bluuebunny/crossref_metadata_2025 huggingface dataset:
Geographical and ecological differentiation in Greek Fagus forest vegetation
Authors: Tsiripidis Ioannis , Bergmeier Erwin , Dimopoulos Panayotis
Published: February 2009

Question: Which are the gradients of floristic differentiation in Greek beech (Fagus sylvatica) forests? Which is the role of geographical and ecological factors in this differentiation?

Location: Beech forests of the plant geographical regions Northeast, North Central and East Central Greece.

Methods: A total of 1404 published and unpublished phytoso‐ciological relevés were used in the analyses. TWINSPAN and DCA were applied to classify and ordinate the relevés. Altitude, Indicator Values of relevés and their X and Y coordinates were used in a posteriori interpretation of the ordination axes. Kendall's correlation coefficients were calculated between DCA relevé scores and explanatory variables. Multiple linear regression was used to partition the variation explained by the first two DCA axes, between the geographical and the ecological variables.

Results: Classification resulted in 14 vegetation units defined by species composition. Two types of gradients, ecological and geographical, were revealed by the DCA of all releves. The partition of the variation accounted for by the first and second DCA axis was attributed mainly to ecological and geographical variables, respectively.

Conclusions: Beech forests of northeast and Central Greece show phytogeographical differences, while ecologically similar vegetation units occur in both regions. A west‐east gradient is revealed in Greek beech forest vegetation. The extent of the study area, its position along regional gradients and the comprehensiveness of the data set that is analysed determine the types of the gradients which can be revealed in a vegetation study.
Transformation of cover‐abundance values for appropriate numerical treatment ‐ Alternatives to the proposals by Podani
Authors: van der Maarel Eddy
Published: February 2009

Abstract. Two alternatives are offered to Podani's proposals, based on the claim that Braun‐Blanquet cover‐abundance estimates cannot be properly analysed by conventional mul‐tivariate methods.

1. The ordinal transform scale, based on an extended Braun‐Blanquet cover‐abundance scale, comes close to a metric cover percentage scale after (1) the abundance values r (very few individuals), + (few ind.), 1 (abundant) and 2m (very abundant, cover < 5%) are replaced by cover percentage estimates and (2) the higher Braun‐Blanquet values, notably 4 and 5, with cover intervals 50‐75% and 75‐100%, respectively, are interpreted as estimates of considerably higher cover values than the usual visual projection on the ground (because of the position of stems and leaves in several layers). I propose the equation ln C= (OTV −2) /a, where C= Cover%, OTV is the 1 to 9 Ordinal Transfer Value and a is a factor weighting the cover values. With this equation cover values in a geometric series are achieved for the nine values in the extended Braun‐Blanquet scale from 0.5 % (OTV 1) to 140% (OTV 9) for a= 1.415, and for a= 1.380 from 0.6 % to 160%.

2. This makes use of an earlier developed ‘optimum‐transformation’ of cover‐abundance values. For each species a frequency distribution of cover‐abundance values is determined for a large data set, i.e. of dune slack vegetation. Tiny species have low values (OTVs 1–3) with high frequencies and hardly occur with higher OTV values; here all scores are considered ‘optimal’. In dominant species OTVs 7 to 9 have the highest frequencies and only these values are considered optimal. Species with intermediate OTV ranges have optimum ranges with low‐bound OTV = 2, 3, 4 and 5, respectively. No species were found in the dune slack data set with a frequency distribution justifying an optimum range with low‐bound OTV = 6.

For mathematically correct numerical treatments’ optimum scores’ can be converted to 1 and sub‐optimal scores to 0 in order to approach a presence/absence situation.

Both alternatives are suggested to be acceptable approximations to a metric basis for numerical analyses.
Radial growth responses to drought of Pinus sylvestris and Quercus pubescens in an inner‐Alpine dry valley
Authors: Weber Pascale , Bugmann Harald , Rigling Andreas
Published: February 2009

Question: Lower montane treeline ecotones such as the inner Alpine dry valleys are regarded as sensitive to climate change. In the dry Valais valley (Switzerland) the composition of the widespread, low altitude Pinus forests is shifting towards a mixed deciduous state. The sub‐boreal P. sylvestris shows high mortality rates, whereas the deciduous sub‐mediterranean Quercus pubescens is spreading. These species may act as early indicators of climate change. We evaluate this hypothesis by focusing on their differences in drought tolerance, which are hardly known, but are likely to be crucial in the current forest shift and also for future forest development.

Methods: We used dendroecological methods to detect species‐specific patterns in the growth response to drought. The relationship between radial growth of 401 trees from 15 mixed stands and drought was analysed by calculating response functions using yearly tree‐ring indices and monthly drought indices. PCA was applied to the response ratios to discover spatial patterns of drought response.

Results: A species‐specific response to moisture as well as a sub‐regional differentiation of the response patterns were found. While Quercus showed a response mainly to the conditions of the previous autumn and those of current spring, Pinus did not start responding before May, but showed responses throughout the whole summer. Quercus may restrict physiological activity to moist periods; growth of Pinus was much more dependent on prior growth.

Conclusions: Given that the climate is changing towards (1) longer summer drought periods, (2) higher mean temperatures and (3) shifted seasonally of moisture availability, Quercus may benefit from adapting better to drier conditions. Pinus may increasingly face problems related to drought stress as it depends on summer moisture and has a smaller adaptive capacity due to its long‐lived photosynthetic tissue.
Afforestation causes changes in post‐fire regeneration in native shrubland communities of northwestern Patagonia, Argentina
Authors: Nuñez Martín Andrés , Raffaele Estela
Published: February 2009

Question: What are the effects of fire in native shrubland communities and in pine plantations established in these shrublands?

