Name: an improved method for rapid intubation of the trachea in mice
Uploaded: 2016-02-22T14:00:00
Description: Watch this Scientific Journal Video about An Improved Method for Rapid Intubation of the Trachea in Mice at JoVE.com

Gli autori ringraziano Brian Johnson del Istologia ed Imaging Nucleo presso l&#39;Università di Washington per aiutare con la colorazione tricromica e l&#39;analisi. Questo lavoro è stato sostenuto da NIH sovvenzioni HL098067 e HL089455.

L&#39;animale cura e l&#39;uso comitati istituzionali (IACUC) presso l&#39;Università di Washington e Cedars-Sinai Medical Center hanno approvato il lavoro degli animali necessari per questi studi. 1. Preparazione <ol><li> Sterilizzare entrambe le pinze punta smussata e il divaricatore tramite autoclave. </li><li> Usando una cappa di sicurezza biologica, preparare uno stock di lavoro di BLM in PBS dalla polvere liofilizzata. Sonicare la soluzione per 10 minuti a 35 Khz per assicurare una miscelazione uniforme...

<ol>
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A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pulmonary+fibrosis:+pathogenesis,+etiology+and+regulation.">Pulmonary fibrosis: pathogenesis, etiology and regulation.</a> Mucosal Immunol. 2, 103-121 (2009).</li><li>Wuyts, W. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+pathogenesis+of+pulmonary+fibrosis:+a+moving+target.">The pathogenesis of pulmonary fibrosis: a moving target.</a> Eur Respir J. 41, 1207-1218 (2013).</li><li>Mouratis, M. A., Aidinis, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Modeling+pulmonary+fibrosis+with+bleomycin.">Modeling pulmonary fibrosis with bleomycin.</a> Curr Opin Pulm Med. 17, 355-361 (2011).</li><li>Moeller, A., Ask, K., Warburton, D., Gauldie, J., Kolb, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+bleomycin+animal+model:+a+useful+tool+to+investigate+treatment+options+for+idiopathic+pulmonary+fibrosis?.">The bleomycin animal model: a useful tool to investigate treatment options for idiopathic pulmonary fibrosis?.</a> Int J Biochem Cell Biol. 40, 362-382 (2008).</li><li>Myll&#228;rniemi, M., Kaarteenaho, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pharmacological+treatment+of+idiopathic+pulmonary+fibrosis+-+preclinical+and+clinical+studies+of+pirfenidone,+nintedanib,+and+N-acetylcysteine.">Pharmacological treatment of idiopathic pulmonary fibrosis - preclinical and clinical studies of pirfenidone, nintedanib, and N-acetylcysteine.</a> European Clinical Respiratory Journal. 2, (2015).</li><li>Reinert, T., Baldotto, C. S. d. R., Nunes, F. A. P., Scheliga, A. A. d. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bleomycin-Induced+Lung+Injury.">Bleomycin-Induced Lung Injury.</a> Journal of Cancer Research. 2013, 1-9 (2013).</li><li>Lakatos, H. F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Oropharyngeal+aspiration+of+a+silica+suspension+produces+a+superior+model+of+silicosis+in+the+mouse+when+compared+to+intratracheal+instillation.">Oropharyngeal aspiration of a silica suspension produces a superior model of silicosis in the mouse when compared to intratracheal instillation.</a> Exp Lung Res. 32, 181-199 (2006).</li><li>Helms, M. N., Torres-Gonzalez, E., Goodson, P., Rojas, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Direct+tracheal+instillation+of+solutes+into+mouse+lung.">Direct tracheal instillation of solutes into mouse lung.</a> J Vis Exp. , (2010).</li><li>Osier, M., Oberdorster, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intratracheal+inhalation+vs+intratracheal+instillation:+differences+in+particle+effects.">Intratracheal inhalation vs intratracheal instillation: differences in particle effects.</a> Fundam Appl Toxicol. 40, 220-227 (1997).</li><li>Driscoll, K. E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intratracheal+instillation+as+an+exposure+technique+for+the+evaluation+of+respiratory+tract+toxicity:+Uses+and+Limitations.">Intratracheal instillation as an exposure technique for the evaluation of respiratory tract toxicity: Uses and Limitations.</a> Toxicol Sci. 55, 24-35 (2000).</li><li>MacDonald, K. D., Chang, H. Y., Mitzner, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+improved+simple+method+of+mouse+lung+intubation.">An improved simple method of mouse lung intubation.</a> J Appl Physiol (1985). 106, 984-987 (1985).</li><li>Spoelstra, E. N., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+novel+and+simple+method+for+endotracheal+intubation+of+mice.">A novel and simple method for endotracheal intubation of mice.</a> Lab Anim. 41, 128-135 (2007).</li><li>Cai, Y., Kimura, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Noninvasive+intratracheal+intubation+to+study+the+pathology+and+physiology+of+mouse+lung.">Noninvasive intratracheal intubation to study the pathology and physiology of mouse lung.</a> J Vis Exp. , e50601 (2013).</li><li>Starcher, B., WIlliams, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+method+for+intratracheal+instillation+of+endotoxin+into+the+lungs+of+mice.">A method for intratracheal instillation of endotoxin into the lungs of mice.</a> Lab Anim. 23, 234-240 (1989).</li><li>Daubeuf, F., Frossard, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Performing+bronchoalveolar+lavage+in+the+mouse.">Performing bronchoalveolar lavage in the mouse.</a> Curr Protoc Mouse Biol. 2, 167-175 (2012).</li><li>Li, Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Severe+lung+fibrosis+requires+an+invasive+fibroblast+phenotype+regulated+by+hyaluronan+and+CD44.">Severe lung fibrosis requires an invasive fibroblast phenotype regulated by hyaluronan and CD44.</a> J Exp Med. 208, 1459-1471 (2011).</li><li>Grazioli, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=CYR61+(CCN1)+overexpression+induces+lung+injury+in+mice.">CYR61 (CCN1) overexpression induces lung injury in mice.</a> Am J Physiol Lung Cell Mol Physiol. 308, L759-L765 (2015).</li><li>Redente, E. 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Nei casi in cui aerosol è impraticabile a causa della limitata disponibilità di reagente, la sicurezza, o il costo, amministrazione diretta tracheale è un metodo superiore per la consegna degli agenti esogeni nei polmoni 16 transtracheale instillazione è stato ampiamente utilizzato per raggiungere questo obiettivo.; Tuttavia, come con tutti gli interventi chirurgici, porta con sé anche il potenziale di complicazioni causate dalla procedura stessa, e non necessariamente l&#39;agente essendo instillato. <s...

