Name: phenotyping mouse pulmonary function in vivo with the lung diffusing capacity
Uploaded: 2015-01-06T10:00:00.000+00:00
Duration: 7 min 13 s
Description: Watch this Scientific Journal Video about Phenotyping Mouse Pulmonary Function In Vivo with the Lung Diffusing Capacity at JoVE.com

This work was supported by NIH HL-10342

NOT: Tüm hayvan protokolleri Johns Hopkins Üniversitesi Hayvan Bakım ve Kullanım Kurulu tarafından kabul edildi. 1. Hayvan hazırlanması <ol><li> Aşağıdaki adımda 2.3 belirtildiği gibi ketamin ve xylazin ile anestezi yoluyla, DFCO ölçümü için 6 C57BL / 6 fareler kontrol hazırlayın. </li><li> Kontroller için olanla aynı yöntem kullanılarak, Tablo 1 &#39;de gösterilen farklı akciğer patolojisi olan diğer tüm fareler hazırlayın. Bu modellerin her biri kurmak için gerekli belirli detaylar ilgili referanslar bulunmaktadır. Kontrol fareleri ve diğer patolojik kohortlarda bu çağın tüm 6-12 haftadır. </li></ol> Karbon monoksit için Difüzyon Faktörü 2. ölçümü (DFCO) <ol><li> Azot, oksijen, neon, ve karbon monoksit için zirveleri ölçmek için makine ile birlikte verilen gaz kromatografisi modülünü kurmak. Bu uygulama için sadece neon ve CO veri. NOT: Bu cihaz bir moleküler s kullanırBir 12.00 mikron film 320.00 mikron kimliği ve 10 metre uzunluğunda taşıyıcı gaz olarak helyum ile ieve sütunu. kromatografi kolonu 0.8 ml bir hacme sahiptir, fakat örnek ile bağlantı boruları yeterli temizleme sağlamak için 2 ml kullanıldı. </li><li> Her deney gününde başlangıcında, küçük farelerden gelen numuneler ölçümleri yapmadan önce, yaklaşık olarak% 0.5 Ne,% 0.5 CO ihtiva eden bir gaz karışımı, torbadan doğrudan 2 ml lik bir numuneyi almak ve hava denge ve kalibre etmek için bu örneği kullanmak Gaz kromatografisi. </li><li> Ketamin (90 mg / kg) ve ksilazin (15 mg / kg) ile fareler anestezi ve refleks hareketinin olmaması ile anestezi teyit etmektedir. Kuruluğunu önlemek için gözleri veteriner merhem sürün. (Çok genç farelerde erişkinlerde 18 G veya 20 G) bir taslaktır iğne kanül ile fareler Tracheostomize. NOT: DFCO anestezi sonrası en az 10 dakika tamamlandı ve herhangi bir mekanik havalandırma veya diğer prosedürler öncesinde olduğu. </li><li> 6 haftalıkken daha büyük farelerde, bir 3 ml&#39;lik bir enjektör t kullanımıo gaz karışımı çantasından gaz 0,8 ml çekilme. Trakeal kanül için şırınga bağlayın ve hızlı bir şekilde akciğer şişirmek. Bir metronom kullanarak, 9 sn saymak, ve sonra hızlı bir şekilde 0.8 ml (üfledi hava) çekilme. </li><li> Bu geri 0.8 mi oda havası ile 2 ml kadar nefes vermeyle oluşan seyreltin, en az 15 saniye boyunca dinlenmeye bırakın. Sonra analiz için gaz kromatografisinde içine bütün örnek enjekte. </li><li> Bu ilk DFCO örneği analiz birlikte, gaz karışımı, torba, ikinci bir 0.8 ml fare akciğer şişirmek ve ilk örneğe benzer bu örnek işlem. İki DFCO ölçümleri ortalama. Not: 0.8 ml hacmi çok küçük farelerin akciğerlerinde ölçümler yapmak için çok büyük olduğu için 2 haftalık olarak genç farelerde ölçümleri için, 0.4 ml bir hacim kullanılır. Bu 6 hafta daha yaşlı fareler için 0.8 ml hacmi kullanmak daha iyidir, ve 0.4 ml hacim bazı fareler için gerekli ise, bu çalışılan kohort tüm fareler için sürekli kullanılması gerektiğini söyledi. </li><li> DFCO hesaplayın(CO 9 / CO c) / (Ne 9 / Ne c), c ve 9 indisler enjekte kalibrasyon gazları konsantrasyonları sırasıyla, 9 saniye nefes tutma süresinden sonra kaldırıldı gazlara bakın - 1 olarak. </li><li> Analiz ve tek yönlü ANOVA ile farklılıkları karşılaştırmak ve tüm kohort farelerde çoklu karşılaştırmalar için Tukey düzeltme ile anlamlılık düzeyini değerlendirmek. P &lt;0.05 anlamlı olarak değerini düşünün. NOT: Burada kullanılan farelerin tamamı burada rapor değil akciğer ventilasyonu, mekanik, akciğer lavaj veya histoloji birkaç sonraki ölçümleri içeren deneysel çalışmaların bir parçası idi. Yöntem, kontrol fareleri, yukarıda yapıldığı gibi her deneysel modellerde aynı olduğu için buna ek olarak, çeşitli patolojik modellerden sadece sonuçlar sunulmaktadır. Bu modellerde daha fazla bilgi ek tabloda yer almaktadır. </li><li> Derin anesthe tarafından hayvanları Euthanizetic aşırı doz servikal dislokasyon veya başları kesilerek izledi. Gerekli yerlerde, daha fazla biyolojik veya histolojik işleme ve analiz için ölü farelerin akciğer hücreleri ve / veya dokuları kaldırın. </li></ol>

