Neuroprotective Effects Of Dimethyl Fumarate Research Articles

Dimethyl Fumarate Exerts a Neuroprotective Effect by Enhancing Mitophagy via the NRF2/BNIP3/PINK1 Axis in the MPP+ Iodide-Induced Parkinson's Disease Mice Model.

Parkinson's disease (PD) is a progressive neurodegenerative disorder linked to the loss of dopaminergic neurons in the substantia nigra. Mitophagy, mitochondrial selective autophagy, is critical in maintaining mitochondrial and subsequently neuronal homeostasis. Its impairment is strongly implicated in PD and is associated with accelerated neurodegeneration. To study the positive effect of dimethyl fumarate (DMF) on mitophagy via the NRF2/BNIP3/PINK1 axis activation in PD disease models. The neuroprotective effect of DMF was explored in in vitro and in vivo PD models. MTT assay was performed to determine the DMF dose followed by JC-1 assay to study its mitoprotective effect in MPP+ exposed SHSY5Y cells. For the in vivo study, C57BL/6 mice were divided into six groups: Normal Control (NC), Disease Control (DC), Sham (Saline i.c.v.), Low Dose (MPP+ iodide+DMF 15 mg/kg), Mid Dose (MPP+ iodide+DMF 30 mg/kg), and High Dose (MPP+ iodide+DMF 60 mg/kg). The neuroprotective effect of DMF was assessed by performing rotarod, open field test, and pole test, and biochemical parameter analysis using immunofluorescence, western blot, and RT-PCR. DMF treatment significantly alleviated the loss of TH positive dopaminergic neurons and enhanced mitophagy by increasing PINK1, Parkin, BNIP3, and LC3 levels in the MPP+ iodide-induced PD mice model. DMF treatment groups showed good locomotor activity and rearing time when compared to the DC group. DMF confers neuroprotection by activating the BNIP3/PINK1/Parkin pathway, enhancing the autophagosome formation via LC3, and improving mitophagy in PD models, and could be a potential therapeutic option in PD.

Neuroprotective effects of dimethyl fumarate against manic-like behavior induced by ketamine in rats.

Medications for treating bipolar disorder (BD) are limited and can cause side effects if used chronically. Therefore, efforts are being made to use new agents in the control and treatment of BD. Considering the antioxidant and anti-inflammatory effects of dimethyl fumarate (DMF), this study was performed to examine the role of DMF on ketamine (KET)-induced manic-like behavior (MLB) in rats. Forty-eight rats were randomly divided into eight groups, including three groups of healthy rats: normal, lithium chloride (LiCl) (45mg/kg, p.o.), and DMF (60mg/kg, p.o.), and five groups of MLB rats: control, LiCl, and DMF (15, 30, and 60mg/kg, p.o.), which received KET at a dose of 25mg/kg, i.p. The levels of total sulfhydryl groups (total SH), thiobarbituric acid reactive substances (TBARS), nitric oxide (NO), and tumor necrosis factor-alpha (TNF-α), as well as the activity of antioxidant enzymes including catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase (GPx) in the prefrontal cortex (PFC) and hippocampus (HPC), were measured. DMF prevented hyperlocomotion (HLM) induced by KET. It was found that DMF could inhibit the increase in the levels of TBARS, NO, and TNF-α in the HPC and PFC of the brain. Furthermore, by examining the amount of total SH and the activity of SOD, GPx, and CAT, it was found that DMF could prevent the reduction of the level of each of them in the brain HPC and PFC. DMF pretreatment improved the symptoms of the KET model of mania by reducing HLM, oxidative stress, and modulating inflammation.

Neuroprotective effects of dimethyl fumarate against depression-like behaviors via astrocytes and microglia modulation in mice: possible involvement of the HCAR2/Nrf2 signaling pathway.

