Effects of opsonization and gamma interferon on growth of Brucella melitensis 16M in mouse peritoneal macrophages in vitro
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Published on: November 16, 2016
M O Eze1, L Yuan, R M Crawford
1Department of Bacterial Diseases, Walter Reed Army Institute of Research, Washington, D.C. 20307-5100, USA.
Abstract:
Entry of opsonized pathogens into phagocytes may benefit or, paradoxically, harm the host. Opsonization may trigger antimicrobial mechanisms such as reactive oxygen or nitric oxide (NO) production but may also provide a safe haven for intracellular replication. Brucellae are natural intramacrophage pathogens of rodents, ruminants, dogs, marine mammals, and humans. We evaluated the role of opsonins in Brucella-macrophage interactions by challenging cultured murine peritoneal macrophages with Brucella melitensis 16M treated with complement- and/or antibody-rich serum. Mouse serum rich in antibody against Brucella lipopolysaccharide (LPS) (aLPS) and human complement-rich serum (HCS) each enhanced the macrophage uptake of brucellae. Combinations of suboptimal levels of aLPS (0. 01%) and HCS (2%) synergistically enhanced uptake. The intracellular fate of ingested bacteria was evaluated with an optimal concentration of gentamicin (2 microg/ml) to control extracellular growth but not kill intracellular bacteria. Bacteria opsonized with aLPS and/or HCS grew equally well inside macrophages in the absence of gamma interferon (IFN-gamma). Macrophage activation with IFN-gamma inhibited replication of both opsonized and nonopsonized brucellae but was less effective in inhibiting replication of nonopsonized bacteria. IFN-gamma treatment of macrophages with opsonized or nonopsonized bacteria enhanced NO production, which was blocked by N(G)-monomethyl L-arginine (MMLA), an NO synthesis inhibitor. MMLA also partially blocked IFN-gamma-mediated bacterial growth inhibition. These studies suggest that primary murine macrophages have limited ability to control infection with B. melitensis, even when activated by IFN-gamma in the presence of highly opsonic concentrations of antibody and complement. Additional cellular immune responses, e.g., those mediated by cytotoxic T cells, may play more important roles in the control of murine brucellosis.
Insights
Opsonization enhances Brucella uptake by macrophages but doesn't prevent intracellular growth. Macrophage activation with gamma interferon (IFN-γ) helps control Brucella, but other immune responses may be crucial for controlling brucellosis.
Area of Science:
- Immunology
- Microbiology
- Cell Biology
Background:
- Opsonization of pathogens can enhance phagocyte uptake but may also facilitate intracellular survival.
- Brucella species are intracellular pathogens that infect macrophages, posing a significant health risk.
- Understanding the role of opsonins in Brucella-macrophage interactions is crucial for developing effective treatments.
Purpose of the Study:
- To evaluate the role of opsonins (antibody and complement) in Brucella melitensis interactions with murine macrophages.
- To investigate the intracellular fate of opsonized and non-opsonized Brucella within macrophages.
- To assess the impact of gamma interferon (IFN-γ) activation on macrophage antimicrobial mechanisms against Brucella.
Main Methods:
- Murine peritoneal macrophages were challenged with Brucella melitensis treated with antibody against Brucella lipopolysaccharide (aLPS) and/or human complement-rich serum (HCS).
- Intracellular bacterial growth was assessed using gentamicin to control extracellular bacteria.
- Macrophage activation with IFN-γ and nitric oxide (NO) production were measured, with NO synthesis inhibition using N(G)-monomethyl L-arginine (MMLA).
Main Results:
- Both aLPS and HCS enhanced macrophage uptake of Brucella, with synergistic effects at suboptimal concentrations.
- Opsonized and non-opsonized Brucella replicated equally well within macrophages without IFN-γ.
- IFN-γ inhibited Brucella replication, enhanced NO production, and partially restored growth inhibition when NO synthesis was blocked.
Conclusions:
- Murine macrophages exhibit limited intrinsic ability to control Brucella melitensis infection, even when activated.
- Opsonization enhances bacterial uptake but does not confer intracellular resistance to macrophages.
- Gamma interferon-induced nitric oxide production plays a role in controlling Brucella replication, but additional cellular immune responses are likely critical for resolving brucellosis.
