Dynamic pathophysiological features of early primary blast lung injury: a novel functional incapacity pig model

Abstract

Introduction

While there is evidence supporting the use of ultrasound for real-time monitoring of primary blast lung injury (PBLI), uncertainties remain regarding the timely detection of early PBLI and the limited data correlating it with commonly used clinical parameters. Our objective is to develop a functional incapacity model for PBLI that better addresses practical needs and to verify the early diagnostic effectiveness of lung ultrasound in identifying PBLI.

Methods

We selected six healthy male pigs to develop an animal model using a bio-shock tube (BST-I). The injuries were induced at a pressure of 4.8 MPa. We monitored the animals before and after the injury using various methods to detect changes in vital signs, lung function, and hemodynamics.

Results

The experimental peak overpressure was measured at 405.89 ± 4.14KPa, with the duration of the first positive peak pressure being 50.01ms. The mortality rate six hours after injury was 50%. The average Military Combat Injury Scale was higher than 3. Significant increases were observed in heart rate (HR), shock index (SI), alveolar-arterial oxygen gradient (AaDO₂), lung ultrasound scores(LUS), and pulmonary vascular permeability index (PVPI) at 0.5 h, 3 h, and 6 h after-injury (p < 0.05). Conversely, there were notable decreases in average arterial pressure(MAP), oxygenation index (OI), stroke volume per heartbeat(SV), cardiac output power index(CPI), global end-diastolic index (GEDI), and intrathoracic blood volume index (ITBI) during the same time periods (p < 0.05). Meanwhile, the extrapulmonary water index (ELWI) showed a significant increase at 0.5 h and 6 h after injury (p < 0.05). At 6 h after injury, pulmonary ultrasound scores were positively correlated with HR (R = 0.731, p < 0.001), AaDO₂ (R = 0.612, p = 0.012), SI (R = 0.661, p = 0.004), ELWI (R = 0.811, p < 0.001), PVPI (R = 0.705, p = 0.002). In contrast, these scores were negatively correlated with SpO₂ (R = -0.583, p = 0.007),OI (R = -0.772, p < 0.001), ITBI (R = -0.637, p = 0.006).

Conclusion

We have successfully developed a novel, and highly reproducible animal model for assessing serious PBLI functional incapacity. This model displays immediate symptoms of hypoxia, decreased cardiac output, decreased blood volume, and abnormal lung ultrasound findings within 0.5 h of injury, with syptoms lasting for up to 6 h. Lung ultrasound evaluation is crucial for the early assessment of injuries, and is comparable to commonly used clinical parameters.

Supplementary Information

The online version contains supplementary material available at 10.1007/s00068-024-02672-y.

Introduction

In modern warfare and workplace accidents, blast injuries have emerged as a leading cause of trauma [1]. These injuries can be difficult to detect, making it challenging to assess them promptly and intervene effectively, which can increase mortality and fatality rates [2–5].

The Military Functional Incapacity Scale (MFIS) is based on an anatomy-focused coding system that has been shown to be effective for diagnosing combat injuries [6–9]. Although the concept of functional incapacity is broad and vague, it is closer to the early treatment needs.Timely detection and medical intervention for serious functional incapacity caused by blast lung injury (BLI) can reduce unnecessary functional incapacity [10–12]. The ability to perform various physical tasks relies on having healthy lungs. A decline or even loss of physical ability can be a direct result of impaired lung function. In cases of primary blast injury, lung injury is the most common type. PBLI detection is challenging due to compensatory mechanisms in the lungs, which may result in delayed symptoms. This accounts for a substantial portion of the higher death rate as well as the challenge of diagnosing PBLI, particularly in the early stages before hospitalization.

While evidence indicates that various physiological parameters associated with blast injuries can serve as benchmarks for pre-hospital injury assessment, accurate diagnosis often requires referral to a trauma center with advanced therapeutic capabilities, such as those of a Level 2 trauma system or higher. In this regard, blast injuries can be accurately diagnosed through ultrasound and CT scans [13–16].

The mechanism of PBLI is complex. By analyzing recent data on war trauma, explosions, and animal experiments, we have developed several highly sensitive injury severity scoring tools. However, these systems rely on anatomical and physiological parameters that are difficult to obtain in a pre-hospital setting. Furthermore, there is a lack of research on how blast injuries progress dynamically and functional incapacity in animal models, which limits our understanding of these types of injuries [17, 18].

The previous team confirmed that the use of 66-partition method ultrasound can be used for assessment and dynamic monitoring of BLI [19]. The MCIS describes the anatomical damage resulting from combat injuries, while the MFIS evaluates the tactical functional impairments affecting a soldier’s ability to perform combat tasks. Given that pulmonary function is the foundation for physical capabilities in tactical operations, our research focuses on the impairment of physical function. We utilize the MCIS for injury assessment and correlate it with relevant MFIS metrics to investigate the extent of functional incapacity [20]. However, the operation is complex, therefore, considering ethical considerations and national circumstances, this model allowed us to collect commonly employed clinical physiological parameters, bedside lung ultrasound in emergency (BLUE)-plus protocol (the 10-partition method), and other relevant data for analysis.

