Objective:To explore the targeted regulation of the inflammatory pathway and its mechanism after AMPK phosphorylation induced by lipopolysaccharide (LPS) in mice and human monocytes induced by THP-1, so as to provide evidence for the clinical application of Mogrol (MO) in the clinical treatment of acute lung injury.Methods:Twenty-four clean C57BL/6 male mice aged 6-8 weeks were randomly (random number) divided into the control group, MO group, LPS group and LPS+ MO group with 6 mice in each group. Mice in the control group were intraperitoneally injected with normal saline (30 mL/kg), mice in the MO group were intraperitoneally injected with MO (30 mg/kg), mice in the lipopolysaccharide group were intraperitoneally injected with lipopolysaccharide (10 mg/kg), mice in the lipopolysaccharide + MO group were intraperitoneally injected with MO (30 mg/kg), and the other side was injected with lipopolysaccharide (10 mg/kg) 30 min later. After 12 h, the mice were sacrificed for sampling and pathology and molecular biological tests were carried out. Cell experiment: THP-1 cells in good condition were cultured in RPMI 1640 medium containing 10% fetal bovine serum for 24 h, and then induced to differentiate into macrophages with 100 ng/mL PMA. The control group, MO group, LPS group and LPS + MO group were established. After drug stimulation, the cell suspension of each group was collected, and the cells and culture medium supernatants were used for subsequent detectionResults:Compared with the control group, the injury degree of the lipopolysaccharide group was obvious, the alveolar cavity structure was destroyed, the inflammatory cell infiltration was increased, and the alveolar septum was obviously thickened in the tissue sections. After MO intervention, the injury degree of lung tissue injury was greatly improved, and MPO and the lung wet/dry weight ratio were also significantly decreased. The mRNA levels of the inflammatory cytokines IL-1β, IL-6 and TNF- α in lung tissues were also significantly decreased under MO intervention [(2.96±0.10) vs. (5.53±0.14), (8.62±0.17) vs. (12.31±0.09), (3.01±0.09) vs. (4.85±0.36)]. The expression levels of NLRP3, caspase-1 p20, GSDMD-N and ASC in the lung tissues of mice in the lipopolysaccharide group were significantly higher than those in the control group, while the phosphorylation level of AMPK in the lipopolysaccharide + MO group was increased, and the expression of scorched death-related proteins was effectively inhibited [(0.58±0.09) vs. (0.89±0.15), (0.19±0.08) vs. (0.93±0.16), (0.65±0.09) vs. (0.86±0.14), (0.30±0.12) vs. (0.47±0.10), all P<0.05]. At the same time, the secretion of the inflammatory factors IL-1β and IL-18, the main markers of scorch death in the tissue measured by ELISA, could also be alleviated by MO. In the cell experiment, MO also promoted the phosphorylation of AMPK, inhibited the expression of proteins related to NLRP3 inflammatory bodies, and significantly improved cell viability. Conclusions:MO attenuates LPS-induced acute lung injury by inhibiting NLRP3-mediated cell pyrogenesis by promoting the phosphorylation of AMPK.

Objective:To investigate the role and mechanism of exogenous derivative 4-octyl itaconate (4-OI) in lipopolysaccharide (LPS)-induced acute lung injury (ALI).Methods:C57BL/6 male mice were randomly divided into the control group, 4-OI group, LPS group, 4-OI+LPS group and deferiprone (DFP)+LPS group, with 6 mice in each group. LPS-induced ALI model was established by intraperitoneal injection of LPS. For the 4-OI+LPS group, mice were pretreated with 4-OI for 2 h before stimulation with LPS. The mice were sacrificed 12 h later and lung tissues were collected for pathological and molecular biological examination. Hematoxylin-eosin and Masson staining were used to detect the level of lung injury and collagen deposition. The expression levels of inflammatory cytokines and ferroptosis associated genes were detected by real-time quantitative PCR, and ferroptosis associated proteins were detected by Western blotting. The chi-square test was performed before the measurement data were counted. One-way analysis of variance was used to compare differences between multiple groups.Results:Compared with the control group, the histopathological damage was aggravated, and collagen deposition and lung injury score and lung wet-dry ratio were significantly increased in the LPS group (all P<0.05), and 4-OI pre-treatment significantly alleviated LPS-induced ALI. 4-OI treatment also significantly reduced the mRNA level of inflammatory cytokines, including IL-1β [(4.38±0.47) vs. (32.65±4.49)], IL-6 [(3.97±0.64) vs. (12.22±0.91)] and TNF-α [(15.06±2.26) vs. (38.53±2.31)]. At the same time, compared with the control group, the levels of lipid peroxidation metabolite 4-hydroxynonenal and malondialdehyde, iron level of lung tissue were significantly increased in the LPS group, and the mRNA level of ferroptosis marker prostaglandin-endoperoxide synthase 2 was also significantly increased (all P<0.05), but these indicators were significantly lower in the 4-OI+LPS group than in the LPS group. The results of immunofluorescence, Western blotting and PCR showed that the protein levels of nuclear factor erythroid 2-related factor 2 (Nrf2), glutathione peroxidase 4 (GPX4) and ferritin heavy chain (FTH1) significantly decreased in the LPS group, while 4-OI treatment significantly increased the Nrf2, GPX4 and FTH1 levels, and showed similar inhibition of ferroptosis with DFP (all P<0.05). Conclusions:4-OI attenuates LPS-induced ALI by increasing Nrf2 and upregulating FTH1 and GPX4, providing a potential drug and its theoretical mechanism for the prevention and treatment of ALI.