Session Type: ACR Concurrent Abstract Session
Session Time: 9:00AM-10:30AM
Background/Purpose: Defining the precise CD4+ T cell subsets that are dysregulated in RA patients is critical to deciphering pathogenesis. Here we present Mixed effects modeling of Associations of Single Cells (MASC), a novel reverse single cell association strategy to determine if a cellular subpopulation is associated with case-control status while controlling for technical confounders and biological covariates. We applied MASC to identify T cell populations expanded in rheumatoid arthritis (RA).
Methods: Cryopreserved PBMCs from 26 RA and 26 osteoarthritis patients were analyzed by a 32-marker mass cytometry panel. All RA patients met ACR 2010 Rheumatoid Arthritis classification criteria. We identified 19 CD4+ T cell populations and tested for case-control associations using our novel association testing method, MASC. MASC uses a logistic regression mixed effects model where the cluster membership of each single cell is the dependent variable. The model estimates fixed effects for donor age, sex, and case-control status, and random effects for each donor and technical batch.
Results: MASC revealed a significantly expanded population of CD4+ T cells, identified as CD27- HLA-DR+ effector memory cells, in RA patients (OR = 1.7; p = 1.1×10-3, Figure 1A). These cells comprised ~2% of CD4+ memory T cells in RA samples compared to 0.8% in OA controls. Compared to peripheral blood, synovial fluid and synovial tissue samples from RA patients were significantly enriched for CD27- HLA-DR+ cells, which comprised ~10-15% of synovial CD4+ T cells (p < 1×10-3, Figure 1B). The abundance of CD4+ CD27Ð HLA-DR+ cells decreased in RA patients who responded to immunosuppressive therapy (p = 0.006). Considered together, mass cytometry and flow cytometry analyses indicated that CD27- HLA-DR+ cells were significantly associated with RA (meta-analysis p = 4.8×10-5). CD27- HLA-DR+ cells express a distinctive effector memory transcriptomic program with Th1- and cytotoxicity-associated features and produce abundant IFN-g and granzyme A upon stimulation (Figure 1C).
Conclusion: MASC is a sensitive and well-calibrated statistical method for analyzing high-dimensional mass cytometry data. We used it in an RA case-control study to identify a novel disease-associated CD4+ T cell population. The expansion of a unique CD27- HLA-DR+ effector memory population in RA periphery may represent disease-driving cells and could be ideal for defining the antigenic repertoire in RA. We propose that MASC is a broadly applicable method to perform association testing with single cell data and can help identify other cellular populations that are critical to rheumatic disease pathogenesis.
To cite this abstract in AMA style:Fonseka C, Rao D, Teslovich N, Korsunsky I, Hannes S, Slowikowski K, Gurish M, Donlin LT, Lederer JA, Weinblatt M, Massarotti E, Coblyn J, Helfgott S, Todd DJ, Bykerk VP, Karlson E, Ermann J, Lee YC, Brenner M, Raychaudhuri S. Single Cell Association Testing Identifies an Expanded Th1-Skewed Cytotoxic Effector CD4+ T Cell Subset in Rheumatoid Arthritis [abstract]. Arthritis Rheumatol. 2018; 70 (suppl 10). https://acrabstracts.org/abstract/single-cell-association-testing-identifies-an-expanded-th1-skewed-cytotoxic-effector-cd4-t-cell-subset-in-rheumatoid-arthritis/. Accessed January 20, 2021.
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/single-cell-association-testing-identifies-an-expanded-th1-skewed-cytotoxic-effector-cd4-t-cell-subset-in-rheumatoid-arthritis/