Dimethyl Fumarate and Monomethyl Fumarate Promote Post-Ischemic Recovery in Mice.

Yao Y ，Miao W ，Liu Z ，Han W ，Shi K ，Shen Y ，Li H ，Liu Q ，Fu Y ，Huang D ，Shi FD ... -  《-》

Neuro-protective effect of monomethyl fumarate on ischemia reperfusion injury in rats: Role of Nrf2/HO1 pathway in peri-infarct region.

Post stroke recanalization has been associated with increased risk of oxidative stress. Stimulating endogenous antioxidant pathway by activation of nuclear factor erythroid-2-related factor-2 (Nrf2) plays a key role in neuronal defense against inflammation and oxidative stress in penumbra. Here, we explored whether monomethyl fumarate (MMF) could produce neuro-protection after ischemia/reperfusion (I/R) injury via Nrf2/HO1 activation. In male SD rats, middle cerebral artery was occluded for 90 min and confirmed using Laser Doppler flowmeter. MMF (10, 20 and 40 mg/kg) was administered in two divided doses at 30 min post ischemia and 5-10 min after reperfusion. After 24 h, effect on neurobehavioral parameters, infarct damage by TTC staining and MRI, oxidative stress and inflammatory cytokines were assessed. Expression studies of nuclear Nrf2 and cytoplasmic HO1 were performed in peri-infarct cortex and striatum; followed by dual immunofluorescence study to check the specific cell type. I/R induced neurobehavioral deficits and infarct damage were significantly (p < 0.05) attenuated by MMF (20 and 40 mg/kg). MMF, 20 mg/kg, significantly normalized I/R induced altered redox status and increased levels of TNF-α, IL-1β in the ipsilateral cortex. MRI data showed significantly reduced infarct in cortex but not in striatum after MMF treatment. Expression of nuclear Nrf2 and cytoplasmic HO1 were significantly (p < 0.05) increased in peri-infarct cortex after treatment with MMF. Additionally, dual immunofluorescence showed increased Nrf2 expression in neurons and HO1 expression in neurons as well as astrocytes in peri-infarct cortex after MMF treatment. Our results show the neuro-protective potential of MMF probably by restricting the progression of damage from striatum to cortex through activation of Nrf2/HO1 pathway in peri-infarct cortex.

Singh D ，Reeta KH ，Sharma U ，Jagannathan NR ，Dinda AK ，Gupta YK ... -  《-》

Fumarate decreases edema volume and improves functional outcome after experimental stroke.

Oxidative stress and inflammation exacerbate tissue damage in the brain after ischemic stroke. Dimethyl-fumarate (DMF) and its metabolite monomethyl-fumarate (MMF) are known to stimulate anti-oxidant pathways and modulate inflammatory responses. Considering these dual effects of fumarates, we examined the effect of MMF treatment after ischemic stroke in mice. Permanent middle cerebral artery occlusion (pMCAO) was performed using adult, male C57BL/6 mice. Thirty minutes after pMCAO, 20mg/kg MMF was administered intravenously. Outcomes were evaluated 6, 24 and 48h after pMCAO. First, we examined whether a bolus of MMF was capable of changing expression of kelch-like erythroid cell-derived protein with CNC homology-associated protein 1 (Keap1) and nuclear factor erythroid 2-related factor (Nrf)2 in the infarcted brain. Next, we studied the effect of MMF on functional recovery. To explore mechanisms potentially influencing functional changes, we examined infarct volumes, edema formation, the expression of heat shock protein (Hsp)72, hydroxycarboxylic acid receptor 2 (Hcar2), and inducible nitric oxide synthase (iNOS) in the infarcted brain using real-time PCR and Western blotting. Concentrations of a panel of pro- and anti-inflammatory cytokines (IFNγ, IL-1β, IL-2, IL-4, IL-5, IL-6, IL-10, IL-12p70, TNF) were examined in both the infarcted brain tissue and plasma samples 6, 24 and 48h after pMCAO using multiplex electrochemoluminiscence analysis. Administration of MMF increased the protein level of Nrf2 6h after pMCAO, and improved functional outcome at 24 and 48h after pMCAO. MMF treatment did not influence infarct size, however reduced edema volume at both 24 and 48h after pMCAO. MMF treatment resulted in increased Hsp72 expression in the brain 6h after pMCAO. Hcar2 mRNA levels increased significantly 24h after pMCAO, but were not different between saline- and MMF-treated mice. MMF treatment also increased the level of the anti-inflammatory cytokine IL-10 in the brain and plasma 6h after pMCAO, and additionally reduced the level of the pro-inflammatory cytokine IL-12p70 in the brain at 24 and 48h after pMCAO. A single intravenous bolus of MMF improved sensory-motor function after ischemic stroke, reduced edema formation, and increased the levels of the neuroprotective protein Hsp72 in the brain. The early increase in IL-10 and reduction in IL-12p70 in the brain combined with changes in systemic cytokine levels may also contribute to the functional recovery after pMCAO.

Clausen BH ，Lundberg L ，Yli-Karjanmaa M ，Martin NA ，Svensson M ，Alfsen MZ ，Flæng SB ，Lyngsø K ，Boza-Serrano A ，Nielsen HH ，Hansen PB ，Finsen B ，Deierborg T ，Illes Z ，Lambertsen KL ... -  《-》

Pretreatment with Korean red ginseng or dimethyl fumarate attenuates reactive gliosis and confers sustained neuroprotection against cerebral hypoxic-ischemic damage by an Nrf2-dependent mechanism.

