Dietary nitrate supplementation reduces the O2 cost of walking and running: a placebo-controlled study

Dietary supplementation with beetroot juice (BR) has been shown to reduce resting blood pressure and the O(2) cost of submaximal exercise and to increase tolerance to high-intensity cycling. We tested the hypothesis that the physiological effects of BR were consequent to its high NO(3)(-) content per se, and not the presence of other potentially bioactive compounds. We investigated changes in blood pressure, mitochondrial oxidative capacity (Q(max)), and physiological responses to walking and moderate- and severe-intensity running following dietary supplementation with BR and NO(3)(-)-depleted BR [placebo (PL)]. After control (nonsupplemented) tests, nine healthy, physically active male subjects were assigned in a randomized, double-blind, crossover design to receive BR (0.5 l/day, containing ～6.2 mmol of NO(3)(-)) and PL (0.5 l/day, containing ～0.003 mmol of NO(3)(-)) for 6 days. Subjects completed treadmill exercise tests on days 4 and 5 and knee-extension exercise tests for estimation of Q(max) (using (31)P-magnetic resonance spectroscopy) on day 6 of the supplementation periods. Relative to PL, BR elevated plasma NO(2)(-) concentration (183 ± 119 vs. 373 ± 211 nM, P < 0.05) and reduced systolic blood pressure (129 ± 9 vs. 124 ± 10 mmHg, P < 0.01). Q(max) was not different between PL and BR (0.93 ± 0.05 and 1.05 ± 0.22 mM/s, respectively). The O(2) cost of walking (0.87 ± 0.12 and 0.70 ± 0.10 l/min in PL and BR, respectively, P < 0.01), moderate-intensity running (2.26 ± 0.27 and 2.10 ± 0.28 l/min in PL and BR, respectively, P < 0.01), and severe-intensity running (end-exercise O(2) uptake = 3.77 ± 0.57 and 3.50 ± 0.62 l/min in PL and BL, respectively, P < 0.01) was reduced by BR, and time to exhaustion during severe-intensity running was increased by 15% (7.6 ± 1.5 and 8.7 ± 1.8 min in PL and BR, respectively, P < 0.01). In contrast, relative to control, PL supplementation did not alter plasma NO(2)(-) concentration, blood pressure, or the physiological responses to exercise. These results indicate that the positive effects of 6 days of BR supplementation on the physiological responses to exercise can be ascribed to the high NO(3)(-) content per se.     

Muscle fiber recruitment and the slow component of O2 uptake: constant work rate vs. all-out sprint exercise

The slow component of pulmonary O(2) uptake (Vo(2)) during constant work rate (CWR) high-intensity exercise has been attributed to the progressive recruitment of (type II) muscle fibers. We tested the following hypotheses: 1) the Vo(2) slow component gain would be greater in a 3-min all-out cycle test than in a work-matched CWR test, and 2) the all-out test would be associated with a progressive decline, and the CWR test with a progressive increase, in muscle activation, as estimated from the electromyogram (EMG) of the vastus lateralis muscle. Eight men (aged 21-39 yr) completed a ramp incremental test, a 3-min all-out test, and a work- and time-matched CWR test to exhaustion. The maximum Vo(2) attained in an initial ramp incremental test (3.97 ± 0.83 l/min) was reached in both experimental tests (3.99 ± 0.84 and 4.03 ± 0.76 l/min for all-out and CWR, respectively). The Vo(2) slow component was greater (P < 0.05) in the all-out test (1.21 ± 0.31 l/min, 4.2 ± 2.2 ml·min(-1)·W(-1)) than in the CWR test (0.59 ± 0.22 l/min, 1.70 ± 0.5 ml·min(-1)·W(-1)). The integrated EMG declined by 26% (P < 0.001) during the all-out test and increased by 60% (P < 0.05) during the CWR test from the first 30 s to the last 30 s of exercise. The considerable reduction in muscle efficiency in the all-out test in the face of a progressively falling integrated EMG indicates that progressive fiber recruitment is not requisite for development of the Vo(2) slow component during voluntary exercise in humans.     

