The authors have no acknowledgements.

This prospective experimental trial with 21 anesthetized male and female domestic pigs (German landrace) at the age of 3&#8211;6 months with a body weight between 45-55 kg was approved by the Governmental Commission on the Care and Use of Animals of the City of Hamburg (Reference-No. 18/17). According to the ARRIVE guidelines, all experiments were carried out and all animals received care in compliance with the &#39;Guide for the Care and Use of Laboratory Animals&#39; (NIH publication No. 86-23, revised 1996)11.
1. Flow Probe Two-point Calibration
<ol>
	<li>Put flow probes in deionized water and connect the probe to the transonic flow probe system by putting the plug into the perivascular flow module.</li>
	<li>Open the data analysis software (e.g., LabChart 8).</li>
	<li>For a two-point calibration, start a measurement by setting the flow probe system to Zero and after a few seconds to Scale.</li>
	<li>In the data analysis software window, go to Units Conversion and choose two-point calibration. Mark a baseline to set to zero. Then, mark an area with 10 L/min and set to 1 V as a preset value.</li>
	<li>Repeat the procedure for the other probe.</li>
</ol>
2. Millar Catheter Calibration
<ol>
	<li>Prior to the zeroing and calibration, pre-soak the tip of the catheter in sterile body temperature warm water for 30 min.</li>
	<li>Connect the Millar catheter to the bridge amplifier by putting the plug into the bridge amplifier module.</li>
	<li>Start the data analysis software.</li>
	<li>Put the tip of the catheter into the pneumatic zeroing tool, set the value to 0 mmHg and start a measurement by clicking Start in the program.</li>
	<li>Keep the measurement running and set the pneumatic zeroing tool to 100 mmHg. Stop the measurement by clicking Stop.</li>
	<li>Run the data analysis software by pressing Start and then press Stop. Click Amplify in the window of bridge and choose Units. Set the baseline of 0 and 100 mmHg, accordingly. According to the preset value for calibration that the software provides, the catheter is now calibrated for all body pressures.</li>
	<li>Repeat the procedure for the other Millar catheter.</li>
</ol>
3. Preparation of the Pig
<ol>
	<li>Medicate the pig by injecting 20 mg/kg of Ketanest, 4 mg/kg of Azaperon and 0.1 mg/kg Midazolam intramuscularly and place a 20 G IV-line into a vein of the ear.</li>
	<li>Place ECG stickers on the chest and oxygen probe on the tail.</li>
	<li>Administer pure oxygen (15&#8211;18 L/min) via the pig&#39;s nose using a mask and surgically prepare down to the trachea.</li>
	<li>Put a loop around the trachea, use a scalpel (11 blade) to make an incision into the trachea and place an 8.5 Mallinckrodt tube into it for a safe airway. Fix the tube with the preset loop and close the skin with sutures.</li>
	<li>Begin anesthesia with Sevoflurane using a MAC of 0.9 (in adolescent pigs equivalent to an endexpiratory concentration of 2.0%)&#160;&#160;and infusion of 0.01 mg/(kg&#8729;h) Fentanyl. Start mechanical ventilation with a tidal volume of 10 mL/kg, a rate of 14/min, and a positive end expiratory pressure (PEEP) of 7 mmHg. Set the inspiratory oxygen rate (fiO2) to 0.3. After 10 min depth of anesthesia is deep enough to perform surgery safely. No elevation of HR and BP should be detected.</li>
	<li>Maintain fluid balance at basal volume rate of 10 mL/(kg&#8729;h) cristalloid using an infusion pump.</li>
	<li>Gently clean the pig&#39;s skin using soap water. Use a skin disinfection solution containing povidone-iodine to decrease skin contamination.</li>
</ol>
4. Vital Parameter Measurements
<ol>
	<li>Use an ultrasound for inserting a 5 F thermistor tipped arterial catheter into the right femoral artery, an 8 F introducer sheath into the left femoral artery, a central venous catheter and an 8 F introducer sheath into the jugular vein (Figure 1).</li>
	<li>Place the catheter placement using Seldinger&#39;s technique12.
	<ol>
		<li>Place a needle into the target vessel under ultrasound vision.</li>
		<li>Put a wire through the needle into the vessel, verify the correct placement of the wire using ultrasound and keep the wire in the vessel throughout the whole procedure. Remove the needle and place a dilator onto the wire.</li>
		<li>With gentle pressure, put the dilator through the skin into the vessel using the wire as guidance. Remove the dilator, put the catheter onto the wire, make sure to the end of the wire is seen at the end of the catheter and place the catheter it into the vessel.</li>
		<li>Remove the wire by gently pulling it out of the catheter.</li>
	</ol>
	</li>
	<li>Insert a 7F pulmonary artery catheter (PAC) into the 8F introducer sheath and place it in the RV. If needed for taking mixed venous blood gas samples, insert the PAC further into the PA until a pulmonary artery curve is shown on to the monitor and pull it back after receiving the samples.</li>
	<li>Insert the first Millar-tip catheter into the 8F introducer sheath in the left femoral artery and placing it in the aorta.</li>
	<li>Perform a mini-laparotomy (approximately 5&#8211;10 cm is enough) above the symphysis by using the electrocautery for prepping down to the linea alba.
	<ol>
		<li>Open the linea alba with scissors and pull out the bladder very gently.</li>
		<li>Put a purse string suture in the bladder using a 3/0 suture and make an incision into the bladder with a scalpel (11 blade).</li>
		<li>Insert a urinary catheter into the bladder, inflate the catheter&#39;s balloon with water and fix it using the purse-string suture. Close the abdomen with a 3/0 suture.</li>
	</ol>
	</li>
</ol>
5. Surgical Preparation of the Heart
<ol>
	<li>Before opening the chest increasing the fiO2 to 1.0 and administer 0.1 mg kg(-1) pancuronium initial bolus intravenously13.</li>
	<li>Perform a median sternotomy.
	<ol>
		<li>Use the electrocautery for prepping down to the sternum. Gently dissect the sternum from the surrounding tissue before dividing the bone with an oscillating saw.</li>
		<li>Use the electrocautery to reduce bleeding and seal the sternum with bone wax. Place a sternal rib spreader between the two halves of the opened sternum and widely open the chest as much as needed for surgery by twisting the handle on the device.</li>
	</ol>
	</li>
	<li>Open the pericardium gently using scissors and forceps and fix it to the skin with a 2/0 suture.</li>
	<li>Dissect down the pulmonary and the artery ascending aorta very gently to avoid bleeding. Carefully place the ultrasound flow probes around both arteries, respectively (Figure 2).</li>
	<li>Place 2 purse string sutures in the pulmonary artery using a 5/0 suture. Use a scalpel (11 blade) to make a small stitch incision (approximately 1 mm) in the middle of the purse strings and place the Millar catheter into the pulmonary artery before fixing it (Figure 3).</li>
	<li>Carefully clamp the LAA and place 2 purse string sutures in it using a 4/0 suture. Make a small incision and place a central venous line into the left atrium before fixing it using the purse string sutures (Figure 3).</li>
	<li>Close the pericardium by suturing a sterile glove onto it, to maintain hemodynamics reliable (Figure 4). Perform the sternal closure with wires and close the skin with a 3/0 suture.</li>
</ol>
6. Assessment and Data Acquisition
<ol>
	<li>Start each measurement with 2 min of AO and PA flow measurements, as well as AO and PA pressure measurements using the data analysis software by clicking Start and Stop button in the program.</li>
	<li>Perform transcardiopulmonary thermodilution to provide cardiac output (CO) as well as pulse pressure variance (PPV) and stroke volume variance (SVV) by using the PiCCO2 system. To start the measurement, click the TD | Start.</li>
	<li>Consecutive inject 15 mL of 10 &#176;C cold saline into a thermistor at the central venous line in the jugular vein three times for thermodilution at each measurement step.</li>
	<li>Take an arterial, central venous and mixed venous blood gas sample after each transcardiopulmonary thermodilution measurement step.</li>
</ol>
7. Volume Optimizing
<ol>
	<li>After a baseline measurement M0 (steps 6.1&#8211;6.4) of all parameters, administer a volume loading step using 5 mL/kg of colloidal infusion (Voluven) using an infusion pump that is connected to the central venous line.</li>
	<li>After 5 min of equilibration, start another measurement step M1 (steps 6.1&#8211;6.4). If the newly generated cardiac output measured by thermodilution using the PICCO2 system (see step 6.2&#8211;6.3) does not increase compared to the formerly measured CO by at least 10%, start another volume loading step (step 7.1).</li>
	<li>Continue with volume loading and equilibration steps until there is no more increase in CO of more than 10%. Now, a balanced fluid status is reached.</li>
</ol>
8. Induction of ARDS with Right Ventricular Dysfunction
<ol>
	<li>Increase the fiO2 to at least 0.5 to 0.8 as required to maintain a spO2 of at least 90%.</li>
	<li>Induce an ARDS with consecutive right ventricular dysfunction by infusion of oleic acid (OA) (0.03&#8211;0.06 mL/kg for about 2 h).</li>
	<li>Use continuous administration of adrenalin using a perfusor (3 mg of adrenalin in 50 mL of saline) to keep hemodynamics stable. Increase the infusion rate as required to maintain a mean arterial pressure of 50 mmHg.</li>
	<li>Add calcium, magnesium and antiarrhythmics (1% Lidocain) as required during the infusion of OA to maintain a stable sinus rhythm.</li>
</ol>
9. Volume Optimizing
<ol>
	<li>After induction of mild to moderate ARDS, perform another measurement of all parameters (M2) by completing steps 6.1&#8211;6.4. 
	NOTE: Now, the baseline model for hemodynamic measurements in ARDS in a pig model is set. For further investigation on volume responsiveness in ARDS and right ventricular dysfunction start to reduce volume load by taking as much blood as need per protocol or increasing volume load by adding a defined amount of infusion.</li>
</ol>
10. Finalization
<ol>
	<li>After finishing the measurements euthanize the pigs while under anesthesia by injecting 1mmol/kg potassium chloride intravenously.</li>
</ol>

