Infusing oleic acid continuously into the pulmonary artery of an anesthetized adult pig induces acute respiratory failure, enabling controlled experimentation during acute respiratory decompensation.
This protocol outlines an acute respiratory distress model utilizing centrally administered oleic acid infusion in Yorkshire pigs. Prior to experimentation, each pig underwent general anesthesia, endotracheal intubation, and mechanical ventilation, and was equipped with bilateral jugular vein central vascular access catheters. Oleic acid was administered through a dedicated pulmonary artery catheter at a rate of 0.2 mL/kg/h. The infusion lasted for 60-120 min, inducing respiratory distress. Throughout the experiment, various parameters including heart rate, respiratory rate, arterial blood pressure, central venous pressure, pulmonary artery pressure, pulmonary capillary wedge pressure, end-tidal carbon dioxide, peak airway pressures, and plateau pressures were monitored. Around the 60 min mark, decreases in partial arterial oxygen pressure (PaO2) and fraction of oxygen-saturated hemoglobin (SpO2) were observed. Periodic hemodynamic instability, accompanied by acute increases in pulmonary artery pressures, occurred during the infusion. Post-infusion, histological analysis of the lung parenchyma revealed changes indicative of parenchymal damage and acute disease processes, confirming the effectiveness of the model in simulating acute respiratory decompensation.
The utilization of porcine models in translational research holds significant importance in advancing our understanding of human medicine1. Porcine models, due to their physiological and anatomical similarities to humans, provide a valuable platform for studying complex diseases and therapeutic interventions1. In the context of acute respiratory failure, porcine models offer a unique opportunity to investigate the pathophysiological mechanisms, evaluate treatment strategies, and assess potential interventions1,2,3. The ability to replicate key aspects of human respiratory physiology and responses to various stimuli in porcine models allows for a comprehensive evaluation of therapeutic modalities before progressing to human trials1,2,3. This research paradigm enables researchers to bridge the gap between preclinical investigations and clinical application, facilitating the development of novel therapies and improving patient outcomes1. Therefore, the establishment of an efficient, effective, and reproducible acute respiratory failure porcine model serves as a crucial tool in advancing the knowledge of respiratory diseases and guiding the development of effective interventions in human medicine1.
Respiratory distress, a critical medical condition, has witnessed limited advancements in its diagnosis and management over the past three decades4. The currently employed evaluation and triage metrics, which include subjective symptoms, physical examination findings, SpO2, and respiratory rate, often exhibit limitations in detecting acute pulmonary conditions at an early stage5,6,7. This inadequacy not only hampers efficient triage and resource allocation but also fails to provide effective, quantitative monitoring of disease progression and treatment response in patients with chronic pulmonary diseases. The emerging landscape of chronic respiratory conditions, such as long COVID, along with the burden of acute respiratory insufficiencies on hospital resources, underscores the urgent need to expand translational research and foster innovation in respiratory disease management.
The direct infusion of oleic acid into a pig's bloodstream has been recognized as a robust method to induce acute respiratory distress8. Oleic acid, a monounsaturated fatty acid, has demonstrated the ability to trigger significant pulmonary injury and compromise respiratory function when introduced into the pulmonary circulation8. Upon infusion, oleic acid provokes vasoconstriction, resulting in increased pulmonary arterial pressure and resistance, leading to impaired gas exchange and oxygenation9. Furthermore, oleic acid promotes the activation of inflammatory pathways, including the release of pro-inflammatory mediators and recruitment of immune cells, which contribute to the development of lung injury and respiratory distress10. All of this results in severe hypoxemia, increases in pulmonary arterial pressures, and the accumulation of extravascular lung water11. Histological evaluation of the lung parenchyma has demonstrated injury that is indistinguishable from human acute respiratory distress9.
This article details a method involving the direct administration of oleic acid into the pulmonary artery to induce acute respiratory distress, avoiding untreatable, severe hemodynamic compromise. The described method is anticipated to be a valuable tool for future researchers exploring the underlying pathophysiological mechanisms of acute respiratory failure and assessing potential therapeutic interventions and innovations.
