Monocyte and Macrophage Transcriptional Phenotypes in Systemic Juvenile Idiopathic Arthritis Reveal TRIM8 as a Mediator of IFN-? Hyper-responsiveness and Risk for Macrophage Activation Syndrome

Systemic juvenile idiopathic arthritis (SJIA) confers high risk for macrophage activation syndrome (MAS), a life-threatening episode of hyperinflammation driven by IFN-?. Monocytes in SJIA display IFN-??hyperresponsiveness, but the molecular basis of this remains unclear. The objective of this study is to identify monocyte and macrophage polarization phenotypes including features of interferon response. Bulk RNA-seq of purified SJIA monocytes revealed marked transcriptional changes in patients with elevated ferritin levels. We identified substantial overlap with multiple polarization states but little evidence of IFN-induced signature. Interestingly, among the most highly upregulated genes was tripartite motif containing 8 (TRIM8), a positive regulator of IFN-? signaling. Single cell RNA-seq of bone marrow macrophages (BMM) from a patient with SJIA and early MAS identified a distinct subpopulation of BMM with altered transcriptomes consistent with hemophagocytes, including upregulated IFN-? response pathways. These BMM also showed significantly increased expression of TRIM8. In vitro knock-down of TRIM8 in macrophages caused significant reductions in IFN-? responsiveness. In conclusions, we identify a clear IFN-? response phenotype in BMM during MAS. TRIM8 is upregulated in both monocytes and macrophages in SJIA, and required for macrophage IFN-? response in vitro, providing a molecular mechanism and novel therapeutic for monocyte hyperresponsiveness to IFN-?. Overall design: Leftover BM aspirates were obtained from patients where no hematologic or oncologic abnormalities were found (control BM) or from patient with suspected SJIA. After red cell lysis, BM cells were stored at -80C in freezing media until sequencing. After thawing, macrophage single cell suspensions were obtained using cell sorting for live (7AAD-) populations expressing the monocyte and macrophage surface markers CD14 and CD163 while excluding cells expressing the granulocyte/monocyte marker CD15. This suspension was then loaded onto the Fluidigm C1 Single-Cell Auto Prep System, which captures up to 96 single cells per chip. These cells were subsequently lysed, and extracted RNA converted into cDNA and sequenced as a pooled library using the Illumina platform, providing approximately 3.5-4 million reads per single cell.