Macrophages drive Tfh cell development through a cell-cell contact-dependent mechanism.

<p>(A, B) In a transwell system, macrophages from infected mice were cultured in triplicate wells in the lower chambers. CD4⁺ T cells from normal mice were cultured in the upper chambers. Expression of CXCR5 versus PD-1 on CD4⁺ T cells (gated as CD3⁺CD4⁺) after co-culture. Data are representative of two independent experiments. Numbers represent the frequency of the boxed population within the CD4⁺ T cell population. ***, P<0.001 (Student's <i>t</i>-test); (C) Flow cytometry of CD4⁺CXCR5⁺F4/80⁺ doublets in the spleens of <i>S. japonicum</i>-infected mice. Data are representative of three experiments with three mice in each group; (D) Gating strategy defining CD4⁺CXCR5⁺F4/80⁻ singlet cells (R3), CD4⁻CXCR5⁻F4/80⁺ singlet cells (R2) or CD4⁺CXCR5⁺F4/80⁺ doublets (R4), (<i>D right</i>). Size (forward scatter (FSC)) of CD4⁺CXCR5⁺F4/80⁻ singlet cells, CD4⁻CXCR5⁻F4/80⁺ singlet cells or CD4⁺CXCR5⁺F4/80⁺ doublets in the spleens of <i>S. japonicum</i>-infected mice (<i>D left</i>). Data are representative of three experiments. (E) Flow cytometry of CD3⁺CD4⁺CXCR5⁺F4/80⁺ doublets sorted from the pooled spleens from three <i>S. japonicum</i>-infected mice and then treated with 2 mM EDTA. Data are representative of three independent experiments.