Our objective is to investigate the progression of physiological and pathological changes in animals subjected to serious BLI and to highlight the value of lung ultrasound examination scores in the early stages of blast injury.

Methods

Experimental animal

The experiment involved six healthy male pigs, each weighing an average of 25.492 ± 6.365 kg, which were obtained from the Animal Experimental Center of the Army Medical Center. The experimental procedures received approval from the Ethics Committee for Animal Experiments at the Third Affiliated Hospital of the Army Medical University (AMUWEC20223478). The animal production license number was SCXK (Chongqing) 2017–0002, and the animal use license number was SYXK (Chongqing) 2017–0002. Animal care and handling were performed strictly by the Guide for the Care and Use of Laboratory Animals.

Device and applications

The research utilized various specialized devices and applications. The primary device was the BST-I shock tube, which is self-developed and simulates an explosion by compressing air to produce a shock wave (Fig. 1). For venous access, we used a 24G closed venous catheter (Intima, China), Monitoring of vital signs (Mindray, China), Intravenous interventions employed a 5 F dual-lumen central venous catheter sourced (Pulsion Medical Systems SE, German), 4-F PICCO catheter (Medical Components, America). For imaging, we utilized a hand-held ultrasound device from (Hua Sheng, China). Blood gas analysis was performed with a portable analyzer (Abbott, USA), and blood collection was facilitated with a 3 ml arterial blood collector (BD, UK). Tissue samples and specimens were preserved and processed using a 10% formalin fixative, polyformaldehyde, hematoxylin-eosin staining, paraffin, etc.

Fig. 1.

The bio-shock tubes (BST-I) (mm). A Drive segment; B Laminate segment; C Expansion segment; D Transition segment; E Experimental segment; F Suspension mechanism

Animal preparation

Food was withheld from the animals for 12 h, and water was allowed for 6 h before the start of the experiment. After acclimatization to the laboratory environment (maintained at a temperature of 25 ± 0.5℃ and humidity of 60 ± 5%), the animals were sedated by an intramuscular injection of ketamine hydrochloride (4.0 mg/kg) and by an intravenous injection of propofol (4.0–6.0 mg/kg) through a catheter placed in the left marginal ear vein. Following anaesthesia induction, the pigs were placed in a supine position, and the tongue was secured using tongue forceps to maintain spontaneous breathing. The concentration of oxygen (FiO₂) equals 21% (We adopted a protocol to preserve spontaneous breathing in animals to reduce the effects of endotracheal intubation, ventilator-assisted breathing, etc., on pulmonary function parameters after lung injury). The operational areas (forehead, thoracic-dorsal region, medial aspect of the right hind limb, and tail) were shaved, followed by a chemical depilatory, and thoroughly wiped. Percutaneous sensors were attached to each area. The depth of sedation and anaesthesia was monitored using the Bispectral Index (BIS). All animals were administrated propofol at a rate of 3.2–6.0 mg/kg/h, sufentanil citrate at 0.4–0.7 µg/kg/h, and ketamine at 0.4–0.7 mg/kg/h. The infusion rates were adjusted to maintain a BIS between 65 and 80 and the Critical-Care Pain Observation Tool (CPOT) scores between 1 and 2 [21, 22]. After ensuring stable anaesthesia, an incision was made in the skin of the animal’s right hind limb. The muscles were bluntly dissected to expose the femoral artery and femoral vein. Using the Seldinger technique, femoral artery catheters and femoral central venous catheters were respectively placed under direct visualization (the tip of the central venous catheter was located near the opening of the right atrium, guided by surface ultrasound). Arterial blood pressure was measured using a PICCO catheter via the femoral artery, while the tip of the deep femoral venous catheter was ultrasound – located at the opening of the right atrium to measure central venous pressure. All transducers were zeroed and positioned at the level of the right atrium.

Animal model construction

After the preparation was completed, all animals were allowed to stabilize for 30 min, and all initial values were used as the before-injury values(baseline). Next, the animals were placed in a BST-I shock tube during the experimental phase. They were supported upright, facing the shockwave source with the distal end sealed (Fig. 2). The animals were exposed to blast pressures of 4.8 MPa. They were continuously sedated and anesthetised, parameters were collected 0.5 h, 3 h, and 6 h after injury (Table 1).

Fig. 2.

The schema of the experiment. A The support frame is used to keep the animal upright; B The demonstration of the monitoring

Table 1.

