Role of Pulmonary Osteoclast-Like Cells in Lung Injury

肺破骨细胞样细胞在肺损伤中的作用

• 批准号: 10620655
• 负责人: Francis Xavier McCormack
• 金额: --
• 依托单位: CINCINNATI VA MEDICAL CENTER RESEARCH
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-05-01 至 2025-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10620655
• 关键词: Acids; Address; Adopted; Adoptive Transfer; Afghanistan; Alveolar; Alveolar Macrophages; Animals; Asbestos; Asbestosis; Asia; Automobile Driving; Bone Marrow; Bone Matrix; CSF1 gene; Calcitonin Receptor; Calcium; Cattle; Cell Differentiation process; Cell Separation; Cell physiology; Cell secretion; Cells; Chloride Channels; Collagen; Complex; Data; Defect; Development; Disease; Enzyme-Linked Immunosorbent Assay; Enzymes; Epithelium; Exposure to; FDA approved; Fiber; Fibrosis; Foreign-Body Giant Cells; Gene Expression; Genes; Genetic; Giant Cells; Gulf War; Health; Histologic; Human; Hydrochloric Acid; Hydroxyapatites; Hydroxyproline; In Vitro; In complete remission; Infiltration; Inflammation; Inhalation; Injections; Injury; Integrins; Irrigation; Knockout Mice; Liquid substance; Lung; Lung Compliance; MMP9 gene; Macrophage; Macrophage Colony-Stimulating Factor; Maps; Measures; Mechanics; Mediating; Microspheres; Middle East; Military Personnel; Minerals; Mission; Mus; Mutation; Myeloid Cells; Occupations; Osteoclasts; Outcome; Particulate; Pathogenesis; Patients; Peptide Hydrolases; Phenotype; Physiological; Poison; Production; Proteins; Proton Pump; Pulmonary Fibrosis; Pulmonary alveolar microlithiasis; Residual state; Risk; Role; Sailor; Slice; Source; Specific qualifier value; Stains; Stereotyping; Structure of parenchyma of lung; TRANCE protein; Tamoxifen; Testing; Tissues; Veterans; Vietnam; War; Zoledronic Acid; autosome; bisphosphonate; bone; burn pit; career; cathepsin K; cell injury; cell transformation; cytokine; experimental study; fibrotic lung; genetic signature; healing; human RNA sequencing; indexing; inorganic phosphate; lung injury; military operation; military service; monocyte; neutralizing antibody; novel; particle; pharmacologic; pre-clinical; prevent; programs; pulmonary function; recruit; respiratory; response; single-cell RNA sequencing; sodium-phosphate cotransporter proteins; surfactant; targeted treatment; tartrate-resistant acid phosphatase; theories

This proposal explores the role of a pulmonary osteoclast-like cell (POLC) in asbestos-induced pulmonary fibrosis. We first discovered POLCs while studying pulmonary alveolar microlithiasis (PAM), a rare, autosomal recessive disorder caused by mutations in the epithelial sodium phosphate co-transporter, Npt2b. Phosphate accumulates in the alveolar lining fluid and complexes with calcium to form spherical hydroxyapatite microliths containing bone matrix proteins and surfactant components. Contact of microliths with alveolar macrophages (AM) and recruited alveolar monocytes (Alv-Mo) induces osteoclastic transformation, with expression of the full repertoire of osteoclast signature genes and proteins in multinucleated giant cells (MNGC) including tartrate resistant acid phosphatase (TRAP), cathepsin K (CTSK), and the proton pump ATP6V0D2. Single cell RNA sequencing of human PAM lung also confirmed a robust osteoclast signature in AM, and IHC confirmed the presence of TRAP and CTSK positive MNGC. Like humans, Npt2b-/- animals develop modest pulmonary fibrosis and a marked restrictive physiologic defect. We found that microliths induce alveolar expression of the requisite osteoclastogenic cytokines, mCSF and RANKL, for POLC differentiation and expression of hydrochloric acid and CTSK that both dissolve stones and damage tissues. We also found that when microliths were adoptively transferred into the lungs of WT animals, they attached to or were degraded by macrophages, and were cleared within 28d without residual inflammation or fibrosis or evidence of lung injury. We noted that hyperdense infiltrates in our index PAM patient progressed rapidly when she was placed on bisphosphonates, and that anti-RANKL therapy slowed the clearance of microliths in Npt2b-/- mice. These data led us to the conclusion that osteoclastic transformation of AM and Alv-Mo is the lung's primary defense against microliths and that it may represent a stereotypic response to other particles, including asbestos. Indeed, we find that asbestos challenge is also associated with TRAP and CTSK expression in myeloid cells and MNGC in the BAL and lung tissue, and that it culminates in destructive remodeling and pulmonary fibrosis. The BAL cells from asbestos but not saline treated mice degrade bone and liberate collagen fragments from the bone matrix when plated on bovine bone slices, the signature function of osteoclasts. The differential tissue responses of complete healing vs. fibrosis to dissolvable (microliths) vs. persistent (asbestos) particulates forms the basis for our hypothesis that acid and matrix degrading enzymes produced by POLCs may be primary drivers of fibrosis when the inhaled particle is invincible. To test this hypothesis in three aims, we will determine; 1) the gene programs that underlie pulmonary osteoclastic specification of recruited monocytes in response to asbestos, 2) the osteoclast functions acquired by monocytes and macrophages upon asbestos challenge, 3) the role of POLCs in the pathogenesis of asbestos-induced pulmonary fibrosis. This proposal directly addresses mechanisms relevant to progressive pulmonary fibrosis that still occurs in the thousands of veterans exposed to asbestos in all branches of the military from the 1930s through the Vietnam, Gulf and Afghanistan Wars, and seeks to develop preclinical evidence supporting anti-osteoclastic strategies such as bisphosphonates of Denosumab for the treatment of asbestosis.

