Csóka Balázs, Németh Zoltán H, Virág László, Gergely Pál, Leibovich S Joseph, Pacher Pál, Sun Chun-Xiao, Blackburn Michael R, Vizi E Sylvester, Deitch Edwin A, Haskó György
Department of Surgery, University of Medicine and Dentistry of New Jersey, New Jersey Medical School, Newark, NJ 07103, USA.
Blood. 2007 Oct 1;110(7):2685-95. doi: 10.1182/blood-2007-01-065870. Epub 2007 May 24.
We recently showed that A(2A) adenosine receptor activation by endogenous adenosine contributes to interleukin-10 (IL-10) production in polymicrobial sepsis. Here we investigated the molecular mechanisms underpinning this interaction between adenosine receptor signaling and infection by exposing macrophages to Escherichia coli. We demonstrated using receptor knockout mice that A(2A) receptor activation is critically required for the stimulatory effect of adenosine on IL-10 production by E coli-challenged macrophages, whereas A(2B) receptors have a minor role. The stimulatory effect of adenosine on E coli-induced IL-10 production did not require toll-like receptor 4 (TLR4) or MyD88, but was blocked by p38 inhibition. Using shRNA we demonstrated that TRAF6 impairs the potentiating effect of adenosine. Measuring IL-10 mRNA abundance and transfection with an IL-10 promoter-luciferase construct indicated that E coli and adenosine synergistically activate IL-10 transcription. Sequential deletion analysis and site-directed mutagenesis of the IL-10 promoter revealed that a region harboring C/EBP binding elements was responsible for the stimulatory effect of adenosine on E coli-induced IL-10 promoter activity. Adenosine augmented E coli-induced nuclear accumulation and DNA binding of C/EBPbeta. C/EBPbeta-deficient macrophages failed to produce IL-10 in response to adenosine and E coli. Our results suggest that the A(2A) receptor-C/EBPbeta axis is critical for IL-10 production after bacterial infection.
我们最近发现，内源性腺苷激活A(2A)腺苷受体有助于多微生物败血症中白细胞介素-10（IL-10）的产生。在此，我们通过将巨噬细胞暴露于大肠杆菌来研究腺苷受体信号传导与感染之间这种相互作用的分子机制。我们使用受体敲除小鼠证明，A(2A)受体激活对于腺苷对大肠杆菌刺激的巨噬细胞产生IL-10的刺激作用至关重要，而A(2B)受体作用较小。腺苷对大肠杆菌诱导的IL-10产生的刺激作用不需要Toll样受体4（TLR4）或MyD88，但被p38抑制所阻断。使用短发夹RNA（shRNA）我们证明肿瘤坏死因子受体相关因子6（TRAF6）损害了腺苷的增强作用。测量IL-10 mRNA丰度并转染IL-10启动子-荧光素酶构建体表明，大肠杆菌和腺苷协同激活IL-10转录。对IL-10启动子进行序列缺失分析和定点诱变表明，含有C/EBP结合元件的区域负责腺苷对大肠杆菌诱导的IL-10启动子活性的刺激作用。腺苷增强了大肠杆菌诱导的C/EBPβ的核积累和DNA结合。C/EBPβ缺陷型巨噬细胞在对腺苷和大肠杆菌的反应中未能产生IL-10。我们的结果表明，A(2A)受体-C/EBPβ轴对于细菌感染后IL-10的产生至关重要。
