Effect and mechanism of total glucosides of paeony protect mice from lipopolysaccharide-induced acute lung injury
Sun Jing , Xin Lingen, Feng Xianzhen , Ding Yuxuan , Zhang Yubiao, Wang Jia
2024, 45(2):
302-311.
doi:10.3969/j.issn.1006-7795.2024.02.019
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Objective To investigate the effect and mechanism of total glucosides of paeony (TGP) in the treatment of acute lung injury in mice. Methods Twenty-four female C57 mice were randomly divided into 4 groups： control group, lipopolysaccharide (LPS) model group, TGP group (LPS+TGP), and dexamethasone (DEX) group (LPS+DEX), with 6 mice in each group. Acute lung injury model was established by nasal inhalation of LPS (4 ng/kg) for 5 consecutive days. The LPS+TGP and LPS+DEX groups were given 50 mg·kg-1·d-1 TGP and 5 mg·kg-1·d-1 DEX by gavage, respectively. The changes in body weight of the mice in each group were observed and recorded. Lung tissue was taken for hematoxylin-eosin (HE) staining to observe lung pathological damage. Flow cytometry was used to detect the changes in the proportions of dendritic cells (DC), neutrophils, and macrophages in peripheral blood, spleen, and lung tissues, as well as the proportions of M1 and M2 macrophages in lung tissues. Enzyme linked immunosorbent assay (ELISA) was used to analyze the levels of interleukin-1β (IL-1β) and interleukin-6 (IL-6) in bronchoalveolar lavage fluid. Real-time quantitative polymerase chain reaction (RT-qPCR) was used to detect the mRNA expression levels of IL-1β, IL-6 and tumor necrosis factor-α (TNF-α) in lung tissue. Western blotting was used to detect the nuclear transcription factor-κB p65 (NF-κB p65) and phospho-NF-κB p65 (pNF-κB p65) protein content. Results TGP and DEX significantly improved the weight loss caused by LPS. HE staining showed that the lung tissue of mice in the LPS group had inflammatory cell infiltration, alveolar septal destruction, and alveolar interstitial thickening, which were mitigated in the LPS+TGP and LPS+DEX groups. The results of flow cytometry of peripheral blood showed that, compared with the LPS group, the proportion of neutrophils in the LPS+TGP and LPS+DEX groups was significantly decreased, while the proportions of DC and macrophages did not change significantly. The results of flow cytometry of spleen tissues showed that, compared with the LPS group, the proportions of neutrophils, DC, and macrophages in the LPS+TGP and LPS+DEX groups were significantly decreased. The results of flow cytometry of lung tissues showed that, compared with the LPS group, the proportion of macrophages in the LPS+TGP and LPS+DEX groups did not change significantly, the proportion of M1 cells decreased significantly, the proportion of M2 macrophages increased, and the proportion of neutrophil is down. ELISA results showed that, compared with the LPS group, the levels of IL-6 and IL-1β in bronchoalveolar lavage fluid were significantly decreased； RT-qPCR results showed that, compared with the LPS group, the expression levels of IL-1β, IL-6 and TNF-α mRNA in lung tissues of the LPS+TGP and LPS+DEX groups were significantly reduced. Western blotting results showed that, the protein expression of NF-κB p65 did not change significantly in the LPS+TGP and LPS+DEX groups compared with the LPS group, and the protein content of pNF-κB p65 was significantly decreased. Conclusion TGP can improve LPS-induced acute lung injury, which may be related to the reduction of the proportion of peripheral blood neutrophils, lung neutrophils and spleen inflammatory cells, the inhibition of M1 polarization of lung macrophages, the inhibition of NF-κB activation, and the reduction of inflammatory signaling pathway and inflammatory factor release.
