Small-fiber painful peripheral neuropathy is one of the long-term complications of alcohol for which there is no reliable successful therapy available. The precise mechanisms by which chronic alcohol consumption produces peripheral nerve fiber damage and loss remain unclear. Emerging data from clinical and preclinical studies suggest that increased oxidative-nitrodative stress mediated release of proinflammatory cytokines from damaged neural tissues may play a central role in the pathogenesis of alcoholic neuropathy. The present study investigated the effect of both the isoforms of vitamin E (α-tocopherol and tocotrienol) against chronic alcohol-induced peripheral neuropathy in rats. Ethanol treated rats showed a significant decrease in paw-withdrawal threshold in both Randall-Selitto and von-Frey hair tests along with a significant reduction in tail flick latency in the tail-immersion test. A decreased pain threshold was associated with significant alterations in oxidative-nitrodative stress markers and an increase in proinflammatory cytokines (TNF-α and IL-1β). The 4-week treatment with tocotrienol significantly ameliorated behavioral, biochemical and molecular alterations in alcohol treated rats. However, α-tocopherol failed to produce any protective effect. The results of the present study suggest that oxidative-nitrodative stress mediated cytokine signaling may be responsible for alcohol-induced peripheral neurotoxicity and tocotrienol treatment might be beneficial in chronic alcoholics exhibiting neuropathy

Vitamin E includes 8 natural compounds (4 tocopherols, 4 tocotrienols) with potential neuroprotective activity. α-Tocopherol has mainly been investigated in relation to cognitive impairment. We examined the relation of all plasma vitamin E forms and markers of vitamin E damage (α-tocopherylquinone, 5-nitro-γ-tocopherol) to mild cognitive impairment (MCI) and Alzheimer’s disease (AD). Within the AddNeuroMed-Project, plasma tocopherols, tocotrienols, α-tocopherylquinone, and 5-nitro-γ-tocopherol were assessed in 168 AD cases, 166 MCI, and 187 cognitively normal (CN) people. Compared with cognitively normal subjects, AD and MCI had lower levels of total tocopherols, total tocotrienols, and total vitamin E. In multivariable-polytomous-logistic regression analysis, both MCI and AD cases had 85% lower odds to be in the highest tertile of total tocopherols and total vitamin E, and they were, respectively, 92% and 94% less likely to be in the highest tertile of total tocotrienols than the lowest tertile. Further, both disorders were associated with increased vitamin E damage. Low plasma tocopherols and tocotrienols levels are associated with increased odds of MCI and AD.

Ethanol-induced damage in the developing brain may result in cognitive impairment including deficits on neuropsychological tests of learning, memory and executive function, yet the underlying mechanisms remain elusive. In the present study we investigated the protective effect of tocotrienol against cognitive deficit, neuroinflammation and neuronal apoptosis in rat pups postnatally exposed to ethanol. Pups were administered ethanol (5g/kg, 12% v/v) by intragastric intubation on postnatal days 7, 8 and 9. Ethanol-exposed pups showed significant memory impairment in Morris water maze task as evident from increase in escape latency and total distance travelled to reach the hidden platform. Time spent in target quadrant, % total distance traversed in target quadrant and frequency of appearance in target quadrant was also significantly decreased in ethanol exposed pups in probe trial. Poor memory retention was exhibited by ethanol-exposed pups in elevated plus maze test also. Impaired cognition was associated with significantly enhanced acetylcholinesterase activity, increased neuroinflammation (oxidative-nitrosative stress, TNF-α, IL-1β and TGF-β1) and neuronal apoptosis (NF-κβ and Caspase-3) in different brain regions of ethanol-exposed pups. Co-administration with tocotrienol significantly ameliorated all the behavioral, biochemical and molecular alterations in the different brain regions of ethanol exposed pups. The current study thus demonstrates the possible involvement of NF-κβ mediated apoptotic signaling in cognitive deficits associated with postnatal ethanol exposure in rats and points to the potential of tocotrienol in the prevention of cognitive deficits in children with fetal alcohol spectrum disorders (FASDs).

