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[This corrects the article DOI: 10.3389/fimmu.2018.02515.].

The process of macrophage polarization is involved in many pathologies such as anti-cancer immunity and autoimmune diseases. Polarized macrophages exhibit various levels of plasticity when M2/M(IL-4) macrophages are reprogrammed into an M1-like phenotype following treatment with IFNγ and/or LPS. At the same time, M1 macrophages are resistant to reprogramming in the presence of M2-like stimuli. The molecular mechanisms responsible for the macrophages polarization, plasticity of M2 macrophages, and lack of plasticity in M1 macrophages remain unknown. Here, we explored the role of Egr2 in the induction and maintenance of macrophage M1 and M2 polarization in the mouse in vitro and in vivo models of inflammation. Egr2 knockdown with siRNA treatment fail to upregulate either M1 or M2 markers upon stimulation, and the overexpression of Egr2 potentiated M1 or M2 marker expression following polarization. Polarisation with M2-like stimuli (IL-4 or IL-13) results in increased Egr2 expression, but macrophages stimulated with M1-like stimuli (IFNγ, LPS, IL-6, or TNF) exhibit a decrease in Egr2 expression. Egr2 was critical for the expression of transcription factors CEBPβ and PPARγ in M2 macrophages, and CEBPβ was highly expressed in M1-polarized macrophages. In siRNA knockdown studies the transcription factor CEBPβ was found to negatively regulate Egr2 expression and is likely to be responsible for the maintenance of the M1-like phenotype and lack plasticity. During thioglycolate-induced peritonitis, adoptively transferred macrophages with Egr2 knockdown failed to become activated as determined by upregulation of MHC class II and CD86. Thus, our study indicates that Egr2 expression is associated with the ability of unstimulated or M2 macrophages to respond to stimulation with inflammatory stimuli, while low levels of Egr2 expression is associated with non-responsiveness of macrophages to their activation.

Recent studies suggest that in addition to their common function in the regulation of thrombosis and hemostasis, platelets also contribute to tissue inflammation affecting adaptive immunity. Platelets have a number of pro-inflammatory and regulatory mediators stored in their α-granules and dense granules, which are promptly released at sites of inflammation or tissue injury. Platelet-derived mediators include cytokines (IL-1α, IL-1β, and TGFβ1), chemokines (CXCL4 and CCL3), immunomodulatory neurotransmitters (serotonin, dopamine, epinephrine, histamine, and GABA), and other low-molecular-weight mediators. In addition, activated platelets synthesize a number of lipid pro-inflammatory mediators such as platelet-activating factor and prostaglandins/thromboxanes. Notably, platelets express multiple toll-like receptors and MHC class I on their surface and store IgG in their α-granules. Platelet-derived factors are highly effective in directly or indirectly modulating the priming and effector function of various subsets of T cells. Besides secreting soluble factors, activated platelets upregulate a number of integrins, adhesion molecules, and lectins, leading to the formation of platelet-T cells aggregates. Activated platelets are able to instantly release neurotransmitters acting similar to neuronal presynaptic terminals, affecting CD4 T cells and other cells in close contact with them. The formation of platelet-T cell aggregates modulates the functions of T cells via direct cell-cell contact interactions and the local release of soluble factors including neurotransmitters. New data suggest an important role for platelets as neuronal and innate-like cells that directly recognize damage- or pathogen- associated molecular patterns and instantly communicate with T cells.

Although it has been demonstrated that cAMP pathway affect both adaptive and innate cell functions, the role of this pathway in the regulation of T-cell-mediated central nervous system (CNS) autoimmune inflammation, such as in experimental autoimmune encephalomyelitis (EAE), remains unclear. It is also unclear how cAMP pathway affects the function of CD4 T cells in vivo at the site of inflammation. We found that adenylyl cyclase activator Forskolin besides inhibition of functions autoimmune CD4 T cells also upregulated microRNA (miR)-124 in the CNS during EAE, which is associated with M2 phenotype of microglia/macrophages. Our study further established that in addition to direct influence of cAMP pathway on CD4 T cells, stimulation of this pathway promoted macrophage polarization toward M2 leading to indirect inhibition of function of T cells in the CNS. We demonstrated that Forskolin together with IL-4 or with Forskolin together with IL-4 and IFNγ effectively stimulated M2 phenotype of macrophages indicating high potency of this pathway in reprogramming of macrophage polarization in Th2- and even in Th1/Th2-mixed inflammatory conditions such as EAE. Mechanistically, Forskolin and/or IL-4 activated ERK pathway in macrophages resulting in the upregulation of M2-associated molecules miR-124, arginase (Arg)1, and Mannose receptor C-type 1 (Mrc1), which was reversed by ERK inhibitors. Administration of Forskolin after the onset of EAE substantially upregulated M2 markers Arg1, Mrc1, Fizz1, and Ym1 and inhibited M1 markers nitric oxide synthetase 2 and CD86 in the CNS during EAE resulting in decrease in macrophage/microglia activation, lymphocyte and CD4 T cell infiltration, and the recovery from the disease. Forskolin inhibited proliferation and IFNγ production by CD4 T cells in the CNS but had rather weak direct effect on proliferation of autoimmune T cells in the periphery and in vitro , suggesting prevalence of indirect effect of Forskolin on differentiation and functions of autoimmune CD4 T cells in vivo . Thus, our data indicate that Forskolin has potency to skew balance toward M2 affecting ERK pathway in macrophages and indirectly inhibit pathogenic CD4 T cells in the CNS leading to the suppression of autoimmune inflammation. These data may have also implications for future therapeutic approaches to inhibit autoimmune Th1 cells at the site of tissue inflammation.