Location: Northern Patagonia, Argentina.

Methods: We surveyed four sites in Chall‐Huaco valley, located in northwest Patagonia. Each site was a vegetation mosaic composed of an unburned Pinus ponderosa plantation, a plantation burned in 1996, and an unburned matorral and a matorral burned by the same fire. We recorded the cover of all vascular plant species. We also analysed species richness, total cover, proportion of exotic species, abundance of woody species and herb species, cover of exotic species, abundance of woody and herb species and differences in composition of species. For both shrubs and tree species we recorded the main strategy of regeneration (by resprouting or by seed).

Results: We found that fire had different effects on native matorral and pine plantations. Five years after fire, plantations came to be dominated by herbs and exotic species, showing differences in floristic composition. In contrast, matorral communities remained very similar to unburned matorral in terms of species richness, proportion of woody species, and herb species and proportion of exotics. Also, pine plantations were primarily colonized by seedlings, while matorrals were primarily colonized by resprouting.

Conclusions: Matorrals are highly fire resilient communities, and the practice of establishing plantations on matorrals produces a strong reduction in the capacity of matorral to return to its original state. The elimination of shrubs owing to the effect of plantations can hinder regeneration of native ecosystems. Burned plantations may slowly develop into ecosystems similar to the native ones, or they may produce a new ecosystem dominated by exotic herbs. This study shows that plantations of exotic conifers affect native vegetation even after they have been removed, as in this case by fire.
Size and heterogeneity rather than landscape context determine plant species richness in semi‐natural grasslands
Authors: Öster Mathias , Cousins Sara A.O. , Eriksson Ove
Published: February 2009

Question:Is plant diversity in fragmented semi‐natural grasslands related to present and historical landscape context?

Location:Southern Sweden.

Methods:Plant diversity was described at 30 semi‐natural grassland sites in terms of total and specialist plant species richness at the site and species density at different scales (0.5–10 m2). These measures are commonly used to assess conservation value of semi‐natural grasslands. Landscape context was measured as contemporary connectivity to other semi‐natural grasslands, historical connectivity 50 years ago, amount of linear elements potentially suitable for dispersal (road verges, power line clearings), and amount of forest (inverse of the openness of the landscape).

Results:The diversity measures were generally correlated with each other, implying that species richness in a subset of the grassland can predict the total richness. Plant species density at three scales (0.5 m2, 10 m2and total) was related to the landscape context using an information theoretic approach. Results showed that total species richness increased with increased size of grasslands, contrary to earlier diversity studies in semi‐natural grasslands. Larger grasslands were more heterogeneous than smaller grasslands, and this is a likely reason for the species‐area relationship. Heterogeneity was also of high importance at the smaller scales (0.5 m2, 10 m2). With increased amount of forest, total species richness increased but species density on 10 m2decreased. There was no influence of connectivity in either the contemporary or the historical landscape, contrary to previous studies.

Conclusions:Grassland size and heterogeneity are of greater importance for plant diversity in semi‐natural grassland, than grassland connectivity in the landscape.
Dispersal and recruitment dynamics in the fleshy‐fruited Persoonia lanceolata (Proteaceae)
Authors: Auld Tony D. , Denham Andrew J. , Turner Ken
Published: February 2009

Question: What is the role of dispersal, persistent soil seed banks and seedling recruitment in population persistence of fleshy‐fruited obligate seeding plant species in fire‐prone habitats?

Location: Southeastern Australia.

Methods: We used a long‐term study of a shrubby, fleshy‐fruited Persoonia species (Proteaceae) to examine (1) seed removal from beneath the canopy of adult plants; (2) seedling recruitment after fire; (3) the magnitude and location of the residual soil seed bank; and (4) the implications for fire management of obligate seeding species. We used demographic sampling techniques combined with Generalised Linear Modelling and regression to quantify population changes over time.

Results: Most of the mature fruits (90%) on the ground below the canopy of plants were removed by Wallabia bicolor (Swamp wallaby) with 88% of seeds extracted from W. bicolor scats viable and dormant. Wallabies play an important role in moving seeds away from parent plants. Their role in occasional long distance dispersal events remains unknown. We detected almost no seed predation in situ under canopies (< 1%). Seedling recruitment was cued to fire, with post‐fire seedling densities 6‐7 times pre‐fire adult densities. After fire, a residual soil seed bank was present, as many seeds (77‐100%) remained dormant and viable at a soil depth where successful future seedling emergence is possible (0‐5 cm). Seedling survival was high (> 80%) with most mortality within 2 years of emergence. Plant growth averaged 17 cm per year. The primary juvenile period of plants was 7–8 years, within the period of likely return fire intervals in the study area. We predicted that the study population increased some five‐fold after the wildfire at the site.

Conclusions: Residual soil seed banks are important, especially in species with long primary juvenile periods, to buffer the populations against the impact of a second fire occurring before the seed bank is replenished.
Filtering processes in the assembly of plant communities: Are species presence and abundance driven by the same traits?
Authors: Cingolani Ana M. , Cabido Marcelo , Gurvich Diego E. , Renison Daniel , Díaz Sandra
Published: February 2009

Question: Is the response of plant traits to environment at the community level similar when considering species abundance and when considering species presence only?