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Ilgi ve hiçbir şey yok çatışmalar ifşa.

Bu çalışmada, biz fare akciğer gaz alışverişi yeteneğini ölçmek için yeni bir metrik tanımladı. Bu metrik difüzyon kapasitesi, yani, akciğer birincil işlevi ölçen bir genel klinik ölçüm, gaz alışverişi yeteneği benzerdir. difüzyon kapasitesi kolayca ve hızlı bir şekilde fareler ve insanlar hem de yapılabilir ancak akciğer fonksiyonel bir ölçüdür. Farelerde akciğer hastalığı tespiti için önemli bir hedef kontrol ve deney kohort arasında akciğer fonksiyon değişiklikleri ölçme...

Fare hemen akciğer hastalıklarının çeşitli modellemek için kullanılan birincil bir hayvandır. Bu tür patolojiler altında yatan mekanizmaları incelemek için, fenotipik yöntemler bunu patolojik değişiklikler ölçmek ki ihtiyaç vardır. Havalandırma mekaniği ölçülür birçok fare çalışma olmasına rağmen, bu ölçümlerin genelde normal insanlarda yapılan akciğer fonksiyonunun standart değerlendirmelere ilgisiz. Bu insan hastalığı için fare modellerinde sonuçların çeviri kolaylaştırabilir fareler ve insan deneklerde eşdeğer ölçümleri gerçekleştirmek için yeteneği beri, talihsiz bir durumdur. Insan deneklerde en yaygın ve en kolay yapılan ölçümlerin bir karbon monoksit (DLCO) 1,2 difüzyon kapasitesi olduğunu, ancak bu ölçüm sadece nadiren fare modellerinde yapılmıştır. Prosedürler genellikle hantal veya req olabilir çünkü 3-7 bildirilmiştir bu çalışmalarda, kısmen, hiçbir izlem çalışmaları olmuşturkarmaşık bir ekipmana uire. Başka bir yaklaşım bilinçli farelerde CO difüzyon ölçümü mümkün olmanın avantajına sahip bir sabit devlet sisteminde, bir CO, yeniden soluk yöntemi kullanmaktır. Ancak bu yöntem çok hantal ve sonuçları farenin havalandırma düzeyi yanı sıra O 2 ve CO 2 konsantrasyonları 8,9 ile değişebilir. Bu zorluklar onun çeşitli avantajlara rağmen, farelerde akciğer patolojileri tespit etmek difüzyon kapasitesi rutin kullanımını engellemiştir gibi görünüyor. Farelerde difüzyon kapasitesi ölçümü ile ilgili sorunları aşmak için, basit araçlarla ayrıntıları farelerde ölçmek için son 10 bildirilmiştir. prosedür hızlı bir şekilde tüm ilham gaza eşit bir hacim örnekleyerek kirlenmemiş alveol gaz örnekleme zor sorunu ortadan kaldırır. Bir çok tekrarlanabilir ölçüm Bu prosedür sonuçları, pa bir dizi duyarlı karbon monoksit (DFCO), difüzyon faktörü olarak adlandırılanAkciğer fenotipinde thologic değişir. (CO 9 / CO c) / (Ne 9 / Ne c), c ve 9 indisler enjekte kalibrasyon gazların yoğunlaşması ve 9 sn nefes tutma süresinden sonra kaldırıldı gazlara bakın - DFCO böylece 1 olarak hesaplanır sırasıyla. DFCO 1 tüm CO tam alımını yansıtan ve 0 CO hiçbir alımını yansıtan, 0 ve 1 arasında değişen bir boyutsuz değişken vardır. Bu sunumda bu difüzyon kapasitesi ölçümü yapmak için nasıl göstermek ve neredeyse tüm amfizem, fibrozis, akut akciğer hasarı, ve viral ve mantar enfeksiyonları dahil olmak üzere mevcut fare akciğer hastalığı modellerinin değişiklikleri belgelemek için nasıl kullanılabilir.

