Integrated Analysis of Tracheobronchial Fluid from Before and After Cardiopulmonary Bypass Reveals Activation of the Integrated Stress Response and Altered Pulmonary Microvascular Permeability.
Integrated Analysis of Tracheobronchial Fluid from Before and After Cardiopulmonary Bypass Reveals Activation of the Integrated Stress Response and Altered Pulmonary Microvascular Permeability.
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DOI：
10.59249/kfyz8002
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发表时间：
2023-03
期刊：
影响因子：
--
通讯作者：
Pierce RW
中科院分区：
文献类型：
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作者：
Habet V;Li N;Qi J;Peng G;Charkoftaki G;Vasiliou V;Sharma L;Pober JS;Dela Cruz C;Yan X;Pierce RW
目的：我们的目的是全面描述的转录活性和信号转导的肺实质和免疫细胞的心肺转流（CPB）之前和之后，通过使用多组学方法结合功能细胞分析。我们推测，来自肺内特定细胞的关键信号通路改变了肺内皮细胞功能，导致疾病恶化或改善。方法：我们收集了一系列气管支气管灌洗样本，来自于2岁以下接受CPB手术的插管患者。立即处理样品用于单细胞RNA测序（10x Genomics）。进行细胞聚类、细胞类型注释和可视化，并鉴定系列样品之间的差异表达基因（DEG）。分别使用质谱法和多重测定法（SomaScan）对上清液进行代谢组学和蛋白质组学分析。使用电细胞基质阻抗传感来进行功能测定以测量穿过人肺微血管内皮细胞（HPMEC）的电阻。结果：对八名患者的分析显示肺实质细胞和免疫细胞的异质混合物。细胞聚类显示转录组特征的时间依赖性变化，表明CPB后细胞表型改变。DEG分析由参与宿主防御、先天免疫和线粒体呼吸运输链的基因表示。损伤途径分析显示CPB后所有细胞类型的整合应激反应均上调。代谢组学分析表明抗坏血酸和醛糖酸代谢的上调。无偏见的蛋白质组学分析显示，参与细胞因子和趋化因子途径的蛋白质上调。CPB后患者上清液改善了HMPEC屏障功能，表明对CPB的保护性细胞反应。结论：接受心脏手术CPB的儿童具有不同的细胞群，转录活性和代谢随时间而变化。儿童下呼吸道对缺血再灌注损伤的反应似乎是保护性的，需要通过未来的研究确定潜在的靶点。
Objective: We aim to comprehensively describe the transcriptional activity and signaling of pulmonary parenchymal and immune cells before and after cardiopulmonary bypass (CPB) by using a multi-omic approach coupled with functional cellular assays. We hypothesize that key signaling pathways from specific cells within the lung alter pulmonary endothelial cell function resulting in worsening or improving disease. Methods: We collected serial tracheobronchial lavage samples from intubated patients less than 2-years-old undergoing surgery with CPB. Samples were immediately processed for single cell RNA sequencing (10x Genomics). Cell clustering, cell-type annotation, and visualization were performed, and differentially expressed genes (DEG) between serial samples were identified. Metabolomic and proteomic analyses were performed on the supernatant using mass spectrometry and a multiplex assay (SomaScan) respectively. Functional assays were done using electric cell-substrate impedance sensing to measure resistance across human pulmonary microvascular endothelial cells (HPMECs). Results: Analysis of eight patients showed a heterogeneous mixture of pulmonary parenchymal and immune cells. Cell clustering demonstrated time-dependent changes in the transcriptomic signature indicating altered cellular phenotypes after CPB. DEG analysis was represented by genes involved in host defense, innate immunity, and the mitochondrial respiratory transport chain. Ingenuity pathway analysis showed upregulation of the integrated stress response across all cell types after CPB. Metabolomic analysis demonstrated upregulation of ascorbate and aldarate metabolism. Unbiased proteomic analysis revealed upregulation of proteins involved in cytokine and chemokine pathways. Post-CPB patient supernatant improved HMPEC barrier function, suggesting a protective cellular response to CPB. Conclusion: Children who undergo CPB for cardiac surgery have distinct cell populations, transcriptional activity, and metabolism that change over time. The response to ischemia-reperfusion injury in the lower airway of children appears to be protective, with the need to identify potential targets through future investigations.