In this study, we have shown that the presence of intact SPI-1 genes enables Salmonella polarization of PAMs towards the less bactericidal M2-related response. This conclusion is supported by the fact that the M2 signaling exhibited a dose-dependent response after infection with the wild-type Salmonella Typhimurium whilst M1 signaling was dose independent. In fact, the M1 signaling after infection with the wild-type Salmonella Typhimurium appeared to be a mere response to LPS since the LPS response, response to heat killed Salmonella Typhimurium and responses to the wild-type strain at different MOI were all similar (Figure ). On the other hand, when SPI-1, but not SPI-2, genes were removed from the Salmonella Typhimurium chromosome, such a mutant was unable to modify macrophage polarization towards the M2. Indeed, infection of PAMs with the ΔSPI-1 mutant resulted in high, MOI dependent, pro-inflammatory signaling. Simultaneously, the SPI-1 mutant was essentially unable to induce the expression of M2 related genes, regardless of the MOI used.
Except for MMP9, for which transcription did not react to any kind of stimulation, the transcriptional profiles of the M2 related genes are clustered into two groups. The first group included IL-10 and arginase-1. These two genes were induced by both strains in a dose-dependent manner regardless of the presence of SPI-1. This can be explained by the fact that, although these genes are related to M2 [7
], IL-10 can be activated by the NF-κB signaling pathway in the late phases of infection to quell the immune response [24
] and arginase-1 is induced following toll like receptor stimulation [25
]. These two genes therefore partially share signaling pathways with the pro-inflammatory genes. The remaining group of mannose receptor, CD163, TfR1, TIMP-1, IL-4R and MMP12 exhibited a highly similar transcriptional pattern. As this pattern was exactly the opposite of that seen for M1 related genes, we propose these genes belong among the M2 polarization markers in PAMs.
The suppression of pro-inflammatory signaling is macrophage specific. The pro-inflammatory signaling in other cells is dependent on Salmonella
invasion, i.e. it is dependent on intact SPI1 [21
]. Interestingly, the alternative function of SPI1 genes in relation to differential cytokine signaling may have a clear evolutionary advantage. It has been shown recently that the inflammation induced by Salmonella
invading tissue results in host responses which either limit the growth of bacteria different from Salmonella
] or result in oxidation of intestinal metabolites, which can be used by Salmonella
for efficient proliferation at the mucosal surfaces [27
]. The invading Salmonella
cells thus provide the noninvasive part of the Salmonella
population a growth advantage in the gut mucus over the rest of the mucosal microbiota. However, once Salmonella
crosses the epithelium and comes in contact with macrophages, bactericidal M1 polarization would result in Salmonella
eradication and a decrease of pro-inflammatory signaling by the epithelial cells. In addition, M2 polarized macrophages may act against the signaling of epithelial cells and keep an appropriate balance between the epithelial cells and macrophages, which may result in the optimal production of Salmonella
required metabolites and limited damage to the host. Interestingly, the ability of Salmonella
to polarize the macrophage response towards the M2 might be further potentiated in host adapted serovars. In such cases, this polarization may enable Salmonella
to escape macrophage killing and, instead, these serovars can be distributed inside the macrophage across the host’s body without being effectively recognized and inactivated.