Blast injury parameters

Classifications	Parameters
Injury scaling	Military Combat Injury Scale (MCIS), Military Functional Incapacity Scale (MFIS)
ECG parameters*	Heart Rate (HR), Blood Pressure: Mean Arterial Pressure (MAP), Systolic Blood Pressure (SBP), Diastolic Blood Pressure (DBP). Respiratory Rate (R), Percutaneous Oxygen Saturation (SpO2), Perfusion Index (Pi), Blood Temperature (T)
Hemodynamic parameters	Central Venous Pressure (CVP), Cardiac Output (CO), Cardiac Index (CI), Global End – Diastolic Volume Index (GEDI), Intrathoracic Blood Volume Index (ITBI), Cardiac Function Index (CFI), Cardiac Power Index (CPI), Global Ejection Fraction (GEF), Venous to arterial carbon dioxide difference (GAP), Oxygen evolution reaction (OER)
Lung function parameters	Arterial Blood (PH), Arterial Partial Pressure of Oxygen (PaO2), Arterial Partial Pressure of Carbon Dioxide (PaCO2), Arterial Oxygen Saturation (SaO2), Lactate (LAC), Central Venous Partial Pressure of Oxygen (PcvO2), Central Venous Partial Pressure of Carbon Dioxide (PcvCO2), Central Venous Oxygen Saturation (ScvO2), Hemoglobin (Hb), Extravascular Lung Water Index (ELWI), Pulmonary Vascular Permeability Index (PVPI), Coalescence Lung Ultrasound Score (cLUSS), Oxygenation Index (OI), alveolar partial pressure of oxygen (PAO2), Alveolar to arterial oxygen tension difference (AaDO2)

*Automated data collection is performed every 5 min. The average values are recorded for 1 h before the injury, as well as 0.5 h, 3 h, and 6 h after the injury. The data within 1 h before and after each time point is averaged for statistical analysis

Hemodynamic parameters were measured using transpulmonary thermodilution. Physiological saline (10 mL, 0.9%) at 4 °C was used as the indicator and was injected via the central femoral venous catheter. Each measurement was performed three times, and the average value was used for the analysis. Arterial blood samples (20 mL) were withdrawn (and reinfused) from the PiCCO catheter. Then, an additional 1 mL sample was collected for arterial blood gas analysis using an arterial blood gas syringe. Similarly, 20 mL of blood was drawn (and reinfused) from the femoral central venous catheter, and an additional 1 mL sample was collected for central venous blood gas analysis using an arterial blood gas syringe.

Equations for parameter calculations:

1. 
2. 
3. 
4. 
5. 

* Oxygen uptake (VO₂), oxygen delivery (DO₂). The tip of the femoral venous catheter was placed at the right atrial junction with ultrasonographic assistance. Therefore, mixed venous oxygen saturation was replaced with ScvO₂ [23, 24].

Injury assessment using blast lung injury scoring systems

The assessment of injury was based on the Organ Injury Scaling(OIS), the Pathology Severity Scale of Lung blast Injury (PSSLBI), the Military Combat Injury Scale (MCIS) and the Military Functional Incapacity Scale (MFIS) (Table 2、Supplementary Table 3) [13, 25, 26]. The final grading criteria for injury severity is PSSLBI.

Table 2.

MCIS and MFIS for Blast lung injury scoring

MCIS				MFIS
Score	Scale	Description	Anatomical Coding	Score	Description
1	Minor	Superficial injuries that can be treated in theater. Minimal or no immediate significant functional impairment. Likely to return to duty within 72 h		1	Able to continue the mission
2	Moderate	No increased risk of death if treatment delayed because of the tactical situation		2	Able to contribute to sustaining the mission
		Likely immediate functional impairment
		Likely able to contribute to sustaining the mission
3	Serious	No shock or airway compromise	1. Single lobe or unspecified lung contusion.	3/4	Loss to mission
		Ideally should be treated within 6 h of injury at a medical treatment facility to avoid increasing	2. Single lobe or unspecified lung laceration.
		Functionally impaired; lost to mission	3.Hemothorax/pneumothorax/hemopneumothorax, unspecified, < 1000 cc hemothorax, < 50% single lung compression, etc.
4	Severe	Injuries that may result in shock or airway compromise;	1. Bilateral or unilateral multi-lobe lung laceration.
		some casualties with injuries in this category will have an increased risk of death or functional incapacity if not treated at a medical treatment facility within 6 h	2. Severe smoke inhalation injury, accompanied by progressive hypoxemia, upper airway edema, and bronchospasm leading to airway obstruction.
		Functionally impaired; lost to mission or military	3. Unilateral, moderate lung blast injury (overpressure); bilateral or unilateral multi-lobe lung contusion.
			4. Tension pneumothorax or bronchopleural fistula with significant air leak.
			5. Severe smoke inhalation injury, accompanied by progressive hypoxemia, upper airway edema, and bronchospasm leading to airway obstruction.
			6. Cardiac contusion, etc.
5	Likely lethal	Injuries that are likely, not survivable in a military setting including catastrophic injuries and those from which the casualty is likely to die within minutes of wounding	Severe, multi-lobe, or bilateral lung blast injury.	4/5	Lost to military

