下面的协议说明，在20-35克鼠效果很好的系统。该方法可以很容易地适应于更大或更小的小鼠通过简单地改变导管的大小。所有动物方案都经约翰霍普金斯大学动物护理和使用委员会。 1.肺灌注<ol><li>选择市售一英寸长的20个克静脉插管以用于插管。 </li><li>修改导管尖端手动弯曲它产生如图1在前端具有微小曲率。 </li><li>麻醉小鼠用氯胺酮（100毫克/千克）和赛拉嗪（15毫克/千克）的混合物中IP注射，并且由于没有反射运动的确认麻醉。麻醉后立即应用兽医软膏的眼睛。麻醉适用兽医药膏对眼睛紧随，并给予卡洛芬（5-10毫克/公斤SQ），用于手术后和滴注镇痛。 </li><li>将谅解备忘录SE仰卧在倾斜的平台。 如图2，一间大办公室粘合剂缝合环上的作品贴得很清楚。 </li><li>剃颈部和清洁的腹侧部分和消毒用70％酒精的颈部区域。用新的latex-和无粉手套，使用消毒用70％酒精手术器械。 </li><li>使用锋利的剪刀做一个小的手术切口在约12毫米以下的下颌切牙颈部。 </li><li>用镊子轻轻拉扯皮肤的脖子直到尾端气管的腹壁可见。 </li><li>轻轻拉回舌和与弯曲尖端的倾斜朝向鼠标的腹面插入插管。正如在1.4，拉轻轻在脖子上的皮肤，然后将导管插入气管。 注：稍加练习，导管将可见向下移动气管。如果它在食道，就会有导管的运动没有视觉瞄准。无切口在气管制成。 </li><li>一旦导管被认为是在颈部气管，其推进约5mm至可靠地传递声带但仍明显高于隆突。 </li><li>准备灌输高达50微升的液体通过与凝胶装载枪头导管注入。放置在鲁尔毂的前端，但注入外表仔细观察在前端同步用鼠标的呼吸的流体的移动之前。然后注入滴注。 </li><li>用1ml注射器，立即做为0.6ml空气进入肺部相对快速充气穿过导管，以帮助分配液体深在肺部。拆下套管。 </li><li>拆下套管。 </li><li>用少量氰基丙烯酸酯类粘合剂的以关闭小外科伤口中为每个VetBond包装插页指示。将小鼠单独的笼子，并直观地监视他们，直到他们醒来，通常表现得没有任何迹象显示discomf的ORT。 </li></ol> 2.肺固定注意：一旦所有的实验过程中的小鼠进行，肺可以通过固定用甲醛（或任何其它期望的固定剂）被准备好进行组织学处理。 <ol><li>牺牲鼠标的IACUC接受的过程。用于在视频显示代表鼠标，麻醉小鼠的颈脱位被使用。 </li><li>通过手术暴露气管腹侧，做一个小切口，并插入18G的存根针头插入气管，用线扎它进行气管造口术（如果尚未完成）。 </li><li>小心打开了中线切开胸腔，切开膈肌，并卸下侧面胸壁揭露肺部。 </li><li>针的接头末端连接到一个环形支架含甲醛水库。 参见图3。 </li><li>将甲醛的顶面25厘米以上的谅解备忘录的水平东南。 参见图3。下一步要确保没有在固定管没有空气通过运行流体出活塞的结束。连接气管套管到储存管的路厄氏端。打开活塞膨胀肺与甲醛。离开肺部压力下至少20分钟。 </li><li>打开活塞膨胀肺与甲醛。离开肺部压力下至少20分钟。 </li><li>接着，打结气管超出存根针的末端。它可以帮助拉回来慢慢针揭露更多uncannulated气管。当捆绑牢固，取出活塞。 </li><li>小心解剖出肺。 </li><li>将甲醛肺部过夜。较长的时间都很好，有的污渍或程序可以指定特定的时间。也任何其它液体固定剂，例如z修复可用于滴注和浸泡。 </li><li>在进一步的组织学处理，测量如下所述固定肺容积。 </li></ol> 3.测量固定肺容积<ol><li>测量使用如图4阿基米德原理的肺容积。从甲醛卸下固定肺和解剖心脏和任何其他非肺组织。 </li><li>使用用于保持肺部完全在水下以前构造简单的自制线支撑装置。 注：此装置需要进行与取平衡正在使用兼容。 图 5中所示的典型装置从塑料移液器和薄（20 G）的金属丝制成。这个系统可以很好地在视频中使用的平衡，但可以很容易地适应大多数实验室天平。 </li><li>放置一个烧杯≈200毫升水对水的平衡和皮与支撑笼到位。 参见图6中取出的金属笼子。将肺在水面上和下水按同笼。 </li><li>记录天平上的权重。这个数字反映了水置换的体积，因此是一个直接测量肺体积。要小心，以确保肺或缝合或部分铁丝笼不接触的侧面或烧杯底部。 </li><li>对于精度，重复这种测量。除去从水肺，并在组织干。再次重重的抽带保持架到位的烧杯中，重复肺容积测量。两个体积测量然后应平均。 注：如果肺部留在福尔马林超过约一周，空气在肺部将溶解在液体中。当这种情况发生时，肺会下沉，所以不再需要使用任何设备作为图 5中，以保持浸没肺。在这种情况下，体积可以通过简单地进行测量通过保持肺由缝线串之一，直到它被完全淹没，如图4。 </li></ol>