Location: Mountain grasslands, central Argentina.

Methods: We used data from 57 floristic samples, ordinated through DCCA along moisture and grazing gradients combined with trait values from 85 species (plant height, leaf area, leaf thickness leaf toughness and SLA). For each sample, we calculated the weighted average (considering species abundance) and the simple average (considering only species presence). Through multiple regressions we analysed how each average (dependent variable) responded to moisture and grazing (DCCA scores along Axes 1 and 2, respectively, as independent variables).

Results: Weighted averages of all traits were significantly associated to both gradients, while simple averages did not always respond. In some cases the responses followed similar but weaker trends than the responses of weighted averages, but in other cases these responses were qualitatively different. Traits more associated with size (plant height, leaf area, leaf thickness) responded more consistently (similar trends for both averages) to grazing than to moisture, while traits more associated with plant resource acquisition (SLA, leaf toughness) responded more consistently to moisture than to grazing.

Conclusion: The trait values and combinations which determine the probability of species presence are not necessary the same as those which determine their probability of becoming abundant. To understand community assembly rules, both species presence and species abundance should be taken into account as the result of different, although closely linked, filtering processes.
Physicochemical properties and distinct DNA binding capacity of the repressor of temperate Staphylococcus aureus phage φ11
Authors: Ganguly Tridib , Das Malabika , Bandhu Amitava , Chanda Palas K. , Jana Biswanath , Mondal Rajkrishna , Sau Subrata
Published: February 2009

The repressor protein and cognate operator DNA of any temperate Staphylococcus aureus phage have not been investigated in depth, despite having the potential to enrich the molecular biology of the staphylococcal system. In the present study, using the extremely pure repressor of temperate Staphylococcus aureus phage φ11 (CI), we demonstrate that CI is composed of α‐helix and β‐sheet to a substantial extent at room temperature, possesses two domains, unfolds at temperatures above 39 °C and binds to two sites in the φ11 cI‐cro intergenic region with variable affinity. The above CI binding sites harbor two homologous 15 bp inverted repeats (O1 and O2), which are spaced 18 bp apart. Several guanine bases located in and around O1 and O2 demonstrate interaction with CI, indicating that these 15 bp sites are used as operators for repressor binding. CI interacted with O1 and O2 in a cooperative manner and was found to bind to operator DNA as a homodimer. Interestingly, CI did not show appreciable binding to another homologous 15 bp site (O3) that was located in the same primary immunity region as O1 and O2. Taken together, these results suggest that φ11 CI and the φ11 CI–operator complex resemble significantly those of the lambdoid phages at the structural level. The mode of action of φ11 CI, however, may be distinct from that of the repressor proteins of λ and related phages.
Stability and fibril formation properties of human and fish transthyretin, and of theEscherichia colitransthyretin‐related protein
Authors: Lundberg Erik , Olofsson Anders , Westermark Gunilla T. , Sauer‐Eriksson A. Elisabeth
Published: February 2009

Human transthyretin (hTTR) is one of several proteins known to cause amyloid disease. Conformational changes in its native structure result in aggregation of the protein, leading to insoluble amyloid fibrils. The transthyretin (TTR)‐related proteins comprise a protein family of 5‐hydroxyisourate hydrolases with structural similarity to TTR. In this study, we tested the amyloidogenic properties, if any, of sea bream TTR (sbTTR) andEscherichia colitransthyretin‐related protein (ecTRP), which share 52% and 30% sequence identity, respectively, with hTTR. We obtained filamentous structures from all three proteins under various conditions, but, interestingly, different structures displayed different tinctorial properties. hTTR and sbTTR formed thin, curved fibrils at low pH (pH 2–3) that bound thioflavin‐T (thioflavin‐T‐positive) but did not stain with Congo Red (CR) (CR‐negative). Aggregates formed at the slightly higher pH of 4.0–5.5 had different morphology, displaying predominantly amorphous structures. CR‐positive material of hTTR was found in this material, in agreement with previous results. ecTRP remained soluble at pH 2–12 at ambient temperatures. By raising of the temperature, fibril formation could be induced at neutral pH in all three proteins. Most of these temperature‐induced fibrils were thicker and straighter than thein vitrofibrils seen at low pH. In other words, the temperature‐induced fibrils were more similar to fibrils seenin vivo. The melting temperature of ecTRP was 66.7 °C. This is approximately 30 °C lower than the melting temperatures of sbTTR and hTTR. Information from the crystal structures was used to identify possible explanations for the reduced thermostability of ecTRP.
Evaluation of the platelet counting by Abbott CELL‐DYN® SAPPHIRE™ haematology analyser compared with flow cytometry
Authors: GRIMALDI E. , DEL VECCHIO L. , SCOPACASA F. , LO PARDO C. , CAPONE F. , PARIANTE S. , SCALIA G. , CATERINA M. DE
Published: February 2009