Distinct profiles of neuromuscular fatigue during muscle contractions below and above the critical torque in humans

Whether the transition in fatigue processes between "low-intensity" and "high-intensity" contractions occurs gradually, as the torque requirements are increased, or whether this transition occurs more suddenly at some identifiable "threshold", is not known. We hypothesized that the critical torque (CT; the asymptote of the torque-duration relationship) would demarcate distinct profiles of central and peripheral fatigue during intermittent isometric quadriceps contractions (3-s contraction, 2-s rest). Nine healthy men performed seven experimental trials to task failure or for up to 60 min, with maximal voluntary contractions (MVCs) performed at the end of each minute. The first five trials were performed to determine CT [~35-55% MVC, denoted severe 1 (S1) to severe 5 (S5) in ascending order], while the remaining two trials were performed 10 and 20% below the CT (denoted CT-10% and CT-20%). Dynamometer torque and the electromyogram of the right vastus lateralis were sampled continuously. Peripheral and central fatigue was determined from the fall in potentiated doublet torque and voluntary activation, respectively. Above CT, contractions progressed to task failure in ~3-18 min, at which point the MVC did not differ from the target torque (S1 target, 88.7 ± 4.3 N·m vs. MVC, 89.3 ± 8.8 N·m, P = 0.94). The potentiated doublet fell significantly in all trials, and voluntary activation was reduced in trials S1-S3, but not trials S4 and S5. Below CT, contractions could be sustained for 60 min on 17 of 18 occasions. Both central and peripheral fatigue developed, but there was a substantial reserve in MVC torque at the end of the task. The rate of global and peripheral fatigue development was four to five times greater during S1 than during CT-10% (change in MVC/change in time S1 vs. CT-10%: -7.2 ± 1.4 vs. -1.5 ± 0.4 N·m·min(-1)). These results demonstrate that CT represents a critical threshold for neuromuscular fatigue development.     

(+/−)-Alashanoid N,Two Enantiomeric Sesquiterpenes from Syringa pinnatifolia

Two enantiomeric humulane sesquiterpenes, namely (+)-alashanoid N ( 1a ) and (−)-alashanoid N ( 1b ), along with two known analogs ((2R,3R,5R)-2,3-epoxy-6,9-humuladien-5-ol-8-one ( 2 ) and (2R,3S,5R)-2,3-epoxy-6,9-humuladien-5-ol-8-one ( 3 )), were described from the peeled stems of a folk Mongolian herbal medicine Syringa pinnatifolia. Their structures were characterized based on UV, IR, NMR, and HR-ESI-MS data analyses, and the absolute configurations were determined by data analysis of X-ray diffraction and quantum chemical calculations. (+/−)-Alashanoid N showed inhibition against NO production in RAW 264.7 macrophage cells with IC₅₀ values of 90.1 μM and 71.7 μM, and protective effect against oxygen-glucose deprivation injury to H9c2 cells at a concentration of 20 μM and 5 μM, respectively.     

Domains I and IV of Annexin A2 Affect the Formation and Integrity of In Vitro Capillary-Like Networks

