Inhibition of myeloperoxidase: evaluation of 2H-indazoles and 1H-indazolones.

Myeloperoxidase (MPO) produces hypohalous acids as a key component of the innate immune response; however, release of these acids extracellularly results in inflammatory cell and tissue damage. The two-step, one-pot Davis-Beirut reaction was used to synthesize a library of 2H-indazoles and 1H-indazolones as putative inhibitors of MPO. A structure-activity relationship study was undertaken wherein compounds were evaluated utilizing taurine-chloramine and MPO-mediated H2O2 consumption assays. Docking studies as well as toxicophore and Lipinski analyses were performed. Fourteen compounds were found to be potent inhibitors with IC50 values <1µM, suggesting these compounds could be considered as potential modulators of pro-oxidative tissue injury pertubated by the inflammatory MPO/H2O2/HOCl/HOBr system.

Alpha-Tocopherol from People to Plants Is an Essential Cog in the Metabolic Machinery.

Significance: Protection from oxygen, a diradical, became a necessity with the evolution of photosynthetic organisms about 2.7 billion years. α-Tocopherol plays an essential, protective role in organisms from plants to people. An overview of human conditions that result in severe vitamin E (α-tocopherol) deficiency is provided. Recent Advances: α-Tocopherol has a critical role in the oxygen protection system by stopping lipid peroxidation, its induced damage, and cellular death by ferroptosis. Recent findings in bacteria and plants support the concept of why lipid peroxidation is so dangerous to life and why the family of tocochromanols are essential for aerobic organisms and for plants. Critical Issues: The hypothesis that prevention of the propagation of lipid peroxidation is the basis for the α-tocopherol requirement in vertebrates is proposed and further that its absence dysregulates energy metabolism, one-carbon metabolism, and thiol homeostasis. By recruiting intermediate metabolites from adjacent pathways to sustain effective lipid hydroperoxide elimination, α-tocopherol function is linked not only to NADPH metabolism and its formation through the pentose phosphate pathway via glucose metabolism but also to sulfur-containing amino acid metabolism and to one-carbon metabolism. Future Directions: Evidence from humans, animals, and plants supports the hypothesis, but future studies are needed to assess the genetic sensors that detect lipid peroxidation and cause the ensuing metabolic dysregulation. Antioxid. Redox Signal. 38, 775-791.

A high-fat diet containing whole walnuts (Juglans regia) reduces tumour size and growth along with plasma insulin-like growth factor 1 in the transgenic adenocarcinoma of the mouse prostate model.

Prostate cancer (PCa) has been linked to fat intake, but the effects of both different dietary fat levels and types remain inconsistent and incompletely characterised. The effects on PCa in the transgenic adenocarcinoma of the mouse prostate (TRAMP) cancer model of an elevated fat (20 % of energy as fat) diet containing 155 g of whole walnuts were compared to those of an elevated fat (20 % of energy as soyabean oil) diet with matched macronutrients, tocopherols as well as a low-fat (8 % of energy as soyabean oil) diet. Mice, starting at 8 weeks of age, consumed one of the three different diets ad libitum; and prostates, livers and blood were obtained after 9, 18 or 24 weeks of feeding. No differences were observed in whole animal growth rates in either high-fat (HF) diet group, but prostate tumour weight and growth rate were reduced in the walnut diet group. Walnut diet group prostate weight, plasma insulin-like growth factor 1, resistin and LDL were lower at 18 weeks, while no statistically significant prostate weight differences by diet were seen at 9 or 24 weeks. Multiple metabolites in the livers differed by diet at 9 and 18 weeks. The walnut diet's beneficial effects probably represent the effects of whole walnuts' multiple constituents and not via a specific fatty acid or tocopherols. Moreover, as the two HF diets had dissimilar effects on prostate tumour growth rate and size, and yet had the same total fat and tocopherol composition and content, this suggests that these are not strongly linked to PCa growth.

α-Tocopherol Pharmacokinetics in Adults with Cystic Fibrosis: Benefits of Supplemental Vitamin C Administration.