<table><tbody><tr><td>Gas Chromatograph</td><td>Inficon</td><td>Micro GC Model 3000A</td><td>Agilent makes a comparable model</td></tr><tr><td>18 G Luer stub needle</td><td>Becton Dickenson</td><td></td><td>Several other possible vendors</td></tr><tr><td>3 ml plastic syringe</td><td>Becton Dickenson</td><td></td><td>Several other possible vendors</td></tr><tr><td>Polypropylene gas sample bags</td><td>SKC</td><td>1 or 2 L capacity works well</td><td>Other gas tight bags will work well</td></tr><tr><td>Gas tank, 0.3% Ne, 0.3% CO, balance air; (size ME)</td><td>Airgas, Inc</td><td>Z04 NI785ME3012</td><td>This is the standard mixture used for DLCO in humans</td></tr><tr><td>25 TCID50/mouse of influenza virus A/PR8 diluted in phosphate buffered saline.</td><td></td><td></td><td></td></tr><tr><td>Porcine pancreatic elastase</td><td>Elastin Products, Owensville, MO</td><td></td><td>5.4 U</td></tr><tr><td>Bleomycin</td><td>APP Pharmaceuticals, Schaumburg, IL</td><td></td><td>0.25 U</td></tr><tr><td>&lt;em&gt;Escherichia coli&lt;/em&gt; LPS</td><td>Sigma L2880</td><td></td><td>3 &amp;mu;g/g body weight; O55:B5</td></tr><tr><td>&lt;i&gt;Aspergillus fumigatus &lt;/i&gt; (isolate Af293) conidia were collected from mature colonies grown on potato dextrose agar.</td><td></td><td></td><td></td></tr></tbody></table>

phenotyping mouse pulmonary function in vivo with the lung diffusing capacity

Fare hemen akciğer hastalıklarının çeşitli modellemek için kullanılan birincil bir hayvandır. Bu tür patolojiler altında yatan mekanizmaları incelemek için, fenotipik yöntemler patolojik değişiklikler ölçmek ki ihtiyaç vardır. Bundan başka, fare modelleri ile translasyon uygunluğu sağlamak için, bu ölçümlerin kolaylıkla hem insanlarda hem farelerde yapılabilir testler olmalıdır. Ne yazık ki, bu literatürde akciğer fonksiyonunun birkaç fenotipik ölçümleri insanlara doğrudan uygulama var. Tek istisna rutin insanlarda yapılan bir ölçüm olan karbon monoksit için difüzyon kapasitesi. Bu yazıda, hızlı ve basit farelerde bu difüzyon kapasitesini ölçmek için bir araç açıklar. Prosedür 1 dk gazı analizi zaman izlediği bir anestezi fare izleyici gazları ile kısa akciğer enflasyon, içerir. Bu amfizem, fibroz, akut akciğer yaralanması ve influenza da dahil olmak üzere pek çok akciğer hastalıkları, tespit etmek için bu yöntem yeteneğini test ettikmantar enfeksiyonları akciğer, yanı sıra genç yavrular izleme akciğer olgunlaşmasının. Sonuçlar önemli tüm akciğer patolojileri düşüşler yanı sıra, akciğer olgunlaşması ile difüzyon kapasitesinde bir artış göstermektedir. Akciğer difüzyon kapasitesi Bu ölçüm, böylece mevcut patolojik akciğer modellerinin çoğu ile fenotipik yapısal değişiklikleri tespit yeteneği ile geniş bir uygulama olan solunum fonksiyon testi sağlar.

