Endotracheal Intubation Using a Flexible Intubation Endoscope as a Standardized Model for Safe Airway Management in Swine

Podsumowanie

The use of pigs in research has increased in recent years. Nevertheless, pigs are characterized by difficult airway anatomy. By demonstrating how to perform endoscopically guided endotracheal intubation, the present protocol aims to further increase laboratory animals' safety to avoid animal suffering and unnecessary death.

Streszczenie

Endotracheal intubation is often a basic requirement for translational research in porcine models for various interventions that require a secured airway or high ventilation pressures. Endotracheal intubation is a challenging skill, requiring a minimum number of successful endotracheal intubations to achieve a high success rate under optimal conditions, which is often unachievable for non-anaesthesiology researchers. Due to the specific porcine airway anatomy, a difficult airway can usually be assumed. The impossibility of establishing a secure airway can result in injury, adverse events, or death of the laboratory animal. Using a prospective, randomized, controlled evaluation approach, it has been shown that fiberoptic-assisted endotracheal intubation takes longer but has a higher first-pass success rate than conventional intubation without causing clinically relevant drops in oxygen saturation. This model presents a standardized regimen for endoscopically guided endotracheal intubation, providing a secured airway, especially for researchers who are inexperienced in the technique of endotracheal intubation via direct laryngoscopy. This procedure is expected to minimize animal suffering and unnecessary animal losses.

Wprowadzenie

Endotracheal intubation is often a basic requirement for translational research in porcine models for various interventions that require a secured airway or high ventilation pressures (such as ventilation during cardiopulmonary resuscitation¹ or acute respiratory distress syndrome²) or require that cerebral blood flow is not compromised through internal compression by supraglottic airway devices³, which are occasionally propagated as alternatives in the context of an anticipated difficult airway in pigs^4,5.

While the lung physiology of pigs shows similar features to that of humans⁶, securing the airway is sometimes significantly more difficult⁷ due to specific differences in the porcine orotracheal anatomy. The snout of a pig has a narrow opening with a very large tongue, the larynx is extremely mobile, and the epiglottis is relatively large, with a free end that extends to the soft palate. Caudally, the larynx forms an obtuse angle with the trachea. The arytenoid cartilages are large⁸. The narrowest part of the airway is at the subglottic level⁹, comparable to the airway anatomy of children¹⁰. Since the larynx in pigs is very mobile, there is a risk that the end of the endotracheal tube will pass through the vocal cords but the larynx will only be displaced caudally by up to several centimeters, which may be mistaken for a correct intubation^8,11. Additionally, esophageal intubation is a common risk when dealing with porcine airway management¹².

The rates of difficult or impossible endotracheal intubations with a corresponding negative impact on the experiment or early mortality have not been systematically recorded, but several case reports have been published^13,14. In humans, there is the possibility of using a flexible intubation endoscope in the context of an unexpectedly difficult conventional intubation¹⁵. Various false intubations often precede this measure. These repeated intubation attempts are associated with adverse events in humans^16,17, especially airway complications¹⁸. Such events are deleterious in test animals since, in the simplest case, they represent a confounder variable in the experiment; in the worst case, they can lead to the unnecessary loss of the animal.

The present study has developed a model based on the guidelines for expected difficult airway management in humans^{15,19,20,21,22,23,24}. Previously, a similar technique has been described for learning fiberoptic intubation in human studies^25,26. The protocol presented in this report aims to provide a standardized and easy-to-adapt intubation model that also allows non-airway specialists to perform successful and safe endotracheal intubation in pigs.

Protokół

The experiments in this protocol were approved by the State and Institutional Animal Care Committee (Landesuntersuchungsamt Rheinland-Pfalz, Koblenz, Germany; approval no. G20-1-135). The experiments were conducted following the ARRIVE guidelines. Overall, 10 anesthetized male pigs (Sus scrofa domestica) with a mean weight of 30 kg ± 2 kg and 12-16 weeks in age were used for the present study.

1. Animal preparation

1. Maintain a normal environment for the animals to minimize stress. Withhold food 6 h before the scheduled experiment to lower the risk of aspiration, but allow access to water.
2. Sedate the pigs with a combined injection of midazolam (0.5 mg/kg) and azaperone (2-3 mg/kg) (see Table of Materials) in the gluteal muscle or neck with a needle (20 G) for intramuscular injection. Leave the animals undisturbed until sedation sets in (15-20 min).
   NOTE: Depending on national regulations, the administration of sedating agents may be subject to scrutiny and may or may not require the supervision of a trained veterinarian. Consult with the local authorities before planning the experiments.
3. Transport the sedated animals from the stables to the laboratory. The transport time must not exceed adequate sedation time (here, 30-60 min). Ensure sufficient heat retention so that the animal does not get hypothermic (i.e., below 38 °C), such as by covering the body with a blanket depending on the outside temperature.
4. Using a sensor (see Table of Materials) clipped to the ear or tail, monitor the peripheral oxygen saturation (SpO₂).
5. Disinfect the skin with a disinfectant (alcoholic) before inserting a peripheral vein cannula (22 G) into an ear vein. Spray the area, wipe once, then spray again, and allow the disinfectant to dry. Secure the ear cannula with a band-aid (See Table of Materials).