The Abbot Cell‐Dyn Sapphire is a new generation haematology analyser. The system uses optical/fluorescence flow cytometry in combination with electronic impedance to produce a full blood count. Optical and impedance are the default methods for platelet counting while automated CD61‐immunoplatelet analysis can be run as selectable test. The aim of this study was to determine the platelet count performance of the three counting methods available on the instrument and to compare the results with those provided by Becton Dickinson FACSCalibur flow cytometer used as reference method. A lipid interference experiment was also performed. Linearity, carryover and precision were good, and satisfactory agreement with reference method was found for the impedance, optical and CD61‐immunoplatelet analysis, although this latter provided the closest results in comparison with flow cytometry. In the lipid interference experiment, a moderate inaccuracy of optical and immunoplatelet counts was observed starting from a very high lipid value.
Evaluation of three automated chromogenic FVIII kits for the diagnosis of mild discrepant haemophilia A
Authors: RODGERS S. E. , DUNCAN E. M. , SOBIERAJ‐TEAGUE M. , LLOYD J. V.
Published: February 2009

In some mild haemophilia A patients (discrepant phenotype), coagulation FVIII levels by one‐stage assay (FVIII‐1st) are more than double those by classical two‐stage coagulation assay (FVIII‐2st), and may fall within the normal range. Our aim was to assess automated two‐stage chromogenic FVIII assays (FVIII‐chr) for diagnosis of mild discrepant haemophilia A. Three chromogenic FVIII kits (Biophen, Coamatic and Dade‐Behring) were evaluated, using recommended and extended incubation times. Samples were tested from patients with discrepant haemophilia (n = 7) and equivalent mild haemophilia (agreement between FVIII‐1st and FVIII‐2st, n = 4). For equivalent haemophilia, FVIII‐chr were consistent with FVIII‐1st and FVIII‐2st for all kits at all incubation times. For discrepant haemophilia, using recommended incubation times, mean FVIII‐chr using Biophen reagents was 22 IU/dl (range 13–31), with Coamatic 26 (17–34) and with Dade‐Behring 41 (33–47), compared with 36 (27–44) for FVIII‐1st and 8 (6–9) for FVIII‐2st. FVIII‐chr decreased as incubation time was increased with Biophen and Coamatic, but decreased less with Dade‐Behring. FVIII‐chr using the Dade‐Behring kit gave similar results to FVIII‐1st and is not suitable for diagnosis of mild discrepant haemophilia A. FVIII‐chr by Biophen and Coamatic kits is suitable for diagnosis of these patients, especially with an extended incubation time.
Auer rod‐like intracytoplasmic inclusions in multiple myeloma. A case report and review of the literature
Authors: HÜTTER G. , NOWAK D. , BLAU I. W. , THIEL E.
Published: February 2009

Several intracytoplasmic morphological changes in the plasma cells of multiple myeloma have been described previously. However, Auer rod‐like inclusions are rarely found in these types of cells. Here, we report a case of multiple myeloma with Auer rod‐like, needle‐shaped intracytoplasmic inclusions in plasma cells. A review of all published cases revealed that this phenomenon is exclusively found in myeloma with kappa‐type paraprotein. The nature of these intracellular inclusions and the relationship to prognostic implications and concomitant illnesses are discussed.
Relationships between platelet counts, platelet volumes and reticulated platelets in patients with ITP: evidence for significant platelet count inaccuracies with conventional instrument methods
Authors: DIQUATTRO M. , GAGLIANO F. , CALABRÒ G. M. , TOMMASI M. , SCOTT C. S. , MANCUSO G. , PALMA B. , MENOZZI I.
Published: February 2009

The platelet count has a primary role in the diagnosis and treatment of idiopathic thrombocytopenic purpura (ITP). This study analysed the accuracy of ITP patient platelet counts determined by Abbott CD‐Sapphire (impedance/optical) and Bayer Advia 120 (optical) analyses, compared with a reference immunoplatelet method. Instrument platelet estimates showed broad equivalence in the higher range of observed values, but significant discrepancies against the immunoplatelet count were seen when platelet counts were <10 × 109/l. CD‐Sapphire mean platelet volume (MPV) results revealed increased (>12 fl) platelet volumes in eight of eight ITP patients with counts of <20 × 109/l compared with 6/6 and 5/13 patients with platelet counts of 20–50 and >50 × 109/l. In contrast, Bayer Advia MPV values showed no relationship with the platelet count. Increased reticulated platelets were associated with an increasing CD‐Sapphire MPV (R2 = 0.61) and a decreasing platelet count. High (>40%) reticulated platelet values were seen in 9/9 patients with immunoplatelet counts of <20 × 109/l compared with 0/19 patients with platelet counts above 20 × 109/l. There may be a need for caution in the interpretation of platelet counts in ITP patients obtained with conventional instrument methods, and therapeutic decisions should ideally be validated by reference immunoplatelet procedures.
Bestrophin expression and function in the human pancreatic duct cell line, CFPAC‐1
Authors: Marsey Laura L. , Winpenny John P.
Published: February 2009