We postulated that dimethyl fumarate (DMF) exerts neuroprotective effects against depression-like behaviors through astrocytes and microglia modulation. To ascertain our hypothesis and define the mechanistic pathways involved in effect of DMF on neuroinflammation, we used the depression model induced by chronic unpredictable mild stress (CUMS), in which, the mice were exposed to stressful events for 28 days and from the 14th day they received DMF in the doses of 50 and 100 mg/kg or fluoxetine 10 mg/kg or saline. On the 29th day, the animals were subjected to behavioral tests. Microglia (Iba1) and astrocyte (GFAP) marker expressions were evaluated by immunofluorescence analyzes and the cytokines TNF-α and IL-Iβ by immunoenzymatic assay. In addition, computational target prediction, 3D protein structure prediction, and docking calculations were performed with monomethyl fumarate (DMF active metabolite) and the Keap1 and HCAR2 proteins, which suggested that these could be the probable targets related protective effects. CUMS induced anxiety- and depressive-like behaviors, cognitive deficit, decreased GFAP, and increased Iba1, TNF-α, and IL-Iβ expression in the hippocampus. These alterations were reversed by DMF. Thus, it is suggested that one of the mechanisms involved in the antidepressant effect of DMF is neuroinflammatory suppression, through the signaling pathway HCAR2/Nrf2. However, more studies must be performed to better understand the molecular mechanisms of this drug.

Anti-Inflammatory Effects of Dimethyl Fumarate in Microglia via an Autophagy Dependent Pathway

Dimethyl fumarate (DMF), which has been approved by the Food and Drug Administration for the treatment of relapsing-remitting multiple sclerosis, is considered to exert anti-inflammatory and antioxidant effects. Microglia maintain homeostasis in the central nervous system and play a key role in neuroinflammation, while autophagy controls numerous fundamental biological processes, including pathogen removal, cytokine production, and clearance of toxic aggregates. However, the role of DMF in autophagy induction and the relationship of this effect with its anti-inflammatory functions in microglia are not well known. In the present study, we investigated whether DMF inhibited neuroinflammation and induced autophagy in microglia. First, we confirmed the anti-neuroinflammatory effect of DMF in mice with streptozotocin-induced diabetic neuropathy. Next, we used in vitro models including microglial cell lines and primary microglial cells to examine the anti-inflammatory and neuroprotective effects of DMF. We found that DMF significantly inhibited nitric oxide and proinflammatory cytokine production in lipopolysaccharide-stimulated microglia and induced the switch of microglia to the M2 state. In addition, DMF treatment increased the expression levels of autophagy markers including microtubule-associated protein light chain 3 (LC3) and autophagy-related protein 7 (ATG7) and the formation of LC3 puncta in microglia. The anti-inflammatory effect of DMF in microglia was significantly reduced by pretreatment with autophagy inhibitors. These data suggest that DMF leads to the induction of autophagy in microglia and that its anti-inflammatory effects are partially mediated through an autophagy-dependent pathway.

The dual reno- and neuro-protective effects of dimethyl fumarate against uremic encephalopathy in a renal ischemia/reperfusion model.

Dimethyl fumarate (DMF), a Nrf2 activator approved for multiple sclerosis (MS) in 2013, showed promising antioxidant and anti-inflammatory effects against cerebral injury. However, its mechanistic maneuver in renal ischemia/reperfusion (I/R) injury and its associated uremic encephalopathy has not been previously highlighted. To fulfill this aim, rats were divided into 4 groups; sham-operated, renal I/R, and 14days pretreated DMF (15 and 25mg/kg/day, orally). The small molecule drug reduced renal I/R-induced elevation in serum creatinine and blood urea nitrogen, the renal content of interleukin (IL)-18 and its pro-activator caspase-1. The DMF antioxidant potential was confirmed by the increased renal Nrf2 mRNA expression/content associated wit an enhanced total antioxidant capacity and an inhibition of lipid peroxidation. This character entailed the suppression of the assessed inflammatory markers, such as nuclear factor (NF)-κB, p38 mitogen-activated protein kinase, and tumor necrosis factor-α. Remotely, DMF protected against uremic encephalopathy signified by the suppressed cortical/hippocampal contents of glial fibrillary acidic protein through suppressing 2 trajectories, the NF-κB/inducible nitric oxide synthase/nitric oxide/guanylyl cyclase/cyclic guanosine monophosphate and IL-6/signal transducer and activator of transcription 3. Moreover, the open field test revealed an enhanced locomotor activity in DMF pretreated rats, reflecting counter ability against functional and behavioral effects of acute uremic encephalopathy. The current study advocates the novel DMF dual protection potential against renal I/R insult and its remote brain injury to compensate uremic encephalopathy and acute kidney injury as well.

