Regulation of type 2 immune responses via neuropeptides and neurotransmitters

先天性淋巴样细胞（ILC）是抵抗感染的第一道防线，并协调组织免疫状态和稳态。ILC 2参与2型免疫反应，包括蠕虫感染和慢性过敏反应。它们响应于广泛的环境刺激产生2型细胞因子（主要是IL-5和IL-13），例如alarmins（IL-33、IL-25、TSLP）和神经调质（NE、Ach、CGRP、NMU）。这些信号中的每一个及其组合独特地控制ILC 2的活性。因此，这些细胞在不同的环境和组织中采用异源表型。血管活性肠肽（VIP）是昼夜节律系统的重要调节剂，是一种神经肽，已被提议作为ILC 2的另一种调节剂。已知VIP释放受进食影响，ILC 2以昼夜节律方式产生IL-5。然而，与其他神经递质不同，VIP介导的ILC 2调节的确切作用、位置和生理相关性仍未完全阐明。此外，缺乏有效和特异性的遗传工具来干扰ILC 2中的VIP信号传导，以及相互矛盾的研究限制了我们对这种潜在的神经免疫回路的理解。在这里，我们建议使用我们的新小鼠模型，该模型允许在ILC 2中产生Vipr 2条件性敲除，以及其他现有的小鼠模型，以研究体内VIP神经元（例如，INTACT，DREADD系统），以及经典（例如，流式细胞术，细胞因子测定），和创新的基因组学方法（例如，单核RNAseq和空间RNAseq）来功能性地表征VIPergic神经元和ILC 2的相互作用。我们将首先确定ILC 2中的VIP信号传导是否参与其异质性、细胞因子产生和随后的嗜酸性粒细胞向组织的募集。我们将使用呼吸和口腔过敏模型进一步确定在2型免疫应答期间是否需要这种神经免疫回路。然后，我们将确定该回路是否驱动昼夜ILC 2活性，包括IL-5的产生和随后的嗜酸性粒细胞的组织动力学。我们将进一步评估是否昼夜肠道吸收，运动和绒毛结构，肠道嗜酸性粒细胞的影响，依赖于VIP-ILC 2轴。最后，我们将解剖VIP神经元的空间和时间的活动，以及它们在稳态和食物过敏期间在小肠中与ILC 2的接近程度。总而言之，这项研究将有助于破译涉及ILC 2的额外神经免疫回路的生理功能，并揭示VIP在ILC 2活性的昼夜节律调节及其对嗜酸性粒细胞稳态和健康和疾病组织功能的影响中的作用。

Innate lymphoid cells (ILC) are the first line of defense against infections and orchestrate tissue immune status and homeostasis. ILC2s are involved in type 2 immune responses, including helminth infections and chronic allergic reactions. They produce type 2 cytokines (mainly IL-5 and IL-13) in response to a wide range of environmental stimuli, such as alarmins (IL-33, IL-25, TSLP) and neuromodulators (NE, Ach, CGRP, NMU). Each of these signals and their combination uniquely controls ILC2s' activity. Hence, these cells adopt a heterogenic phenotype in different contexts and tissues. The vasoactive intestinal peptide (VIP), an essential regulator of the circadian system, is a neuropeptide that has been proposed as another regulator of ILC2s. VIP release is known to be influenced by feeding, and ILC2s produce IL-5 in a circadian fashion. However, unlike other neurotransmitters, the precise action, location, and physiological relevance of VIP-mediated regulation of ILC2s are still not fully elucidated. Besides, the lack of efficient and specific genetic tools to interfere with VIP signaling in ILC2s and conflicting studies limit our understanding of this potential neuro-immune circuit. Here, we propose to use our new mouse model that allows the generation of Vipr2 conditional knockout in ILC2s, as well as other existing mouse models, to study VIP neurons in vivo (e.g., INTACT, DREADD systems ), together with a combination of classical (e.g., flow cytometry, cytokines assays), and innovative genomics approaches (e.g., single nuclei RNAseq, and spatial RNAseq) to functionally characterize the interplay of VIPergic neurons and ILC2s. We will first determine whether VIP signaling in ILC2s participates in their heterogeneity, cytokine production, and subsequent eosinophil recruitment to tissues. We will further determine whether this neuro-immune circuit is required during the type 2 immune response using respiratory and oral allergy models. Then we will determine whether this circuit drives circadian ILC2 activity, including IL-5 production and subsequent eosinophils' tissue dynamic. We will further evaluate whether circadian intestinal absorption, motility, and villi architecture, influenced by intestinal eosinophils, rely on the VIP-ILC2 axis. Finally, we will dissect VIP neurons' spatial and temporal activity and their proximity to ILC2s in the small intestine both at steady-state and during food allergy. Altogether, this study will help decipher the physiological functions of an additional neuro-immune circuit involving ILC2s and shed light on a yet elusive role of VIP in the circadian regulation of ILC2s' activity and its impact on eosinophil homeostasis and tissue functions in health and disease.