Macrophage-intrinsic regulators of tissue type 2 inflammation

巨噬细胞在2型免疫应答过程中发挥多种调节和效应功能，但控制这种功能可塑性的因素定义不清。巨噬细胞的转录谱由转录因子和表观遗传机制/修饰剂形成，其允许快速反应和-在某些情况下-稳定的重编程和“先天记忆”。酶转氨酶-2（TG 2）和转录因子芳烃受体（AhR）已经涉及2型免疫应答的调节，但是它们在“2型”巨噬细胞活化和功能中的作用仍然不清楚。我们已经观察到，来自屋尘螨（HDM）过敏性哮喘患者的血液单核细胞或来自HDM致敏小鼠的骨髓细胞的巨噬细胞在体外培养7天后显示出增加的炎症基因表达和反应性。引人注目的是，HDM致敏小鼠的骨髓祖细胞显示出与来自幼稚小鼠的骨髓祖细胞相比TG 2的表达增加。这表明外周巨噬细胞祖细胞在2型炎症过程中稳定重编程，TG 2可能在此过程中发挥作用。鉴于最近报道的作用，TG 2组蛋白（H3 K4 me 3Q 5）的乙酰化和转录激活（神经元），我们的目标是调查是否髓/造血TG 2缺陷导致改变基因表达在2型激活的巨噬细胞在体内蠕虫感染或过敏性气道炎症（AAI）HDM触发。我们还旨在确定来自哮喘患者和来自HDM致敏或蠕虫感染小鼠的巨噬细胞中基因表达的差异是否与H3 K4 me 3Q 5和其他组蛋白标记和/或染色质可及性相关。此外，我们发现，缺乏AhR或其直接下游靶酶细胞色素P450 CYP 1B 1导致更严重的AAI在两个过敏性哮喘的临床前模型。尽管CYP 1B 1主要通过其在鼠和人肺中的非造血（上皮）细胞中的表达来调节AAI，但AhR在刺激时调节上皮和骨髓细胞中的基因表达。引人注目的是，在用屋尘螨或LPS刺激时，缺乏AhR的肺泡样巨噬细胞不能上调抗炎性前列腺素（PGE 2）和白三烯（LTB 4、LTC 4、LTD 4、LTE 4）。因此，我们现在的目标是在全球范围内的AhR在调节巨噬细胞内的脂质介质合成的作用，并验证这种功能的AAI和蠕虫感染的设置在体内/体外。这两个目标都将通过下一代测序方法（RNAseq，ChIPseq，ATACseq）来解决，这些方法已经在几个FOR 2599项目的框架内针对巨噬细胞进行了优化。总之，我们将确定AhR和TG 2作为2型免疫中巨噬细胞活化和重编程的关键调节因子的作用。

Macrophages fulfil a multitude of regulatory and effector functions during type 2 immune responses, but the factors that govern this functional plasticity are poorly defined. Transcriptional profiles of macrophages are shaped by transcription factors and epigenetic mechanisms/ modifiers that allow for rapid responses and – in some contexts - stable reprogramming and “innate memory”. The enzyme transglutaminase-2 (TG2) and the transcription factor aryl hydrocarbon receptor (AhR) have been implicated in the regulation of type 2 immune responses, but their role in “type 2” macrophage activation and function has remained obscure. We have observed that macrophages derived from blood monocytes of house dust mite (HDM) allergic asthma patients or from bone marrow cells of HDM-sensitized mice show increased inflammatory gene expression and responsiveness after 7 days of in vitro culture. Strikingly, bone marrow progenitors of HDM-sensitized mice showed an increased expression of TG2 compared to progenitors from the bone marrow of naïve mice. This suggested that peripheral macrophage progenitors are stably reprogrammed during type 2 inflammation and that TG2 may play a role in this process. Given the recently reported role of TG2 in histone (H3K4me3Q5) serotonylation and transcriptional activation (in neurons), we aim to investigate whether a myeloid/ hematopoietic TG2 deficiency results in altered gene expression in type 2 activated macrophages in vivo during helminth infection or allergic airway inflammation (AAI) triggered by HDM. We also aim to determine whether differences in gene expression in macrophages from asthma patients and from HDM-sensitized or helminth infected mice correlate with H3K4me3Q5 and other histone marks and/ or chromatin accessibility. Moreover, we have found that lack of the AhR or its immediate downstream target enzyme cytochrome P450 CYP1B1 leads to more severe AAI in two preclinical models of allergic asthma. Whereas CYP1B1 regulates AAI primarily through its expression in non-hematopoietic (epithelial) cells in both murine and human lung the AhR regulates gene expression in epithelial and myeloid cells upon stimulation. Strikingly, alveolar-like macrophages deficient for AhR fail to upregulate anti-inflammatory prostaglandins (PGE2) and leukotrienes (LTB4, LTC4, LTD4, LTE4) upon stimulation with house dust mite or LPS. Therefore, we now aim to globally profile the role of the AhR in the regulation of lipid mediator synthesis within macrophages and validate this function in settings of AAI and helminth infection in vivo/ex vivo. Both of these objectives will be addressed by next generation sequencing approaches (RNAseq, ChIPseq, ATACseq) that have been optimized for macrophages within the frame of several FOR2599 projects. In summary, we will determine the role of AhR and TG2 as key regulator of macrophage activation and reprogramming in type 2 immunity.