All animal experiments were performed with the approval of Animal Ethics Committee of Kunming Medical University. Animals were managed in accordance with Chinese Association for Laboratory Animal Sciences guidelines and current legal requirements. Our animal facilities meet the standards of Laboratory Animal-Requirements of Environment and Housing Facilities (GB 14925-2010).
2.1. Baseline data
Fifteen healthy miniature pigs (~16 weeks old, indicating sexual maturity), both male and female, weighing (28±5) kg on average, were provided by the animal laboratory of Kunming Medical University. The pigs used in the experiment were accustomed and fed in the laboratory for more than one day until all physiological parameters tended to be stable and be prepared for the subsequent experiment. The experimental animals were subject to dietary fasting but had free access to water on the night before surgery.
2.2. Self-designed inspiratory impedance threshold device (SIITD)
The SIITD was designed based on the principle of negative pressure suction under this background. This SIITD can intermittently block the inhalation of gas into lungs during the elastic expansion of chest wall after each CPR compression, and hence increase intrathoracic negative pressure and returned blood volume. Therefore, the blood supply of important organs such as heart coronary arteries, brain, liver, kidney can be increased and eventually improve the success rate of CPR. As illustrated in Figure 1, the SIITD was equipped with an oxygen supply end (10) connected with an oxygen supply pipe on the bottom (5) with a cavity (4). It had an air supply exhalation end (9) connected with a patient’s mask or a throat catheter and an evacuation end (6) connected with the atmosphere. A single control valve (2) and the first one-way valve (3) were set on the oxygen supply end (10). The second one-way valve (7) was designed on the evacuation end. One end of the column body (5) was fixed with the bent pipe (1). The vertical port (10) of the bent pipe (1) was downward to form an oxygen supply end connected with the oxygen supply pipe. The other end of the column body (5) was fixed with a straight pipe (8) with a vertical upper port (6) and a vertical lower port (9), which was an air supply exhalation end to be connected with a mask or a laryngeal airway of a patient. The vertical upper port (6) of the straight pipe was an evacuation end, and the second one-way valve (7) was designed on the evacuation end to allow for exhalation, whereas prevent inhalation.
2.3. Establishment of pig models
The fifteen pigs that we chose for experiments were similar in age, weight and physical appearance. Therefore, the grouping in our study were totally random. Fifteen healthy pigs were randomly assigned into three groups and each group includes five pigs. Anesthesia was performed on all pigs during all experimental procedures, all unnecessary suffering was avoided, and research was terminated if unnecessary pain or fear resulted in our pigs.
All pigs were subject to basic anesthesia via intramuscular administration of ketamine at a dose of 15 mg/kg, atropine 0.02 mg/kg, and midazolam 0.2 mg/kg, respectively. When pigs stood unstably after anesthetic administration, the animals were transferred and fixed on the operating table. Electrocardiogram was performed to measure the blood oxygen saturation (SPO2). The auricular vein was cut open and supplemented with the balance salt solution at a dose of 20 ml/kg. Intravenous injection of propofol 3 mg/kg and fentanyl 2 μg/kg were administered. Spontaneous breathing was retained and ID6.5-7.5 endotracheal tube with bursa was inserted with a depth of 16-18 cm. After successful intubation, the breathing machine was connected to perform intermittent positive pressure mechanical ventilation (IPPV). The breathing parameters were adjusted with a breathing frequency of 15-20 times/min, a tidal volume of 8-10 ml/kg, and an inspiratory-to-expiratory ratio of 1:2. The end-expiratory carbon dioxide partial pressure (ETPCO2) was kept at 35-40 mmHg, which was connected to the Datex. Ohmeda monitoring machine. The femoral artery puncture was performed and was connected to the transducer on the monitoring machine to measure the dynamic blood pressure. Throughout the entire experiment, the esophageal TEE probe was inserted into the esophagus by approximately 30 cm to measure the cardiac output and returned blood volume of the experimental animals.
Following anesthesia induction, the three groups of pigs respectively received intravenous injection of ketamine (model 1), MgSO4 (model 2) and KCl (model 3). Each pig in model 1 received intravenous injection of 200 mg ketamine after anesthesia procedures for cardiac arrest induction, and cardiac arrest was established in about 11 minutes after ketamine administration. Each pig in model 2 received intravenous injection of 100 mg MgSO4 after anesthesia procedures, and then 100 mg MgSO4 were injected in every minutes until cardiac arrest was established. It took overall about four minutes and 300 mg MgSO4 to cause cardiac arrest. Each pig in model 3 received intravenous injection of 1 g KCI after anesthesia procedures and cardiac arrest was established in about 30 seconds. Establishment of cardiac arrest was confirmed when the electrocardiogram hinted the signs of cardiac arrest and ventricular fibrillation occurred prior to cardiac arrest. By observing the ventricular fibrillation waveform of electrocardiogram, ventricular fibrillation was defined when the arterial blood pressure was lower than 40 mmHg. The mechanical ventilation was terminated upon the incidence of ventricular fibrillation.
The SIITD was connected and manual closed-chest CPR was initiated at the presence of cardiac arrest. The depth of chest was 25% (approximately 5 cm) of the anterior and posterior diameter of the chest. The compression frequency was 100 times/min to ensure the chest to fully rebound. The proportion of compression relaxation period was 1:1 to minimize the interruption as possible. To occlude the flow of gas into the lungs, SIITD was closed after each compression of CPR when the chest wall was pressed to the lowest point. The effect of increasing the blood flow to the heart was achieved by using the principle of negative pressure suction generated by increasing the elastic retraction of chest wall. After 2-, 6- and 10-min CPR, the heart rate and hemodynamic parameters including arterial blood pressure, blood oxygen saturation, end-diastolic volume and cardiac output were quantitatively measured. The Esophageal echocardiography and blood-gas analyses were performed.
CPR was terminated after pigs were confirmed dead. No of pigs was successfully rescued after experiments. Carcass of pigs were handed over to the Animal Laboratory of Kunming Medical University for unified handling.
2.5. Data collection
After the chest compression was started, the values of the heart rate (HR), mean arterial blood pressure (MAP), pulse blood oxygen saturation (SPO2), end diastolic volume (EDV), cardiac output (CO) and alternative parameters were recorded at 2 min, 4 min and 10 min, respectively.
2.6. Statistical analysis
SPSS 19.0 software was utilized for statistical analysis (SPSS Inc., Chicago, IL). Measurement data were expressed as mean ± standard deviation (x±s). Hemodynamics and blood gases during CPR were analyzed with single factor ANOVA. A P value of less than 0.05 was considered as statistical significance.