Name: Mouse Pneumonectomy Model of Compensatory Lung Growth (Video) | JoVE Journal
Uploaded: 2014-12-17T14:00:00.000+00:00
Duration: 9 min 22 s
Description: Learn the mouse pneumonectomy model to study compensatory lung growth and alveolar regeneration mechanisms. This surgical technique enables researchers to investigate lung development and tissue repair processes in vivo.

ai_agent_IP

ai_agent

maintenance

instantTrialDurationMinutes

isBiopharma

enableNewMyJoveSectionDesign

educationFeatures

seoMetaApi

ai_copilot_institution_blacklist

ai_chat_institution_blacklist

freeAccessWindow

ssoRoleCapture

scheduledMaintenanceBanner

joveLabsFeatures

disable_autoplay_for_bots

extended_trial_duration

5 minutes

The authors would like to acknowledge the Cincinnati Children&#8217;s Hospital Division of Veterinary Services for their assistance. This manuscript was supported by the National Institutes of Health K12 HD028827. Anna Perl PhD taught the authors this surgical procedure.

NOTE: Animal use statement: All procedures in this study were conducted with approval and following the guidelines of the Institutional Animal Use and Care Committee (IACUC) at Cincinnati Children&#8217;s Hospital. Eight week-old C57BL/6J male mice were obtained from Jackson Laboratories (Bar Harbor, ME) and allowed to acclimate for one week prior to use. Up until surgery, animals were housed in a pathogen-free barrier facility and provided autoclaved chow and filtered water ad libdium. Each mouse cage was supplied with a dedicated air and water, and rooms were maintained on a 12 hr day-night cycle. Following recovery from surgery, mice were maintained in cages with filtered tops, provided autoclaved chow ad libidum, and provided filtered water from a water bottle.
1. Preparation of Instruments
<ol>
	<li>Make 6 skin retractors using paper clips and pins. Twist straightened paper clips on the shanks of paper pins, leave a 5 cm straight steel wire on one end and make a single 0.5 cm &#8220;U&#8221; shaped hook at the end of the wire.</li>
	<li>Make some 15 x 15 cm square surgical drapes using plastic wrap. Prepare one dressing per mouse. Put a paper tower in between each wrap.</li>
	<li>Sterilize all surgical tools along with a stack of 12 x 12 inch cork tiles, gauze, and cotton-tipped swabs.</li>
</ol>
2. Mouse Preparation
<ol>
	<li>Induce anesthesia with 2% isoflurane. Weigh animal.</li>
	<li>In a dedicated surgical preparation area shave left thorax and neck area with electrical shaver.</li>
	<li>Apply a drop of the artificial tear ointment to the mouse&#8217;s eyes.</li>
	<li>Decontaminate neck and left thorax with chlorhexidine and isopropyl alcohol. Repeat twice more.</li>
</ol>
3. Mouse Oro-tracheal Intubation and Mechanical Ventilation
<ol>
	<li>Have a non-sterile surgical technician place the mouse supine in the pre-warmed surgical area.</li>
	<li>Confirm depth of anesthesia by documenting lack of a response to paw pinch.</li>
	<li>After washing hands and donning surgical attire, mask, and hat, don sterile surgical gloves.</li>
	<li>After draping and using aseptic technique, make a 1 cm vertical incision over the anterior mid-neck to expose the larynx. Lightly retract the strap muscles with curved, serrated 10 cm forceps and expose the larynx and trachea by spreading the strap muscles with the tip of a straight scissors.</li>
	<li>Orally insert a 22 G blunt-tip angiocatheter into the mid-trachea (Figure 1A) and visually confirm placement (Figure 1B).&#160;Maintain anesthesia and ventilate using 1-3% isoflurane through rodent ventilator (225 &#181;l per stroke; 200 stokes per min). Employ a pressure limit of 15 cm H2O.</li>
</ol>
4. Mouse Pneumonectomy
<ol>
	<li>Lay the mouse in the right lateral decubitus position with the mouse&#8217;s back facing the operator (left side up). Use a self-sealing plastic wrap as a sterile drape. Cutting through the drape, use blunt tipped curved scissors to make a 2 cm long cut parallel to the ribs at the 4th and 5th intercostal space. Insert the blunt tip curved scissors and dissect the skin away from the underlying ribs and intercostal muscles.</li>