Neuroprotective effect of dimethyl fumarate on cognitive impairment induced by ischemic stroke.

BackgroundOxidative damage may contribute to post-stroke cognitive impairment (PSCI), but the underlying mechanisms are not fully elucidated. Dimethyl fumarate (DMF) has been used as an antioxidant in multiple sclerosis and psoriasis patients. We hypothesized that redox state was associated with PSCI, and DMF might exert neuroprotective effect against PSCI via anti-oxidative actions.MethodsTo confirm this hypothesis, we first conducted a clinical study (NCT03519828) that enrolled patients diagnosed with acute ischemic stroke within 48 hours. Data were analyzed based on demographic characteristics, disease history, clinical data and redox state. Logistic regression was used to identify the factors associated with PSCI. Next, a middle cerebral artery occlusion (MCAO) rat model was used to explore the antioxidant capacity and neuroprotective effect of DMF. Furthermore, behavioural experiments, histology and immunostaining, and transmission electron microscopy were also performed.ResultsHigher baseline NIHSS score, lower GSH/GSSG and T-AOC levels were found in the PSCI patients. Better performance in Morris water maze and shuttle box testing, more regular arranged neurons and Nissl bodies, less TUNEL-positive cells and autophagosomes, lower expression of 4-HNE, and higher expression of GCLM and NQO1 were found in the (DMF + MCAO) rats compared with the MCAO rats.ConclusionsThese findings suggest that DMF may alleviate PSCI via neuroprotective actions, providing a new therapeutic strategy for PSCI.

Neuroprotective effect of dimethyl fumarate in stroke: The role of nuclear factor erythroid 2-related factor 2

Background: There is evidence that supports the neuroprotective effects of dimethyl fumarate (DMF) in stroke. Nuclear factor erythroid 2-related factor 2 (Nrf2) has both anti-oxidant and anti-inflammatory mechanisms. We investigated the neuroprotective effects of DMF via Nrf2 activation in the cortex, striatum, and diencephalon in a middle cerebral artery occlusion (MCAO) model of stroke. Methods: 22 Sprague-Dawley male rats were randomized into 3 groups. In DMF-treated group (n = 8), rats received 15 mg/kg oral DMF twice daily by gavage from day 0 to 14 after a 60-minute MCAO. The vehicle group (n = 7) underwent MCAO and were given methocel/H2O, using the same method and schedule. In the sham group (n = 7), neck was opened, but neither middle cerebral artery (MCA) was occluded nor any drug was administered. After 14 days, the animals were sacrificed. The infarct volume were assessed by stereology method. Nrf2 expression was evaluated in the cortex, striatum, and diencephalon by immunohistochemistry method. Results: Ratio of infarct to total brain volume was significantly lower in the DMF-treated group (5.76%) in comparison with the vehicle group (22.39%) (P < 0.0001). Nrf2 expression was higher in DMF-treated group in comparison with both the vehicle and sham groups in cortex, striatum, diencephalon, and total brain (P < 0.0001). In the DMF-treated group, significant negative correlation between Nrf2 expression and infarct volume was observed in cortex, striatum, diencephalon, and total brain. Conclusion: DMF induces Nrf2 expression and its neuroprotective effects in different brain anatomical regions.

Fumaric Acids Do Not Directly Influence Gene Expression of Neuroprotective Factors in Highly Purified Rodent Astrocytes.