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K., 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=Mechanical+Circulatory+Support+Devices+for+Acute+Right+Ventricular+Failure.">Mechanical Circulatory Support Devices for Acute Right Ventricular Failure.</a> Circulation. 136, 314-326 (2017).</li><li>Zochios, V., Jones, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Acute+right+heart+syndrome+in+the+critically+ill+patient.">Acute right heart syndrome in the critically ill patient.</a> Heart Lung Vessel. 6 (3), 157-170 (2014).</li><li>Ranucci, M., 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=Fluid+responsiveness+and+right+ventricular+function+in+cardiac+surgical+patients.+A+multicenter+study.">Fluid responsiveness and right ventricular function in cardiac surgical patients. A multicenter study.</a> HSR Proceedings in Intensive Care and Cardiovascular Anesthesia. 1 (1), 21-29 (2009).</li><li>Mehta, S. R., 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=Impact+of+right+ventricular+involvement+on+mortality+and+morbidity+in+patients+with+inferior+myocardial+infarction.">Impact of right ventricular involvement on mortality and morbidity in patients with inferior myocardial infarction.</a> Journal of the American College of Cardiology. 37, 37-43 (2001).</li><li>Vieillard-Baron, A., Charron, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Preload+responsiveness+or+right+ventricular+dysfunction.">Preload responsiveness or right ventricular dysfunction.</a> Critical Care Medicine. 37 (9), 2662-2663 (2009).</li><li>Marik, P. 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B., Quintel, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Relationship+among+cardiac+index,+inspiration/expiration+ratio,+and+perfluorocarbon+dose+during+partial+liquid+ventilation+in+an+oleic+acid+model+of+acute+lung+injury+in+sheep.">Relationship among cardiac index, inspiration/expiration ratio, and perfluorocarbon dose during partial liquid ventilation in an oleic acid model of acute lung injury in sheep.</a> Journal of Pediatric Surgery. 40 (9), 1395-1403 (2005).</li><li>Zhu, Y. B., 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=Atrial+natriuretic+peptide+attenuates+inflammatory+responses+on+oleic+acid-induced+acute+lung+injury+model+in+rats.">Atrial natriuretic peptide attenuates inflammatory responses on oleic acid-induced acute lung injury model in rats.</a> Chinese Medical Journal (English. 126 (4), 747-750 (2013).</li><li>Gould, D. A., Baun, M. 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+Role+of+the+Pulmonary+Afferent+Receptors+in+Producing+Hemodynamic+Changes+during+Hyperinflation+and+Endotracheal+Suctioning+in+an+Oleic+Acid-Injured+Animal+Model+of+Acute+Respiratory+Failure.">The Role of the Pulmonary Afferent Receptors in Producing Hemodynamic Changes during Hyperinflation and Endotracheal Suctioning in an Oleic Acid-Injured Animal Model of Acute Respiratory Failure.</a> Biology Research for Nursing. 1 (3), 179-189 (2000).</li><li>Galie, 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=2015+ESC/ERS+Guidelines+for+the+diagnosis+and+treatment+of+pulmonary+hypertension.">2015 ESC/ERS Guidelines for the diagnosis and treatment of pulmonary hypertension.</a> European Heart Journal. 37, 67-119 (2015).</li><li>Galie, 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=2015+ESC/ERS+Guidelines+for+the+diagnosis+and+treatment+of+pulmonary+hypertension:+The+Joint+Task+Force+for+the+Diagnosis+and+Treatment+of+Pulmonary+Hypertension+of+the+European+Society+of+Cardiology+(ESC)+and+the+European+Respiratory+Society+(ERS),+Endorsed+by:+Association+for+European+Paediatric+and+Congenital+Cardiology+(AEPC),+International+Society+for+Heart+and+Lung+Transplantation+(ISHLT).">2015 ESC/ERS Guidelines for the diagnosis and treatment of pulmonary hypertension: The Joint Task Force for the Diagnosis and Treatment of Pulmonary Hypertension of the European Society of Cardiology (ESC) and the European Respiratory Society (ERS), Endorsed by: Association for European Paediatric and Congenital Cardiology (AEPC), International Society for Heart and Lung Transplantation (ISHLT).</a> European Respiratory Journal. 46, 903-975 (2015).</li><li>Oliveira, R. K., 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=Usefulness+of+pulmonary+capillary+wedge+pressure+as+a+correlate+of+left+ventricular+filling+pressures+in+pulmonary+arterial+hypertension.">Usefulness of pulmonary capillary wedge pressure as a correlate of left ventricular filling pressures in pulmonary arterial hypertension.</a> Journal of Heart and Lung Transplantation. 33 (4), 459 (2014).</li><li>Hoeper, M. M., 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+global+view+of+pulmonary+hypertension.">A global view of pulmonary hypertension.</a> Lancet Respiratory Medicine. 4, 306-322 (2016).</li><li>Nagy, A. I., 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+pulmonary+capillary+wedge+pressure+accurately+reflects+both+normal+and+elevated+left+atrial+pressure.">The pulmonary capillary wedge pressure accurately reflects both normal and elevated left atrial pressure.</a> American Heart Journal. 167 (6), 876-883 (2014).</li><li>Daughters, G. T., 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=Effects+of+the+pericardium+on+left+ventricular+diastolic+filling+and+systolic+performance+early+after+cardiac+operations.">Effects of the pericardium on left ventricular diastolic filling and systolic performance early after cardiac operations.</a> Journal of Thoracic and Cardiovascular Surgery. 104 (4), 1084-1091 (1992).</li><li>Zimmerman, R., 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=Posttransfusion+Increase+of+Hematocrit+per+se+Does+Not+Improve+Circulatory+Oxygen+Delivery+due+to+Increased+Blood+Viscosity.">Posttransfusion Increase of Hematocrit per se Does Not Improve Circulatory Oxygen Delivery due to Increased Blood Viscosity.</a> Anesthesia &amp; Analgesia. 124 (5), 1547-1554 (2017).</li><li>Giglioli, C., 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=Hemodynamic+effects+in+patients+with+atrial+fibrillation+submitted+to+electrical+cardioversion.">Hemodynamic effects in patients with atrial fibrillation submitted to electrical cardioversion.</a> International Journal of Cardiology. 168 (4), 4447-4450 (2013).</li></ol>