Şekil 1, var Aspergillus ve influenza enfeksiyonu hem önemli düşüşler, hem de fibrotik amfizematöz önemli düşüş vardı ve akut grupları A, B, C, D, E ve F yetişkin farelerden DFCO ölçümlerini göstermektedir Akciğer hasarı modelleri. Şekil 2 2-6 hafta fareler yaş olarak zamanla DFCO&#39;da G Grubu gelişimsel değişiklikleri gösterir. Bu sefer boyunca akciğer gelişimi ile hafif fakat anlamlı bir artış vardı. küçük bir enflasyon hacmi kullanılarak etkisi ...

Watch this Scientific Journal Video about Phenotyping Mouse Pulmonary Function In Vivo with the Lung Diffusing Capacity at JoVE.com

Phenotyping Mouse Pulmonary Function In Vivo with the Lung Diffusing Capacity

We describe a means to quickly and simply measure the lung diffusing capacity in mice and show that it is sufficiently sensitive to phenotype changes in multiple common lung pathologies. This metric thus brings direct translational relevance to the mouse models, since diffusing capacity is also easily measured in humans.

Fenotiplendirme Fare Solunum Fonksiyon<em&gt; İn Vivo</em&gt; Akciğer Yayıcı Kapasite

The mouse is now the primary animal used to model a variety of lung diseases. To study the mechanisms that underlie such pathologies, phenotypic methods ...

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Biology

Phenotyping Mouse Pulmonary Function In Vivo with the Lung Diffusing Capacity

10.5K Görüntüleme.  Johns Hopkins University Bloomberg School of Public Health. We describe a means to quickly and simply measure the lung diffusing capacity in mice and show that it is sufficiently sensitive to phenotype changes in multiple common lung pathologies. This metric thus brings direct translational relevance to the mouse models, since diffusing capacity is also easily measured in humans.

Video: Phenotyping Mouse Pulmonary Function In Vivo with the Lung Diffusing Capacity

A Standardized Method for Measuring Internal Lung Surface Area via Mouse Pneumonectomy and Prosthesis Implantation

Pulmonary morphology, physiology, and respiratory functions change in both physiological and pathological conditions. Internal lung surface area (ISA), representing the gas-exchange capacity of the lung, is a critical criterion to assess respiratory function. However, observer bias can significantly influence measured values for lung morphological parameters. The protocol that we describe here minimizes variations during measurements of two morphological parameters used for ISA calculation: internal lung volume (ILV) and mean linear intercept (MLI). Using ISA as a morphometric and functional parameter to determine the outcome of alveolar regeneration in both pneumonectomy (PNX) and prosthesis implantation mouse models, we found that the increased ISA following PNX treatment was significantly blocked by implantation of a prosthesis into the thoracic cavity1. The ability to accurately quantify ISA is not only expected to improve the reliability and reproducibility of lung function studies in injured-induced alveolar regeneration models, but also to promote mechanistic discoveries of multiple pulmonary diseases.

Internal lung surface area (ISA) is a critical criterion for assessing lung morphology and physiology in lung diseases and injury-induced alveolar regeneration. We describe here a standardized method that can minimize the measurement bias for ISA in both lung pneumonectomy and prosthesis implantation mouse models.

Fare Pnömonektomi ve Protez İmplantasyonu ile İç Akciğer Yüzey Alanının Ölçümü için Standart Bir Yöntem

Pulmonary morphology, physiology, and respiratory functions change in both physiological and pathological conditions. Internal lung surface area (ISA), ...

Gelişim Biyolojisi

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.

Sigara Duman maruz kalan Fare Akciğer Amfizem ve Küçük Havayolu Yaptırma otomatik ölçüm

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 ...

