The authors want to thank Dagmar Dirvonskis for her excellent technical support.

The experiments in this protocol were approved by the State and Institutional Animal Care Committee (Landesuntersuchungsamt Rheinland-Pfalz, Koblenz, Germany, TVA G21-1-080). The experiments were conducted in accordance with the ARRIVE guidelines. For this study, six healthy male German Landrace pigs aged 2-3 months and weighing 30-35 kg were used. The experimental timeline is summarized in Figure 1. The details related to all materials and instruments used in this protocol are listed in the...

Dosage-Dependent Induction of Endotoxemia in Pigs via LPS Infusion

<ol>
	<li>Vincent, J. -. L., Jones, G., David, S., Olariu, E., Cadwell, K. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Frequency+and+mortality+of+septic+shock+in+Europe+and+North+America:+a+systematic+review+and+meta-analysis.">Frequency and mortality of septic shock in Europe and North America: a systematic review and meta-analysis.</a> Critical Care. 23 (1), 196 (2019).</li><li>Reinhart, 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=Recognizing+sepsis+as+a+Global+Health+Priority+-+A+WHO+Resolution.">Recognizing sepsis as a Global Health Priority - A WHO Resolution.</a> New England Journal of Medicine. 377 (5), 414-417 (2017).</li><li>Cecconi, M., Evans, L., Levy, M., Rhodes, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sepsis+and+septic+shock.">Sepsis and septic shock.</a> The Lancet. 392 (10141), 75-87 (2018).</li><li>Font, M. D., Thyagarajan, B., Khanna, A. 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S., Lowry, S. F., Calvano, S. E., Remick, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Inflammation+and+the+host+response+to+injury+investigators+acute+inflammatory+response+to+endotoxin+in+mice+and+humans.">Inflammation and the host response to injury investigators acute inflammatory response to endotoxin in mice and humans.</a> Clinical and Diagnostic Laboratory Immunology. 12 (1), 60-67 (2005).</li><li>Dickson, K., Lehmann, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Inflammatory+response+to+different+toxins+in+experimental+sepsis+models.">Inflammatory response to different toxins in experimental sepsis models.</a> International Journal of Molecular Sciences. 20 (18), 4341 (2019).</li><li>Bassols, A., Costa, C., Eckersall, P. D., Osada, J., Sabri&#224;, J., Tibau, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+pig+as+an+animal+model+for+human+pathologies:+A+proteomics+perspective.">The pig as an animal model for human pathologies: A proteomics perspective.</a> PROTEOMICS - Clinical Applications. 8 (9-10), 715-731 (2014).</li><li>Meurens, F., Summerfield, A., Nauwynck, H., Saif, L., Gerdts, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+pig:+a+model+for+human+infectious+diseases.">The pig: a model for human infectious diseases.</a> Trends in Microbiology. 20 (1), 50-57 (2012).</li><li>Ali, J., Cody, J., Maldonado, Y., Ramakrishna, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Near-infrared+spectroscopy+(NIRS)+for+cerebral+and+tissue+oximetry:+analysis+of+evolving+applications.">Near-infrared spectroscopy (NIRS) for cerebral and tissue oximetry: analysis of evolving applications.</a> Journal of Cardiothoracic and Vascular Anesthesia. 36, 2758-2766 (2022).</li><li>. Getinge Deutschland GmbH PiCCO Technologie Erweitertes h&#228;modynamisches Monitoring auf h&#246;chstem Niveau Available from: <a target="_blank" href="https://www.getinge.com/dam/hospital/documents/german/picco_haemodynamisches_monitoring_broschuere-de-non_us.pdf">https://www.getinge.com/dam/hospital/documents/german/picco_haemodynamisches_monitoring_broschuere-de-non_us.pdf</a> (2023)</li><li>Breslow, M. J., Miller, C. F., Parker, S. D., Walman, A. T., Traystman, R. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+vasopressors+on+organ+blood+flow+during+endotoxin+shock+in+pigs.">Effect of vasopressors on organ blood flow during endotoxin shock in pigs.</a> American Journal of Physiology-Heart and Circulatory Physiology. 252 (2), H291-H300 (1987).</li><li>Fink, M. P., 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=Systemic+and+mesenteric+O2+metabolism+in+endotoxic+pigs:+effect+of+ibuprofen+and+meclofenamate.">Systemic and mesenteric O2 metabolism in endotoxic pigs: effect of ibuprofen and meclofenamate.</a> Journal of Applied Physiology. 67 (5), 1950-1957 (1989).</li><li>Lado-Abeal, J., 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=Lipopolysaccharide+(LPS)-induced+septic+shock+causes+profound+changes+in+myocardial+energy+metabolites+in+pigs.">Lipopolysaccharide (LPS)-induced septic shock causes profound changes in myocardial energy metabolites in pigs.</a> Metabolomics. 14 (10), 131 (2018).</li><li>Park, 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=Characterization+of+fecal+peritonitis-induced+sepsis+in+a+porcine+model.">Characterization of fecal peritonitis-induced sepsis in a porcine model.</a> The Journal of Surgical Research. 244, 492-501 (2019).</li><li>Jarkovska, D., 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=Heart+rate+variability+in+porcine+progressive+peritonitis-induced+sepsis.">Heart rate variability in porcine progressive peritonitis-induced sepsis.</a> Frontiers in Physiology. 6, 412 (2015).</li><li>Kohoutova, 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=Vagus+nerve+stimulation+attenuates+multiple+organ+dysfunction+in+resuscitated+porcine+progressive+sepsis.">Vagus nerve stimulation attenuates multiple organ dysfunction in resuscitated porcine progressive sepsis.</a> Critical Care Medicine. 47 (6), e461-e469 (2019).</li><li>Vintrych, P., 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=Modeling+sepsis,+with+a+special+focus+on+large+animal+models+of+porcine+peritonitis+and+bacteremia.">Modeling sepsis, with a special focus on large animal models of porcine peritonitis and bacteremia.</a> Frontiers in Physiology. 13, 1094199 (2022).</li><li>Stengl, 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=Reduced+L-type+calcium+current+in+ventricular+myocytes+from+pigs+with+hyperdynamic+septic+shock.">Reduced L-type calcium current in ventricular myocytes from pigs with hyperdynamic septic shock.</a> Critical Care Medicine. 38 (2), 579-587 (2010).</li></ol>