Daniel A. Reuter is a member of Pulsion Medical Advisory Board. Constantin J.C. Trepte has received honorary award for lectures by Maquet. All other authors declare no conflicts of interest.

ARDS, complicated by pulmonary hypertension, is a very deadly disease. For patients suffering from this condition, further information about treating it is necessary. When working and researching with living creatures, it is very important to be as sensible as possible. In this case it is necessary to gather as much information as possible in one experiment.
There are some critical surgical steps in an open-beating heart model like this. To not use pigs unnecessarily, there must be an experienced surgeon to dissect the heart skeleton between the ascending aorta and the pulmonary artery while hemodynamics are unstable due to the pressure on the RV and RA. Another critical step is putting the Millar tip catheter into the pulmonary artery. To get a better exposure of the surgical field, the right ventricular outflow tract (RVOT) needs to be pushed away very gently. With the right amount of pressure, it is possible have good visualibility and stability of the PA. This makes it easier to take small bites with the 5.0 suture and decreases the risk of PA bleeding or injury.
When measuring hemodynamics, losing a great amount of blood and thus changing the hematocrit significantly can influence the measurements and the results20. When placing the catheter into the artery, using a tourniquet first and making a very small incision to fix the catheter quickly could prevent any blood loss. Ensure that all small bleedings are stopped before insertion of the Millar catheter, because electrocautery can damage the catheter (as described in the catheters manual). After closing of the pericardium and the sternum small bleedings can accumulate over time and cause changes in hematocrit or cause a pericardial tamponade with significant changes in hemodynamics. This could cause a termination of the experiment.
When cutting into the LA, one must be careful. The LA is the pacemaker of the heart and it can react with heart rhythm disturbances when touching it with cold metal instruments. Before putting the clamp gently around the LAA, administration of magnesium could prevent atrial fibrillation (AF). Rhythm disturbances like AF have great impact on left as well as right ventricular hemodynamics21.