2. Anesthesia and mechanical ventilation

1. Administer analgesia through an intravenous injection of 4 µg/kg of fentanyl. Induce anesthesia with an intravenous injection of 3 mg/kg of propofol (see Table of Materials).
   NOTE: Due to the bolus application, the drug quickly floods into the active compartment, providing a fast onset of deep anesthesia.
2. Place the pig on a stretcher in a supine position and fix it with bandages. Apply muscle relaxant through an intravenous injection of 0.5 mg/kg of atracurium (see Table of Materials).
3. Instantaneously start noninvasive ventilation via a dog ventilation mask (see Table of Materials) or similar models. To ensure a tight fit of the mask, place the thenar eminence and the thumbs of both hands on top of the mask while performing jaw thrust with the remaining fingers.
   NOTE: Ventilation parameters: FiO₂ (inspiratory oxygen fraction) = 100%, peak inspiratory pressure = <20 cmH₂0, respiratory rate = 18-20 breaths/min, PEEP (positive end-expiratory pressure) = 5 cmH₂0.
4. Maintain anesthesia through continuous infusion of 0.1-0.2 mg/kg/h of fentanyl and 8-12 mg/kg/h of propofol. Start infusing with 5 mL/kg/h of balanced electrolyte solution (see Table of Materials) continuously. Constantly maintain an adequate depth of anesthesia.
   ​NOTE: Surrogate parameters for this are the absence of movement, the lack of own respiratory efforts after intubation, and the absence of a sudden increase in heart rate. If possible, avoid permanent muscle relaxation to enable motor reactions as a sign of insufficient depth of anesthesia.

3. Endotracheal intubation

1. Have an assistant stand on the left side of the head. Have the assistant's left hand open the mouth and pinch the tongue outward and left with a compress. Ask the assistant to press down on the right upper lip with the right index finger to provide a better mouth opening.
2. Perform direct laryngoscopy. To do this, insert the laryngoscope (see Table of Materials) into the right side of the mouth and push it forward while pushing the tongue to the left. Advance the tip of the laryngoscope until it rests in the epiglottic vallecula.
   NOTE: The epiglottis usually obscures the glottis by sticking to the soft palate.
3. Carefully push the epiglottis aside with a tube guiding wire (see Table of Materials) with a gentle scooping motion from the right piriform recess to the left along the soft palate.
4. Pass the handle of the laryngoscope to the assistant to fix it in the current position.
5. Now, take the flexible intubation endoscope onto which an endotracheal tube has already been mounted and that is connected to a video monitor. Insert the endoscope orally and advance it over the base of the tongue until the glottis is visualized.
   NOTE: To avoid fogging of the camera, the prior application of anti-fogging agents (see Table of Materials) is recommended.
6. Advance the endoscope between the vocal ligaments into the trachea. Confirm the anatomy of the trachea by visually identifying the cartilaginous rings and the pars membranacea. Advance the endoscope until it rests above the carina. Try not to touch the sensitive mucosa with the tip of the endoscope to avoid swelling and bleeding.
7. While maintaining the position of the endoscope, advance the endotracheal tube until it becomes visible in the camera image.
   NOTE: If the endotracheal tube cannot be advanced through the glottic plane, there is a possibility that it has become caught on the arytenoid cartilage. In this case, the endotracheal tube must be withdrawn 1 cm and rotated by 90° before gently advancing again. If necessary, this maneuver can be repeated. Similar calibers of flexible intubation endoscope and endotracheal tube can minimize the risk of this issue occurring. If the endotracheal tube cannot be advanced despite this maneuver, it is likely that the subglottic narrowness-the narrowest part of the porcine larynx-cannot be passed. In this case, a smaller endotracheal tube size needs to be selected. Regular commercially available endotracheal tubes in sizes 6.5 cm or 7.0 cm ID should be able to pass the glottis as long as no anatomic abnormalities are present. Endotracheal tube size requirements vary depending on the piglet size and breed.
8. Withdraw the flexible intubation endoscope while maintaining the position of the endotracheal tube.
9. Using a 10 mL syringe, inflate the cuff with 10 mL of air. Control the cuff pressure with a cuff manager (target value: 30 cmH₂O, see Table of Materials).
10. Confirm the correct placement of the endotracheal tube and adequate ventilation by periodic and regular exhalation of carbon dioxide via capnography²⁴ and double-sided ventilation via auscultation¹⁵.
11. Start mechanical ventilation after connecting the tube with a ventilator (PEEP = 5 cmH₂O, respiratory rate = variable to achieve an end-tidal CO₂ of <6 kPa, usually 30-50 min⁻¹, FiO₂ = 0.4, I:E (inspiration to expiration ratio) = 1:2, tidal volume = 6-8 mL/kg).
12. Expand the monitoring (e.g., the establishment of an intra-arterial blood pressure measurement, the installation of a central venous or pulmonary arterial catheter²⁷) or continue with the intervention.
    NOTE: Depending on the question of the further experiments, define limit values for the vital parameters and intervention options and establish the monitoring accordingly in the study protocol.