We present a protocol for inducing experimental endotoxemia in pigs through LPS infusion, aiming to reliably induce changes commonly observed in sepsis and septic shock. Several critical steps need to be considered in this protocol. Adequate sedation of pigs prior to transport is crucial to prevent stress-induced elevation of catecholamine levels, which could potentially compromise the results. Intubation of pigs may pose challenges compared to humans due to the anatomical features of their elongated snouts. To address t...

Sepsis and septic shock rank among the leading causes of mortality in patients receiving intensive care treatment1,2,3. Sepsis arises when an infection triggers a dysregulated immune response resulting in multiorgan failure. It is characterized by life-threatening symptoms, including hemodynamic instability, respiratory distress, hepatic and renal failure, as well as cognitive impairment4,5. Septic shock represents a subset of sepsis with particularly severe symptoms that significantly increase mortality. These symptoms include persistent hypotension requiring vasopressor therapy and a serum lactate level exceeding 2 mmol&#8729;L-1&#160;4,5. Mortality rates in patients with septic shock have been estimated as high as 40%, even with hospital treatment1,3,5.
Gram-negative bacteria, such as Pseudomonas and Escherichia coli, often cause infections triggering this dysregulated immune response4. The underlying pathophysiological mechanisms are complex and not yet fully understood. One well-described aspect involves the activation of Toll-like receptors on immune cells by pathogen-associated molecular patterns (PAMPs), leading to the release of cytokines such as tumor necrosis factor-alpha (TNF&#945;) or Interleukin 1 (IL 1)4. One of these PAMPs is lipopolysaccharide (LPS), which constitutes a component of the cell membrane in gram-negative bacteria6. LPS has been employed in animal models to induce endotoxemia and endotoxemic shock7,8.
Animal models provide a controlled and standardized setting to develop and investigate novel treatment strategies. Due to its similar anatomy, immunological physiology, and comparable hemodynamic parameters, the pig model is particularly well-suited for studying the effects of endotoxemic shock9,10. Furthermore, standard medical equipment commonly used in human patients can be readily applied in pigs due to the similar size of their airways and blood vessels, facilitating instrumentation and hemodynamic monitoring.
With this protocol, we provide an experimental model for endotoxemic shock in pigs by intravenously infusing LPS derived from E. coli. To monitor the effects, we measured hemodynamic and pulmonary parameters, including arterial blood pressure, heart rate, peripheral oxygen saturation, pulmonary arterial pressure, and airway pressure. To evaluate the influence of endotoxemia on cerebral oxygen supply, we used near-infrared spectrometry (NIRS). With this method, the cerebral oxygen saturation can be evaluated via an adhesive electrode applied to the forehead11.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Atracurium Hikma 50 mg/5mL</td>