<ol>
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此处描述的方法有若干优点。第一所需设备是简单且廉价的。第二，该插管可迅速用很少的错误进行。第三，要修复肺以恒定压力，然后测量所述固定肺容积的能力允许结构或细胞在肺10正确定量。 一个可能的缺点插管是小手术。这可能会限制重复该过程如果需要2次滴注的能力。然而，随着粘接剂的精心手术和申请中，我们已经能够每周定期做第二次滴注或?...

由于一些原因，肺癌还没有被广泛研究，在小鼠。其中一个原因是，获得的肺是非常困难的体内 ，并固定肺部定量分析操作并不常见。本文介绍的方法旨在改变这种状况。这里的目标是描述简单的方法，将在设置和小鼠肺分析肺癌或其它病理极大地帮助。而所有这些方法是全新的，它们没有被一起呈现如在这里所描述的简化方式的独立的方法。 已经有许多已描述的方法进行小鼠肺插管主要做重复肺功能或支气管肺泡灌洗在纵向研究单个老鼠的目的手稿。由于该原始文件，但已在该所描述的不同的方法来谅解备忘录几个其他文件Ë插管1-9。虽然所有这些方法都可以成功地被使用，它们通常需要相当大的训练，并且通常不是没有一个非平凡的故障率。另外，在为了进行肺功能测量中，套管需要适应气管得足够紧以便没有空气泄漏。然而，另一个实际使用气管插管是直接提供特定的药物（如癌细胞或其他侮辱），或治疗药物到肺。这样的程序并不需要紧密配合插管，也没有任何复杂的肺功能设备。此处示出了该方法的新颖特征包括一个小的外科手术，其允许插管而不进入食道插管的任何可能性。这个简单的方法使插管成功相对较少的培训或经验。多达30只小鼠/小时可以使用此方法具有故障率接近零对待。 一旦吨他老鼠是准备来牺牲，受伤或癌肺然后可以进行组织学和病理分析中删除。然而，为了正确地量化任何组织学变量与其它的肺相比，有必要标准化的固定程序和适当地量化固定肺容积10。本文详细介绍了简单的程序，使标准化的固定程序，以及一个方法来衡量固定的肺容积。该卷在组织学的量化的重要指标，因为如果没有这样的容积测定，只有相对密度可以测量10。一旦肺容积是已知的，然而，细胞和肺中的其他结构的测量绝对测量然后可以定量。