Right ventricular (RV) dysfunction is a major cause of morbidity and mortality in patients with heart failure1, especially if the underlying cause is pulmonary hypertension2. The RV pumps blood into the low-resistance pulmonary system, which is normally associated with high compliance. Therefore, the RV is characterized by low peak systolic pressure. It also generates one sixth the stroke work compared with the left ventricle (LV)3. Due to its thinner muscle, the RV is very vulnerable to a change in pre- and afterload4,5. The isovolumic phases of contraction and relaxation during systole and diastole in the RV are not as distinct as in the LV. The examination of left and right ventricular hemodynamic parameters is highly important in therapy of critically ill patients with acute right heart distress4,7, because RV failure increases short-term mortality significantly6.
Preload parameters like the central venous pressure (CVP) and left ventricular preload parameters like pulmonary capillary wedge pressure (PCWP) have been used for a long time to determine volume status of patients. Lately, it has been shown that these parameters alone are not suitable to detect a patient&#39;s need of fluids8,9,10. Recognizing fluid responsiveness is essential to detect and treat volume deprivation and volume overload in patients with RV dysfunction. Avoiding volume overload is essential to decrease the mortality and length of intensive care unit (ICU) stay in these patients.
With this study, we established a pig model of right ventricular dysfunction that is consistent and replicable. Due to its similarity to humans, it is necessary to establish consistent and reproducible experimental large animal models of cardiac hemodynamics and measurements for research purpose.

<table>
	<tbody>
		<tr>
			<td>Animal Bio Amp</td>
			<td>ADInstruments</td>
			<td>FE136</td>
			<td></td>
		</tr>
		<tr>
			<td>Quad BridgeAmp</td>
			<td>ADInstruments</td>
			<td>FE224</td>
			<td></td>
		</tr>
		<tr>
			<td>Power Lab 16/35</td>
			<td>ADInstruments</td>
			<td>5761-E</td>
			<td></td>
		</tr>
		<tr>
			<td>LabChart 8.1.8 Windows</td>
			<td>ADInstruments</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Pulmonary artery catheter 7 F</td>
			<td>Edwards Lifesciences Corporation</td>
			<td>&#160;&#160;131F7&#160;</td>
			<td></td>
		</tr>
		<tr>
			<td>Prelude Sheath Introducer 8 F</td>
			<td>Merit Medical Systems, Inc.</td>
			<td>SI-8F-11-035</td>
			<td></td>
		</tr>
		<tr>
			<td>COnfidence Cardiac Output Flowprobes</td>
			<td>Transonic</td>
			<td>AU-IFU-PAUProbes-EN Rev. A 4/13</td>
			<td></td>
		</tr>
		<tr>
			<td>Adrenalin</td>
			<td>Sanofi</td>
			<td>6053210</td>
			<td></td>
		</tr>
		<tr>
			<td>Oleic acid</td>
			<td>Sigma Aldrich</td>
			<td>112-80-1</td>
			<td></td>
		</tr>
		<tr>
			<td>Magnesium Verla</td>
			<td>Verla</td>
			<td>7244946</td>
			<td></td>
		</tr>
		<tr>
			<td>Ketamin</td>
			<td>Richter Pharma AG</td>
			<td>BE-V433246</td>
			<td></td>
		</tr>
		<tr>
			<td>Azaperon</td>
			<td>Sanochemia Pharmazeutika AG</td>
			<td>QN05AD90</td>
			<td></td>
		</tr>
		<tr>
			<td>Midazolam</td>
			<td>Roche Pharma AG</td>
			<td>3085793</td>
			<td></td>
		</tr>
	</tbody>
</table>

invasive hemodynamic monitoring of aortic and pulmonary artery hemodynamics in a large animal model of ards

One of the leading causes of morbidity and mortality in patients with heart failure is right ventricular (RV) dysfunction, especially if it is due to pulmonary hypertension. For a better understanding and treatment of this disease, precise hemodynamic monitoring of left and right ventricular parameters is important. For this reason, it is essential to establish experimental pig models of cardiac hemodynamics and measurements for research purpose. 
This article shows the induction of ARDS by using oleic acid (OA) and consequent right ventricular dysfunction, as well as the instrumentation of the pigs and the data acquisition process that is needed to assess hemodynamic parameters. To achieve right ventricular dysfunction, we used oleic acid (OA) to cause ARDS and accompanied this with pulmonary artery hypertension (PAH). With this model of PAH and consecutive right ventricular dysfunction, many hemodynamic parameters can be measured, and right ventricular volume load can be detected. 
All vital parameters, including respiratory rate (RR), heart rate (HR) and body temperature were recorded throughout the whole experiment. Hemodynamic parameters including femoral artery pressure (FAP), aortic pressure (AP), right ventricular pressure (peak systolic, end systolic and end diastolic right ventricular pressure), central venous pressure (CVP), pulmonary artery pressure (PAP) and left arterial pressure (LAP) were measured as well as perfusion parameters including ascending aortic flow (AAF) and pulmonary artery flow (PAF). Hemodynamic measurements were performed using transcardiopulmonary thermodilution to provide cardiac output (CO). Furthermore, the PiCCO2 system (Pulse Contour Cardiac Output System 2) was used to receive parameters such as stroke volume variance (SVV), pulse pressure variance (PPV), as well as extravascular lung water (EVLW) and global end-diastolic volume (GEDV). Our monitoring procedure is suitable for detecting right ventricular dysfunction and monitoring hemodynamic findings before and after volume administration.

