Calcitonin gene related peptide negatively regulates alarmin-driven type 2 innate lymphoid cell responses. Calcitonin gene related peptide negatively regulates alarmin-driven type 2 innate lymphoid cell responses

Neuroimmune interactions have emerged as critical modulators of allergic inflammation, and type 2 innate lymphoid cells (ILC2s) are an important cell type for mediating these interactions. Here, we show that ILC2s expressed both the neuropeptide CGRP (Calcitonin Gene-Related Peptide) and its receptor. CGRP potently inhibited alarmin-driven type 2 cytokine production and proliferation by lung ILC2s both in vitro and in vivo. CGRP induced marked changes in ILC2 expression programs in vivo and in vitro, attenuating alarmin-driven proliferative and effector responses. A distinct subset of ILCs scored highly for a CGRP-specific gene signature after in vivo alarmin stimulation, suggesting CGRP regulated this response. Finally, we observed increased ILC2 proliferation and type 2 cytokine production and exaggerated responses to alarmins in mice lacking the CGRP receptor. Together, these data indicate that endogenous CGRP is a critical negative regulator of ILC2 responses in vivo. Overall design: For in vitro bulk RNA-seq experiments, there are 32 samples total, representing 2 biological replicates, each split into 2 technical replicates, of 4 conditions at 2 time points. For in vivo scRNA-seq experiments, there are 8 samples total, representing 2 biological replicates of 4 conditions at 1 time point. For in vitro ATAC-seq experiments, there are 6 samples total, representing 3 replicates of 2 conditions at 1 time point.

神经免疫相互作用已成为过敏性炎症的关键调控因子，而2型先天淋巴细胞（type 2 innate lymphoid cells，ILC2s）是介导此类相互作用的重要细胞类群。本研究发现，ILC2s可同时表达神经肽CGRP（Calcitonin Gene-Related Peptide）及其受体。CGRP可在体外及体内环境中强效抑制肺脏ILC2s由警报素介导的2型细胞因子产生与增殖过程。CGRP可在体内及体外环境中显著改变ILC2s的基因表达程序，削弱警报素介导的增殖与效应应答。在体内经警报素刺激后，一类独特的ILC亚群的CGRP特异性基因特征富集度较高，这提示CGRP可调控该应答过程。最后，我们在缺失CGRP受体的小鼠体内观察到ILC2s增殖与2型细胞因子产生增强，且对警报素的应答更为亢进。综上，上述实验数据表明，内源性CGRP是体内ILC2s应答过程的关键负调控因子。实验整体设计：体外批量RNA测序（bulk RNA-seq）实验共包含32份样本，涵盖4种实验条件、2个时间点，每个条件设置2次生物学重复，且每个生物学重复下设2次技术重复。体内单细胞RNA测序（scRNA-seq）实验共包含8份样本，涵盖4种实验条件、1个时间点，每个条件设置2次生物学重复。体外ATAC测序（ATAC-seq）实验共包含6份样本，涵盖2种实验条件、1个时间点，每个条件设置3次重复。