	<li>Retract the skin with four retractors to expose a square 1.5 x 1.5 cm surgical window (Figure 2A). Secure the retractors to the cork board.</li>
	<li>Dissect down to ribs using curved forceps, and use one tip of the curved forceps to enter the thoracic cavity.</li>
	<li>Using the blunt tip micro-scissors, use the lower blade to enter the thoracic cavity. Make a 0.5 cm incision between the ribs and repeat in the opposite direction.</li>
	<li>Using the two remaining retractors, open the thorax in the anterior-posterior axis and the secure the retractors to the cork board (Figure 2B).</li>
	<li>Using curved blunt-tipped forceps in the left hand, grasp the left lung and displace the upper portion of the left lung laterally and inferiorly through the thoracotomy until the left pulmonary artery and bronchus are exposed (Figure 3A,B).</li>
	<li>Holding the loaded titanium vascular microclip applicator in the right hand with the body of the applicator in the palm and curved tip pointing away from the palm (Figure 3C), slide the applicator tip into the thorax along the curvature of the posterior aspect of the left lung and clip the left bronchus and pulmonary artery (Figure 3D).</li>
	<li>Remove the applicator but keep the left lung retracted. Grasp the blunt tip micro-scissors with the right hand and cut the bronchus and pulmonary artery distal to the clip and remove left lung (Figure 3E).</li>
	<li>Remove the rib retractors.</li>
	<li>Use the curved blunt forceps to pinch up 1 cm of skin inferior to the incision but above the level of the diaphragm and insert a 24 G angiocatheter through the skin and into the left thoracic cavity (Figure&#160;4A,B).</li>
	<li>Use 5-0 prolene suture to place two interrupted sutures around the 4th and 5th ribs to close the thoracic cavity.</li>
	<li>Remove the skin retractors. Use two sets of forceps to approximate the skin along the length of the incision and glue the skin closed.</li>
	<li>Connect a 3 ml luer-lock syringe to the angiocatheter and remove residual air by applying gentle suction and withdrawing the angiocatheter.</li>
	<li>Glue the neck incision closed using two sets of forceps as before.</li>
</ol>
5. Mouse Sham Pneumonectomy
<ol>
	<li>Expose the left lung as noted in &#8220;Mouse Pneumonectomy&#8221; protocol. Lift the rib cage with curved blunt forceps to allow air into the left chest cavity (Figure 5A,B).</li>
	<li>Place a 24 G angiocatheter into the left thoracic cavity as above being careful not to injure the left lung.</li>
	<li>Using 5-0 prolene suture and being careful not to puncture the lung (Figure 5C), place two lengths of suture material into the 3rd/4th and 5th/6th rib interspaces (Figure 5D). Place both lengths of suture material before tying to lessen the risks of left lung herniation. Tie the suture material to make two interrupted stitches (Figure 5E).</li>
	<li>Glue the skin over the thoracic incision, remove residual air with the angiocatheter, and glue the neck incision as above.</li>
</ol>
6. Resuscitation, Analgesia, and Recovery
<ol>
	<li>Turn off the isoflurane, and administer 0.1 mg/kg of buprenorphine and 0.5 ml of normal saline subcutaneously.</li>
	<li>When spontaneous respirations resume, remove the endotracheal tube.</li>
	<li>Observe mouse until it is again ambulatory. Walking typically resumes several minutes after removal of the endotracheal tube.</li>
	<li>Place the mouse in a 27 &#176;C incubator (humidified, 25% oxygen) to recover O/N. 
	NOTE: We place several pellets of chow wetted with water on the cage floor for the first 24 hr after surgery.</li>
	<li>Administer 0.1 mg/kg of buprenorphine by intraperitoneal injection twice daily for three days after surgery. Take care not to open the surgical site when handling animals.</li>
</ol>
7. Mouse Monitoring
<ol>
	<li>Weigh mice at 1, 3, 5, and 7 days after surgery.</li>
</ol>