(1) Background: Dimethylfumarate (DMF) has been approved for the treatment of relapsing remitting multiple sclerosis. However, the mode of action of DMF and its assumed active primary metabolite monomethylfumarate (MMF) is still not fully understood. Former reports suggest a neuroprotective effect of DMF mediated via astrocytes by reducing pro-inflammatory activation of these glial cells. We investigated potential direct effects of DMF and MMF on neuroprotective factors like neurotrophic factors and growth factors in astrocytes to elucidate further possible mechanisms of the mode of action of fumaric acids; (2) Methods: highly purified cultures of primary rat astrocytes were pre-treated in vitro with DMF or MMF and incubated with lipopolysaccharides (LPS) or a mixture of interferon gamma (IFN-γ) plus interleukin 1 beta (IL-1β) in order to simulate an inflammatory environment. The gene expression of neuroprotective factors such as neurotrophic factors (nuclear factor E2-related factor 2 (NGF), brain-derived neurotrophic factor (BDNF), glial cell-derived neurotrophic factor (GDNF)) and growth factors (fibroblast growth factor 2 (FGF2), platelet-derived growth factor subunit A (PDGFa), ciliary neurotrophic factor (CNTF)) as well as cytokines (tumor necrosis factor alpha (TNFα), interleukin 6 (IL-6), IL-1β, inducible nitric oxide synthase (iNOS)) was examined by determining the transcription level with real-time quantitative polymerase chain reaction (qPCR); (3) Results: The stimulation of highly purified astrocytes with either LPS or cytokines changed the expression profile of growth factors and pro- inflammatory factors. However, the expression was not altered by either DMF nor MMF in unstimulated or stimulated astrocytes; (4) Conclusions: There was no direct influence of fumaric acids on neuroprotective factors in highly purified primary rat astrocytes. This suggests that the proposed potential neuroprotective effect of fumaric acid is not mediated by direct stimulation of neurotrophic factors in astrocytes but is rather mediated by other pathways or indirect mechanisms via other glial cells like microglia as previously demonstrated.

The Neuroprotective Effect of Dimethyl Fumarate in MPTP‐mouse Model of Parkinson's Disease via ROS/NF‐κB/Nrf‐2

Oxidative stress plays a key role in Parkinson's disease (PD) and nuclear transcription factor related to NF‐E2 (Nrf‐2) and nuclear factor kappa‐light‐chain‐enhancer of activated B cells (NF‐κB) are involved in neuroprotection against PD. The aim of the present study was to investigate the neurotherapeutic action, NF‐κB/Nrf‐2 dependent, of DMF in a mouse model of PD and in vitro SHSY‐5Y cells. Daily oral gavage of DMF (10, 30 and 100 mg/kg) significantly reduced neuronal degeneration of dopaminergic tract and behavioral impairments induced by four injections of the dopaminergic neurotoxin 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine (MPTP). Moreover, treatment with DMF prevented dopamine depletion increasing tyrosine hydroxylase (TH) and dopamine transporter (DAT) and also reduced α‐synuclein‐positive neurons. Furthermore, DMF treatment up‐regulated Nrf‐2 pathway, increasing NeuN+/Nrf‐2+ cells in the striatum and inducing activation of manganese superoxide dismutase (Mn‐SOD) and heme‐oxygenase‐1 (HO‐1), regulating glutathione levels too. Also, DMF reduced interleukin1 (Il‐1β), cyclooxygenase 2 (COX‐2), lowered nitrotyrosine (NT) and neuronal nitrite oxide synthase (nNOS) expression, modulated microglia activation, restored nerve growth factor (NGF) levels and preserved by microtubule‐associated protein 2 (MAP‐2) alterations. The protective effects of DMF treatment, via Nrf‐2, were confirmed in in vitro studies, trough the inhibition of Nrf‐2 by Trigonelline (TR). These findings underline that DMF, used in vivo at low dose (30mg/kg) and in vitro at the concentration of 30 μM, as NF‐κB/Nrf‐2 based therapy, offers novel cytoprotective modalities that further augment the natural antioxidant response in neurodegenerative and inflammatory disease models. These results support the thesis that DMF may constitute a promising therapeutic target for the treatment of PD.

The Neuroprotective Effect of Dimethyl Fumarate in an MPTP-Mouse Model of Parkinson's Disease: Involvement of Reactive Oxygen Species/Nuclear Factor-κB/Nuclear Transcription Factor Related to NF-E2.

