query: Omnivores produce less trimethylamine N-oxide from dietary I-carnitine than vegetarians.

positive: Intestinal microbiota metabolism of L-carnitine, a nutrient in red meat, promotes atherosclerosis Intestinal microbiota metabolism of choline and phosphatidylcholine produces trimethylamine (TMA), which is further metabolized to a proatherogenic species, trimethylamine-N-oxide (TMAO). We demonstrate here that metabolism by intestinal microbiota of dietary L-carnitine, a trimethylamine abundant in red meat, also produces TMAO and accelerates atherosclerosis in mice. Omnivorous human subjects produced more TMAO than did vegans or vegetarians following ingestion of L-carnitine through a microbiota-dependent mechanism. The presence of specific bacterial taxa in human feces was associated with both plasma TMAO concentration and dietary status. Plasma L-carnitine levels in subjects undergoing cardiac evaluation (n = 2,595) predicted increased risks for both prevalent cardiovascular disease (CVD) and incident major adverse cardiac events (myocardial infarction, stroke or death), but only among subjects with concurrently high TMAO levels. Chronic dietary L-carnitine supplementation in mice altered cecal microbial composition, markedly enhanced synthesis of TMA and TMAO, and increased atherosclerosis, but this did not occur if intestinal microbiota was concurrently suppressed. In mice with an intact intestinal microbiota, dietary supplementation with TMAO or either carnitine or choline reduced in vivo reverse cholesterol transport. Intestinal microbiota may thus contribute to the well-established link between high levels of red meat consumption and CVD risk.

negative: Transmission of atherosclerosis susceptibility with gut microbial transplantation. Recent studies indicate both clinical and mechanistic links between atherosclerotic heart disease and intestinal microbial metabolism of certain dietary nutrients producing trimethylamine N-oxide (TMAO). Here we test the hypothesis that gut microbial transplantation can transmit choline diet-induced TMAO production and atherosclerosis susceptibility. First, a strong association was noted between atherosclerotic plaque and plasma TMAO levels in a mouse diversity panel (n = 22 strains, r = 0.38; p = 0.0001). An atherosclerosis-prone and high TMAO-producing strain, C57BL/6J, and an atherosclerosis-resistant and low TMAO-producing strain, NZW/LacJ, were selected as donors for cecal microbial transplantation into apolipoprotein e null mice in which resident intestinal microbes were first suppressed with antibiotics. Trimethylamine (TMA) and TMAO levels were initially higher in recipients on choline diet that received cecal microbes from C57BL/6J inbred mice; however, durability of choline diet-dependent differences in TMA/TMAO levels was not maintained to the end of the study. Mice receiving C57BL/6J cecal microbes demonstrated choline diet-dependent enhancement in atherosclerotic plaque burden as compared with recipients of NZW/LacJ microbes. Microbial DNA analyses in feces and cecum revealed transplantation of donor microbial community features into recipients with differences in taxa proportions between donor strains that were transmissible to recipients and that tended to show coincident proportions with TMAO levels. Proportions of specific taxa were also identified that correlated with plasma TMAO levels in donors and recipients and with atherosclerotic lesion area in recipients. Atherosclerosis susceptibility may be transmitted via transplantation of gut microbiota. Gut microbes may thus represent a novel therapeutic target for modulating atherosclerosis susceptibility.

negative: Vegetarian dietary patterns and mortality in Adventist Health Study 2. IMPORTANCE Some evidence suggests vegetarian dietary patterns may be associated with reduced mortality, but the relationship is not well established. OBJECTIVE To evaluate the association between vegetarian dietary patterns and mortality. DESIGN Prospective cohort study; mortality analysis by Cox proportional hazards regression, controlling for important demographic and lifestyle confounders. SETTING Adventist Health Study 2 (AHS-2), a large North American cohort. PARTICIPANTS A total of 96,469 Seventh-day Adventist men and women recruited between 2002 and 2007, from which an analytic sample of 73,308 participants remained after exclusions. EXPOSURES Diet was assessed at baseline by a quantitative food frequency questionnaire and categorized into 5 dietary patterns: nonvegetarian, semi-vegetarian, pesco-vegetarian, lacto-ovo-vegetarian, and vegan. MAIN OUTCOME AND MEASURE The relationship between vegetarian dietary patterns and all-cause and cause-specific mortality; deaths through 2009 were identified from the National Death Index. RESULTS There were 2570 deaths among 73,308 participants during a mean follow-up time of 5.79 years. The mortality rate was 6.05 (95% CI, 5.82-6.29) deaths per 1000 person-years. The adjusted hazard ratio (HR) for all-cause mortality in all vegetarians combined vs nonvegetarians was 0.88 (95% CI, 0.80-0.97). The adjusted HR for all-cause mortality in vegans was 0.85 (95% CI, 0.73-1.01); in lacto-ovo-vegetarians, 0.91 (95% CI, 0.82-1.00); in pesco-vegetarians, 0.81 (95% CI, 0.69-0.94); and in semi-vegetarians, 0.92 (95% CI, 0.75-1.13) compared with nonvegetarians. Significant associations with vegetarian diets were detected for cardiovascular mortality, noncardiovascular noncancer mortality, renal mortality, and endocrine mortality. Associations in men were larger and more often significant than were those in women. CONCLUSIONS AND RELEVANCE Vegetarian diets are associated with lower all-cause mortality and with some reductions in cause-specific mortality. Results appeared to be more robust in males. These favorable associations should be considered carefully by those offering dietary guidance.

negative: Circulating trimethylamine N‐oxide and the risk of cardiovascular diseases: a systematic review and meta‐analysis of 11 prospective cohort studies Circulating trimethylamine N-oxide (TMAO), a canonical metabolite from gut flora, has been related to the risk of cardiovascular disorders. However, the association between circulating TMAO and the risk of cardiovascular events has not been quantitatively evaluated. We performed a systematic review and meta-analysis of all available cohort studies regarding the association between baseline circulating TMAO and subsequent cardiovascular events. Embase and PubMed databases were searched for relevant cohort studies. The overall hazard ratios for the developing of cardiovascular events (CVEs) and mortality were extracted. Heterogeneity among the included studies was evaluated with Cochran's Q Test and I2 statistics. A random-effect model or a fixed-effect model was applied depending on the heterogeneity. Subgroup analysis and meta-regression were used to evaluate the source of heterogeneity. Among the 11 eligible studies, three reported both CVE and mortality outcome, one reported only CVEs and the other seven provided mortality data only. Higher circulating TMAO was associated with a 23% higher risk of CVEs (HR = 1.23, 95% CI: 1.07-1.42, I2 = 31.4%) and a 55% higher risk of all-cause mortality (HR = 1.55, 95% CI: 1.19-2.02, I2 = 80.8%). Notably, the latter association may be blunted by potential publication bias, although sensitivity analysis by omitting one study at a time did not significantly change the results. Further subgroup analysis and meta-regression did not support that the location of the study, follow-up duration, publication year, population characteristics or the samples of TMAO affect the results significantly. Higher circulating TMAO may independently predict the risk of subsequent cardiovascular events and mortality.