			<td>Hikma Pharma GmbH, Martinsried</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Azaperone (Stresnil) 40 mg/mL</td>
			<td>Lilly Deutschland GmbH, Bad Homburg, Germany</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>BD Discardit II Spritze 2, 5, 10, 20 mL</td>
			<td>Becton Dickinson S.A. Carretera, Mequinenza Fraga, Spain</td>
			<td>syringe</td>
			<td></td>
		</tr>
		<tr>
			<td>BD Luer Connecta&#160;</td>
			<td>Becton Dickinson Infusion Therapy, AB Helsingborg, Schweden</td>
			<td>3-way-stopcock</td>
			<td></td>
		</tr>
		<tr>
			<td>Curafix i.v. classics</td>
			<td>Lohmann &#38; Rauscher International GmbH &#38; Co. KG, Rengsdorf, Germany</td>
			<td>Cannula retention dressing</td>
			<td></td>
		</tr>
		<tr>
			<td>Datex Ohmeda S5</td>
			<td>GE Healthcare Finland Oy, Helsinki, Finland</td>
			<td>hemodynamic monitor</td>
			<td></td>
		</tr>
		<tr>
			<td>Engstr&#246;m Carestation</td>
			<td>GE Heathcare, Madison USA</td>
			<td>ventilator</td>
			<td></td>
		</tr>
		<tr>
			<td>Fentanyl-Janssen 0.05 mg/mL</td>
			<td>Janssen-Cilag GmbH, Neuss</td>
			<td>fentanyl</td>
			<td></td>
		</tr>
		<tr>
			<td>F&#252;hrungsstab, Durchmesser 4.3</td>
			<td>R&#252;sch</td>
			<td>endotracheal tube introducer</td>
			<td></td>
		</tr>
		<tr>
			<td>Incetomat-line 150 cm</td>
			<td>Fresenius, Kabi Deutschland, GmbH</td>
			<td>perfusor line</td>
			<td></td>
		</tr>
		<tr>
			<td>Intrafix Primeline</td>
			<td>B. Braun Melsungen AG, Melsungen, Germany</td>
			<td>Infusion line</td>
			<td></td>
		</tr>
		<tr>
			<td>Introducer sheath 5 Fr.</td>
			<td>Terumo Healthcare</td>
			<td>arterial introducer&#160;</td>
			<td></td>
		</tr>
		<tr>
			<td>INVOS</td>
			<td>Medtronic, Dublin, Ireland</td>
			<td>near infrared spectrometry</td>
			<td></td>
		</tr>
		<tr>
			<td>JOZA Einmal Nitril Untersuchungshandschuhe&#160;</td>
			<td>JOZA, M&#252;nchen, Germany</td>
			<td>disposable gloves</td>
			<td></td>
		</tr>
		<tr>
			<td>Laryngoscope, 45.48.50, KL 2000</td>
			<td>Medicon</td>
			<td>Laryngoscope handle</td>
			<td></td>
		</tr>
		<tr>
			<td>Littmann Classic III Stethoscope</td>
			<td>3M Deutschland GmbH, Neuss, Germany</td>
			<td>stethoscope</td>
			<td></td>
		</tr>
		<tr>
			<td>LPS (E. coli; Serotype O111:B4)</td>
			<td>Sigma-Aldrich, Switzerland</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>MAC Two-Lumen Central venous access set</td>
			<td>Arrow international inc. Reading, PA, USA</td>
			<td>venous introducer</td>
			<td></td>
		</tr>
		<tr>
			<td>Maimed Vlieskompresse</td>
			<td>Maimed GmbH, Neuenkirchen, Germany</td>
			<td>Fleece compress to fix the tongue</td>
			<td></td>
		</tr>
		<tr>
			<td>Masimo LNCS Adtx SpO2 sensor</td>
			<td>Masimo Corporation Irvine, Ca 92618 USA</td>
			<td>saturation clip for the tail</td>
			<td></td>
		</tr>
		<tr>
			<td>Masimo LNCS TC-I SpO2 ear clip sensor</td>
			<td>Masimo Corporation Irvine, Ca 92618 USA</td>