Oxidative stress plays a key role in Parkinson disease (PD), and nuclear transcription factor related to NF-E2 (Nrf-2) is involved in neuroprotection against PD. The aim of the present study was to investigate a role for nuclear factor-κB (NF-κB)/Nrf-2 in the neurotherapeutic action of dimethyl fumarate (DMF) in a mouse model of PD and in vitro in SHSY-5Y cells. Daily oral gavage of DMF (10, 30, and 100 mg/kg) significantly reduced neuronal cell degeneration of the dopaminergic tract and behavioral impairments induced by four injections of the dopaminergic neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine. Moreover, treatment with DMF prevented dopamine depletion, increased tyrosine hydroxylase and dopamine transporter activities, and also reduced the number of α-synuclein-positive neurons. Furthermore, DMF treatment upregulated the Nrf-2 pathway, increased NeuN+/Nrf-2+ cell number in the striatum, induced activation of manganese superoxide dismutase and heme oxygenase-1, and regulated glutathione levels. Moreover, DMF reduced interleukin 1 levels, cyclooxygenase 2 activity, and nitrotyrosine neuronal nitrite oxide synthase expression. This treatment also modulated microglia activation, restored nerve growth factor levels, and preserved microtubule-associated protein 2 alterations. The protective effects of DMF treatment, via Nrf-2, were confirmed in in vitro studies, through inhibition of Nrf-2 by trigonelline. These findings demonstrate that DMF, both in a mouse model of PD and in vitro, provides, via regulation of the NF-κB/Nrf-2 pathway, novel cytoprotective modalities that further augment the natural antioxidant response in neurodegenerative and inflammatory disease models. These results support the thesis that DMF may constitute a promising therapeutic target for the treatment of PD. Antioxid. Redox Signal. 27, 453-471.

Dimethyl Fumarate Protects Neural Stem/Progenitor Cells and Neurons from Oxidative Damage through Nrf2-ERK1/2 MAPK Pathway.

Multiple sclerosis (MS) is the most common multifocal inflammatory demyelinating disease of the central nervous system (CNS). Due to the progressive neurodegenerative nature of MS, developing treatments that exhibit direct neuroprotective effects are needed. Tecfidera™ (BG-12) is an oral formulation of the fumaric acid esters (FAE), containing the active metabolite dimethyl fumarate (DMF). Although BG-12 showed remarkable efficacy in lowering relapse rates in clinical trials, its mechanism of action in MS is not yet well understood. In this study, we reported the potential neuroprotective effects of dimethyl fumarate (DMF) on mouse and rat neural stem/progenitor cells (NPCs) and neurons. We found that DMF increased the frequency of the multipotent neurospheres and the survival of NPCs following oxidative stress with hydrogen peroxide (H2O2) treatment. In addition, utilizing the reactive oxygen species (ROS) assay, we showed that DMF reduced ROS production induced by H2O2. DMF also decreased oxidative stress-induced apoptosis. Using motor neuron survival assay, DMF significantly promoted survival of motor neurons under oxidative stress. We further analyzed the expression of oxidative stress-induced genes in the NPC cultures and showed that DMF increased the expression of transcription factor nuclear factor-erythroid 2-related factor 2 (Nrf2) at both levels of RNA and protein. Furthermore, we demonstrated the involvement of Nrf2-ERK1/2 MAPK pathway in DMF-mediated neuroprotection. Finally, we utilized SuperArray gene screen technology to identify additional anti-oxidative stress genes (Gstp1, Sod2, Nqo1, Srxn1, Fth1). Our data suggests that analysis of anti-oxidative stress mechanisms may yield further insights into new targets for treatment of multiple sclerosis (MS).

Fumarates modulate microglia activation through a novel HCAR2 signaling pathway and rescue synaptic dysregulation in inflamed CNS.

