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The many faces of macrophages : polarization, endotoxin priming, and Salmonella resistance Sedivy-Haley, Katharine Jana-Marie

Abstract

Different forms of macrophage activation or polarization are relevant in the pathogenesis of a variety of diseases from inflammatory conditions to infections. It has been previously established that classically activated or M1 macrophages such as those produced by IFNγ stimulation are non-permissive for intracellular Salmonella infection, while alternatively activated or M2 macrophages such as those produced by IL-4 stimulation are permissive for Salmonella growth. It is not known whether endotoxin tolerant macrophages (primed with endotoxin stimulation), such as those observed in sepsis, are permissive for Salmonella growth. A gentamicin protection assay was performed for these three types of differently polarized human monocyte-derived macrophages (MDM) in vitro, and bacterial load measured through colony counts and microscopy. Endotoxin primed MDM (MEP) had a similar bacterial load to M1 macrophages at the initial and 2-hour time-points, but became more susceptible to Salmonella by the 4- and 24-hour time-points. Transcriptomic comparisons using RNA-Seq were performed to generate hypotheses regarding mechanisms for the differences observed between these polarization types, based on differential gene expression. Key immune pathways including JAK-STAT were enriched in uninfected M1 and MEP compared to uninfected M2 macrophages, suggesting a priming effect on these pathways due to polarization. Meanwhile, Salmonella-infected M1 showed increased expression of key inflammasome genes and Salmonella resistance genes compared to M2 and MEP macrophages. These effects were also observed in similarly treated human induced-pluripotent stem cell derived macrophages (iPSDM), further validating the usefulness of iPSDM as a macrophage model in polarization and infection experiments. In order to investigate the mechanistic relevance of these observations, Ruxolitinib was applied to inhibit JAK1-2 during the polarization phase of the experiment. This increased Salmonella permissiveness at the 4-hour time point in resistant M1 macrophages, but not in M2 or MEP macrophages, which are susceptible at this time point. This is consistent with an important role for JAK-STAT priming and resistance to Salmonella infection. These observations provide insights into the effects of polarization on Salmonella resistance in macrophages, and the suitability of iPSDM for macrophage study.

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