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Investigation of Macrophage Polarization Using Bone Marrow Derived Macrophages
Published on: June 23, 2013
A transcriptomic analysis of the effects of macrophage polarization and endotoxin tolerance on the response to
Katharine Sedivy-Haley1, Travis Blimkie1, Reza Falsafi1
1Department of Microbiology and Immunology, University of British Columbia, Vancouver, British Columbia, Canada.
Abstract:
Salmonella is an intracellular pathogen causing significant morbidity and mortality. Its ability to grow inside macrophages is important to virulence, and is dependent on the activation state of the macrophages. Classically activated M1 macrophages are non-permissive for Salmonella growth, while alternatively activated M2 macrophages are permissive for Salmonella growth. Here we showed that endotoxin-primed macrophages (MEP), such as those associated with sepsis, showed similar levels of Salmonella resistance to M1 macrophages after 2 hr of intracellular infection, but at the 4 hr and 24 hr time points were susceptible like M2 macrophages. To understand this mechanistically, transcriptomic sequencing, RNA-Seq, was performed. This showed that M1 and MEP macrophages that had not been exposed to Salmonella, demonstrated a process termed here as primed activation, in expressing relatively higher levels of particular anti-infective genes and pathways, including the JAK-STAT (Janus kinase-signal transducer and activator of transcription) pathway. In contrast, in M2 macrophages these genes and pathways were largely expressed only in response to infection. Conversely, in response to infection, M1 macrophages, but not MEP macrophages, modulated additional genes known to be associated with susceptibility to Salmonella infection, possibly contributing to the differences in resistance at later time points. Application of the JAK inhibitor Ruxolitinib before infection reduced resistance in M1 macrophages, supporting the importance of early JAK-STAT signalling in M1 resistance to Salmonella.
Insights
Endotoxin-primed macrophages (MEP) initially resist Salmonella like M1 cells but become susceptible later. This occurs because M1 and MEP cells show early anti-infective gene expression, unlike M2 cells, with JAK-STAT signaling crucial for M1 resistance.
Area of Science:
- Immunology
- Microbiology
- Pathogen-Host Interactions
Background:
- Salmonella is an intracellular pathogen whose virulence depends on macrophage permissiveness.
- M1 macrophages resist Salmonella, while M2 macrophages are permissive.
- Endotoxin-primed macrophages (MEP), relevant to sepsis, exhibit dynamic Salmonella resistance.
Purpose of the Study:
- To investigate the differential permissiveness of M1, M2, and MEP macrophages to Salmonella intracellular growth.
- To elucidate the molecular mechanisms underlying Salmonella resistance and susceptibility in different macrophage subtypes.
Main Methods:
- Macrophage infection assays with Salmonella.
- Transcriptomic sequencing (RNA-Seq) to analyze gene expression profiles.
- Pharmacological inhibition of the JAK-STAT pathway using Ruxolitinib.
Main Results:
- MEP macrophages mimic M1 resistance early but show M2-like susceptibility later.
- M1 and MEP macrophages exhibit 'primed activation' with higher baseline anti-infective gene expression, including the JAK-STAT pathway.
- M1 macrophages modulate additional susceptibility genes upon infection, unlike MEP macrophages.
- JAK inhibition (Ruxolitinib) diminishes M1 macrophage resistance to Salmonella.
Conclusions:
- Macrophage activation state critically influences Salmonella intracellular growth.
- Early JAK-STAT pathway activation contributes to M1 macrophage resistance.
- MEP macrophages display a unique temporal response to Salmonella infection, distinct from both M1 and M2 phenotypes.