Our animal model shows a broad variety of hemodynamic parameters in pigs. Due to its similarity in size and hemodynamics, one can easily use the exact same equipment as used in humans to get similar results.&#160;However, anesthesia values are based on experience and may change based on weight/ age/ strain of pig.&#160; A veterinarian should be consulted to evaluate anesthetic plan.
Results of previous OA induced acute lung injury (ALI) models were inconsistent13,14,15,16. Former protocols stated that OA has been administered mixing it with blood, normal saline, or purely administering it into the heart, a central vein or a peripheral vein in doses of 0.6&#8211;2 mL/kg bodyweight17,18. We tried all of the above methods and found out that purely administering low doses of OA (0.03&#8211;0.06 mL/kg for about 2 h) achieved the most consistent results of ARDS without losing any animals due to respiratory failure or severe acute right heart failure.
First, we were able to show that the intravenous infusion of OA is an easy and good model to induce ARDS as shown before. Depending on the amount of OA administered, one gets a mild to severe lung injury up to death13. It has been shown that an amount of about 0.1 mL/kg OA is mostly used to have a moderate ALI16,18.
To get a mild to moderate ARDS that can be used for further investigation, it is sufficient to inject 0.03-0.06 mL/kg OA. After the administration of this small amount of OA, the oxygenation index decreased from 516.83 &#177; 50.25 mmHg to 181.19 &#177; 32.25 mmHg (p = 0.0006) (Figure 6). The decrease of oxygenated blood is accompanied with a statistically significant increase in carboxylated blood from 36.71 &#177; 4.51 mmHg to 46.50 &#177; 6.87 mmHg (p = 0.008) (Figure 7).
Pulmonary hypertension is defined as a PAP of more than 25 mmHg, a PCWP (which equals the left arterial pressure) of &#8804; 15 mmHg and a pulmonary vascular resistance (PVR) &#62; 240 dyn &#215; s &#215; cm&#8722;5 19,20,21. There is a prevalence of about 1%17 with this common disease worldwide. The PCWP accurately reflects both normal and elevated LAP and vice versa18. In our open-heart animal model, we used a catheter placed in the left atrial to measure this value, because a PAC placed in the pulmonary artery through the pulmonary flow probe could cause incorrect flow measurements (Figure 5).
For a correct and especially consistent measurement of the PAP, we used a Millar catheter, which is put directly in the PA and placed in the main pulmonary artery (MPA) about 2 cm after the pulmonary valve.
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/57405/57405fig1.jpg" /> 
Figure 1: For safe and easy airway management during the whole surgery, perform a tracheotomy and placement of an 8.5 tube directly in the trachea. The bigger the inner diameter of the tube, the better for mechanical ventilation during ARDS. The catheters in the right jugular vein and both femoral arteries are placed by ultrasound using Seldinger&#39;s technique. <a href="/files/ftp_upload/57405/57405fig1large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/57405/57405fig2.jpg" /> 
Figure 2: After opening the pericardium, push the RV and RAA away gently for better visualization of the aorta and pulmonary artery. Hemodynamics must be monitored closely during these steps due to a decreased cardiac output. Dissect the connective tissue of the cardiac skeleton between the PA and the aorta gently, especially as the PA is very predisposed towards bleeding due to its thinner wall. Choose the right sized chronic lined low probes (mostly 18&#8211;20 mm) to put around the aorta and the pulmonary artery. <a href="/files/ftp_upload/57405/57405fig2large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 3" class="xfigimg" src="/files/ftp_upload/57405/57405fig3.jpg" /> 
Figure 3: Use a vascular clamp to fix the LAA and to avoid bleeding. For safe and secure surgery, place two purse string sutures around an edge of the LAA, making a small incision and putting the catheter into the heart. Quickly open the clamp to position the catheter approximately 5 cm deep into the left atrium while monitoring the pressure curve. Reposition the catheter as needed. Fix the catheter by using the purse string sutures. <a href="/files/ftp_upload/57405/57405fig3large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 4" class="xfigimg" src="/files/ftp_upload/57405/57405fig4.jpg" /> 
Figure 4: Very gently put two purse string sutures into the PA. To avoid unnecessary bleeding, use a tourniquet on one of the purse strings. Make a small incision and put the Millar catheter into the pulmonary artery and immediately pull down the tourniquet. Fix it using both sutures. Impose the probe shell on both aortal and pulmonary flow probes. <a href="/files/ftp_upload/57405/57405fig4large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 5" class="xfigimg" src="/files/ftp_upload/57405/57405fig5.jpg" /> 
Figure 5: Use a patch to close the pericardium. Because opening of the pericardium during cardiac surgery goes along with an increase in CO and stroke work index, we chose to close the pericardium using a patch to maintain hemodynamic conditions similar to the ones prior to surgery19. <a href="/files/ftp_upload/57405/57405fig5large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 6" class="xfigimg" src="/files/ftp_upload/57405/57405fig6.jpg" /> 
Figure 6: Since we increased the oxygen fraction due to the pulmonary impairment of the ARDS, the oxygenation index was calculated for each measurement step. We were able to see a decrease from 516.83 &#177; 50.25 mmHg at the baseline measurement (1) to 181.19 &#177; 32.25 mmHg (p = 0.0006) after administration of OA (5). <a href="/files/ftp_upload/57405/57405fig6large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 7" class="xfigimg" src="/files/ftp_upload/57405/57405fig7.jpg" /> 
Figure 7: Along with the decrease of oxygenated arterial blood goes a statistically significant increase in carboxylated blood after induction of ARDS. The baseline measurement was at 36.71 &#177; 4.51 mmHg and increased to 46.50 &#177; 6.87 mmHg (p = 0.008) after administration of OA. <a href="/files/ftp_upload/57405/57405fig7large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>

Watch this Scientific Journal Video about Invasive Hemodynamic Monitoring of Aortic and Pulmonary Artery Hemodynamics in a Large Animal Model of ARDS at JoVE.com

Invasive Hemodynamic Monitoring of Aortic and Pulmonary Artery Hemodynamics in a Large Animal Model of ARDS

We present a protocol of creating right ventricular dysfunction in a pig model by inducing ARDS. We demonstrate invasive monitoring of left and right ventricular cardiac output using flow probes around the aorta and the pulmonary artery, as well as blood pressure measurements in the aorta and pulmonary artery.