<ol><li>Strueby, L., Thebaud, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Advances+in+bronchopulmonary+dysplasia%5BTitle%5D)+AND+%22Expert+review+of+respiratory+medicine%22%5BJournal%5D)">Advances in bronchopulmonary dysplasia.</a> Expert review of respiratory medicine. , (2014).</li>
<li>Donn, S. M., Sinha, S. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Recent+advances+in+the+understanding+and+management+of+bronchopulmonary+dysplasia%5BTitle%5D)+AND+%22Seminars+in+fetal+%26+neonatal+medicine%22%5BJournal%5D)">Recent advances in the understanding and management of bronchopulmonary dysplasia.</a> Seminars in fetal & neonatal medicine. 14, 332(2009).</li>
<li>Molen, T., Miravitlles, M., Kocks, J. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((COPD+management%3A+role+of+symptom+assessment+in+routine+clinical+practice%5BTitle%5D)+AND+%22International+journal+of+chronic+obstructive+pulmonary+disease%22%5BJournal%5D)">COPD management: role of symptom assessment in routine clinical practice.</a> International journal of chronic obstructive pulmonary disease. 8, 461-471 (2013).</li>
<li>Seifert, A. W., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Skin+shedding+and+tissue+regeneration+in+African+spiny+mice+%28Acomys%29%5BTitle%5D)+AND+%22Nature%22%5BJournal%5D)">Skin shedding and tissue regeneration in African spiny mice (Acomys).</a> Nature. 489, 561-565 (2012).</li>
<li>Vidal, P., Dickson, M. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Regeneration+of+the+distal+phalanx.+A+case+report%5BTitle%5D)+AND+%22Journal+of+hand+surgery%22%5BJournal%5D)">Regeneration of the distal phalanx. A case report.</a> Journal of hand surgery. 18, 230-233 (1993).</li>
<li>Potter, P. C., Levine, M. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Bone+Regeneration+Following+Chronic+Suppurative+Osteitis+of+the+Distal+Phalanx%5BTitle%5D)+AND+%22Annals+of+surgery%22%5BJournal%5D)">Bone Regeneration Following Chronic Suppurative Osteitis of the Distal Phalanx.</a> Annals of surgery. 80, 728-729 (1924).</li>
<li>McKim, L. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Regeneration+of+the+Distal+Phalanx%5BTitle%5D)+AND+%22Canadian+Medical+Association+journal%22%5BJournal%5D)">Regeneration of the Distal Phalanx.</a> Canadian Medical Association journal. 26, 549-550 (1932).</li>
<li>Brown, L. M., Rannels, S. R., Rannels, D. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Implications+of+post-pneumonectomy+compensatory+lung+growth+in+pulmonary+physiology+and+disease%5BTitle%5D)+AND+%22Respir+Res%22%5BJournal%5D)">Implications of post-pneumonectomy compensatory lung growth in pulmonary physiology and disease.</a> Respir Res. 2, 340-347 (2001).</li>
<li>Holder, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Regeneration+and+compensatory+growth%5BTitle%5D)+AND+%22British+medical+bulletin%22%5BJournal%5D)">Regeneration and compensatory growth.</a> British medical bulletin. 37, 227-232 (1981).</li>
<li>Hsia, C. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Lessons+from+a+canine+model+of+compensatory+lung+growth%5BTitle%5D)+AND+%22Curr+Top+Dev+Biol%22%5BJournal%5D)">Lessons from a canine model of compensatory lung growth.</a> Curr Top Dev Biol. 64, 17-32 (2004).</li>
<li>Butler, J. P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Evidence+for+adult+lung+growth+in+humans%5BTitle%5D)+AND+%22N+Engl+J+Med%22%5BJournal%5D)">Evidence for adult lung growth in humans.</a> N Engl J Med. 367, 244-247 (2012).</li>
