Regulation of human dendritic cell activation

Toll样受体（TLR）配体的识别导致宿主免疫的机制仍然不清楚。现在认为，为了诱导有效的免疫应答，微生物必须刺激TLR家族内外的复杂的模式识别受体。这些不同受体的联合激活可导致调节先天性和适应性免疫的互补、协同或拮抗作用。因此，完整理解TLR在宿主抗感染中的作用需要“解码”这些多受体相互作用。我们过去的工作已经揭示了不同T细胞受体的配体之间在树突状细胞活化中的显著协同作用，以及I型干扰素在调节树突状细胞和组织及肿瘤细胞中TLR下游信号传导中的作用。特别地，我们表明，TLR配体的细胞反应不仅是促炎介质的产生，而且它们还参与组织稳态的控制并调节细胞分化、增殖和凋亡。MyD 88和TRIF信号传导与I型IFN的产生之间的平衡决定了组织和肿瘤细胞中的增殖与凋亡以及树突细胞中的活化与存活。我们还发现，β-葡聚糖（酵母和真菌的一种成分）通过dectin 1受体刺激树突状细胞有助于诱导人类Th 17应答，并与TLR受体刺激协同作用，以激活树突状细胞和促炎细胞因子产生。在过去的一年中，我们发现通过dectin 1（一种在其细胞质部分具有ITAM样基序的受体）的信号传导导致仅诱导少量早期应答细胞因子，如IL-1 β、IL-6和TNF，并且仅非常适度地激活NF-κ B。然而，在这方面，β-葡聚糖刺激诱导的IL-1 β（并通过炎性小体处理，也通过dectin-1刺激激活）发挥有效的正反馈机制，这是最佳NF-κ B活化和产生迟反应细胞因子如IL-12和IL-23所必需的。1家族细胞因子在含ITAM受体的树突状细胞和其他免疫细胞的活化中的作用。特别是，我们想了解这些受体激活NF-κ B的能力有多大程度是直接通过BCL-10激活的，还是依赖于内源性IL-1提供的放大信号。我们还想了解不同的ITAM受体正或负调节树突状细胞和骨髓细胞对TLR配体的反应性的分子机制。特别是，我们希望了解Akt/mTOR通路的作用及其受I型干扰素的调节。β-葡聚糖刺激而非LPS刺激诱导人单DC分泌高水平IL-1 β的观察结果（6）使我们研究IL-1 β是否在响应β-葡聚糖的细胞因子分泌中起反馈积极作用。IL-1的产生依赖于SYK介导的NALP 3的活化，NALP 3反过来活化IL-1 β前体剪接和分泌所需的caspase-1。用IL-1 RA阻断内源性IL-1 β的作用（或用IL-1 β的抗体或通过阻断caspase-1），早期基因诱导不受影响，但几个晚期基因的表达几乎完全消失。 诱导基因可分为三大类：早期，IL-1非依赖性基因（诱导4小时，但在12小时较少）;晚期IL-1依赖性基因（诱导12小时比4小时更好）;和晚期IL-1非依赖性基因。总体而言，我们的分析确定，β-葡聚糖诱导的基因子集严格依赖于内源性IL-1来维持单DC中的表达，这些基因包括许多免疫相关基因，如IL-12、IL-23、IL-10和TNF。这些基因中的许多也被LPS诱导，但它们中的大多数仅具有瞬时动力学。向LPS中加入外源性IL-1 β可维持这些基因的表达，表明IL-1 R和TLR可类似地为这些基因的转录发出信号，但它们的长期表达需要内源性IL-1的正反馈。TNF还对所有NF-κ B依赖性基因的基因表达具有一些正反馈效应，但对IL-1依赖性基因，TNF的效应是适度的，并且在不存在IL-1的情况下不足以维持这些基因响应于β-葡聚糖或LPS的表达。两种配体均诱导I型IFN依赖性基因的早期表达，当内源性I型IFN信号传导被中和抗体阻止时，所述I型IFN依赖性基因被阻断。然而，LPS对IFN-β的诱导作用是适度的，而β-葡聚糖几乎检测不到。 这些研究清楚地揭示了炎症和稳态基因调控的新机制，这些机制可能在炎症，免疫和癌症中发挥重要作用。树突状细胞、巨噬细胞和其他骨髓细胞在对感染的适应性和先天性免疫应答的调节中以及在炎症的调节和失调中发挥作用，因为它涉及感染、退行性和自身免疫性疾病和癌症。通过这些机制调节树突细胞和骨髓细胞功能在免疫治疗（例如疫苗、癌症免疫治疗）的规划中以及在疾病治疗和癌症预防/治疗中靶向基于炎症的病理学中是重要的。

The mechanisms by which the recognition of Toll-like receptor (TLR) ligands leads to host immunity remain poorly defined. It is now thought that to induce an effective immune response, microorganisms must stimulate complex sets of pattern-recognition receptors, both within and outside of the TLR family. The combined activation of these different receptors can result in complementary, synergistic or antagonistic effects that modulate innate and adaptive immunity. Therefore, a complete understanding of the role of TLRs in host resistance to infection requires 'decoding' of these multiple receptor interactions.Our past work has uncovered the significant synergy in dendritic cell activation between ligands of different T cell receptors and the role of type I interferon in regulating TLR downstream signaling in both dendritic cells and tissue and tumor cells. In particular we showed that the cellular response to TLR ligands is not only production of pro-inflammatory mediators but they are also involved in control of tissue homeostasis and regulate cellular differentiation, proliferation, and apoptosis. The balance between MyD88 and TRIF signaling and the production of type I IFN determine proliferation versus apoptosis in tissue and tumor cells and activation versus survival in dendritic cells. We also have found that the stimulation of the dendritic cells by beta-glucan (a component of yeast and fungi) through the dectin 1 receptor facilitate the induction of an Th17 response in human and synergize with TLR receptor stimulation for activation of dendritic cells and pro-inflammatory cytokine production. In the last year we have discovered that the signaling through dectin 1, a receptor with an ITAM-like motif in its cytoplasmic portion, results in the induction of only a small number of early responding cytokine such as Il-1beta, Il-6, and TNF and in only a very modest activation of NF-kappaB. However, IL-1beta induced