One of the leading causes of morbidity and mortality in patients with heart failure is right ventricular (RV) dysfunction, especially if it is due to ...

invasive-hemodynamic-monitoring-aortic-pulmonary-artery-hemodynamics

Nanyang Technological University

Research

JoVE Journal

Medicine

10.0K Views.  Universitatsklinikum Hamburg-Eppendorf. We present a protocol of creating right ventricular dysfunction in a pig model by inducing ARDS. We demonstrate invasive monitoring of left and right ventricular cardiac output using flow probes around the aorta and the pulmonary artery, as well as blood pressure measurements in the aorta and pulmonary artery.

Video: Invasive Hemodynamic Monitoring of Aortic and Pulmonary Artery Hemodynamics in a Large Animal Model of ARDS

An in vivo Rodent Model of Contraction-induced Injury and Non-invasive Monitoring of Recovery

Muscle strains are one of the most common complaints treated by physicians. A muscle injury is typically diagnosed from the patient history and physical exam alone, however the clinical presentation can vary greatly depending on the extent of injury, the patient's pain tolerance, etc. In patients with muscle injury or muscle disease, assessment of muscle damage is typically limited to clinical signs, such as tenderness, strength, range of motion, and more recently, imaging studies. Biological markers, such as serum creatine kinase levels, are typically elevated with muscle injury, but their levels do not always correlate with the loss of force production. This is even true of histological findings from animals, which provide a "direct measure" of damage, but do not account for all the loss of function. Some have argued that the most comprehensive measure of the overall health of the muscle in contractile force. Because muscle injury is a random event that occurs under a variety of biomechanical conditions, it is difficult to study. Here, we describe an in vivo animal model to measure torque and to produce a reliable muscle injury. We also describe our model for measurement of force from an isolated muscle in situ. Furthermore, we describe our small animal MRI procedure.

An in vivo Rodent Model of Contraction-induced Injury and Non-invasive Monitoring of Recovery

An in vivo animal model of injury is described. The method takes advantage of the subcutaneous position of the fibular nerve. Velocity, timing of muscle activation, and arc of motion are all pre-determined and synchronized using commercial software. Post injury changes are monitored in vivo using MR imaging/spectroscopy.

Muscle strains are one of the most  common complaints treated by physicians. A muscle injury is typically diagnosed from the patient history and  physical ...

Non-invasive Optical Measurement of Cerebral Metabolism and Hemodynamics in Infants

Perinatal brain injury remains a significant cause of infant mortality and morbidity, but there is not yet an effective bedside tool that can accurately screen for brain injury, monitor injury evolution, or assess response to therapy. The energy used by neurons is derived largely from tissue oxidative metabolism, and neural hyperactivity and cell death are reflected by corresponding changes in cerebral oxygen metabolism (CMRO2). Thus, measures of CMRO2 are reflective of neuronal viability and provide critical diagnostic information, making CMRO2 an ideal target for bedside measurement of brain health.
Brain-imaging techniques such as positron emission tomography (PET) and single-photon emission computed tomography (SPECT) yield measures of cerebral glucose and oxygen metabolism, but these techniques require the administration of radionucleotides, so they are used in only the most acute cases.
Continuous-wave near-infrared spectroscopy (CWNIRS) provides non-invasive and non-ionizing radiation measures of hemoglobin oxygen saturation (SO2) as a surrogate for cerebral oxygen consumption. However, SO2 is less than ideal as a surrogate for cerebral oxygen metabolism as it is influenced by both oxygen delivery and consumption. Furthermore, measurements of SO2 are not sensitive enough to detect brain injury hours after the insult 1,2, because oxygen consumption and delivery reach equilibrium after acute transients 3. We investigated the possibility of using more sophisticated NIRS optical methods to quantify cerebral oxygen metabolism at the bedside in healthy and brain-injured newborns. More specifically, we combined the frequency-domain NIRS (FDNIRS) measure of SO2 with the diffuse correlation spectroscopy (DCS) measure of blood flow index (CBFi) to yield an index of CMRO2 (CMRO2i) 4,5.
With the combined FDNIRS/DCS system we are able to quantify cerebral metabolism and hemodynamics. This represents an improvement over CWNIRS for detecting brain health, brain development, and response to therapy in neonates. Moreover, this method adheres to all neonatal intensive care unit (NICU) policies on infection control and institutional policies on laser safety. Future work will seek to integrate the two instruments to reduce acquisition time at the bedside and to implement real-time feedback on data quality to reduce the rate of data rejection.

We combined frequency-domain near-infrared spectroscopy measures of cerebral hemoglobin oxygenation with diffuse correlation spectroscopy measures of cerebral blood flow index to estimate an index of oxygen metabolism. We tested the utility of this measure as a bedside screening tool to evaluate the health and development of the newborn brain.

Perinatal brain injury remains a significant cause of infant mortality  and morbidity, but there is not yet an effective bedside tool that can  accurately ...

Monitoring of Systemic and Hepatic Hemodynamic Parameters in Mice

The use of mouse models in experimental research is of enormous importance for the study of hepatic physiology and pathophysiological disturbances. However, due to the small size of the mouse, technical details of the intraoperative monitoring procedure suitable for the mouse were rarely described. Previously we have reported a monitoring procedure to obtain hemodynamic parameters for rats. Now, we adapted the procedure to acquire systemic and hepatic hemodynamic parameters in mice, a species ten-fold smaller than rats. This film demonstrates the instrumentation of the animals as well as the data acquisition process needed to assess systemic and hepatic hemodynamics in mice. Vital parameters, including body temperature, respiratory&#160;rate&#160;and heart&#160;rate were recorded throughout the whole procedure. Systemic hemodynamic parameters consist of carotid artery pressure (CAP) and central venous pressure (CVP). Hepatic perfusion parameters include portal vein pressure (PVP), portal flow rate as well as the flow rate of the common hepatic artery (table 1). Instrumentation and data acquisition to record the normal values was completed within 1.5 h. Systemic and hepatic hemodynamic parameters remained within normal ranges during this procedure.
This procedure is challenging but feasible. We have already applied this procedure to assess hepatic hemodynamics in normal mice as well as during 70% partial hepatectomy and in liver lobe clamping experiments. Mean PVP after resection (n= 20), was 11.41&#177;2.94 cmH2O which was significantly higher (P&#60;0.05) than before resection (6.87&#177;2.39 cmH2O). The results of liver lobe clamping experiment indicated that this monitoring procedure is sensitive and suitable for detecting small changes in portal pressure and portal flow rate. In conclusion, this procedure is reliable in the hands of an experienced micro-surgeon but should be limited to experiments where mice are absolutely needed.