Pancreatic duct epithelial cells (PDECs) have been shown to express calcium activated chloride channels (CaCCs) and there is evidence for their involvement in fluid secretion from these cells. The molecular identity of the CaCC in PDECs remains unknown. Recently, the bestrophin family of proteins have been proposed as a potential molecular candidate for CaCCs. Expression of bestrophins is strongly correlated with the function of CaCCs in a variety of tissues. In the present study, the expression of bestrophins has been investigated in the cystic fibrosis pancreatic duct cell line, CFPAC‐1. Iodide efflux analysis was used to characterise native CaCCs in CFPAC‐1 cell monolayers. Efflux was induced with the addition of UTP (100 μm, 10.2 ± 1.5 nmol min−1), which was blocked by the chloride channel blockers niflumic acid (81%) and DIDS (90%). The UTP‐stimulated iodide efflux was shown to be Ca2+ dependent and cAMP independent. RT‐PCR analysis of RNA isolated from CFPAC‐1 cells demonstrated positive identification of all four human bestrophin mRNAs. Western blot of CFPAC‐1 cell protein isolates with antibodies specific to human bestrophin 1 (hBest1) showed that hBest1 protein was expressed in this cell line. HBest1 was present on the cell surface, demonstrated using biotinylation and confocal imaging, as well as in the cytoplasm. SiRNA‐mediated silencing of hBest1 in CFPAC‐1 cells reduced the UTP‐stimulated iodide efflux by around 40%. This study provides evidence that the bestrophins are expressed in pancreatic duct cells and, more specifically, that hBest1 plays a role in the CaCCs found in these cells.
Carbohydrate sensing in the human mouth: effects on exercise performance and brain activity
Authors: Chambers E. S. , Bridge M. W. , Jones D. A.
Published: February 2009

Exercise studies have suggested that the presence of carbohydrate in the human mouth activates regions of the brain that can enhance exercise performance but direct evidence of such a mechanism is limited. The first aim of the present study was to observe how rinsing the mouth with solutions containing glucose and maltodextrin, disguised with artificial sweetener, would affect exercise performance. The second aim was to use functional magnetic resonance imaging (fMRI) to identify the brain regions activated by these substances. In Study 1A, eight endurance‐trained cyclists ( 60.8 ± 4.1 ml kg−1 min−1) completed a cycle time trial (total work = 914 ± 29 kJ) significantly faster when rinsing their mouths with a 6.4% glucose solution compared with a placebo containing saccharin (60.4 ± 3.7 and 61.6 ± 3.8 min, respectively, P= 0.007). The corresponding fMRI study (Study 1B) revealed that oral exposure to glucose activated reward‐related brain regions, including the anterior cingulate cortex and striatum, which were unresponsive to saccharin. In Study 2A, eight endurance‐trained cyclists ( 57.8 ± 3.2 ml kg−1 min−1) tested the effect of rinsing with a 6.4% maltodextrin solution on exercise performance, showing it to significantly reduce the time to complete the cycle time trial (total work = 837 ± 68 kJ) compared to an artificially sweetened placebo (62.6 ± 4.7 and 64.6 ± 4.9 min, respectively, P= 0.012). The second neuroimaging study (Study 2B) compared the cortical response to oral maltodextrin and glucose, revealing a similar pattern of brain activation in response to the two carbohydrate solutions, including areas of the insula/frontal operculum, orbitofrontal cortex and striatum. The results suggest that the improvement in exercise performance that is observed when carbohydrate is present in the mouth may be due to the activation of brain regions believed to be involved in reward and motor control. The findings also suggest that there may be a class of so far unidentified oral receptors that respond to carbohydrate independently of those for sweetness.
Muscle‐specific VEGF deficiency greatly reduces exercise endurance in mice
Authors: Olfert I. Mark , Howlett Richard A. , Tang Kechun , Dalton Nancy D. , Gu Yusu , Peterson Kirk L. , Wagner Peter D. , Breen Ellen C.
Published: February 2009

Vascular endothelial growth factor (VEGF) is required for vasculogenesis and angiogenesis during embryonic and early postnatal life. However the organ‐specific functional role of VEGF in adult life, particularly in skeletal muscle, is less clear. To explore this issue, we engineered skeletal muscle‐targeted VEGF deficient mice (mVEGF−/−) by crossbreeding mice that selectively express Cre recombinase in skeletal muscle under the control of the muscle creatine kinase promoter (MCKcre mice) with mice having a floxed VEGF gene (VEGFLoxP mice). We hypothesized that VEGF is necessary for regulating both cardiac and skeletal muscle capillarity, and that a reduced number of VEGF‐dependent muscle capillaries would limit aerobic exercise capacity. In adult mVEGF−/− mice, VEGF protein levels were reduced by 90 and 80% in skeletal muscle (gastrocnemius) and cardiac muscle, respectively, compared to control mice (P < 0.01). This was accompanied by a 48% (P < 0.05) and 39% (P < 0.05) decreases in the capillary‐to‐fibre ratio and capillary density, respectively, in the gastrocnemius and a 61% decrease in cardiac muscle capillary density (P < 0.05). Hindlimb muscle oxidative (citrate synthase, 21%; β‐HAD, 32%) and glycolytic (PFK, 18%) regulatory enzymes were also increased in mVEGF−/− mice. However, this limited adaptation to reduced muscle VEGF was insufficient to maintain aerobic exercise capacity, and maximal running speed and endurance running capacity were reduced by 34% and 81%, respectively, in mVEGF−/− mice compared to control mice (P < 0.05). Moreover, basal and dobutamine‐stimulated cardiac function, measured by transthoracic echocardiography and left ventricular micromanomtery, showed only a minimal reduction of contractility (peak +dP/dt) and relaxation (peak –dP/dt, τE). Collectively these data suggests adequate locomotor muscle capillary number is important for achieving full exercise capacity. Furthermore, VEGF is essential in regulating postnatal muscle capillarity, and that adult mice, deficient in cardiac and skeletal muscle VEGF, exhibit a major intolerance to aerobic exercise.
NADPH oxidase activity is necessary for acute intermittent hypoxia‐induced phrenic long‐term facilitation
Authors: MacFarlane P. M. , Satriotomo I. , Windelborn J. A. , Mitchell G. S.
Published: February 2009