The transcriptional factor Nrf2, a master regulator of oxidative stress and inflammation that are tightly linked to the development and progression of cerebral ischemia pathology, plays a vital role in inducing the endogenous neuroprotective process. Here, hypoxic-ischemia (HI) was performed in adult Nrf2 knockout and wildtype mice that were orally pretreated either with standardized Korean red ginseng extract (Ginseng) or dimethyl fumarate (DMF), two candidate Nrf2 inducers, to determine whether the putative protection was through an Nrf2-dependent mechanism involving the attenuation of reactive gliosis. Results show that Nrf2 target cytoprotective genes were distinctly elevated following HI. Pretreatment with Ginseng or DMF elicited robust neuroprotection against the deterioration of acute cerebral ischemia damage in an Nrf2-dependent manner as revealed by the reductions of neurological deficits score, infarct volume and brain edema, as well as enhanced expression levels of Nrf2 target antioxidant proteins and anti-inflammation mediators. In both ischemic striatum and cortex, the dynamic pattern of attenuated reactive gliosis in astrocytes and microglia, including affected astrocytic dysfunction in glutamate metabolism and water homeostasis, correlated well with the Nrf2-dependent neuroprotection by Ginseng or DMF. Furthermore, such neuroprotective benefits extended to the late phase of ischemic brain damage after HI, as evidenced by improvements in neurobehavioral outcomes, infarct volume and brain edema. Overall, pretreatment with Ginseng or DMF identically attenuates reactive gliosis and confers long-lasting neuroprotective efficacy against ischemic brain damage through an Nrf2-dependent mechanism. This study also provides new insight into the profitable contribution of reactive gliosis in the Nrf2-dependent neuroprotection in acute brain injury.

Liu L ，Vollmer MK ，Ahmad AS ，Fernandez VM ，Kim H ，Doré S ... -  《-》

Distinct Nrf2 Signaling Mechanisms of Fumaric Acid Esters and Their Role in Neuroprotection against 1-Methyl-4-Phenyl-1,2,3,6-Tetrahydropyridine-Induced Experimental Parkinson's-Like Disease.

A promising approach to neurotherapeutics involves activating the nuclear-factor-E2-related factor 2 (Nrf2)/antioxidant response element signaling, which regulates expression of antioxidant, anti-inflammatory, and cytoprotective genes. Tecfidera, a putative Nrf2 activator, is an oral formulation of dimethylfumarate (DMF) used to treat multiple sclerosis. We compared the effects of DMF and its bioactive metabolite monomethylfumarate (MMF) on Nrf2 signaling and their ability to block 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced experimental Parkinson's disease (PD). We show that in vitro DMF and MMF activate the Nrf2 pathway via S-alkylation of the Nrf2 inhibitor Keap1 and by causing nuclear exit of the Nrf2 repressor Bach1. Nrf2 activation by DMF but not MMF was associated with depletion of glutathione, decreased cell viability, and inhibition of mitochondrial oxygen consumption and glycolysis rates in a dose-dependent manner, whereas MMF increased these activities in vitro However, both DMF and MMF upregulated mitochondrial biogenesis in vitro in an Nrf2-dependent manner. Despite the in vitro differences, both DMF and MMF exerted similar neuroprotective effects and blocked MPTP neurotoxicity in wild-type but not in Nrf2 null mice. Our data suggest that DMF and MMF exhibit neuroprotective effects against MPTP neurotoxicity because of their distinct Nrf2-mediated antioxidant, anti-inflammatory, and mitochondrial functional/biogenetic effects, but MMF does so without depleting glutathione and inhibiting mitochondrial and glycolytic functions. Given that oxidative damage, neuroinflammation, and mitochondrial dysfunction are all implicated in PD pathogenesis, our results provide preclinical evidence for the development of MMF rather than DMF as a novel PD therapeutic. Almost two centuries since its first description by James Parkinson, Parkinson's disease (PD) remains an incurable disease with limited symptomatic treatment. The current study provides preclinical evidence that a Food and Drug Administration-approved drug, dimethylfumarate (DMF), and its metabolite monomethylfumarate (MMF) can block nigrostriatal dopaminergic neurodegeneration in a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine mouse model of PD. We elucidated mechanisms by which DMF and its active metabolite MMF activates the redox-sensitive transcription factor nuclear-factor-E2-related factor 2 (Nrf2) to upregulate antioxidant, anti-inflammatory, mitochondrial biosynthetic and cytoprotective genes to render neuroprotection via distinct S-alkylating properties and depletion of glutathione. Our data suggest that targeting Nrf2-mediated gene transcription using MMF rather than DMF is a promising approach to block oxidative stress, neuroinflammation, and mitochondrial dysfunction for therapeutic intervention in PD while minimizing side effects.

Ahuja M ，Ammal Kaidery N ，Yang L ，Calingasan N ，Smirnova N ，Gaisin A ，Gaisina IN ，Gazaryan I ，Hushpulian DM ，Kaddour-Djebbar I ，Bollag WB ，Morgan JC ，Ratan RR ，Starkov AA ，Beal MF ，Thomas B ... -  《-》