by beta-glucan stimulation (and processed through the inflammasome also activated through dectin-1 stimulation) exert a potent positive feedback mechanisms that it is necessary for optimal NF-kappaB activation and production of late responsive cytokines such as IL-12 and IL-23.We now plan to extend the study of the IL-1 family of cytokines in the activation of dendritic cells and other immune cells by ITAM-containing receptors. In particular we want to understand how much the ability of these receptors to activate NF-kappaB is direct through BCL10 activation or it is dependent through the amplifying signal provided by endogenous IL-1. We also want to understand the molecular mechanisms by which different ITAM-containing receptors positively or negatively regulate the responsiveness of dendritic cells and myeloid cells to TLR ligands. In particular we want to undersatnd the role of the Akt/mTOR pathway and its modulation by type I interferon.The observation that beta-glucan stimulation but not LPS stimulation induced high levels of IL-1beta secretion by human mono-DC (6) led us to investigate whether IL-1beta plays a feedback positive role in the secretion of cytokines in response to beta-glucan. IL-1 production was dependent on SYK mediated activation of NALP3 that in turn activated caspase-1 required for pro-IL-1beta splicing and secretion. Blocking the effect of endogenous IL-1beta with IL-1RA (or with antibodies to IL-1beta or by blocking caspase-1) the early gene induction was not affected but the expression of several of the late genes was almost completely abolished. Induced genes could be divided into three major groups; early, IL-1-independent genes (induced well at 4 h but less so at 12 h); late IL-1-dependent genes (induced better at 12 than at 4 h); and late IL-1-independent genes. Overall our analysis established that a subset of genes induced by beta-glucan is strictly dependent on endogenous IL-1 for maintained expression in mono-DC and these genes include many immunologically relevant genes such as IL-12, IL-23, IL-10, and TNF. Many of these genes are also induced by LPS but most of them with only a transient kinetics. Addition of exogenous IL-1beta to LPS maintains the expression of these genes, suggesting that the IL-1R and TLR can similarly signal for the transcription of these genes but that their long term expression requires the positive feedback of endogenous IL-1. TNF also has some positive feedback effects on gene expression of all NF-kB dependent genes but on the IL-1-dependent gene the effect of TNF is modest and insufficient in the absence of IL-1 to maintain the expression of these genes in response to either beta-glucan or LPS. Both ligands induce the early expression of type I IFN dependent genes that it is blocked when endogenous type I IFN signaling is prevented by neutralizing antibodies. However, the induction of IFN-beta is modest with LPS and almost undetectable with beta-glucan. These studies are clearly unveiling new mechanisms of inflammatory and homeostatic gene regulation that are likely to play an important role in inflammation, immunity, and cancer. Dendritic cells, macrophages, and other myeloid cells play a role both in the regulation of the adaptive and innate immune response to infection as well as in the regulation and dysregulation of inflammation as it relates to infections, degenerative and autoimmune diseases and cancer. Regulating through these mechanisms dendritic cell and myeloid cell functions is important in the planning of immunotherapy (e.g. vaccine, cancer immunotherapy) as well in targeting inflammation-based pathology in disease therapy and cancer prevention/treatment.