negative: Risk of hospitalization or death from ischemic heart disease among British vegetarians and nonvegetarians: results from the EPIC-Oxford cohort study. BACKGROUND Few previous prospective studies have examined differences in incident ischemic heart disease (IHD) risk between vegetarians and nonvegetarians. OBJECTIVE The objective was to examine the association of a vegetarian diet with risk of incident (nonfatal and fatal) IHD. DESIGN A total of 44,561 men and women living in England and Scotland who were enrolled in the European Prospective Investigation into Cancer and Nutrition (EPIC)-Oxford study, of whom 34% consumed a vegetarian diet at baseline, were part of the analysis. Incident cases of IHD were identified through linkage with hospital records and death certificates. Serum lipids and blood pressure measurements were available for 1519 non cases, who were matched to IHD cases by sex and age. IHD risk by vegetarian status was estimated by using multivariate Cox proportional hazards models. RESULTS After an average follow-up of 11.6 y, there were 1235 IHD cases (1066 hospital admissions and 169 deaths). Compared with nonvegetarians, vegetarians had a lower mean BMI [in kg/m(2); -1.2 (95% CI: -1.3, -1.1)], non-HDL-cholesterol concentration [-0.45 (95% CI: -0.60, -0.30) mmol/L], and systolic blood pressure [-3.3 (95% CI: -5.9, -0.7) mm Hg]. Vegetarians had a 32% lower risk (HR: 0.68; 95% CI: 0.58, 0.81) of IHD than did nonvegetarians, which was only slightly attenuated after adjustment for BMI and did not differ materially by sex, age, BMI, smoking, or the presence of IHD risk factors. CONCLUSION Consuming a vegetarian diet was associated with lower IHD risk, a finding that is probably mediated by differences in non-HDL cholesterol, and systolic blood pressure.

negative: Effect of lipid restriction on mitochondrial free radical production and oxidative DNA damage. Many studies have shown that caloric restriction (40%) decreases mitochondrial reactive oxygen species (ROS) generation in rodents. Moreover, we have recently found that 7 weeks of 40% protein restriction without strong caloric restriction also decreases ROS production in rat liver. This is interesting since it has been reported that protein restriction can also extend longevity in rodents. In the present study we have investigated the possible role of dietary lipids in the effects of caloric restriction on mitochondrial oxidative stress. Using semipurified diets, the ingestion of lipids in male Wistar rats was decreased by 40% below controls, while the other dietary components were ingested at exactly the same level as in animals fed ad libitum. After 7 weeks of treatment the liver mitochondria of lipid-restricted animals showed significant increases in oxygen consumption with complex I-linked substrates (pyruvate/malate and glutamate/malate). Neither mitochondrial H(2)O(2) production nor oxidative damage to mitochondrial or nuclear DNA was modified in lipid-restricted animals. Oxidative damage to mitochondrial DNA was one order of magnitude higher than that of nuclear DNA in both dietary groups. These results deny a role for lipids and reinforce the possible role of dietary proteins as being responsible for the decrease in mitochondrial ROS production and DNA damage in caloric restriction.

negative: Vegetarian vs. conventional diabetic diet – A 1-year follow-up Abstract Objective Our previous 6-month, randomized study demonstrated the beneficial effect of a vegetarian (V) compared to a conventional diet (C) with similar caloric restriction on cardiovascular risk factors for patients with type 2 diabetes (T2D), namely increased insulin sensitivity, reduced body weight, reduced volume of visceral and subcutaneous fat, decreased LDL-cholesterol and improved oxidative stress markers and chosen adipokines. We conducted post-trial monitoring to determine whether the improved outcomes persisted 1 year after the end of the study. Methods 62 subjects with T2D who completed the study were asked to come for a 1-year follow-up to measure weight, waist circumference, HbA1c and blood lipids. No attempts were made to maintain their previously assigned diets. Results 44 patients (71%) attended the post-trial monitoring. Hypoglycemic agents were increased by 14% in V and by 26% in C; insulin therapy was introduced in 5% in V and in 13% in C one year after the end of the intervention. Neither weight nor waist circumference changed significantly in either group. HbA1c increased ( p  ≤ 0.05) similarly in both groups (+0.49 ± 1.04% in V vs. +0.42 ± 0.8% in C). Blood lipids did not change in either group. Conclusion One year after the end of the intervention, the positive effects of a vegetarian diet on cardiovascular risk factors compared to a conventional diet were partially maintained.

negative: Metabolic modifiers as performance-enhancing technologies for livestock production With the current and predicted increase in world population, growing global demand and consumption of food will result in increasing competition for land, water, and energy. In turn, this will severely affect our ability to produce food as will the urgent requirement to reduce the impact of food systems on the environment (Godfray et al., 2010). Globalization has boosted trade in livestock inputs and products and resulted in industry growth, and concurrently livestock production has undergone a complex process of technical and geographical change (Gerber et al., 2007). This has resulted in a challenge to livestock producers; growing demand for their produce with a dwindling supply of resources, with the only solution being a significant increase in efficiency. Also, as incomes increase in the burgeoning economies, so does the demand for high-quality animal proteins such as meat, milk, and eggs; thus the Food and Agriculture Organization of the United Nations suggests that food requirements will increase by 70% by 2050 (Anonymous, 2009). The recent success in developed and emerging technologies suggests that the animal industries are well placed to prosper through these new challenges. To ensure these technologies—such as metabolic modifiers including somatotropin, immunization against gonadotropin-releasing factor, and orally active dietary additives like ractopamine, zilpaterol, cysteamine, chromium, betaine, and dietary neuroleptics—can be effectively utilized throughout the animal industries, further emphasis is required on their acceptance and development. As a result of these technological advancements, producers have benefited because of improved production efficiencies while meat packers have improved processing efficiencies because of increased lean meat yield. Ostensibly, the consumer has also benefited because meat is leaner and less expensive to purchase. However, there have been some concerns that the focus on increasing production efficiency and lean meat yield has been to the detriment of meat quality (Dunshea et al., 2005). It is also interesting that at a time of apparently greater need for these technologies, there are external influences such as market differentiation and trade barriers as well as consumer resistance that challenge the use of technologies. A tenet of this article is that if we are to meet the increased global demand for animal protein, then we must continue to develop and adopt technologies to improve livestock efficiency, but we must be cognizant of the potential barriers affecting acceptance.

