Ringraziamo Francesca Polverino MD, Research Fellow presso il Brigham and Women Hospital per il suo contributo a questo articolo, e anche Monica Yao, BS, e Kate Rydell, BS per la loro assistenza con allevamento murina ed esponendo i topi al fumo di sigaretta. Questo lavoro è stato sostenuto dal servizio Public Health, National Heart, Lung, and Blood Institute Grants HL111835, HL105339, HL114501, assistenti di volo Medical Research Institute di Grant # CIA123046, il Brigham and Institute Consorzio Ospedale-Lovelace Respiratory Research delle donne, e la Cambridge NIHR biomedica Centro di Ricerca.

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We have no conflicts of interest to disclose.

Most researcher use mice to model the main chronic lung pathologies and abnormal lung physiology in COPD (airspace enlargement, SAR, and increases in lung compliance) present in the human disease. A comprehensive approach to assess the effect of molecules of interest on both emphysema development and SAR is needed in mice in order to comprehensively assess the activities of molecules of interest in these chronic disease processes. 
There are several critical steps in this protocol. First, dur...

L&#39;uso di modelli animali per lo studio BPCO è impegnativo, perché nessun modello può riprodurre perfettamente tutte le caratteristiche della malattia umana (2). La maggior parte dei ricercatori utilizzano topi per modellare la BPCO a causa delle somiglianze tra i topi e gli esseri umani nella loro polmonari fisiologia, patologia, genetica, e metaboliti. Inoltre, i topi sono relativamente poco costoso per studiare, e sia l&#39;enfisema e la piccola rimodellamento delle vie aeree si sviluppano entro 6 mesi di esposizione CS (5,7-9). Il fumo di sigaretta indotta BPCO: Diversi metodi possono indurre BPCO nei topi. Molti ricercatori espongono topi per CS, che è il fattore eziologico principale per BPCO umana. Esposizione CS per 6 mesi provoca lo sviluppo di enfisema e piccole rimodellamento delle vie aeree (SAR) nei topi, ma la gravità della malattia che viene indotta varia a seconda del ceppo murino studiata. Per esempio, i topi NZWLacZ sono resistenti allo sviluppo di enfisema CS-indotta che AKR / J topi sono extremely sensibili (10). La maggior parte dei ricercatori studiano C57BL / 6 topi deformazione del modello di esposizione CS come tanti topi del gene targeting sono disponibili in questo ceppo. Dopo 6 mesi di esposizione CS, enfisema e fibrosi piccole vie aeree sviluppano in wild type (WT) C57BL / 6 topi, ed entrambe le lesioni sono relativamente mite a gravità (5,10). I ricercatori utilizzano due tipi di CS dell&#39;esposizione: esposizioni naso-solo e tutto il corpo. I principali svantaggi del naso di sola tecnica di esposizione sono che: 1) si tratta di un metodo più alta intensità di manodopera; e 2) topi devono essere trattenuto in piccole sezioni che possono indurre una risposta allo stress e ipertermia negli animali (11). Il principale svantaggio di esposizione al corpo intero (qui descritto) è che gli animali possono ingerire (così come inalare) nicotina e catrame prodotti quando hanno pulito la loro pelliccia. I topi esposti a corpo intero CS hanno livelli di carbossiemoglobina inferiori e ridotto la perdita di peso corporeo rispetto animali esposti a naso sola CS (12). test di funzionalità polmonare (PFT): Misure di compliance polmonare e elastanza solito sono simili in C57BL / 6 wild type (WT) topi esposti all&#39;aria o CS per 6 mesi a causa della enfisema relativamente mite che si sviluppa quando il ceppo è esposto CS (10). Tuttavia, quando la distruzione enfisematosa è più grave, aumenti di compliance polmonare e si sposta a sinistra del volume pressione (PV) flusso loop possono essere rilevati. Quest&#39;ultimo può essere osservato, per esempio, in ceppi murini che sono più sensibili agli effetti di CS (10), in CS-esposti / 6 ceppo topi C57BL gene-targeting che hanno un tipo di enfisema più grave di C57BL / 6 topi WT (13), o in topi CS-esposti soggetti a cambiamenti ambientali che li rendono più sensibili agli effetti di CS (14). Questo protocollo utilizza un piccolo ventilatore animali per misurare la riduzione del ritorno elastico del polmone (aumento della compliance polmonare quasi statica [Cost] e riduzioni nel tessutoelastanza [H]), flussi di loop PV, e cambiamenti nella resistenza delle vie aeree e dei tessuti in topi anestetizzati (15,16). Misure di enfisema polmonare: Analisi dello sviluppo dell&#39;enfisema in CS-esposti C56BL / 6 ceppo topi è impegnativo, perché la sua distribuzione è spazialmente eterogenea. Diversi metodi differenti quantificano l&#39;allargamento dello spazio aereo nei topi. Il primo metodo è stato utilizzato il mezzo di intercettazione lineare (L m) (17). Tuttavia, il metodo L m è un lento processo manuale che non può catturare l&#39;eterogeneità della malattia (a meno che tutte le sezioni del polmone vengono campionati a caso) e il suo impiego può quindi introdurre bias osservatore nell&#39;analisi. L&#39;indice distruttivo [DI, (18)] quantifica anche l&#39;allargamento dello spazio aereo utilizzando un foglio trasparente con 50 punti equamente distribuiti posizionati sopra un&#39;immagine digitalizzata stampata di un ematossilina e la sezione del polmone eosina macchiato. Il metodo PI punteggi zona circostante ogni punto accondo la misura in cui i condotti alveolari e pareti alveolari all&#39;interno di questa zona sono distrutti. Lo svantaggio principale del metodo DI è che richiede tempo e non più preciso di altri metodi (19,20). Questo misure protocollo significano lunghezza della corda alveolare e zona alveolare su sezioni polmonari in paraffina colorate con macchia di Gill. Morfometria software converte le immagini di sezioni polmonari alle immagini binarie (in cui il tessuto è bianco e spazio aereo è nero), e poi sovrappone una griglia uniforme di linee orizzontali e verticali (corde) e il software poi quantifica la lunghezza di ciascuna corda all&#39;interno di aree individuate dalla software come spazio aereo. Utilizzando questo metodo, è possibile misurare le dimensioni degli alveoli in tutte le parti del polmone in modo standardizzato e relativamente automatizzato (21). Piccolo rimodellamento delle vie aeree (SAR): La maggiore deposizione di proteine ​​ECM (soprattutto interstitial collagene) intorno piccole vie aeree tra gli animali CS-esposti e contribuisce a ostruzione delle vie aeree. I ricercatori non studiano SAR in modelli animali di BPCO frequentemente come sviluppo enfisema (22). Per quantificare SAR in topi CS-esposti, questo protocollo utilizza il software di analisi di immagine per misurare lo spessore dello strato di proteine ​​ECM che si deposita intorno piccole vie aeree (airways avente un diametro medio compreso tra 300 e 899 m) nelle sezioni polmonari paraffina colorati con colorazione tricromica di Masson.