该提案探讨了肺破骨细胞样细胞（POLC）在石棉诱导的肺纤维化中的作用。 我们在研究肺泡微石症 (PAM) 时首次发现 POLC，这是一种罕见的常染色体隐性遗传病 由上皮磷酸钠协同转运蛋白 Npt2b 突变引起的疾病。磷酸盐积累 在肺泡衬里液中并与钙复合形成含有骨的球形羟基磷灰石微石 基质蛋白和表面活性剂成分。微石与肺泡巨噬细胞（AM）接触并招募 肺泡单核细胞 (Alv-Mo) 诱导破骨细胞转化，表达全部破骨细胞 多核巨细胞 (MNGC) 中的破骨细胞特征基因和蛋白质，包括抗酒石酸 磷酸酶 (TRAP)、组织蛋白酶 K (CTSK) 和质子泵 ATP6V0D2。单细胞 RNA 测序 人 PAM 肺也证实了 AM 中强大的破骨细胞特征，IHC 证实了 TRAP 的存在 CTSK 阳性 MNGC。与人类一样，Npt2b-/- 动物会出现轻度肺纤维化，并出现明显的肺纤维化。 限制性生理缺陷。我们发现微石诱导肺泡表达必需的 破骨细胞因子、mCSF 和 RANKL，用于 POLC 分化和盐酸表达 CTSK 既可以溶解结石，又可以损伤组织。我们还发现，当细石器被采用时 转移到WT动物的肺部，它们附着在巨噬细胞上或被巨噬细胞降解，并被清除 28 天内无残留炎症或纤维化或肺损伤证据。我们注意到高密度渗透 在我们的索引 PAM 患者中，当她接受双磷酸盐治疗时，她的病情进展迅速，并且抗 RANKL 治疗减缓了 Npt2b-/- 小鼠中微石的清除速度。这些数据使我们得出这样的结论：破骨细胞 AM 和 Alv-Mo 的转化是肺对抗微石的主要防御，它可能代表了 对其他颗粒（包括石棉）的刻板反应。事实上，我们发现石棉挑战也 与骨髓细胞中的 TRAP 和 CTSK 表达以及 BAL 和肺组织中的 MNGC 表达相关，并且 最终导致破坏性重塑和肺纤维化。来自石棉但未经盐水处理的 BAL 细胞 当小鼠铺在牛骨切片上时，它们会降解骨骼并从骨基质中释放胶原蛋白碎片， 破骨细胞的特征功能。完全愈合与纤维化对可溶解物质的不同组织反应 （微石）与持久性（石棉）颗粒构成了我们假设的基础，即酸和基质降解 当吸入颗粒无敌时，POLC 产生的酶可能是纤维化的主要驱动因素。测试 我们将确定这个假设的三个目标； 1）肺破骨细胞的基因程序 招募的单核细胞响应石棉的规范，2）单核细胞获得的破骨细胞功能 和巨噬细胞在石棉攻击下，3）POLCs在石棉诱发的发病机制中的作用 肺纤维化。该提案直接解决了与进行性肺纤维化相关的机制， 从 20 世纪 30 年代到 20 世纪 越南、海湾和阿富汗战争，并寻求开发支持抗破骨作用的临床前证据 用于治疗石棉沉滞症的策略，例如狄诺塞麦的双磷酸盐。