Phrenic long‐term facilitation (pLTF) following acute intermittent hypoxia (AIH) is a form of spinal, serotonin‐dependent synaptic plasticity that requires reactive oxygen species (ROS) formation. We tested the hypothesis that spinal NADPH oxidase activity is a necessary source of ROS for pLTF. Sixty minutes post‐AIH (three 5‐min episodes of 11% O2, 5 min intervals), integrated phrenic and hypoglossal (XII) nerve burst amplitudes were increased from baseline, indicative of phrenic and XII LTF. Intrathecal injections (∼C4) of apocynin or diphenyleneiodonium chloride (DPI), two structurally and functionally distinct inhibitors of the NADPH oxidase complex, attenuated phrenic, but not XII, LTF. Immunoblots from soluble (cytosolic) and particulate (membrane) fractions of ventral C4 spinal segments revealed predominantly membrane localization of the NADPH oxidase catalytic subunit, gp91phox, whereas membrane and cytosolic expression were both observed for the regulatory subunits, p47phox and RAC1. Immunohistochemical analysis of fixed tissues revealed these same subunits in presumptive phrenic motoneurons of the C4 ventral horn, but not in neighbouring astrocytes or microglia. Collectively, these data demonstrate that NADPH oxidase subunits localized within presumptive phrenic motoneurons are a major source of ROS necessary for AIH‐induced pLTF. Thus, NADPH oxidase activity is a key regulator of spinal synaptic plasticity, and may be a useful pharmaceutical target in developing therapeutic strategies for respiratory insufficiency in patients with, for example, cervical spinal injury.
Nitric oxide generated by muscle corrects defects in hippocampal neurogenesis and neural differentiation caused by muscular dystrophy
Authors: Deng Bo , Glanzman David , Tidball James G.
Published: February 2009

Duchenne muscular dystrophy (DMD) results from null mutation of dystrophin, a membrane‐associated structural protein that is expressed in skeletal muscle. Dystrophin deficiency causes muscle membrane lesions, muscle degeneration and eventually death in afflicted individuals. However, dystrophin deficiency also causes cognitive defects that are difficult to relate to the loss of dystrophin. We assayed neurogenesis in the dentate gyrus (DG) in the mdx mouse model of DMD, using bromodeoxyuridine incorporation as a marker of proliferation and NeuN expression as a marker of differentiation. Our findings show that dystrophin mutation disrupts adult neurogenesis by promoting cell proliferation in the DG and suppressing neuronal differentiation. Because loss of dystrophin from muscle results in the secondary loss of neuronal nitric oxide synthase (nNOS), and NO is able to modulate neurogenesis, we assayed whether the genetic restoration of nNOS to mdx muscles corrected defects in adult, hippocampal neurogenesis. Assays of NO in the sera of active mice showed significant reductions in NO caused by the dystrophin mutation. However, over‐expression of nNOS in the muscles of mdx mice increased serum NO and normalized cell proliferation and neuronal differentiation in the DG. These findings indicate that muscle‐derived NO regulates adult neurogenesis in the brain and loss of muscle nNOS may underlie defects in the central nervous system in DMD.
Is there an optimum endurance polygenic profile?
Authors: Ruiz Jonatan R. , Gómez‐Gallego Félix , Santiago Catalina , González‐Freire Marta , Verde Zoraida , Foster Carl , Lucia Alejandro
Published: February 2009

We analysed seven genetic polymorphisms that are candidates to explain individual variations in human endurance phenotypic traits, at least in Caucasian people (ACE Ins/Del, ACTN3 Arg577Ter, AMPD1 Gln12Ter, CKMM 1170 bp/985 + 185 bp, HFE His63Asp, GDF‐8 Lys153Arg and PPARGC1A Gly482Ser) in 46 world‐class endurance athletes and 123 controls (all Spanish Caucasians). Using the model developed by Williams & Folland we determined (1) the ‘total genotype score’ (TGS, from the accumulated combination of the seven polymorphisms, with a maximum value of ‘100’ for the theoretically optimal polygenic score) in the non‐athlete (control) group, in the athlete group and in the total Spanish population, and (2) the probability for the occurrence of Spanish individuals with the ‘perfect’ polygenic endurance profile (i.e. TGS = 100). The probability of a Spanish individual possessing a theoretically optimal polygenic profile for up to the seven candidate genetic polymorphisms we studied was very small, i.e. ∼0.07% (or 1 in 1351 Spanish individuals). The mean TGS was higher in athletes (70.22 ± 15.58) than in controls (62.43 ± 11.45) and also higher than predicted for the total Spanish population (60.80 ± 12.1), suggesting an overall more ‘favourable’ polygenic profile in the athlete group. However, only three of the best Spanish endurance athletes (who are also amongst the best in the world) had the best possible score for up to six genes and none of them had the optimal profile. Other polymorphisms yet undiscovered as well as several factors independent of genetic endowment may explain why some individuals reach the upper end of the endurance performance continuum.
Menstrual cycle effects on sympathetic neural responses to upright tilt
Authors: Fu Qi , Okazaki Kazunobu , Shibata Shigeki , Shook Robin P. , VanGunday Tiffany B. , Galbreath M. Melyn , Reelick Miriam F. , Levine Benjamin D.
Published: February 2009