<table border="0" cellpadding="0" cellspacing="0" width="579">
	<tbody>
		<tr height="34">
			<td height="34" style="height:34px;width:223px;">Whole-body smoke exposure device</td>
			<td style="width:103px;">Teague Enterprise</td>
			<td style="width:101px;">TE-10z</td>
			<td rowspan="2" style="width:153px;">Chronic Smoke exposures to induce chronic lung disease in mice</td>
		</tr>
		<tr height="34">
			<td height="34" style="height:34px;width:223px;">Research Cigarette</td>
			<td style="width:103px;">University of Kentucky</td>
			<td style="width:101px;">3R4F reference cigarettes</td>
		</tr>
		<tr height="34">
			<td height="34" style="height:34px;width:223px;">Pallflex&#174; Air Monitoring Filters, Emfab Filters TX40HI20WW, 25 mm</td>
			<td style="width:103px;">Pall Corporation</td>
			<td style="width:101px;">7219</td>
			<td rowspan="4" style="width:153px;">For measurement of TPMs</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;width:223px;">25 mm filter holder</td>
			<td style="width:103px;">Pall Corporation</td>
			<td style="width:101px;"></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;width:223px;">Filter sampler</td>
			<td style="width:103px;">Intermatic</td>
			<td style="width:101px;">Metal T100</td>
		</tr>
		<tr height="34">
			<td height="34" style="height:34px;width:223px;">Gas meter</td>
			<td style="width:103px;">AEM</td>
			<td style="width:101px;">Gas meters G1.6; G2.5; G4</td>
		</tr>
		<tr height="34">
			<td height="34" style="height:34px;width:223px;">Tracheal Cannula for mouse 18 gauge</td>
			<td style="width:103px;">Labinvention</td>
			<td style="width:101px;"></td>
			<td rowspan="2" style="width:153px;">Analysis of pulmonary function</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;width:223px;">Mechanical ventilator</td>
			<td style="width:103px;">Scireq</td>
			<td style="width:101px;">FlexiVent</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;width:223px;">Gill&#39;s hematoxylin solution&#160;</td>
			<td style="width:103px;">Sigma-Aldrich</td>
			<td style="width:101px;">GSH316</td>
			<td rowspan="3" style="width:153px;">For Gill staining, work under a fume hood</td>
		</tr>
		<tr height="34">
			<td height="34" style="height:34px;width:223px;">Hematoxylin solution, Harris modified</td>
			<td style="width:103px;">Sigma-Aldrich</td>
			<td style="width:101px;">HHS16</td>
		</tr>
		<tr height="34">
			<td height="34" style="height:34px;width:223px;">Cytoseal-60</td>
			<td style="width:103px;">Thermo Scientific</td>
			<td>8310-16</td>
		</tr>
		<tr height="34">
			<td height="34" style="height:34px;width:223px;">Micro-Slide-Field-Finder</td>
			<td style="width:103px;">Andwin Scientific INC</td>
			<td>50-949-582</td>
			<td rowspan="2" style="width:153px;">For analysis of emphysema</td>
		</tr>
		<tr height="34">
			<td height="34" style="height:34px;width:223px;">Scion Image Program</td>
			<td style="width:103px;">Scion Corporation</td>
			<td style="width:101px;"></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;width:223px;">Mason&#39;s trichrome stain</td>
			<td style="width:103px;">Sigma-Aldrich</td>
			<td style="width:101px;">HT15</td>
			<td rowspan="2" style="width:153px;">For analysis of small airway fibrosis</td>
		</tr>
		<tr height="34">
			<td height="34" style="height:34px;width:223px;">MetaMorp Offline version 7.0</td>
			<td style="width:103px;">Molecullar Devices LLC</td>
			<td style="width:101px;">31032</td>
		</tr>
	</tbody>
</table>