BACKGROUND: Numerous abnormalities in cystic fibrosis (CF) could influence tocopherol absorption, transportation, storage, metabolism and excretion. We hypothesized that the oxidative distress due to inflammation in CF increases vitamin E utilization, which could be positively influenced by supplemental vitamin C administration. METHODS: Immediately before and after receiving vitamin C (500 mg) twice daily for 3.5 weeks, adult CF patients (n = 6) with moderately advanced respiratory tract (RT) disease consumed a standardized breakfast with 30% fat and a capsule containing 50 mg each hexadeuterium (d6)-α- and dideuterium (d2)-γ-tocopheryl acetates. Blood samples were taken frequently up to 72 h; plasma tocopherol pharmacokinetics were determined. During both trials, d6-α- and d2-γ-tocopherols were similarly absorbed and reached similar maximal plasma concentrations ~18-20 h. As predicted, during vitamin C supplementation, the rates of plasma d6-α-tocopherol decline were significantly slower. CONCLUSIONS: The vitamin C-induced decrease in the plasma disappearance rate of α-tocopherol suggests that vitamin C recycled α-tocopherol, thereby augmenting its concentrations. We conclude that some attention should be paid to plasma ascorbic acid concentrations in CF patients, particularly to those individuals with more advanced RT inflammatory disease and including those with severe exacerbations.

Alpha-1 antitrypsin deficiency and recombinant protein sources with focus on plant sources: Updates, challenges and perspectives.

Alpha-1 antitrypsin deficiency (A1ATD) is an autosomal recessive disease characterized by low plasma levels of A1AT, a serine protease inhibitor representing the most abundant circulating antiprotease normally present at plasma levels of 1-2 g/L. The dominant clinical manifestations include predispositions to early onset emphysema due to protease/antiprotease imbalance in distal lung parenchyma and liver disease largely due to unsecreted polymerized accumulations of misfolded mutant A1AT within the endoplasmic reticulum of hepatocytes. Since 1987, the only FDA licensed specific therapy for the emphysema component has been infusions of A1AT purified from pooled human plasma at the 2020 cost of up to US $200,000/year with the risk of intermittent shortages. In the past three decades various, potentially less expensive, recombinant forms of human A1AT have reached early stages of development, one of which is just reaching the stage of human clinical trials. The focus of this review is to update strategies for the treatment of the pulmonary component of A1ATD with some focus on perspectives for therapeutic production and regulatory approval of a recombinant product from plants. We review other competitive technologies for treating the lung disease manifestations of A1ATD, highlight strategies for the generation of data potentially helpful for securing FDA Investigational New Drug (IND) approval and present challenges in the selection of clinical trial strategies required for FDA licensing of a New Drug Approval (NDA) for this disease.

Modulation of ozone-sensitive genes in alpha-tocopherol transfer protein null mice.

Alpha-tocopherol transfer protein (ATTP) null mice (ATTP-/-) have a systemic alpha-tocopherol (AT) deficiency, with their lung AT levels being < 10% of those in AT-replete ATTP(+/+) mice when fed a standard rodent chow diet. ATTP(+/+) and ATTP(-/-) mice (4 wk old male mice, n = 16 per group) were fed a standard diet (35 IU AT/kg diet) for 8-12 wk, exposed 6 h/day for 3 days to either to O(3) (0.5 ppm) or filtered air, then sacrificed. No significant differences in plasma or lung AT concentrations were observed in response to this level of O(3) exposure. Lung genomic responses of the lungs to O(3) were determined using Affymetrix 430A 2.0 arrays containing over 22,600 probe sets representing 14,000 well-characterized mouse genes. As compared with filtered air exposure, O(3) exposure resulted in 99 genes being differentially expressed in ATTP(-/-) mice, as compared to 52 differentially expressed genes in ATTP(+/+) mice. The data revealed an O(3)-induced upregulation of genes related to cell proliferation/DNA repair and inflammatory-immune responses in both ATTP(+/+) and ATTP(-/-) mice, with the expression of 22 genes being common to both, whereas 30 and 77 genes were unique to ATTP(+/+) and ATTP(-/-) mice, respectively. The expressions of O(3) sensitive genes-Timp1, Areg, Birc5 and Tnc-were seen to be further modulated by AT status. The present study reveals AT modulation of adaptive response of lung genome to O(3) exposure.

