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Dimethyl fumarate (DMF) administration promotes retinal ganglion cell (RGC) survival after optic nerve crush (ONC). (A) Photomicrographs of tubulin β3 (TUBB3)-positive cells with or without DMF administration on day 7 after ONC. The concentration of DMF was 100 mg/kg. The photomicrographs show temporal retinal areas at a 1 mm distance from the optic disc. (B) Quantitative analysis of surviving TUBB3-positive cells. The y-axis represents the mean density of TUBB3-positive cells. Sham indicates a sham operation without ONC. The asterisks indicate p < 0.01 (n = 6), and each circle denotes the values of individual animals. Statistical analysis was performed using one-way analysis of variance (ANOVA) with Bonferroni's test for post-hoc analysis compared to ONC with the vehicle. Scale bar = 50 µm.
Source publication
This study aimed to verify whether dimethyl fumarate (DMF) promotes the survival of retinal ganglion cells (RGCs) after optic nerve crush (ONC) accompanied by activation of the NF-E2-related factor 2 (Nrf2)/heme oxygenase-1 (HO-1) pathway. We examined changes in the densities of tubulin β3 (TUBB3)-positive RGCs and the amplitudes of the positive sc...
Citations
... Two in vivo studies of optic nerve injury (Mori et al., 2020) and light-induced retinal degeneration (Dietrich et al., 2021) further highlight pro-survival effects of DMF on different cellular layers within the retina. In particular, the former study reports that DMF administration (100 mg/kg) protected retinal ganglion cells by degeneration, while the latter study shows a significant reduction in photoreceptors loss after DMF treatment (15 or 30 mg/kg). ...
Dimethyl fumarate (DMF) is an FDA-approved drug for the treatment of psoriasis and multiple sclerosis. DMF is known to stabilize the transcription factor Nrf2, which in turn induces the expression of antioxidant response element genes. It has also been shown that DMF influences autophagy and participates in the transcriptional control of inflammatory factors by inhibiting NF-κB and its downstream targets. DMF is receiving increasing attention for its potential to be repurposed for several diseases. This versatile molecule is indeed able to exert beneficial effects on different medical conditions through a pleiotropic mechanism, in virtue of its antioxidant, immunomodulatory, neuroprotective, anti-inflammatory, and anti-proliferative effects. A growing number of preclinical and clinical studies show that DMF may have important therapeutic implications for chronic diseases, such as cardiovascular and respiratory pathologies, cancer, eye disorders, neurodegenerative conditions, and systemic or organ specific inflammatory and immune-mediated diseases. This comprehensive review summarizes and highlights the plethora of DMF’s beneficial effects and underlines its repurposing opportunities in a variety of clinical conditions.
... MMF significantly induced antioxidative genes and reduced inflammatory gene expression, decreased cell loss, and improved retinal function after retinal damage in wild-type mice but not in Nrf2 KO mice, suggesting that Nrf2 modulation plays a major role in retinal protection. Recently, Mori et al. [39] showed that DMF ameliorates RGC survival after optic nerve crush, possibly through the Nrf2/HO-1 axis. DMF treatment prevented retinal injury in a model of light-induced photoreceptor loss, which resulted in retinal degeneration tightly linked to oxidative stress and made the damage sensitive to the antioxidant mode of action of the drug [37]. ...
... The KO models for Nrf2/HO-1 exhibited retinal damage [12,16], with an increase in RGC death and increased levels of inflammatory markers (IL-6, TNF-α, cyclooxygenase 2, COX-2, iNOS, and monocyte chemoattractant protein 1, MCP-1), reverted by the administration of HO-1 inducers [33]. Mori et al. [39] showed that DMF promotes the survival of RGCs after optic nerve crush, possibly via the Nrf2/HO-1 pathway. Moreover, the drug is under clinical investigation (www.clinicaltrial.gov ...