<li>Konerding, M. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Spatial+dependence+of+alveolar+angiogenesis+in+post-pneumonectomy+lung+growth%5BTitle%5D)+AND+%22Angiogenesis%22%5BJournal%5D)">Spatial dependence of alveolar angiogenesis in post-pneumonectomy lung growth.</a> Angiogenesis. 15, 23-32 (2012).</li>
<li>Kho, A. T., Liu, K., Visner, G., Martin, T., Boudreault, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Identification+of+dedifferentiation+and+redevelopment+phases+during+postpneumonectomy+lung+growth%5BTitle%5D)+AND+%22Am+J+Physiol+Lung+Cell+Mol+Physiol%22%5BJournal%5D)">Identification of dedifferentiation and redevelopment phases during postpneumonectomy lung growth.</a> Am J Physiol Lung Cell Mol Physiol. 305, 542-554 (2013).</li>
<li>Wang, W., Nguyen, N. M., Guo, J., Longitudinal Woods, J. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Noninvasive+Monitoring+of+Compensatory+Lung+Growth+in+Mice+after+Pneumonectomy+via+%283%29He+and+%281%29H+Magnetic+Resonance+Imaging%5BTitle%5D)+AND+%22Am+J+Respir+Cell+Mol+Biol%22%5BJournal%5D)">Noninvasive Monitoring of Compensatory Lung Growth in Mice after Pneumonectomy via (3)He and (1)H Magnetic Resonance Imaging.</a> Am J Respir Cell Mol Biol. 49, 697-703 (2013).</li>
<li>Gibney, B. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Detection+of+murine+post-pneumonectomy+lung+regeneration+by+18FDG+PET+imaging%5BTitle%5D)+AND+%22EJNMMI+research%22%5BJournal%5D)">Detection of murine post-pneumonectomy lung regeneration by 18FDG PET imaging.</a> EJNMMI research. 2, 48(2012).</li>
<li>Rawlins, E. L., Perl, A. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((The+a%27MAZE%27ing+world+of+lung-specific+transgenic+mice%5BTitle%5D)+AND+%22Am+J+Respir+Cell+Mol+Biol%22%5BJournal%5D)">The a'MAZE'ing world of lung-specific transgenic mice.</a> Am J Respir Cell Mol Biol. 46, 269-282 (2012).</li>
<li>Ochs, M., Muhlfeld, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Quantitative+microscopy+of+the+lung%3A+a+problem-based+approach.+Part+1%3A+basic+principles+of+lung+stereology%5BTitle%5D)+AND+%22Am+J+Physiol+Lung+Cell+Mol+Physiol%22%5BJournal%5D)">Quantitative microscopy of the lung: a problem-based approach. Part 1: basic principles of lung stereology.</a> Am J Physiol Lung Cell Mol Physiol. 305, L15-22 (2013).</li>
<li>Ysasi, A. B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Effect+of+unilateral+diaphragmatic+paralysis+on+postpneumonectomy+lung+growth%5BTitle%5D)+AND+%22Am+J+Physiol+Lung+Cell+Mol+Physiol%22%5BJournal%5D)">Effect of unilateral diaphragmatic paralysis on postpneumonectomy lung growth.</a> Am J Physiol Lung Cell Mol Physiol. 305, L439-445 (2013).</li>
<li>Dane, D. M., Yilmaz, C., Estrera, A. S., Hsia, C. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Separating+in+vivo+mechanical+stimuli+for+postpneumonectomy+compensation%3A+physiological+assessment%5BTitle%5D)+AND+%22Journal+of+applied+physiology%22%5BJournal%5D)">Separating in vivo mechanical stimuli for postpneumonectomy compensation: physiological assessment.</a> Journal of applied physiology. 114, 99-106 (2013).</li>
<li>Mortola, J. P., Magnante, D., Saetta, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Expiratory+pattern+of+newborn+mammals%5BTitle%5D)+AND+%22Journal+of+applied+physiology%22%5BJournal%5D)">Expiratory pattern of newborn mammals.</a> Journal of applied physiology. 58, 528-533 (1985).</li>
<li>MacDonald, K. D., Chang, H. Y., Mitzner, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((An+improved+simple+method+of+mouse+lung+intubation%5BTitle%5D)+AND+%22Journal+of+applied+physiology%22%5BJournal%5D)">An improved simple method of mouse lung intubation.</a> Journal of applied physiology. 106, 984-987 (2009).</li>
</ol>