Annexin A2 (AnxA2) is a widely expressed multifunctional protein found in different cellular compartments. In spite of lacking a hydrophobic signal peptide, AnxA2 is found at the cell surface of endothelial cells, indicative of a role in angiogenesis. Increased extracellular levels of AnxA2 in tumours correlate with neoangiogenesis, metastasis and poor prognosis. We hypothesised that extracellular AnxA2 may contribute to angiogenesis by affecting endothelial cell-cell interactions and motility. To address this question, we studied the effect of heterotetrameric and monomeric forms of AnxA2, as well as its two soluble domains on the formation and maintenance of capillary-like structures by using an in vitro co-culture system consisting of endothelial and smooth muscle cells. In particular, addition of purified domains I and IV of AnxA2 potently inhibited the vascular endothelial growth factor (VEGF)-dependent formation of the capillary-like networks in a dose-dependent manner. In addition, these AnxA2 domains disrupted endothelial cell-cell contacts in preformed capillary-like networks, resulting in the internalisation of vascular endothelial (VE)-cadherin and the formation of VE-cadherin-containing filopodia-like structures between the endothelial cells, suggesting increased cell motility. Addition of monoclonal AnxA2 antibodies, in particular against Tyr23 phosphorylated AnxA2, also strongly inhibited network formation in the co-culture system. These results suggest that extracellular AnxA2, most likely in its Tyr phosphorylated form, plays a pivotal role in angiogenesis. The exogenously added AnxA2 domains most likely mediate their effects by competing with endogenous AnxA2 for extracellular factors necessary for the initiation and maintenance of angiogenesis, such as those involved in the formation/integrity of cell-cell contacts.     

Stem emissions of monoterpenes,acetaldehyde and methanol from Scots pine (Pinus sylvestris L.) affected by tree–water relations and cambial growth

Tree stems are an overlooked source of volatile organic compounds (VOCs). Their contribution to ecosystem processes and total VOC fluxes is not well studied, and assessing it requires better understanding of stem emission dynamics and their driving processes. To gain more mechanistic insight into stem emission patterns, we measured monoterpene, methanol and acetaldehyde emissions from the stems of mature Scots pines (Pinus sylvestris L.) in a boreal forest over three summers. We analysed the effects of temperature, soil water content, tree water status, transpiration and growth on the VOC emissions and used generalized linear models to test their relative importance in explaining the emissions. We show that Scots pine stems are considerable sources of monoterpenes, methanol and acetaldehyde, and their emissions are strongly regulated by temperature. However, even small changes in water availability affected the emission potentials: increased soil water content increased the monoterpene emissions within a day, whereas acetaldehyde and methanol emissions responded within 2–4 days. This lag corresponded to their transport time in the xylem sap from the roots to the stem. Moreover, the emissions of monoterpenes, methanol and acetaldehyde were influenced by the cambial growth rate of the stem with 6–10-day lags.     

Yeast cytochrome c peroxidase. Coordination and spin states of heme prosthetic group

Electronic absorption and electron paramagnetic resonance (EPR) spectroscopic examinations revealed that a freshly prepared cytochrome c peroxidase (CCP) contains a penta-coordinated high spin ferric protoheme group. The penta-coordinated high spin state of fresh CCP is maintained in a remarkably wide range of pH (4-8). The freezing of fresh CCP induces the reversible coordination of an internal strong field ligand to the heme iron to form a hexa-coordinated low spin compound, which shows EPR extrema at gx = 2.70, gy = 2.20 and gz = 1.78. In the presence of glycerol the freezing-induced artifacts are eliminated and the fresh enzyme exhibits an EPR spectrum of rhombically distorted axial symmetry with EPR extrema at gx = 6.4, gy = 5.3, and gz = 1.97 at 10 K, characteristic of the penta-coordinated high spin enzyme. Upon aging CCP is converted to a hexa-coordinated high spin state due to the coordination of an internal weak field ligand to the heme iron. This conversion is accelerated at acidic pH values, and its reversibility varies from fully reversible to irreversible depending on the degree of enzyme aging. The aging-induced hexa-coordinated CCP is unreactive with hydrogen peroxide and exhibits an EPR spectrum of purely axial symmetry with extrema at g = 6 and g = 2 and an electronic absorption spectrum with an intensified Soret band at 408 nm (epsilon 408 nm = 120 mM-1 cm-1) and a blue-shifted charge-transfer band at 620 nm. Spectroscopic properties of different coordination and spin states of fresh and aged CCPs are compiled in order to formulate a generalized spectroscopic characterization of penta- and hexa-coordinated high spin ferric hemoproteins.     