In the area of drug discovery, repurposing strategies represent an approach to discover new uses of approved drugs besides their original indications. We used this approach to investigate the effects of dimethyl fumarate (DMF), a drug approved for relapsing–remitting multiple sclerosis and psoriasis treatment, on early injury associated with diabetic retinopathy (DR). We used an in vivo streptozotocin (STZ)-induced diabetic rat model. Diabetes was induced by a single injection of STZ in rats, and after 1 week, a group of animals was treated with a daily intraperitoneal injection of DMF or a vehicle. Three weeks after diabetes induction, the retinal expression levels of key enzymes involved in DR were evaluated. In particular, the biomarkers COX-2, iNOS, and HO-1 were assessed via Western blot and immunohistochemistry analysis. Diabetic rats showed a significant retinal upregulation of COX-2 and iNOS compared to the retina of normal rats (non-diabetic), and an increase in HO-1 was also observed in the STZ group. This latter result was due to a mechanism of protection elicited by the pathological condition. DMF treatment significantly induced the retinal expression of HO-1 in STZ-induced diabetic animals with a reduction in iNOS and COX-2 retinal levels. Taken together, these results suggested that DMF might be useful to counteract the inflammatory process and the oxidative response in DR. In conclusion, we believe that DMF represents a potential candidate to treat diabetic retinopathy and warrants further in vivo and clinical evaluation.
... Dimethyl fumarate (DMF) is a fumaric acid-derived small molecule that exhibits potent antioxidant and anti-inflammatory properties; DMF is a disease-modifying agent under the brand name "Tecfidera" that has been FDA-approved to be used to treat patients with severe psoriasis and relapsing-remitting multiple sclerosis (RRMS) (9). DMF's antioxidant and anti-inflammatory mechanism of action is thought to involve the activation of the nuclear factor erythroid 2-related factor 2 (Nrf2), which controls the expression of various genes that regulates antioxidant and detoxification processes (10). The activation of the Nrf2 pathway by DMF has been demonstrated in several studies; DMF treatment showed increased Nrf2 protein levels and gene expression, which was accompanied by increased expression of downstream target genes, including heme oxygenase 1 (HO-1) and NAD(P)H: quinone oxidoreductase 1 (NQO1), reduced Nf-kB, TGF-β signalling and cell senescence (11)(12)(13) (14). ...
Background and Purpose Doxorubicin is a broad-spectrum antineoplastic agent; however, however, its genotoxic/cytotoxic effects limit its clinical application. Dimethyl fumarate (DMF) is an FDA-approved oral drug shown to have antioxidant, anti-inflammatory and antimutagenic effects via activating Nrf2 antioxidant pathway. The present study aimed to investigate the possible protective effect of DMF against doxorubicin-induced chromosomal and DNA damage in rat bone marrow cells. Experimental Approach Wistar Albino rats of both sexes were administered DMF orally (15mg/kg once daily for 14 days) alone or with doxorubicin which was injected as a single dose (90 mg/kg at day 14) to induce toxicity. The blood samples were collected 24 hours after doxorubicin’s injection from all groups to measure the serum levels of MDA, GSH, SOD, and GPx1 and bone marrow was harvested to assess chromosomal aberration, micronucleus, and comet assays. Key Results The rats in the doxorubicin-only group exhibited a significant decrease in mitotic index and depleted GSH and antioxidants enzymes serum levels with a significant elevation in MDA serum level, % DNA in Tail, micronucleus appearance and chromosomal aberrations compared to the control group; DMF pretreatment prior to doxorubicin exposure, significantly-reduced % DNA in Tail, micronucleus appearance, and chromosomal aberrations, improved mitotic index, restored GSH level and antioxidant enzymes activity compared doxorubicin-only group. Conclusion and Implication This study revealed that DMF alone has no DNA-damaging or clastogenic activities; DMF has protective effects against the genotoxicity induced by doxorubicin; thus, DMF might be a potential chemoprotective agent against doxorubicin-induced toxicity in cancer chemotherapy
... Particularly, treatment with increasing concentrations of DMF was able to ameliorate the outcome of C57BL/6J mice subjected to optic nerve crush (ONC). An increase in Tubβ3 + RGCs and in the amplitude of positive scotopic threshold response (pSTR) was detected, together with an increase in Nrf2 and HO-1 protein levels [107]. This evidence, together with the evidence collected by You and colleagues [97], suggests a potential application of DMF in the context of glaucoma. ...