Dimethyl fumarate (DMF), recently approved as an oral immunomodulatory treatment for relapsing-remitting multiple sclerosis (MS), metabolizes to monomethyl fumarate (MMF) which crosses the blood–brain barrier and has demonstrated neuroprotective effects in experimental studies. We postulated that MMF exerts neuroprotective effects through modulation of microglia activation, a critical component of the neuroinflammatory cascade that occurs in neurodegenerative diseases such as MS. To ascertain our hypothesis and define the mechanistic pathways involved in the modulating effect of fumarates, we used real-time PCR and biochemical assays to assess changes in the molecular and functional phenotype of microglia, quantitative Western blotting to monitor activation of postulated pathway components, and ex vivo whole-cell patch clamp recording of excitatory post-synaptic currents in corticostriatal slices from mice with experimental autoimmune encephalomyelitis (EAE), a model for MS, to study synaptic transmission. We show that exposure to MMF switches the molecular and functional phenotype of activated microglia from classically activated, pro-inflammatory type to alternatively activated, neuroprotective one, through activation of the hydroxycarboxylic acid receptor 2 (HCAR2). We validate a downstream pathway mediated through the AMPK–Sirt1 axis resulting in deacetylation, and thereby inhibition, of NF-κB and, consequently, of secretion of pro-inflammatory molecules. We demonstrate through ex vivo monitoring of spontaneous glutamate-mediated excitatory post-synaptic currents of single neurons in corticostriatal slices from EAE mice that the neuroprotective effect of DMF was exerted on neurons at pre-synaptic terminals by modulating glutamate release. By exposing control slices to untreated and MMF-treated activated microglia, we confirm the modulating effect of MMF on microglia function and, thereby, its indirect neuroprotective effect at post-synaptic level. These findings, whereby DMF-induced activation of a new HCAR2-dependent pathway on microglia leads to the modulation of neuroinflammation and restores synaptic alterations occurring in EAE, represent a possible novel mechanism of action for DMF in MS.Electronic supplementary materialThe online version of this article (doi:10.1007/s00401-015-1422-3) contains supplementary material, which is available to authorized users.

Dimethyl fumarate attenuates 6-OHDA-induced neurotoxicity in SH-SY5Y cells and in animal model of Parkinson’s disease by enhancing Nrf2 activity

Oxidative stress is central to the pathology of several neurodegenerative diseases, including Parkinson’s disease (PD), and therapeutics designed to enhance antioxidant potential could have clinical value. In this study, we investigated whether dimethyl fumarate (DMF) has therapeutic effects in cellular and animal model of PD, and explore the role of nuclear transcription factor related to NF-E2 (Nrf2) in this process. Treatment of animals and dopaminergic SH-SY5Y cells with DMF resulted in increased nuclear levels of active Nrf2, with subsequent upregulation of antioxidant target genes. The cytotoxicity of 6-hydroxydopamine (6-OHDA) was reduced by pre-treatment with DMF in SH-SY5Y cells. The increase in the reactive oxygen species caused by 6-OHDA treatment was also attenuated by DMF in SH-SY5Y cells. The neuroprotective effects of DMF against 6-OHDA neurotoxicity were dependent on Nrf2, since treatment with Nrf2 siRNA failed to block against 6-OHDA neurotoxicity and induce Nrf2-dependent cytoprotective genes in SH-SY5Y cells. In vivo, DMF oral administration was shown to upregulate mRNA and protein levels of Nrf2 and Nrf2-regulated cytoprotective genes, attenuate 6-OHDA induced striatal oxidative stress and inflammation in C57BL/6 mice. Moreover, DMF ameliorated dopaminergic neurotoxicity in 6-OHDA-induced PD animal models as evidenced by amelioration of locomotor dysfunction, loss in striatal dopamine, and reductions in dopaminergic neurons in the substantia nigra and striatum. Taken together, these data strongly suggest that DMF may be beneficial for the treatment of neurodegenerative diseases like PD.

Fumarates Promote Cytoprotection of Central Nervous System Cells against Oxidative Stress via the Nuclear Factor (Erythroid-Derived 2)-Like 2 Pathway

Oxidative stress is central to the pathology of several neurodegenerative diseases, including multiple sclerosis, and therapeutics designed to enhance antioxidant potential could have clinical value. The objective of this study was to characterize the potential direct neuroprotective effects of dimethyl fumarate (DMF) and its primary metabolite monomethyl fumarate (MMF) on cellular resistance to oxidative damage in primary cultures of central nervous system (CNS) cells and further explore the dependence and function of the nuclear factor (erythroid-derived 2)-like 2 (Nrf2) pathway in this process. Treatment of animals or primary cultures of CNS cells with DMF or MMF resulted in increased nuclear levels of active Nrf2, with subsequent up-regulation of canonical antioxidant target genes. DMF-dependent up-regulation of antioxidant genes in vivo was lost in mice lacking Nrf2 [Nrf2(-/-)]. DMF or MMF treatment increased cellular redox potential, glutathione, ATP levels, and mitochondrial membrane potential in a concentration-dependent manner. Treating astrocytes or neurons with DMF or MMF also significantly improved cell viability after toxic oxidative challenge in a concentration-dependent manner. This effect on viability was lost in cells that had eliminated or reduced Nrf2. These data suggest that DMF and MMF are cytoprotective for neurons and astrocytes against oxidative stress-induced cellular injury and loss, potentially via up-regulation of an Nrf2-dependent antioxidant response. These data also suggest DMF and MMF may function through improving mitochondrial function. The clinical utility of DMF in multiple sclerosis is being explored through phase III trials with BG-12, which is an oral therapeutic containing DMF as the active ingredient.