This film demonstrates how to acquire systemic and hepatic hemodynamics in mice. The whole monitoring includes acquisition of vital parameters, systemic blood pressure, central venous pressure, common hepatic artery flow rate, and portal vein pressure as well as the portal flow rate in mice.

The use of mouse models in experimental research is of enormous importance for the study of hepatic physiology and pathophysiological disturbances. ...

Murine Echocardiography of Left Atrium, Aorta, and Pulmonary Artery

The present methodology teaches the investigator how to measure and use the LAV as a surrogate of chronic elevations in Left Ventricular diastolic pressure through echocardiography, as well as to obtain measurements of the Aorta and PA diameter in mice.
Mice older than 10 d of age can be analyzed using the present technique. The technique is composed of 3 main steps: set-up, image acquisition, and image analysis. The set-up step consists of getting the mouse anesthetized with 1% isoflurane, shaving it, and taping it in a supine position to a heated EKG board where the image acquisition will take place. The image acquisition step consists of learning to identify the cardiac structures and obtaining all the required images with its correspondent probe and axis in order to be able to calculate volumes and diameters. The image analysis step consists of measuring the previously acquired images with the aid of computer software.
Advantages of the proposed technique include a fast (15 min) procedure that would allow the researcher to evaluate interventions in a non-invasive, non-terminal approach and therefore follow the same mouse over time; each mouse can be used as its own control. This fact plus having the same operator perform all the acquisition and analysis for the entire experiment minimizes the limitation of operator-dependency. The present methodology is useful for mouse researchers in cardiovascular and pulmonary medicine.

The following protocol describes the methodology for the acquisition and analysis of echocardiographic images used to obtain the Left Atrial Volume (LAV), Aorta (Ao) diameter, and Pulmonary Artery (PA) diameter in mice. This technique is a non-invasive, non-terminal procedure that allows assessment of the cardiopulmonary function.

The present methodology teaches the investigator how to measure and use the LAV as a surrogate of chronic elevations in Left Ventricular diastolic ...

Assessment of Plasma Coagulation on Liver Tissue in a Large Animal Model In Vivo

Plasma coagulation as a form of electrocautery is used in liver surgery for decades to seal the large liver cut surface after major hepatectomy to prevent hemorrhages at a later stage. The exact effects of plasma coagulation on liver tissue are only poorly examined. In our porcine model, the coagulation effects can be examined close to the clinical application. A combined laser Doppler flowmeter and spectrophotometer documents microcirculation changes during coagulation at 8 mm tissue depth noninvasively, providing quantifiable information about hemostasis beyond the subjective clinical impression. The temperature at coagulation site is assessed with an infrared thermometer prior and post coagulation and with a thermographic camera during coagulation, a measurement of the gas beam temperature is not possible due to the upper threshold of the devices. The depth of coagulation is measured microscopically on hematoxylin/eosin stained sections after calibration with an object micrometer and gives an exact information about the power setting-coagulation depth-relation. The sealing effect is examined on the bile ducts as it is not possible for a plasma coagulator to seal larger blood vessels. Burst pressure experiments are carried out on explanted organs to rule out blood pressure related effects.

Assessment of Plasma Coagulation on Liver Tissue in a Large Animal Model In Vivo

Here we present a protocol to experimentally assess plasma coagulation in liver tissue in vivo. In a porcine model, microcirculation is examined by laser Doppler, coagulation depth is measured histologically, temperature at coagulation site by infrared thermometer and thermographic camera, and duct sealing effect is documented by burst pressure experiments.

Plasma coagulation as a form of electrocautery is used in liver surgery for decades to seal the large liver cut surface after major hepatectomy to prevent ...

Ferric Chloride-induced Canine Carotid Artery Thrombosis: A Large Animal Model of Vascular Injury

Occlusive arterial thrombosis leading to cerebral ischemic stroke and myocardial infarction contributes to ~13 million deaths every year globally. Here, we have translated a vascular injury model from a small animal into a large animal (canine), with slight modifications that can be used for pre-clinical screening of prophylactic and thrombolytic agents. In addition to the surgical methods, the modified protocol describes the step-by-step methods to assess carotid artery canalization by angiography, detailed instructions to process both the brain and carotid artery for histological analysis to verify carotid canalization and cerebral hemorrhage, and specific parameters to complete an assessment of downstream thromboembolic events by utilizing magnetic resonance imaging (MRI). In addition, specific procedural changes from the previously well-established small animal model necessary to translate into a large animal (canine) vascular injury are discussed.

Here, we present the modifications necessary to a well characterized and commonly used small animal ferric chloride-induced (FeCl3) carotid artery injury model for use in a large animal vascular injury model. The resulting model can be utilized for pre-clinical trial assessment of both prophylactic and thrombolytic pharmacological and mechanical interventions.

Occlusive arterial thrombosis leading to cerebral ischemic stroke and myocardial infarction contributes to ~13 million deaths every year globally. Here, ...