negative: tBid induces alterations of mitochondrial fatty acid oxidation flux by malonyl-CoA-independent inhibition of carnitine palmitoyltransferase-1 Recent studies suggest a close relationship between cell metabolism and apoptosis. We have evaluated changes in lipid metabolism on permeabilized hepatocytes treated with truncated Bid (tBid) in the presence of caspase inhibitors and exogenous cytochrome c. The measurement of β-oxidation flux by labeled palmitate demonstrates that tBid inhibits β-oxidation, thereby resulting in the accumulation of palmitoyl-coenzyme A (CoA) and depletion of acetyl-carnitine and acylcarnitines, which is pathognomonic for inhibition of carnitine palmitoyltransferase-1 (CPT-1). We also show that tBid decreases CPT-1 activity by a mechanism independent of both malonyl-CoA, the key inhibitory molecule of CPT-1, and Bak and/or Bax, but dependent on cardiolipin decrease. Overexpression of Bcl-2, which is able to interact with CPT-1, counteracts the effects exerted by tBid on β-oxidation. The unexpected role of tBid in the regulation of lipid β-oxidation suggests a model in which tBid-induced metabolic decline leads to the accumulation of toxic lipid metabolites such as palmitoyl-CoA, which might become participants in the apoptotic pathway.

negative: Dietary status of trained female cyclists. Dietary status was evaluated in eight highly trained female cyclists. Each cyclist kept a 3-day weighed food record. Diets were analyzed for nutrient content using a computerized software package. Blood was also obtained and evaluated for hemoglobin, hematocrit, and albumin. For an athletic group, the cyclists' diets were found to be low in energy (85% RDA) and carbohydrate (4.4 gm/kg body weight per day). Mean daily dietary intakes were well below the RDAs for folacin (76% RDA), magnesium (81%), iron (59%), and zinc (48%). In addition, more than one-third of the cyclists failed to consume 67% of the RDA for the following micronutrients: pyridoxine, folacin, cobalamin, vitamin E, magnesium, iron, and zinc. Hemoglobin (135 gm/L), hematocrit (0.39), and albumin (45 gm/L) values were all normal, although most hemoglobin values were in the lower 50% of normal range. Foods such as meats, poultry, fish, beans, peas, and nuts were low or absent from the diets of most athletes. Dietary quality in this group of female cyclists could have been greatly improved with the addition of more of those foods. These athletes could benefit from nutrition education and diet counseling.

negative: Microbiome-driven allergic lung inflammation is ameliorated by short-chain fatty acids The mammalian gastrointestinal tract harbors a microbial community with metabolic activity critical for host health, including metabolites that can modulate effector functions of immune cells. Mice treated with vancomycin have an altered microbiome and metabolite profile, exhibit exacerbated T helper type 2 cell (Th2) responses, and are more susceptible to allergic lung inflammation. Here we show that dietary supplementation with short-chain fatty acids (SCFAs) ameliorates this enhanced asthma susceptibility by modulating the activity of T cells and dendritic cells (DCs). Dysbiotic mice treated with SCFAs have fewer interleukin-4 (IL4)-producing CD4+ T cells and decreased levels of circulating immunoglobulin E (IgE). In addition, DCs exposed to SCFAs activate T cells less robustly, are less motile in response to CCL19 in vitro, and exhibit a dampened ability to transport inhaled allergens to lung draining nodes. Our data thus demonstrate that gut dysbiosis can exacerbate allergic lung inflammation through both T cell- and DC-dependent mechanisms that are inhibited by SCFAs.

negative: Arginase I: a limiting factor for nitric oxide and polyamine synthesis by activated macrophages? Because arginase hydrolyzes arginine to produce ornithine and urea, it has the potential to regulate nitric oxide (NO) and polyamine synthesis. We tested whether expression of the cytosolic isoform of arginase (arginase I) was limiting for NO or polyamine production by activated RAW 264.7 macrophage cells. RAW 264.7 cells, stably transfected to overexpress arginase I or beta-galactosidase, were treated with interferon-gamma to induce type 2 NO synthase or with lipopolysaccharide or 8-bromo-cAMP (8-BrcAMP) to induce ornithine decarboxylase. Overexpression of arginase I had no effect on NO synthesis. In contrast, cells overexpressing arginase I produced twice as much putrescine after activation than did cells expressing beta-galactosidase. Cells overexpressing arginase I also produced more spermidine after treatment with 8-BrcAMP than did cells expressing beta-galactosidase. Thus endogenous levels of arginase I are limiting for polyamine synthesis, but not for NO synthesis, by activated macrophage cells. This study also demonstrates that it is possible to alter arginase I levels sufficiently to affect polyamine synthesis without affecting induced NO synthesis.

negative: H2S and HS− donor NaHS releases nitric oxide from nitrosothiols, metal nitrosyl complex, brain homogenate and murine L1210 leukaemia cells Nitrosoglutathione [(GSNO), 500 nmol/l] relaxed the norepinephrine precontracted rat aortic rings. The relaxation effect was pronouncedly enhanced by H2S- and HS−-donor NaHS (30 μmol/l) at 7.5 pH but not at 6.3 pH. To study molecular mechanism of this effect, we investigated whether NaHS can release NO from NO donors. Using an electron paramagnetic resonance spectroscopy method of spin trap and by measuring the NO oxidation product, which is nitrite, by the Griess reaction, we report that NaHS released NO from nitrosothiols, namely from GSNO, S-nitroso-N-acetyl-dl-penicillamine (SNAP), from metal nitrosyl complex nitroprusside (SNP) and from rat brain homogenate and murine L1210 leukaemia cells. From the observation that the releasing effect was more pronounced at 8.0 pH than 6.0 pH, we suppose that HS−, rather than H2S, is responsible for the NO-releasing effect. Since in mammals, H2S and HS− are produced endogenously, we assume that their effect to release NO from nitrosothiols and from metal nitrosyl complexes are responsible for some of their biological activities and that this mechanism may be involved in S-nitrosothiol-signalling reactions.

