Dietary Omega-3 Polyunsaturated Fatty-Acid Supplementation Upregulates Protective Cellular Pathways in Patients with Type 2 Diabetes Exhibiting Improvement in Painful Diabetic Neuropathy.

BACKGROUND: Omega-3 polyunsaturated fatty acids (PUFAs) have been proposed to improve chronic neuroinflammatory diseases in peripheral and central nervous systems. For instance, docosahexaenoic acid (DHA) protects nerve cells from noxious stimuli in vitro and in vivo. Recent reports link PUFA supplementation to improving painful diabetic neuropathy (pDN) symptoms, but cellular mechanisms responsible for this therapeutic effect are not well understood. The objective of this study is to identify distinct cellular pathways elicited by dietary omega-3 PUFA supplementation in patients with type 2 diabetes mellitus (T2DM) affected by pDN. METHODS: Forty volunteers diagnosed with type 2 diabetes were enrolled in the "En Balance-PLUS" diabetes education study. The volunteers participated in weekly lifestyle/nutrition education and daily supplementation with 1000 mg DHA and 200 mg eicosapentaenoic acid. The Short-Form McGill Pain Questionnaire validated clinical determination of baseline and post-intervention pain complaints. Laboratory and untargeted metabolomics analyses were conducted using blood plasma collected at baseline and after three months of participation in the dietary regimen. The metabolomics data were analyzed using random forest, hierarchical clustering, ingenuity pathway analysis, and metabolic pathway mapping. RESULTS: The data show that metabolites involved in oxidative stress and glutathione production shifted significantly to a more anti-inflammatory state post supplementation. Example of these metabolites include cystathionine (+90%), S-methylmethionine (+9%), glycine cysteine-glutathione disulfide (+157%) cysteinylglycine (+19%), glutamate (-11%), glycine (+11%), and arginine (+13.4%). In addition, the levels of phospholipids associated with improved membrane fluidity such as linoleoyl-docosahexaenoyl-glycerol (18:2/22:6) (+253%) were significantly increased. Ingenuity pathway analysis suggested several key bio functions associated with omega-3 PUFA supplementation such as formation of reactive oxygen species (p = 4.38 × 10-4, z-score = -1.96), peroxidation of lipids (p = 2.24 × 10-5, z-score = -1.944), Ca2+ transport (p = 1.55 × 10-4, z-score = -1.969), excitation of neurons (p = 1.07 ×10-4, z-score = -1.091), and concentration of glutathione (p = 3.06 × 10-4, z-score = 1.974). CONCLUSION: The reduction of pro-inflammatory and oxidative stress pathways following dietary omega-3 PUFA supplementation is consistent with the promising role of these fatty acids in reducing adverse symptoms associated with neuroinflammatory diseases and painful neuropathy.

Effects of omega-3 polyunsaturated fatty-acid supplementation on neuropathic pain symptoms and sphingosine levels in Mexican-Americans with type 2 diabetes.

PURPOSE: To determine whether dietary supplementation with omega-3 polyunsaturated fatty acids (PUFAs) reduces neuropathic pain symptoms in Mexican-Americans with type 2 diabetes. METHODS: Forty volunteers with type 2 diabetes enrolled in the "En Balance-PLUS" program, which provided weekly nutrition-diabetes education and daily supplementation with 1,000 mg docosahexaenoic acid (DHA)-200 mg eicosapentaenoic acid over 3 months. The study assessed self-reported neuropathic pain symptoms pre/postintervention using the short-form McGill Pain Questionnaire (SF-MPQ), monitored clinical laboratory values at baseline and 3 months, and performed baseline and 3-month metabolomic analysis of plasma samples. RESULTS: A total of 26 participants self-reported neuropathic pain symptoms at baseline. After 3 months of omega-3 PUFA supplementation, participants reported significant improvement in SF-MPQ scores (sensory, affective, and visual analogue scale; P<0.001, P=0.012, and P<0.001, respectively). Untargeted metabolomic analysis revealed that participants in the moderate-high SF-MPQ group had the highest relative plasma sphingosine levels at baseline compared to the low SF-MPQ group (P=0.0127) and the nonpain group (P=0.0444). Omega-3 PUFA supplementation increased plasma DHA and reduced plasma sphingosine levels in participants reporting neuropathic pain symptoms (P<0.001 and P<0.001, respectively). Increased plasma DHA levels significantly correlated with improved SF-MPQ sensory scores (r=0.425, P=0.030). Improved SF-MPQ scores, however, did not correlate with clinical/laboratory parameters. CONCLUSION: The data suggest that omega-3 PUFAs dietary supplementation may reduce neuropathic pain symptoms in individuals with type 2 diabetes and correlates with sphingosine levels in the plasma.

