Antifibrotic Drug Nintedanib Inhibits CSF1R to Promote IL-4-associated Tissue Repair Macrophages.

Profibrotic and prohomeostatic macrophage phenotypes remain ill-defined, both in vivo and in vitro, impeding the successful development of drugs that reprogram macrophages as an attractive therapeutic approach to manage fibrotic disease. The goal of this study was to reveal profibrotic and prohomeostatic macrophage phenotypes that could guide the design of new therapeutic approaches targeting macrophages to treat fibrotic disease. This study used nintedanib, a broad kinase inhibitor approved for idiopathic pulmonary fibrosis, to dissect lung macrophage phenotypes during fibrosis-linked inflammation by combining in vivo and in vitro bulk and single-cell RNA-sequencing approaches. In the bleomycin model, nintedanib drove the expression of IL-4/IL-13-associated genes important for tissue regeneration and repair at early and late time points in lung macrophages. These findings were replicated in vitro in mouse primary bone marrow-derived macrophages exposed to IL-4/IL-13 and nintedanib. In addition, nintedanib promoted the expression of IL-4/IL-13 pathway genes in human macrophages in vitro. The molecular mechanism was connected to inhibition of the colony stimulating factor 1 (CSF1) receptor in both human and mouse macrophages. Moreover, nintedanib counterbalanced the effects of TNF on IL-4/IL-13 in macrophages to promote expression of IL-4/IL-13-regulated tissue repair genes in fibrotic contexts in vivo and in vitro. This study demonstrates that one of nintedanib's antifibrotic mechanisms is to increase IL-4 signaling in macrophages through inhibition of the CSF1 receptor, resulting in the promotion of tissue repair phenotypes.

Application of human iPSC-derived macrophages in a miniaturized high-content-imaging-based efferocytosis assay.

Macrophages play a pivotal role in drug discovery due to their key regulatory functions in health and disease. Overcoming the limited availability and donor variability of human monocyte-derived macrophages (MDMs), human induced pluripotent stem cell (iPSC)-derived macrophages (IDMs) could provide a promising tool for both disease modeling and drug discovery. To access large numbers of model cells for medium- to high-throughput application purposes, an upscaled protocol was established for differentiation of iPSCs into progenitor cells and subsequent maturation into functional macrophages. These IDM cells resembled MDMs both with respect to surface marker expression and phago- as well as efferocytotic function. A statistically robust high-content-imaging assay was developed to quantify the efferocytosis rate of IDMs and MDMs allowing for measurements both in the 384- and 1536-well microplate format. Validating the applicability of the assay, inhibitors of spleen tyrosine kinase (Syk) were shown to modulate efferocytosis in IDMs and MDMs with comparable pharmacology. The miniaturized cellular assay with the upscaled provision of macrophages opens new routes to pharmaceutical drug discovery in the context of efferocytosis-modulating substances.

Senescent cells suppress macrophage-mediated corpse removal via upregulation of the CD47-QPCT/L axis.

Progressive accrual of senescent cells in aging and chronic diseases is associated with detrimental effects in tissue homeostasis. We found that senescent fibroblasts and epithelia were not only refractory to macrophage-mediated engulfment and removal, but they also paralyzed the ability of macrophages to remove bystander apoptotic corpses. Senescent cell-mediated efferocytosis suppression (SCES) was independent of the senescence-associated secretory phenotype (SASP) but instead required direct contact between macrophages and senescent cells. SCES involved augmented senescent cell expression of CD47 coinciding with increased CD47-modifying enzymes QPCT/L. SCES was reversible by interfering with the SIRPα-CD47-SHP-1 axis or QPCT/L activity. While CD47 expression increased in human and mouse senescent cells in vitro and in vivo, another ITIM-containing protein, CD24, contributed to SCES specifically in human epithelial senescent cells where it compensated for genetic deficiency in CD47. Thus, CD47 and CD24 link the pathogenic effects of senescent cells to homeostatic macrophage functions, such as efferocytosis, which we hypothesize must occur efficiently to maintain tissue homeostasis.