Abstract Rationale Chrysotile asbestos inhalation drives a progressive fibrotic response in the lung, yet the cellular programs that mediate injury progression remain incompletely defined. Recent work has identified pulmonary osteoclast-like cells (POLCs) as rare, macrophage-derived effector cells that emerge in particulate lung injury and promote matrix remodeling. Whether POLCs are induced by asbestos and contribute to asbestosis pathogenesis has not been established. Here, we combined single-cell transcriptomics with histologic, molecular, and physiologic readouts to define asbestos-induced POLCs and test whether their modulation alters disease severity. Methods C57BL/6J mice received intratracheal instillation of chrysotile asbestos, and replicate whole lungs were collected at follow-up timepoints from Day 0 (control) to Day 56 post-instillation. Pulmonary function testing, lung histology, and qPCR for osteoclast-associated genes were performed in parallel. Whole-lung single-nucleus RNA sequencing was used to model longitudinal transcriptional dynamics across the injury time course. In a separate cohort, whole-lung single-cell RNA sequencing was performed to characterize cell-specific responses to treatment with zoledronic acid, a bisphosphonate that inhibits osteoclast activity. Monocle3 supported dimensionality reduction, cell state annotation, pathway analysis, and pseudotime modeling, and a beta-binomial model quantified relative changes in cell type proportions. Results Asbestos exposure results in a progressive decline in pulmonary function and produces histologic evidence of early inflammatory infiltration and fibrotic deposition by Day 28. scRNA-seq identifies more than 30 lung cell states, with strong early expansion of myeloid populations. Alveolar macrophages show increased expression of canonical osteoclast-associated genes, including Acp5, Ctsk, and Atp6v0d2, which rise sharply by Day 7 and persist through later timepoints. qPCR confirms upregulation of osteoclast-related transcripts, acquisition of capacity for bone pitting and protein staining for TRAP5b and CTSK demonstrates robust induction of osteoclast programs. TRAP+ multinucleated cells are significantly increased in BALF from asbestos-exposed mice. Pseudotime analyses indicate differentiation of POLCs along a trajectory marked by acquisition of bone-resorption, matrix-remodeling, and catabolic signatures. Intraperitoneal (i.p.) treatment with the bisphosphonate, zoledronic acid, reduced POLC abundance, and attenuated parenchymal fibrosis due to asbestos based on multiple gene expression, histologic, and physiologic measures relative to i.p. vehicle control, supporting a functional role for POLCs in disease progression. Conclusions Chrysotile asbestos induces the emergence of pulmonary osteoclast-like cells that arise from macrophage lineages and display robust osteoclast gene programs. Pharmacologic inhibition of POLCs reduces lung fibrosis, identifying POLCs as a potential therapeutic target in asbestosis. This abstract is funded by: I01 BX005128-01a (VA Merit)
Franks et al. (Fri,) studied this question.