Mechanisms of immunomodulation in lung injury by mesenchymal stromal cell exosomes

总结 由于没有单一有效的治疗方法来预防或治疗支气管肺发育不良（BPD）， 早产儿的慢性肺病，需要新的工具来治疗和减少进一步并发症的风险， 紧急间充质干细胞/基质细胞（MSC）疗法在BPD的临床前模型中显示出前景， 证明了组织学和功能益处。我们已经证明了MSC的治疗能力 包含在它们的分泌体中，并且其中的主要治疗载体由外泌体代表。 外泌体是亚微米囊泡，其含有多种生物活性货物（例如脂质、多种蛋白质， 和小的非编码RNA）。最近，使用高氧诱导的BPD小鼠模型，我们证明， 推注剂量的人MSC-外泌体（称为MEx），显著改善肺形态， 发育和肺功能，减少肺纤维化，恢复肺血管损失，改善 肺血管重构和肺动脉高压。我们还证明了MEx很容易 在体外和体内都被巨噬细胞（Mφ）吸收，结果使Mφ表型转变为抗- 炎症、抗纤维化和促调节状态。然而，尽管MEx具有很好的治疗潜力， 我们对它们的生物活性特性和导致这种效应的分子机制的理解 仍然不清楚。外来体是异质EV群体。已知外来体亚群在以下方面不同： 生物物理学、蛋白质组学和RNA库。因此，不同的MEx亚群介导替代生物学特性， 功能协调发展的在这个提议中，我们预测只有某种MEx亚型负责治疗效果， 并且这种“生物活性”MEx亚群通过调节Mφ表型来抑制肺中的炎症信号传导 并防止高氧诱导的血管和肺泡损伤的发展。为了验证这些假设，我们 提出以下具体目标（SA）。SA #1：分离、表征和定义治疗性（“生物活性”）MEx 子集SA #2：评估MEx的体内生物分布和抗炎/免疫调节能力。SA #3： 研究MEx如何影响我们实验BPD模型中靶细胞的表观遗传景观。 总的来说，该提案将为MEx生物学和MEx-靶细胞相互作用提供重要见解， 开发有效的新型BPD治疗模式。

Summary With no single effective therapy for either the prevention or treatment of bronchopulmonary dysplasia (BPD), a chronic lung disease of preterm infants, the need for new tools to treat and reduce risk of further complication is urgent. Mesenchymal stem/stromal cell (MSC) therapy has shown promise in preclinical models of BPD, demonstrating both histological and functional benefits. We have shown that the therapeutic capacity of MSCs is comprised in their secretome, and that the major therapeutic vector therein is represented by the exosomes. Exosomes are submicron vesicles that harbor a diverse array of bioactive cargo (e.g. lipids, diverse proteins, and small non-coding RNAs). Recently, using a murine model of hyperoxia-induced BPD, we demonstrated that a bolus dose of human MSC-exosomes (termed MEx), significantly improved lung morphology, pulmonary development and lung function, decreased lung fibrosis, restored pulmonary blood vessel loss and ameliorated pulmonary vascular remodeling and pulmonary hypertension. We have also demonstrated that MEx are readily taken up by macrophages (Mφ) both in vitro and in vivo and, as a result, shift the Mφ phenotype to an anti- inflammatory, anti-fibrotic, and pro-regulatory state. However, despite the promising therapeutic potential of MEx, our understanding of their bioactive properties and the molecular mechanism(s) responsible for such effects remain unclear. Exosomes are a heterogeneous EV population. Exosome subpopulations are known to differ in biophysical, proteomic and RNA repertoire. Consequently, different MEx subsets mediate alternative biological functions. In this proposal we predict that only a certain subtype of MEx is responsible for the therapeutic effects, and that this ‘bioactive’ MEx subset dampens inflammatory signaling in the lung via modulation of Mφ phenotype and prevents the development of hyperoxia–induced vascular and alveolar injury. To test these hypotheses, we propose the following specific aims (SA). SA#1: To isolate, characterize and define therapeutic (‘bioactive’) MEx subsets. SA#2: To assess MEx in vivo biodistribution and anti-inflammatory/immunomodulatory capacity. SA#3: Investigate how MEx impacts the epigenetic landscape of target cells in our experimental BPD model. Collectively, this proposal will provide important insights in MEx biology and MEx-target cell interaction, that can be leveraged to develop effective, novel therapeutic modalities for BPD.