MetS confers an increased risk of future diabetes and CVD [1
]. Several studies have shown that MetS is also associated with increased inflammation and oxidative stress. The NCEP has recommended that therapeutic lifestyle changes are the primary treatment strategy for MetS. Dietary micronutrients with anti-oxidant and anti-inflammatory potential could be alternative strategies to reduce the cardiovascular burden in MetS. AT is a potent antioxidant and anti-inflammatory agent; however, prospective CVD clinical trials have not shown promising results [3
]. This could in part be explained by the fact that AT supplementation results in decreased levels of GT, the main form of dietary vitamin E. While previous supplementation studies with mixed tocopherols that were enriched with GT have yielded significant reductions in biomarkers of oxidative and nitrative stress, no studies have been conducted with purified GT alone in MetS [7
]. Thus, in this study, we provide novel data with regard to purified GTsupplementation compared to ATor AT+GT alone on bioavailability of AT and GTas well as on biomarkers of oxidative stress and inflammation in MetS subjects.
In accordance with other studies, we show that plasma AT levels increased significantly with AT supplementation following 8 weeks of supplementation with ATalone or in combination with GT. Furthermore, previous studies have shown that AT supplementation decreases plasma GT levels as observed in our study (37% decrease) [6
]. It has been suggested that AT can replace GT in lipid membranes, resulting in decreased plasma GT levels and thus mixed tocopherol supplementation could potentially avoid adverse biological effects. We show here that the decrease in GT levels with AT supplementation is abrogated with the combination of AT and GT. The concentration of tocopherols appears to be saturable, resulting in increased metabolism to CEHCs. In healthy subjects, previously, it has been suggested that urinary CEHC can be used as a biomarker of adequate tocopherol stores [19
]. In this study, we have performed comprehensive analyses of both plasma and urinary CEHCs and show that AT supplementation resulted in increased levels A-CEHC and, for the first time, show that with purified GT supplementation, there is increased G-CEHC with no effect on A-CEHC. It is also interesting to note that the combination of AT+GT resulted in significantly increased A- and G-CEHCs; however, the level of A-CEHCs was significantly decreased compared to AT supplementation alone, indicating the importance of the hepatic AT transport protein in regulating the metabolic fate of the tocopherols and the preference for GT metabolism to G-CEHC potentially interfering with AT metabolism.
CRP is a prototypic marker of inflammation and a risk marker for CVD [20
]. In addition, several lines of evidence indicate that CRP may be an active participant in atherosclerosis. Also, studies have reported that AT supplementation (≥800 IU/day) results in significant decreases in hsCRP levels [21
]. However, there is a paucity of data examining tocopherol supplementation in MetS subjects and no data with regard to purified GT supplementation on biomarkers of inflammation in MetS. In this placebo-controlled study, we report that only the combination of AT+ GT resulted in significant reduction in hsCRP compared to placebo and either AT or GT supplementation significantly decreased HsCRP levels compared to baseline but these were not significantly different from those of placebo. In a previous small study, there was a significant reduction in hsCRP in hemodialysis patients following administration of GT-enriched mixed tocopherols (600 mg/day for 14 days) but not AT supplementation alone [11
]. Furthermore, the effect of GT supplementation alone was not studied. Recently, Wu et al. [13
] reported that neither AT or mixed tocopherol supplementation (500 mg/day for 6 weeks) resulted in any significant changes in CRP, MCP-1, IL-6, or TNF-α levels. In accordance with these studies, we also failed to observe any significant change in whole blood release of cytokines, except TNF-α, which was significantly different with AT alone or AT+GT therapy.
Several studies have demonstrated that AT has antioxidant effects. In this study, we show that GT supplementation alone significantly reduces biomarkers of oxidative stress, plasma MDA and HNE, and lipid peroxides, similar to AT alone or the combination. Previous studies in vitro have shown that GT is also highly potent in quenching reactive nitrogen species compared to AT [23
]. Thus, we also examined urinary nitrotyrosine levels following GT supplementation alone and in combination with AT. While AT alone failed to affect urinary nitrotyrosine, GT alone as well as in combination with AT resulted in significant reduction in urinary nitrotyrosine. Previously, Cooney et al. [24
] have shown that GT reacts with nitrogen dioxide to produce nitric oxide while AT forms an intermediate analog, such as the quinone in vitro. Also, AT and nitric oxide cooperatively inhibit lipid peroxidation better than AT/ascorbate combination in vitro [25
]. However, it is important to note that these studies were performed in an vitro system. In the present study, in vivo, it appears that GT is more potent than AT in decreasing nitrosative stress, and this needs to be confirmed in future larger studies, also examining mechanisms for these observations.
Thus, in conclusion, we report for the first time in MetS subjects that the combination of AT + GT therapy results in significant increases in AT and GT concentrations as well as their metabolites in plasma and urine, as well as significant reductions in hsCRP, urinary nitrotyrosine, and lipid peroxides. These results point to the superiority of combined AT + GT supplementation in ameliorating both oxidative and nitrative stress and inflammation in MetS subjects. Future studies will be directed at examining mechanisms for these changes and testing the effect of combined supplementation on cardiovascular events in high risk populations such as chronic kidney disease and metabolic syndrome.