Figure 1.

Tfr Cells Are Present at Reduced Numbers in Cxcr5^fl/flFoxp3^cre-yfp Mice

Mice were immunized with NP-KLH/alum i.p., and the GC response was analyzed 14 days after immunization.

(A) Histogram of CXCR5 expression in Foxp3⁺CD4⁺ Treg cells, naive T cells as a CXCR5-negative control population, and wild-type B cells as a CXCR5-positive population.

(B) CXCR5 mean fluorescence intensity (MFI; geometric mean) in Foxp3⁺CD4⁺ Treg cells from mice of the indicated genotypes.

(C) Analysis of Tfr and Tfh cells 14 days after influenza A virus (HKx31) infection in Cxcr5^fl/flFoxp3^cre mice and Cxcr5^+/+Foxp3^cre controls. Representative confocal images of splenic cryosections stained for Foxp3 (magenta), Ki67 (blue), CD3 (green), and IgD (orange); Foxp3⁺ cells are indicated by arrows. Scale bar, 40 μm.

(D) Average GC size in square micrometers measured as the IgD⁻Ki67⁺ area. Each dot represents the average size of 2–6 GCs per mouse.

(E) Quantification of the average number of Tfr cells per mouse, defined as CD3⁺Foxp3⁺ cells within the GC, per 5,000 μm². Each dot represents the average number of Tfr cells per 5,000 μm² of GC area per mouse, from 2–6 GCs.

(F) Representative flow cytometry contour plots of CXCR5⁺PD-1⁺ Tfh cells from Foxp3⁻CD4⁺ cells.

(G and H) Quantification of the (G) frequency and (H) absolute number of CXCR5⁺PD-1⁺ Tfh cells.

(I) Representative flow cytometry contour plots of Bcl6⁺PD-1⁺ Tfh cells of Foxp3⁻CD4⁺ cells.

(J and K) Quantification of the (J) percentage and (K) absolute number of Bcl6⁺PD-1⁺Foxp3⁻CD4⁺ Tfh cells.

Each symbol represents one mouse, the horizontal bars represent mean values, and the error bars show the SD. The p values were determined using a Mann-Whitney U test. Data represent two independent experiments.