Miranda G. Gurra

Rheumatoid arthritis (RA) is a systemic autoimmune disease currently with no universally highly effective prevention strategies. Identifying pathogenic immune phenotypes in at-risk populations prior to clinical onset is crucial to establishing effective prevention strategies. Here, we applied multimodal single-cell technologies (mass cytometry and CITE-Seq) to characterize the immunophenotypes in blood from at-risk individuals (ARIs) identified through the presence of serum antibodies against citrullinated protein antigens (ACPAs) and/or first-degree relative (FDR) status, as compared with patients with established RA and people in a healthy control group. We identified significant cell expansions in ARIs compared with controls, including CCR2+CD4+ T cells, T peripheral helper (Tph) cells, type 1 T helper cells, and CXCR5+CD8+ T cells. We also found that CD15+ classical monocytes were specifically expanded in ACPA-negative FDRs, and an activated PAX5lo naive B cell population was expanded in ACPA-positive FDRs. Further, we uncovered the molecular phenotype of the CCR2+CD4+ T cells, expressing high levels of Th17- and Th22-related signature transcripts including CCR6, IL23R, KLRB1, CD96, and IL22. Our integrated study provides a promising approach to identify targets to improve prevention strategy development for RA.

Introduction Macrophage function is determined by microenvironment and origin. Brain and retinal microglia are both derived from yolk sac progenitors, yet their microenvironments differ. Utilizing single-cell RNA sequencing (scRNA-seq) data from mice, we tested the hypothesis that retinal and brain microglia exhibit distinct transcriptional profiles due to their unique microenvironments. Methods Eyes and brains from 2-4 month wildtype mice were combined (20 eyes; 3 brains) to yield one biologically diverse sample per organ. Each tissue was digested into single cell suspensions, enriched for immune cells, and sorted for scRNA-seq. Analysis was performed in Seurat v3 including clustering, integration, and differential expression. Multi-parameter flow cytometry was used for validation of scRNA-seq results. Lymphocytic choriomeningitis virus (LCMV) Clone 13, which produces a systemic, chronic, and neurotropic infection, was used to validate scRNA-seq and flow cytometry results in vivo . Results Cluster analysis of integrated gene expression data from eye and brain identified 6 Tmem119 + P2ry12 + microglial clusters. Differential expression analysis revealed that eye microglia were enriched for more pro-inflammatory processes including antigen processing via MHC class I (14.0-fold, H2-D1 and H2-K1 ) and positive regulation of T-cell immunity (8.4-fold) compared to brain microglia. Multi-parameter flow cytometry confirmed that retinal microglia expressed 3.2-fold greater H2-Db and 263.3-fold more H2-Kb than brain microglia. On Day 13 and 29 after LCMV infection, CD8 + T-cell density was greater in the retina than the brain. Discussion Our data demonstrate that the microenvironment of retina and brain differs, resulting in microglia-specific gene expression changes. Specifically, retinal microglia express greater MHC class I by scRNA-seq and multi-parameter flow cytometry, resulting in a possibly enhanced capability to stimulate CD8 + T-cell inflammation during LCMV infection. These results may explain tissue-specific differences between retina and brain during systemic viral infections and CD8 + T-cell driven autoimmune disease.

TR-MCs. These findings show a facet of mononuclear cell biology that could be imperative to understanding tissue-resident myeloid cell function in RA.

BACKGROUND: Cord blood (CB) leptin is positively associated with adiposity at birth, but the association with child adiposity is unclear. OBJECTIVES: We hypothesized that CB leptin is positively associated with adiposity in peripubertal children and with childhood leptin. METHODS: Leptin was measured in 986 CB and 931 childhood stored samples from a prospective birth cohort. Adiposity measures were collected at birth and mean age 11.5 years. Linear and logistic regression analyses were used to evaluate associations between log-transformed CB leptin and neonatal and childhood adiposity measures as continuous and categorical variables, respectively. RESULTS: CB leptin was positively associated with neonatal and childhood adiposity. Childhood associations were attenuated when adjusted for maternal body mass index (BMI) and glucose, but remained statistically significant for childhood body fat percentage (β = 1.15%, confidence interval [CI] = 0.46-1.84), body fat mass (β = 0.69 kg, 95% CI = 0.16-1.23), sum of skin-folds (β = 1.77 mm, 95% CI = 0.31-3.24), log-transformed child serum leptin (β = 0.13, 95% CI = 0.06-0.20), overweight/obesity (OR = 1.21, 95% CI = 1.03-1.42), obesity (OR = 1.31, 95% CI = 1.04-1.66) and body fat percentage >85th percentile (OR = 1.38, 95% CI = 1.12-1.73). Positive associations between newborn adiposity measures and CB leptin confirmed previous reports. CONCLUSION: CB leptin is positively associated with neonatal and childhood adiposity and child leptin levels, independent of maternal BMI and maternal hyperglycemia. CB leptin may be a biomarker of future adiposity risk.

Macrophages are a pivotal cell type within the synovial lining and sub-lining of the joint, playing a crucial role in maintaining homeostasis of synovium. Although fate-mapping techniques have been employed to differentiate synovial macrophages from other synovial myeloid cells, no comprehensive study has yet been conducted within the mouse synovial macrophage compartment. In this study, we present, for the first time, lineage tracing results from 18 myeloid-specific fate-mapping models in mouse peripheral blood (PB) and synovial tissue. The identification of synovial macrophages and monocyte-lineage cells through flow cytometry was further validated using cellular indexing of transcriptomes and epitopes by sequencing (CITE-seq) datasets. These findings provide a valuable methodological tool for researchers to select appropriate models for studying the function of synovial myeloid cells and serve as a reference for investigations in other tissue types.