Results
H. polygyrus infection induces blood monocytosis and increases lung mononuclear phagocytes .
The early response to RSV infection in mice drives the recruitment of monocytes to the lung causing an accumulation of mononuclear phagocytes13. In the initial hours of infection these cells are predominantly Ly6Chi, later downregulating Ly6C and becoming predominantly CD11c+. Flow cytometric analysis was used to assess if these populations were also affected by H. polygyrus infection. Female Balb/c mice were infected with H. polygyrus and lungs assessed for innate immune cell composition through the later stages of infection. Cells of the monocyte-macrophage lineage were identified by their expression of the high affinity Fc receptor, CD64 (also known as FcγR1), and subdivided on the basis of SiglecF expression. SiglecF expressing CD64+ cells represent alveolar macrophages, whereas the SiglecF compartment will contain interstitial macrophages and some monocytes and their progeny. Compared with sham infection controls there was an increase in the numbers of mononuclear phagocytes (CD45+, Ly6G-, SiglecF-, CD11b+, CD64+, Fig. 1A) in the lungs of H. polygyrusinfected mice from ten days post infection, subsiding yet still elevated at 14 days. (Fig. 1B). In comparison, the resident alveolar macrophage compartment (CD45+, Ly6G-, SiglecF+, CD11b-, CD64+, Fig. 1A) remained unchanged (Fig. 1C). The expansion was predominantly due to Ly6C+ macrophages with a variable increase in CD11c+ only at day ten (Fig. 1D-F).
A large proportion of the recruited macrophages seen in response to lung inflammatory stimuli are derived from circulating monocytes23,24. It is therefore likely that the vascular monocyte reservoir would reflect the observed influx of mononuclear phagocytes to the lung. Peripheral blood was sampled from mice throughout H. polygyrus infection to assess circulatory monocyte counts by flow cytometry (CD45+, Lin-, CD11b+, Ly6G-, CD115+, Fig. 2A). Elevated total monocyte numbers were observed from seven to ten days post infection (Fig. 2B) and their numbers returned to baseline by 14 days post infection. Circulatory monocytes in the mouse can be distinguished into three phenotypic subtypes; classical, intermediate and non-classical monocytes25,26. Using the markers Ly6C and Treml4 to separate these three populations (Fig. 2A), the H. polygyrus induced expansion of monocytes was found to be predominantly in Ly6C+Treml4- classical monocytes (Fig. 2C).