Pulmonary ultrasound examination

Lung ultrasound can analyze the location and severity of lung injury based on morphological changes. Using the BLUE-plus protocol, we collected the videos of the pulmonary ultrasound (more than 10 s) from the upper BLUE point, lower BLUE point, diaphragmatic point, PLAPs point (approximately located in the lower lobe), and posterior blue point, respectively, at 0.5 h, 3 h, and 6 h before and after injury [27]. The cLUSS (coalescence-traditional Lung ultrasound score), totaling the left and right cLUSS, is defined as follows: 0 points for A-line or ≤ 2 separate B-lines; 1 point for ≥ 3 B-lines; 2 points for coalescing - lines; 3 points for a tissue-like pattern (Fig. 3) [28].

Fig. 3.

Ultrasonic examination of pig lungs using the BLUE-plus scheme for zoning and different score images. In A, B, C and D, the rectangular grid represents the ultrasound probe scanning area of the pig’s bilateral lungs. The probe is oriented vertically to the direction of the ribs. The upper blue point (U), lower blue point (L), diaphragmatic point (D), PLAPS point (P), and posterior blue point (P’) should be marked on the image. E, F, G, and H correspond to ultrasound scores of 0 (A-line, red arrow), 1 (B-line, red arrow), 2 (diffuse B-line, red arrow), and 3 (tissue-like sign, red arrow), respectively

Lung pathological examination and wet-to-dry weight measurement

All animals were dissected after injury. Dead animals were dissected within 24 h, and those that survived for more than 24 h were dissected after being euthanized using an overdose of anaesthesia. Macroscopic anatomical features such as lung haemorrhages, tears, the percentage of the bleeding area relative to the total area, and the presence of hemothorax were recorded.

To determine the lung wet-to-dry weight ratio (W/D), lung tissue (with trachea and main bronchi removed) was wrapped in aluminum foil and weighed to obtain the “wet weight”. The tissue was then baked at a constant temperature of 60 °C for 72 h and re-weighed to get the “dry weight”. The W/D ratio was calculated as W/D = wet weight/dry weight. The lung coefficient was calculated as lung wet weight (g) divided by body weight (kg) [29]. Tissue samples were retained, stained with HE, and examined under a light microscope. Features examined included intravascular obstruction, in flammatory cell infiltration, pulmonary congestion, thickening of the alveolar septa, presence of amorphous material and detachment of the bronchiole lining as detailed in the Lung histology scoring system (Supplementary Table 4) [30].

Statistical analysis

Experimental data were analyzed using SPSS 26.0, and graphs were created using GraphPad Prism 8. The Shapiro-Wilk test was used to test the normality of the data. Normally distributed continuous variables are presented as mean ± standard error of the mean. Repeated one-way ANOVA was performed to compare the multimodal parameters. Quantitative data with a non-normal distribution were represented by the median. The Friedman test was employed to compare the pre-injury data with data from various post-injury periods pairwise. Parameters showing statistical differences were then evaluated for correlation using the Pearson correlation coefficient (R), and linear regression curve fitting was conducted. After Bonferroni correction for multiple comparisons, P < 0.05 was defined as statistically significant.

Results

The severity of blast lung injury in pathophysiology

Under the driving pressure of the shock tube was 4.8 MPa, the peak of the reflected wave was 405.89 ± 4.14KPa, and the duration was 50.01ms. After the injury, 2 out of 6 pigs survived for less than 1 h (22 min and 44 min), 1 survived for 85 min, and 3 survived for more than 360 min (Fig. 4). After the injury, all pigs exhibited varying degrees of bleeding from the mouth and nose. Pigs that died within 1 h showed, upon dissection, blood clots obstructing the bronchus. Before the pigs died, there were widespread petechiae on their chest and abdomen, cyanosis around the mouth and nose, and signs of respiratory distress. The lung injury severity score OIS was at least level IV, and the PSSLBI was serious or above. The wet-to-dry weight ratio of the lungs was 3.57 ± 0.72; the lung coefficient was 17.41 ± 2.72.

Fig. 4.

Survival curve within 3 h

Analysis of lung tissue that appeared normal on the surface revealed a slight thickening of the alveolar walls, infiltration by inflammatory cells, a reduction in the number of alveoli, and enlargement of the remaining alveoli. Grossly abnormal lung tissue showed pronounced thickening of the alveolar walls, infiltration by neutrophils, alveolar collapse, and even red blood cells diffusely filling the entire alveolus (Fig. 5). Tissue samples from six points on each animal’s left and right lungs were examined under a microscope after HE staining, resulting in a pathological score of 6.3 ± 1 score.

Fig. 5.