The authors have nothing to disclose.

We have provided the most detailed description of the mouse PNX and mouse sham PNX procedures reported to date. We have made the reader aware of several of the common pitfalls that investigators learning the procedure commonly encounter, and we have outlined several techniques developed by our laboratory to mitigate against these pitfalls. Other laboratories utilizing this model may have developed other technique modifications or use different instruments. When evaluating differences in techniques, individual investigato...

The principal function of the lung is to provide for oxygen and carbon dioxide exchange between an organism and the atmosphere. In humans, a host of congenital and acquired conditions lead to reduced lung surface area which results in impaired lung function. Although a host of therapies such as inhaled corticosteroids, bronchodilators, supplemental oxygen, and chronic mechanical ventilation are used to mitigate the consequences of impaired lung function1-3, the ideal therapy for these conditions would promote regrowth of functional lung tissue &#8211; i.e., lung regeneration.
Mammalian tissue regeneration has been well documented. The African Spiny Mouse can regenerate large areas of skin without scar formation4. The distal phalanx in humans can regenerate following injury or amputation5-7. Following pneumonectomy (PNX), compensatory lung growth occurs in mice8, rats9, dogs10, and humans11. By definition, compensatory lung growth involves not only expansion of existing airspaces, but re-septation of these enlarged airspaces with expansion of the associated microcirculation12. Gene expression analysis has demonstrated that this model recapitulates many of the signaling events of lung development13. Four weeks after mouse PNX, alveolar surface area is equivalent to that of sham operated animals14. In this manuscript, we describe the mouse PNX and sham PNX procedures.

