Contribution of MINCLE-SYK Signaling to Activation of Primary Human APCs by Mycobacterial Cord Factor and the Novel Adjuvant TDB
Contribution of MINCLE-SYK Signaling to Activation of Primary Human APCs by Mycobacterial Cord Factor and the Novel Adjuvant TDB
复制标题
DOI：
10.4049/jimmunol.1500102
复制
发表时间：
2015-09-01
影响因子：
4.4
通讯作者：
Lang, Roland
中科院分区：
文献类型：
--
作者：
Ostrop, Jenny;Jozefowski, Katrin;Lang, Roland
海藻糖-6，6-二霉菌酸酯（TDM）是分枝杆菌索状因子，是一种丰富的细胞壁糖脂，也是结核分枝杆菌的主要毒力因子。其合成类似物海藻糖-6，6-二山嵛酸酯（TDB）是一种新的佐剂，目前正在进行I期临床试验。在啮齿动物中，C型凝集素受体Mincle和Mcl结合TDB/TDM并通过Syk-Card 9途径激活巨噬细胞和树突状细胞（DC）。然而，目前还不清楚这些糖脂是否通过相同的机制激活人类先天免疫细胞。我们进行了体外分析TDB/TDM刺激的原代人单核细胞，巨噬细胞和DC;确定C型凝集素受体的表达;并测试了SYK，MINCLE和MCL的小干扰RNA敲低和遗传互补的贡献。我们观察到响应于TDB或TDM的稳健的趋化因子和细胞因子释放。MCSF驱动的巨噬细胞在TDM刺激后分泌更高水平的IL-8、IL-6、CCL 3、CCL 4和CCL 2，而DC对TDB的反应更强烈，GM-CSF驱动的巨噬细胞对TDB和TDM的反应相同。SYK激酶和衔接蛋白CARD 9对于糖脂诱导的IL-8产生至关重要。单核细胞和巨噬细胞中MINCLE和MCL的mRNA表达较高，静息条件下MINCLE和MCL蛋白定位于细胞内。小干扰RNA介导的MINCLE或MCL敲低平均导致TDB或TDM诱导的IL-8产生减少。相反，逆转录病毒在鼠Mincle缺陷型DC中的表达表明，人MINCLE而不是MCL足以赋予对TDB/TDM的响应性。我们的研究表明，SYK-CARD 9信号转导在TDB/TDM诱导的人和小鼠先天免疫细胞活化中起关键作用，可能是通过MINCLE的参与。
Trehalose-6,6-dimycolate (TDM), the mycobacterial cord factor, is an abundant cell wall glycolipid and major virulence factor of Mycobacterium tuberculosis. Its synthetic analog trehalose-6,6-dibehenate (TDB) is a new adjuvant currently in phase I clinical trials. In rodents, the C-type lectin receptors Mincle and Mcl bind TDB/TDM and activate macrophages and dendritic cells (DC) through the Syk-Card9 pathway. However, it is unknown whether these glycolipids activate human innate immune cells through the same mechanism. We performed in vitro analysis of TDB/TDM-stimulated primary human monocytes, macrophages, and DC; determined C-type lectin receptor expression; and tested the contribution of SYK, MINCLE, and MCL by small interfering RNA knockdown and genetic complementation. We observed a robust chemokine and cytokine release in response to TDB or TDM. MCSF-driven macrophages secreted higher levels of IL-8, IL-6, CCL3, CCL4, and CCL2 after stimulation with TDM, whereas DC responded more strongly to TDB and GM-CSF-driven macrophages were equally responsive to TDB and TDM. SYK kinase and the adaptor protein CARD9 were essential for glycolipid-induced IL-8 production. mRNA expression of MINCLE and MCL was high in monocytes and macrophages, with MINCLE and MCL proteins localized intracellularly under resting conditions. Small interfering RNA-mediated MINCLE or MCL knockdown caused on average reduced TDB- or TDM-induced IL-8 production. Conversely, retroviral expression in murine Mincle-deficient DC revealed that human MINCLE, but not MCL, was sufficient to confer responsiveness to TDB/TDM. Our study demonstrates that SYK-CARD9 signaling plays a key role in TDB/TDM-induced activation of innate immune cells in man as in mouse, likely by engagement of MINCLE.