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.

Questo protocollo inizia con l&#39;esposizione del corpo intero di topi per CS. Un&#39;adeguata sorveglianza e la manutenzione del dispositivo e monitoraggio dei TPM conta garantire esposizioni fumo costanti (Figura 1). È importante che il ricercatore pratica la tecnica di gonfiaggio polmonare utilizzando il dispositivo di gonfiaggio Questo protocollo inizia con l&#39;esposizione del corpo intero di topi per CS. Un&#39;adeguata sorveglianza e la manutenzione del dispositivo...

Watch this Scientific Journal Video about Automated Measurement of Pulmonary Emphysema and Small Airway Remodeling in Cigarette Smoke-exposed Mice at JoVE.com

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

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.

Misurazione automatica di enfisema polmonare e piccole vie aeree ritocca in sigaretta Mice-fumo esposti

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 ...

automated-measurement-pulmonary-emphysema-small-airway-remodeling

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JoVE Journal

Medicine

13.2K Views.  Brigham and Women's Hospital - Harvard Medical School. 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. 

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

Protocol for Long Duration Whole Body Hyperthermia in Mice

Hyperthermia is a general term used to define the increase in core body temperature above normal. It is often used to describe the increased core body temperature that is observed during fever. The use of hyperthermia as an adjuvant has emerged as a promising procedure for tumor regression in the field of cancer biology. For this purpose, the most important requirement is to have reliable and uniform heating protocols. We have developed a protocol for hyperthermia (whole body) in mice. In this protocol, animals are exposed to cycles of hyperthermia for 90 min followed by a rest period of 15 min. During this period mice have easy access to food and water. High body temperature spikes in the mice during first few hyperthermia exposure cycles are prevented by immobilizing the animal. Additionally, normal saline is administered in first few cycles to minimize the effects of dehydration. This protocol can simulate fever like conditions in mice up to 12-24 hr. We have used 8-12 weeks old BALB/Cj female mice to demonstrate the protocol.

This paper describes a protocol for whole body hyperthermia in mice that can stimulate fever like conditions up to 12-24 hr.

Protocollo per lungo Ipertermia Corpo intera durata in Mouse

Hyperthermia is a general term used to define the increase in core body temperature above normal. It is often used to describe the increased core body ...

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In vivo Measurement of the Mouse Pulmonary Endothelial Surface Layer

The endothelial glycocalyx is a layer of proteoglycans and associated glycosaminoglycans lining the vascular lumen. In vivo, the glycocalyx is highly hydrated, forming a substantial endothelial surface layer (ESL) that contributes to the maintenance of endothelial function. As the endothelial glycocalyx is often aberrant in vitro and is lost during standard tissue fixation techniques, study of the ESL requires use of intravital microscopy. To best approximate the complex physiology of the alveolar microvasculature, pulmonary intravital imaging is ideally performed on a freely-moving lung. These preparations, however, typically suffer from extensive motion artifact. We demonstrate how closed-chest intravital microscopy of a freely-moving mouse lung can be used to measure glycocalyx integrity via ESL exclusion of fluorescently-labeled high molecular weight dextrans from the endothelial surface. This non-recovery surgical technique, which requires simultaneous brightfield and fluorescent imaging of the mouse lung, allows for longitudinal observation of the subpleural microvasculature without evidence of inducing confounding lung injury.

