Abstract:
Objective To investigate the interventional effects of BAY11-7082 on lung inflammatory response at the early stage and acute lung injury of rats with severe burns.
Methods (1) Experiment 1. Twelve Sprague-Dawley (SD) rats were divided into control (C) group and burn (B) group according to the random number table, with 3 rats in group C and 9 rats in group B. Rats in group C did not receive any special treatment. Rats in group B were inflicted with 30% total body surface area full-thickness burn on the back. Immediately after injury, rats in group B were intraperitoneally injected with normal saline in the dosage of 50 mL/kg. Abdominal aorta blood and lung tissue samples were collected from three rats in group B at post injury hour (PIH) 12, 24, and 48, respectively. The interleukin-1β (IL-1β) and the IL-18 content of serum were determined with enzyme-linked immunosorbent assay. The mRNA expressions of IL-1β and IL-18 in lung tissue were determined with real-time fluorescent quantitative reverse transcription polymerase chain reaction (RT-PCR). Sample collection and determination in rats of group C were performed as above. (2) Experiment 2. Eighteen SD rats were divided into control (C) group, simple burn (SB) group, and BAY11-7082 intervention (BI) group according to the random number table, with 6 rats in each group. Rats in group C did not receive any special treatment. Rats in groups SB and BI were inflicted with injury as in experiment 1. Immediately after injury, rats in group SB were intraperitoneally injected with normal saline in the dosage of 50 mL/kg, and those in group BI with 8 mg/mL (final mass concentration) BAY11-7082 solution in the dosage of 50 mL/kg. Lung tissue and bronchoalveolar lavage fluid (BALF) of rats with burns were collected at the optimal observation time point concluded from experiment 1. The morphology of lung tissue was observed with hematoxylin-eosin staining, and the pathological damage of lung tissue was graded. The myeloperoxidase (MPO) content of lung tissue and the total protein content of BALF were detected by microplate reader. The protein expressions of nucleotide-binding oligomerization domain-like receptor-3 (NLRP3) and cysteine-aspartic proteases 1 (caspase-1) in lung tissue were determined with Western-blotting. The mRNA expressions of IL-1β, IL-18, NLRP3, and caspase-1 in lung tissue were determined with real-time fluorescence quantitative RT-PCR. Sample collection and determination in rats of group C were performed as above. Data were processed with one-way analysis of variance and LSD-
t test.
Results (1) The IL-1β and IL-18 content of serum in rats of group B at PIH 12, 24, and 48 were significantly higher than those of group C (
t=10.55, 22.05, 12.47, 10.60, 15.22, 11.94,
P<0.01). The mRNA expressions of IL-1β and IL-18 in rats of group B at PIH 12, 24, and 48 were significantly higher than those of group C (
t=3.62, 7.19, 5.28, 3.20, 12.62, 7.31,
P<0.05 or
P<0.01). PIH 24 was the optimal observation time point for the following experiment. (2) At PIH 24, compared with those in group SB, the inflammatory cell infiltration and erythrocyte exudates of alveolar in group BI were obviously reduced, and the pulmonary interstitial edema obviously subsided. The pathological damage score of lung tissue in rats of group SB was (9.00±1.00) points, significantly higher than (1.10±0.26) points of group C (
t=13.23,
P<0.01). The pathological damage score of lung tissue in rats of group BI was (4.93±0.70) points, which was significantly lower than that of group SB (
t=5.76,
P<0.01) but still significantly higher than that of group C (
t=8.84,
P<0.01). At PIH 24, the MPO content of lung tissue and the total protein content of BALF in rats of group SB were (1.83±0.15) U/mg and (1.39±0.20) mg/mL, respectively, significantly higher than (0.51±0.10) U/mg and (0.44±0.05) mg/mL of group C (
t=12.50, 7.86,
P<0.01). The MPO content of lung tissue and the total protein content of BALF in rats of group BI were (0.91±0.12) U/mg and (0.60±0.10) mg/mL, respectively, significantly lower than those of group SB (
t=8.36, 6.06,
P<0.01). At PIH 24, the protein expressions of NLRP3 and caspase-1 in lung tissue of rats of group SB were 3.10±0.09 and 2.99±0.30, respectively, significantly higher than 1.00 and 1.00 of group C (
t=9.06, 11.28,
P<0.01). The protein expressions of NLRP3 and caspase-1 in lung tissue of rats of group BI were 1.13±0.08 and 1.81±0.11, respectively, significantly lower than those of group SB (
t=7.24, 3.91,
P<0.05 or
P<0.01). At PIH 24, the mRNA expressions of IL-1β, IL-18, NLRP3, and caspase-1 in lung tissue of rats in group SB were 5.0±0.4, 3.32±0.21, 3.54±0.42, and 6.3±1.0, respectively, significantly higher than 1.0, 1.00, 1.00, and 1.0 of group C (
t=13.97, 14.14, 11.78, 7.13,
P<0.01). The mRNA expressions of IL-1β, IL-18, NLRP3, and caspase-1 in lung tissue of rats in group BI were 2.6±0.5, 2.00±0.28, 1.39±0.21, and 2.5±0.5, respectively, significantly lower than those of group SB (
t=7.11, 5.80, 9.99, 4.65,
P<0.05 or
P<0.01).
Conclusions Applying BAY11-7082 at the early stage of acute lung injury of rats with severe burn can reduce the expression of caspase-1, decrease the levels of IL-1β and IL-18, and decrease the MPO content of lung tissue and the total protein content of BALF through inhibiting NLRP3, thus alleviating the lung inflammatory response and lung injury.
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