Mechanism Of IL-33Expression In Relation With Lung Injury In Humans And Mice With Burn-blast Combined Injury

Objective:Burn-blast combined injury is one of the most common injuries in military conflict, terrorist attacks, traffic accidents, and other emergencies. Lung injury after burn-blast combined injury results in immediate death in up to47%of cases. Acute lung injury (ALI) and acute respiratory distress syndrome (ARDS) are the main reasons for abrupt death in such injuries. Hypovolemic shock develops immediately after such injuries, initiating the generalized inflammatory cascade. Prior research in ALI and ARDS has demonstrated the importance of cytokines and neutrophils. Therefore, we focused on investigating cytokine-mediated neutrophils in the lungs after burn-blast combined injury. IL-33is a recently described member of the IL-1family, which also includes IL-1α/β, IL-1Ra, and IL-18. IL-33mRNA is expressed by multiple organs and cell types in humans and mice. IL-33protein is mainly expressed by epithelial cells, endothelial cells, and fibroblasts. IL-33attenuates sepsis by enhancing neutrophil influx to the site of infection. However, the function of IL-33-enhanced neutrophil influx in lung injury is unclear, specifically after burn-blast combined injury. Therefore, we established a burn-blast combined injury mice model. We investigated that IL-33would promote the participation of neutrophil migration in lung injury.Methods:Burn-blast combined injury was induced in wild-type (WT) mice and IL-33transgenic mice. Mice were randomly divided into control group, burn group, blast group and burn-blast combined group. Immunohistochemistry assay was used to detect myeloperoxidase (MPO), IL-33, GRK2and CXCR2in lung after burn-blast combined group. Transmission electron microscopy (TEM) was used for observation of changes in lung. Plasma IL-6and TNF-a levels were measured by ELISA kits. Micro-CT was used for observation of changes in lung. In addition, Immunohistochemistry assay was used to detect myeloperoxidase (MPO) and IL-33in humans with burn-blast combined (BBLP). Chest radiograph scans was used to detect lung in humans with burn-blast combined.Results: 1.We found that the mice at a distance of53cm, which exposed to94Ka peak overpressure, was the moderate blast injury distance when compared to mice at a distance of43cm and63cm, in these distances they were exposed200Ka and88.3Ka peak-overpressure, respectively.2. The lungs of mice in burn-blast combined group (BBL) were severely congestive and most mice had more than5hemorrhage spots. However, the lungs of mice in burn (BU) or blast group (BL) showed less degree of congestion. In addition, the lungs of mice in control group (CO) were normal. The burn-blast combined injury group showed a significantly greater wet-to-dry weight ratio than the blast group and burn groups. The lung hemorrhage area in the burn-blast combined injury group was significantly higher than that for the blast injury and burn groups.3. Burn-blast combined injury mice showed concomitantly reduced respiratory rate (Rr), tidal volume (Tv), minute ventilation volume (Mv), peak expiratory flow (PEF) as compared to burn injured mice, blast injured mice, and control mice at24hours. However, after burn-blast combined injury, WT mice exhibited significant lengthening of expiratory time (Te) and inspiratory time (Ti) as compared to each experimental group of WT mice at24hours.4. We found moderate ground-glass shadows were seen in the left lung tissue in the burn-blast combined injury group on performing micro-CT. However, ground glass slightly appeared in various regions of left lung in the blast group and no lesions in the lung tissue of the control group and burn group.5. MPO-, IL-33-and GRK2-positive cells significantly increased in the lung after burn-blast combined injury. However, the number of CXCR2-positive cells in WT mice was lower than those in the lungs of IL-33transgenic mice after burn-blast combined injury; Burn-blast combined injury mice had elevated plasma concentrations of IL-6and TNF-α.6. In burn-blast combined injured IL-33transgenic mice, blast-injured IL-33transgenic mice, and burn-injured IL-33transgenic mice., some significant positive correlations were observed between IL-33-positive cells and both GRK2-positive cells (r=0.65, p=0.042; r=0.638, p=0.0047; r=0.764, p=0.001, respectively) and MPO-positive cells (r=0.716, p=0.02; r=0.661, p=0.0037; r=0.755, p= 0.012, respectively). The correlation coefficients suggested a strong association between IL-33-positive and Mv in burn-injured mice (r=-0.677, p=0.031), blast-injured mice (r=-0.68, p=0.03), and burn-blast combined injury mice(r=-0.816, p=0.004).7. A large area of alveolar hemorrhage was seen in the patient with burn-blast combined injury. Interestingly, an increased number of MPO cells accumulated in the same area that IL-33is typically expressed.8. Lung chest radiograph scans of the patient with burn-blast combined injury showed moderate ground-glass shadows in the left lung, while other areas appeared normal or nearly normal.Conclusions:1. We successfully established a stable mice model with burn-blast combined injury, which is inflicted with moderate blast injury by placing C57BL/6mice53cm away from explosive source, generating pressure peak of94Kpa, followed by induction of25%TBSA full-thickness burns on the back with90℃water for9s.2. Our results suggest IL-33increases neutrophil influx to the site of infection and inflammation through GRK2pathway to block the downregulation of CXCR2with burn-blast combined injury.3. Our results suggest that increased IL-33is involved in lung injury in humans with burn-blast combined injury.