In vivo Measurement of the Mouse Pulmonary Endothelial Surface Layer

The endothelial glycocalyx/endothelial surface layer is ideally studied using intravital microscopy. Intravital microscopy is technically challenging in a moving organ such as the lung. We demonstrate how simultaneous brightfield and fluorescent microscopy may be used to estimate endothelial surface layer thickness in a freely-moving in vivo mouse lung.

Nella misura in vivo della superficie polmonare strato endoteliale mouse

The endothelial glycocalyx is a layer of proteoglycans and associated glycosaminoglycans lining the vascular lumen. In vivo, the glycocalyx is highly ...

Echocardiographic Assessment of the Right Heart in Mice

Transgenic and toxic models of pulmonary arterial hypertension (PAH) are widely used to study the pathophysiology of PAH and to investigate potential therapies. Given the expense and time involved in creating animal models of disease, it is critical that researchers have tools to accurately assess phenotypic expression of disease. Right ventricular dysfunction is the major manifestation of pulmonary hypertension. Echocardiography is the mainstay of the noninvasive assessment of right ventricular function in rodent models and has the advantage of clear translation to humans in whom the same tool is used. Published echocardiography protocols in murine models of PAH are lacking.
In this article, we describe a protocol for assessing RV and pulmonary vascular function in a mouse model of PAH with a dominant negative BMPRII mutation; however, this protocol is applicable to any diseases affecting the pulmonary vasculature or right heart. We provide a detailed description of animal preparation, image acquisition and hemodynamic calculation of stroke volume, cardiac output and an estimate of pulmonary artery pressure.

This article provides a protocol for the echocardiographic assessment of right ventricular size and pulmonary hypertension in mice. Applications include phenotype determination and serial assessment in transgenic and toxin-induced mouse models of cardiomyopathy and pulmonary vascular disease.

La valutazione ecocardiografica del cuore destro in topi

Transgenic and toxic models of pulmonary arterial hypertension (PAH) are widely used to study the pathophysiology of PAH and to investigate potential ...

Automated Measurement of Microcirculatory Blood Flow Velocity in Pulmonary Metastases of Rats

Because the lung is a major target organ of metastatic disease, animal models to study the physiology of pulmonary metastases are of great importance. However, very few methods exist to date to investigate lung metastases in a dynamic fashion at the microcirculatory level, due to the difficulty to access the lung with a microscope. Here, an intravital microscopy method is presented to functionally image and quantify the microcirculation of superficial pulmonary metastases in rats, using a closed-chest pulmonary window and automated analysis of blood flow velocity and direction. The utility of this method is demonstrated to measure increases in blood flow velocity in response to pharmacological intervention, and to image the well-known tortuous vasculature of solid tumors. This is the first demonstration of intravital microscopy on pulmonary metastases in a closed-chest model. Because of its minimized invasiveness, as well as due to its relative ease and practicality, this technology has the potential to experience widespread use in laboratories that specialize on pulmonary tumor research.

A method is presented to measure microcirculatory blood flow velocity in pulmonary cancer metastases of the pleural surface in rats in an automated fashion, using closed-chest pulmonary intravital microscopy. This model has potential to be used as a widespread tool to perform physiologic research on pulmonary metastases in rodents.

Automated Misura di microcircolatorio Portata in polmonare Metastasi di Ratti

Because the lung is a major target organ of metastatic disease, animal models to study the physiology of pulmonary metastases are of great importance. ...

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.

Un metodo migliore per Rapid intubazione della trachea nei topi

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

A Model of Cardiac Remodeling Through Constriction of the Abdominal Aorta in Rats

Heart failure is one of the leading causes of death worldwide. It is a complex clinical syndromethat includes fatigue, dyspnea, exercise intolerance, and fluid retention. Changes in myocardial structure, electrical conduction, and energy metabolism develop with heart failure, leading to contractile dysfunction, increased risk of arrhythmias, and sudden death. Hypertensive heart disease is one of the key contributing factors of cardiac remodeling associated with heart failure. The most commonly-used animal model mimicking hypertensive heart disease is created via surgical interventions, such as by narrowing the aorta. Abdominal aortic constriction is a useful experimental technique to induce a pressure overload, which leads to heart failure. The surgery can be easily performed, without the need for chest opening or mechanical ventilation. Abdominal aortic constriction-induced cardiac pathology progresses gradually, making this model relevant to clinical hypertensive heart failure. Cardiac injury and remodeling can be observed 10 weeks after the surgery. The method described here provides a simple and effective approach to produce a hypertensive heart disease animal model that is suitable for studying disease mechanisms and for testing novel therapeutics.

