Memory CD4 T cells express Tfh factors after activation by TSLP-DC. CD4 memory T cells were cultured with DCs, TSLP-DC, or LPS-DC, and cytokines were measured at the indicated days after 24 h of restimulation with anti-CD3/CD28 beads. ELISA assay for IL-21 at day 6 of culture in A and at day 2, 4, and 6 in B. Mean ± SEM for 13 and 5 donors is shown, from four and two independent experiments, respectively. ELISA assay for CXCL13 in C and CBA assay for Th cytokines in D in the same experimental settings as in B. (E and F) Memory CD4 T cells were separated into CXCR5⁺ and CXCR5⁻ cells by FACS sorting, and cultured with DCs or TSLP-DC for 6 d. IL-21 and CXCL13 quantification after 24 h of anti-CD3/CD28 stimulation is shown as mean ± SEM from nine independent experiments. FACS staining for ICOS, PD1, and CXCR5 at day 6 of co-culture. BCL6 was quantified on the CXCR5^hi/PD1^hipopulation. The geometric MFI is plotted for three or two independent experiments. *, P < 0.05; **, P < 0.01; ***, P < 0.001, paired Student’s t test.