The protocol received approval from the Vanderbilt University Institutional Animal Care and Use Committee (protocol M1800176-00) and strictly adhered to the National Institute of Health Guidelines for the Care and Use of Laboratory Animals. Male and female Yorkshire pigs, weighing approximately 40-45 kg, were utilized in this experiment. The animals were obtained from a commercial source (see Table of Materials). The current practice does not involve screening for any pre-existing medical conditions in the acquired swine. While it is acknowledged that this practice could potentially interfere with or mask intended results, it is considered unlikely according to the vendor, and this limitation is accepted.
1. Animal preparation
2. Oleic acid infusion
3. Venous waveform analysis and ventilator management procedure
Early single pig, pilot data demonstrates an increase in RIVA-RI prior to changes in other respiratory monitoring measures (RR and SpO2), in line with changes in PaO2 (Figure 3). The drop in PaO2 is the "positive" result this model intends to achieve. Preliminary data also shows that RIVA-RI increases and the PaO2 decreases with disease progression starting at the 30-min mark (Figure 3; red arrow). PaO2 is the hemodynamic parameter of most interest in the setting of monitoring the respiratory condition secondary to the fact it reflects the balance between oxygen delivery and consumption no matter the clinical state of a patient28,29. It also drives the diagnosis of acute respiratory distress and the implementation of treatments30. Both values changed well before decreases in SpO2 in the porcine model, which was delayed to the 60-min mark31 (Figure 3). The preliminary data also supports the previously reported limitation in the singular use of SpO2 as the clinical indicator of pulmonary disease severity29,32,33,34. These data support that this described model is successful in achieving an acute respiratory failure using oleic acid delivery directly through a PAC.
In addition to successfully achieving acute respiratory failure using gas exchange definitions, histological evaluation of porcine lung tissue5,6 (formalin-fixed, paraffin-embedded and stained with hematoxylin and eosin) after successful achievement of respiratory distress demonstrated changes consistent with human acute respiratory distress (Figure 2). This supports previous literature of oleic acid use for acute respiratory distress9.
Figure 1: Timeline of a porcine acute respiratory distress study in Yorkshire pigs. RIVA-RI values, hemodynamic data (heart rate, respiratory rate, blood pressure, SpO2, pulmonary artery pressures, pulmonary capillary wedge pressure, central venous pressure, pleural pressure, plateau pressure, and peak pressure), and arterial blood gas analysis (PaO2, PaCO2, Lactate, Base Excess, and pH) were obtained every 30 min for the first 1.5 h and then every 15 min thereafter until sacrifice. The NHLBI ARDS Clinical Network Mechanical Ventilation Protocol will be initiated when the P/F ratio is <100. Ventilator adjustments will follow this protocol with goal oxygenation of PaO2 55-80 mmHg or SpO2 88%-95%. A minimum of 2 min for venous waveform capture will occur prior to any escalation of FiO2 and/or PEEP. Abbreviations: ABG = arterial blood gas; RIVA = Respiratory non-Invasive Venous waveform Analysis; min = minute; PaO2 = partial pressure of oxygen; PaCO2 = partial pressure of carbon dioxide; mL = millimeter; kg = kilogram; h = hour; FiO2 = fraction of inspired oxygen; PEEP = positive end-expiratory pressure. Please click here to view a larger version of this figure.
Figure 2: Histology of porcine lung post oleic acid infusion with PaO2/FiO2 <100. Histological changes of porcine lung tissue validates this oleic acid model with demonstration of alveolar expanded alveolar septa by myxoid-appearing fibrous tissue, pneumocyte hyperplasia, and lymphocytic infiltration (red arrows)-classical findings of acute respiratory distress. Scale bar = 500 µm; magnified part = 100 µm. Please click here to view a larger version of this figure.