Nonostante alcune differenze anatomiche e fisiologiche, 1 modelli murini continuano ad essere prezioso per la modellazione di biologia umana e patogenesi della malattia. 2 Dal punto di vista allevamento, i topi sono facili da gestire, hanno un basso tempo di allevamento, una durata di vita accelerato, e sono relativamente poco costoso a casa. Con lo sviluppo di diversi ceppi genetici e strategie (ad es., Condizionali knock-out, topi reporter, approcci lineage tracing, ecc), così come l&#39;ampia gamma di reagenti disponibili (ad es., Anticorpi, proteine ​​ricombinanti, inibitori, ecc), i topi sono diventati un elemento essenziale organismo modello vertebrato per scoprire i processi di omeostasi e malattie umane. 3 I topi sono stati particolarmente preziosi per lo studio le malattie polmonari, tra cui lesione acuta polmonare (ALI) e fibrosi polmonare. 4 ALI negli esseri umani può essere causata da traumi, lesioni, o sepsi ed è caratterizzata da epiteliale eperdita endoteliale (es., edema), l&#39;infiammazione e la fibrosi nascente. In molti pazienti, ALI progredisce nella sua forma grave, sindrome da distress respiratorio acuto (ARDS), che si traduce spesso in fibrosi e la morte a causa di insufficienza respiratoria. 5,6 fibrosi polmonare è una progressiva, una patologia fatale caratterizzata dall&#39;eccesso deposizione di matrice extracellulare , in particolare collagene di tipo I, portando a deficit della funzionalità respiratoria. 7,8 somministrazione di bleomicina (BLM) è il modello più diffuso e meglio caratterizzato per indurre ALI e fibrosi negli animali da esperimento. 9 Sebbene BLM-indotta fibrosi polmonare nei roditori fa non ricapitolare completamente i fenotipi umani fibrotiche, 10 studi di topo con questo modello hanno portato alla scoperta di importanti fattori che influenzano l&#39;insorgenza e la progressione della malattia. 11 Mentre l&#39;esatto meccanismo (s) dietro fibrogenesi BLM-indotta sono sconosciuti, il danno avvioè pensato per sorgere dal contatto-dipendente rotture del DNA nelle cellule epiteliali che rivestono le vie aeree di conduzione e alveoli, in particolare, di tipo 1 pneumociti. 12 Il bisogno di contatto diretto tra BLM e l&#39;epitelio polmonare mette in evidenza l&#39;importanza di un robusto percorso di consegna , e queste preoccupazioni sono anche germano a una vasta gamma di trattamenti mirati per le vie aeree distali, tra cui proteine ​​ricombinanti, anticorpi, siRNA, virus, batteri, particelle, e altro ancora. Aspirazione orofaringeo (OPA) è stato ampiamente utilizzato per questo scopo 13, ma un importante una lacuna di OPA è che una parte dell&#39;agente erogata sia assorbito nel tratto gastrointestinale, determinando così imprecisione della dose somministrata. Un altro approccio ampiamente utilizzato è instillazione transtracheale, che coinvolge tracheostomia sotto forte anestesia per esporre la trachea e instillazione di un agente direttamente nelle vie respiratorie. 14 Tuttavia, non solo può taleuna procedura sia indesiderabile a causa della sua invasività, ma è anche in termini di tempo, richiede un bel po &#39;di allenamento, e provoca una lesione potente per le vie respiratorie sono stati sviluppati. 15,16 Diversi protocolli che coinvolgono l&#39;amministrazione diretta di agenti in trachea senza la necessità di un intervento chirurgico, 16,17,18,19,20 tempi di recupero ma questi metodi comportano estesi causato da anestetici potenti, l&#39;uso di attrezzature costose (es., otoscope / laringoscopio, disponibili in commercio tavole procedura, fibra ottica fili, ecc), un eccesso di manipolazione nella cavità orale, l&#39;incertezza del dosaggio. Questo documento descrive un metodo relativamente semplice di somministrazione tramite intubazione che permette un ricercatore modo rapido, economico e affidabile infondere un reagente nel polmone murino con limitato rischio di danno residuo ai tessuti circostanti.