			<td>Saturation clip for the ear</td>
			<td></td>
		</tr>
		<tr>
			<td>Masimo Radical 7</td>
			<td>Masimo Corporation Irvine, Ca 92618 USA</td>
			<td>periphereal oxygen saturation&#160;&#160;&#160;</td>
			<td></td>
		</tr>
		<tr>
			<td>Midazolam 15 mg/3 mL</td>
			<td>B.Braun Melsungen AG, Germany</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Midmark Canine Mask Small Plastic with Diaphragm FRSCM-0005</td>
			<td>Midmark Corp., Dayton, Ohio, USA</td>
			<td>dog ventilation mask</td>
			<td></td>
		</tr>
		<tr>
			<td>Monocryl surgical suture</td>
			<td>Johnson &#38; Johnson, Belgium</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td></td>
			<td>B.Braun Melsungen AG, Germany</td>
			<td>saline solution</td>
			<td></td>
		</tr>
		<tr>
			<td>NaCl 0.9 %</td>
			<td>Sanofi- Aventis, Seutschland GmbH</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Octeniderm farblos</td>
			<td>Sch&#252;lke &#38; Mayr GmbH, Nordenstedt, Germany</td>
			<td>Alcoholic disinfectant</td>
			<td></td>
		</tr>
		<tr>
			<td>Original Perfusor syringe 50 mL</td>
			<td>B.Braun Melsungen AG, Germany</td>
			<td>perfusor syringe</td>
			<td></td>
		</tr>
		<tr>
			<td>PA-Katheter Swan Ganz 7.5 Fr 110 cm</td>
			<td>Edwards Lifesciences LLC, Irvine CA, USA</td>
			<td>Swan-Ganz catheter</td>
			<td></td>
		</tr>
		<tr>
			<td>Perfusor FM Braun</td>
			<td>B.Braun Melsungen AG, Germany</td>
			<td>syringe pump</td>
			<td></td>
		</tr>
		<tr>
			<td>PiCCO catheter</td>
			<td>PULSION Medical Systems SE, Feldkirchen, DE</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Potassium chloride 1 M</td>
			<td>Fresenius, Kabi Germany GmbH</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Propofol 2% 20 mg/mL (50 mL flasks)</td>
			<td>Fresenius, Kabi Deutschland, GmbH</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Pulse-contour continous cardiac output System PiCCO2</td>
			<td>PULSION Medical Systems SE, Feldkirchen, DE</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>R&#252;schelit Super Safety Clear &#62;ID 6/6.5 /7.0 mm</td>
			<td>Teleflex Medical Sdn. Bhd, Malaysia</td>
			<td>endotracheal tube</td>
			<td></td>
		</tr>
		<tr>
			<td>Sonosite Micromaxx Ultrasoundsystem</td>
			<td>Sonosite Bothell, WA, USA&#160;</td>
			<td>ultrasound&#160;</td>
			<td></td>
		</tr>
		<tr>
			<td>Stainless Macintosh Gr&#246;&#223;e 4</td>
			<td>Welch Allyn69604</td>
			<td>blade for laryngoscope</td>
			<td></td>
		</tr>
		<tr>
			<td>Sterofundin</td>
			<td>B.Braun Melsungen AG, Melsungen, Germany</td>
			<td>Balanced electrolyte solution</td>
			<td></td>
		</tr>
		<tr>
			<td>Vasco OP sensitive&#160;</td>
			<td>B.Braun Melsungen AG, Germany</td>
			<td>sterile gloves</td>
			<td></td>
		</tr>
		<tr>
			<td>Vasofix Safety 22 G-16 G</td>
			<td>B.Braun Melsungen AG, Germany</td>
			<td>venous catheter</td>
			<td></td>
		</tr>
		<tr>
			<td>VBM Cuff Manometer</td>
			<td>VBM Medizintechnik GmbH, Sulz a.N., Germany&#160;</td>
			<td>cuff pressure gauge</td>
			<td></td>
		</tr>
	</tbody>
</table>

