A CCL20+ macrophage subpopulation is the major producer of TNF in axial spondyloarthritis synovium and is driven by CD40L and a myeloid autocrine circuit

Objectives Tumour necrosis factor (TNF) and interleukin (IL)-17 are central drivers of axial spondyloarthritis (AxSpA). We previously identified activated CD4+ tissue-resident memory Th17 cells as the principal source of IL-17A/F in AxSpA synovium. We sought to define the cellular source of TNF and mechanisms driving the CCL20 + myeloid phenotype in joint and blood. Methods Single-cell RNA sequencing (scRNA-seq) of synovial tissue from 5 patients with AxSpA was combined with immunofluorescence, synovial-fluid scRNA-seq, and flow cytometry, as well as comparisons with rheumatoid arthritis datasets. Monocyte-derived macrophages from healthy controls (HCs) and patients with AxSpA were cocultured with activated autologous CD4 + T cells, with blocking and cytokine experiments defining mechanisms of CCL20 + induction. Circulating monocytes were assessed by scRNA-seq and functional assays. Results CCL20 + macrophages were the predominant source of TNF in AxSpA synovium, with TNF protein confirmed in tissue and synovial fluid. Activated CD4 + T cells induced the CCL20 + phenotype through CD40L-CD40 signalling and an IL-1β/TNF-α autocrine circuit in HC-derived and AxSpA-derived cells. Baseline AxSpA monocytes showed a higher CCL20 + signature than HC monocytes, but this difference largely disappeared after T-cell-mediated induction. Under stimulation, AxSpA monocytes also produced more TNF, IL-6, IL-1β, and IL-23 than HC monocytes, a difference attenuated following IL-1/TNF priming. Conclusions CCL20 + macrophages are a major CD40L- and IL-1β/TNF-α-driven source of TNF in AxSpA synovium. AxSpA circulating monocytes exhibit heightened basal CCL20 + signatures and inflammatory responsiveness that converge towards HC responses after inflammatory priming, supporting circulating CCL20 + monocytes as a marker of IL-1β/TNF-α-driven myeloid inflammation.