<table border="0" cellpadding="0" cellspacing="0" width="1528">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:481px;">Bleomycin For Injection, 30 units/vial</td>
			<td style="width:304px;">APP Pharmaceuticals, LLC</td>
			<td style="width:139px;">103720</td>
			<td style="width:604px;">For best results, BLM should be suspended in PBS, aliquoted, and stored as single use lyophilzed aliquots</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Blunt End Forceps</td>
			<td style="width:304px;">N/A</td>
			<td style="width:139px;">N/A</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Tongue Depressor (i.e. bent Valleylab Blade Electrode, 2.4&#34;)&#160;</td>
			<td style="width:304px;">Covidien</td>
			<td>E1551G</td>
			<td>Before use, create a 45 degree bend 1.5 cm&#160; from the blade tip. A suitable depressor can also be created from any metal implement of similar dimensions.&#160;</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Exel Safelet Catheter 22G X 1&#34;</td>
			<td style="width:304px;">Exel International</td>
			<td>26746</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">1 mL Slip-tip Disposable Tuberculin Syringe (200/sp, 1600/ca)</td>
			<td style="width:304px;">BD</td>
			<td>309659</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">0.2ml Pipettor and Filter Tips</td>
			<td style="width:304px;">N/A</td>
			<td style="width:139px;">N/A</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:481px;">Fiber-Lite Illuminator. Model 181-1: Model 180 mated with Standard Dual Gooseneck illuminator</td>
			<td style="width:304px;">Dolan Jenner Industries, Inc.</td>
			<td style="width:139px;">181-1</td>
			<td>Lower output LED illuminators are not recommended as they fail to suficiently illuminate the trachea.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Intubation Board</td>
			<td style="width:304px;">N/A</td>
			<td style="width:139px;">N/A</td>
			<td>See Diagram 1.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Colored Label Tape: 0.5&#160;in. Wide</td>
			<td style="width:304px;">Fisherbrand</td>
			<td>15-901-15A</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Oxygen&#160;</td>
			<td style="width:304px;">N/A</td>
			<td style="width:139px;">N/A</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Phosphate-Buffered Saline, 1X</td>
			<td style="width:304px;">Corning</td>
			<td>21-040-CV</td>
			<td>Product should be sterile</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Non-Sterile Silk Black Braided Suture Spool, 91.4 m, Size 4-0</td>
			<td style="width:304px;">Harvard Apparatus</td>
			<td>517698</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Table Top Anesthesia Machine Isoflurane</td>
			<td style="width:304px;">Highland Medical Equipment</td>
			<td>N/A</td>
			<td>http://www.highlandmedical.net/</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Slide Top Induction Mouse Isoflurane Chamber</td>
			<td>MIP / Anesthesia Technologies</td>
			<td>AS-01-0530-SM</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">FORANE (isoflurane, USP) Liquid For Inhalation 100 mL&#160;</td>
			<td style="width:304px;">Baxter</td>
			<td>1001936040</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">&#160;Nanozoomer Digital Pathology system</td>
			<td style="width:304px;">Hamamatsu</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">IgM ELISA Quantification Kit&#160;</td>
			<td style="width:304px;">Bethyl Laboratories</td>
			<td style="width:139px;">E90-101</td>
			<td></td>
		</tr>
	</tbody>
</table>