Tıp

Measurement of the Pressure-volume Curve in Mouse Lungs

In recent decades the mouse has become the primary animal model of a variety of lung diseases. In models of emphysema or fibrosis, the essential phenotypic changes are best assessed by measurement of the changes in lung elasticity. To best understand specific mechanisms underlying such pathologies in mice, it is essential to make functional measurements that can reflect the developing pathology. Although there are many ways to measure elasticity, the classical method is that of the total lung pressure-volume (PV) curve done over the whole range of lung volumes. This measurement has been made on adult lungs from nearly all mammalian species dating back almost 100 years, and such PV curves also played a major role in the discovery and understanding of the function of pulmonary surfactant in fetal lung development. Unfortunately, such total PV curves have not been widely reported in the mouse, despite the fact that they can provide useful information on the macroscopic effects of structural changes in the lung. Although partial PV curves measuring just the changes in lung volume are sometimes reported, without a measure of absolute volume, the nonlinear nature of the total PV curve makes these partial ones very difficult to interpret. In the present study, we describe a standardized way to measure the total PV curve. We have then tested the ability of these curves to detect changes in mouse lung structure in two common lung pathologies, emphysema and fibrosis. Results showed significant changes in several variables consistent with expected structural changes with these pathologies. This measurement of the lung PV curve in mice thus provides a straightforward means to monitor the progression of the pathophysiologic changes over time and the potential effect of therapeutic procedures.

Here we present a protocol to simply and reliably measure the lung pressure-volume curve in mice, showing that it is sufficiently sensitive to detect phenotypic parenchymal changes in two common lung pathologies, pulmonary fibrosis and emphysema. This metric provides a means to quantify the lung&#8217;s structural changes with developing pathology.

Fare Akciğerler Basınç-hacim Eğrisi Ölçümü

In recent decades the mouse has become the primary animal model of a variety of lung diseases. In models of emphysema or fibrosis, the essential ...

Intraperitoneal Glucose Tolerance Test, Measurement of Lung Function, and Fixation of the Lung to Study the Impact of Obesity and Impaired Metabolism on Pulmonary Outcomes

Obesity and respiratory disorders are major health problems. Obesity is becoming an emerging epidemic with an expected number of over 1 billion obese individuals worldwide by 2030, thus representing a growing socioeconomic burden. Simultaneously, obesity-related comorbidities, including diabetes as well as heart and chronic lung diseases, are continuously on the rise. Although obesity has been associated with increased risk for asthma exacerbations, worsening of respiratory symptoms, and poor control, the functional role of obesity and perturbed metabolism in the pathogenesis of chronic lung disease is often underestimated, and underlying molecular mechanisms remain elusive. This article aims to present methods to assess the effect of obesity on metabolism, as well as lung structure and function. Here, we describe three techniques for mice studies: (1) assessment of intraperitoneal glucose tolerance (ipGTT) to analyze the effect of obesity on glucose metabolism; (2) measurement of airway resistance (Res) and respiratory system compliance (Cdyn) to analyze the effect of obesity on lung function; and (3) preparation and fixation of the lung for subsequent quantitative histological assessment. Obesity-related lung diseases are probably multifactorial, stemming from systemic inflammatory and metabolic dysregulation that potentially adversely influence lung function and the response to therapy. Therefore, a standardized methodology to study molecular mechanisms and the effect of novel treatments is essential.

The incidence of obesity is rising and increases the risk of chronic lung diseases. To establish the underlying mechanisms and preventive strategies, well-defined animal models are needed. Here, we provide three methods (glucose-tolerance-test, body plethysmography, and lung fixation) to study the effect of obesity on pulmonary outcomes in mice.

Mayi glikoz tolerans testi, akciğer işlev ölçümü ve akciğer fiksasyonu obezite ve pulmoner sonuçlar üzerine Engelli metabolizma etkisini incelemek için

Obesity and respiratory disorders are major health problems. Obesity is becoming an emerging epidemic with an expected number of over 1 billion obese ...

İmmünoloji ve Enfeksiyon

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.