Dimethyl fumarate (DMF) is a small molecule currently approved and used in the treatment of psoriasis and multiple sclerosis due to its immuno-modulatory, anti-inflammatory, and antioxidant properties. As an Nrf2 activator through Keap1 protein inhibition, DMF unveils a potential therapeutical use that is much broader than expected so far. In this comprehensive review we discuss the state-of-art and future perspectives regarding the potential repositioning of this molecule in the panorama of eye pathologies, including Age-related Macular Degeneration (AMD). The DMF’s mechanism of action, an extensive analysis of the in vitro and in vivo evidence of its beneficial effects, together with a search of the current clinical trials, are here reported. Altogether, this evidence gives an overview of the new potential applications of this molecule in the context of ophthalmological diseases characterized by inflammation and oxidative stress, with a special focus on AMD, for which our gene–disease (KEAP1-AMD) database search, followed by a protein–protein interaction analysis, further supports the rationale of DMF use. The necessity to find a topical route of DMF administration to the eye is also discussed. In conclusion, the challenge of DMF repurposing in eye pathologies is feasible and worth scientific attention and well-focused research efforts.
... The evidence from studies of animal models of eye diseases shows that DMF also displays beneficial effects at the ocular level. For instance, a recent in vivo study showed that DMF has survival-promoting effects in retinal ganglion cells after optic nerve crush, possibly through the Nrf2/HO-1 pathway [19]. A case report describes a successful therapeutic attempt with DMF for macular edema, an ocular pathology characterized by vascular dysfunction and inflammation [20]. ...
Dimethyl fumarate (DMF) is a well-known activator of Nrf2 (NF-E2-related factor 2), used in the treatment of psoriasis and multiple sclerosis. The mechanism of action consists in the modification of the cysteine residues on the Nrf2-inhibitor Keap1, thus leading to the dissociation of these two proteins and the consequent activation of Nrf2. Considering the paucity of evidence of DMF effects in the context of retinal endothelium, this in vitro study investigated the role of DMF in human retinal endothelial cells (HREC). Here, we show for the first time in HREC that DMF activates the Nrf2 pathway, thus leading to an increase in HO-1 protein levels and a decrease in intracellular ROS levels. Furthermore, this molecule also shows beneficial properties in a model of hyperglucose stress, exerting cytoprotective prosurvival effects. The overall collected results suggest that DMF-mediated activation of the Nrf2 pathway may also be a promising strategy in ocular diseases characterized by oxidative stress. This study opens a new perspective on DMF and suggests its potential repositioning in a broader therapeutical context.
... Nrf2 activation by over-expression protected RGCs in mice from the optic nerve crush (ONC) injury [167]. Pharmacological activators that stabilize Nrf2, such as CDDO-Im [168] and dimethyl fumarate [169], have been found to be RGC protective in mice with ONC injury. Nrf2 activators bardoxolone methyl (RTA-402) [170] and sulforaphane [171] are shown to be RGC protective for retinal ischemia-reperfusion injury in rats. ...
Inherited retinal degeneration is a group of blinding disorders afflicting more than 1 in 4000 worldwide. These disorders frequently cause the death of photoreceptor cells or retinal ganglion cells. In a subset of these disorders, photoreceptor cell death is a secondary consequence of retinal pigment epithelial cell dysfunction or degeneration. This manuscript reviews current efforts in identifying targets and developing small molecule-based therapies for these devastating neuronal degenerations, for which no cures exist. Photoreceptors and retinal ganglion cells are metabolically demanding owing to their unique structures and functional properties. Modulations of metabolic pathways, which are disrupted in most inherited retinal degenerations, serve as promising therapeutic strategies. In monogenic disorders, great insights were previously obtained regarding targets associated with the defective pathways, including phototransduction, visual cycle, and mitophagy. In addition to these target-based drug discoveries, we will discuss how phenotypic screening can be harnessed to discover beneficial molecules without prior knowledge of their mechanisms of action. Because of major anatomical and biological differences, it has frequently been challenging to model human inherited retinal degeneration conditions using small animals such as rodents. Recent advances in stem cell-based techniques are opening new avenues to obtain pure populations of human retinal ganglion cells and retinal organoids with photoreceptor cells. We will discuss concurrent ideas of utilizing stem-cell-based disease models for drug discovery and preclinical development.