Effects of Dietary Vitamin E Supplementation in Bladder Function and Spasticity during Spinal Cord Injury.

Traumatic spinal cord injury (SCI) results in debilitating autonomic dysfunctions, paralysis and significant sensorimotor impairments. A key component of SCI is the generation of free radicals that contributes to the high levels of oxidative stress observed. This study investigates whether dietary supplementation with the antioxidant vitamin E (alpha-tocopherol) improves functional recovery after SCI. Female adult Sprague-Dawley rats were fed either with a normal diet or a dietary regiment supplemented with vitamin E (51 IU/g) for eight weeks. The rats were subsequently exposed either to a contusive SCI or sham operation, and evaluated using standard functional behavior analysis. We report that the rats that consumed the vitamin E-enriched diet showed an accelerated bladder recovery and significant improvements in locomotor function relative to controls, as determined by residual volumes and Basso, Beatie, and Bresnaham BBB scores, respectively. Interestingly, the prophylactic dietary intervention did not preserve neurons in the ventral horn of injured rats, but it significantly increased the numbers of oligodendrocytes. Vitamin E supplementation attenuated the depression of the H-reflex (a typical functional consequence of SCI) while increasing the levels of supraspinal serotonin immunoreactivity. Our findings support the potential complementary use of vitamin E to ameliorate sensory and autonomic dysfunctions associated with spinal cord injury, and identified promising new cellular and functional targets of its neuroprotective effects.

Neurorestorative targets of dietary long-chain omega-3 fatty acids in neurological injury.

Long-chain omega-3 polyunsaturated fatty acids (LC-O3PUFAs) exhibit therapeutic potential for the treatment and prevention of the neurological deficits associated with spinal cord injury (SCI). However, the mechanisms implicated in these protective responses remain unclear. The objective of the present functional metabolomics study was to identify and define the dominant metabolic pathways targeted by dietary LC-O3PUFAs. Sprague-Dawley rats were fed rodent purified chows containing menhaden fish oil-derived LC-O3PUFAs for 8 weeks before being subjected to sham or spinal cord contusion surgeries. We show, through untargeted metabolomics, that dietary LC-O3PUFAs regulate important biochemical signatures associated with amino acid metabolism and free radical scavenging in both the injured and sham-operated spinal cord. Of particular significance, the spinal cord metabolome of animals fed with LC-O3PUFAs exhibited reduced glucose levels (-48 %) and polar uncharged/hydrophobic amino acids (less than -20 %) while showing significant increases in the levels of antioxidant/anti-inflammatory amino acids and peptides metabolites, including ß-alanine (+24 %), carnosine (+33 %), homocarnosine (+27 %), kynurenine (+88 %), when compared to animals receiving control diets (p < 0.05). Further, we found that dietary LC-O3PUFAs impacted the levels of neurotransmitters and the mitochondrial metabolism, as evidenced by significant increases in the levels of N-acetylglutamate (+43 %) and acetyl CoA levels (+27 %), respectively. Interestingly, this dietary intervention resulted in a global correction of the pro-oxidant metabolic profile that characterized the SCI-mediated sensorimotor dysfunction. In summary, the significant benefits of metabolic homeostasis and increased antioxidant defenses unlock important neurorestorative pathways of dietary LC-O3PUFAs against SCI.