lipopolysaccharide infusion as a porcine endotoxemic shock model

Sepsis and septic shock are frequently encountered in patients treated in intensive care units (ICUs) and are among the leading causes of death in these patients. It is caused by a dysregulated immune response to an infection. Even with optimized treatment, mortality rates remain high, which makes further insights into the pathophysiology and new treatment options necessary. Lipopolysaccharide (LPS) is a component of the cell membrane of gram-negative bacteria, which are often responsible for infections causing sepsis and septic shock. 
The severity and high mortality of sepsis and septic shock make standardized experimental studies in humans impossible. Thus, an animal model is needed for further studies. The pig is especially well suited for this purpose as it closely resembles humans in anatomy, physiology, and size. 
This protocol provides an experimental model for endotoxemic shock in pigs by LPS infusion. We were able to reliably induce changes frequently observed in septic shock patients, including hemodynamic instability, respiratory failure, and acidosis. This will allow researchers to gain valuable insight into this highly relevant condition and evaluate new therapeutic approaches in an experimental setting.

Author Spotlight: Induction of Experimental Endotoxemic Shock in Pigs for Studying Hemodynamic and Respiratory Failure

Lipopolysaccharide Infusion as a Porcine Endotoxemic Shock Model

With this protocol, we induce experimental endotoxemic shock in pigs. We were able to reliably cause symptoms of endotoxemia, such as hemodynamic instability and respiratory failure. Lipopolysaccharide is a toxic substance.When working with lipopolysaccharide, it is essential always to wear gloves, protective goggles, a mask, and a lab coat. This protocol provides an easy and reliable way to induce endotoxemia. Because of their similar anatomy and physiology, pigs are particularly well-suited for this kind of research as standard medical equipment may be used.Using this protocol, we will be able to study the effects of different ventilation strategies in respiratory failure and lung damage during endotoxemic shock.

For this study, six healthy male pigs aged 2-3 months and weighing 30-35 kg were anesthetized and received an infusion of lipopolysaccharide (LPS) to induce endotoxemia. To determine the appropriate dosage of LPS required to consistently induce symptoms of shock, the pigs were administered various induction doses of LPS ranging from 100 &#181;g kg-1 to 200 &#181;g kg-1 over a 30-min period, followed by a maintenance dose of 1/10 of the initial dose per hour for the remainder of the experiment. All a...

Watch this Scientific Journal Video about Lipopolysaccharide Infusion as a Porcine Endotoxemic Shock Model at JoVE.com

We provide a protocol for an experimental endotoxemic shock model in pigs by infusion of lipopolysaccharide.

Sepsis and septic shock are frequently encountered in patients treated in intensive care units (ICUs) and are among the leading causes of death in these ...

lipopolysaccharide-infusion-as-a-porcine-endotoxemic-shock-model

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503 vues.  University Medical Center of the Johannes Gutenberg-University. Avec ce protocole, nous induisons un choc endotoxémique expérimental chez les porcs. Nous avons pu provoquer de manière fiable des symptômes d’endotoxémie, tels que l’instabilité hémodynamique et l’insuffisance respiratoire. Le lipopolysaccharide est une substance toxique.Lorsque vous travaillez avec des lipopolysaccharides, il est essentiel de toujours porter des gants, des lunettes de protection, un masque et une blouse de laboratoire. Ce protocole offre un moyen simple ...

Vidéo: Pleins feux sur l’auteur : Induction d’un choc endotoxémique expérimental chez le porc pour l’étude de l’insuffisance hémodynamique et respiratoire