negative: Metabolic effects of isoenergetic nutrient exchange over 24 hours in relation to obesity in women. Twenty-four hour whole body indirect calorimetry has been used to study the effects of feeding, during a sedentary test day, isoenergetic diets which varied in fat (3 or 40 per cent of total energy) and carbohydrate (82 or 45 per cent) content. Three groups of women were studied: lean, obese and 'post-obese' after slimming. Energy expenditure was greater in absolute terms in the obese women. Twenty-four hour energy expenditure was lower by only 3-7 per cent when fasting compared to that when fed to achieve energy balance. There were no large differences in energy expenditure between the two diets or between the groups but the thermogenic effect of the high carbohydrate diet was significantly greater than that of the high fat diet (5.8 vs 3.5 per cent of energy expenditure: P less than 0.01). The post-obese tended to have lower energy expenditure per kg FFM than controls when fasting and when high-fat fed, but this pattern was not shown by the obese. Sleeping energy expenditure was particularly low in the post-obese group when high-fat fed. Dirunal variations in RQ appear to show more marked rise in morning RQ from the nocturnal minimum in the obese and post-obese, which might be evidence for an energy-saving mechanism through greater availability of stored dietary carbohydrate.

negative: Spare respiratory capacity rather than oxidative stress regulates glutamate excitotoxicity after partial respiratory inhibition of mitochondrial complex I with rotenone. Partial inhibition of mitochondrial respiratory complex I by rotenone reproduces aspects of Parkinson's disease in rodents. The hypothesis that rotenone enhancement of neuronal cell death is attributable to oxidative stress was tested in an acute glutamate excitotoxicity model using primary cultures of rat cerebellar granule neurons. As little as 5 nM rotenone increased mitochondrial superoxide (O2*-) levels and potentiated glutamate-induced cytoplasmic Ca2+ deregulation, the first irreversible stage of necrotic cell death. However, the potent cell-permeant O2*- trap manganese tetrakis (N-ethylpyridinium-2yl) porphyrin failed to prevent the effects of the inhibitor. The bioenergetic consequences of rotenone addition were quantified by monitoring cell respiration. Glutamate activation of NMDA receptors used the full respiratory capacity of the in situ mitochondria, and >80% of the glutamate-stimulated respiration was attributable to increased cellular ATP demand. Rotenone at 20 nM inhibited basal and carbonyl cyanide p-trifluoromethoxyphenylhydrazone-stimulated cell respiration and caused respiratory failure in the presence of glutamate. ATP synthase inhibition by oligomycin was also toxic in the presence of glutamate. We conclude that the cell vulnerability in the rotenone model of partial complex I deficiency under these specific conditions is primarily determined by spare respiratory capacity rather than oxidative stress.

negative: Vegetarian diet improves insulin resistance and oxidative stress markers more than conventional diet in subjects with Type 2 diabetes AIMS The aim of this study was to compare the effects of calorie-restricted vegetarian and conventional diabetic diets alone and in combination with exercise on insulin resistance, visceral fat and oxidative stress markers in subjects with Type 2 diabetes. METHODS A 24-week, randomized, open, parallel design was used. Seventy-four patients with Type 2 diabetes were randomly assigned to either the experimental group (n = 37), which received a vegetarian diet, or the control group (n = 37), which received a conventional diabetic diet. Both diets were isocaloric, calorie restricted (-500 kcal/day). All meals during the study were provided. The second 12 weeks of the diet were combined with aerobic exercise. Participants were examined at baseline, 12 weeks and 24 weeks. Primary outcomes were: insulin sensitivity measured by hyperinsulinaemic isoglycaemic clamp; volume of visceral and subcutaneous fat measured by magnetic resonance imaging; and oxidative stress measured by thiobarbituric acid reactive substances. Analyses were by intention to treat. RESULTS Forty-three per cent of participants in the experimental group and 5% of participants in the control group reduced diabetes medication (P < 0.001). Body weight decreased more in the experimental group than in the control group [-6.2 kg (95% CI -6.6 to -5.3) vs. -3.2 kg (95% CI -3.7 to -2.5); interaction group × time P = 0.001]. An increase in insulin sensitivity was significantly greater in the experimental group than in the control group [30% (95% CI 24.5-39) vs. 20% (95% CI 14-25), P = 0.04]. A reduction in both visceral and subcutaneous fat was greater in the experimental group than in the control group (P = 0.007 and P = 0.02, respectively). Plasma adiponectin increased (P = 0.02) and leptin decreased (P = 0.02) in the experimental group, with no change in the control group. Vitamin C, superoxide dismutase and reduced glutathione increased in the experimental group (P = 0.002, P < 0.001 and P = 0.02, respectively). Differences between groups were greater after the addition of exercise training. Changes in insulin sensitivity and enzymatic oxidative stress markers correlated with changes in visceral fat. CONCLUSIONS A calorie-restricted vegetarian diet had greater capacity to improve insulin sensitivity compared with a conventional diabetic diet over 24 weeks. The greater loss of visceral fat and improvements in plasma concentrations of adipokines and oxidative stress markers with this diet may be responsible for the reduction of insulin resistance. The addition of exercise training further augmented the improved outcomes with the vegetarian diet.

negative: Kinetic evidence for rapid oxidation of (-)-epicatechin by human myeloperoxidase. Apocynin has been reported to require dimerization by myeloperoxidase (MPO) to inhibit leukocyte NADPH oxidase. (-)-Epicatechin, a dietary flavan-3-ol, has been identified as a 'prodrug' of apocynin-like metabolites that inhibit endothelial NADPH oxidase activity and elevate the cellular level of nitric oxide. Since (-)-epicatechin has tentatively been identified as substrate of MPO, we studied the one-electron oxidation of (-)-epicatechin by MPO. By using multi-mixing stopped-flow technique, we demonstrate that (-)-epicatechin is one of the most efficient electron donors for heme peroxidases investigated so far. Second order rate constants for the (-)-epicatechin-mediated conversion of MPO-compound I to compound II and compound II to resting enzyme were estimated to be 1.9 x 10(7) and 4.5 x 10(6) M(-1)s(-1), respectively (pH 7, 25 degrees C). The data indicate that (-)-epicatechin is capable of undergoing fast MPO-mediated one-electron oxidation.

