Effect of Neonatal Hyperoxia on Alveolar Development and Infection

项目概要（摘要） 越来越多的人认识到产前和产后的环境因素会影响我们以后的健康 生活中发育中的肺将经历的最深刻的环境因素之一是氧气 因为它在出生时从胎儿环境过渡到空气环境。当术语肺准备好呼吸氧气时， 早产儿的肺过快地过渡到空气中，并且经常暴露于用于维持适当的 氧饱和度出生时这种异常的氧气暴露会增加长期肺部疾病的风险 通过不为人知的机制。我们的实验室开发并使用了一种独特的小鼠模型， 了解出生时高水平的氧气如何增加甲型流感病毒（IAV）感染的严重程度， 成年人了在上一个资助期进行的研究确定，新生儿高氧加速 肺泡上皮2型细胞（AEC 2）的增殖，当小鼠返回时，这些细胞缓慢耗尽， 室内空气中，并且当小鼠感染IAV时，AEC 2的损失有助于纤维化肺病。 这项提议是建立在这些发现的基础上，通过研究AEC 2的氧依赖性消耗如何影响细胞的生长， 增加了IAV感染的严重性。通过遗传谱系研究，我们发现新生儿高氧血症 通过刺激AEC 2分化为肺泡上皮1型细胞（AEC 1）来消耗AEC 2， 在IAV感染过程中，高氧可增强AEC 1的死亡，暴露于高氧的成年小鼠中的许多AEC 1 新生儿高氧表达Ki67，Ki67是传统上用于标记癌症进展的增殖标记物。最近 然而，研究表明Ki67并不调节细胞增殖，而是通过其 组织异染色质的能力。因为增殖细胞对基因毒性作用过敏， 高氧和受损细胞可以传递的经验，彻底表观遗传，我们将测试 新生儿高氧通过诱导Ki67依赖性增强对IAV感染敏感性假说 增殖的AEC 2在分化成AEC 1时保持的表观遗传变化。目的 1使用新的Ki67报告基因和Ki67缺失小鼠来确定在新生儿期增殖的AEC 2是否 高氧产生以持续Ki67表达为标志的AEC 1。目标2感染这些小鼠， IAV表达荧光蛋白的重组菌株用于确定Ki67是否修饰了 AEC1对IAV感染有反应。Aim 3使用细胞特异性深度RNA测序和染色质 免疫沉淀以确定Ki67是否修饰基因中氧依赖性变化的子集 这些变化有助于解释为什么AEC 1对IAV感染易感。理解 新生儿高氧如何影响小鼠AEC对IAV感染的反应是很重要的，因为科学 这些发现将刺激旨在改善早产儿健康的治疗方法的发展。

PROJECT SUMMARY (ABSTRACT) There is growing appreciation that prenatal and postnatal environmental factors shape our health later in life. One of the most profound environmental factors that the developing lung will ever experience is oxygen as it transitions at birth from a fetal to air environment. While the term lung is prepared to breathe oxygen, the preterm lung transitions into air too soon and is often exposed to excess oxygen used to maintain appropriate oxygen saturations. This aberrant oxygen exposure at birth increases the risk for long-term lung disease through poorly understood mechanisms. Our laboratory developed and uses a unique mouse model designed to understand how high levels of oxygen at birth increases the severity of influenza A virus (IAV) infections in adults. Studies conducted during the previous funding period established that neonatal hyperoxia accelerates proliferation of alveolar epithelial type 2 cells (AEC2s), that these cells are slowly depleted when mice return to room air, and that the loss of AEC2s contributes to fibrotic lung disease when the mice are infected with IAV. This proposal builds on these findings by investigating how the oxygen-dependent depletion of AEC2s enhances the severity of IAV infections. Using genetic lineage studies, we found that neonatal hyperoxia depletes AEC2s by stimulating their differentiation into alveolar epithelial type 1 cells (AEC1s), neonatal hyperoxia enhances death of AEC1s during IAV infection, and that many AEC1s in adult mice exposed to neonatal hyperoxia express Ki67, a proliferation marker traditionally used to mark cancer progression. Recent studies however show Ki67 does not regulate cell proliferation but rather modifies gene expression via its ability to organize heterochromatin. Because proliferating cells are hypersensitive to the genotoxic effects of hyperoxia and damaged cells can transmit that experience thorough epigenetic inheritance, we will test the hypothesis that neonatal hyperoxia enhances sensitivity to IAV infection by inducing Ki67-dependent epigenetic changes in proliferating AEC2s that are maintained when they differentiate into AEC1s. Aim 1 uses novel Ki67-reporter and Ki67-null mice to determine whether AEC2s that proliferate during neonatal hyperoxia produce AEC1s that are marked by persistent Ki67 expression. Aim 2 infects these mice with recombinant strains of IAV-expressing fluorescent protein used to determine whether Ki67 modifies how AEC1s respond to IAV infection. Aim 3 uses cell-specific deep RNA sequencing and chromatin immunoprecipitation to determine whether Ki67 modifies a subset of oxygen-dependent changes in gene expression and that these changes help explain why AEC1s are susceptible to IAV infection. Understanding how neonatal hyperoxia shapes how AECs respond to IAV infection in mice is important because the scientific discoveries will stimulate development of therapies designed to improve the health of people born preterm.