Development of PNEC innervation and neuroplasticity after early life insult

摘要 肺神经内分泌细胞(PNECs)如何被神经支配以及神经对PNEC的调节程度 在发育和疾病中的作用是该领域的开放和基本问题。这样做的目的是 建议解决这些基本问题，并评估神经营养素4(NT4)在这些 事件。使用NT4-/-小鼠品系和暴露于过敏原的新生小鼠模型，我们产生了 初步数据导致了本提案中将研究的3个假设：1)PNEC需要NT4 发育过程中的神经支配和早期生命侮辱后PNEC神经支配的增加；2)早期 生活变应原暴露改变支配PNECs的感觉传入和传出神经的功能 从而导致γ-氨基丁酸分泌失调和长期杯状细胞化生；3) 肺肥大细胞是早期生活变应原暴露后NT4水平升高的候选来源。至 综上所述，我们发现NT4在出生后发育的PNECs中表达，并作为营养因子发挥作用 神经性神经建立连接的因素。暴露于发育中的、出生后肺的过敏原 使NT4水平异常升高。在这种病理情况下，我们发现PNEC的神经支配 随着杯状细胞化生时间的延长而增加。值得注意的是，PNEC是唯一的细胞来源 肺中的GABA，过敏原诱导的小鼠模型中杯状细胞化生及其相关的信号 与人类哮喘患者和吸烟者的粘液过度产生有关。此外，PNEC还需要NT4 神经过度支配和GABA分泌失控，与其他 神经调节内分泌的神经内分泌系统。这些初步发现表明， NT4在PNEC神经发育和早期生命损伤后神经可塑性中的重要作用 它将通过比较PNEC的神经支配方式和程度来广泛表征 使用不同类型神经标记物的野生型和NT4-/-小鼠，有无过敏原暴露(目的 1)。NT4诱导的PNEC过度神经支配与早期杯状细胞化生延长的关系 过敏原暴露，AIM 2中拟议的实验将评估感觉传入和 PNECs分泌GABA的传出信号及其与NT4的关系。最后，考虑到 NT4在异常PNEC神经支配中的中心作用在病理条件下，我们检测了NT4的表达 在受伤的肺部。我们发现，在生命早期的过敏原中，有一个扩大的、激活的肥大细胞群表达NT4。 曝光。肺肥大细胞是否通过产生NT4参与PNEC的超神经支配 评估(目标3)。为了进一步增强疾病相关性，我们将验证小鼠工作中的关键发现 婴幼儿灵长类损伤模型和人肺标本。总而言之，这项提案调查的是复杂的 出生后发育和损伤过程中神经、PNECs和炎症之间的相互作用。我们的发现 提示慢性呼吸道疾病的发病机制，如哮喘，可能涉及干扰。 在早期的侮辱事件之后的发育过程。我们预计拟议的 研究将提供有关肺神经内分泌系统如何形成和 功能。黏膜过度分泌的神经-PNEC轴机制的鉴定 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.