Zimmermann Katrin, van Phi Valerie D, Brase Angela, Phi-van Loc
Friedrich-Loeffler-Institut, Institute of Animal Welfare and Animal Husbandry, Celle, Germany.
Institute of Radiology, Klinikum rechts der Isar, Technische Universität München, München, Germany.
Innate Immun. 2015 May;21(4):406-15. doi: 10.1177/1753425914547434. Epub 2014 Sep 11.
Serotonin (5-hydroxytryptamine; 5-HT) transporter (5-HTT) is involved in inflammation and the stress response. In this study, we examined the regulation of 5-HTT expression in macrophage HD11 cells in response to bacterial LPS. Long-term exposure of cells to LPS (6-18 h) produced a decrease in 5-HTT mRNA expression. Accordingly, reduced 5-HTT activity measured by 5-HT uptake was also observed in LPS-treated HD11 cells. Moreover, LPS treatment, as well as co-transfection with an expression vector encoding the chicken CCAAT/enhancer binding protein beta (C/EBPβ), resulted in inhibition of 5-HTT promoter activity. Indeed, sequence analysis revealed several C/EBPβ binding motifs in the upstream region of the 5-HTT gene, which specifically interacted with C/EBPβ both in an in vitro band shift assay and in living HD11 cells. The C/EBPβ binding was activated in cells treated with LPS. The role of C/EBPβ in LPS inhibition of 5-HTT expression was further confirmed by small interfering RNA interference, which demonstrated that knockdown of endogenous C/EBPβ attenuated the inhibition of 5-HTT expression in LPS-treated cells. Taken together, the results suggest that C/EBPβ plays a critical role in regulating the 5-HTT gene in macrophages in response to pro-inflammatory stimuli.
血清素（5-羟色胺；5-HT）转运体（5-HTT）参与炎症反应和应激反应。在本研究中，我们检测了巨噬细胞HD11细胞中5-HTT表达对细菌脂多糖（LPS）的反应调控。细胞长期暴露于LPS（6 - 18小时）会导致5-HTT mRNA表达下降。相应地，在LPS处理的HD11细胞中也观察到通过5-HT摄取测量的5-HTT活性降低。此外，LPS处理以及与编码鸡CCAAT/增强子结合蛋白β（C/EBPβ）的表达载体共转染，均导致5-HTT启动子活性受到抑制。实际上，序列分析揭示了5-HTT基因上游区域存在几个C/EBPβ结合基序，它们在体外凝胶迁移试验和活的HD11细胞中均与C/EBPβ特异性相互作用。在LPS处理的细胞中，C/EBPβ结合被激活。小干扰RNA干扰进一步证实了C/EBPβ在LPS抑制5-HTT表达中的作用，这表明敲低内源性C/EBPβ可减弱LPS处理细胞中5-HTT表达的抑制。综上所述，结果表明C/EBPβ在巨噬细胞中响应促炎刺激调控5-HTT基因方面发挥关键作用。
