Activation and differentiation of CD4+ T cells by HEL-specific B-cells exposed to HEL-VLPs. CD4+ T-cells from non-immunized (naive) or GagPol immunized (Ad-Hgp) mice were co-cultured with naïve B-cells from SW-HEL mice and splenic DCs in the presence of HEL-VLPs or control VLPs lacking HEL. (A) After one day, cells were stained for CD4 and the CD-154 activation marker. The experiment was performed three times with similar results, data of one representative experiment are shown. (B) After 6 days of incubation, cells were stained for CD4, intracellular BCL-6, and CXCR-5. Follicular-helper cells were defined by co-expression of BCL-6 and CXCR-5. Numbers in upper right corners indicate mean percentage ± standard deviation of follicular-helper cells among total CD4+ T cells from three independent experiments. Each of the three experiments was performed in duplicates and the mean of the duplicates was used for statistical analysis of the mean of the three experiments. P values of unpaired t-tests for the indicated group comparisons are shown.