GM-CSF Down-Regulates TLR Expression via the Transcription Factor PU.1 in Human Monocytes

Toll-like receptors (TLR) are crucial sensors of microbial agents such as bacterial or viral compounds. These receptors constitute key players in the induction of inflammation, e.g. in septic or chronic inflammatory diseases. Colony-stimulating factors (CSFs) such as granulocyte-macrophage-CSF (GM-CSF) or granulocyte-CSF (G-CSF) have been extensively investigated in their capacity to promote myelopoiesis in febrile neutropenia or to overcome immunosuppression in patients suffering from sepsis-associated neutropenia or from monocytic immunoincompetence. We report here that GM-CSF, downregulates TLR1, TLR2 and TLR4 in a time- and dose-dependent fashion in human monocytes. Diminished pathogen recognition receptor expression was accompanied by reduced downstream p38 and extracellular-signal-regulated kinase (ERK) signaling upon lipoteichoic acid (LTA) and lipopolysaccharide (LPS) binding—and accordingly led to impaired proinflammatory cytokine production. Knockdown experiments of the transcription factors PU.1 and VentX showed that GM-CSF driven effects on TLR regulation is entirely PU.1 but not VentX dependent. We further analysed monocyte TLR and CD14 expression upon exposure to the IMID® immunomodulatory drug Pomalidomide (CC-4047), a Thalidomide analogue known to downregulate PU.1. Indeed, Pomalidomide in part reversed the GM-CSF-mediated effects. Our data indicate a critical role of PU.1 in the regulation of TLR1, 2, 4 and of CD14, thus targeting PU.1 ultimately results in TLR modulation. The PU.1 mediated immunomodulatory properties of GM-CSF should be taken into consideration upon usage of GM-CSF in inflammatory or infection-related conditions.

Toll样受体（Toll-like receptors, TLR）是识别细菌或病毒组分等微生物病原体的关键感受器。此类受体在炎症诱导过程中发挥核心作用，例如在脓毒症或慢性炎症疾病中。集落刺激因子（Colony-stimulating factors, CSFs）如粒细胞-巨噬细胞集落刺激因子（granulocyte-macrophage-CSF, GM-CSF）与粒细胞集落刺激因子（granulocyte-CSF, G-CSF）已被广泛研究，其可在发热性中性粒细胞减少症患者中促进髓系造血，或可改善脓毒症相关中性粒细胞减少症及单核细胞免疫功能低下患者的免疫抑制状态。本研究报道，粒细胞-巨噬细胞集落刺激因子可在人单核细胞中以时间和剂量依赖的方式下调Toll样受体1（TLR1）、Toll样受体2（TLR2）与Toll样受体4（TLR4）的表达。病原识别受体表达的降低，伴随脂磷壁酸（lipoteichoic acid, LTA）与脂多糖（lipopolysaccharide, LPS）结合后下游p38及细胞外调节蛋白激酶（extracellular-signal-regulated kinase, ERK）信号通路活性的减弱，进而导致促炎细胞因子生成受损。针对转录因子PU.1与VentX的敲低实验显示，粒细胞-巨噬细胞集落刺激因子对Toll样受体调控的作用完全依赖于PU.1，而非VentX。本研究进一步分析了人单核细胞在暴露于免疫调节药物（IMID®）泊马度胺（Pomalidomide, CC-4047）——一种已知可下调PU.1表达的沙利度胺类似物——后的Toll样受体与CD14表达情况。实验结果证实，泊马度胺可部分逆转粒细胞-巨噬细胞集落刺激因子介导的上述效应。本研究数据表明，PU.1在调控Toll样受体1、2、4及CD14的表达中发挥关键作用，因此靶向PU.1可最终实现对Toll样受体的调控。在炎症或感染相关疾病中使用粒细胞-巨噬细胞集落刺激因子时，应考虑到PU.1介导的粒细胞-巨噬细胞集落刺激因子的免疫调节特性。