Anatomical illustrations of a pig’s lungs, heart, and brain after PBLI (40 μm). A. the anatomical image of the dorsal side of the lungs. B the anatomical image of the ventral side of the lungs. C shows pale, bloody foam sputum inside the main bronchus. D depicts a thrombus tree peeled from the segmental bronchus. E shows multiple haemorrhages beneath the endocardium of the left ventricle. F shows scattered haemorrhages beneath the endocardium of the right ventricle. G shows the dissected abdomen with bleeding and edema in the small intestine. H reveals that the dissected brain shows no apparent abnormalities. Images I, J, and K correspond to the markers ①, ② and ③in image A, respectively. Image L corresponds to the marker ④ in image E

All 6 pigs had scattered, multifocal haemorrhages on the endocardium of the left ventricle. 4 of them had a few scattered haemorrhages beneath the endocardium of the right ventricle, and 1 had a small patchy haemorrhage on the epicardium. Upon microscopic examination of the haemorrhages, diffuse bleeding was confined to the endocardium or epicardium, and there were no signs of oedema or bleeding in the myocardial layer. All six brain dissections revealed normal gyri and sulci, with no evident haemorrhages or swelling. Under microscopic HE pathology, no abnormalities were observed. Dissections of 3 abdomens revealed varying degrees of haemorrhage and oedema in the small and large intestines (Fig. 5).

Dynamic vital signs in pigs post blast lung injury

After the injury, HR and SI significantly increased at 0.5 h, 3 h, and 6 h (RM ANOVA, HR: F = 8.438, p = 0.016; SI: F = 24.512, p = 0.016), while MAP (RM ANOVA, F = 13.891, p = 0.001) decreased at these time points. In addition, the T, R, and Pi did not show significant change at 6 h after injury (T: Friedman test p = 0.818, R: F = 0.338, p = 0.799; Pi: F = 3.355, p = 0.132) (Fig. 6).

Fig. 6.

The 24-hour trend of HR, MAP, and SI changes after the BLI. HR: Before injury vs. 0.5 h (p = 0.021), 3 h (p = 0.026), 6 h (p = 0.047) after injury; MAP: Before injury vs. 0.5 h (p = 0.031), 3 h (p = 0.018), 6 h (p = 0.008) after injury; SI: Before injury vs. 0.5 h (p = 0.011), 3 h (p = 0.010), 6 h (p = 0.010) after injury

Dynamic hemodynamic parameters in pigs post-blast lung injury

After the blast injury, the SV (RM ANOVA, F = 22.548, p = 0.001), CI (RM ANOVA, F = 14.984, p = 0.044), CPI (RM ANOVA, F = 11.213, p = 0.012), GEDI (RM ANOVA, F = 21.721, p = 0.010), GEF (Friedman test, p = 0.040), and ITBI (RM ANOVA, F = 10.156, p = 0.045) significantly decreased. Significant differences in the hemodynamic parameters, including CFI, dpmx, SVRI, pH, LAC, CVP, GAP, and OER, were not observed pre-injury and post-injury (CFI: F = 1.065, p = 0.417, dpmx: F = 3.293, p = 0.10, SVRI: F = 1.109, p = 0.405, pH: F = 3.112, p = 0.175, LAC: Friedman test, p = 0.172, CVP: F = 2.505, p = 0.244, GAP: F = 1.481, p = 0.318, and OER: F = 1.406, p = 0.349) (Fig. 7).

Fig. 7.

Changes in hemodynamic parameters within 6 h post-injury. A SV decreased at 0.5 h (p = 0.041), 3 h (p = 0.003), and 6 h (p = 0.016) after injury compared with before injury. GEF did not significantly change at 0.5 h, but decreased at 3 h (p = 0.002) and 6 h (p = 0.025) post-injury compared with pre-injury. B No change in CI at 0.5 h (p = 0.055) and 6 h (p = 0.050) after injury. There was a decrease in CI at 3 h after injury (p = 0.007). CPI decreased at 0.5 h (p = 0.008), 3 h (p = 0.010), and 6 h (p = 0.036) after injury. C GEDI decreased at 0. 5 h (p = 0.021), 3 h (p = 0.017), and 6 h (p = 0.045) after injury. ITBI decreased at 0.5 h (p = 0.021), 3 h (p = 0.017), and 6 h (p = 0.045) after injury. *P < 0.05, †P < 0.01, Data are presented as the mean ± standard error of the mean. *P < 0.05, †P < 0.01 vs. baseline. Before injury: baseline

Dynamic lung function in pigs post blast lung injury

After the blast injury, significant decreases were observed in SpO₂ (RM ANOVA, F = 8.744, p = 0.027), OI (RM ANOVA, F = 6.473, p = 0.013), SaO₂ (RM ANOVA, F = 3.876, p = 0.038), and ScvO₂ (RM ANOVA, F = 3.534, p = 0.048) at 0.5 h, 3 h, and 6 h; however, AaDO₂ (RM ANOVA, F = 9.902, p = 0.003), cLUSS (RM ANOVA, F = 18.936, p = 0.044), PVPI (RM ANOVA, F = 6.248, p = 0.028), ELWI (RM ANOVA, F = 6.019, p = 0.016), and R-cLUSS (RM ANOVA, F = 56.543, p<0.001) significantly increased at the same time points (Fig. 8). Moreover, PaCO₂ and L-cLUSS did not change substantially at all time points (RM ANOVA, PaCO₂: F = 0.554, p>0.05; L-cLUSS: p>0.05).