Kronik obstrüktif akciğer hastalığı manken tekrarlanan ozon maruz farelerde nesil

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

Evaluation of Respiratory System Mechanics in Mice using the Forced Oscillation Technique

The forced oscillation technique (FOT) is a powerful, integrative and translational tool permitting the experimental assessment of lung function in mice in a comprehensive, detailed, precise and reproducible manner. It provides measurements of respiratory system mechanics through the analysis of pressure and volume signals acquired in reaction to predefined, small amplitude, oscillatory airflow waveforms, which are typically applied at the subject's airway opening. The present protocol details the steps required to adequately execute forced oscillation measurements in mice using a computer-controlled piston ventilator (flexiVent; SCIREQ Inc, Montreal, Qc, Canada). The description is divided into four parts: preparatory steps, mechanical ventilation, lung function measurements, and data analysis. It also includes details of how to assess airway responsiveness to inhaled methacholine in anesthetized mice, a common application of this technique which also extends to other outcomes and various lung pathologies. Measurements obtained in na&iuml;ve mice as well as from an oxidative-stress driven model of airway damage are presented to illustrate how this tool can contribute to a better characterization and understanding of studied physiological changes or disease models as well as to applications in new research areas.

The present protocol provides a detailed step-by-step description of the procedures required to execute measurements of respiratory system mechanics as well as the assessment of airway responsiveness to inhaled methacholine in mice using the forced oscillation technique (flexiVent; SCIREQ Inc, Montreal, Qc, Canada).

Zorla Salınım Tekniği kullanarak Fare Solunum Sistemi Mekanik Değerlendirilmesi

The  forced oscillation technique (FOT) is a powerful, integrative and translational  tool permitting the experimental assessment of lung function in mice ...

Biyoloji

Assessment of Respiratory Function in Conscious Mice by Double-chamber Plethysmography

Air volume changes created by a conscious subject breathing spontaneously within a body box are at the basis of plethysmography, a technique used to non-invasively assess some features of the respiratory function in humans as well as in laboratory animals. The present article focuses on the application of the double-chamber plethysmography (DCP) in small animals. It provides background information on the methodology as well as a detailed step-by-step procedure to successfully assess respiratory function in conscious, spontaneously breathing animals in a non-invasive manner. The DCP can be used to monitor the respiratory function of multiple animals in parallel, as well as to identify changes induced by aerosolized substances over a chosen time period and in a repeated manner. Experiments on control and allergic mice are used herein to demonstrate the utility of the technique, explain the associated outcome parameters, as well as to discuss the related advantages and shortcomings. Overall, the DCP provides valid and theoretically sound readouts that can be trusted to evaluate the respiratory function of conscious small animals both at baseline and after challenges with aerosolized substances.

The objective of the present article is to provide a detailed description of the recommended procedures to evaluate respiratory function in conscious mice by double-chamber plethysmography.

Solunum fonksiyon çift-odası pletismografisi tarafından bilinçli farelerde değerlendirilmesi

Air volume changes created by a conscious subject breathing spontaneously within a body box are at the basis of plethysmography, a technique used to ...

Hyperpolarized 129Xe Lung MRI and Spectroscopy in Mechanically Ventilated Mice

Hyperpolarized (HP) xenon-129 (129Xe) is an inhaled magnetic resonance imaging (MRI) contrast agent with unique spectral and physical properties that can be exploited to quantify pulmonary physiology, including ventilation, restricted diffusion (alveolar-airspace size), and gas exchange. In humans, it has been used to evaluate disease severity and progression in a variety of pulmonary disorders and is approved for clinical use in the United States and United Kingdom. Beyond its clinical applications, the ability of 129Xe MRI to noninvasively assess pulmonary pathophysiology and provide spatially resolved information is valuable for preclinical research. Among animal models, mice are the most widely used due to the accessibility of genetically modified disease models. Here, 129Xe MRI is promising as a minimally invasive, radiation-free, and sensitive technique to longitudinally monitor lung disease progression and therapy response (e.g., in drug discovery). This technique can extend to preclinical applications by incorporating an MRI-triggered, free-breathing apparatus or mechanical ventilator to deliver gas. Here, we describe the steps and provide checklists to ensure robust data collection and analysis, including creating a thermally polarized xenon gas phantom for quality control, optimizing polarization, animal handling (sedation, intubation, ventilation, and care for mice), and protocols for ventilation, restricted diffusion, and gas exchange data. While preclinical 129Xe MRI can be applied in various animal models (e.g., rats, pigs, sheep), this protocol focuses on mice due to the challenges posed by their small anatomy, which are balanced by their affordability and the availability of many disease models.

