Rationale: The contributions of diverse cell populations in the human lung to pulmonary fibrosis pathogenesis are poorly understood. Single-cell RNA sequencing can reveal changes within individual cell populations during pulmonary fibrosis that are important for disease pathogenesis. Objectives: To determine whether single-cell RNA sequencing can reveal disease-related heterogeneity within alveolar macrophages, epithelial cells, or other cell types in lung tissue from subjects with pulmonary fibrosis compared with control subjects. Methods: We performed single-cell RNA sequencing on lung tissue obtained from eight transplant donors and eight recipients with pulmonary fibrosis and on one bronchoscopic cryobiospy sample from a patient with idiopathic pulmonary fibrosis. We validated these data using in situ RNA hybridization, immunohistochemistry, and bulk RNA-sequencing on flow-sorted cells from 22 additional subjects. Measurements and Main Results: We identified a distinct, novel population of profibrotic alveolar macrophages exclusively in patients with fibrosis. Within epithelial cells, the expression of genes involved in Wnt secretion and response was restricted to nonoverlapping cells. We identified rare cell populations including airway stem cells and senescent cells emerging during pulmonary fibrosis. We developed a web-based tool to explore these data. Conclusions: We generated a single-cell atlas of pulmonary fibrosis. Using this atlas, we demonstrated heterogeneity within alveolar macrophages and epithelial cells from subjects with pulmonary fibrosis. These results support the feasibility of discovery-based approaches using next-generation sequencing technologies to identify signaling pathways for targeting in the development of personalized therapies for patients with pulmonary fibrosis.

Misharin et al. elucidate the fate and function of monocyte-derived alveolar macrophages during the course of pulmonary fibrosis. These cells persisted throughout the life span, were enriched for the expression of profibrotic genes, and their genetic ablation ameliorated development of pulmonary fibrosis.

Ontologically distinct populations of macrophages differentially contribute to organ fibrosis through unknown mechanisms.We applied lineage tracing, single-cell RNA sequencing and single-molecule fluorescence in situ hybridisation to a spatially restricted model of asbestos-induced pulmonary fibrosis.We demonstrate that tissue-resident alveolar macrophages, tissue-resident peribronchial and perivascular interstitial macrophages, and monocyte-derived alveolar macrophages are present in the fibrotic niche. Deletion of monocyte-derived alveolar macrophages but not tissue-resident alveolar macrophages ameliorated asbestos-induced lung fibrosis. Monocyte-derived alveolar macrophages were specifically localised to fibrotic regions in the proximity of fibroblasts where they expressed molecules known to drive fibroblast proliferation, including platelet-derived growth factor subunit A. Using single-cell RNA sequencing and spatial transcriptomics in both humans and mice, we identified macrophage colony-stimulating factor receptor (M-CSFR) signalling as one of the novel druggable targets controlling self-maintenance and persistence of these pathogenic monocyte-derived alveolar macrophages. Pharmacological blockade of M-CSFR signalling led to the disappearance of monocyte-derived alveolar macrophages and ameliorated fibrosis.Our findings suggest that inhibition of M-CSFR signalling during fibrosis disrupts an essential fibrotic niche that includes monocyte-derived alveolar macrophages and fibroblasts during asbestos-induced fibrosis.

Alveolar macrophages orchestrate the response to viral infections. Age-related changes in these cells may underlie the differential severity of pneumonia in older patients. We performed an integrated analysis of single-cell RNA-Seq data that revealed homogenous age-related changes in the alveolar macrophage transcriptome in humans and mice. Using genetic lineage tracing with sequential injury, heterochronic adoptive transfer, and parabiosis, we found that the lung microenvironment drove an age-related resistance of alveolar macrophages to proliferation that persisted during influenza A viral infection. Ligand-receptor pair analysis localized these changes to the extracellular matrix, where hyaluronan was increased in aged animals and altered the proliferative response of bone marrow-derived macrophages to granulocyte macrophage colony-stimulating factor (GM-CSF). Our findings suggest that strategies targeting the aging lung microenvironment will be necessary to restore alveolar macrophage function in aging.

Abstract. Since 2004, an East African genotype of Chikungunya virus (CHIKV) has emerged, causing significant epidemics of an arthralgic syndrome. In addition, this virus has been associated for the first time with neonatal transmission and neurological complications. In the current study, pregnant Rhesus macaques were inoculated with an enzootic or epidemic strain of CHIKV to compare pathogenesis and transplacental transmission potential. Viremias were similar for both strains and peaked at 2-3 days post-inoculation (dpi). Viral RNA was detected at necropsy at 21 dpi in maternal lymphoid, joint-associated, and spinal cord tissues. The absence of detectable viral RNA and the lack of germinal center development in fetuses indicated that transplacental transmission did not occur. Neutralizing antibodies were detected in all dams and fetuses. Our study establishes a non-human primate model for evaluating vaccines and antiviral therapies and indicates that Rhesus macaques could serve as a competent enzootic reservoir.

Graphical Abstract Highlights d Metformin prevents IL-6-dependent thrombosis induced by urban particulate matter d Metformin inhibits mitochondrial complex I to prevent ROSmediated IL-6 release d Metformin inhibits mitochondrial ROS to prevent CRAC channel activation d Mitochondrial ROS and CRAC channel inhibition in vivo prevent thrombosis

Three studies are described that examined the relation between performance-based (PB) feedback delivered via e-mail and preschool teachers' use of recommended practices. The authors conducted the first two studies in the same classroom with different classroom staff. The third study was conducted with three different teachers employed in three separate classrooms. In Study I, the teachers demonstrated initial increases in use of targeted practices; however, the increases did not maintain over time. In Study II, the teachers demonstrated initial increases with some variability across behaviors. In Study III, a functional relation was established for three teachers across three self-selected behaviors, which generalized and maintained. The discussion includes implications for future PD research and practice, and provides recommendations for designing effective PB feedback systems.

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