Flow cytometry.
Peritoneal lavage cells were surface stained with the antibodies listed
in Supplementary Table 1. Flow cytometry data were acquired in a FACS
Canto II Cytometer (BD), at the Instituto de Higiene, Universidad de la
República, and then analysed using the FlowJo software package. Single
cells were selected on the basis of FSC-H and FSC-A, and then viable
cells selected using the Live/Dead GreenTM viability
probe. Within live cell singlets, eosinophils and neutrophils were
defined as SiglecFhi and Ly6G+ cells
respectively. B cells (CD19+) were excluded from
SiglecF- Ly6G- cells.
SiglecF- Ly6G-CD19- cells were divided into monocytes
(Ly6Chi) and non-monocytes (Ly6C-).
The second category was further divided into small peritoneal
macrophages and dendritic cells (SPM/DC; MHCII+F4/80-), MHCII-F4/80med macrophages, and large peritoneal macrophages
(LPM; MHCII- F4/80high) (26, 27).
The gating strategy is summarised in supplementary Figure S1.
Statistical analysis .
Data were analysed by the non-parametric method of Kruskal-Wallis (28),
followed by the post-hoc multiple comparison test described by Conover
(29) and the Benjamini-Hochberg correction for controlling the false
discovery rate (30). The symbols *, ** and *** represent p-values less
than 0.05, 0.01 and 0.001 respectively.