Strictosamide alleviates acute lung injury via regulating T helper 17 cells, regulatory T cells, and gut microbiota.

BACKGROUND: Nauclea officinalis (Pierre ex Pit.) Merr. & Chun (Rubiaceae) is widely used to treat respiratory diseases in China. Strictosamide is its main active component and has significant anti-inflammatory activity. However, the effects and molecular mechanisms of strictosamide in the treatment of acute lung injury (ALI) remain largely unknown. PURPOSE: This study aimed to examine the regulatory effects of strictosamide on T helper 17 cells (Th17 cells)/Regulatory T cells (Treg cells) and gut microbiota in ALI-affected mice. MATERIALS AND METHODS: The ALI model was induced using lipopolysaccharide (LPS) intraperitoneal injection. Hematoxylin-eosin (H&E) staining, the number of inflammatory cells in broncho-alveolar lavage fluid (BALF), the Wet/Dry (W/D) ratio, and myeloperoxidase (MPO) activity were utilized as evaluation indices for the therapeutic efficacy of strictosamide on ALI. Flow cytometry (FCM), enzyme-linked immune sorbent assay (ELISA), quantitative reverse transcription polymerase chain reaction (qRT-PCR), and western blotting were used to determine the regulation of strictosamide on the Th17/Treg cells and the STAT3/STAT5 signaling pathway. The analysis of gut microbiota was conducted using 16S rDNA sequencing. The verification of the relationship between the gut microbiome and immune function was conducted using Spearman analysis. RESULTS: Strictosamide attenuated inflammation on ALI induced by LPS, which reduced the levels of Th17-related factors interleukin (IL)-6 and IL-17 and increased Treg-related factors IL-10 and transforming growth factor (TGF)-ß. In the spleens and whole blood, strictosamide reduced the proportion of Th17 cells and increased the proportion of Treg cells. Furthermore, strictosamide increased Forkhead/winged helix transcription factor 3 (Foxp3) and p-STAT5 protein expression while inhibiting Retinoid-related orphan nuclear receptors-γt (RORγt) and p-STAT3 expression. Moreover, strictosamide reshaped the diversity and structure of the gut microbiota, and influence the associations between immune parameters and gut microbiota in ALI mice. CONCLUSIONS: In summary, the results of the current investigation showed that strictosamide has a therapeutic impact on LPS-induced ALI. The mechanism of action of this effect may be associated with the modulation of Th17 and Treg cells differentiation via the SATA signaling pathway, as well as the impact of the gut microbiota.

Circ_0138960 knockdown alleviates lipopolysaccharide-induced inflammatory response and injury in human dental pulp cells by targeting miR-545-5p/MYD88 axis in pulpitis.

Background/purpose: Circular RNAs (circRNAs) have been shown to play important regulatory roles in many human diseases, yet their functions in pulpitis remain to be clarified. This study was designed to investigate the function of circ_0138960 in pulpitis progression and its underlying mechanism. Material and methods: Cell viability and proliferation were analyzed by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and 5-Ethynyl-2'-deoxyuridine (EdU) assay. Flow cytometry and enzyme-linked immunosorbent assay (ELISA) were conducted to analyze cell apoptosis rate and the release of inflammatory cytokines. The activity of superoxide dismutase (SOD) was analyzed using a SOD assay kit. Dual-luciferase reporter and RNA-pull down assays were performed to verify the interaction between microRNA-545-5p (miR-545-5p) and circ_0138960 or myeloid differentiation primary response gene 88 (MYD88). Results: Lipopolysaccharide (LPS) treatment restrained the proliferation and promoted the apoptosis, inflammation, and oxidative stress of human dental pulp cells (hDPCs). LPS treatment dose-dependently up-regulated circ_0138960 expression in hDPCs. Circ_0138960 knockdown overturned LPS-induced inflammation and injury in hDPCs. Circ_0138960 could act as a molecular sponge for miR-545-5p, and circ_0138960 knockdown protected hDPCs from LPS-induced effects by up-regulating miR-545-5p. miR-545-5p directly interacted with the 3' untranslated region (3'UTR) of MYD88, and MYD88 overexpression reversed miR-545-5p-mediated effects in LPS-treated hDPCs. Circ_0138960 positively regulated MYD88 expression by sponging miR-545-5p in hDPCs. LPS could activate nuclear factor kappa-B (NF-κB) signaling by targeting circ_0138960/miR-545-5p/MYD88 axis in hDPCs. Conclusion: Circ_0138960 knockdown attenuated LPS-induced inflammatory response and injury in hDPCs by targeting the miR-545-5p/MYD88/NF-κB axis.

Integrated metabolomics, network pharmacology and biological verification to reveal the mechanisms of Nauclea officinalis treatment of LPS-induced acute lung injury.

BACKGROUND: Acute lung injury (ALI) is a severe inflammatory disease, underscoring the urgent need for novel treatments. Nauclea officinalis Pierre ex Pitard (Danmu in Chinese, DM) is effective in treating inflammatory respiratory diseases. However, there is still no evidence of its protective effect against ALI. METHODS: Metabolomics was applied to identify the potential biomarkers and pathways in ALI treated with DM. Further, network pharmacology was introduced to predict the key targets of DM against ALI. Then, the potential pathways and key targets were further verified by immunohistochemistry and western blot assays. RESULTS: DM significantly improved lung histopathological characteristics and inflammatory response in LPS-induced ALI. Metabolomics analysis showed that 16 and 19 differential metabolites were identified in plasma and lung tissue, respectively, and most of these metabolites tended to recover after DM treatment. Network pharmacology analysis revealed that the PI3K/Akt pathway may be the main signaling pathway of DM against ALI. The integrated analysis of metabolomics and network pharmacology identified 10 key genes. These genes are closely related to inflammatory response and cell apoptosis of lipopolysaccharide (LPS)-induced ALI in mice. Furthermore, immunohistochemistry and western blot verified that DM could regulate inflammatory response and cell apoptosis by affecting the PI3K/Akt pathway, and expression changes in Bax and Bcl-2 were also triggered. CONCLUSION: This study first integrated metabolomics, network pharmacology and biological verification to investigate the potential mechanism of DM in treating ALI, which is related to the regulation of inflammatory response and cell apoptosis. And the integrated analysis can provide new strategies and ideas for the study of traditional Chinese medicines in the treatment of ALI.