Inflammatory mediators play a key role in the pathogenesis of acute respiratory distress syndrome (ARDS). In this study, we aimed to explore the involvement of the Kcnq1 opposite strand/antisense transcript 1 (Kcnq1ot1)/miR-381-3p/E26 transformation-specific proto-oncogene 2 (ETS2) axis in inflammation of lipopolysaccharide (LPS)-induced ARDS. Microarray analysis revealed ETS2 as an upregulated gene in ARDS. Then, a LPS-induced ARDS mouse model was constructed, with a series of gain- or loss-of-function experiments conducted to evaluate the lung function and neutrophil extracellular trap (NET) formation in lung tissue and determine the neutrophil number, myeloperoxidase (MPO) activity, and inflammatory factor levels in bronchoalveolar lavage fluid (BALF). As the results revealed, downregulated expression of ETS2 resulted in improved lung function, decreased NETs, MPO activity, and levels of interleukin (IL)-6 and tumor necrosis factor alpha (TNF-α), as well as increased IL-10 level. Then, the assays of dual-luciferase reporter, RNA-binding protein immunoprecipitation (RIP), and RNA pull-down were performed to validate that Kcnq1ot1 promoted ETS2 expression by competitively binding to miR-381-3p. Meanwhile, it was also found that Kcnq1ot1 silencing reversed the promotive effect of EST2 on ARDS. Our results provide evidence that Kcnq1ot1 silencing may reduce the inflammatory response in LPS-induced ARDS via inhibition of miR-381-30-dependent ETS2, thereby presenting new molecular understanding for the development of ARDS.

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Background To investigate the effects of serum amyloid A1 (SAA1) on lipopolysaccharide (LPS) -induced inflammation in vascular smooth muscle cells (VSMCs). SAA1 expression was detected in LPS induced VSMCs at different concentrations for different time by using Western blotting. After pre-incubation with recombinant SAA1 protein, VSMCs were treated with 1 μg/ml LPS for 24 h. The VSMCs were then divided into Control, SAA1 siRNA, Nox4 siRNA, LPS, LPS + SAA1 siRNA, LPS + Nox4 siRNA and LPS + SAA1 siRNA + Nox4 groups. MTT was performed to observe the toxicity of VSMCs. Lucigenin-enhanced chemiluminescence method was used to detect superoxide anion (O 2 − ) production and NADPH oxidase activity. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to determine expressions of inflammatory factors. Western blotting was used to determine expressions of NOX-4 and p38MAPK/NF-κB pathway related proteins. Results LPS promoted SAA1 protein expression in a concentration−/time-dependent manner. Recombinant SAA1 protein could increase NOX4/ROS production and promote the release of inflammatory factors ( IL-1β , IL-6 , IL-8 , IL-17 , TNF-α and MCP-1 ) in LPS (1 μg/ml) - induced VSMCs. Besides, both SAA1 siRNA and NOX-4 siRNA could not only enhance the O 2 − production and NADPH oxidase activity, but also up-regulate the protein expression of NOX4, the release of inflammatory factors, and the levels of p-p38 and p-NF-κB p65 in LPS-induced VSMCs. However, no significant differences in each index were observed between LPS group and LPS + SAA1 siRNA + Nox4 group. Conclusion SAA1-mediated NOX4/ROS pathway could activate p38MAPK/NF-κB pathway, thereby contributing to the release of inflammatory factors in LPS-induced VSMCs.

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