with little expense and minimal post-procedure care.

This article presents a rapid and simple method for administering bleomycin directly into the mouse trachea via intubation. Key advantages of this method are that it is highly reproducible, easy to master, and does not require specialized equipment or lengthy recovery times.

在小鼠气管插管快速的改进方法

Despite some anatomical and physiological differences, mouse models continue to be an essential tool for studying human lung disease. Bleomycin toxicity ...

Comparing the Effects of Electronic Cigarette Vapor and Cigarette Smoke in a Novel In Vivo Exposure System

Electronic cigarettes (E-cigarettes) are being widely used, and growing in popularity. It is estimated that more than 9 million adults use them regularly. The potential adverse health effects of electronic cigarette vapor (E-vapor) exposure are poorly defined. While several animal models of E-vapor exposure have been developed, few models expose rodents to clinically relevant quantities of nicotine and make direct comparisons to cigarette smoke within the same exposure system. Here, we present a method for constructing and operating an E-vapor chamber and cigarette smoke chamber. The chambers are constructed by outfitting anesthesia chambers with a computer controlled pumping system that delivers consistent amounts of E-vapor or cigarette smoke to rodents. Nicotine exposure is measured indirectly by quantifying pre and post-exposure serum cotinine levels. This exposure system can be modified to accommodate various types of E-cigarettes&#160;and tobacco cigarettes, and can be used to compare the effects of E-vapor&#160;and cigarette smoke in vivo.

Comparing the Effects of Electronic Cigarette Vapor and Cigarette Smoke in a Novel In Vivo Exposure System

This protocol describes a method for exposing rodents to electronic cigarette vapor (E-vapor) and cigarette smoke. Exposure chambers are constructed by modifying anesthesia chambers with an automated pumping system that delivers E-vapor or cigarette smoke to rodents. This system can easily be modified to accommodate many experimental endpoints.

比较电子烟和烟雾在小说中的影响<em&gt;体内</em&gt;曝光系统

Electronic cigarettes (E-cigarettes) are being widely used, and growing in popularity. It is estimated that more than 9 million adults use them regularly. ...

Generation of a Chronic Obstructive Pulmonary Disease Model in Mice by Repeated Ozone Exposure

Chronic obstructive pulmonary disease (COPD) is characterized by persistent airflow limitation and lung parenchymal destruction. It has a very high incidence in aging populations. The current conventional therapies for COPD focus mainly on symptom-modifying drugs; thus, the development of new therapies is urgently needed. Qualified animal models of COPD could help to characterize the underlying mechanisms and can be used for new drug screening. Current COPD models, such as lipopolysaccharide (LPS) or the porcine pancreatic elastase (PPE)-induced emphysema model, generate COPD-like lesions in the lungs and airways but do not otherwise resemble the pathogenesis of human COPD. A cigarette smoke (CS)-induced model remains one of the most popular because it not only simulates COPD-like lesions in the respiratory system, but it is also based on one of the main hazardous materials that causes COPD in humans. However, the time-consuming and labor-intensive aspects of the CS-induced model dramatically limit its application in new drug screening. In this study, we successfully generated a new COPD model by exposing mice to high levels of ozone. This model demonstrated the following: 1) decreased forced expiratory volume 25, 50, and 75/forced vital capacity (FEV25/FVC, FEV50/FVC, and FEV75/FVC), indicating the deterioration of lung function; 2) enlarged lung alveoli, with lung parenchymal destruction; 3) reduced fatigue time and distance; and 4) increased inflammation. Taken together, these data demonstrate that the ozone exposure (OE) model is a reliable animal model that is similar to humans because ozone overexposure is one of the etiological factors of COPD. Additionally, it only took 6 - 8 weeks, based on our previous work, to create an OE model, whereas it requires 3 - 12 months to induce the cigarette smoke model, indicating that the OE model might be a good choice for COPD research.

This study describes the successful generation of a new chronic obstructive pulmonary disease (COPD) animal model by repeatedly exposing mice to high concentrations of ozone.

反复臭氧暴露对小鼠慢性阻塞性肺疾病模型的产生

Chronic obstructive pulmonary disease (COPD) is characterized by persistent airflow limitation and lung parenchymal destruction. It has a very high ...