Figure 3: Graphical relationship between RIVA-RI and PaO2 and SpO2 over time of oleic acid infusion in a porcine model of one pig. As acute respiratory distress is induced through the direct infusion of oleic acid into the pulmonary artery, RIVA-RI increases as PaO2 (A) and SpO2 (B) decrease. When RIVA-RI was visualized in the color spectrum (C) over the entire course of oleic acid infusion, an increase in the RIVA-RI value was observed after 40 min of infusion and increased throughout the development of respiratory distress. Red arrows indicate the point at 30 min where RIVA-RI and PaO2 started to change, but SpO2 remained the same. Abbreviations = RIVA-RI = Respiratory non-Invasive Venous waveform Analysis Respiratory Index, PaO2 = partial pressure of oxygen, SpO2 = oxygen hemoglobin saturation, min= minutes. Please click here to view a larger version of this figure.
The key element in this protocol is to closely monitor the hemodynamic condition of the pig during the administration of oleic acid to induce respiratory distress15. It is of the utmost importance for researchers to take the necessary time to appropriately position the hemodynamic monitoring devices. One specific drawback of this model is the potential hemodynamic instability that may arise as a result of inflammation and injury to the respiratory vasculature8,10,18. Although the direct administration of oleic acid into the pulmonary artery leads to less instability than its administration at the jugular/atrial junction, the possibility of such instability should still be acknowledged. Typically, around 45-60 min after the initiation of the procedure, a significant increase in MPAP occurs. Therefore, it may be necessary to provide hemodynamic support in order to achieve respiratory failure in pigs. Normally, administering 50-70 mcg of norepinephrine and/or 10-30 mcg of epinephrine for SBP below 60 mmHg and/or HR below 50 bpm will rectify hemodynamic deterioration and allow for the continuation of the experiment. Development of hemodynamic instability has been identified as the primary limitation of this model.
Another limitation of this model is the duration required for the experiment. Based on previous experience, it typically takes an average of 60 min for the early signs of respiratory distress to manifest. In most cases, the PaO2 continues to decline; however, there have been isolated instances in which some form of presumed adaptation occurs, leading to a temporary rebound in PaO2. To date, this rebound has not persisted for longer than 30 min during ongoing oleic acid infusion. It is important to appreciate that the use of oleic acid is directly toxic to the endothelial cells, and this injury is followed by an increased pulmonary microvascular permeability and intrapulmonary shunt11. For this reason, it is believed a more direct administration leads to a more stable and isolated pulmonary injury and acute respiratory failure – the goal of this protocol.
This method was conceived based on the fusion of multiple reports using oleic acid to induce porcine respiratory distress. Boker et. al. used oleic acid-induced lung injury to investigate changes in PaO2, lung compliance, and proinflammatory cytokines for comparing mechanical ventilation using ARDSnet low tidal volumes20. They used a continuous infusion of oleic acid but defined respiratory failure as a PaO2 less than or equal to 65 mmHg for two consecutive measurements, 5 min apart20. They supported their hemodynamics with a continuous dopamine infusion (5-12 μg/kg/min), titrated to control the MAP greater than 50 mmHg20. Mutch et. al.19 also investigated mechanical ventilation strategies after acute respiratory distress using an oleic acid model. They placed a 5 Fr catheter into the femoral vein but their catheter ended within the right atria and oleic acid was administered at a rate of 0.2 mL/kg/h19. Dopamine was also used for hemodynamic support with MAP goals of 60 mmHg19. Respiratory failure was defined as a PaO2 equal to or less than 60 mmHg for two consecutive measurements, 5 min apart19. Multiple differences exist between the model described herein and these two prior examples of oleic acid use. First, there was an obvious difference in the maintenance of hemodynamic control. Prophylactic vasoactive infusion was not desired, and a more reactive administration at the first sign of hemodynamic distress was chosen for this protocol to spare the pig of vasoactive medications if not required. A second difference was observed in oleic acid delivery. They chose to administer them more systemically through a 5 Fr single-lumen catheter that ended approximately 1-2 cm above the diaphragm through the femoral vein and more hemodynamic instability with systemic administration has been observed. In contrast, it is significantly less when administered directly into the pulmonary artery through a dedicated pulmonary artery catheter.