<table><tbody><tr><td>6 inch Vascular clip applicator</td><td>Teleflex Medical (WECK)</td><td>137062</td><td></td></tr><tr><td>Horizon small titanium red clip</td><td>Teleflex Medical (WECK)</td><td>1201</td><td></td></tr><tr><td>Narrow pattern 12 cm curved forceps</td><td>Fine Science Tools</td><td>11003-12</td><td></td></tr><tr><td>Curved serrated 10 cm Graefe forceps</td><td>Fine Science Tools</td><td>11052-10</td><td></td></tr><tr><td>Castroviejo needle holder</td><td>Fine Science Tools</td><td>12565-14</td><td></td></tr><tr><td>Straight 9 cm Strabismus scissors (blunt tip)</td><td>Fine Science Tools</td><td>14075-09</td><td></td></tr><tr><td>Straight 8.5 cm hardened fine scissors</td><td>Fine Science Tools</td><td>14090-09</td><td></td></tr><tr><td>Straight, blunt tip Cohan-Vannas spring scissors</td><td>Fine Science Tools</td><td>15000-12</td><td></td></tr><tr><td>Skin glue</td><td>Gluture</td><td>32046</td><td></td></tr><tr><td>22 G Angiocatheter</td><td></td><td></td><td></td></tr><tr><td>24 G Angiocatheter</td><td></td><td></td><td></td></tr><tr><td>3 ml Luer lock syringe</td><td></td><td></td><td></td></tr><tr><td>4 Short retractors</td><td></td><td></td><td></td></tr><tr><td>2 Long retractors</td><td></td><td></td><td></td></tr><tr><td>5-0 Prolene on curved cutting needle</td><td>Ethicon</td><td>8698G</td><td></td></tr><tr><td>0.5 ml Syringe on 27 G needle</td><td></td><td></td><td></td></tr><tr><td>Normal saline</td><td></td><td></td><td></td></tr><tr><td>Buprenorphine</td><td></td><td></td><td></td></tr><tr><td>Press-n-Seal wrap</td><td>Glad Products Company</td><td></td><td></td></tr><tr><td>12 x 12 inch Cork board stack</td><td>Office Depot</td><td></td><td></td></tr><tr><td>70% Ethanol</td><td></td><td></td><td></td></tr><tr><td>Betadine</td><td></td><td></td><td></td></tr><tr><td>Mouse ventilator</td><td>Hugo Sachs Elektronnik</td><td>Minivent Type 845</td><td></td></tr><tr><td>Isoflurane vaporizer</td><td>OHMEDA</td><td>Excel 210 SE</td><td></td></tr><tr><td>Artificial tear ointment</td><td>Puralube</td><td>NDC: 17033-211-38</td><td></td></tr></tbody></table>

mouse pneumonectomy model of compensatory lung growth

Mouse Pneumonectomy Model of Compensatory Lung Growth

In humans, disrupted repair and remodeling of injured lung contributes to a host of acute and chronic lung disorders which may ultimately lead to disability or death. Injury-based animal models of lung repair and regeneration are limited by injury-specific responses making it difficult to differentiate changes related to the injury response and injury resolution from changes related to lung repair and lung regeneration. However, use of animal models to identify these repair and regeneration signaling pathways is critical to the development of new therapies aimed at improving pulmonary function following lung injury. The mouse pneumonectomy model utilizes compensatory lung growth to isolate those repair and regeneration signals in order to more clearly define mechanisms of alveolar re-septation. Here, we describe our technique for performing mouse pneumonectomy and sham pneumonectomy. This technique may be utilized in conjunction with lineage tracing or other transgenic mouse models to define molecular and cellular mechanism of lung repair and regeneration.

A plot of PNX and sham operated mouse weights is provided in Figure 6. In our hands, survival is consistently 95 - 100% for both PNX and sham pneumonectomy. For descriptions of how the right lung re-grows in this model and the expected time course, we refer the reader to manuscripts of Gibney et al.15 and Wang et al.14
Several common pitfalls must be avoided to successfully perform the mouse PNX and mouse sham pneumonectomy procedures.</...

Mouse pneumonectomy is a commonly employed model of compensatory lung growth. This procedure can be used in conjunction with lineage tracing or transgenic mouse models to elucidate underlying mechanisms.

In humans, disrupted repair and remodeling of injured lung contributes to a host of acute and chronic lung disorders which may ultimately lead to ...