Fig. 8.

Trend of changes in lung function parameters over 6 h post-injury. A SpO₂ decreased at 0.5 h (p = 0.013) and 6 h (p = 0.008) after blast injury, while no change was found at 3 h (p = 0.350). SaO₂ dropped at 0.5 h (p = 0.005) after injury and showed no change at 3 h (p = 0.093) and 6 h (p = 0.065). ScvO₂ decreased at 0.5 h (p = 0.007) and 3 h (p = 0.015) after injury, with no change at 6 h (p = 0.258). B OI decreased at 0.5 h (p = 0.002), 3 h (p = 0.023), and 6 h (p = 0.015) after injury. AaDO₂ increased at 0.5 h (p = 0.01), 3 h (p = 0.01), and 6 h (p = 0.01) after injury. C cLUSS increased at 0.5 h (p = 0.008), 3 h (p = 0.011), and 6 h (p = 0.009) after injury. PVPI increased at 0.5 h (p = 0.024), 3 h (p = 0.044), and 6 h (p = 0.007) after injury. ELWI increased at 0.5 h (p = 0.005) and 6 h (p = 0.013) after injury, with no change at 3 h (p = 0.084). D R-cLUSS increased at 0.5 h (p = 0.029), 3 h (p = 0.002), and 6 h (p = 0.007) after injury. *P < 0.05, †P < 0.01, Data are presented as the mean ± standard error of the mean. *P < 0.05, †P < 0.01 vs. baseline. Before: baseline

Lung ultrasound score of pigs post blast lung injury

In the early stages of the injury, there were varying degrees of ultrasonic abnormalities at different time points, primarily manifested as diffuse merging B-lines. Within 6 h after the injury, the lung ultrasound examination showed no signs of pneumothorax or fluid accumulation in the thoracic cavity, but cLUSS has a very strong correlation with OI (R = -0.772) and ELWI (R = 0.811) and a strong correlation with HR (R = 0.731) and PVPI (R = 0.705). It also has a moderate correlation with SI (R = 0.661), SpO₂ (R = -0.583), AaDO₂ (R = 0.612), and ITBI (R = -0.637) (Fig. 9).

Fig. 9.

The linear regression at 6 h post-injury. The correlation between cLUSS and parameters: HR and cLUSS (R = 0.731, p<0.001); SpO₂ and cLUSS (R = -0.583, p = 0.007); OI and cLUSS (R = -0.772, p<0.001); AaDO₂ and cLUSS (R = 0.612, p = 0.012); SI and cLUSS (R = 0.661, p = 0.004); ITBI and cLUSS (R = 0.637, p = 0.006); ELWI and cLUSS (R = 0.811, p<0.001); PVPI and cLUSS (R = 0.705, p = 0.002)

Discussion

Shockwaves are the primary cause of injuries from explosions, with the lung being the most commonly affected area. The BLI is a crucial criterion for assessing combat injuries in the US military. An MCIS score of 4 is considered critical, and its associated MFIS score is three or higher. However, there is still no available research on the pathophysiological dynamics of serious PBLI on an international scale [29, 31–36]. BST-I has been proven to establish stable animal injury models [30, 37]. Therefore, we used BST-I to creat a serious injury model and conducted animal experiments using handheld ultrasound and other multimodal monitoring methods [38]. The results showed that while there were no visible external injuries, the annimals exhibited such as increased HR, SI, Lung capillary permeability, and decreased cardiac output, blood volume. These symptoms were accompanied by respiratory failure, which persisted for up to 6 h. Ultrasound imaging of the lungs was able to detect lung injuries immediately after the blast, and it shows a strong correlationg with commonly used heart and lung function parameters within 6 h post-injury.

Based on our previous research, we selected a driving segment pressure of 4.8 Mpa for the shock wave tube to induce injuries. At this pressure, the peak overpressure and duration of the shock wave remained stable. The damage caused by shock waves is closely related to the explosive equivalent, distance, environment, and posture of the personnel. Our model creates injuries in a closed-end biological shock tube. Due to factors such as reflected waves and synthetic waves, the shock wave causes more severe injuries on the side of the pig farther from the shock wave source compared to the nearer side, manifesting as inconsistent lung injuries between the left and right lungs. Therefore, during the pulmonary ultrasound examination, it is essential to conduct a bilateral lung assessment to avoid missing any missed diagnoses.

