MyD88-dependent influx of monocytes and neutrophils impairs lymph node B cell responses to chikungunya virus infection via Irf5, Nos2 and Nox2
C57BL/6 mice were treated with 300–500 μg IgG2b isotype control mAb or anti-Gr-1 mAb via intraperitoneal injection one day prior to inoculation with 103 PFU of CHIKV AF15561 in the left footpad. (A) At 14 dpi, dLN sections were stained for ERTR7+ stromal cells, B220+ B cells, and GL7+ GC B cells. (B) Area per GC, (C) Number of GCs per LN section, and (D) total GC area per LN were determined. (E) Representative flow cytometry plots of GL7+CD95+ GC B cells (gated on CD19+B220+ cells), (F) percentage and (G) total number of GC B cells in the dLN at 14 dpi. (H) Representative flow cytometry plots of CD138+IgD- plasma cells (gated on CD19+), (I) percentage and (J) total number of plasma cells in the dLN at 14 dpi. (K) Number of CHIKV-specific IgG+ antibody secreting cells (ASCs) in the dLN at 14 dpi. Serum collected at the indicated timepoints was assayed for (L) total CHIKV-specific IgG by ELISA (expressed as fold change in IgG endpoint over the IgG2b group) and (M) neutralizing activity of CHIKV by focus reduction neutralization test (FRNT, expressed as fold change in FRNT50 over the IgG2b group). Errors bars represent mean ± SEM. Data in (A-D) are derived from 5–6 LNs per group with 7–12 LN sections analyzed per group. Data in (E-M) are derived from 2–3 independent experiments with 7–10 mice per group. Statistical significance was determined by Student’s t-test in B-D (note that significance is only displayed for comparison of IgG2b and α-Gr-1 groups), F-G, I-J, and K; or by two-way ANOVA with Bonferroni’s post-test in L-M (*, P < 0.05; **, P < 0.01; ***, P < 0.001).