Whole-Body Nanoparticle Aerosol Inhalation Exposures

Inhalation is the most likely exposure route for individuals working with aerosolizable engineered nano-materials (ENM). To properly perform nanoparticle inhalation toxicology studies, the aerosols in a chamber housing the experimental animals must have: 1) a steady concentration maintained at a desired level for the entire exposure period; 2) a homogenous composition free of contaminants; and 3) a stable size distribution with a geometric mean diameter &lt; 200 nm and a geometric standard deviation &sigma;g &lt; 2.5 5. The generation of aerosols containing nanoparticles is quite challenging because nanoparticles easily agglomerate. This is largely due to very strong inter-particle forces and the formation of large fractal structures in tens or hundreds of microns in size 6, which are difficult to be broken up. Several common aerosol generators, including nebulizers, fluidized beds, Venturi aspirators and the Wright dust feed, were tested; however, none were able to produce nanoparticle aerosols which satisfy all criteria 5.
A whole-body nanoparticle aerosol inhalation exposure system was fabricated, validated and utilized for nano-TiO2 inhalation toxicology studies. Critical components: 1) novel nano-TiO2 aerosol generator; 2) 0.5 m3 whole-body inhalation exposure chamber; and 3) monitor and control system. Nano-TiO2 aerosols generated from bulk dry nano-TiO2 powders (primary diameter of 21 nm, bulk density of 3.8 g/cm3) were delivered into the exposure chamber at a flow rate of 90 LPM (10.8 air changes/hr). Particle size distribution and mass concentration profiles were measured continuously with a scanning mobility particle sizer (SMPS), and an electric low pressure impactor (ELPI). The aerosol mass concentration (C) was verified gravimetrically (mg/m3). The mass (M) of the collected particles was determined as M = (Mpost-Mpre), where Mpre and Mpost are masses of the filter before and after sampling (mg). The mass concentration was calculated as C = M/(Q*t), where Q is sampling flowrate (m3/min), and t is the sampling time (minute). The chamber pressure, temperature, relative humidity (RH), O2 and CO2 concentrations were monitored and controlled continuously. Nano-TiO2 aerosols collected on Nuclepore filters were analyzed with a scanning electron microscope (SEM) and energy dispersive X-ray (EDX) analysis.
In summary, we report that the nano-particle aerosols generated and delivered to our exposure chamber have: 1) steady mass concentration; 2) homogenous composition free of contaminants; 3) stable particle size distributions with a count-median aerodynamic diameter of 157 nm during aerosol generation. This system reliably and repeatedly creates test atmospheres that simulate occupational, environmental or domestic ENM aerosol exposures.

A whole-body nanoparticle aerosol inhalation exposure facility was constructed for nano-sized titanium dioxide (TiO2) inhalation toxicology studies. This system provides nano-TiO2 aerosol test atmospheres that have: 1) a steady mass concentration; 2) a homogenous composition free of contaminants; and 3) a stable particle size distribution during aerosol generation.

全身纳米气溶胶吸入暴露

Inhalation is the most likely exposure route for individuals working with aerosolizable engineered nano-materials (ENM). To properly perform nanoparticle ...

生物学

Lung Fixation under Constant Pressure for Evaluation of Emphysema in Mice

Emphysema is a significant feature of chronic obstructive pulmonary disease (COPD). Studies involving an emphysematous mouse model require optimal lung fixation to produce reliable histological specimens of the lung. Due to the nature of the lung&rsquo;s structural composition, which consists largely of air and tissue, there is a risk that it collapses or deflates during the fixation process. Various lung fixation methods exist, each of which has its own advantages and disadvantages. The lung fixation method presented here utilizes constant pressure to enable optimal tissue evaluation for studies using an emphysematous mouse lung model. The main advantage is that it can fix many lungs with the same condition at one time. Lung specimens are obtained from chronic cigarette smoke-exposed mice. Lung fixation is performed using specialized equipment that enables the production of constant pressure. This constant pressure maintains the lung in a reasonably inflated state. Thus, this method generates a histological specimen of the lung that is suitable to evaluate cigarette smoke-induced mild emphysema.

Presented here is a useful protocol for lung fixation that creates a stable condition for histological evaluate of lung specimens from a mouse model of emphysema. The main advantage of this model is that it can fix many lungs with the same constant pressure without lung collapse or deflation.

小鼠肺气肿评价持续压力下的肺固定

Emphysema is a significant feature of chronic obstructive pulmonary disease (COPD). Studies involving an emphysematous mouse model require optimal lung ...

Using Nicotine in a Silica-Exposed Mouse Model to Promote Lung Epithelial-Mesenchymal Transition

Smoking and exposure to silica are common among occupational workers, and silica is more likely to injure the lungs of smokers than non-smokers. The role of nicotine, the primary addictive ingredient in cigarettes, in silicosis development is unclear. The mouse model employed in this study was simple and easily controlled, and it effectively simulated the effects of chronic nicotine ingestion and repeated exposure to silica on lung fibrosis through epithelial-mesenchymal transition in human beings. In addition, this model can help in the direct study of the effects of nicotine on silicosis while avoiding the effects of other components in cigarette smoke.
After environmental adaptation, mice were injected subcutaneously with 0.25 mg/kg nicotine solution into the loose skin over the neck every morning and evening at 12 h intervals over 40 days. Additionally, crystalline silica powder (1-5 &#181;m) was suspended in normal saline, diluted to a suspension of 20 mg/mL, and dispersed evenly using an ultrasonic water bath. The isoflurane-anesthetized mice inhaled 50 &#181;L of this silica dust suspension through the nose and were awoken via chest massage. Silica exposure was administrated daily on days 5-19.
The double-exposed mouse model was exposed to nicotine and then silica, which matches the exposure history of workers who are exposed to both harmful factors. In addition, nicotine promoted pulmonary fibrosis through epithelial-mesenchymal transformation (EMT) in mice. This animal model can be used to study the effects of multiple factors on the development of silicosis.

This study describes a mouse model to study the synergistic effect of nicotine on the progression of pulmonary fibrosis in experimental silicosis mice. The dual-exposure mouse model simulates the pathological progression in the lung after simultaneous exposure to nicotine and silica. The methods described are simple and highly reproducible.

在二氧化硅暴露小鼠模型中使用尼古丁促进肺上皮-间充质转化

Smoking and exposure to silica are common among occupational workers, and silica is more likely to injure the lungs of smokers than non-smokers. The role ...