The use of Raman microscopy to determine and localize vitamin E in biological samples.

Alpha-tocopherol (aT), the predominant form of vitamin E in mammals, is thought to prevent oxidation of polyunsaturated fatty acids. In the lung, aT is perceived to be accumulated in alveolar type II cells and secreted together with surfactant into the epithelial lining fluid. Conventionally, determination of aT and related compounds requires extraction with organic solvents. This study describes a new method to determine and image the distribution of aT and related compounds within cells and tissue sections using the light-scattering technique of Raman microscopy to enable high spatial as well as spectral resolution. This study compared the nondestructive analysis by Raman microscopy of vitamin E, in particular aT, in biological samples with data obtained using conventional HPLC analysis. Raman spectra were acquired at spatial resolutions of 2-0.8 microm. Multivariate analysis techniques were used for analyses and construction of corresponding maps showing the distribution of aT, alpha-tocopherol quinone (aTQ), and other constituents (hemes, proteins, DNA, and surfactant lipids). A combination of images enabled identification of colocalized constituents (heme/aTQ and aT/surfactant lipids). Our data demonstrate the ability of Raman microscopy to discriminate between different tocopherols and oxidation products in biological specimens without sample destruction. By enabling the visualization of lipid-protein interactions, Raman microscopy offers a novel method of investigating biological characterization of lipid-soluble compounds, including those that may be embedded in biological membranes such as aT.

Mice lacking alpha-tocopherol transfer protein gene have severe alpha-tocopherol deficiency in multiple regions of the central nervous system.

Ataxia with vitamin E deficiency is caused by mutations in alpha-tocopherol transfer protein (alpha-TTP) gene and it can be experimentally generated in mice by alpha-TTP gene inactivation (alpha-TTP-KO). This study compared alpha-tocopherol (alpha-T) concentrations of five brain regions and of four peripheral organs from 5 months old, male and female, wild-type (WT) and alpha-TTP-KO mice. All brain regions of female WT mice contained significantly higher alpha-T than those from WT males. alpha-T concentration in the cerebellum was significantly lower than that in other brain regions of WT mice. These sex and regional differences in brain alpha-T concentrations do not appear to be determined by alpha-TTP expression which was undetectable in all brain regions. All the brain regions of alpha-TTP-KO mice were severely depleted in alpha-T. The concentration of another endogenous antioxidant, total glutathione, was unaffected by gender but was decreased slightly but significantly in most brain regions of alpha-TTP-KO mice. The results show that both gender and the hepatic alpha-TTP, but not brain alpha-TTP gene expression are important in determining alpha-T concentrations within the brain. Interestingly, functional abnormality (ataxia) develops only very late in alpha-TTP-KO mice in spite of the severe alpha-tocopherol deficiency in the brain starting at an early age.

An oxidation-resistant, recombinant alpha-1 antitrypsin produced in Nicotiana benthamiana.

