Finding a role for PNECs in asthma Pulmonary neuroendocrine cells (PNECs) are a rare cell type located in airway and alveolar epithelia and are often in contact with sensory nerve fibers. They have a wide phylogenic distribution and are found even in the relatively primitive lungs of amphibia and reptiles, suggesting a critical function. Sui et al. found that mice lacking PNECs have suppressed type 2 (allergic) immune responses. PNECs were observed in close proximity to group 2 innate lymphoid cells (ILC2s) around airway branch points. The PNECs enhanced ILC2 activity by secreting CGRP (calcitonin gene-related peptide). They also induced goblet-cell hyperplasia via the neurotransmitter GABA (γ-aminobutyric acid). Interestingly, human asthma patients were found to have increased PNEC numbers, suggesting a potential therapeutic target for the treatment of asthma. Science, this issue p. eaan8546 PNECs, a rare population of cells in the airways, are critical for amplifying the airway allergen signal into mucosal responses in the lungs. INTRODUCTION The lung, with its vast surface area, senses and responds to signals in inhaled air. Aberrant interactions between the lung and the environment underlie many diseases, including asthma. In vitro data show that pulmonary neuroendocrine cells (PNECs), a rare airway epithelial cell population, can act as chemosensors. Once stimulated in culture, they release dense core vesicles rich in neuropeptides, amines, and neurotransmitters. These bioactive molecules are capable of eliciting immune and physiological responses. A recent in vivo study by our group revealed that the proper development of PNECs into self-clustering units called neuroepithelial bodies is essential for restricting the number of immune cells in the naïve lung. However, whether PNECs can function in vivo to translate exogenous airway signals such as allergens into the cascade of downstream responses is unknown. RATIONALE To test the hypothesis that PNECs act as sensors in the lung, we generated mouse mutants that lack PNECs by inactivating Ascl1 in the airway epithelium—i.e., mutants that were depleted of PNECs starting at development. We exposed these mutants to either ovalbumin or house dust mites, following regimes of existing asthma models. We determined whether the mutants showed different asthmatic responses than controls. We elucidated the underlying mechanisms by identifying molecular effectors and cellular targets of PNECs. To complement the functional tests in mice, we investigated whether human asthma patients showed pathological changes in their PNECs. RESULTS Although normal at baseline, Ascl1-mutant mice exhibited severely reduced goblet cell hyperplasia and immune cell numbers compared with controls after allergen challenge. In investigating possible molecular effectors, we found that several PNEC products were decreased in mutants relative to controls after allergen challenge, including calcitonin gene-related peptide (CGRP) and γ-aminobutyric acid (GABA). In exploring possible cellular targets, we found that innate lymphoid group 2 cells (ILC2s) were enriched at airway branch points, similar to PNECs. The PNEC product CGRP stimulated ILC2 production of interleukin-5 in culture. Conversely, inactivation of the CGRP receptor gene Calcrl in ILC2s led to dampened immune responses to allergens. In contrast to CGRP, GABA did not increase ILC2 cytokine secretion. Rather, inactivation of GABA biogenesis led to defective goblet cell hyperplasia after allergen challenge, suggesting that GABA is required for this response in the airway epithelium. The instillation of a mixture of CGRP and GABA in Ascl1 mutants restored both immune cell increases and goblet cell hyperplasia after allergen challenge, indicating that these products are the primary molecular effectors of PNECs in vivo. Consistent with these results from mice, we found increased PNEC numbers and cluster sizes in human asthma patients, which may underlie the heightened response to allergens in these individuals. CONCLUSION Our results demonstrate that PNECs, despite being a rare population of cells in the airway, are critical for amplifying the airway allergen signal into mucosal type 2 responses. Specifically, PNECs act through their product GABA to stimulate airway epithelial mucus production. In parallel, PNECs act through another product, CGRP, to stimulate ILC2 production of cytokines, which in turn recruit downstream immune cells. PNECs and ILC2s form neuroimmunological modules at the airway branch points, which are also the sites where airway particles are enriched. Our findings indicate that the PNEC-ILC2 axis functions to sense inhaled inputs, such as allergens, and amplify them into lung outputs, such as the allergic asthma response. PNECs are preferentially localized at branch points. A mouse airway stained by antibody against CGRP, to label PNECs (magenta) and antibody against SCGB1A1 to label club cells (green) (200× magnification). PNECs often cluster into neuroepithelial bodies and are preferentially localized at branch points. Pulmonary neuroendocrine cells (PNECs) are rare airway epithelial cells whose function is poorly understood. Here we show that Ascl1-mutant mice that have no PNECs exhibit severely blunted mucosal type 2 response in models of allergic asthma. PNECs reside in close proximity to group 2 innate lymphoid cells (ILC2s) near airway branch points. PNECs act through calcitonin gene-related peptide (CGRP) to stimulate ILC2s and elicit downstream immune responses. In addition, PNECs act through the neurotransmitter γ-aminobutyric acid (GABA) to induce goblet cell hyperplasia. The instillation of a mixture of CGRP and GABA in Ascl1-mutant airways restores both immune and goblet cell responses. In accordance, lungs from human asthmatics show increased PNECs. These findings demonstrate that the PNEC-ILC2 neuroimmunological modules function at airway branch points to amplify allergic asthma responses.