It is well known that reactive oxygen species (ROS) attack several living tissues and increase the risk of development and progression of serious diseases. In neuronal level, ROS induce cell death in concentration-dependent fashion. However, little is known about the mechanisms of neuronal changes by ROS prior to induction of cell death. Here we found that treatment of cerebellar granule neurons (CGCs) with 0.5 μM hydrogen peroxide induced axonal injury, but not cell death. The number of dendrites remarkably decreased in hydrogen peroxide-treated CGCs, and extensive beading was observed on survival dendrites. In addition, an abnormal band of the original collapsin response mediator protein (CRMP)-2 was detected by Western blotting in hydrogen peroxide-treated CGCs. Treatment with each tocotrienol isoform prevented axonal and dendrite degeneration and induction of the abnormal band of the original band of CRMP-2 in hydrogen peroxide-treated CGCs. These results indicate that treatment with tocotrienols may therefore be neuroprotective in the presence of hydrogen peroxide by preventing changes to the CRMP-2 that occur before neuron death.

Background: Down syndrome (DS) neurons are more susceptible to oxidative stress and previous studies have shown that vitamin E was able to reduce oxidative stress and improve DS neurons’ viability. Therefore, this study was done to investigate the protective role of γ-tocotrienol (γT3) in DS neurons from hydrogen peroxide (H2O2) -induced oxidative stress. The pro-apoptosis tendency of γT3 was compared to α-tocopherol (αT) in non-stress condition as well.

Methods: Primary culture of DS and euploid neurons were divided into six groups of treatment: control, H2O2, γT3 pre-treatment with H2O2, γT3 only, αT pre-treatment with H2O2 and αT only. The treatments were assessed by MTS assay and apoptosis assay by single-stranded DNA (ssDNA) apoptosis ELISA assay, Hoechst and Neu-N immunofluorescence staining. The cellular uptake of γT3 and αT was determined by HPLC while protein expressions were determined by Western blot. Comparison between groups was made by the Student’s t test, one-way ANOVA and Bonferroni adjustment as well as two-way ANOVA for multiple comparisons.

Results: One day incubation of γT3 was able to reduced apoptosis of DS neurons by 10%, however γT3 was cytotoxic at longer incubation period (14 days) and at concentrations ≥ 100 μM. Pre-treatment of αT and γT3 only attenuate apoptosis and increase cell viability in H2O2-treated DS and euploid neurons by 10% in which the effects were minimal to maintain most of the DS cells’ morphology. γT3 act as a free radical scavenger by reducing ROS generated by H2O2. In untreated controls, DS neurons showed lower Bcl-2/Bax ratio and p53 expression compared to normal neurons, while cPKC and PKC-δ expressions were higher in DS neurons. On the other hand, pre-treatment of γT3 in H2O2-treated DS neurons have reduced Bcl-2/Bax ratio, which was not shown in euploid neurons. This suggests that pre-treatment of γT3 did not promote DS cell survival. Meanwhile γT3 and αT treatments without H2O2 as well as pre-treatment of γT3 and αT induced changes in cPKC and PKC-δ expression in DS neurons suggesting interaction of γT3 and αT with PKC activity.

Conclusion: Our study suggests that γT3 pre-treatment are not sufficient to protect DS neurons from H2O2-induced oxidative assault, instead induced the apoptosis process.