negative: Moderate regular exercise increases basal production of nitric oxide in elderly women. Vascular endothelial cells produce nitric oxide (NO), which is a potent vasodilator substance and is thought to have antiatherosclerotic properties. Therefore, it has also been proposed that NO may be useful to regulate vascular tonus and prevent progression of atherosclerosis. On the other hand, NO activity reduces with aging. We previously reported that the plasma nitrite/nitrate (NOx: the stable end product of NO) concentration was significantly increased by intense aerobic exercise training in healthy young humans. We hypothesized that lifestyle modification (e.g., even mild regular exercise training) can increase NO production in previously sedentary older humans. We measured the plasma NOx concentration before and after a mild aerobic exercise training regimen (cycling on a leg ergometer at 80% ventilatory threshold for 30 min, 5 days/week) for 3 months in elderly women. In addition, we assessed the plasma concentration of cyclic guanosine monophosphate (cGMP), a second messenger of NO, in the same samples. The individual ventilatory threshold increased significantly after the 3-month exercise training. The blood pressure at rest significantly decreased after exercise training. These results suggest that the 3-month exercise training in the older women produced favorable physiological effects. The plasma concentration of NOx significantly increased by the exercise training, and the plasma concentration of cGMP also increased by the exercise training. The present study suggests that even a mild regular aerobic-endurance exercise increases NO production in previously sedentary older humans, which may have beneficial effects (i.e., antihypertensive and antiatherosclerotic effects by endogenous NO) on the cardiovascular system.

negative: gamma-tocopherol, the major form of vitamin E in the US diet, deserves more attention. gamma-tocopherol is the major form of vitamin E in many plant seeds and in the US diet, but has drawn little attention compared with alpha-tocopherol, the predominant form of vitamin E in tissues and the primary form in supplements. However, recent studies indicate that gamma-tocopherol may be important to human health and that it possesses unique features that distinguish it from alpha-tocopherol. gamma-Tocopherol appears to be a more effective trap for lipophilic electrophiles than is alpha-tocopherol. gamma-Tocopherol is well absorbed and accumulates to a significant degree in some human tissues; it is metabolized, however, largely to 2,7,8-trimethyl-2-(beta-carboxyethyl)-6-hydroxychroman (gamma-CEHC), which is mainly excreted in the urine. gamma-CEHC, but not the corresponding metabolite derived from alpha-tocopherol, has natriuretic activity that may be of physiologic importance. Both gamma-tocopherol and gamma-CEHC, but not alpha-tocopherol, inhibit cyclooxygenase activity and, thus, possess antiinflammatory properties. Some human and animal studies indicate that plasma concentrations of gamma-tocopherol are inversely associated with the incidence of cardiovascular disease and prostate cancer. These distinguishing features of gamma-tocopherol and its metabolite suggest that gamma-tocopherol may contribute significantly to human health in ways not recognized previously. This possibility should be further evaluated, especially considering that high doses of alpha-tocopherol deplete plasma and tissue gamma-tocopherol, in contrast with supplementation with gamma-tocopherol, which increases both. We review current information on the bioavailability, metabolism, chemistry, and nonantioxidant activities of gamma-tocopherol and epidemiologic data concerning the relation between gamma-tocopherol and cardiovascular disease and cancer.

negative: Agreement between an online dietary assessment tool (myfood24) and an interviewer-administered 24-h dietary recall in British adolescents aged 11-18 years. myfood24 Is an online 24-h dietary assessment tool developed for use among British adolescents and adults. Limited information is available regarding the validity of using new technology in assessing nutritional intake among adolescents. Thus, a relative validation of myfood24 against a face-to-face interviewer-administered 24-h multiple-pass recall (MPR) was conducted among seventy-five British adolescents aged 11-18 years. Participants were asked to complete myfood24 and an interviewer-administered MPR on the same day for 2 non-consecutive days at school. Total energy intake (EI) and nutrients recorded by the two methods were compared using intraclass correlation coefficients (ICC), Bland-Altman plots (using between and within-individual information) and weighted κ to assess the agreement. Energy, macronutrients and other reported nutrients from myfood24 demonstrated strong agreement with the interview MPR data, and ICC ranged from 0·46 for Na to 0·88 for EI. There was no significant bias between the two methods for EI, macronutrients and most reported nutrients. The mean difference between myfood24 and the interviewer-administered MPR for EI was -230 kJ (-55 kcal) (95 % CI -490, 30 kJ (-117, 7 kcal); P=0·4) with limits of agreement ranging between 39 % (3336 kJ (-797 kcal)) lower and 34 % (2874 kJ (687 kcal)) higher than the interviewer-administered MPR. There was good agreement in terms of classifying adolescents into tertiles of EI (κ w =0·64). The agreement between day 1 and day 2 was as good for myfood24 as for the interviewer-administered MPR, reflecting the reliability of myfood24. myfood24 Has the potential to collect dietary data of comparable quality with that of an interviewer-administered MPR.

negative: Role of circulating nitrite and S-nitrosohemoglobin in the regulation of regional blood flow in humans. To determine the relative contributions of endothelial-derived nitric oxide (NO) vs. intravascular nitrogen oxide species in the regulation of human blood flow, we simultaneously measured forearm blood flow and arterial and venous levels of plasma nitrite, LMW-SNOs and HMW-SNOs, and red cell S-nitrosohemoglobin (SNO-Hb). Measurements were made at rest and during regional inhibition of NO synthesis, followed by forearm exercise. Surprisingly, we found significant circulating arterial-venous plasma nitrite gradients, providing a novel delivery source for intravascular NO. Further supporting the notion that circulating nitrite is bioactive, the consumption of nitrite increased significantly with exercise during the inhibition of regional endothelial synthesis of NO. The role of circulating S-nitrosothiols and SNO-Hb in the regulation of basal vascular tone is less certain. We found that low-molecular-weight S-nitrosothiols were undetectable and S-nitroso-albumin levels were two logs lower than previously reported. In fact, S-nitroso-albumin primarily formed in the venous circulation, even during NO synthase inhibition. Whereas SNO-Hb was measurable in the human circulation (brachial artery levels of 170 nM in whole blood), arterial-venous gradients were not significant, and delivery of NO from SNO-Hb was minimal. In conclusion, we present data that suggest (i) circulating nitrite is bioactive and provides a delivery gradient of intravascular NO, (ii) S-nitroso-albumin does not deliver NO from the lungs to the tissue but forms in the peripheral circulation, and (iii) SNO-Hb and S-nitrosothiols play a minimal role in the regulation of basal vascular tone, even during exercise stress.