<table border="0" cellpadding="0" cellspacing="0">
	<tbody>
		<tr>
			<td>Laboratory Balance</td>
			<td>Ohaus</td>
			<td>Adventurer Pro Model AV 313&#160;</td>
			<td>Other balances can be used if they have a range of 1 - 300g</td>
		</tr>
		<tr>
			<td>20 g Luer intravenous catheter</td>
			<td>Insylte</td>
			<td></td>
			<td>Several other possible vendors, e.g., Jelco Optiva</td>
		</tr>
		<tr>
			<td>500 mL laboratory bottle</td>
			<td>Various</td>
			<td></td>
			<td>Several other possible vendors</td>
		</tr>
	</tbody>
</table>

instillation and fixation methods useful in mouse lung cancer research

灌输活剂，细胞或化学物质直接进入肺而不伤害或杀死小鼠的能力是在肺癌的研究的一个重要工具。虽然有许多方法，这些方法已经发表，显示如何插管老鼠肺功能的测量，没有一个是不为小鼠大同伙快速气管滴注的潜在问题。在本文中，一个简单快捷的方法进行说明，使调查员进行这种滴注以有效的方式。该方法不需要任何特殊的工具或照明，并且可以用很少的实施而得知。它涉及麻醉小鼠，使一个小切口，在颈部以可视化的气管，然后直接插入静脉导管。小切口被快速关闭用组织粘合剂，并且将小鼠允许恢复。一个熟练的学生或技术人员可以在2分钟/小鼠的平均速率做滴注。一旦吨他癌症成立时，经常有必要在肺部的定量组织学分析。传统病理学家平时也懒得固定在规范肺胀，并分析往往是基于一个计分制度，可以说是相当主观的。虽然这有时可能是足够足够的肺肿瘤的大小，总的估计，肺结构或细胞的任何适当的体视学定量分析需要一个可重复的固定程序和后续的肺容积测量。在这里，我们描述了两个固定在压力下的肺部，然后精确测量固定肺容积简单可靠的方法。唯一的要求是一个实验室的平衡，是准确的范围内为1mg-300克。这里提出这样的程序可以大大提高创建，处理和分析肺癌小鼠的能力。

该过程描述在第一协议本身不导致任何广义的结果。这里只描述一个非常可靠的手段来灌输物质直接进入气管。 图7示出了其中锥虫蓝滴注用这里描述的方法的肺的一个例子。有染料的广泛分布，类似于已经看到的直接给出进入气管或小鼠11,12其他染料或示踪剂。我们也用这种方法来提供或者博莱霉素或弹性蛋白酶对肺，这导致广泛纤维化或肺气肿，分别。 <p class="jove_conten...

Watch this Scientific Journal Video about Instillation and Fixation Methods Useful in Mouse Lung Cancer Research at JoVE.com

Instillation and Fixation Methods Useful in Mouse Lung Cancer Research

本文的目的是描述简单的方法，将在设置和小鼠肺分析肺癌或其它病理极大地帮助。我们提出3个协议能够简单，可靠地进行肺滴注，固定，和肺容积的测量。

灌输和固定方法有用的小鼠肺癌研究

The ability to instill live agents, cells, or chemicals directly into the lung without injuring or killing the mice is an important tool in lung cancer ...

instillation-and-fixation-methods-useful-in-mouse-lung-cancer-research

Research

JoVE Journal

Medicine

28.1K 次观看.  Johns Hopkins University. 本文的目的是描述简单的方法，将在设置和小鼠肺分析肺癌或其它病理极大地帮助。我们提出3个协议能够简单，可靠地进行肺滴注，固定，和肺容积的测量。 

视频: Instillation and Fixation Methods Useful in Mouse Lung Cancer Research

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 ...