... The anti-inflammatory capacity of DMFu showed beneficial effects in an autoimmune uveitis rat model (Labsi et al., 2021) and optic neuritis murine model (Zyla et al., 2019). Further, DMFu promotes the survival of retinal ganglion cells (RGCs) after optic nerve crush (Mori et al., 2020) and protects against retinal degeneration in a light-induced photoreceptor loss mouse model (Dietrich et al., 2020). Mechanistically, studies have shown that DMFu activates Nrf2/heme oxygenase-1 (HO-1) (Zyla et al., 2019;Mori et al., 2020) and increases glutathione (Nelson et al., 1999;Dietrich et al., 2020) to mediate its anti-inflammatory and anti-oxidative effects. ...
... Further, DMFu promotes the survival of retinal ganglion cells (RGCs) after optic nerve crush (Mori et al., 2020) and protects against retinal degeneration in a light-induced photoreceptor loss mouse model (Dietrich et al., 2020). Mechanistically, studies have shown that DMFu activates Nrf2/heme oxygenase-1 (HO-1) (Zyla et al., 2019;Mori et al., 2020) and increases glutathione (Nelson et al., 1999;Dietrich et al., 2020) to mediate its anti-inflammatory and anti-oxidative effects. Intriguingly, in this study, DMFu alone induces significant metabolic changes compared to DMSOtreated control cells. ...
The retinal pigment epithelium (RPE) acts as a metabolic gatekeeper between photoreceptors and the choroidal vasculature to maintain retinal function. RPE dysfunction is a key feature of age-related macular degeneration (AMD), the leading cause of blindness in developed countries. Inflammation is a key pathogenic mechanism in AMD and tumor necrosis factor-alpha (TNFα) has been implicated as a pro-inflammatory cytokine involved in AMD. While mitochondrial dysfunction has been implicated in AMD pathogenesis, the interplay between inflammation and cellular metabolism remains elusive. The present study explores how the pro-inflammatory cytokine, TNFα, impacts mitochondrial morphology and metabolic function in RPE. Matured human primary RPE (H-RPE) were treated with TNFα (10 ng/ml) for up to 5 days. TNFα-induced upregulation of IL-6 secretion and inflammatory genes (IL-6, IL-8, MCP-1) was accompanied by increased oxidative phosphorylation (OXPHOS) and reduced glycolysis, leading to an increase in cellular adenosine triphosphate (ATP) content. Transmission electron microscopy (TEM) revealed defects in mitochondrial morphology with engorged mitochondria and loss of cristae integrity following TNFα treatment. Pre-treatment with the anti-inflammatory drug, 80 μM dimethyl fumarate (DMFu), blocked TNFα-induced inflammatory activation of RPE (IL-6, IL-8, MCP-1, CFH, CFB, C3) and normalized their bioenergetic profile to control levels by regulating PFKFB3 and PKM2 gene expression. Furthermore, DMFu prevented TNFα-induced mitochondrial dysfunction and morphological anomalies. Thus, our results indicate that DMFu serves as a novel therapeutic avenue for combating inflammatory activation and metabolic dysfunction of RPE in AMD.
... Dimethyl fumarate (DMF) is known to prevent pro-oxidant insults through the extracellular signal-regulated kinase ERK1/2-dependent activation of Nrf2/HO-1 pathway, restoring the functions of mitochondria in different models such as cardiomyocytes [26], retinal pigment epithelium (RPE) [27] and retinal ganglion cells [28]. Moreover, when tested in diabetic conditions, DMF ameliorated epithelial wound healing and reduced both oxidative damage and inflammation in vitro and in vivo, producing the activation of Nrf2 signaling [29,30]. ...