Young women are more susceptible to orthostatic intolerance than men, though the sex‐specific pathophysiology remains unknown. As blood pressure (BP) is regulated through the baroreflex mechanism, we tested the hypothesis that baroreflex control of muscle sympathetic nerve activity (MSNA) during orthostasis is impaired in women and can be affected by the menstrual cycle. MSNA and haemodynamics were measured supine and during a graded upright tilt (30 deg for 6 min, 60 deg for 45 min or till presyncope) in 11 young men and 11 women during the early follicular (EFP) and mid‐luteal phase (MLP) of the menstrual cycle. Sympathetic baroreflex sensitivity was quantified using the slope of the linear correlation between total activity and diastolic BP during spontaneous breathing. Baroreflex function was further assessed during a Valsalva manoeuvre (VM). Although MSNA burst frequency responses during tilting were similar between sexes and menstrual phases, increases in total activity were lower in women during EFP than MLP (P= 0.030), while total peripheral resistance and plasma noradrenaline were not similarly lower; upright total activity tended to be lower in women during EFP than men (P= 0.102). Sympathetic baroreflex sensitivity did not differ between sexes (P= 0.676) supine (−281 ± 46 (s.e.m.) units beat−1 mmHg−1 in men vs−252 ± 52 in EFP and −272 ± 40 in MLP in women), at 30 deg tilt (−648 ± 129 vs−611 ± 79 and −487 ± 94), and at 60 deg tilt (−792 ± 135 vs−831 ± 92 and −814 ± 142); this sensitivity was not affected by the menstrual cycle (P= 0.747). Similar sympathetic baroreflex sensitivity between sexes and phases was also observed during the VM. Cardiovagal baroreflex sensitivity assessed during decreasing BP (i.e. early phase II of the VM) was comparable between sexes, but it was greater in men than women during increasing BP (i.e. phase IV); the menstrual cycle had no influences on cardiovagal baroreflex sensitivity. We conclude that the menstrual cycle affects sympathetic neural responses but not sympathetic baroreflex sensitivity during orthostasis, though upright vasomotor sympathetic activity is not clearly different between men and women. Not only sympathetic but also cardiovagal baroreflex sensitivity is similar between sexes and menstrual phases during a hypotensive stimulus. However, cardiovagal baroreflex‐mediated bradycardia during a hypertensive stimulus is different between sexes but not affected by the menstrual cycle. Thus, other factors rather than sympathetic baroreflex control mechanisms contribute to sex differences in orthostatic tolerance in young humans.
Rats selectively bred for low aerobic capacity have reduced hepatic mitochondrial oxidative capacity and susceptibility to hepatic steatosis and injury
Authors: Thyfault John P. , Rector R. Scott , Uptergrove Grace M. , Borengasser Sarah J. , Morris E. Matthew , Wei Yongzhong , Laye Matt J. , Burant Charles F. , Qi Nathan R. , Ridenhour Suzanne E. , Koch Lauren G. , Britton Steve L. , Ibdah Jamal A.
Published: February 2009

Fatty liver has been linked to low aerobic fitness, but the mechanisms are unknown. We previously reported a novel model in which rats were artificially selected to be high capacity runners (HCR) and low capacity runners (LCR) that in a sedentary condition have robustly different intrinsic aerobic capacities. We utilized sedentary HCR/LCR rats (generation 17; max running distance equalled 1514 ± 91 vs. 200 ± 12 m for HCR and LCR, respectively) to investigate if low aerobic capacity is associated with reduced hepatic mitochondrial oxidative capacity and increased susceptibility to hepatic steatosis. At 25 weeks of age, LCR livers displayed reduced mitochondrial content (reduced citrate synthase activity and cytochrome c protein) and reduced oxidative capacity (complete palmitate oxidation in hepatic mitochondria (1.15 ± 0.13 vs. 2.48 ± 1.1 nm g−1 h, P < 0.0001) and increased peroxisomal activity (acyl CoA oxidase and catalase activity) compared to the HCR. The LCR livers also displayed a lipogenic phenotype with higher protein content of both sterol regulatory element binding protein‐1c and acetyl CoA carboxylase. These differences were associated with hepatic steatosis in the LCR including higher liver triglycerides (6.00 ± 0.71 vs. 4.20 ± 0.39 nmol g−1, P= 0.020 value), >2‐fold higher percentage of hepatocytes associated with lipid droplets (54.0 ± 9.2 vs. 22.0 ± 3.5%, P= 0.006), and increased hepatic lipid peroxidation compared to the HCR. Additionally, in rats aged to natural death, LCR livers had significantly greater hepatic injury (fibrosis and apoptosis). We provide novel evidence that selection for low intrinsic aerobic capacity causes reduced hepatic mitochondrial oxidative capacity that increases susceptibility to both hepatic steatosis and liver injury.
Molecular Genetics of
para
-Aminosalicylic Acid Resistance in Clinical Isolates and Spontaneous Mutants of
Mycobacterium tuberculosis
Authors: Mathys Vanessa , Wintjens René , Lefevre Philippe , Bertout Julie , Singhal Amit , Kiass Mehdi , Kurepina Natalia , Wang Xiao-Ming , Mathema Barun , Baulard Alain , Kreiswirth Barry N. , Bifani Pablo
Published: February 2009

