Background CD4+ T cells are important mediators of inflammation in rheumatoid arthritis; however, the specific CD4+ T cell populations most important in driving disease pathology remain unclear. CD4+ T cells are often divided into subsets based on effector functions (e.g. Th1, Th2, Th17). Here we describe an alternative strategy of defining T cell subpopulations in blood and synovial fluid based on differential expression of integrins and other migratory receptors that help cells localise to specific target tissues. Blockade of integrin-dependent migration is already employed clinically; therefore, understanding patterns of migratory receptor expression that allow infiltration into the joint and other target organs is of significant interest.
Methods We developed multiparametric flow cytometry panels to characterise expression of integrins and other migratory receptors on blood and synovial fluid CD4+ T cells. Dimensional reduction was performed using Spanning tree Progression of Density normalised Events (SPADE) to identify CD4+ T cell subpopulations that express specific combinations of migratory receptors.
Results More than half of circulating memory CD4+ T cells coordinately express α3, α5, and α6 integrins. α4 integrin is expressed on ˜25%–50% of circulating memory CD4+ T cells, but in a distinct pattern such that memory CD4+ T cells can be divided into 3 groups based on the differential expression of α4 and α6 integrin (α4+α6-, α4-α6+, α4+α6+). Both central memory and effector memory CD4+ T cell subsets contain these populations, while naïve T cells lack significant expression of either α chain. Interestingly, both CD4+CD25+CD127- regulatory T cells and CD4+ CLA+’skin-homing’ cells fall predominantly within the α4-α6+ population. Both α4- α6+ and α4+α6+ cells co-express β1, while α4+α6- cells co-express β7 rather than β1. α4β7+ cells constitute ˜10%–20% of circulating memory CD4+ T cells; however, these cells are rare in inflammatory synovial fluid, in which almost all CD4+ T cells express β1. Small populations of circulating memory CD4+ cells also express α1, α2, αV, and αE integrins and CD146, with certain subsets substantially enriched in synovial fluid. SPADE analyses allowed for visual demonstration of cell subpopulations defined by migratory receptor expression.
Conclusion Differential integrin expression identifies CD4+ T subpopulations in a manner non- redundant with traditional methods of classifying T cells. Specific integrin-defined memory CD4+ subpopulations are enriched in synovial fluid compared to blood, suggesting that certain integrins, in particular β1 integrins, may promote CD4+ T localisation to the joint. Further characterisation of integrin-defined T cell subpopulations that can infiltrate the joint may lend new insights into mechanisms of synovial inflammation.
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