Background/Purpose: Systemic juvenile idiopathic arthritis (SJIA) is a severe and distinct subtype of childhood arthritis. Children with SJIA are at risk for macrophage activation syndrome (MAS), a life-threatening episode of hyperinflammation driven by interferon-gamma (IFNγ). Previous work has suggested that monocytes in SJIA display hyperresponsiveness to IFNγ, 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.

Methods: Bulk RNA-sequencing (RNA-seq) was performed on purified monocytes from 26 patients with SJIA without overt MAS. In addition, single-cell RNA-seq was performed on isolated bone marrow macrophages (BMM). THP-1 monocytic cells were transfected with TRIM8-specific small-interfering RNA (siRNA) prior to stimulation with IFNγ.

Results: Bulk RNA-seq of purified SJIA monocytes revealed marked transcriptional changes between cells from patients with high vs low serum ferritin levels including upregulated gene pathways Response to External Stimulus (p=2.73×10^-17) Defense Response (p=2.66×10^-14), and Inflammatory Response (p=1.95×10^-11). When comparing the SJIA monocyte signature to well-characterized polarization phenotypes, we identified little evidence of IFNγ-induced signature but substantial overlap with multiple polarization states, reflecting either a mixed phenotype or multiple distinct cell populations. Interestingly, among the most highly upregulated genes in SJIA monocytes was tripartite motif containing 8 (TRIM8), an E3 ubiquitin-ligase involved in activation of IFNγ through promoting degradation of the suppressor of cytokine signaling 1 (SOCS1). Elevated TRIM8 expression was found in monocytes from both active and inactive SJIA patients, with the highest levels in those with subclinical MAS (n=3). Furthermore, we utilized scRNA-seq to determine gene expression profiles of BMM from a patient with subclinical MAS. This single cell approach identified a distinct subpopulation of BMM which exhibited markedly altered transcriptional profiles, with the most significantly upregulated pathways being cellular response to IFNγ (p=1.35e-14), endocytic vesicle membranes (p=8.44E-14), and phagosome (p=2.98e-9). These BMM also showed significantly increased expression of TRIM8 (6.4-fold), IFNAR1 (10.5-fold), and IFNGR2 (13.8-fold). To confirm the role of TRIM8 in augmenting macrophage responses to IFNγ, TRIM8 expression was knocked-down in THP-1 using siRNA. TRIM8 knock-down macrophages showed significant reductions in both early (4 hour) and late (24-48 hours) response to IFNγ, as determined by production of CXCL9, a biomarker for MAS activity in both mouse models and patients.

Conclusion: Peripheral monocytes in SJIA display markers of multiple polarization states, while during MAS BMM demonstrate a clear IFNγ response phenotype. TRIM8 is highly expressed in both monocytes and macrophages in SJIA, and in vitro knockdown of TRIM8 reduces macrophage IFNγ response. These data provide a molecular mechanism for monocyte hyperresponsiveness to IFNγ in SJIA, as well as a novel therapeutic target for MAS.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/monocyte-and-macrophage-transcriptional-phenotypes-in-systemic-juvenile-idiopathic-arthritis-reveal-trim8-as-a-mediator-of-ifn%ce%b3-hyperresponsiveness-and-risk-for-macrophage-activation-syndrome/