Development of PNEC innervation and neuroplasticity after early life insult

摘要 肺神经内分泌细胞（PNEC）如何受到神经支配以及神经在多大程度上调节PNEC 在发育和疾病中的功能是该领域的开放和基本问题。的目的 我们的建议是解决这些基本问题，并评估神经营养因子4（NT4）在这些方面的关键作用。 事件使用NT4-/-小鼠系和新生小鼠过敏原暴露模型，我们产生了 初步数据导致本提案将研究3个假设：1）PNEC需要NT4 神经支配在发展过程中和增加PNEC神经支配后，早期生活的侮辱; 2）早期 生活变应原暴露改变了支配PNEC的感觉传入和传出神经的功能 从而导致γ-氨基丁酸（GABA）分泌失调和长期杯状细胞化生; 3） 肺肥大细胞是在生命早期暴露于过敏原后升高的NT 4水平的候选来源。到 总之，我们发现NT4在出生后发育过程中由PNEC表达，并作为营养因子发挥作用， 使支配神经建立连接的因素。过敏原暴露于发育中的出生后肺部 异常地升高了NT4的水平在这种病理状态下，我们发现PNEC的神经支配 与杯状细胞化生延长相关。值得注意的是，PNEC是唯一的细胞来源， 肺中的GABA是小鼠模型中过敏原诱导杯状细胞化生所必需的信号， 与人类哮喘患者和吸烟者的粘液分泌过剩有关。此外，PNEC需要NT4 神经支配过度和GABA分泌失调，与其他研究中建立的范式一致。 神经内分泌系统，即神经调节内分泌。这些初步调查结果表明， NT4在发育过程中PNEC神经支配和早期生命损伤后的神经可塑性中的重要作用， 这将通过比较PNEC神经支配的模式和程度来广泛表征， 使用不同类型神经的标记物，在有和没有变应原暴露的情况下，野生型和NT4-/-小鼠（Aim 1）。将NT4诱导的PNEC神经支配过度与生命早期延长的杯状细胞化生联系起来 过敏原暴露，目标2中提出的实验将评估感觉传入的功能变化， 传出信号，诱导GABA分泌的PNEC和他们的关系NT4。最后，鉴于 在病理条件下，NT4在异常PNEC神经支配中的中心作用，我们检测了NT4的表达 受伤的肺部。我们发现在生命早期过敏原中，扩大、活化的肥大细胞群表达NT 4 exposure.肺肥大细胞是否通过产生NT 4而促进PNEC的神经支配， 评价（目标3）。为了进一步增强疾病的相关性，我们将验证小鼠工作的关键发现， 婴儿灵长类动物损伤模型和人肺样本。这项提案共同探讨了复杂的 在出生后发育和损伤过程中，神经、PNEC和炎症之间的相互作用。我们的研究结果 表明慢性气道疾病如哮喘的发病机制可能涉及干扰， 在早期侮辱事件之后的发育过程。我们期望完成拟议的 研究将提供有关肺神经内分泌系统如何形成的基础知识， 功能协调发展的粘液分泌过多的神经-PNEC轴沿着机制的鉴定 可能为发现新的治疗策略奠定基础。

ABSTRACT How pulmonary neuroendocrine cells (PNECs) become innervated and to what extent nerves regulate PNEC function in development and diseases are open and basic questions for the field. The objective of this proposal is to address these fundamental issues and to assess the key role of neurotrophin 4 (NT4) in these events. Using a NT4-/- mouse line and a neonatal mouse model of allergen exposure, we generated preliminary data leading to 3 hypotheses that will be studied in this proposal: 1) NT4 is required for PNEC innervation during development and for the increases in PNEC innervation following early life insults; 2) early life allergen exposure alters the function of sensory afferents and efferent nerves that innervate PNECs thereby causing deregulated γ-aminobutyric acid (GABA) secretion and long-term goblet cell metaplasia; 3) pulmonary mast cells are a candidate source of elevated NT4 levels following early life allergen exposure. To summarize, we found that NT4 was expressed by PNECs during postnatal development and acted as a trophic factor for the innervating nerves to establish connection. Allergen exposure to developing, postnatal lungs aberrantly elevated the levels of NT4. Under this pathological condition, we discovered that PNEC innervation was increased associated with prolonged goblet cell metaplasia. Notably, PNECs were the only cell source of GABA in lungs, a signal essential for allergen-induced goblet cell metaplasia in mouse models and associated with mucous overproduction in human asthmatics and smokers. In addition, NT4 was required for PNEC hyperinnervation and deregulated GABA secretion, consistent with the established paradigm in other neuroendocrine systems that nerves regulate endocrine secretion. These preliminary findings point to an essential role for NT4 in PNEC innervation during development and neuroplasticity following early life injury, which will be extensively characterized by comparing the pattern and degree of PNEC innervation between wild type and NT4-/- mice with and without allergen exposure using markers for different types of nerves (Aim 1). To connect NT4-induced PNEC hyperinnervation to prolonged goblet cell metaplasia following early life allergen exposure, proposed experiments in Aim 2 will assess functional changes in sensory afferents and efferent signals that induce GABA secretion from PNECs and their relationships to NT4. Lastly, given the central role of NT4 in aberrant PNEC innervation under pathological conditions, we examined NT4 expression in injured lungs. We found an enlarged, activated mast cell population expresses NT4 during early life allergen exposure. Whether pulmonary mast cells contribute to PNEC hyperinnervation by producing NT4 will be evaluated (Aim 3). To further enhance disease relevance, we will validate key findings from the mouse work in infant primate models of injury and human lung samples. Together, this proposal investigates complex interactions between nerves, PNECs, and inflammation during postnatal development and injury. Our findings indicate that the pathogenesis of chronic airway diseases, such as asthma, may involve disrupted developmental processes following early episodes of insults. We expect that completion of the proposed studies will provide fundamental knowledge about how the pulmonary neuroendocrine system forms and functions. Identification of the mechanisms along the nerve-PNEC axis underlying mucous overproduction may lay the foundation for the discovery of new treatment strategies.