Supplementary Figure 3: Immune response to NP-OVA in wild-type and Pik3cdE1020K/+ mice. | Nature Immunology

Supplementary Figure 3: Immune response to NP-OVA in wild-type and Pik3cdE1020K/+ mice.

From: Hyperactivated PI3Kδ promotes self and commensal reactivity at the expense of optimal humoral immunity

Supplementary Figure 3

Wild-type and Pik3cdE1020K/+ mice were immunized subcutaneously, at different age (2-months, 4-months, 1-year-old), with NP-OVA in alum in the hock and sacrificed on day+10. Analyses performed on draining (dLN) and resting (rLN) popliteal lymph nodes. a, Analyses of 2-month-old mice: frequency of PD-1+CXCR5+Foxp3 TFH cells (percent of CD4+B220 T cells, left panel) and GL-7+FAS+ GC B cells (of B220+CD19+ B cells, right panel) in rLN and dLN (n=5 per group). (b-c) Analyses of 4-month-old mice. b, cellularity of rLN and dLN (n=6 per group). c, Numbers of NP and NP+ GC B cells in dLN (wild-type n=6, Pik3cdE1020K/+ n=5). d, Analyses of 1-year-old mice: percentage and numbers of NP+ GC B cells in dLN (wild-type n=7, Pik3cdE1020K/+ n=9). e, Analyses of 2-month-old mice: percentage of NP+ GC B cells and ratios between numbers of NP+ and NP GC B cells in dLN (n=5 per group). f, Analyses of 4-month-old mice: ELISA for serum IgG1 (wild-type n=8, Pik3cdE1020K/+ n=6). (g-m) Analyses based on Figure 3g. g, Proportion of FDC networks with a GC having greater than 10 BCL-6+ cells (wild-type rLN, n=3; wild-type dLN, n=2; Pik3cdE1020K/+ rLN and dLN n=3). Each symbol represents an entire LN section corresponding to 5-8 FDC networks. h, Area of BCL-6+ GCs per FDC region. Each symbol corresponds to a GC (wild-type rLN, n=3; wild-type dLN, n=5; Pik3cdE1020K/+ rLN, n=13; Pik3cdE1020K/+ dLN n=6). i, Histo-cytometric representation of GCs from wild-type dLN depicted in Figure 3g, II. The distribution of PD-1+CD4+ T cells within the LZ and DZ of the GC was determined using the density of CD35 and BCL-6 to mark the LZ and DZ, respectively. j, Percentage of TFH cells (PD-1+CD4+ surfaces) in the DZ as determined by histo-cytometry. k, Normalized numbers of TFH cells (identified as in j) per DZ GC area (BCL-6+CD35). Each symbol corresponds to a GC (j,k, wild-type dLN, n=4; Pik3cdE1020K/+ rLN, n=12; Pik3cdE1020K/+ dLN n=6). IF analyses are representative of 2-3 independent lymph nodes analyzed. l, Representative FACS plots and histograms of DZ (CXCR4hiCD86lo) and LZ (CXCR4loCD86hi) gated on NP+ GC B cells (wild-type n=8, Pik3cdE1020K/+ n=7). m, Percentage (of live cells) and numbers of CD4+B220CXCR5PD1Foxp3+ Treg cells, and CD4+B220PD-1+CXCR5+Foxp3+ TFR cells in rLN and dLN (wild-type n=8, Pik3cdE1020K/+ n=7). Data in (a-f, l, m) are representative of two independent experiments. Shown is the mean ± SEM. Significance analyzed by Mann-Whitney U test. *P < 0.05; **P < 0.01; ***P < 0.001