Hyperpolarized xenon MRI can quantify regional lung microstructure (air-space dimensions) and physiology (ventilation and gas exchange) in translational research and clinical care. Although challenging, it can provide comparable pulmonary insights in preclinical studies. This protocol describes the infrastructure and procedures needed to perform routine xenon lung MRI in mice.

Hyperpolarized (HP) xenon-129 (129Xe) is an inhaled magnetic resonance imaging (MRI) contrast agent with unique spectral and physical properties that can ...

Biyomühendislik

Measuring Respiratory Function in Mice Using Unrestrained Whole-body Plethysmography

Respiratory dysfunction is one of the leading causes of morbidity and mortality in the world and the rates of mortality continue to rise. Quantitative assessment of lung function in rodent models is an important tool in the development of future therapies. Commonly used techniques for assessing respiratory function including invasive plethysmography and forced oscillation. While these techniques provide valuable information, data collection can be fraught with artefacts and experimental variability due to the need for anesthesia and/or invasive instrumentation of the animal. In contrast, unrestrained whole-body plethysmography (UWBP) offers a precise, non-invasive, quantitative way by which to analyze respiratory parameters. This technique avoids the use of anesthesia and restraints, which is common to traditional plethysmography techniques. This video will demonstrate the UWBP procedure including the equipment set up, calibration and lung function recording. It will explain how to analyze the collected data, as well as identify experimental outliers and artefacts that results from animal movement. The respiratory parameters obtained using this technique include tidal volume, minute volume, inspiratory duty cycle, inspiratory flow rate and the ratio of inspiration time to expiration time. UWBP does not rely on specialized skills and is inexpensive to perform. A key feature of UWBP, and most appealing to potential users, is the ability to perform repeated measures of lung function on the same animal.

The assessment of respiratory physiology has traditionally relied upon techniques, which require restraint or sedation of the animal. Unrestrained whole-body plethysmography, however, provides precise, non-invasive, quantitative analysis of respiratory physiology in animal models. In addition, the technique allows repeated respiratory assessment of mice allowing for longitudinal studies.

Yarı Tüm vücut pletismografisi kullanılarak Farelerde Solunum Fonksiyon Ölçme

Respiratory dysfunction is one of the leading causes of morbidity and mortality in the world and the rates of mortality continue to rise. Quantitative ...

In vivo Evaluation of Mucociliary Clearance in Mice

Respiratory motile cilia, specialized organelles of the cell, line the apical surface of epithelial cells lining the respiratory tract. By beating in a metachronal, synchronal fashion, these multiple, motile, actin-based organelles generate a cephalad fluid flow clearing the respiratory tract of inhaled pollutants and pathogens. With increasing environmental pollution, novel viral pathogens and emerging multi-drug resistant bacteria, cilia generated mucociliary clearance (MCC) is essential for maintaining lung health. MCC is also depressed in multiple congenital disorders like primary ciliary dyskinesia, cystic fibrosis as well as acquired disorders like chronic obstructive pulmonary disease. All these disorders have established, in some case multiple, mouse models. In this publication, we detail a method using a small amount of radioactivity and dual-modality SPECT/CT imaging to accurately and reproducibly measure MCC in mice in vivo. The method allows for recovery of mice after imaging, making serial measurements possible, and testing potential therapeutics longitudinally over time. The data in wild-type mice demonstrates the reproducibility of the MCC measurement as long as adequate attention to detail is paid, and the protocol strictly adhered to.

In this publication, we describe protocols for assessing airway mucociliary clearance (MCC) in mice in vivo utilizing dual-modality radionuclide imaging. This protocol is designed for a single photon emission computed tomography (SPECT) and computed tomography (CT) acquisition protocol using mouse whole body (MWB) collimators in a dual SPECT/CT system.

Farelerde Mukozdiliyel Temizliğin In vivo Değerlendirilmesi

Respiratory motile cilia, specialized organelles of the cell, line the apical surface of epithelial cells lining the respiratory tract. By beating in a ...

Fenotiplendirme Fare Solunum Fonksiyon

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