negative: Nicotinamide Phosphoribosyltransferase/Visfatin Does Not Catalyze Nicotinamide Mononucleotide Formation in Blood Plasma Nicotinamide (Nam) phosphoribosyltransferase (NAMPT) is the rate-limiting enzyme in mammalian NAD synthesis, catalyzing nicotinamide mononucleotide (NMN) formation from Nam and 5-phosphoribosyl 1-pyrophosphate (PRPP). NAMPT has also been described as an adipocytokine visfatin with a variety of actions, although physiological significance of this protein remains unclear. It has been proposed that possible actions of visfatin are mediated through the extracellular formation of NMN. However, we did not detect NMN in mouse blood plasma, even with a highly specific and sensitive liquid chromatography/tandem mass spectrometry. Furthermore, there is no or little ATP, the activator of NAMPT, in extracellular spaces. We thus questioned whether visfatin catalyzes the in situ formation of NMN under such extracellular milieus. To address this question, we here determined K(m) values for the substrates Nam and PRPP in the NAMPT reaction without or with ATP using a recombinant human enzyme and found that 1 mM ATP dramatically decreases K(m) values for the substrates, in particular PRPP to its intracellular concentration. Consistent with the kinetic data, only when ATP is present at millimolar levels, NAMPT efficiently catalyzed the NMN formation at the intracellular concentrations of the substrates. Much lower concentrations of Nam and almost the absence of PRPP and ATP in the blood plasma suggest that NAMPT should not efficiently catalyze its reaction under the extracellular milieu. Indeed, NAMPT did not form NMN in the blood plasma. From these kinetic analyses of the enzyme and quantitative determination of its substrates, activator, and product, we conclude that visfatin does not participate in NMN formation under the extracellular milieus. Together with the absence of NMN in the blood plasma, our conclusion does not support the concept of "NAMPT-mediated systemic NAD biosynthesis. " Our study would advance current understanding of visfatin physiology.

negative: Steroid hormone signalling links reproduction to lifespan in dietary-restricted Caenorhabditis elegans. Dietary restriction (DR) increases healthspan and longevity in many species, including primates, but it is often accompanied by impaired reproductive function. Whether signals associated with the reproductive system contribute to or are required for DR effects on lifespan has not been established. Here we show that expression of the cytochrome P450 DAF-9/CYP450 and production of the steroid hormone Δ(7)-dafachronic acid (DA) are increased in C. elegans subjected to DR. DA signalling through the non-canonical nuclear hormone receptor NHR-8/NHR and the nutrient-responsive kinase let-363/mTOR is essential for DR-mediated longevity. Steroid signalling also affects germline plasticity in response to nutrient deprivation and this is required to achieve lifespan extension. These data demonstrate that steroid signalling links germline physiology to lifespan when nutrients are limited, and establish a central role for let-363/mTOR in integrating signals derived from nutrients and steroid hormones.

negative: Mitochondrial respiration in subcutaneous and visceral adipose tissue from patients with morbid obesity. Adipose tissue exerts important endocrine and metabolic functions in health and disease. Yet the bioenergetics of this tissue is not characterized in humans and possible regional differences are not elucidated. Using high resolution respirometry, mitochondrial respiration was quantified in human abdominal subcutaneous and intra-abdominal visceral (omentum majus) adipose tissue from biopsies obtained in 20 obese patients undergoing bariatric surgery. Mitochondrial DNA (mtDNA) and genomic DNA (gDNA) were determined by the PCR technique for estimation of mitochondrial density. Adipose tissue samples were permeabilized and respirometric measurements were performed in duplicate at 37 degrees C. Substrates (glutamate (G) + malate (M) + octanoyl carnitine (O) + succinate (S)) were added sequentially to provide electrons to complex I + II. ADP ((D)) for state 3 respiration was added after GM. Uncoupled respiration was measured after addition of FCCP. Visceral fat contained more mitochondria per milligram of tissue than subcutaneous fat, but the cells were smaller. Robust, stable oxygen fluxes were found in both tissues, and coupled state 3 (GMOS(D)) and uncoupled respiration were significantly (P < 0.05) higher in visceral (0.95 +/- 0.05 and 1.15 +/- 0.06 pmol O(2) s(1) mg(1), respectively) compared with subcutaneous (0.76 +/- 0.04 and 0.98 +/- 0.05 pmol O(2) s(1) mg(1), respectively) adipose tissue. Expressed per mtDNA, visceral adipose tissue had significantly (P < 0.05) lower mitochondrial respiration. Substrate control ratios were higher and uncoupling control ratio lower (P < 0.05) in visceral compared with subcutaneous adipose tissue. We conclude that visceral fat is bioenergetically more active and more sensitive to mitochondrial substrate supply than subcutaneous fat. Oxidative phosphorylation has a higher relative activity in visceral compared with subcutaneous adipose tissue.

negative: Nitric oxide synthase generates superoxide and nitric oxide in arginine-depleted cells leading to peroxynitrite-mediated cellular injury. Besides synthesizing nitric oxide (NO), purified neuronal NO synthase (nNOS) can produce superoxide (.O2-) at lower L-Arg concentrations. By using electron paramagnetic resonance spin-trapping techniques, we monitored NO and .O2- formation in nNOS-transfected human kidney 293 cells. In control transfected cells, the Ca2+ ionophore A23187 triggered NO generation but no .O2- was seen. With cells in L-Arg-free medium, we observed .O2- formation that increased as the cytosolic L-Arg levels decreased, while NO generation declined. .O2- formation was virtually abolished by the specific NOS blocker, N-nitro-L-arginine methyl ester (L-NAME). Nitrotyrosine, a specific nitration product of peroxynitrite, accumulated in L-Arg-depleted cells but not in control cells. Activation by A23187 was cytotoxic to L-Arg-depleted, but not to control cells, with marked lactate dehydrogenase release. The cytotoxicity was largely prevented by either superoxide dismutase or L-NAME. Thus, with reduced L-Arg availability NOS elicits cytotoxicity by generating .O2- and NO that interact to form the potent oxidant peroxynitrite. Regulating arginine levels may provide a therapeutic approach to disorders involving .O2-/NO-mediated cellular injury.

