Metabotropic glutamate receptor 5 may be involved in macrophage plasticity

Abstract Background Macrophages are a functionally heterogeneous cell population and depending on microenvironments they polarize in two main groups: M1 and M2. Glutamic acid and glutamate receptors may participate in the regulation of macrophage plasticity. To investigate the role of glutamatergic systems in macrophages physiology, we performed the transfection of mGluR5 cDNAs into RAW-264.7 cells. Results Comparative analysis of modified (RAW-mGluR5 macrophages) and non-modified macrophages (RAW-macrophages) has shown that the RAW-mGluR5 macrophages absorbed more glutamate than control cells and the amount of intracellular glutamate correlated with the expression of excitatory amino acid transporters -2 (EAAT-2). Besides, our results have shown that RAW-mGluR5 macrophages expressed a higher level of peroxisome proliferator-activated receptor γ (PPAR-γ) and secreted more IL-10, high mobility group box 1 proteins (HMGB1) and Galectin-3 than control RAW-macrophages. Conclusions We propose that elevation of intracellular glutamate and expression of mGluR5 may initiate the metabolic rearrangement in macrophages that could contribute to the formation of an immunosuppressive phenotype

MOESM1 of Metabotropic glutamate receptor 5 may be involved in macrophage plasticity

Additional file 1: Figure S1. Expression of mGluR5, EAAT-2, PPAR-γ and HMGB1 proteins in control and mGluR5-transfected macrophages. RAW 264.7 cells (RAW-NT) and mGluR5-transfected macrophages (RAW-mGluR5) (5–105 cells per well) were incubated with LPS (100 ng ⁄ ml) or IL-10 (20 nM) for 24 h, followed by the determination of EAAT-2 (b), PPAR-γ (d) and HMGB1 (f) expression by western blot analysis, as described in the “Methods” section. (h) β-Actin was also visualized by Western blotting to confirm equal loading of the fractions. Data shown are representative of three independent experiments. Quantification of EAAT2 blots shown in c, *P < 0.05, vs corresponding RAW-NT cells. Quantification of PPAR-γ shown in e, *P < 0.05, vs corresponding RAW-NT cells, **P < 0.05, vs mGluR5 control. Quantification of HMGB1 blots shown in g, *P < 0.05, vs corresponding RAW-NT cells

MOESM4 of Metabotropic glutamate receptor 5 may be involved in macrophage plasticity

Additional file 4: Figure S4. Effect of mGluR5 transfection on the HMGB1 and Gal-3 secretion. RAW 264.7 cells (RAW-NT) and mGluR5-transfected macrophages (RAW-mGluR5) (5–105 cells per well) were incubated with LPS (100 ng⁄ml) or IL-10 (20 nM) for 24 h, followed by the determination of HMGB1 proteins (a) and Gal-3 (b), as described in the “Methods” section. Data represented are mean ± SEM of results from four separate experiments performed in duplicate. *P < 0.05, vs RAW-NT control cells. **P < 0.05, vs RAW-mGlur5 control cells

MOESM2 of Metabotropic glutamate receptor 5 may be involved in macrophage plasticity

Additional file 2: Figure S2. Effect of LPS and glutamate on NO and IL-10 secretion. RAW 264.7 cells (RAW-NT) and mGluR5-transfected macrophages (RAW-mGluR5) (5–105 cells per well) were incubated with LPS (100 ng ⁄ ml) or glutamate (40 μM) for 24 h, followed by determination of NO (a) and IL-10 (b) secretion, as described in the “Methods” section. Data represented are mean ± SEM of results from four. separate experiments performed in duplicate. *P < 0.05, vs corresponding RAW-NT cells. **P < 0.05, vs RAW-mGluR5 control cells