The goal of this procedure is to perform a left pneumonectomy to induce compensatory growth of the right lung. This is accomplished by first performing a left thoracotomy and exposing the left hilum. The second step of the procedure is to ligate the left helium and left pulmonary artery.The third step is to remove the left lung. The final step is to close the thoracotomy. The mouse pneumonectomy model of compensatory lung growth can be utilized in conjunction with other biometric or biochemical techniques to elucidate mechanisms of alveolar growth and regeneration.This method can answer some key questions in the lung development and regeneration field, such as what are the key cellular processes involved in septation and what are the critical cellular molecular processes necessary to reinitiate septation? Generally, investigators who are new to this technique will struggle with one of several key steps. They may have difficulty with tracheal cannulation.They may have difficulty creating an adequate surgical window. They may have difficulty identifying the hilum and clipping it, and they also may have difficulty with avoiding injury to the left lung during sham thoracotomy. Demonstrating this procedure will be Shang Lu, a senior laboratory assistant for my laboratory To begin after inducing anesthesia in the mouse with 2%isof fluorine, shave the left thorax and neck area with an electrical shaver, apply artificial tear ointment to the eyes of the mouse and decontaminate the area by swabbing with chlorhexidine three times followed by isopropyl alcohol three times followed by Chlorhexidine.Three times. Confirm the depth of anesthesia by documenting the lack of a response to a poor pinch after making a one centimeter vertical incision over the anterior mid neck to expose the larynx, use serrated 10 centimeter forceps to lightly retract the strap muscles. Then use the tips of a straight scissors to spread the strap muscles.Next orally, insert a 22 gauge blunt tip angio catheter into the mid trachea and visually confirm placement through the incision. Place the mouse on a rodent ventilator and maintain anesthesia with 2%isof fluorine, and a pressure limit of 15 centimeters.Water. Position the still anesthetized mouse in the right lateral decubitus position with the back left side facing the operator.Place self-sealing plastic wrap over the body of the mouse as a sterile drape. Use blunt tipped curved scissors to make a two centimeter long cut parallel to the ribs at the fourth and fifth intercostal space. Then insert the tips of the scissors and dissect the skin away from the underlying ribs and intercostal muscles.Place four retractors on the skin to form a 1.5 by 1.5 centimeter surgical window. Secure each retractor to the cork board. Use curved forceps to dissect down to the ribs and then bluntly dissect the intercostal.To expose the fourth and fifth ribs, use one tip of a curved forceps to make a hole to enter the thorax cavity. Now use the lower blade of a pair of blunt tip micro scissors to depress along the incision until the lower blade enters the thoracic cavity. Make a 0.5 centimeter incision between the ribs and then repeat in the opposite direction.Then open the thorax along the anterior posterior axis by inserting two retractors and securing them to the corkboard. Now load the micro clip applicator. Next, using curved blunt tipped forceps held in the non-dominant hand, grasp the left lung and displace the upper portion laterally and inferiorly through the thoracotomy until the left pulmonary artery and bronchus are exposed.Then hold the loaded titanium vascular micro clip replicator in the dominant hand with the body of the applicator in the palm and the curve tip pointing away from the palm. Slide the applicator tip into the thorax along the curvature of the posterior aspect of the left lung, and clip the left bronchus and pulmonary artery. Remove the applicator but keep the left lung retracted.Grasp the blunt tip micro scissors with the right hand and cut the bronchus and pulmonary artery distal to the clip and remove the left lung. After removing the lung, take out the rib retractors. Then use the curved blunt forceps to pinch up one centimeter of skin inferior to the incision but above the level of the diaphragm, and insert a 24 gauge angio catheter through the skin and into the left thoracic cavity.Next place, two interrupted five zero proline sutures around the fourth and fifth ribs. To close the thoracic cavity, remove the skin retractors and use two sets of forceps to approximate the skin along the length of the incision and use tissue adhesive to glue the skin closed. Finally, connect a three milliliter lu lock syringe to the angio catheter and remove residual air by applying gentle suction and withdrawing the angio catheter.Finished by gluing the neck, incision closed using two sets of forceps as for the closure of the thoracic incision. After exposing the left lung of the anesthetized mouse as previously shown, use curved blunt forceps to lift the rib cage and allow air into the left chest cavity Laser 24 gauge angio catheter into the left, the thoracic cavity as previously shown, being careful not to injure the left lung. Using five zero proline suture and being careful not to puncture the lung.Place two lengths of suture material into the third and fourth and fifth and sixth rib into spaces. Place both lengths of suture material before tying to lessen the risks of left lung herniation. Tie the suture material to make two interrupted stitches.Glue the skin over the thoracic incision. Remove residual air with the angio catheter and glue the neck incision as previously shown. Turn off the isof fluorine and administer 0.1 milligrams per kilogram of buprenorphine and 0.5 milliliters of normal saline subcutaneously.When spontaneous respiration resumes, remove the endotracheal tube. Observe the mouse until it is ambulatory. Walking typically resumes several minutes after removal of the endotracheal tube.Administer 0.1 milligrams per kilogram of buprenorphine by intraperitoneal injection twice daily for three days after surgery. Carefully monitor the animal during these three days and take care not to open the surgical site when handling animals. This image shows weight loss after surgery.24 hours after surgery, pneumo somy mice will lose an average of 1.5 grams and sham mice will lose an average of 1.1 grams. Both groups regain their preoperative weight by postoperative day three. This fluorescent image shows tropo elastin immunofluorescence staining of the accessory lobe under control conditions.Here, tropo elastin immunofluorescence of the accessory lobe is shown 72 hours post pneumo omy existing air spaces are increased. This final TRO Elastin immunofluorescence image shows the accessory lobe four weeks post Pneumo Otomy. Note the Loba growth and Reation.After watching this video, you should have a good understanding of how to perform the mouse pneumonectomy model of compensatory lung growth and how to perform the sham thoracotomy, which is used as a control for this procedure.