Dietary omega-3 polyunsaturated fatty acids improve the neurolipidome and restore the DHA status while promoting functional recovery after experimental spinal cord injury.

Omega-3 polyunsaturated fatty acids (ω-3 PUFAs) confer multiple health benefits and decrease the risk of neurological disorders. Studies are needed, however, to identify promising cellular targets and to assess their prophylactic value against neurodegeneration. The present study (1) examined the efficacy of a preventive diet enriched with ω-3 PUFAs to reduce dysfunction in a well-established spinal cord injury (SCI) animal model and (2) used a novel metabolomics data analysis to identify potential neurolipidomic targets. Rats were fed with either control chow or chow enriched with ω-3 PUFAs (750 mg/kg/day) for 8 weeks before being subjected to a sham or a contusion SCI operation. We report new evidence showing that rats subjected to SCI after being pre-treated with a diet enriched with ω-3 PUFAs exhibit significantly better functional outcomes. Pre-treated animals exhibited lower sensory deficits, autonomic bladder recovery, and early improvements in locomotion that persisted for at least 8 weeks after trauma. We found that SCI triggers a robust alteration in the cord PUFA neurolipidome, which was characterized by a marked docosahexaenoic acid (DHA) deficiency. This DHA deficiency was associated with dysfunction and corrected with the ω-3 PUFA-enriched diet. Multivariate data analyses revealed that the spinal cord of animals consuming the ω-3 PUFA-enriched diet had a fundamentally distinct neurolipidome, particularly increasing the levels of essential and long chain ω-3 fatty acids and lysolipids at the expense of ω-6 fatty acids and its metabolites. Altogether, dietary ω-3 PUFAs prophylaxis confers resiliency to SCI mediated, at least in part, by generating a neuroprotective and restorative neurolipidome.

Expression of E-FABP in PC12 cells increases neurite extension during differentiation: involvement of n-3 and n-6 fatty acids.

Epidermal fatty acid-binding protein (E-FABP), a member of the family of FABPs, exhibits a robust expression in neurons during axonal growth in development and in nerve regeneration following nerve injury. This study examines the impact of E-FABP expression in normal neurite extension in differentiating pheochromocytoma cell (PC12) cultures supplemented with selected long chain free fatty acids (LCFFA). We found that E-FABP binds to a broad range of saturated and unsaturated LCFFAs, including those with potential interest for neuronal differentiation and axonal growth such as C22:6n-3 docosahexaenoic acid (DHA), C20:5n-3 eicosapentaenoic acid (EPA), and C20:4n-6 arachidonic acid (ARA). PC12 cells exposed to nerve growth factor (NGFDPC12) exhibit high E-FABP expression that is blocked by mitogen-activated protein kinase kinase (MEK) inhibitor U0126. Nerve growth factor-differentiated pheochromocytoma cells (NGFDPC12) antisense clones (NGFDPC12-AS) which exhibit low E-FABP expression have fewer/shorter neurites than cells transfected with vector only or NGFDPC12 sense cells (NGFDPC12-S). Replenishing NGFDPC12-AS cells with biotinylated recombinant E-FABP (biotin-E-FABP) protein restores normal neurite outgrowth. Cellular localization of biotin-E-FABP in NGFDPC12 was detected mostly in the cytoplasm and in the nuclear region. Treatment of NGFDPC12 with DHA, EPA, or ARA further enhances neurite length but it does not trigger further induction of TrkA or MEK phosphorylation or E-FABP mRNA observed in differentiating PC12 cells without LCFFA supplementation. Significantly, DHA and EPA neurite stimulating effects are higher in NGFDPC12-S than in NGFDPC12-AS cells. These findings are consistent with the scenario that neurite extension of differentiating PC12 cells, including further stimulation by DHA and EPA, requires sufficient cellular levels of E-FABP.