Skip to main page content
Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
, 16 (1), 130

Macrophage Galactose-Type Lectin (MGL) Is Induced on M2 Microglia and Participates in the Resolution Phase of Autoimmune Neuroinflammation

Affiliations

Macrophage Galactose-Type Lectin (MGL) Is Induced on M2 Microglia and Participates in the Resolution Phase of Autoimmune Neuroinflammation

Juan M Ilarregui et al. J Neuroinflammation.

Abstract

Background: Multiple sclerosis (MS) involves a misdirected immune attack against myelin in the brain and spinal cord, leading to profound neuroinflammation and neurodegeneration. While the mechanisms of disease pathogenesis have been widely studied, the suppression mechanisms that lead to the resolution of the autoimmune response are still poorly understood. Here, we investigated the role of the C-type lectin receptor macrophage galactose-type lectin (MGL), usually expressed on tolerogenic antigen-presenting cells (APCs), as a negative regulator of autoimmune-driven neuroinflammation.

Methods: We used in silico, immunohistochemical, immunofluorescence, quantitative real-time polymerase chain reaction (qRT-PCR) and flow cytometry analysis to explore the expression and functionality of MGL in human macrophages and microglia, as well as in MS post-mortem tissue. In vitro, we studied the capacity of MGL to mediate apoptosis of experimental autoimmune encephalomyelitis (EAE)-derived T cells and mouse CD4+ T cells. Finally, we evaluated in vivo and ex vivo the immunomodulatory potential of MGL in EAE.

Results: MGL plays a critical role in the resolution phase of EAE as MGL1-deficient (Clec10a-/-) mice showed a similar day of onset but experienced a higher clinical score to that of WT littermates. We demonstrate that the mouse ortholog MGL1 induces apoptosis of autoreactive T cells and diminishes the expression of pro-inflammatory cytokines and inflammatory autoantibodies. Moreover, we show that MGL1 but not MGL2 induces apoptosis of activated mouse CD4+ T cells in vitro. In human settings, we show that MGL expression is increased in active MS lesions and on alternatively activated microglia and macrophages which, in turn, induces the secretion of the immunoregulatory cytokine IL-10, underscoring the clinical relevance of this lectin.

Conclusions: Our results show a new role of MGL-expressing APCs as an anti-inflammatory mechanism in autoimmune neuroinflammation by dampening pathogenic T and B cell responses, uncovering a novel clue for neuroprotective therapeutic strategies with relevance for in MS clinical applications.

Keywords: C-type lectin receptors; Experimental autoimmune encephalomyelitis; Inflammation; MGL; Microglia; Multiple sclerosis; Tolerance.

Conflict of interest statement

The authors declare that they have no competing interests.