mouse-pneumonectomy-model-of-compensatory-lung-growth

Research

JoVE Journal

Medicine

Obliterative bronchiolitis is the key impediment to the long-term survival of lung transplant recipients and the lack of a robust preclinical model precludes examining obliterative bronchiolitis immunopathogenesis. Unlike other solid organ transplants, vascularized mouse lung transplantation has only recently been developed. Here we show our independently developed obliterative bronchiolitis model after murine orthotopic single-lung transplantation.

Development of Obliterative Bronchiolitis in a Murine Model of Orthotopic Lung Transplantation

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’s structural changes with developing pathology.

Measurement of the Pressure-volume Curve in Mouse Lungs

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.

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

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.

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

The rodent left pneumonectomy is a valuable technique in pulmonary hypertension research. Here, we present a protocol to describe the rat pneumonectomy procedure and postoperative care to ensure minimal morbidity and mortality.

The Left Pneumonectomy Combined with Monocrotaline or Sugen as a Model of Pulmonary Hypertension in Rats

Here, we present optimizations to a rat lung transplantation model that serve to improve outcomes. We provide a size guide for cuffs based on body weight, a measurement strategy to ascertain the 4th intercostal space, and methods of wound closure and BAL (bronchoalveolar lavage) fluid and tissue collection.

A Rat Lung Transplantation Model of Warm Ischemia/Reperfusion Injury: Optimizations to Improve Outcomes

We present an ex vivo pig lung model for the demonstration of pulmonary mechanics and alveolar recruitment maneuvers for teaching purposes. The lungs can be used for more than one day (up to five days) with minimal changes in pulmonary mechanics variables.

Ex Vivo Porcine Experimental Model for Studying and Teaching Lung Mechanics

This protocol describes the development of a neonatal piglet model of acute lung injury that models early pathogenic events occurring in the preterm lung, including sub-adequate surfactant quantity, hyperoxia, high-pressure ventilation, and inflammation, to facilitate understanding of molecular triggers of bronchopulmonary dysplasia and enhance therapeutic translation.

Development of a Neonatal Piglet Acute Lung Injury Model Recreating the Early Environment of Preterm Infant Lungs

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.

A Simple Method of Mouse Lung Intubation

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.

Diagnostic Ultrasound Imaging of Mouse Diaphragm Function

Learn the mouse pneumonectomy model to study compensatory lung growth and alveolar regeneration mechanisms. This surgical technique enables researchers to investigate lung development and tissue repair processes in vivo.

Mouse Pneumonectomy Model of Compensatory Lung Growth

Explore More Videos