科研

医学

A Simple Method of Mouse Lung Intubation

A simple procedure to intubate mice for pulmonary function measurements would have several advantages in longitudinal studies with limited numbers or expensive animal. One of the reasons that this is not done more routinely is that it is relatively difficult, despite there being several published studies that describe ways to achieve it. In this paper we demonstrate a procedure that eliminates one of the major hurdles associated with this intubation, that of visualizing the trachea during the entire time of intubation. The approach uses a 0.5 mm fiberoptic light source that serves as an introducer to direct the intubation cannula into the mouse trachea. We show that it is possible to use this procedure to measure lung mechanics in individual mice over a time course of at least several weeks. The technique can be set up with relatively little expense and expertise, and it can be routinely accomplished with relatively little training. This should make it possible for any laboratory to routinely carry out this intubation, thereby allowing longitudinal studies in individual mice, thereby minimizing the number of mice needed and increasing the statistical power by using each mouse as its own control.

This paper describes a striaghforward and efficient method of intubating mice for pulmonary function measurements or pulmonary instillation, that allows the mice to recover and be studied at later times. The procedure involves an inexpensive fiberoptic light source that directly illuminates the trachea.

一个简单的方法小鼠肺插管

A  simple procedure to intubate mice for pulmonary function measurements would  have several advantages in longitudinal studies with limited numbers or  ...

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 ...

Diagnostic Ultrasound Imaging of Mouse Diaphragm Function

Function analysis of rodent respiratory skeletal muscles, particularly the diaphragm, is commonly performed by isolating muscle strips using invasive surgical procedures. Although this is an effective method of assessing in vitro diaphragm activity, it involves non-survival surgery. The application of non-invasive ultrasound imaging as an in vivo procedure is beneficial since it not only reduces the number of animals sacrificed, but is also suitable for monitoring disease progression in live mice. Thus, our ultrasound imaging method may likely assist in the development of novel therapies that alleviate muscle injury induced by various respiratory diseases. Particularly, in clinical diagnoses of obstructive lung diseases, ultrasound imaging has the potential to be used in conjunction with other standard tests to detect the early onset of diaphragm muscle fatigue. In the current protocol, we describe how to accurately evaluate diaphragm contractility in a mouse model using a diagnostic ultrasound imaging technique.

Diagnostic ultrasound imaging has proven to be effective in diagnosing various respiratory diseases in human and animal subjects. We demonstrate a comprehensive ultrasound protocol utilized by Dr. Zuo's lab to analyze diaphragm kinetics specifically in mouse models. This is also a non-invasive research technique which can provide quantitative information on mouse respiratory muscle function.

小鼠膈肌功能诊断超声成像

Function analysis of rodent respiratory skeletal muscles, particularly the diaphragm, is commonly performed by isolating muscle strips using invasive ...

The Bovine Lung in Biomedical Research: Visually Guided Bronchoscopy, Intrabronchial Inoculation and In Vivo Sampling Techniques

There is an ongoing search for alternative animal models in research of respiratory medicine. Depending on the goal of the research, large animals as models of pulmonary disease often resemble the situation of the human lung much better than mice do. Working with large animals also offers the opportunity to sample the same animal repeatedly over a certain course of time, which allows long-term studies without sacrificing the animals.
The aim was to establish in vivo sampling methods for the use in a bovine model of a respiratory Chlamydia psittaci infection. Sampling should be performed at various time points in each animal during the study, and the samples should be suitable to study the host response, as well as the pathogen under experimental conditions.
Bronchoscopy is a valuable diagnostic tool in human and veterinary medicine. It is a safe and minimally invasive procedure. This article describes the intrabronchial inoculation of calves as well as sampling methods for the lower respiratory tract. Videoendoscopic, intrabronchial inoculation leads to very consistent clinical and pathological findings in all inoculated animals and is, therefore, well-suited for use in models of infectious lung disease. The sampling methods described are bronchoalveolar lavage, bronchial brushing and transbronchial lung biopsy. All of these are valuable diagnostic tools in human medicine and could be adapted for experimental purposes to calves aged 6-8 weeks. The samples obtained were suitable for both pathogen detection and characterization of the severity of lung inflammation in the host.