Another method of inducing acute lung injury in swine was described by Rissel et. al.11, where they performed both bronchoalveolar lavage and an injection of oleic acid. Their double-hit method clearly causes severe lung injury by mimicking the two central elements of the pathomechanism of Acute Respiratory Distress Syndrome (ARDS)11 and showed similar PaO2/FiO2 at 1 h mark11. However, their described goal is to cause severe lung injury in pigs that is suitable to study different treatment options in ARDS11. The goal of the described protocol is to create isolated acute respiratory failure; therefore, the addition of another step to ensure surfactant depletion as well as alveolar collapse has not been found to be necessary and has not been performed to date in this described model.
In light of the necessity for innovation and research in the field of respiratory diseases, the establishment of an efficient, reliable, and reproducible animal model is of paramount significance. The use of oleic acid to induce acute respiratory failure has been extensively studied8,10,11,18,19,20. The present report provides a detailed account of cannulation, hemodynamic monitoring, and oleic acid administration strategies employed to induce acute respiratory failure in adult pigs. Together with existing literature, this comprehensive model description aims to facilitate the investigation of pivotal hypotheses within the field by future scientists.
The authors have nothing to disclose.
The authors would like to thank Dr. José A. Diaz, Jamie Adcock, Mary Susan Fultz and the S.R. Light Laboratory at Vanderbilt University Medical Center for their assistance and support. This work was supported by a grant from the National Heart, Lung, and Blood Institute of the National Institutes of Health (BA; R01HL148244). The content is the sole responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
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Arterial Catheter | Merit Medical, South Jordan, UT, USA | MAK401 | MAK Mini Access Kit 4F |
Blood Pressure Amp | AD Instruments, Colorado Springs, CO, USA | FE117 | https://www.adinstruments.com/products/bp-blood-pressure-amp |
Central Venous Catheter | Arrow International, Cleveland, OH, USA | AK-09800 | 8.5 Fr. x 4" (10 cm) Arrow-Flex |
Disposable Pressure Transducers | AD Instruments, Colorado Springs, CO, USA | MLT0670 | https://www.adinstruments.com/products/disposable-bp-transducers |
Edwards Lifesciences Triple Stage Venous Cannulas | Edwards Life Sciences, Irvine, CA | TF293702 | https://www.graylinemedical.com/products/edwards-lifesciences-triple-stage-venous-cannulas-venous-dual-stage-cannula-tf293702?variant=31851942576185&gad=1& gclid=Cj0KCQiAr8eqBhD3ARIsAIe -buNdmkzavUBaIx-1be7boWn2kW hbUR6QCjaobB08uuK9qJW66JvY TM4aAufGEALw_wcB |
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LabChart 8 software | AD Instruments, Colorado Springs, CO, USA | N/A | https://www.adinstruments.com/products/labchart |
Lahey Retractor | BOSS Instruments LTD, Gordonsville, VA 22942 | 18-1210 | https://bossinstruments.com/product/7-3-4-lahey-thyroid-retractor-6mmx28mm/ |
Oleic Acid | Sigma-Aldrich, Merck, Darmstadt, Germany | O1008 | https://www.sigmaaldrich.com/US/en/product/sial/o1008?gclid=CjwKCAjwzJmlBhBBEiwAEJy Lu2047wRpXqF_Z2BegUyhgZJ _WygsWfErhgrGCIyMp8PxwNH sTZ8qARoCl1QQAvD_BwE&gcl src=aw.ds |
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Pulmonary Artery Catheter | Edwards Life Sciences, Irvine, CA | 131F7 | Swan Ganz 7F x 110cm |
Standard Endotracheal Tubes | Teleflex, Morrisville, NC 27560 | 5-10313 | https://www.teleflex.com/usa/en/product-areas/anesthesia/airway-management/endotracheal-tubes/standard-tubes/index.html |
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Xylazine HCl 100 mg/mL, Injectable Solution, 50 mL | Patterson Veterinary, Loveland, CO 80538 | 07-894-5244 | https://www.pattersonvet.com/ProductItem/078945244 |
Yorkshire Pigs | Oak Hill Genetics, Ewing, IL, USA | 138274 | Female/Male Swine- Yorkshire/Landrace 81-100lbs |
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