探索乙酰丙炭酸乙酯在脂多醣（LPS）刺激的RAW 264.7小鼠来源巨噬细胞和斑马鱼中的抗炎作用及其潜在机制。采用3-[4,5-二甲基噻唑-2-基]-2,5-二苯基四唑溴化物（MTT）法进行实验，研究乙酰丙炭酸乙酯在不同浓度（12.5-100 µmol/L）对RAW 264.7细胞的毒性。细胞经LPS（100 ng/mL）刺激12小时，在体外建立炎症模型，通过酶联免疫吸附检测法（ELISA）检测前炎性细胞因子白细胞介素（IL）-6和肿瘤坏死因子 α（TNF-α）的产生。采用Western blot确定信号转导与激活转录因子3（STAT3）、核因子 kappa B（NF-κB） p65、磷酸化STAT3（p-STAT3, Tyr705）、核因子 kappa B的抑制因子（IκB）α和磷酸化I κB α（p-IκB α, Ser32）的蛋白表达，并采用共聚焦成像技术鉴定NF-κB p65和磷酸化STAT3（Tyr705）的核转位。此外，对斑马鱼（受精后3天）的脐囊注射2纳升LPS（0.5 mg/mL）来诱导体内炎症模型。通过生存分析、苏木精-伊红（HE）染色、中性粒细胞迁移观察和定量实时聚合酶链反应（qRT-PCR）进一步研究乙酰丙炭酸乙酯的抗炎作用及其可能的机制。乙酰丙炭酸乙酯的非毒性浓度范围为12.5至100 µmol/L。乙酰丙炭酸乙酯抑制LPS诱导的RAW 264.7细胞释放IL-6和TNF-α（P<0.05或P<0.01），降低了IκBα的降解和磷酸化（P<0.05），并减少了NF-κB p65和磷酸化STAT3（Tyr705）的核转位（P<0.01）。乙酰丙炭酸乙酯还降低了LPS诱导的斑马鱼炎症细胞浸润和中性粒细胞迁移，同时增加了其存活率（P<0.05或P<0.01）。此外，乙酰丙炭酸乙酯还抑制了LPS刺激的斑马鱼IL-6、TNF-α、IκBα、STAT3和NF-κB的mRNA表达水平（P<0.01）。乙酰丙炭酸乙酯通过抑制RAW 264.7巨噬细胞和斑马鱼中的NF-κB和STAT3信号通路发挥抗炎作用。©2023.中国中西医结合杂志社和Springer-Verlag GmbH Germany, part of Springer Nature.

To explore the anti-inflammatory effects of ethyl lithospermate in lipopolysaccharide (LPS)-stimulated RAW 264.7 murine-derived macrophages and zebrafish, and its underlying mechanisms.3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazoliumbromide (MTT) assays were performed to investigate the toxicity of ethyl lithospermate at different concentrations (12.5-100 µ mol/L) in RAW 264.7 cells. The cells were stimulated with LPS (100 ng/mL) for 12 h to establish an inflammation model in vitro, the production of pro-inflammatory cytokines interleukin (IL)-6 and tumor necrosis factor α (TNF-α) were assessed by enzyme linked immunosorbent assay (ELISA). Western blot was used to ascertain the protein expressions of signal transducer and activator of transcription 3 (STAT3), nuclear factor kappa B (NF-κB) p65, phospho-STAT3 (p-STAT3, Tyr705), inhibitor of NF-κB (IκB) α, and phospho-I κB α (p-IκB α, Ser32), and confocal imaging was used to identify the nuclear translocation of NF-κB p65 and p-STAT3 (Tyr705). Additionally, the yolk sacs of zebrafish (3 days post fertilization) were injected with 2 nL LPS (0.5 mg/mL) to induce an inflammation model in vivo. Survival analysis, hematoxylin-eosin (HE) staining, observation of neutrophil migration, and quantitative real-time polymerase chain reaction (qRT-PCR) were used to further study the anti-inflammatory effects of ethyl lithospermate and its probable mechanisms in vivo.The non-toxic concentrations of ethyl lithospermate have been found to range from 12.5 to 100 µ mol/L. Ethyl lithospermate inhibited the release of IL-6 and TNF-α(P<0.05 or P<0.01), decreased IκBα degradation and phosphorylation (P<0.05) as well as the nuclear translocation of NF-κB p65 and p-STAT3 (Tyr705) in LPS-induced RAW 264.7 cells (P<0.01). Ethyl lithospermate also decreased inflammatory cells infiltration and neutrophil migration while increasing the survival rate of LPS-stimulated zebrafish (P<0.05 or P<0.01). In addition, ethyl lithospermate also inhibited the mRNA expression levels of of IL-6, TNF-α, IκBα, STAT3, and NF-κB in LPS-stimulated zebrafish (P<0.01).Ethyl lithospermate exerts anti-Inflammatory effected by inhibiting the NF-κB and STAT3 signal pathways in RAW 264.7 macrophages and zebrafish.© 2023. The Chinese Journal of Integrated Traditional and Western Medicine Press and Springer-Verlag GmbH Germany, part of Springer Nature.