negative: Vegan proteins may reduce risk of cancer, obesity, and cardiovascular disease by promoting increased glucagon activity. Amino acids modulate the secretion of both insulin and glucagon; the composition of dietary protein therefore has the potential to influence the balance of glucagon and insulin activity. Soy protein, as well as many other vegan proteins, are higher in non-essential amino acids than most animal-derived food proteins, and as a result should preferentially favor glucagon production. Acting on hepatocytes, glucagon promotes (and insulin inhibits) cAMP-dependent mechanisms that down-regulate lipogenic enzymes and cholesterol synthesis, while up-regulating hepatic LDL receptors and production of the IGF-I antagonist IGFBP-1. The insulin-sensitizing properties of many vegan diets--high in fiber, low in saturated fat--should amplify these effects by down-regulating insulin secretion. Additionally, the relatively low essential amino acid content of some vegan diets may decrease hepatic IGF-I synthesis. Thus, diets featuring vegan proteins can be expected to lower elevated serum lipid levels, promote weight loss, and decrease circulating IGF-I activity. The latter effect should impede cancer induction (as is seen in animal studies with soy protein), lessen neutrophil-mediated inflammatory damage, and slow growth and maturation in children. In fact, vegans tend to have low serum lipids, lean physiques, shorter stature, later puberty, and decreased risk for certain prominent 'Western' cancers; a vegan diet has documented clinical efficacy in rheumatoid arthritis. Low-fat vegan diets may be especially protective in regard to cancers linked to insulin resistance--namely, breast and colon cancer--as well as prostate cancer; conversely, the high IGF-I activity associated with heavy ingestion of animal products may be largely responsible for the epidemic of 'Western' cancers in wealthy societies. Increased phytochemical intake is also likely to contribute to the reduction of cancer risk in vegans. Regression of coronary stenoses has been documented during low-fat vegan diets coupled with exercise training; such regimens also tend to markedly improve diabetic control and lower elevated blood pressure. Risk of many other degenerative disorders may be decreased in vegans, although reduced growth factor activity may be responsible for an increased risk of hemorrhagic stroke. By altering the glucagon/insulin balance, it is conceivable that supplemental intakes of key non-essential amino acids could enable omnivores to enjoy some of the health advantages of a vegan diet. An unnecessarily high intake of essential amino acids--either in the absolute sense or relative to total dietary protein--may prove to be as grave a risk factor for 'Western' degenerative diseases as is excessive fat intake.

negative: Impaired mitochondrial fatty acid oxidation and insulin resistance in aging: novel protective role of glutathione. Aging is associated with impaired fasted oxidation of nonesterified fatty acids (NEFA) suggesting a mitochondrial defect. Aging is also associated with deficiency of glutathione (GSH), an important mitochondrial antioxidant, and with insulin resistance. This study tested whether GSH deficiency in aging contributes to impaired mitochondrial NEFA oxidation and insulin resistance, and whether GSH restoration reverses these defects. Three studies were conducted: (i) in 82-week-old C57BL/6 mice, the effect of naturally occurring GSH deficiency and its restoration on mitochondrial (13) C1 -palmitate oxidation and glucose metabolism was compared with 22-week-old C57BL/6 mice; (ii) in 20-week C57BL/6 mice, the effect of GSH depletion on mitochondrial oxidation of (13) C1 -palmitate and glucose metabolism was studied; (iii) the effect of GSH deficiency and its restoration on fasted NEFA oxidation and insulin resistance was studied in GSH-deficient elderly humans, and compared with GSH-replete young humans. Chronic GSH deficiency in old mice and elderly humans was associated with decreased fasted mitochondrial NEFA oxidation and insulin resistance, and these defects were reversed with GSH restoration. Acute depletion of GSH in young mice resulted in lower mitochondrial NEFA oxidation, but did not alter glucose metabolism. These data suggest that GSH is a novel regulator of mitochondrial NEFA oxidation and insulin resistance in aging. Chronic GSH deficiency promotes impaired NEFA oxidation and insulin resistance, and GSH restoration reverses these defects. Supplementing diets of elderly humans with cysteine and glycine to correct GSH deficiency could provide significant metabolic benefits.

negative: Mechanism of NO-mediated oxidative and nitrative DNA damage in hamsters infected with Opisthorchis viverrini: a model of inflammation-mediated carcinogenesis. Inflammation mediated by infection is an important factor causing carcinogenesis. Opisthorchis viverrini (OV) infection is a risk factor of cholangiocarcinoma (CHCA), probably through chronic inflammation. Formation of 8-nitroguanine and 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG), and expression of inducible nitric oxide synthase (iNOS) and heme oxygenase-1 (HO-1) were assessed in the liver of hamsters infected with OV. We newly produced specific anti-8-nitroguanine antibody without cross-reaction. Double immunofluorescence staining revealed that 8-oxodG and 8-nitroguanine were formed mainly in the same inflammatory cells and epithelium of bile ducts from day 7 and showed the strongest immunoreactivity on days 21 and 30, respectively. It is noteworthy that 8-oxodG and 8-nitroguanine still remained in epithelium of bile ducts on day 180, although amount of alanine aminotransferase activity returned to normal level. A time course of 8-nitroguanine was associated with iNOS expression. Furthermore, this study demonstrated that HO-1 expression and subsequent iron accumulation may be involved in enhancement of oxidative DNA damage in epithelium of small bile ducts. In conclusion, nitrative and oxidative DNA damage via iNOS expression in hamsters infected with OV may participate in CHCA carcinogenesis.

negative: Relation of plasma leptin concentrations to sex, body fat, dietary intake, and peak oxygen uptake in young adult women and men. The purpose of this study was to examine the relation of leptin to metabolic and dietary factors in college-age adults. Young adult women and men (n = 32) were recruited and underwent testing for measurement of body mass index, body composition, peak oxygen consumption (VO2peak), dietary intake, and plasma levels of leptin and insulin. Ln leptin was significantly greater for women than for men (2.1 versus 1.2 ng/mL, respectively). This difference remained significant even after adjusting ln leptin for fat mass and fat-free mass as covariates in separate analyses. VO2peak was higher for men than for women and this remained significant after adjustment for differences in fat-free mass and total body mass. Significant correlations were found between ln leptin and indicators of fat mass in women and men, with higher correlations for similar variables observed in men (r = 0.548, 0.674, and 0.732 for body mass index, percentage of body fat, and fat mass [kg] for women, respectively, and r = 0.740, 0.888, 0.858 for body mass index, percentage of body fat, and fat mass [kg] for men, respectively). Ln leptin showed a significant inverse relationship with VO2peak (r = -0.751) in men only. After adjusting ln leptin for body fat mass using partial correlations, ln leptin was not significantly associated with any of the measured variables. Alternatively, after normalization of ln leptin using fat mass as the divisor, a less adequate statistical analysis method, men showed statistical significant correlations between ln leptin and dietary intake and VO2peak. Although plasma leptin values were higher in women, stronger associations were evident for men than for women between leptin and metabolic and dietary factors.