Six animals had a 50% survival rate at 6 h, exhibiting symptoms like bleeding from the mouth and nose, dyspnea, cLUSS > 6, and continuous Type I respiratory failure. We successfully and reliably established a serious PBLI (Pulmonary Blast Injury) animal model using a combination of shock tube mechanical parameters, MCIS and MFIS [28, 39]. The associated MFIS score was 3, providing a model for studying blast injury functional incapacity evaluations.

Considering the long-term effects of anaesthesia on physiological parameters, we used Bis and CPOT to standardise analgesia and sedation depth before and after injury. Under the same conditions, before and after injury parameters analysis revealed high heart rates and low blood pressure in animals within 6 h after injury. Furthermore, hemodynamic parameters like SV, CPI, ITBI, and GEDI showed a decline at 0.5 h after the injury and persisted for 6 h, but CI and GEF only started declining after 3 h. This indicates that the decreased work by the heart, due to cardiac injury and reduced circulating blood volume, may be contributing factors. The combined evidence suggests that the heart increases its rate to enhance systemic perfusion and improve cardiac output per minute. [40–41]. Dissections revealed scattered hemorrhagic spots under the left ventricular endocardium, suggesting blast-induced cardiac dysfunction, reduced left ventricular output capacity, and reduced blood volume, leading to an increased shock index. Although PICCO indicated no statistically significant difference in peripheral vascular resistance, the continued low blood pressure and GEDI suggest that this wasn’t caused by a decrease in peripheral vascular resistance but by reduced cardiac function and circulating volume. Previous research on combat injuries also indicates [42–45]. That such injuries often manifest as hemorrhagic shock types with increased shock index. Our study didn’t test for cardiac electrophysiological function, but we observed frequent arrhythmias in animals within 6 h post-injury, suggesting possible electrophysiological dysfunction influencing hemodynamic changes [5, 46, 47].

In order to be closer to the situation of early treatment of war wounds, we do not make any additional treatment interventions, including tracheal intubation, except for appropriate analgesia and sedation. After injury, animals showed symptoms of bleeding from the mouth and nose, difficulty breathing, and an oxygenation index of around 200mmHg. Lung function parameters like SpO₂, SaO₂, and ScvO₂ decreased, while cLUSS, PVPI, ELWI, and AaDO₂ increased. There were no significant differences in PaCO₂. Lung dissections revealed diffuse bleeding and oedema in the lungs, with the right lung being more damaged than the left. The lung ultrasound score also indicated that the right side was more severely affected than the left, and the score gradually increased within 6 h. Moderate amounts of bloody secretions were found in the airways. Pathological manifestations included interstitial bleeding, oedema, lung inflammatory cell infiltration, and red blood cells filling the alveoli. There were no complete obstructions or blockages in the large airways, indicating that the primary cause of respiratory failure was a dysfunction in lung diffusion. The physiological changes after injury were consistent with previous pathological studies on blast lung injuries [36, 37, 48–53]. Considering the changes in lung function, we believe that after injury, the body exhibits decompensation. Oxygen transport function declines, not meeting the body’s oxygen consumption needs. Gradually, there’s a mismatch in large and small circulation, sacrificing small circulation like microcirculation to ensure perfusion and oxygen consumption in primary organs. The condition worsens without intervention, leading to irrecoverable decompensation and death [54–56].

Primary blast injury manifest not merely as inflammatory injuries to the lung capillaries and resulting oedema but also include damages like tearing of the lung capillaries, rupture of the alveoli, and increased pulmonary hydrostatic pressure [45, 57–59]. Changes in the parameters of organs like the heart and lungs at different phases reflect the body’s responses under various influences. Hence, we compared lung ultrasound scores with other parameters, revealing that the lung ultrasound score significantly increased at 0.5 h after primary blast lung injury. The score trend was consistent with the severity of the injuries. In correlation analyses, cLUSS showed a good correlation with HR, OI, ELWI, PVPI, SpO₂, SI, and AaDO₂, followed by ELWI and PVPI. Although ELWI recovered at 3 h post-injury, it increased again at 6 h post-injury. This phenomenon suggests that after the blast injury, the body attempts to reabsorb third-space fluid to increase effective circulating blood volume. However, the improved lung permeability results in reabsorbed fluid again seeping into lung interstitial spaces. The BLUE-plus protocol plays a significant role in early diagnosis and treatment of acute and critical conditions. Our experiments used handheld ultrasound to conduct lung examinations following the BLUE-plus protocol. We found that when PVPI > 3, all six animals had a cLUSS of > 6. Even though cLUSS increased 0.5 h post-injury and persisted within 6 h, whether we can predict the severity and trend of the injury will be the future direction.