中文翻译：

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肺神经内分泌细胞放大过敏性哮喘反应


寻找 PNEC 在哮喘中的作用 肺神经内分泌细胞 (PNEC) 是一种罕见的细胞类型，位于气道和肺泡上皮细胞中，经常与感觉神经纤维接触。它们具有广泛的系统发育分布，甚至在两栖动物和爬行动物相对原始的肺部中也发现了它们，这表明它们具有重要的功能。隋等人。发现缺乏 PNEC 的小鼠抑制了 2 型（过敏性）免疫反应。在气道分支点周围观察到 PNEC 靠近第 2 组先天淋巴细胞 (ILC2)。 PNEC 通过分泌 CGRP（降钙素基因相关肽）增强 ILC2 活性。他们还通过神经递质 GABA（γ-氨基丁酸）诱导杯状细胞增生。有趣的是，人类哮喘患者被发现有增加的 PNEC 数量，这表明哮喘治疗的潜在治疗靶点。科学，本期第 14 页。 eaan8546 PNEC 是气道中的一种罕见细胞群，对于将气道过敏原信号放大为肺部粘膜反应至关重要。简介 肺部具有巨大的表面积，可以感知吸入空气中的信号并做出反应。肺部与环境之间的异常相互作用是许多疾病的根源，包括哮喘。体外数据表明，肺神经内分泌细胞（PNEC）是一种罕见的气道上皮细胞群，可以充当化学传感器。一旦在培养物中受到刺激，它们就会释放出富含神经肽、胺和神经递质的致密核心囊泡。这些生物活性分子能够引发免疫和生理反应。我们小组最近的一项体内研究表明，PNEC 正确发育成称为神经上皮体的自聚集单元对于限制幼稚肺中免疫细胞的数量至关重要。 然而，PNEC 是否可以在体内发挥作用，将过敏原等外源气道信号转化为下游反应级联尚不清楚。基本原理 为了检验 PNEC 作为肺部传感器的假设，我们通过灭活气道上皮中的 Ascl1 生成了缺乏 PNEC 的小鼠突变体，即从发育开始就耗尽 PNEC 的突变体。我们按照现有哮喘模型的方案，将这些突变体暴露于卵清蛋白或屋尘螨。我们确定突变体是否表现出与对照组不同的哮喘反应。我们通过识别 PNEC 的分子效应器和细胞靶点来阐明其潜在机制。为了补充小鼠的功能测试，我们研究了人类哮喘患者的 PNEC 是否表现出病理变化。结果 尽管基线时正常，但与对照组相比，Ascl1 突变小鼠在过敏原攻击后表现出杯状细胞增生和免疫细胞数量严重减少。在研究可能的分子效应物时，我们发现在过敏原攻击后，突变体中的几种 PNEC 产物相对于对照组有所减少，包括降钙素基因相关肽 (CGRP) 和 γ-氨基丁酸 (GABA)。在探索可能的细胞靶标时，我们发现先天淋巴 2 组细胞 (ILC2) 在气道分支点富集，与 PNEC 类似。 PNEC 产物 CGRP 刺激培养物中 ILC2 产生白细胞介素 5。相反，ILC2 中 CGRP 受体基因 Calcrl 的失活会导致对过敏原的免疫反应减弱。与 CGRP 相比，GABA 不会增加 ILC2 细胞因子的分泌。 相反，GABA 生物发生的失活导致过敏原攻击后杯状细胞增生缺陷，这表明气道上皮的这种反应需要 GABA。在 Ascl1 突变体中注入 CGRP 和 GABA 混合物可恢复过敏原攻击后免疫细胞的增加和杯状细胞的增生，表明这些产品是体内 PNEC 的主要分子效应器。与小鼠的这些结果一致，我们发现人类哮喘患者的 PNEC 数量和簇大小增加，这可能是这些个体对过敏原反应增强的基础。结论 我们的结果表明，尽管 PNEC 是气道中的稀有细胞群，但它对于将气道过敏原信号放大为粘膜 2 型反应至关重要。具体来说，PNEC 通过其产物 GABA 刺激气道上皮粘液的产生。与此同时，PNEC 通过另一种产物 CGRP 发挥作用，刺激 ILC2 产生细胞因子，进而招募下游免疫细胞。 PNEC和ILC2在气道分支点形成神经免疫模块，也是气道颗粒富集的部位。我们的研究结果表明，PNEC-ILC2 轴的功能是感知吸入输入（例如过敏原），并将其放大为肺部输出（例如过敏性哮喘反应）。 PNEC 优先位于分支点。小鼠气道经 CGRP 抗体染色，用于标记 PNEC（洋红色），经 SCGB1A1 抗体染色，用于标记俱乐部细胞（绿色）（200 倍放大倍数）。 PNEC 通常聚集成神经上皮体，并且优先位于分支点。肺神经内分泌细胞（PNEC）是罕见的气道上皮细胞，其功能尚不清楚。 在这里，我们发现没有 PNEC 的 Ascl1 突变小鼠在过敏性哮喘模型中表现出严重减弱的 2 型粘膜反应。 PNEC 位于气道分支点附近的第 2 组先天淋巴细胞 (ILC2) 附近。 PNEC 通过降钙素基因相关肽 (CGRP) 刺激 ILC2 并引发下游免疫反应。此外，PNEC通过神经递质γ-氨基丁酸（GABA）发挥作用，诱导杯状细胞增生。将 CGRP 和 GABA 混合物滴注到 Ascl1 突变体气道中可以恢复免疫和杯状细胞反应。与此相一致的是，人类哮喘患者的肺部显示 PNEC 增加。这些发现表明 PNEC-ILC2 神经免疫模块在气道分支点发挥作用，放大过敏性哮喘反应。