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Tocotrienols (T(3)) belong to the family of vitamin E compounds (α-, β-, γ-, δ-tocopherols and -tocotrienols) and have unique biological properties that make them potential neuroprotective dietary factors. In addition to their antioxidant activity, T(3) at micromolar concentrations exert cholesterol-lowering activities in cells, animal models and some, but not all, human studies by means of inhibition of the activity of the rate-limiting enzyme in cholesterol biosynthesis, 3-hydroxy-3-methylglutaryl coenzyme A reductase. At lower concentrations (∼10nmol/L), T(3) modulate signalling pathways involved in neuronal cell death in cell culture experiments. Targets of T(3) include prenyl transferases, non-receptor tyrosine kinase, phospholipase A(2), 12-lipoxygenase, cyclooxygenase-2, and nuclear factor κB. The low bioavailability and rapid excretion of T(3) represents a major hurdle in their preventive use. Fasting plasma concentrations, even after supplementation with high doses, are below 1μmol/L. T(3) bioavailability may be enhanced by ingestion with a high-fat meal, self-emulsifying drug delivery systems, or phytochemicals that inhibit T(3) metabolism and excretion. T(3) have no known adverse effects when consumed as part of a normal diet and the studies reviewed here support the notion that they may have potential as neuroprotective agents. However, experiments in relevant animal models and randomised human intervention trials addressing the neuroprotection mediated   by T(3) are scarce and, thus, highly warranted.

Vitamin E consists of tocopherols and tocotrienols, in which α-tocotrienol is the most potent neuroprotective form that is also effective in protecting against stroke in rodents. As neuroprotective agents alone are insufficient to protect against stroke, we sought to test the effects of tocotrienol on the cerebrovascular circulation during ischemic stroke using a preclinical model that enables fluoroscopy-guided angiography. Mongrel canines (mean weight=26.3±3.2 kg) were supplemented with tocotrienol-enriched (TE) supplement (200 mg b.i.d, n=11) or vehicle placebo (n=9) for 10 weeks before inducing transient middle cerebral artery (MCA) occlusion. Magnetic resonance imaging was performed 1 hour and 24 hours post reperfusion to assess stroke-induced lesion volume. Tocotrienol-enriched supplementation significantly attenuated ischemic stroke-induced lesion volume (P<0.005). Furthermore, TE prevented loss of white matter fiber tract connectivity after stroke as evident by probabilistic tractography. Post hoc analysis of cerebral angiograms during MCA occlusion revealed that TE-supplemented canines had improved cerebrovascular collateral circulation to the ischemic MCA territory (P<0.05). Tocotrienol-enriched supplementation induced arteriogenic tissue inhibitor of metalloprotease 1 and subsequently attenuated the activity of matrix metalloproteinase-2. Outcomes of the current preclinical trial set the stage for a clinical trial testing the effects of TE in patients who have suffered from transient ischemic attack and are therefore at a high risk for stroke.

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Background & Purpose: α-Tocotrienol (TCT) represents the most potent neuroprotective form of natural vitamin E that is Generally Recognized As Safe certified by the U.S. Food and Drug Administration. This work addresses a novel molecular mechanism by which α-TCT may be protective against stroke in vivo. Elevation of intracellular oxidized glutathione (GSSG) triggers neural cell death. Multidrug resistance-associated protein 1 (MRP1), a key mediator of intracellular oxidized glutathione efflux from neural cells, may therefore possess neuroprotective functions.

Methods: Stroke-dependent brain tissue damage was studied in MRP1-deficient mice and α-TCT-supplemented mice.

Results: Elevated MRP1 expression was observed in glutamate-challenged primary cortical neuronal cells and in stroke-affected brain tissue. MRP1-deficient mice displayed larger stroke-induced lesions, recognizing a protective role of MRP1. In vitro, protection against glutamate-induced neurotoxicity by α-TCT was attenuated under conditions of MRP1 knockdown; this suggests the role of MRP1 in α-TCT-dependent neuroprotection. In vivo studies demonstrated that oral supplementation of α-TCT protected against murine stroke. MRP1 expression was elevated in the stroke-affected cortical tissue of α-TCT-supplemented mice. Efforts to elucidate the underlying mechanism identified MRP1 as a target of microRNA (miR)-199a-5p. In α-TCT-supplemented mice, miR-199a-5p was downregulated in stroke-affected brain tissue.

Conclusion: This work recognizes MRP1 as a protective factor against stroke. Furthermore, findings of this study add a new dimension to the current understanding of the molecular bases of α-TCT neuroprotection in 2 ways: by identifying MRP1 as a α-TCT-sensitive target and by unveiling the general prospect that oral α-TCT may regulate miR expression in stroke-affected brain tissue.

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