Typical clinical manifestations of blast injury include mild external injuries and severe internal injuries. Lung damage is common, and early diagnosis is difficult, which can lead to misdiagnosis and death. Currently, pre-hospital trauma assessments rely heavily on various clinical monitoring aids, and there is a lack of essential examination and testing methods during early treatment. However, with the advancement of medical technology, using handheld ultrasound for non-invasive monitoring of the lungs allows for the rapid identification of pulmonary injuries in the early stages of blast injuries [60]. Moreover, its correlation with lung function parameters, such as ELWI, is relatively good. It can replace PICCO parameters within the first 6 h and can be used with other obtained parameters to dynamically monitor and assess injury progression [61–63]. This can guide our strategies for early treatment of blast injuries.

Our study has several limitations, which will be addressed in subsequent research. First, a small sample size might increase the risk of Type I errors. Second, to minimise heterogeneity and the influence of gender in blast injuries, this study only included male animals. Based on animal experimentation ethics and early trauma care standards, we employed multi-modal analgesia and sedation for our experimental study. However, we did not set up a control group to ascertain the potential impact of analgesic and sedative drugs on animals. Lastly, this study only focused on animals with a functional incapacity score of 3, and the pig model may not reveal an excellent non-human surrogate for the MFIS. In subsequent studies, we will expand the sample size and include animal experimental data across different functional incapacity levels.

Conclusion

Using a novel, stable, reproducible, and relevant pig model simulating PBLI to the heart and lungs, we can more effectively cater to the requirements of onsite forensic evaluations of injuries sustained in combat. The recent findings of this study systematically assessed parameters that exhibited significant disparities during the initial phases of PBLI and elucidated the temporal patterns of their evolution. Notably, cLUSS can serve as rapid and sensitive forensic indicators for assessing PBLI. To gauge their early forensic efficacy, it is imperative to establish a quantifiable correlation between these findings in large animal models and their applicability to humans.

Electronic supplementary material

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Abbreviations

MCIS

Military Combat Injury Scale

MFIS

Military Functional Incapacity Scale

HR

Heart Rate

MAP

Mean Arterial Pressure

SBP

Systolic Blood Pressure

DBP

Diastolic Blood Pressure

R

Respiratory Rate

SpO₂

Percutaneous Oxygen Saturation

Pi

Perfusion Index

Blood T

Temperature

CVP

Central Venous Pressure

CO

Cardiac Output

CI

Cardiac Index

GEDI

Global End–Diastolic Volume Index

ITBI

Intrathoracic Blood Volume Index

CFI

Cardiac Function Index

CPI

Cardiac Power Index

GEF

Global Ejection Fraction

GAP

Venous to arterial carbon dioxide difference

OER

Oxygen evolution reaction

PH

Arterial Blood

PaO₂

Arterial Partial Pressure of Oxygen

PaCO₂

Arterial Partial Pressure of Carbon Dioxide

SaO₂

Arterial Oxygen Saturation

LAC

Lactate

PcvO₂

Central Venous Partial Pressure of Oxygen

PcvCO₂

Central Venous Partial Pressure of Carbon Dioxide

ScvO₂

Central Venous Oxygen Saturation

Hb

Hemoglobin

ELWI

Extravascular Lung Water Index

PVPI

Pulmonary Vascular Permeability Index

cLUSS

Coalescence Lung Ultrasound Score

OI

Oxygenation Index

PAO₂

Alveolar partial pressure of oxygen

AaDO₂

Alveolar to arterial oxygen tension difference

Author contributions

Shifeng Shao: original draft, Validation, Project administration, Methodology, Investigation, Formal analysis, Data curation. Shasha Wu: Writing – review & editing, Methodology, Investigation, Data curation. Jun Liu: Writing – review & editing, Methodology, Investigation, Data curation. Zhikang Liao: Writing – review & editing, Methodology, Investigation, Data curation. Pengfei Wu: Writing – review & editing, Methodology, Investigation, Data curation. Yuan Yao: Writing – review & editing, Methodology, Investigation, Data curation. Zhen Wang: Methodology, Investigation, Funding acquisition, Data curation, Conceptualization. Liang Zhang: Writing – review & editing, Software, Methodology, Formal analysis. Yaoli Wang: Writing – review & editing, Supervision, Funding acquisition, Formal analysis, Conceptualization. Hui Zhao: Writing – review & editing, Supervision, Resources, Project administration, Methodology, Formal analysis, Conceptualization.

Funding

This work was supported by the Chongqing Municipality Special Project for Technological Innovation and Application Development (Grant number 318-084063), and Key specialty of the army’s clinical focus.

Data availability

No datasets were generated or analysed during the current study.

Declarations

Ethics approval and consent to participate

All experimental procedures were conducted in strict accordance with the Guide for the Care and Use of Laboratory Animals established by the Animal Ethics Committee of the Army Medical University (AMUWEC20223478).

Consent for publication

Not applicable.

Competing interests

The authors declare no competing interests.