Figure 4
From: Lung B cells promote early pathogen dissemination and hasten death from inhalation anthrax
Bacillus anthracis (BA) spores rapidly associate with B cells in the lungs of infected mice. (a) A/J mice were infected intratracheally (i.t.) with 1 × 108 DyLight 649-labeled BA spores. At 2 and 5 hours post injection (h.p.i.), bronchoalveolar lavage (BAL) was performed. Lavaged lungs were perfused, excised, and digested with collagenase to prepare single-cell suspensions. Lung digests were stained and analyzed by fluorescence-activated cell sorting (FACS). CD45+ cells were selected from the live gate and analyzed for cell populations as previously reported,30 including B cells (IgM+ CD19+), alveolar macrophages (highly autofluorescent, CD11c+, side scatterhi and MHCIIlo), lung dendritic cells (CD11c+, SSClo, MHCII+), and neutrophils (Ly6G+). The number of each gated population that was spore+ was calculated and graphed (mean±s.e.m.). Data are representative of three independent experiments with n=3 per group. (b, c) Cryosections of lower right lobes from lungs of naive (left panel) and infected (right panel) mice were analyzed for the distribution of BA spores (green), immunoglobulin M (IgM; blue), CD11c (red), and 4,6-diamidino-2-phenylindole (DAPI; gray). Airways (AW) are labeled and insets show spore association with either IgM+ cells or CD11c+ cells. Arrows in panel b highlight B-cell locations. All images were taken at original magnification × 25. Similar observations were made in two independent experiments with n=2–3 per group.
