JFNE-A isolated from Jing-Fang n-butanol extract attenuates lipopolysaccharide-induced acute lung injury by inhibiting oxidative stress and the NF-κB signaling pathway via promotion of autophagy

Abstract

BackgroundJing-Fang powder consists of Jingjie (Nepeta tenuifolia Benth, (Lamiaceae)). and Fangfeng (Saposhnikovia divaricata (Turcz.) Schischk, (Apiaceae)) Previous studies have revealed that the Jing-Fang powder n-butanol extract (JFNE) has anti-acute lung injury (ALI) and anti-inflammatory properties; however, the active ingredient and mechanism remain unknown. PurposeIn the present study, we investigated the anti-inflammatory effect of a bioactive fraction obtained from JFNE(JFNE-A) on lipopolysaccharide (LPS)-induced ALI in mice and explored the underlying mechanism. Study designThe anti-acute lung injury effect and mechanism of JFNE-A was investigated by prophylactic administration of JFNE-A in mice with LPS-induced acute lung injury. MethodsThe expression levels of myeloperoxidase(MPO) in lung tissues of mice and interleukin(IL)-6, tumor necrosis factor(TNF)-α, IL-1β, IL-5, interferon (IFN)-γ, monocyte chemotactic protein (MCP)-1, macrophage colony stimulating factor (M-CSF), macrophage inflammatory protein (MIP)-1α, and MIP-1β in bronchi alveolar lavage fluid (BALF) were detected by reagent kit and the histological changes were examined by hematoxylin and eosin (H & E) for general histopathological conditions under a light microscope. In addition, the ultrastructure of the cells in lung tissues were observed and photographed under a transmission electron microscope. The expression levels of protein were detected via Western blotting and the mRNA expression of relative genes were determined of via reverse transcriptase polymerase chain reaction (RT-PCR). What's more, we also further clarified the potential targets of JFNE-A through network pharmacology analysis, which could be utilized in ALI treatment. ResultsOur results showed that pretreatment with JFNE-A for 7 days significantly reduced the lung pathological injury score, alleviated pulmonary edema, and decreased the lung tissue MPO level. Mechanistically, JFNE-A dramatically downregulated the protein levels of IL-6, TNF-α, IL-1β, M-CSF, and IFN-γ in BALF and mRNA expression levels of IL-6, TNF-α, IL-1β, and IFN-γ in lung tissues. JFNE-A also significantly lowered the protein levels of iNOS and phosphorylated NF-κB (p65) and mRNA expression levels of iNOS, Rela, CHUK, and NF-κB1, and also elevated the protein expression levels of Nrf2, HO-1, and SOD1 and the mRNA expression levels of Nrf2, Hmox1, and Keap-1 in the lungs. Moreover, JFNE-A significantly decreased the protein expression of p62 and increased the ratio of LC3II/LC3I. It also upregulated the mRNA expression levels of Atg5 and Beclin-1, whereas it reduced the mRNA expression level of SQSTM1 and increased autophagosome structures. ConclusionOverall, treatment with JFNE-A ameliorated LPS-induced ALI in mice by suppressing the NF-κB signaling pathways and promoting Nrf2 signaling pathways by accelerating autophagy.