Proteases and reactive oxygen species (ROS) have long been implicated in playing key roles in host tissue injury at sites of inflammation dominated by macrophage activations and/or neutrophil infiltrations. Imbalances between proteases/antiproteases and ROS/antioxidants are recognized to contribute to amplification of inflammatory-based host tissue injury. This has been especially well-documented in such respiratory tract diseases as chronic obstructive pulmonary disease, cystic fibrosis, and acute respiratory distress syndrome. Inflammation-related protease/ROS disequilibria are further confounded by recognition that proteases can increase ROS by several different mechanisms and that ROS can inactivate proteases. The major human antiprotease, alpha-1 antitrypsin (AAT), is dramatically inactivated by ROS. AAT deficiency is the most prevalent genetic predisposing factor leading to emphysema, a condition treated by replacement infusions of plasma-derived AAT (hAAT) at a cost of up to $200,000 per year per patient. An updated method for production of a plant-made recombinant AAT (prAAT) engineered for enhanced oxidation resistance compared to hAAT is presented. Plant-made recombinant AAT shows comparable antiprotease activity to hAAT, and retains full activity under oxidative conditions that would deactivate hAAT. Additionally, we show that prAAT has similar effectiveness in preventing neutrophil elastase-induced cell death in an in vitro human bronchial epithelial cell culture model. We conclude that prAAT is potentially a "biobetter" AAT product that could be made available to individuals with a wide spectrum of inflammatory disorders characterized by overly aggressive neutrophilic infiltrations.

Quantitative assessment of cyanide in cystic fibrosis sputum and its oxidative catabolism by hypochlorous acid.

RATIONALE: Cystic fibrosis (CF) patients are known to produce cyanide (CN-) although challenges exist in determinations of total levels, the precise bioactive levels, and specificity of its production by CF microflora, especially P. aeruginosa. Our objective was to measure total CN- levels in CF sputa by a simple and novel technique in P. aeruginosa positive and negative adult patients, to review respiratory tract (RT) mechanisms for the production and degradation of CN-, and to interrogate sputa for post-translational protein modification by CN- metabolites. METHODS: Sputa CN- concentrations were determined by using a commercially available CN- electrode, measuring levels before and after addition of cobinamide, a compound with extremely high affinity for CN-. Detection of protein carbamoylation was measured by Western blot. MEASUREMENTS AND MAIN RESULTS: The commercial CN- electrode was found to overestimate CN- levels in CF sputum in a highly variable manner; cobinamide addition rectified this analytical issue. Although P. aeruginosa positive patients tended to have higher total CN- values, no significant differences in CN- levels were found between positive and negative sputa. The inflammatory oxidant hypochlorous acid (HOCl) was shown to rapidly decompose CN-, forming cyanogen chloride (CNCl) and the carbamoylating species cyanate (NCO-). Carbamoylated proteins were found in CF sputa, analogous to reported findings in asthma. CONCLUSIONS: Our studies indicate that CN- is a transient species in the inflamed CF airway due to multiple biosynthetic and metabolic processes. Stable metabolites of CN-, such as cyanate, or carbamoylated proteins, may be suitable biomarkers of overall CN- production in CF airways.

Genome-wide screening of alpha-tocopherol sensitive genes in heart tissue from alpha-tocopherol transfer protein null mice (ATTP(-/-)).

Alpha-tocopherol transfer protein (ATTP) null mice (ATTP(-/-)) have a systemic deficiency of alpha-tocopherol (AT). The heart AT levels of ATTP(-/-) are <10% of those in ATTP(+/+) mice. The genomic responses of heart to AT deficiency were determined in 3 months old male ATTP(-/-) mice and compared with their ATTP(+/+) littermate controls using Affymetrix 430A 2.0 high density oligonucleotide arrays. Differential analysis of approximately 13000 genes identified repression of genes related to immune system and activation of genes related to lipid metabolism and inflammation with no significant change in the expression of classical antioxidant genes (catalase, superoxide dismutase, glutathione peroxidase) in ATTP(-/-) as compared to ATTP(+/+) mice. The present data identifies novel classes of AT sensitive genes in heart tissue.

Antioxidants in cystic fibrosis. Conclusions from the CF antioxidant workshop, Bethesda, Maryland, November 11-12, 2003.

Although great strides are being made in the care of individuals with cystic fibrosis (CF), this condition remains the most common fatal hereditary disease in North America. Numerous links exist between progression of CF lung disease and oxidative stress. The defect in CF is the loss of function of the transmembrane conductance regulator (CFTR) protein; recent evidence that CFTR expression and function are modulated by oxidative stress suggests that the loss may result in a poor adaptive response to oxidants. Pancreatic insufficiency in CF also increases susceptibility to deficiencies in lipophilic antioxidants. Finally the airway infection and inflammatory processes in the CF lung are potential sources of oxidants that can affect normal airway physiology and contribute to the mechanisms causing characteristic changes associated with bronchiectasis and loss of lung function. These multiple abnormalities in the oxidant/antioxidant balance raise several possibilities for therapeutic interventions that must be carefully assessed.

