To explore the protective effect and mechanism of ghrelin on acute lung injury in mice.

Sixty mice were randomly divided into 6 groups, including control group, model group, low-dose ghrelin group, middle-dose group, high-dose group and dexamethasone group. The mice in control group and model group were intraperitoneally injected with 0.2 mL normal saline, the mice in ghrelin group were injected with 400 μg/kg, 200 μg/kg and 100 μg/kg respectively, and the mice in dexamethasone group were injected with 2 mg/kg. One hour after administration, 10 μg (20 μL) lipopolysaccharide (LPS) was instilled into the nasal cavity in model group as well as treatment groups, and the equal volume of saline was injected into the control group. After 24 hours, lung wet/dry weight (W/D) was detected. The contents of tumor necrosis factor-α (TNF-α) , interleukin-6 (IL-6) in bronchoalveolar lavage fluid (BALF) and the contents of TNF-α, IL-6 and IL-1β in serum were analyzed by ELISA. The pathological morphology of lung tissue was analyzed by HE staining. The expression of IL-1β p17, cysteinyl aspartate specific proteinase-1 (Caspase-1) p20, Nod-like receptor protein 3 (NLRP3) and GSDMD protein were detected by western blot. Alveolar macrophages were cultured in vitro and divided into control group, LPS+ATP group, low dose group, middle dose group and high dose group. The pyroptosis was analyzed by PI staining. And the expression of IL-1β p17, caspase-1 p20, NLRP3 and GSDMD proteins were observed by western blot. One-way analysis of variance was used for comparison between multiple groups, and LSD-t test was used for pairwise comparison between groups.

Compared with the control group, the lung W/D (4.03±0.46 vs 12.71±0.68) , TNF-α in BALF (3.92±0.59 vs 12.83±0.66) , IL-6 (23.94±3.51 vs 159.03±5.21) , serum levels of TNF-α (2.67±0.29 vs 13.23±0.76) , IL-6 (26.73±2.61 vs 141.64±3.86) and IL-1β (43.89±4.19 vs 249.03±5.38) in lung tissue of model group were increased, and IL-1β p17 (0.67±0.02 vs 0.93±0.02) , caspase-1 p20 (0.67±0.04 vs 1.02±0.08) , NLRP3 (0.58±0.04 vs 0.91±0.03) and GSDMD (0.46±0.06 vs 1.06±0.09) proteins expression were up-regulated significantly (P < 0.05) . Compared with the model group, W/D (12.71±0.68 vs 11.13±0.53, 7.56±0.31, 6.12±0.32, 6.14±0.34) and the contents of TNF-α (12.83±0.66 vs 9.89±0.47, 9.78±0.53, 7.33±0.27, 6.27±0.38) , IL-6 (159.03±5.21 vs 130.32±2.49, 122.87±3.31, 67.42±1.70, 56.45±3.33) in BALF and the contents of TNF-α (13.23±0.76 vs 10.14±0.52, 9.04±0.46, 6.43±0.38, 6.35±0.26) , IL-6 (141.64±3.86 vs 121.89±3.34, 116.42±2.68, 71.23±3.02, 78.54±5.13) and IL-1β (249.03±5.38 vs 230.14±5.53, 196.53±6.41, 100.67±3.50, 91.56±4.29) were dose-dependently decreased in serum in low dose group, middle dose group, high dose group and dexamethasone group, and the expression of IL-1β p17 (0.93±0.02 vs 0.84±0.01, 0.71±0.02, 0.61±0.04, 0.60±0.02) , caspase-1 p20 (1.02±0.08 vs 0.90±0.03, 0.81±0.02, 0.63±0.03, 0.61±0.03) , NLRP3 (0.91±0.03 vs 0.85±0.03, 0.68±0.05, 0.64±0.02, 0.68±0.03) and GSDMD (1.06±0.09 vs 0.71±0.02, 0.75±0.02, 0.67±0.03, 0.61±0.01) protein in lung tissue was dose-dependently downregulated (P < 0.05) . Compared with the control group, the number of PI-positive cells in the LPS+ATP group was increased, the cells were swelled and the membrane was ruptured, and IL-1β p17 (0.44±0.01 vs 0.99±0.03) and caspase-1 p20 (0.37±0.01 vs 1.32±0.02) , NLRP3 (0.39±0.02 vs 1.31±0.01) and GSDMD (0.39±0.01 vs 0.83±0.02) in alveolar macrophages were significantly up-regulated, and the difference was statistically significant (P < 0.05) . Compared with LPS+ATP group, the number of PI positive cells in low, middle and high dose groups was dose-dependently decreased, cell swelling and membrane rupture were relieved, and the expression of IL-1β p17 (0.99±0.03 vs 0.55±0.02, 0.45±0.02, 0.31±0.02) , caspase-1 p20 (1.32±0.02 vs 0.45±0.02, 0.42±0.02, 0.09±0.01) , NLRP3 (1.31±0.01 vs 0.90±0.02, 0.82±0.02, 0.33±0.01) and GSDMD (0.83±0.02 vs 0.67±0.04, 0.49±0.01, 0.35±0.02) in alveolar macrophages was dose-dependently downregulated (P < 0.05) .

Ghrelin has protective effect on acute lung injury in mice, which may be related to NLRP3 inflammasome mediated inflammatory response and pyroptosis in alveolar macrophages.