Dimethylfumarate inhibits microglial and astrocytic inflammation by suppressing the synthesis of nitric oxide, IL-1beta, TNF-alpha and IL-6 in an in-vitro model of brain inflammation.

BackgroundBrain inflammation plays a central role in multiple sclerosis (MS). Dimethylfumarate (DMF), the main ingredient of an oral formulation of fumaric acid esters with proven therapeutic efficacy in psoriasis, has recently been found to ameliorate the course of relapsing-remitting MS. Glial cells are the effector cells of neuroinflammation; however, little is known of the effect of DMF on microglia and astrocytes. The purpose of this study was to use an established in vitro model of brain inflammation to determine if DMF modulates the release of neurotoxic molecules from microglia and astrocytes, thus inhibiting glial inflammation.MethodsPrimary microglial and astrocytic cell cultures were prepared from cerebral cortices of neonatal rats. The control cells were treated with LPS, an accepted inducer of pro-inflammatory properties in glial cells, and the experimental groups with LPS and DMF in different concentrations. After stimulation/incubation, the generation of nitric oxide (NO) in the cell culture supernatants was determined by measuring nitrite accumulation in the medium using Griess reagent. After 6 hours of treatment RT-PCR was used to determine transcription levels of iNOS, IL-1β, IL-6 and TNF-α mRNA in microglial and astrocytic cell cultures initially treated with DMF, followed after 30 min by LPS treatment. Moreover, we investigated possible involvement of the ERK and Nrf-2 transduction pathway in microglia using western blot analysis.ResultsPretreatment with DMF decreased synthesis of the proinflammatory mediators iNOS, TNF-α, IL-1β and IL-6 at the RNA level in activated microglia and astrocytes in vitro, associated with a decrease in ERK phosphorylation in microglia.ConclusionsCollectively, these results suggest that the neuroprotective effects of DMF may be in part functionally attributable to the compound's ability to inhibit expression of multiple neuroinflammatory mediators in brain of MS patients.

Use of dichloro-difluoromethane aerosols in preparing frozen sections.

Oxidative stress is central to the pathology of several neurodegenerative diseases, including multiple sclerosis (MS), and therapeutics designed to enhance antioxidant potential could have clinical value. The objective of this study was to characterize the potential direct neuroprotective effects of dimethyl fumarate (DMF) and its primary metabolite monomethyl fumarate (MMF) on cellular resistance to oxidative damage in primary cultures of central nervous system (CNS) cells and further explore the dependence and function of the Nuclear factor (erythroid-derived 2)-like 2 (Nrf2) pathway in this process. Treatment of animals or primary cultures of CNS cells with DMF or MMF resulted in increased nuclear levels of active Nrf2, with subsequent upregulation of canonical antioxidant target genes. DMF-dependent upregulation of antioxidant genes in vivo was lost in mice lacking Nrf2 (Nrf2-/-). DMF or MMF treatment increased cellular redox potential, glutathione, ATP levels and mitochondrial membrane potential in a concentration-dependent manner. Treating astrocytes or neurons with DMF or MMF also significantly improved cell viability after toxic oxidative challenge in a concentration-dependent manner. This effect on viability was lost in cells that had eliminated or reduced Nrf2. These data suggest that DMF and MMF are cytoprotective for neurons and astrocytes against oxidative stress-induced cellular injury and loss, potentially via upregulation of an Nrf2-dependent antioxidant response. These data also suggest DMF and MMF may function through improving mitochondrial function. The clinical utility of DMF in multiple sclerosis is being explored through phase 3 trials with BG-12, which is an oral therapeutic containing DMF as the active ingredient.