Genome wide responses of murine lungs to dietary alpha-tocopherol.

Alpha-tocopherol (alpha-T) may affect biological processes by modulating mRNA concentrations. This study screened the responses of approximately 15,000 lung mRNAs to dietary alpha-T in mice. The lung was chosen as the target organ because it is subjected to cyclical variations in oxidant and inflammatory stressors and alpha-T has been implicated in their modulations. The analysis identified approximately 400 mRNAs sensitive to alpha-T status of lungs determined by dietary alpha-T. The female lung transcriptome appears to be more sensitive to the alpha-T status than that of the male lungs. Here, we focus on the induction of 13 cytoskeleton genes by dietary alpha-T because they were similarly induced in the male and the female lungs. Their inductions were confirmed by quantitative-real-time-polymerase chain reaction (qRT-PCR). Immunohistochemical analyses of three of the encoded proteins suggest that they are expressed in lung vasculature and alveolar regions. The data suggest that the lung alpha-T status may modulate cytoarchitecture of lungs.

Evidence of health impacts of sulfate-and nitrate-containing particles in ambient air.

Ambient particulate matter (PM) is a complex mixture of inorganic and organic compounds. The U.S. Environmental Protection Agency (EPA) regulates PM as a criteria pollutant and promulgates National Ambient Air Quality Standards for it. The PM indicator is based on mass concentration, unspecified as to chemical composition, for specific size fractions. The numerical standards are based on epidemiologic evidence of associations between the various size-related particle mass concentrations as indicators and excess mortality and cardiorespiratory health effects as endpoints. The U.S. National Research Council has stated that more research is needed to differentiate the apparent health effects associated with different particle chemical constituents. Sulfate and nitrate constitute a significant portion of the particle mass in the atmosphere, but are accompanied by similar amounts of carbonaceous material, along with low concentrations of various species, including bioactive organic compounds and redox cycling metals. Extensive animal and human toxicology data show no significant effects for particles consisting only of sulfate and nitrate compounds at levels in excess of ambient air concentrations. A few epidemiologic studies, including both short-term time-series studies and long-term cohort studies, have included the sulfate content of PM as a specific variable in health effect analyses. There are much less data for nitrate. The results from the epidemiologic studies with PM sulfate are inconsistent. A detailed analysis of the time-series epidemiological studies shows that PM sulfate has a weaker "risk factor" than PM2.5 for health effects. Since sulfate is correlated with PM2.5, this result is inconsistent with sulfate having a strong health influence. However, there are many limitations with these types of studies that warrant caution for any comparison between a chemical component and mass concentration. In total, the epidemiologic and toxicologic evidence provide little or no support for a causal association of PM sulfate and health risk at ambient concentrations. For nitrate-containing PM, virtually no epidemiological data exist. Limited toxicological evidence does not support a causal association between particulate nitrate compounds and excess health risks. There are some possible indirect processes through which sulfate and nitrate in PM may affect health-related endpoints, including interactions with certain metal species and a linkage with production of secondary organic matter. There is insufficient evidence to include or exclude these processes as being potentially important to PM-associated health risk.

Modulation of cutaneous wound healing by ozone: differences between young and aged mice.

