Gating strategy of CD4+ T cell subsets. The gating strategy (gating result from a representative female RA patient) was as follows: (a) singlet PBMCs were gated for lymphocytes and then further gated for CD4+ T cells. CD4+ cells where then divided into naïve (CD45RA+) and memory (CD45RAneg) subsets. From naïve cells, CCR4negCCR6negCXCR3neg cells were defined as Th0. Memory cells were divided into four subsets based on CCR4 and CCR6 expression, each of which was the further divided based on CXCR3 expression; Th1 (CCR4negCCR6negCXCR3+), Th2 (CCR4+CCR6negCXCR3neg), CXCR3+Th2 (CCR4+CCR6negCXCR3+), Th17 (CCR4+CCR6+CXCR3neg), CXCR3+Th17 (CCR4+CCR6+CXCR3+), Th1Th17 (CCR4negCCR6+CXCR3+), and CCR6+ only (CCR4negCCR6+CXCR3neg). (b) The cutoff for CTLA-4 positivity on CD4+ T cells were determined using fluorescence minus one (FMO) and cutoff for FOXP3 positivity in CD4+ cells was based on FOXP3 expression in CD25neg gated CD4+ cells. (c) Regulatory T cells (Tregs) were defined by CD25+CD127low expression, while the remaining cells were defined as non-Tregs. CXCR5+ Tregs were defined as follicular regulatory T cells (TFregs) and CXCR5+ non-Tregs as follicular helper T cells (TFh).