Differential infection of mononuclear phagocytes by Francisella tularensis: role of the macrophage mannose receptor

Abstract

Francisella tularensis (Ft) is a Gram-negative bacterium and the causative agent of tularemia. It is well established that this organism replicates inside macrophages, but we are only beginning to understand this interface at the molecular level. Herein, we compared directly the ability of Ft subspecies holarctica live-vaccine strain to infect freshly isolated human peripheral blood monocytes, monocyte-derived macrophages (MDM), and cells of the murine macrophage cell line J774A.1 (J774). We now show that unopsonized bacteria infected human MDM fivefold more efficiently than monocytes or J774 cells in standard media. Moreover, enhanced infection of MDM was mediated, in part, by the macrophage mannose receptor (MR). Forming Ft phagosomes accumulated MR, and infection was inhibited by MR-blocking antibody or soluble mannan but not by the dectin-1 ligand laminarin. Up-regulation of MR in MDM (by exposure to interleukin-4) increased Ft phagocytosis, as did expression of MR in J774 cells. Conversely, opsonized Ft were ingested readily by monocytes and MDM. Medium supplementation with 2.5% fresh autologous serum was sufficient to confer opsonophagocytosis and CD11b accumulated in the membrane at sites of Ft engulfment. Infection of monocytes by opsonized Ft was nearly ablated by complement receptor 3 (CR3) blockade. Conversely, MDM used MR and CD11b/CD18 to ingest opsonized organisms. Altogether, our data demonstrate differential infection of mononuclear phagocytes by Ft and define distinct roles for MR and CR3 in phagocytosis.

Francisella Targets Cholesterol-Rich Host Cell Membrane Domains for Entry into Macrophages

“…than by colony count, may have contributed to these findings. Other studies, which have found less efficient entry into macrophages by the F. tularensis subspecies holarctica vaccine strain (8,9), also did not use the particular army lot 11 strain used herein, or the derivation of the strain was not detailed (39). Metabolic studies of various lots of the Francisella vaccine strain (Ref.…”

“…Multiplicities of infection (MOI) Ͼ10 -60 were required to achieve efficient infection. A role for the mannose receptor was further confirmed in an infection model using F. tularensis LVS and J774 macrophages, where induction of expression of the mannose receptor on human monocyte-derived macrophages by IL-4 led to a significant increase in the uptake of Francisella (9). The authors also acknowledged that because neither blockage of the mannose receptor nor of the complement receptor could ablate infection, there are probably more receptors involved.…”

“…Protease treatment of bacteria does not affect this entry pathway, whereas peroxidation and crosslinking of bacterial surface carbohydrates and/or lipopolysaccharides lead to conventional phagocytosis of Francisella. Using F. novicida and specifically looking at primary human monocytes, contributions from complement receptor 3, Fc␥ receptors, mannose receptors, and surfactant protein A have been described (7)(8)(9). Interestingly, these authors also described a 60% reduction of association of Francisella LVS with primary human monocytes in comparison to F. novicida.…”

A Novel Role for the Bactericidal/Permeability Increasing Protein in Interactions of Gram-Negative Bacterial Outer Membrane Blebs with Dendritic Cells

“…Thereafter, the cells were washed twice in PBS to remove any unbound material, and MDDC were allowed to adhere to an eight-chamber microscopy slide for 20 min. Adherent MDDCs were washed in PBS, fixed in 10% formalin for 15 min at 25°C, and then permeabilized in 50% methanol/50% acetone for 5 min at 4°C as previously described (31). Fixed and permeabilized cells were rinsed in PBS supplemented with 0.5 g/L sodium azide and 5 g/L BSA, and then blocked in PBS containing 10% heat-inactivated normal horse serum (blocking buffer) for 1 h at 25°C (31).…”

Resident microglia die and infiltrated neutrophils and monocytes become major inflammatory cells in lipopolysaccharide‐injected brain

“…The antibodies, OX-42, and Mac-1 that recognize CD11b generally used to detect microglia in rat and mouse, respectively, are unable to distinguish among microglia, neutrophils, and monocytes, since CD11b is expressed in all three cell types (Akiyama and McGeer, 1990;Jones et al, 1988;Schulert and Allen, 2006). In this study, we examined the behavior of these inflammatory cells in lipopolysaccharide (LPS)-injected brain using the additional markers, myeloperoxidase (MPO), and ionized calcium binding adaptor molecule-1 (Iba-1).…”