Figures

Fig. 1
Fig. 1
MGL expression is upregulated in MS lesions. a Box plot showing the z-transformed expression values of MGL in NAWM and rim from chronic inactive (n = 7) and active (n = 7) MS lesions. *P < 0.05 (Kruskal-Wallis test followed by Dunn’s multiple comparisons test). b Expression of MGL in normal-appearing white matter (NAWM) and MS chronic inactive, chronic active, and active lesions. Consecutive brain sections from MS patients were stained for MGL and imaged by standard microscopy (representative of 3 donors). Scale bars, 50 μm. Insets, double staining showing the expression of MGL (red) and MHC II (green). Blue is a nuclear staining with DAPI. Images were collected from the same section with the same exposure time between different areas to allow comparison. Scale bars, 15 μm
Fig. 2
Fig. 2
MGL is expressed on alternatively activated microglia. a, b Monocyte-derived macrophages were polarized to the M0, M1, and M2 phenotypes. a Flow cytometry histograms. Representative plots from three independent experiments. b M0, M1, and M2 macrophages were stimulated with 10 ng/ml LPS in the presence or absence of GalNAc dendrimer (MGL agonist) or Gal-dendrimer (control dendrimer). After ON incubation, IL-10 was measured in the supernatants by ELISA. Values (fold change) are relative to that of control medium (medium without dendrimers). Mean + SEM, Mann Whitney U test, *P < 0.05 vs control treatment. Data are from two independent experiments with cells from four different donors. c Primary human microglia was isolated from freshly obtained post-mortem brain tissue and further polarized to the M0, M1, and M2 phenotypes. Cells were then lysed for mRNA isolation and the expression levels of MGL assayed by real-time quantitative RT-PCR. Each color identifies the cells from one donor. Values (relative expression, RE) are relative to that of GAPDH mRNA. *P < 0.05; ***P < 0.001 (Friedman test followed by Dunn’s multiple comparisons test). d Confocal microscopy in MS active lesion showing the merged expression of MGL (red) and P2Y12R (green). Nuclear staining (blue). Images were collected from the same section as in Fig. 1b. Scale bar, 15 μm
Fig. 3
Fig. 3
MGL1 but not MGL2 induces apoptosis of activated mouse T helper cells. Naïve CD4+ T cells from wild-type C57Bl/6 mice were stimulated for 3 days with CD3 and CD28 mAbs. Activated cells were washed and left untreated or incubated ON with 10 μg/ml of the indicated Fc-chimeras in the presence (white bars) or absence (black bars) of the Ca2+-chelator EDTA (5 mM). Then, cells were washed and stained with 7-AAD and AV and analyzed by flow cytometry. Cell death was determined as the difference in the percentage of AV+ cells between polarized cells treated with or without MGL-Fcs. Data are from three independent experiments with cells from three mice. One-way ANOVA followed by Tukey’s multiple comparisons test; *P < 0.05; **P < 0.01; ***P < 0.001
Fig. 4
Fig. 4
MGL1 deficiency results in enhanced susceptibility to EAE. Wild-type (WT) and Clec10a−/− mice were immunized with MOG35–55 and examined for disease progression (a) and body weight changes (b). Two independent experiments with 6–8 mice per group are shown. Values represent the mean + SEM; *P < 0.05; **P < 0.01; ***P < 0.001 (one-tailed unpaired Student’s t test of the AUC); #P < 0.05; ##P < 0.01 (one-tailed Mann-Whitney of the mean maximum scores)
Fig. 5
Fig. 5
Increased pro-inflammatory immune response in Clec10a−/− mice. MOG35–55-specific proliferation (a, b) and IL-17 production (c, d) by DLN cells (a, c) and splenocytes (b, d) from WT and Clec10−/− mice 27 days post-immunization (dpi; endpoint of experiments in Fig. 2). Proliferation was assessed by [3H]-thymidine incorporation, and cytokine secretion was analyzed by ELISA after 72 h of MOG35–55 restimulation. Values represent the mean ± SEM; *P < 0.05; **P < 0.01; ***P < 0.001 vs. cells from WT mice stimulated with MOG35–55 (Student t test). e Serum anti-MOG35–55 IgG1 and IgG2a antibody titers from WT and Clec10−/− mice obtained at 27 dpi. Mean ± SEM. Mann Whitney U test, *P < 0.05. f IgG1/IgG2c ratio per mouse from the titers shown in e. Mean ± SEM. Mann Whitney U test, **P < 0.01. af One representative experiment out of two is shown
Fig. 6
Fig. 6
Cells from Clec10a−/− mice are more susceptible to MGL1-induced cell death. a, b Wild-type (WT) and MGL1-deficient (Clec10a−/−) mice were immunized with MOG35–55 for EAE induction. At 27 dpi, DLN cells were isolated and incubated in the presence or absence of MGL1-Fc or MGL2-Fc. a After 30 min of incubation, cells were stained with anti-human IgG Fc to determine MGL1/2 binding and gated on MHC II cells. b Cell death was evaluated by AV binding after ON incubation. Mean ± SEM. Student t test, **P < 0.01

Similar articles

See all similar articles

References

    1. Reich DS, Lucchinetti CF, Calabresi PA. Multiple sclerosis. N Engl J Med. 2018;378:169–180. doi: 10.1056/NEJMra1401483. - DOI - PubMed
    1. Dendrou CA, Fugger L, Friese MA. Immunopathology of multiple sclerosis. Nat Rev Immunol. 2015;15:545–558. doi: 10.1038/nri3871. - DOI - PubMed
    1. Schwartz M, Baruch K. The resolution of neuroinflammation in neurodegeneration: leukocyte recruitment via the choroid plexus. EMBO J. 2014;33:7–22. doi: 10.1002/embj.201386609. - DOI - PMC - PubMed
    1. Joller N, Peters A, Anderson AC, Kuchroo VK. Immune checkpoints in central nervous system autoimmunity. Immunol Rev. 2012;248:122–139. doi: 10.1111/j.1600-065X.2012.01136.x. - DOI - PMC - PubMed
    1. van Kooyk Y, Ilarregui JM, van Vliet SJ. Novel insights into the immunomodulatory role of the dendritic cell and macrophage-expressed C-type lectin MGL. Immunobiology. 2015;220:185–192. doi: 10.1016/j.imbio.2014.10.002. - DOI - PubMed

LinkOut - more resources

Feedback