Figure 1.

VHL deficiency results in defective Tfh cell development and function. (A) Representative flow-cytometric plots of activated CD44⁺CD4⁺ T cells from WT and CD4^CreVhl^fl/fl (VHL cKO) mice 8 d after infection with LCMV. Numbers adjacent to outlined areas indicate frequency of Tfh (CXCR5⁺ICOS⁺ or CXCR5⁺SLAM^lo) cells or GC-Tfh (CXCR5⁺PD-1^hi or CXCR5⁺Bcl-6^hi) cells. (B) Quantification of frequency (among CD44⁺CD4⁺ T cells) and number of Tfh cells and GC-Tfh cells of mice as in A (n = 6 per group). (C) RT-PCR analysis of mRNA of Tfh cell–related genes in CD44⁺CD4⁺ and CD44⁻CD4⁺ T cells from WT and VHL cKO mice 8 d after LCMV infection; results were normalized to those of Actb mRNA (encoding β-actin). (D) Representative flow-cytometric plots of total B220⁺ B cells from WT and VHL cKO mice 8 d after LCMV infection. Numbers adjacent to outlined areas indicate frequency of GC B (GL7⁺CD95⁺; top row) or plasma (CD138⁺IgD^lo; bottom row) cells in the spleen. (E) Quantification of frequency (among B220⁺ B cells) and number of GC B and plasma cells in the spleen of mice as in D (n = 6 per group). (F) ELISA of LCMV-specific IgG in the sera from infected mice as in D (n = 6 per group), presented as absorbance at 490 nm (A490). (G) Representative flow-cytometric plots of CD44⁺CD4⁺ T cells from WT and VHL cKO mice 7 d after immunization with SRBC. Numbers adjacent to outlined areas indicate frequency of GC-Tfh (CXCR5⁺PD-1^hi or CXCR5⁺Bcl-6^hi) cells. (H) Quantification of frequency (among CD44⁺CD4⁺ T cells) and number of GC-Tfh cells of mice as in G (n = 3–4 per group). Each symbol (B, E, and H) represents an individual mouse; small horizontal lines indicate the mean (± SD). *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001; ns, nonsignificant (Student’s t test). Data are representative of three independent experiments.