Glucose induces corneal epithelial dysfunctions characterized by delayed wound repair. Nuclear erythroid 2-related factor 2 (Nrf2) mediates cell protection mechanisms even through the Heme oxygenase-1 (HO-1) up-regulation. Here, we synthesized new HO-1 inducers by modifying dimethyl fumarate (DMF) and used docking studies to select VP13/126 as a promising compound with the best binding energy to Kelch-like ECH-associated protein 1 (keap1), which is the the regulator of Nrf2 nuclear translocation. We verified if VP13/126 protects SIRC cells from hyperglycemia compared to DMF. SIRC were cultured in normal (5 mM) or high glucose (25 mM, HG) in presence of DMF (1–25 μM) or VP13/126 (0.1–5 μM) with or without ERK1/2 inhibitor PD98059 (15 μM). VP13/126 was more effective than DMF in the prevention of HG-induced reduction of cell viability and proliferation. Reduction of wound closure induced by HG was similarly counteracted by 1 μM VP13/126 and 10 μM DMF. VP13/126 strongly increased phospho/total ERK1/2 and restored HO-1 protein in HG-treated SIRC; these effects are completely counteracted by PD98059. Moreover, high-content screening analysis showed a higher rate of Nrf2 nuclear translocation induced by VP13/126 than DMF in HG-stimulated SIRC. These data indicate that VP13/126 exerts remarkable pro-survival properties in HG-stimulated SIRC, promoting the Nrf2/HO-1 axis.
The aim of this study was to investigate the effects of 7,8-dihydroxyflavone (7,8-DHF) in protecting retinal ganglion cells (RGCs) and promoting axonal regeneration, and to explore its potential molecular mechanisms. We used three-dimensional retinal culture system and optic nerve crush (ONC) rat models in this study. The pro-axonal regenerative effect of 7,8-DHF was determined with light microscopy observation and immunofluorescence staining of Thy1.1 and GAP43. The RGC protective function of 7,8-DHF was detected by RBPMS immunofluorescent staining and TUNEL staining. The inhibition effect of 7,8-DHF on astrocyte activation was measured using GFAP immunofluorescence and Western blotting. The protein levels of p-TrkB, p-AKT and p-ERK was examined by Western blotting and immunohistochemistry. Our results revealed that 7,8-DHF significantly promoted the average density and length of regenerated neurites and suppressed the apoptosis of GCL cells in three-dimensional culture system and significantly increased the number of RBPMS-positive cells and inhibited the GFAP expression and apoptosis of GCL cells in ONC rats. Our results also revealed that 7,8-DHF activates TrkB, AKT and ERK proteins in vivo, however, these activations can be inhibited byANA-12. In conclusion, 7,8-DHF protects RGCs and promotes axonal regeneration through the TrkB signaling pathway followed by AKT and ERK activation.
Background
Experimental studies on retinal vasculature and retinal ganglion cells (RGCs) have investigated developmental and pathological conditions of the retina; they rely on whole-mount retinal immunostaining. Methanol, an auxiliary fixed medium for retinal whole-mount preparations, has been used in some studies; however, its application in short- and long-term storage of retinas for further study has not been well described. We aimed to evaluate methanol use as a preservation treatment for further immunostaining of the retina.
Methods
We generated the oxygen-induced retinopathy (OIR) and optic nerve crush (ONC) mouse models and used their retinas for analysis. We pipetted cold methanol (−20 °C) on the surface of the retina to help fix the tissues while promoting permeability, after which the retinas were stored in cold methanol (−20 °C) for 1, 6, or 12 months before being evaluated using various optical techniques. Thereafter, retinal whole-mount immunostaining was performed to analyse retinal neovascularisation and retinal hypoxia in OIR model, and retinal ganglion cell survival rate in ONC model.
Results
Quantitative analysis revealed no significant differences in these fixed retinas after long-term storage in terms of retinal vasculature or retinal hypoxia in OIR model. Similarly, no significant difference was found in RGC survival rate after long-term storage in methanol. These results suggest that methanol can be used as a storage medium when preserving retinal whole-mount samples.
Conclusions
Cold (−20 °C) methanol is a good medium for storing fixed retinas for long periods, which is useful when retinas must be stored for further analysis.