Induction and Phenotyping of Acute Right Heart Failure in a Large Animal Model of Chronic Thromboembolic Pulmonary Hypertension

The development of acute right heart failure (ARHF) in the context of chronic pulmonary hypertension (PH) is associated with poor short-term outcomes. The morphological and functional phenotyping of the right ventricle is of particular importance in the context of hemodynamic compromise in patients with ARHF. Here, we describe a method to induce ARHF in a previously described large animal model of chronic PH, and to phenotype, dynamically, right ventricular function using the gold standard method (i.e., pressure-volume PV loops) and with a non-invasive clinically available method (i.e., echocardiography). Chronic PH is first induced in pigs by left pulmonary artery ligation and right lower lobe embolism with biological glue once a week for 5 weeks. After 16 weeks, ARHF is induced by successive volume loading using saline followed by iterative pulmonary embolism until the ratio of the systolic pulmonary pressure over systemic pressure reaches 0.9 or until the systolic systemic pressure decreases below 90 mmHg. Hemodynamics are restored with dobutamine infusion (from 2.5 &#181;g/kg/min to 7.5 &#181;g/kg/min). PV-loops and echocardiography are performed during each condition. Each condition requires around 40 minutes for induction, hemodynamic stabilization and data acquisition. Out of 9 animals, 2 died immediately after pulmonary embolism and 7 completed the protocol, which illustrates the learning curve of the model. The model induced a 3-fold increase in mean pulmonary artery pressure. The PV-loop analysis showed that ventriculo-arterial coupling was preserved after volume loading, decreased after acute pulmonary embolism and was restored with dobutamine. Echocardiographic acquisitions allowed to quantify right ventricular parameters of morphology and function with good quality. We identified right ventricular ischemic lesions in the model. The model can be used to compare different treatments or to validate non-invasive parameters of right ventricular morphology and function in the context of ARHF.

We present a protocol to induce and phenotype an acute right heart failure in a large animal model with chronic pulmonary hypertension. This model can be used to test therapeutic interventions, to develop right heart metrics&#160;or to improve the understanding of acute right heart failure pathophysiology.

The development of acute right heart failure (ARHF) in the context of chronic pulmonary hypertension (PH) is associated with poor short-term outcomes. The ...

Isometric Contractility Measurement of the Mouse Mesenteric Artery Using Wire Myography

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and drugs. It is widely used in many studies on the physiological functions of vascular smooth muscle, the pathogenesis of vascular diseases such as hypertension, and the development of smooth muscle relaxant drugs. The mouse is a widely used model animal with a large pool of disease models and genetically modified strains. We introduced this method to measure the isometric contraction of mouse mesenteric artery in detail. A 1.4-mm segment of mouse mesenteric resistance artery was isolated and mounted on a myograph chamber by passing two steel wires through its lumen. After equilibration and normalization steps, the vessel segment was potentiated by a high-K+ solution twice prior to the contraction assay. As an example of the application of this method in drug development, we measured the relaxant effect of a novel natural substance, neoliensinine, isolated from a Chinese herb, embryos of the lotus seed (Nelumbo nucifera Gaertn.) on mouse mesenteric arteries. The vessel segments mounted on the myograph chamber were stimulated with a high-K+ solution. When the force tension reached a stable sustained phase, cumulative doses of neoliensinine were added to the chamber. We found that neoliensinine had a dose-dependent relaxant effect on smooth muscle contraction, thus suggesting that it bears potential activity against hypertension. In addition, as the vessel segment can survive at least 4 hours after mounting and maintain contractility induced by the high-K+ solution for many times, we suggest that the wire myograph system may be used for the time-consuming process of drug screening.

The wire myograph technique is used to investigate vascular smooth muscle functions and screen new drugs. We report a detailed protocol for measuring the isometric contractility of the mouse mesenteric artery and for screening new relaxants of vascular smooth muscle.

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and ...

Invasive Hemodynamic Assessment for the Right Ventricular System and Hypoxia-Induced Pulmonary Arterial Hypertension in Mice

Pulmonary arterial hypertension (PAH) is a chronic and severe cardiopulmonary disorder. Mice are a popular animal model used to mimic this disease. However, the evaluation of right ventricular pressure (RVP) and pulmonary artery pressure (PAP) remains technically challenging in mice. RVP and PAP are more difficult to measure than left ventricular pressure because of the anatomical differences between the left and right heart systems. In this paper, we describe a stable right heart hemodynamic measurement method and its validation using healthy and PAH mice. This method is based on open-chest surgery and mechanical ventilation support. It is a complicated procedure compared to closed chest procedures. While a well-trained surgeon is required for this surgery, the advantage of this procedure is that it can generate both RVP and PAP parameters at the same time, so it is a preferable procedure for the evaluation of PAH models.

Here, we present a protocol to perform an invasive hemodynamic assessment of the right ventricle and pulmonary artery in mice using an open-chest surgery approach.

Pulmonary arterial hypertension (PAH) is a chronic and severe cardiopulmonary disorder. Mice are a popular animal model used to mimic this disease. ...

A Pre-clinical Rat Model for the Study of Ischemia-reperfusion Injury in Reconstructive Microsurgery

Ischemia-reperfusion injury is the main cause of flap failure in reconstructive microsurgery. The rat is the preferred preclinical animal model in many areas of biomedical research due to its cost-effectiveness and its translation to humans. This protocol describes a method to create a preclinical free skin flap model in rats with ischemia-reperfusion injury. The described 3 cm x 6 cm rat free skin flap model is easily obtained after the placement of several vascular ligatures and the section of the vascular pedicle. Then, 8 h after the ischemic insult and completion of the microsurgical anastomosis, the free skin flap develops the tissue damage. These ischemia-reperfusion injury-related damages can be studied in this model, making it a suitable model for evaluating therapeutic agents to address this pathophysiological process. Furthermore, two main monitoring techniques are described in the protocol for the assessment of this animal model: transit-time ultrasound technology and laser speckle contrast analysis.

Here, we describe a pre-clinical animal model for studying the pathophysiology of ischemia-reperfusion injury in reconstructive microsurgery. This free skin flap model based on the superficial caudal epigastric vessels in the rat may also allow for the evaluation of different therapies and compounds to counteract ischemia-reperfusion injury-related damage.

Ischemia-reperfusion injury is the main cause of flap failure in reconstructive microsurgery. The rat is the preferred preclinical animal model in many ...