negative: Do Montelukast Sodium and N-Acetylcysteine Have a Nephroprotective Effect on Unilateral Ureteral Obstruction? A Placebo Controlled Trial in a Rat Model. PURPOSE We assessed the nephroprotective effects of montelukast sodium and N-acetylcysteine on secondary renal damage due to unilateral ureteral obstruction in a rat model. MATERIALS AND METHODS In this study 30 Wistar albino male rats were randomized into 3 groups, including placebo, N-acetylcysteine and montelukast sodium. Three rats served as the control group. The left ureter of the rats was sutured with 4-zero polyglactin sutures. Medications were given 3 days before obstruction and continued for 15 days. Dimercaptosuccinic acid renal scintigraphy was performed before obstruction and on day 15. Rats were sacrificed on day 15 and histopathological examinations were done. We biochemically assessed oxidative stress markers (myeloperoxidase and malondialdehyde), sulfhydryl and total nitrite for lipid peroxidation, oxidative protein damage and antioxidant levels, respectively. RESULTS On pathological examination inflammation and tubular epithelial damage in the N-acetylcysteine and montelukast sodium groups were less than in the placebo group (p <0.05). No difference was seen in normal kidneys. Myeloperoxidase, malondialdehyde and total nitrite levels in the N-acetylcysteine group, and myeloperoxidase and malondialdehyde levels in the montelukast sodium group were lower than in the placebo group (p <0.05). No statistical difference was seen in sulfhydryl levels (p >0.05) or among the N-acetylcysteine, montelukast sodium and placebo groups on scintigraphy (p >0.05). No pathological, chemical and scintigraphic differences were seen among the N-acetylcysteine, montelukast sodium and sham treated groups (p >0.05). CONCLUSIONS N-acetylcysteine and montelukast sodium have a protective effect against obstructive damage of the kidney. However, further investigations are needed.

negative: The pecking order of free radicals and antioxidants: lipid peroxidation, alpha-tocopherol, and ascorbate. Free radicals vary widely in their thermodynamic properties, ranging from very oxidizing to very reducing. These thermodynamic properties can be used to predict a pecking order, or hierarchy, for free radical reactions. Using one-electron reduction potentials, the predicted pecking order is in agreement with experimentally observed free radical electron (hydrogen atom) transfer reactions. These potentials are also in agreement with experimental data that suggest that vitamin E, the primary lipid soluble small molecule antioxidant, and vitamin C, the terminal water soluble small molecule antioxidant, cooperate to protect lipids and lipid structures against peroxidation. Although vitamin E is located in membranes and vitamin C is located in aqueous phases, vitamin C is able to recycle vitamin E; i.e., vitamin C repairs the tocopheroxyl (chromanoxyl) radical of vitamin E, thereby permitting vitamin E to function again as a free radical chain-breaking antioxidant. This review discusses: (i) the thermodynamics of free radical reactions that are of interest to the health sciences; (ii) the fundamental thermodynamic and kinetic properties that are associated with chain-breaking antioxidants; (iii) the unique interfacial nature of the apparent reaction of the tocopherol free radical (vitamin E radical) and vitamin C; and (iv) presents a hierarchy, or pecking order, for free radical electron (hydrogen atom) transfer reactions.

negative: Dietary saturated and unsaturated fats as determinants of blood pressure and vascular function. The amount and type of dietary fat have long been associated with the risk of CVD. Arterial stiffness and endothelial dysfunction are important risk factors in the aetiology of CHD. A range of methods exists to assess vascular function that may be used in nutritional science, including clinic and ambulatory blood pressure monitoring, pulse wave analysis, pulse wave velocity, flow-mediated dilatation and venous occlusion plethysmography. The present review focuses on the quantity and type of dietary fat and effects on blood pressure, arterial compliance and endothelial function. Concerning fat quantity, the amount of dietary fat consumed habitually appears to have little influence on vascular function independent of fatty acid composition, although single high-fat meals postprandially impair endothelial function compared with low-fat meals. The mechanism is related to increased circulating lipoproteins and NEFA which may induce pro-inflammatory pathways and increase oxidative stress. Regarding the type of fat, cross-sectional data suggest that saturated fat adversely affects vascular function whereas polyunsaturated fat (mainly linoleic acid (18 : 2n-6) and n-3 PUFA) are beneficial. EPA (20 : 5n-3) and DHA (22 : 6n-3) can reduce blood pressure, improve arterial compliance in type 2 diabetics and dyslipidaemics, and augment endothelium-dependent vasodilation. The mechanisms for this vascular protection, and the nature of the separate physiological effects induced by EPA and DHA, are priorities for future research. Since good-quality observational or interventional data on dietary fatty acid composition and vascular function are scarce, no further recommendations can be suggested in addition to current guidelines at the present time.

negative: Pyruvate dehydrogenase activity and acetyl group accumulation during exercise after different diets. Pyruvate dehydrogenase activity (PDHa) and acetyl group accumulation were examined in human skeletal muscle at rest and during exercise after different diets. Five males cycled at 75% of maximal O2 uptake (VO2 max) to exhaustion after consuming a low-carbohydrate diet (LCD) for 3 days and again 1-2 wk later for the same duration after consuming a high-carbohydrate diet (HCD) for 3 days. Resting PDHa was lower after a LCD (0.20 +/- 0.04 vs. 0.69 +/- 0.05 mmol.min-1.kg wet wt-1; P < 0.05) and coincided with a greater intramuscular acetyl-CoA-to-CoASH ratio, acetyl-CoA content, and acetylcarnitine content. PDHa increased during exercise in both conditions but at a lower rate in the LCD condition compared with the HCD condition (1.46 +/- 0.25 vs. 2.65 +/- 0.23 mmol.min-1.kg wet wt-1 at 16 min and 1.88 +/- 0.20 vs. 3.11 +/- 0.14 at the end of exercise; P < 0.05). During exercise muscle acetyl-CoA and acetylcarnitine content and the acetyl-CoA-to-CoASH ratio decreased in the LCD condition but increased in the HCD condition. Under resting conditions PDHa was influenced by the availability of fat or carbohydrate fuels acting through changes in the acetyl-CoA-to-CoASH ratio. However, during exercise the activation of PDHa occurred independent of changes in the acetyl-CoA-to-CoASH ratio, suggesting that other factors are more important.