GlaxoSmithKline Research Centre Zagreb Limited, Zagreb, Croatia.
Br J Pharmacol. 2012 Mar;165(5):1348-60. doi: 10.1111/j.1476-5381.2011.01576.x.
Azithromycin has been reported to modify activation of macrophages towards the M2 phenotype. Here, we have sought to identify the mechanisms underlying this modulatory effect of azithromycin on human monocytes, classically activated in vitro.
Human blood monocytes were primed with IFN-γ for 24 h and activated with LPS for 24 h. Azithromycin, anti-inflammatory and lysosome-affecting agents were added 2 h before IFN-γ. Cytokine and chemokine expression was determined by quantitative PCR and protein release by ELISA. Signalling molecules were determined by Western blotting and transcription factor activation quantified with a DNA-binding ELISA kit.
Azithromycin (1.5-50 µM) dose-dependently inhibited gene expression and/or release of M1 macrophage markers (CCR7, CXCL 11 and IL-12p70), but enhanced CCL2, without altering TNF-α or IL-6. Azithromycin also enhanced the gene expression and/or release of M2 macrophage markers (IL-10 and CCL18), and the pan-monocyte marker CD163, but inhibited that of CCL22. The Toll-like receptor (TLR) 4 signalling pathway was modulated, down-regulating NF-κB and STAT1 transcription factors. The inhibitory profile of azithromycin differed from that of dexamethasone, the phosphodiesterase-4 inhibitor roflumilast and the p38 kinase inhibitor SB203580 but was similar to that of the lysosomotropic drug chloroquine. Effects of concanamycin and NH4Cl, which also act on lysosomes, differed significantly.
Azithromycin modulated classical activation of human monocytes by inhibition of TLR4-mediated signalling and possible effects on lysosomal function, and generated a mediator expression profile that differs from that of monocyte/macrophage phenotypes so far described.
阿奇霉素已被报道可调节巨噬细胞向 M2 表型的激活。在此，我们试图确定阿奇霉素对体外经典激活的人单核细胞的这种调节作用的机制。
用 IFN-γ 对人血单核细胞进行预刺激 24 小时，然后用 LPS 激活 24 小时。在 IFN-γ 前 2 小时加入阿奇霉素、抗炎和溶酶体影响剂。通过定量 PCR 测定细胞因子和趋化因子的表达，通过 ELISA 测定蛋白释放。通过 Western 印迹测定信号分子，通过 DNA 结合 ELISA 试剂盒定量测定转录因子的激活。
阿奇霉素（1.5-50 μM）剂量依赖性地抑制 M1 巨噬细胞标志物（CCR7、CXCL11 和 IL-12p70）的基因表达和/或释放，但增强 CCL2，而不改变 TNF-α 或 IL-6。阿奇霉素还增强了 M2 巨噬细胞标志物（IL-10 和 CCL18）和泛单核细胞标志物 CD163 的基因表达和/或释放，但抑制了 CCL22 的表达。Toll 样受体（TLR）4 信号通路被调节，下调 NF-κB 和 STAT1 转录因子。阿奇霉素的抑制谱与地塞米松、磷酸二酯酶-4 抑制剂罗氟司特和 p38 激酶抑制剂 SB203580 不同，但与溶酶体药物氯喹相似。同样作用于溶酶体的康纳霉素和 NH4Cl 的作用差异显著。
阿奇霉素通过抑制 TLR4 介导的信号转导和可能对溶酶体功能的影响，调节了人单核细胞的经典激活，并产生了与迄今为止描述的单核细胞/巨噬细胞表型不同的介质表达谱。