The Bovine Lung in Biomedical Research: Visually Guided Bronchoscopy, Intrabronchial Inoculation and In Vivo Sampling Techniques

This article describes bronchoscopic techniques in the bovine lung under experimental conditions, i.e. bronchoscopically guided inoculation, bronchoalveolar lavage, bronchial brushing, and transbronchial lung biopsy.

牛龙在生物医学研究：视觉引导支气管镜，支气管内接种和<em&gt;在体内</em&gt;抽样技术

There is an ongoing search for alternative animal models in research of respiratory medicine. Depending on the goal of the research, large animals as ...

Intubation-mediated Intratracheal (IMIT) Instillation: A Noninvasive, Lung-specific Delivery System

Respiratory disease studies typically involve the use of murine models as surrogate systems. However, there are significant physiologic differences between the murine and human respiratory systems, especially in their upper respiratory tracts (URT). In some models, these differences in the murine nasal cavity can have a significant impact on disease progression and presentation in the lower respiratory tract (LRT) when using intranasal instillation techniques, potentially limiting the usefulness of the mouse model to study these diseases. For these reasons, it would be advantageous to develop a technique to instill bacteria directly into the mouse lungs in order to study LRT disease in the absence of involvement of the URT. We have termed this lung specific delivery technique intubation-mediated intratracheal (IMIT) instillation. This noninvasive technique minimizes the potential for instillation into the bloodstream, which can occur during more invasive traditional surgical intratracheal infection approaches, and limits the possibility of incidental digestive tract delivery. IMIT is a two-step process in which mice are first intubated, with an intermediate step to ensure correct catheter placement into the trachea, followed by insertion of a blunt needle into the catheter to mediate direct delivery of bacteria into the lung. This approach facilitates a &#62;98% efficacy of delivery into the lungs with excellent distribution of reagent throughout the lung. Thus, IMIT represents a novel approach to study LRT disease and therapeutic delivery directly into the lung, improving upon the ability to use mice as surrogates to study human respiratory disease. Furthermore, the accuracy and reproducibility of this delivery system also makes it amenable to Good Laboratory Practice Standards (GLPS), as well as delivery of a wide range of reagents which require high efficiency delivery to the lung.

Intubation-mediated Intratracheal IMIT Instillation: A Noninvasive, Lung-specific Delivery System

Intubation-mediated intratracheal (IMIT) instillation of reagents is an excellent, noninvasive method for studying respiratory disease, as well as a method for instilling therapeutic reagents directly into the lung. It is a rapid and highly reproducible method which is suitable for preclinical testing.

插管介导气管内（IMIT）灌输：一种无创，龙专用输送系统

Respiratory disease studies typically involve the use of murine models as surrogate systems. However, there are significant physiologic differences ...

Studying Pancreatic Cancer Stem Cell Characteristics for Developing New Treatment Strategies

Pancreatic ductal adenocarcinoma (PDAC) contains a subset of exclusively tumorigenic cancer stem cells (CSCs) which have been shown to drive tumor initiation, metastasis and resistance to radio- and chemotherapy. Here we describe a specific methodology for culturing primary human pancreatic CSCs as tumor spheres in anchorage-independent conditions. Cells are grown in serum-free, non-adherent conditions in order to enrich for CSCs while their more differentiated progenies do not survive and proliferate during the initial phase following seeding of single cells. This assay can be used to estimate the percentage of CSCs present in a population of tumor cells. Both size (which can range from 35 to 250 micrometers) and number of tumor spheres formed represents CSC activity harbored in either bulk populations of cultured cancer cells or freshly harvested and digested tumors 1,2. Using this assay, we recently found that metformin selectively ablates pancreatic CSCs; a finding that was subsequently further corroborated by demonstrating diminished expression of pluripotency-associated genes/surface markers and reduced in vivo tumorigenicity of metformin-treated cells. As the final step for preclinical development we treated mice bearing established tumors with metformin and found significantly prolonged survival. Clinical studies testing the use of metformin in patients with PDAC are currently underway (e.g., NCT01210911, NCT01167738, and NCT01488552). Mechanistically, we found that metformin induces a fatal energy crisis in CSCs by enhancing reactive oxygen species (ROS) production and reducing mitochondrial transmembrane potential. In contrast, non-CSCs were not eliminated by metformin treatment, but rather underwent reversible cell cycle arrest. Therefore, our study serves as a successful example for the potential of in vitro sphere formation as a screening tool to identify compounds that potentially target CSCs, but this technique will require further in vitro and in vivo validation to eliminate false discoveries.

