Single-cell RNA sequencing in juvenile idiopathic arthritis

Abstract

Juvenile idiopathic arthritis (JIA) is one of the most common chronic inflammatory rheumatic diseases in children, with onset before age 16 and lasting for more than 6 weeks. JIA is a highly heterogeneous condition with various consequences for health and quality of life. For some JIA patients, early detection and intervention remain challenging. As a result, further investigation of the complex and unknown mechanisms underlying JIA is required. Advances in technology now allow us to describe the biological heterogeneity and function of individual cell populations in JIA. Through this review, we hope to provide novel ideas and potential targets for the diagnosis and treatment of JIA by summarizing the current findings of single-cell RNA sequencing studies and understanding how the major cell subsets drive JIA pathogenesis.

Keywords: Juvenile idiopathic arthritis; Macrophage; Macrophage activation syndrome; Monocyte; Single-cell RNA sequencing; Synoviocyte; T cell.

Figure 1

Flow chart of single-cell transcriptome sequencing. Its process focuses on examining isolated cells for quality control, library construction, data analysis, and visualization to reveal the characteristics of cell populations and functions. The figure was drawn by utilizing Figdraw software.

Figure 2

Identification of monocytes or macrophages in sJIA patients and a mouse model of MAS by using flow cytometry combined with scRNA-seq. (A) The mechanisms associated with bone marrow-derived macrophages in a patient with sJIA-MAS. IFN-γ induction possibly regulates the JAK-STAT1 signaling pathway via TRIM8. AMϕ expressed markers are related to intracellular granule movement, cytokine response, and intrinsic immune response. The overexpression of the AHRR and AIP genes may down-regulate the AHR pathway, which is associated with MAS transition (B) The immune response in blood monocytes and AMϕ in a mouse model of MAS induced by TLR9. The acute MAS model mice showed high expression of inflammatory factors in serum, lung tissues, and lavage fluid. AMϕs were mainly classically activated. Flow cytometry identified a unique AMϕ phenotype in BAL fluid, namely, CD11c⁺ CD11b^variable CD64⁺ AMϕs. ScRNA-seq identified two distinct subpopulations of MAS AMϕ in lung tissues and showed extensive high expression of adaptive immunity and persistent low expression of hypoxic genes. The acute MAS model mice exhibited M2 polarization after remission, but IL-18, IL-12, and RANTES expression was barely affected, suggesting an incomplete effect of IFN-γ blockers. In recurrent MAS model mice, reprogrammed AMϕ toward classic polarization activated inflammatory responses. The red arrows represent up-regulation. The green arrows represent down-regulation. A short straight line indicates an inhibitory or blocking effect. A tapering arrow at the end indicates remission-related features. The figure was drawn by utilizing Figdraw software. AHR, aryl hydrocarbon receptor; AHRR, aryl hydrocarbon receptor repressor; AIP, aryl hydrocarbon receptor interacting protein; AMϕ, alveolar macrophage; ARNT, aryl hydrocarbon receptor nuclear translocator; BAL, bronchoalveolar lavage; CCR5, C–C motif chemokine receptor 5; CXCL9, C-X-C motif chemokine ligand 9; CXCL10, C-X-C motif chemokine ligand 10; IL-1β, interleukin-1β; IL-6, interleukin-6; IL-10, interleukin-10; IL-12, interleukin-12; IL-12p70, interleukin-12p70; IL-17, interleukin-17; IL-18, interleukin-18; IFN-γ, interferon-gamma; JAK, Janus kinase; KLF13, Kruppel-like factor 13; MAFB, MAF BZIP transcription factor B; MHC II, major histocompatibility complex II; NFKB1, nuclear factor Kappa B subunit 1; P, phosphorylation; STXBP2, syntaxin binding protein 2; STAT1, signal transducer and activator of transcription 1; RANTES, regulated upon activation, normal T-cell expressed and presumably secreted; SOCS1, suppressor of cytokine signaling 1; TGFβ, transforming growth factor-beta; TLR9, Toll-like receptor 9; TNF, tumor necrosis factor; TRIM8, tripartite motif containing 8.

Figure 3

Description of T-cell subpopulations in SF at the single-cell level. The figure demonstrates a predominantly Th1 phenotype in oJIA SF. CD4⁺, CD8⁺, and γδ T cells mainly expressed IFN-γ and CXCR3 but not IL-17, suggesting classic Th1 polarization. Although CD4⁺ Tmem cells expressed the surface markers IL-17 and CD161, these cells did not significantly secrete IL-17. IFN-γ⁺ CXCR3⁺ CD4⁺ Tmem cells cannot be accurately identified due to experimental factors. Single-cell sequencing detected CD4+ Treg cells in SF that also had a Th1 phenotype, but the presence of anti-inflammatory and pro-inflammatory properties suggested heterogeneity of the disease and immune response, while the presence and function of unstable Treg cells were unknown. In addition, a group of Tph-like phenotype cells in CD4⁺ Teff cells may be involved in the disease process in ANA-positive patients, but the relationship between B cells and disease severity is also unknown. The figure was drawn by using Figdraw software. BATF, basic leucine zipper ATF-like transcription factor; CCR6, C–C motif chemokine receptor 6; CTLA4, cytotoxic T-lymphocyte associated protein 4; CXCR3, C-X-C motif chemokine receptor 3; FOXP3, forkhead box P3; ICOS, inducible T-cell costimulator; IFNG, interferon gamma; IKZF2, IKAROS family zinc finger 2; IL2RA, interleukin 2 receptor subunit alpha; IL12RB2, interleukin 12 receptor subunit beta 2; KLRB1, killer cell lectin-like receptor B1; MAF, MAF BZIP transcription factor; PMDR1, PR/SET domain 1; RORC, RAR related orphan receptor C; SF, synovial fluid; TBX21, T-box transcription factor 21; Teff, effector T cell; Tmem, memory T cell; Treg, regulatory T cell; TNFRSF18, TNF receptor superfamily member 18.