Cutaneous tissues are frequently exposed to prooxidative environments, including UV radiation and air pollutants. Among the latter, ozone (O(3)) is of particular concern because of its high and dominating presence in photochemical smog. It is well known that O(3) depletes small molecular weight antioxidants, oxidizes proteins, induces lipid peroxidation and activates cellular responses in various tissues. Using an in vivo model (SKH-1 hairless mice), the interaction between O(3) exposure (0.5ppmx6h/day) and age was examined in relation to cutaneous wound healing. Compared to younger (8 weeks) mice, older (18 months) mice exposed to O(3) (day 0 to day 9 after wounding) exhibited delayed wound closure, increased lipid peroxidation (measured as 4-HNE protein adducts) and protein oxidation (measured as carbonyls concentration) and decreased levels of P-IkappaBalpha and TGFbeta protein. These findings support the hypothesis that oxidant pollutant exposure and age interact so as to disrupt normal wound healing processes.

Regulation of the alpha-tocopherol transfer protein in mice: lack of response to dietary vitamin E or oxidative stress.

The alpha-tocopherol transfer protein (TTP) plays an important role in the regulation of plasma alpha-tocopherol concentrations. We hypothesized that hepatic TTP levels would be modulated by dietary vitamin E supplementation and/or by oxidative stress. Mice were fed either a High E (1150 mg RRR-alpha-tocopheryl acetate/kg diet) or a Low E (11.5 mg/kg diet) diet for 2 wk. High E increased plasma and liver alpha-tocopherol concentrations approximately 8- and 40-fold, respectively, compared with Low E-fed mice, whereas hepatic TTP increased approximately 20%. Hepatic TTP concentrations were unaffected by fasting (24 h) in mice fed either diet. To induce oxidative stress, chow-fed mice were exposed for 3 d to environmental tobacco smoke (ETS) for 6 h/d (total suspended particulate, 57.4 +/- 1.8 mg/m3). ETS exposure, while resulting in pulmonary and systemic oxidative stress, had no effect on hepatic alpha-tocopherol concentrations or hepatic TTP. Overall, changes in hepatic TTP concentrations were minimal in response to dietary vitamin E levels or ETS-related oxidative stress. Thus, hepatic TTP concentrations may be at sufficient levels such that they are unaffected by either modulations of dietary vitamin E or by the conditions of environmentally related oxidative stress used in the present studies.

The Diagnosis and Management of Alpha-1 Antitrypsin Deficiency in the Adult.

Background: The diagnosis and clinical management of adults with alpha-1 antitrypsin deficiency (AATD) have been the subject of ongoing debate, ever since the publication of the first American Thoracic Society guideline statement in 1989.1 In 2003, the "American Thoracic Society (ATS)/European Respiratory Society (ERS) Statement: Standards for the Diagnosis and Management of Individuals with Alpha-1 Antitrypsin Deficiency" made a series of evidence-based recommendations, including a strong recommendation for broad-based diagnostic testing of all symptomatic adults with chronic obstructive pulmonary disease (COPD).2 Even so, AATD remains widely under-recognized. To update the 2003 systematic review and clinical guidance, the Alpha-1 Foundation sponsored a committee of experts to examine all relevant, recent literature in order to provide concise recommendations for the diagnosis and management of individuals with AATD. Purpose: To provide recommendations for: (1) the performance and interpretation of diagnostic testing for AATD, and (2) the current management of adults with AATD and its associated medical conditions. Methods: A systematic review addressing the most pressing questions asked by clinicians (clinician-centric) was performed to identify citations related to AATD that were published since the 2003 comprehensive review, specifically evaluating publications between January 2002 and December 2014. Important, more recent publications were solicited from the writing committee members as well. The combined comprehensive literature reviews of the 2003 document and this current review comprise the evidence upon which the committee's conclusions and recommendations are based. Results: Recommendations for the diagnosis and management of AATD were formulated by the committee. Conclusions: The major recommendations continue to endorse and reinforce the importance of testing for AATD in all adults with symptomatic fixed airflow obstruction, whether clinically labeled as COPD or asthma. Individuals with unexplained bronchiectasis or liver disease also should be tested. Family testing of first-degree relatives is currently the most efficient detection technique. In general, individuals with AATD and emphysema, bronchiectasis, and/or liver disease should be managed according to usual guidelines for these clinical conditions. In countries where intravenous augmentation therapy with purified pooled human plasma-derived alpha-1 antitrypsin is available, recent evidence now provides strong support for its use in appropriate individuals with lung disease due to AATD.

