Identification of a broadly fibrogenic macrophage subset induced by type 3 inflammation

Abstract

Macrophages are central orchestrators of the tissue response to injury, with distinct macrophage activation states playing key roles in fibrosis progression and resolution. Identifying key macrophage populations found in human fibrotic tissues could lead to new treatments for fibrosis. Here, we used human liver and lung single-cell RNA sequencing datasets to identify a subset of CD9 + TREM2 + macrophages that express SPP1 , GPNMB , FABP5 , and CD63 . In both human and murine hepatic and pulmonary fibrosis, these macrophages were enriched at the outside edges of scarring and adjacent to activated mesenchymal cells. Neutrophils expressing MMP9, which participates in the activation of TGF-β1, and the type 3 cytokines GM-CSF and IL-17A coclustered with these macrophages. In vitro, GM-CSF, IL-17A, and TGF-β1 drive the differentiation of human monocytes into macrophages expressing scar-associated markers. Such differentiated cells could degrade collagen IV but not collagen I and promote TGF-β1–induced collagen I deposition by activated mesenchymal cells. In murine models blocking GM-CSF, IL-17A or TGF-β1 reduced scar-associated macrophage expansion and hepatic or pulmonary fibrosis. Our work identifies a highly specific macrophage population to which we assign a profibrotic role across species and tissues. It further provides a strategy for unbiased discovery, triage, and preclinical validation of therapeutic targets based on this fibrogenic macrophage population.

Reciprocal inflammatory signals establish profibrotic cross-feeding metabolism

“…Interestingly, analysis of a published scRNAseq time course following bleomycin lung injury 18 showed maximal Arg1 in cells annotated to be similar to the C2 transitional macrophage compartment we previously demonstrated to localize to the fibrotic niche after injury 4 ( Figure 2c ; Figure S3b ). C2 cells not only expressed higher Arg1 but also expressed higher levels of the Fab5 markers that were recently found to be associated with pro-fibrotic macrophages in both mouse and human fibrotic lung and liver datasets 19 ( Figure S3b ). Similar to our previous findings for these transitional cells 4 , Arg1+ cells were found in proximity to clusters of activated fibroblasts that form after injury ( Figure 2d ), and flow cytometry confirmed Arg1 expression in CD11b+CD64+ macrophages as opposed to monocytes, alveolar macrophages, or dendritic cells ( Figure S4 ).…”

“…To further test the clinical significance of Arg1 + macrophages, we performed scRNAseq analysis of aged mice compared to young given that aging is a major risk factor for the development of lung fibrosis in patients. Remarkably, Arg1 + macrophages were not only greatly increased in bleomycin-injured aged mice but also co-expressed the Fab5 profibrotic macrophage genes 19 ( Figure 3c ; Fig S5b ). We then labeled clinical lung samples from IPF by immunofluorescence.…”

“…Cells expressing Arg1 were defined as cells having >1 counts after scaling and centering 44 . Cells expressing Fab5 genes were defined as cells whose average expression of Trem2 , Cd9 , Spp1 , Gpnmb , and Fabp5 19 was >1 after scaling and centering. N=2 mice per condition.…”

“…(b) Top: Feature plot of macrophages from Strunz et al 18 with Arg1 and Spp1 expression as well as Fab5 profibrotic macrophage genes 19 indicated. Bottom: proportions of expression by C1, C2, or C3 annotation.…”

“…(b) Dot plot of Fab5 19 gene expression in old and young mouse lung macrophages after bleomycin injury (corresponding to Figure 3c). UI=uninjured.…”