DMF (dimethyl fumarate) exerts anti-inflammatory and pro-metabolic effects in a variety of cell types, and a formulation (BG-12) is being evaluated for monotherapy in multiple sclerosis patients. DMF modifies glutathione (GSH) levels that can induce expression of the anti-inflammatory protein HO-1 (haem oxygenase-1). In primary astrocytes and C6 glioma cells, BG-12 dose-dependently suppressed nitrite production induced by either LI [LPS (lipopolysaccharide) at 1 μg/ml plus IFNγ (interferon γ) at 20 units/ml] or a mixture of pro-inflammatory cytokines, with greater efficacy in C6 cells. BG-12 reduced NOS2 (nitric oxide synthase 2) mRNA levels and activation of a NOS2 promoter, reduced nuclear levels of NF-κB (nuclear factor κB) p65 subunit and attenuated loss of IκBα (inhibitory κBα) in both cell types, although with greater effects in astrocytes. In astrocytes, LI decreased mRNA levels for GSHr (GSH reductase) and GCL (c-glutamylcysteine synthetase), and slightly suppressed GSHs (GSH synthetase) mRNAs. Co-treatment with BG-12 prevented those decreased and increased levels above control values. In contrast, LI reduced GSHp (GSH peroxidase) and GCL in C6 cells, and BG-12 had no effect on those levels. BG-12 increased nuclear levels of Nrf2 (nuclear factor-erythroid 2 p45 subunit-related factor 2), an inducer of GSH-related enzymes, in astrocytes but not C6 cells. In astrocytes, GSH was decreased by BG-12 at 2 h and increased at 24 h. Prior depletion of GSH using buthionine-sulfoximine increased the ability of BG-12 to reduce nitrites. In astrocytes, BG-12 increased HO-1 mRNA levels and effects on nitrite levels were blocked by an HO-1 inhibitor. These results demonstrate that BG-12 suppresses inflammatory activation in astrocytes and C6 glioma cells, but with distinct mechanisms, different dependence on GSH and different effects on transcription factor activation.

Background: The purinergic receptor P2x7 is expressed on myeloid cells as well as on CNS glial cells, and P2x7 activation has been shown to increase both glial and T-cell activation. These properties suggest a role in the development of autoimmune disease including multiple sclerosis.

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The endogenous neurotransmitter noradrenaline (NA) is known to exert potent anti-inflammatory effects in glial cells, as well as provide neuroprotection against excitatory and inflammatory stimuli. These properties raise the possibility that increasing levels of NA in the central nervous system (CNS) could provide benefit in neurological diseases and conditions containing an inflammatory component. In the current study, we tested this possibility by examining the consequences of selectively modulating CNS NA levels on the development of clinical signs in experimental autoimmune encephalomyelitis (EAE). In mice immunized with myelin oligodendrocyte glycoprotein peptide to develop a chronic disease, pretreatment to selectively deplete CNS NA levels exacerbated clinical scores. Elevation of NA levels using the selective NA reuptake inhibitor atomoxetine did not affect clinical scores, while treatment of immunized mice with the synthetic NA precursor L-threo-3,4-dihydroxyphenylserine (L-DOPS) prevented further worsening. In contrast, treatment of mice with a combination of atomoxetine and L-DOPS led to significant improvement in clinical scores as compared to the control group. The combined treatment reduced astrocyte activation in the molecular layer of the cerebellum as assessed by staining for glial fibrillary protein but did not affect Th1 or Th17 type cytokine production from splenic T cells. These data suggest that selective elevation of CNS NA levels could provide benefit in EAE and multiple sclerosis without influencing peripheral immune responses.