Pancreatic cancer stem cells (CSCs) can be expanded in vitro using the anchorage-independent sphere culture technique, which represents a powerful tool to study CSC biology and can serve as the first step to develop novel CSC-targeting therapies. Here the methodology for expanding, analyzing and targeting of pancreatic CSCs is provided.

研究胰腺癌干细胞特性开发新的治疗策略

Pancreatic ductal adenocarcinoma (PDAC) contains a subset of exclusively tumorigenic cancer stem cells (CSCs) which have been shown to drive tumor ...

Echocardiographic Measurement of Right Ventricular Diastolic Parameters in Mouse

Diastolic dysfunction is a prominent feature of right ventricular (RV) remodeling associated with conditions of pressure overload. However, the RV diastolic function is rarely quantified in experimental studies. This might be due to technical difficulties in the visualization of the RV in the apical four-chamber view in rodents. Here we describe two positions facilitating the visualization of the apical four-chamber view in mice to assess the RV diastolic function.
The apical four-chamber view is enabled by tilting the mouse fixation platform to the left and caudally (LeCa) or to the right and cranially (RiCr). Both positions provide images of comparable quality. The results of the RV diastolic function obtained from two positions are not significantly different. Both positions are comparably easy to perform. This protocol can be incorporated into published protocols and enables detailed investigations of the RV function.

Here we describe and compare two positions for obtaining the apical four-chamber view in mice. These positions enable the quantification of the right ventricular function, provide comparable results, and can be used interchangeably.

小鼠右心室舒张参数的超声心电图测量

Diastolic dysfunction is a prominent feature of right ventricular (RV) remodeling associated with conditions of pressure overload. However, the RV ...

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 ...

Inducing Acute Lung Injury in Mice by Direct Intratracheal Lipopolysaccharide Instillation

Airway administration of lipopolysaccharide (LPS) is a common way to study pulmonary inflammation and acute lung injury (ALI) in small animal models. Various approaches have been described, such as the inhalation of aerosolized LPS as well as nasal or intratracheal instillation. The presented protocol describes a detailed step-by-step procedure to induce ALI in mice by direct intratracheal LPS instillation and perform FACS analysis of blood samples, bronchoalveolar lavage (BAL) fluid, and lung tissue. After intraperitoneal sedation, the trachea is exposed and LPS is administered via a 22 G venous catheter. A robust and reproducible inflammatory reaction with leukocyte invasion, upregulation of proinflammatory cytokines, and disruption of the alveolo-capillary barrier is induced within hours to days, depending on the LPS dosage used. Collection of blood samples, BAL fluid, and lung harvesting, as well as the processing for FACS analysis, are described in detail in the protocol. Although the use of the sterile LPS is not suitable to study pharmacologic interventions in infectious diseases, the described approach offers minimal invasiveness, simple handling, and good reproducibility to answer mechanistic immunological questions. Furthermore, dose titration as well as the use of alternative LPS preparations or mouse strains allow modulation of the clinical effects, which can exhibit different degrees of ALI severity or early vs. late onset of disease symptoms.

Presented here is a step-by-step procedure to induce acute lung injury in mice by direct intratracheal lipopolysaccharide instillation and to perform FACS analysis of blood samples, bronchoalveolar lavage fluid, and lung tissue. Minimal invasiveness, simple handling, good reproducibility, and titration of disease severity are advantages of this approach.

通过直接内切内脂多糖灌输诱导小鼠急性肺损伤

Airway administration of lipopolysaccharide (LPS) is a common way to study pulmonary inflammation and acute lung injury (ALI) in small animal models. ...