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.

Topi intubati sono stati monitorati giornalmente per la perdita di peso e di angoscia, e si sono sacrificati 4, 10 o 17 giorni dopo tramite iniezione intraperitoneale di 2,5% 2,2,2- tribromoethanol. Il lavaggio broncoalveolare (BAL) è stato raccolto in tre lavaggi di PBS come descritto altrove 21 , e il polmone destro è stato fissato in formalina 10%, inclusi in paraffina, e colorati con tricromica di Masson dalla University of Washington Istologia ed Imagi...

Watch this Scientific Journal Video about An Improved Method for Rapid Intubation of the Trachea in Mice at JoVE.com

An Improved Method for Rapid Intubation of the Trachea in Mice

Un metodo migliore per Rapid intubazione della trachea nei topi

Questo articolo presenta un metodo rapido e semplice per l&#39;amministrazione bleomicina direttamente nella trachea del mouse tramite intubazione. I principali vantaggi di questo metodo sono che è altamente riproducibile, facile da padroneggiare, e non richiede attrezzature specializzate o tempi di recupero lunghi.

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

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Guidelines for Elective Pediatric Fiberoptic Intubation

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

Endotracheal Intubation in Mice via Direct Laryngoscopy Using an Otoscope

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Mice, both wildtype and transgenic, are the principal mammalian model in biomedical research currently. Intubation and mechanical ventilation are ...

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Semi-Minimal Invasive Method to Induce Myocardial Infarction in Rats and the Assessment of Cardiac Function by an Isolated Working Heart System

Metodo semi-minimo invasivo per indurre l'infarto miocardico nei ratti e la valutazione della funzione cardiaca da parte di un sistema cardiaco funzionante isolato

Myocardial infarction (MI) remains the main contributor to morbidity and mortality worldwide. Therefore, research on this topic is mandatory. An easily ...

Blind Endotracheal Intubation in Neonatal Rabbits

Intubazione endotracheale cieca nei conigli neonatale

The newborn rabbit is a useful animal model for various pathologies and procedures. Airway management of the rabbit is complex due to its anatomical ...