Figure 7. Tfh versus effector Th cells are distinguishable by the second cell division, in association with differential early IL-2Rα expression levels.

Early virus-specific Tfh vs. effector Th cell differentiation was analyzed at 48 hrs after acute LCMV infection. (A) Representative FACS plots of SM CD4⁺ T cells from LCMV infected (“Day 2”) and uninfected (“Naïve”) mice. Gates identify IL2Rα^hiCXCR5^lo vs. IL2Rα^intCXCR5^hi SM CD4⁺ T cells. (B) CXCR5 expression by IL2Rα^hi (black) vs. IL2Rα^int (red) SM CD4⁺ T cells. Isotype control = gray filled histogram. (C) Bcl6 expression by IL2Rα^hi vs. IL2Rα^int SM CD4⁺ T cells. (D) Expression of IL2Rα and CXCR5 by SM CD4⁺ T cells in MD4-µMT recipients, day 2 p.i. (E) Bcl6 expression by SM CD4⁺ T cells in MD4-µMT recipients, day 2 p.i. (F) IL2Rα and Blimp1 expression by Blimp1-YFP SM CD4⁺ T cells in B6 recipients, day 2 p.i. Representative SM CD4⁺ T cell FACS plot and quantitation are shown. (G) IL2R α expression (MFI) by SM CD4⁺ T cell populations gated in (F). (H) Bcl6 expression by SM CD4⁺ T cells in B6 recipients at day 2 p.i. Gates identify IL2Rα^hi vs. IL2Rα^int SM CD4⁺ T cells. Bcl6 protein expression (MFI) by each population was calculated. (I, J) CFSE-labeled SM CD4⁺ T cells were transferred into B6 recipient mice subsequently infected with LCMV. Cell divisions and differentiation were analyzed at day 2 p.i. (I) Cell divisions. Gray filled histogram = SM CD4⁺ T cells in uninfected mice. (J) Representative FACS plots are shown of SM CD4⁺ T cells that have undergone 2 (left) and 3 (right) cell divisions. Gates identify IL2Rα^hiBcl6⁻ and IL2Rα^intBcl6⁺ SM CD4⁺ T cells. Data in all panels are representative of three or more independent experiments; n = 3–4 per group. **P<0.01, ***P<0.001.