Decoding temporal epithelial signaling programs to restore homeostasis in acute lung injury

摘要 急性肺损伤/急性呼吸窘迫综合征（Acute lung injury/acute respiratory distress syndrome，ALI/ARDS）是一种常见的肺损伤， 综合征，死亡率高。ALI/ARDS源于肺与肺之间的沟通障碍， 上皮细胞和免疫细胞，并且是严重呼吸道感染病例中的主要因素， 包括新冠肺炎ALI/ARDS的治疗目前受到不充分的模型的限制。 信号通路与肺上皮细胞行为之间的复杂关系。核心 信号通路网络控制上皮结构和细胞因子释放，是一种 通过现有靶向激酶抑制剂对ALI/ARDS进行干预的重要未开发点，或 代谢调节剂。该核心网络包括NF-kB，一种主要的转录因子， 细胞因子释放和炎症，以及激酶ERK、AKT、JNK、p38、mTOR和AMPK， 其响应细胞生长和代谢状态调节细胞因子的产生。 值得注意的是，这种信号网络也在肺癌中起着因果作用，并参与其他癌症的发生。 肺上皮疾病，包括纤维化和COPD。我们的实验室开发了一种 广泛的活细胞技术平台，用于跟踪信号事件的时间程序， 这些途径之间的时间、强度和协调的不同模式， 决定上皮细胞的命运。我们的总体目标是定量解码细胞因子如何 分泌由多激酶活性程序指定。我们的假设是 在单细胞水平上的信号传导活动的重合阶段将显著改善细胞的免疫功能。 预测细胞因子分泌的整体和局部变异性，并将确定代谢 减轻炎症的手法。我们的方法将开发所需的技术， 检测和操纵时间信号程序，以恢复肺上皮细胞的稳态， ALI/ARDS。AIM 1将开发一个高内容的数据驱动模型， 在多种条件下，细胞逐个细胞地释放细胞因子。目标2将开发 表征肺上皮亚区中局部炎症环境的能力 通过检查10-100个相邻细胞的组中激酶信号传导的动态程序。 在此过程中，我们将提供使用固定免疫荧光板的原理证明 标记来捕捉动态信号模式。目标3将研究以下方面的潜力： 用现有的药理学试剂操纵时间信号程序，专注于 AMPK激活剂作为调节较大信号网络的途径的新兴益处。我们 多学科团队将跨越肺部护理，细胞生物学和 计算系统建模，以创建一个技术平台，连接现代单一的， 细胞信号转导分析在肺部疾病的紧迫挑战。我们预计， 该项目将确定潜在的新的ALI/ARDS干预途径，以及揭示 细胞因子信号传导途径的一般概念，与治疗广泛相关， 肺恶性肿瘤和炎性疾病。

Abstract Acute lung injury/acute respiratory distress syndrome (ALI/ARDS) is a common pulmonary syndrome with high mortality. ALI/ARDS stems from a communication breakdown between lung epithelial cells and immune cells and is a major factor in severe cases of respiratory infection, including COVID-19. Treatments for ALI/ARDS are currently limited by inadequate models of the complex relationships between signaling pathways and lung epithelial cell behavior. A core network of signaling pathways control epithelial structure and cytokine release and are an important unexploited point of intervention in ALI/ARDS via existing targeted kinase inhibitors or metabolic modulators. This core network includes NF-kB, a primary transcription factor for cytokine release and inflammation, and the kinases ERK, AKT, JNK, p38, mTOR and AMPK, which modulate cytokine production in response to cellular growth and metabolic status. Notably, this signaling network also plays a causal role in lung cancer and is involved in other diseases of the lung epithelium, including fibrosis and COPD. Our lab has developed an extensive live-cell technology platform for tracking temporal programs of signaling events – different patterns of timing, intensity, and coordination between these pathways – that govern epithelial cell fate decisions. Our overall objective is to quantitatively decode how cytokine secretion is specified by multi-kinase activity programs. Our hypothesis is that accounting for coinciding phases of signaling activity at the single cell level will significantly improve the prediction of both overall and local variability in cytokine secretion and will identify metabolic manipulations that reduce inflammation. Our approach will develop the technology needed to detect and manipulate temporal signaling programs to restore lung epithelial homeostasis in ALI/ARDS. Aim 1 will develop a high-content data-driven model correlating signaling programs to cytokine release on a cell-by-cell basis across multiple conditions. Aim 2 will develop the capacity to characterize the localized inflammatory environment in subregions of lung epithelium by examining the dynamic programs of kinase signaling in groups of 10-100 neighboring cells. In the process, we will provide proof of principle for using panels of fixed immunofluorescence markers to capture dynamic signaling patterns. Aim 3 will investigate the potential for manipulating temporal signaling programs with existing pharmacological agents, focusing on the emerging benefits of AMPK activators as a route to modulate the larger signaling network. Our multi-disciplinary team will work across the boundaries of pulmonary care, cell biology, and computational systems modeling to create a technology platform that connects modern single- cell signal transduction analysis to pressing challenges in lung disease. We anticipate that our project will identify potential new routes of intervention for ALI/ARDS, as well as revealing general concepts in cytokine signaling pathways that are broadly relevant for the treatment of lung malignancies and inflammatory disease.