Wyniki

Endotracheal intubation was performed on 10 male pigs (age 12-16 weeks, weight 30 kg ± 3 kg) in a prospective, randomized, controlled study setting. The pigs were randomized into two groups: one was conventionally laryngoscopically intubated (CI group), and the other group was intubated assisted via a flexible intubation endoscope as described in the protocol (FIE group). The group assignment was done by pulling sealed envelopes. The investigator was assigned randomly on a daily basis.

Dyskusje

In previous studies, our research group has already described specific details regarding the translational benefits of the porcine model^2,27,32,33. Generally, reducing the stress level of the animal and unnecessary pain should be an integral part of any study protocol and is paramount for generating reliably reproducible data. Therefore, awake endoscopically guided intubation of the pig with an...

Materiały

Name	Company	Catalog Number	Comments
Ambu aScope Regular	Ambu GmbH, Medizinprodukte, Bad Nauheim, Germany		Disposable fiber optic outer diameter 5 mm
Ambu aView Monitor	Ambu GmbH, Medizinprodukte, Bad Nauheim, Germany		monitor
Atracurium Hikma 50 mg/5mL	Hikma Pharma GmbH, Martinsried		atracurium
Azaperone (Stresnil) 40mg/mL	Lilly Deutschland GmbH, Bad Homburg, Germany		azaperone
BD Discardit II Spritze 2, 5, 10, 20 mL	Becton Dickinson S.A. Carretera, Mequinenza Fraga, Spain		syringe
BD Luer Connecta	Becton Dickinson Infusion Therapy, AB Helsingborg, Schweden		3-way-stopcock
BD Microlance 3 20 G	Becton Dickinson S.A. Carretera, Mequinenza Fraga, Spain		cannula
Curafix i.v. classics	Lohmann & Rauscher International GmbH & Co. KG, Rengsdorf, Germany		Cannula retention dressing
Engström Carestation	GE Heathcare, Madison USA		ventilator
Fentanyl-Janssen 0.05 mg/mL	Janssen-Cilag GmbH, Neuss		fentanyl
Führungsstab, Durchmesser 4.3	Rüsch		endotracheal tube introducer
IBM SPSS Statistics for Windows, Version 20	IBM SPSS Statistics for Windows, Version 20.0. Armonk, NY: IBM Corp.)		Statistical software
Incetomat-line 150 cm	Fresenius, Kabi Deutschland, GmbH		perfusor line
Intrafix Primeline	B. Braun Melsungen AG, Melsungen, Germany		Infusion line
JOZA Einmal Nitril Untersuchungshandschuhe	JOZA, München, Germany		disposable gloves
Laryngoscope, 45.48.50, KL 2000	Medicon		Laryngoscope handle
Littmann Classic III Stethoscope	3M Deutschland GmbH, Neuss, Germany		stethoscope
Luer Lock	B.Braun Melsungen AG, Germany
Maimed Vlieskompresse	Maimed GmbH, Neuenkirchen, Germany		Fleece compress to fix the tongue
Masimo LNCS Adtx SpO2 sensor	Masimo Corporation Irvine, Ca 92618 USA		saturation clip for the tail
Masimo LNCS TC-I SpO2 ear clip sensor	Masimo Corporation Irvine, Ca 92618 USA		Saturation clip for the ear
Masimo Radical 7	Masimo Corporation Irvine, Ca 92618 USA		periphereal oxygen saturation
Midazolam 15 mg/3 mL	Hameln Pharma GmbH, Hameln, Germany		midazolam
Midmark Canine Mask Small Plastic with Diaphragm FRSCM-0005	Midmark Corp., Dayton, Ohio, USA		dog ventilation mask
Octeniderm farblos	Schülke & Mayr GmbH, Nordenstedt, Germany		Alcoholic disinfectant
Original Perfusor syringe 50 mL	B.Braun Melsungen AG, Germany		perfusor syringe
Perfusor FM Braun	B.Braun Melsungen AG, Germany		syringe pump
Propofol 2% 20 mg/mL (50 mL flasks)	Fresenius, Kabi Deutschland, GmbH		propofol
RÜSCH Führungsstab für Endotrachealtubus (ID 5.6 mm)	Teleflex Medical Sdn. Bhd, Malaysia		PVC coated tube guiding wire
Rüschelit Super Safety Clear >ID 6/6.5 /7.0 mm	Teleflex Medical Sdn. Bhd, Malaysia		endotracheal tube
Stainless Macintosh Größe 4	Welch Allyn69604		blade for laryngoscope
Sterofundin	B.Braun Melsungen AG, Melsungen, Germany		Balanced electrolyte solution
Ultrastop Antibeschlagmittel bottle with dropper 25 mL	Sigmapharm Arzneimittel GmbH, Wien, Austria		Antifog agent
Vasofix Safety 22 G-16 G	B.Braun Melsungen AG, Germany		venous catheter
VBM Cuff Manometer	VBM Medizintechnik GmbH, Sulz a.N., Germany		cuff pressure gauge
Zelette	Lohmann & Rauscher International GmbH & Co. KG, Rengsdorf, Germany		Tissue swab