A rat model of abdominal aortic constriction that induces cardiac hypertrophy and remodeling is described. An efficient, highly-reproducible, and minimally-invasive method is used to provide a simple yet useful platform for research in myocardial hypertrophy and dysfunction.

Un modello di rimodellamento cardiaco Attraverso Costrizione del Aorta addominale in Rats

Heart failure is one of the leading causes of death worldwide. It is a complex clinical syndromethat includes fatigue, dyspnea, exercise intolerance, and ...

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.

Confronto degli effetti di fumo elettronico di sigarette e sigaretta in un romanzo<em&gt; In Vivo</em&gt; Sistema di esposizione

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

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.

Fissazione polmonare sotto pressione costante per la valutazione dell'enfisema nei topi

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

Transdermal Measurement of Glomerular Filtration Rate in Mice

Transdermal analysis of glomerular filtration rate (GFR) is an established technique that is used to assess renal function in mouse and rat models of acute kidney injury and chronic kidney disease. The measurement system consists of a miniaturized fluorescence detector that is directly attached to the skin on the back of conscious, freely moving animals, and measures the excretion kinetics of the exogenous GFR tracer, fluorescein-isothiocyanate (FITC) conjugated sinistrin (an inulin analog). This system has been described in detail in rats. However, because of their smaller size, measurement of transcutaneous GFR in mice presents additional technical challenges. In this paper we therefore provide the first detailed practical guide to the use of transdermal GFR monitors in mice based on the combined experience of three different investigators who have been performing this assay in mice over a number of years.

Here we describe a protocol to measure glomerular filtration rate (GFR) in conscious, freely moving mice using a transdermal GFR monitor.

Transdermico misurazione del tasso di filtrazione glomerulare in topi

Transdermal analysis of glomerular filtration rate (GFR) is an established technique that is used to assess renal function in mouse and rat models of ...

Design and Development of a Model to Study the Effect of Supplemental Oxygen on the Cystic Fibrosis Airway Microbiome

Airway microbial communities are thought to play an important role in the progression of cystic fibrosis (CF) and other chronic pulmonary diseases. Microbes have traditionally been classified based on their ability to use or tolerate oxygen. Supplemental oxygen is a common medical therapy administered to people with cystic fibrosis (pwCF); however, existing studies on oxygen and the airway microbiome have focused on how hypoxia (low oxygen) rather than hyperoxia (high oxygen) affects the predominantly aerobic and facultative anaerobic lung microbial communities. To address this critical knowledge gap, this protocol was developed using an artificial sputum medium that mimics the composition of sputum from pwCF. The use of filter sterilization, which yields a transparent medium, allows optical methods to follow the growth of single-celled microbes in suspension cultures. To create hyperoxic conditions, this model system takes advantage of established anaerobic culturing techniques to study hyperoxic conditions; instead of removing oxygen, oxygen is added to cultures by daily sparging of serum bottles with a mixture of compressed oxygen and air. Sputum from 50 pwCF underwent daily sparging for a 72-h period to verify the ability of this model to maintain differential oxygen conditions. Shotgun metagenomic sequencing was performed on cultured and uncultured sputum samples from 11 pwCF to verify the ability of this medium to support the growth of commensal and pathogenic microbes commonly found in cystic fibrosis sputum. Growth curves were obtained from 112 isolates obtained from pwCF to verify the ability of this artificial sputum medium to support the growth of common cystic fibrosis pathogens. We find that this model can culture a wide variety of pathogens and commensals in CF sputum, recovers a community highly similar to uncultured sputum under normoxic conditions, and creates different culture phenotypes under varying oxygen conditions. This new approach might lead to a better understanding of unanticipated effects induced by the use of oxygen in pwCF on airway microbial communities and common respiratory pathogens.

The goal of this protocol is to develop a model system for the effect of hyperoxia on cystic fibrosis airway microbial communities. Artificial sputum medium emulates the composition of sputum, and hyperoxic culture conditions model the effects of supplemental oxygen on lung microbial communities.

Progettazione e sviluppo di un modello per studiare l'effetto dell'ossigeno supplementare sul microbioma delle vie aeree della fibrosi cistica

Airway microbial communities are thought to play an important role in the progression of cystic fibrosis (CF) and other chronic pulmonary diseases. ...