Purpose: Pathogens consist of a wide variety of evolutionarily conserved molecular structures that are recognized by pattern recognition receptors (PRRs) of innate immunity. Reasonably assuming that no single PRR is ever likely to be the sole trigger of the immune response during infection, a great deal remains unknown about collaborative mechanisms and consequential crosstalk effects between multiple PRRs belonging to different families. Here, we aimed to investigate inflammatory response to combined stimulation of cytosolic nucleotide-binding oligomerization domain (NOD) receptors: NOD1, NOD2 and membrane-bound C-type lectin receptors (CLRs): Mincle and Dectin-1 in comparison to individual stimulation both in vitro and in vivo.
Materials and Methods: For in vitro studies, we used human monocytic THP-1 cells endogenously expressing NOD1,2, as well as Mincle and Dectin-1 receptors. Using reporter gene and immunoassay approaches, we measured activity of key proinflammatory transcription factors (NF-κB and AP-1) and cytokine production after addition of specific PRR agonists or their pairwise combinations. In vivo NF-κB activity (bioluminescent detection in NF-κB-Luc transgenic mice), as well as cytokine levels in mouse blood serum, was measured 3 hours after intramuscular injection of PRR agonists.
Results: We detected that combined stimulation of NOD1/2 and C-type lectin receptors (Dectin-1, Mincle) strongly potentiates NF-κB and AP-1 transcription factor activity in human monocytic THP-1 cells, as well as resulting in enhanced levels of IL-8 cytokine production. We demonstrated that RIP2- and Syk-dependent signaling pathways downstream of NOD1/2 and Dectin-1/Mincle, respectively, are essential for the potentiated proinflammatory cell response. Lastly, we confirmed that synergy between NOD and C-type lectin receptors resulting in potentiated levels of NF-κB activation and cytokine (IL-6, KC) production also occurs in vivo.
Conclusion: These findings originally indicate cooperation between NODs and CLRs, leading to potentiated levels of proinflammatory immune response both in vitro and in vivo.

Keywords: pattern recognition receptors, innate immunity, synergy, collaboration


中文翻译：

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NOD1/2 和 C 型凝集素受体 Dectin-1 和 Mincle 在体外和体内协同增强促炎反应。

目的：病原体由多种进化上保守的分子结构组成，这些分子结构被先天免疫的模式识别受体 (PRR) 识别。合理地假设没有单一的 PRR 可能是感染期间免疫反应的唯一触发因素，但对于属于不同家族的多个 PRR 之间的协作机制和随之而来的串扰效应，仍有很多未知之处。在这里，我们旨在研究对细胞溶质核苷酸结合寡聚化结构域 (NOD) 受体的联合刺激的炎症反应：NOD1、NOD2 和膜结合 C 型凝集素受体 (CLR)：Mincle 和 Dectin-1，与单独的刺激相比体外和体内。
材料和方法：对于体外研究，我们使用了内源性表达 NOD1、2 以及 Mincle 和 Dectin-1 受体的人类单核细胞 THP-1 细胞。使用报告基因和免疫测定方法，我们测量了添加特定 PRR 激动剂或其成对组合后关键促炎转录因子（NF-κB 和 AP-1）的活性和细胞因子的产生。在肌肉内注射 PRR 激动剂 3 小时后，测量体内 NF-κB 活性（在 NF-κB-Luc 转基因小鼠中的生物发光检测）以及小鼠血清中的细胞因子水平。
结果：我们检测到 NOD1/2 和 C 型凝集素受体 (Dectin-1, Mincle) 的联合刺激可强烈增强人单核细胞 THP-1 细胞中 NF-κB 和 AP-1 转录因子的活性，并导致增强的IL-8 细胞因子的产生。我们证明了分别位于 NOD1/2 和 Dectin-1/Mincle 下游的 RIP2 和 Syk 依赖性信号通路对于增强的促炎细胞反应至关重要。最后，我们证实 NOD 和 C 型凝集素受体之间的协同作用导致 NF-κB 活化和细胞因子（IL-6，KC）产生水平的增强也在体内发生。
结论：这些发现最初表明 NOD 和 CLR 之间的合作，导致体外和体内促炎免疫反应水平增强。

关键词：模式识别受体，先天免疫，协同作用，协作