Probes for CSF2

Myeloid cell activation by antineutrophil cytoplasmic antibody (ANCA) is pivotal for necrotising vasculitis, including necrotising crescentic glomerulonephritis (NCGN). In contrast to neutrophils, the contribution of classical monocyte (CM) and non-classical monocyte (NCM) remains poorly defined. We tested the hypothesis that CMs contribute to antineutrophil cytoplasmic antibody-associated vasculitis (AAV) and that colony-stimulating factor-2 (CSF2, granulocyte-macrophage colony-stimulating factor (GM-CSF)) is an important monocyte-directed disease modifier.Myeloperoxidase (MPO)-immunised MPO-/- mice were transplanted with haematopoietic cells from wild-type (WT) mice, C-C chemokine receptor 2 (CCR2)-/- mice to abrogate CM, or transcription factor CCAAT-enhancer-binding protein beta (C/EBPβ)-/- mice to reduce NCM, respectively. Monocytes were stimulated with CSF2, and CSF2 receptor subunit beta (CSF2rb)-deficient mice were used. Urinary monocytes and CSF2 were quantified and kidney Csf2 expression was analysed. CSF2-blocking antibody was used in the nephrotoxic nephritis (NTN) model.Compared with WT mice, CCR2-/- chimeric mice showed reduced circulating CM and were protected from NCGN. C/EBPβ-/- chimeric mice lacked NCM but developed NCGN similar to WT chimeric mice. Kidney and urinary CSF2 were upregulated in AAV mice. CSF2 increased the ability of ANCA-stimulated monocytes to generate interleukin-1β and to promote TH17 effector cell polarisation. CSF2rb-/- chimeric mice harboured reduced numbers of kidney TH17 cells and were protected from NCGN. CSF2 neutralisation reduced renal damage in the NTN model. Finally, patients with active AAV displayed increased urinary CM numbers, CSF2 levels and expression of GM-CSF in infiltrating renal cells.CMs but not NCMs are important for inducing kidney damage in AAV. CSF2 is a crucial pathological factor by modulating monocyte proinflammatory functions and thereby TH17 cell polarisation.

Hyperinflammation contributes to lung injury and subsequent acute respiratory distress syndrome (ARDS) with high mortality in patients with severe coronavirus disease 2019 (COVID-19). To understand the underlying mechanisms involved in lung pathology, we investigated the role of the lung-specific immune response. We profiled immune cells in bronchoalveolar lavage fluid and blood collected from COVID-19 patients with severe disease and bacterial pneumonia patients not associated with viral infection. By tracking T cell clones across tissues, we identified clonally expanded tissue-resident memory-like Th17 cells (Trm17 cells) in the lungs even after viral clearance. These Trm17 cells were characterized by a a potentially pathogenic cytokine expression profile of IL17A and CSF2 (GM-CSF). Interactome analysis suggests that Trm17 cells can interact with lung macrophages and cytotoxic CD8+ T cells, which have been associated with disease severity and lung damage. High IL-17A and GM-CSF protein levels in the serum of COVID-19 patients were associated with a more severe clinical course. Collectively, our study suggests that pulmonary Trm17 cells are one potential orchestrator of the hyperinflammation in severe COVID-19.

Maturation of the definitive adrenal cortex occurs between 3 and 6 weeks post-partum and involves onset of CYP11B2 expression and establishment of the laminin-encased 3D structure of glomeruli that contain rosettes of 10 to 15 zona glomerulosa (zG) cells that work in coordination to produce optimal amounts of aldosterone. Although this process is dependent on canonical WNT/b-catenin signaling, cellular sources of WNT ligands remain elusive and the mechanisms involved in the extensive extra-cellular matrix remodeling associated with rosette/glomeruli morphogenesis are unknown. Beyond their role in innate immunity, macrophages are involved in extra-cellular matrix remodeling under a wide variety of pathophysiological conditions and have the capacity to produce WNT ligands. This, together with the presence of macrophages within the zG cells, strongly suggest that macrophages may play a role in zG morphogenesis and differentiation. Supporting this idea, a recent publication has shown that intra-tissular aldosterone concentration was reduced in the absence of macrophages under stress conditions. However, whether macrophages play a direct role in controlling aldosterone secretion or an indirect role by remodelling the postnatal zG is unknown. The presence of tissue resident macrophages in a specific zone is dependent on the production of trophic factors such as IL34, CSF1, CSF2 or CX3CL1 by nearby, tissue resident ‘niche’ cells. In return, macrophages are thought to provide positive ‘feedback’ signals to their niche, generating mutually beneficial circuits between the niche and its macrophages. To gain insight into the role of macrophages in zG morphogenesis and homeostasis, we used single cell sequencing and RNAscope analyses to show expression of CX3CL1 in the zG and of CX3CR1 in macrophages. Interestingly, CX3CL1 expression in the zG was downstream of WNT signalling, suggesting existence of a bi-directional interaction between macrophages and zG. To further study the role of macrophages during the maturation of the zG, we pharmacologically depleted macrophages by the small molecule inhibitor Pexidartinib at different time points between 3- and 12-weeks post-partum, when maturation of the zG occurs. Short-term depletion of macrophages resulted in a more disorganized and elongated zG, suggesting a delay in maturation. However, long-term depletion of macrophages resulted in exacerbated maturation of the rosettes, suggesting that the short-term zG defect was followed by establishment of a compensatory mechanism to allow formation of rosettes even in the absence of macrophages. Whether these perturbations of the zG are correlated with an altered production of aldosterone is still under study.