SRs mediate efferocytosis of PITs in vivo. (A–G and L) Mice were infected with 1:1 ratio of FliCind AmpR and WT KanR S. Typhimurium i.p. for 17 h, and treated with doxycycline. Spleens were harvested 7 h later. (C) Mice were injected with isotype or anti-Ly6G antibodies and infected 12 h after antibody treatment. (L) Mice were treated with Fucoidan at the same time as doxycycline. (H and I) WT BMMs (CellTracker Green) and Casp-1−/−Casp-11−/− BMMs (CellTracker Blue) were co-cultured, imaged and analyzed as in Fig. 4 C, and treated with serum (H), Annexin V (H), or fucoidan (I). (J) WT BMMs were infected with SPI1-induced S. Typhimurium as in Fig. 1 and treated with PBS or fucoidan for 2 h. Percentage dead cells was determined as in Fig. 1 B. (K and M) Spleens from mice (six animals per group) infected i.p with control or FliCind GFP–S. Typhimurium for 24 h and treated with doxycycline and fucoidan for 3.5 h, were harvested, prepared for flow cytometry and stained. Neutrophils with intracellular macrophage markers (CD45+ CD11b+ Ly6G+high CD68+ F4/80+) were identified by flow cytometry. All gating was performed using FlowJo. Data are representative of three individual experiments (A–J and L) or was pooled from two individual experiments (K and M). All error bars represent SE. P-values were determined by a two-tailed Student’s t test; *, P < 0.05.