Studies in humans using deuterium-labeled alpha- and gamma-tocopherols demonstrate faster plasma gamma-tocopherol disappearance and greater gamma-metabolite production.

We hypothesized that human plasma alpha- and gamma-tocopherol concentrations reflect differences in their kinetics, especially influenced by gamma-tocopherol metabolism. Vitamin E kinetics were evaluated in humans (n=14) using approximately 50 mg each of an equimolar ratio of d6-alpha- and d2-gamma-tocopheryl acetates administered orally. Mass spectrometry was used to measure deuterated plasma tocopherols, as well as plasma and urinary vitamin E metabolites, alpha- and gamma-carboxyethylhydroxychromans (CEHCs). Plasma d2-gamma-tocopherol fractional disappearance rates (FDR; 1.39+/-0.44 pools/day, mean+/-SD) were more than three times greater than those of d6-alpha-tocopherol (0.33+/-0.11, p<0.001). The d2-gamma-tocopherol half-life was 13+/-4 h compared with 57+/-19 for d6-alpha-tocopherol. Whereas neither plasma nor urinary d6-alpha-CEHC was detectable (limit of detection 1 nmol/L), gamma-CEHC (labeled plus unlabeled) increased from 129+/-20 to 258+/-40 nmol/L by 12 h and returned to baseline by 48 h; at 12 h d2-gamma-CEHC represented 54+/-4% of plasma gamma-CEHC. Women compared with men had a greater d2-gamma-tocopherol FDR (p<0.004) and a greater maximal plasma d2-gamma-CEHC concentration (p<0.02) and CEHC FDR (p<0.007), as well as excreting four times as much d2-gamma-CEHC (p<0.04) in urine. Thus, gamma-tocopherol is rapidly metabolized to gamma-CEHC, and to a greater degree in women than in men, whereas alpha-tocopherol is maintained in the plasma and little is metabolized to alpha-CEHC.

Alpha-tocopherol modulates Cyp3a expression, increases gamma-CEHC production, and limits tissue gamma-tocopherol accumulation in mice fed high gamma-tocopherol diets.

Although all forms of vitamin E are absorbed, the liver preferentially secretes alpha-, but not gamma-tocopherol, into plasma. Liver alpha-tocopherol secretion is under the control of the alpha-tocopherol transfer protein (TTP). Therefore, to assess gamma-tocopherol bioactivities Ttpa-/-, +/- and +/+ mice were fed for 5 weeks diets containing gamma-tocopherol 550 (gamma-T550), gamma-tocopherol 60 (gamma-T60) mg/kg that also contained trace amounts of alpha-tocopherol, a vitamin E-deficient diet, or a control diet. Plasma and tissues from mice fed gamma-T550 diets were found to contain similar gamma- and alpha-tocopherol concentrations despite the high dietary gamma-tocopherol content; nervous tissues contained almost no gamma-tocopherol. Liver vitamin E metabolites (carboxyethyl hydroxychromans, CEHCs) were also measured. In mice with widely ranging liver alpha- (from 0.7 to 16 nmol/g) and gamma-tocopherol concentrations (0 to 13 nmol/g), hepatic alpha-CEHC was undetectable, but gamma-CEHC concentrations (0.1 to 0.8 nmol/g) were correlated with both alpha- and gamma-tocopherol concentrations (P < 0.004). Hepatic cytochrome P450s (CYPs) involved in vitamin E metabolism, Cyp4f and Cyp3a, were also measured. There were no variations in Cyp4f protein expression as related to diet or mouse genotype. However, Cyp3a was correlated (P < 0.0001) with liver alpha-, but not gamma-tocopherol concentrations. These data support the hypothesis that alpha-tocopherol modulates xenobiotic metabolism by increasing Cyp3a expression, gamma-CEHC formation, and the excretion of both gamma-tocopherol and gamma-CEHC.