Phylogenetic analyses of two mitochondrial metabolic genes sampled in parallel from angiosperms find fundamental interlocus incongruence

Plant molecular phylogeneticists have supported an analytical approach of combining loci from different genomes, but the combination of mitochondrial sequences with chloroplast and nuclear sequences is potentially problematic. Low substitution rates in mitochondrial genes should decrease saturation, which is especially useful for the study of deep divergences. However, individual mitochondrial loci are insufficiently informative, so that combining congruent loci is necessary. For this study atp1 and cox1 were selected, which are of similar lengths, encode components of the respiratory pathway, and generally lack introns. Thus, these genes might be expected to have similar functional constraints, selection pressures, and evolutionary histories. Strictly parallel sampling of 52 species was achieved as well as six additional composite terminals with representatives from the major angiosperm clades. However, analyses of the separate loci produced strongly incongruent topologies. The source of the incongruence was investigated by validating sequences with questionable affinities, excluding RNA-edited nucleotides, deleting taxa with unexpected phylogenetic associations, and comparing different phylogenetic methods. However, even after potential artifacts were addressed and sites and taxa putatively associated with conflict were excluded, the resulting gene trees for the two mitochondrial loci were still substantially incongruent by all measures examined. Therefore, combining these loci in phylogenetic analysis may be counterproductive to the goal of fully resolving the angiosperm phylogeny.     

Weight loss and the effect on stature in children during a residential intervention program

Weight loss is generally high in residential weight-loss programs but the effect of a large weight loss on linear growth is not known. We report the weight loss and the influence on linear growth in a large group of children during a residential weight-loss program focusing on nutrition and physical activity. In a longitudinal noncontrolled intervention study of 990 overweight children (540 girls) attending the weight reduction program from 1990 to 2001 for about 11 weeks (age: 10-14 years, mean BMI-standard deviation score (SDS) at enrollment: 2.83) weight and height were measured initially and after end of treatment. Weekly measurements of height and weight were performed on 138 children. The children lost on average 9.4 kg, reduced their BMI by 4.5 kg/m(2) and BMI-SDS by 0.98. In a multiple regression analysis (P < or = 0.001) weight loss was higher in boys than girls (1.7 kg), higher if the weight was higher at admission (-0.192 kg/kg at baseline) and was positively associated with duration of stay (-80 g/day). Initially the boys' BMI-SDS was higher than the girls' BMI-SDS (P < or = 0.05) but after 8 weeks of treatment the boys had lower BMI-SDS than the girls. There was no negative effect on linear growth during the treatment; on the contrary, linear growth accelerated during the stay as the average increase in height was 2.38 cm corresponding to 11.4 cm/year. In conclusion the children lost close to 1 kg/week during the stay without any negative effect on linear growth. The cause of the linear growth acceleration needs further investigation.     

Assembly and structural analysis of a covalently closed nano-scale DNA cage

The inherent properties of DNA as a stable polymer with unique affinity for partner molecules determined by the specific Watson–Crick base pairing makes it an ideal component in self-assembling structures. This has been exploited for decades in the design of a variety of artificial substrates for investigations of DNA-interacting enzymes. More recently, strategies for synthesis of more complex two-dimensional (2D) and 3D DNA structures have emerged. However, the building of such structures is still in progress and more experiences from different research groups and different fields of expertise are necessary before complex DNA structures can be routinely designed for the use in basal science and/or biotechnology. Here we present the design, construction and structural analysis of a covalently closed and stable 3D DNA structure with the connectivity of an octahedron, as defined by the double-stranded DNA helices that assembles from eight oligonucleotides with a yield of ~30%. As demonstrated by Small Angle X-ray Scattering and cryo-Transmission Electron Microscopy analyses the eight-stranded DNA structure has a central cavity larger than the apertures in the surrounding DNA lattice and can be described as a nano-scale DNA cage, Hence, in theory it could hold proteins or other bio-molecules to enable their investigation in certain harmful environments or even allow their organization into higher order structures.