Lung transplant injury drives chronic lung allograft dysfunction via recruitment ofmonocyte-derived alveolar macrophages

项目概述：对于许多晚期肺部疾病患者，肺移植仍然是唯一的选择。 延长生命的可行治疗方案。然而，肺移植患者的5年生存率为54%， 在实体器官移植接受者中。慢性肺移植物功能障碍（CLAD）是导致 肺或心肺移植后的发病率和晚期死亡率。因此，生物标志物的鉴定和 新的治疗靶点对于预防或治疗CLAD和延长肺移植后的存活是必不可少的。 虽然多种因素导致CLAD，但肺移植手术后的早期事件对CLAD的发生起着至关重要的作用。 在为随后的CLAD奠定基础方面发挥作用。这些包括手术创伤和缺血再灌注 激活循环和驻留免疫细胞的损伤，随后是内皮损伤和免疫细胞 外渗。我们已经报道，单核细胞，招募到受伤的肺，可以建立一个长期的， 驻留并分化为致病性单核细胞衍生的肺泡巨噬细胞。我们已经找到了 单核细胞衍生的肺泡巨噬细胞的组织重塑和纤维化（类似CLAD）使用遗传 删除策略此外，使用无偏的单细胞转录组学分析（RNA-seq）， 来自肺纤维化患者的组织和供体肺的活组织检查（两者都是在肺移植期间获得的 肺移植外科医生- Ankit Bharat，该提案的主要贡献者），我们确定了一个 肺纤维化患者中特有的致病性肺泡巨噬细胞群。 我们对单细胞RNA-seq数据的计算分析表明，单核细胞来源的肺泡巨噬细胞 通过丛蛋白D1/信号传导被引导至其新的致病性小生境，并由M-CSF/M- CSFR信令。与这一假设一致，靶向M-CSF/M-CSFR信号传导特异性地消除了 单核细胞衍生的肺泡巨噬细胞和改善的病理学。我们提供了小鼠的初步数据， 慢性肺移植物功能障碍的模型和患者支持这种机制与CLAD的相关性。 因此，我们将使用小鼠模型和来自肺移植患者的样本来检验以下假设： 病原性单核细胞衍生的肺泡巨噬细胞，在最初的移植周围损伤期间募集， 通过丛蛋白D1/信号蛋白信号传导建立长期驻留，并通过M-CSF/M- CSFR信号传导在三个相互关联的目标中驱动CLAD： 目的1：确定单核细胞衍生的肺泡巨噬细胞是否在移植肺内募集到移植肺。 在移植当天，通过M-CSF/M-CSFR信号传导维持。 目的2：确定致病性单核细胞衍生的肺泡巨噬细胞是否定位于 丛蛋白D1/脑信号蛋白介导的肺纤维化 目的3：确定异常肺泡巨噬细胞的出现是否与表达增加有关， PLXND 1和自分泌M-CSF/M-CSFR信号可以在早期CLAD患者的BAL液中鉴定。

Project Summary: For many patients with advanced lung disease, lung transplantation remains the only viable therapeutic option to extend life. Yet the 5-year survival for lung transplant patients is 54%, the worst among solid organ transplant recipients. Chronic lung allograft dysfunction (CLAD) is the leading cause of morbidity and late mortality after lung or heart-lung transplantation. Hence, identification of biomarkers and novel therapeutic targets is essential to prevent or treat CLAD and extend survival after lung transplantation. While multiple factors contribute to CLAD, early events after the lung transplant procedure play a crucial role in setting the stage for subsequent CLAD. These include surgical trauma and ischemia-reperfusion injury that activates circulating and resident immune cells followed by the endothelial injury and immune cell extravasation. We have reported that monocytes, recruited to the injured lung, can establish a long-term residency and differentiate into pathogenic monocyte-derived alveolar macrophages. We have causally linked monocyte-derived alveolar macrophages to tissue-remodeling and fibrosis (resembling CLAD) using a genetic deletion strategy. Moreover, using unbiased single-cell transcriptomic profiling (RNA-seq) of explanted lung tissue from the patients with pulmonary fibrosis and biopsies of the donor lung (both obtained during lung transplantation by lung transplant surgeon – Ankit Bharat, key contributor to this proposal), we identified a distinct population of pathogenic alveolar macrophages exclusively present in patients with pulmonary fibrosis. Our computational analyses of single-cell RNA-seq data suggest that monocyte-derived alveolar macrophages are guided to their new pathogenic niches via plexin D1/signaling and are uniquely maintained by M-CSF/M- CSFR signaling. Consistent with this hypothesis, targeting M-CSF/M-CSFR signaling specifically eliminated monocyte-derived alveolar macrophages and ameliorated pathology. We present preliminary data from mouse models and patients with chronic lung allograft dysfunction supporting relevance of this mechanism for CLAD. We will thus use mouse models and samples from lung transplant patients to test the hypothesis that pathogenic monocyte-derived alveolar macrophages, recruited during the initial peri-transplant injury, establish long term residency via plexin D1/semaphorin signaling and are maintained via M-CSF/M- CSFR signaling to drive CLAD in three interrelated aims: Aim 1: To determine whether monocyte-derived alveolar macrophages recruited to the transplanted lung within day of the transplant are maintained by M-CSF/M-CSFR signaling. Aim 2: To determine whether pathogenic monocyte-derived alveolar macrophages are localized to regions of lung fibrosis lung via plexin D1/semaphorin signaling. Aim 3: To determine whether the emergence of aberrant alveolar macrophages with increased expression of PLXND1 and autocrine M-CSF/M-CSFR signaling can be identified in BAL fluid from patients with early CLAD.