The emergence of
Mycobacterium tuberculosis
resistant to first-line antibiotics has renewed interest in second-line antitubercular agents. Here, we aimed to extend our understanding of the mechanisms underlying para-aminosalicylic acid (PAS) resistance by analysis of six genes of the folate metabolic pathway and biosynthesis of thymine nucleotides (
thyA, dfrA, folC, folP1, folP2
, and
thyX
) and three
N
-acetyltransferase genes [
nhoA, aac(1)
, and
aac(2)
] among PAS-resistant clinical isolates and spontaneous mutants. Mutations in
thyA
were identified in only 37% of the clinical isolates and spontaneous mutants. Overall, 24 distinct mutations were identified in the
thyA
gene and 3 in the
dfrA
coding region. Based on structural bioinformatics techniques, the altered ThyA proteins were predicted to generate an unfolded or dysfunctional polypeptide. The MIC was determined by Bactec/Alert and dilution assay. Sixty-three percent of the PAS-resistant isolates had no mutations in the nine genes considered in this study, revealing that PAS resistance in
M. tuberculosis
involves mechanisms or targets other than those pertaining to the biosynthesis of thymine nucleotides. The alternative mechanism(s) or pathway(s) associated with PAS resistance appears to be PAS concentration dependent, in marked contrast to
thyA
-mutated PAS-resistant isolates.
Substrate-Induced Inactivation of the
Escherichia coli
AmiD
N
-Acetylmuramoyl-
l
-Alanine Amidase Highlights a New Strategy To Inhibit This Class of Enzyme
Authors: Pennartz Anne , Généreux Catherine , Parquet Claudine , Mengin-Lecreulx Dominique , Joris Bernard
Published: February 2009

In the eubacterial cell, the peptidoglycan is perpetually hydrolyzed throughout the cell cycle by different enzymes such as lytic transglycosylases, endopeptidases, and amidases. In
Escherichia coli
, four
N
-acetylmuramoyl-
l
-alanine amidases, AmiA, -B, -C, and -D, are present in the periplasm. AmiA, -B, and -C are soluble enzymes, whereas AmiD is a lipoprotein anchored in the outer membrane. To determine more precisely the specificity and the kinetic parameters of AmiD, we overproduced and purified the native His-tagged AmiD in the presence of detergent and a soluble truncated form of this enzyme by removing its signal peptide and the cysteine residue responsible for its lipidic anchorage. AmiD is a zinc metalloenzyme and is inactivated by a metal chelator such as EDTA. Native His-tagged and truncated AmiD hydrolyzes peptidoglycan fragments that have at least three amino acids in their peptide chains, and the presence of an anhydro function on the
N
-acetylmuramic acid is not essential for its activity. The soluble truncated AmiD exhibits a biphasic kinetic time course that can be explained by the inactivation of the enzyme by the substrate. This behavior highlights a new strategy to inhibit this class of enzymes.
Binding of Faropenem and Other β-Lactam Agents to Penicillin-Binding Proteins of Pneumococci with Various β-Lactam Susceptibilities
Authors: Kosowska-Shick Klaudia , McGhee Pamela , Appelbaum Peter C.
Published: February 2009

Faropenem demonstrated low MICs (≤1 μg/ml) for all penicillin-susceptible and nonsusceptible pneumococci and exhibited very strong abilities to bind to
Streptococcus pneumoniae
penicillin-binding proteins (PBPs), except for PBP2X. The lower faropenem affinity for PBP2X did not affect MICs for any strains tested, and only imipenem had lower MICs, with much lower binding affinities for all PBPs tested, than faropenem.
Transcription of Inflammatory Genes in the Lung after Infection with Community-Associated Methicillin-Resistant
Staphylococcus aureus
: a Role for Panton-Valentine Leukocidin?
Authors: Montgomery Christopher P. , Daum Robert S.
Published: February 2009

Necrotizing pneumonia caused by community-associated methicillin-resistant
Staphylococcus aureus
(CA-MRSA) isolates is increasingly common and frequently severe. The early inflammatory response in the lung after CA-MRSA infection remains largely undefined. Additionally, many workers have hypothesized that the Panton-Valentine leukocidin (PVL) is a key virulence determinant in CA-MRSA necrotizing pneumonia. We hypothesized that intratracheal inoculation of rats with a USA300 CA-MRSA isolate would result in early expression of genes involved in the immune response and that this would correlate with inflammation and tissue destruction characteristic of necrotizing pneumonia. In addition, we hypothesized that infection with a PVL deletion mutant would result in an attenuated early host response. Infection of rats with a sublethal inoculum of USA300 (strain LAC) resulted in rapid increased expression of most cytokine, chemokine, and inflammatory receptor gene transcripts studied, as assessed by quantitative real-time reverse transcriptase PCR (qRT-PCR). The increased gene transcription was followed by inflammation, increased bacterial survival in the lungs, and necrotizing pneumonia. Infection with strain LAC and infection with strain LAC Δ
pvl
(
lukSF-PV
deletion mutant) resulted in indistinguishable diseases, as assessed by mortality, in vivo bacterial recovery, and pulmonary pathology. Assessment of the transcription of inflammatory genes by qRT-PCR also revealed little difference after infection with LAC and after infection with LAC Δ
pvl
, either in animals that died or in animals that survived to 24 h after inoculation. We conclude that in a rat model of necrotizing pneumonia, there was an early, brisk inflammatory transcriptional response associated with neutrophil recruitment and tissue destruction. Deletion of
lukSF-PV
did not alter the early immune response to CA-MRSA in the lung.
Cholera Caused by
Vibrio cholerae
O1 Induces T-Cell Responses in the Circulation
Authors: Bhuiyan Taufiqur Rahman , Lundin Samuel B. , Khan Ashraful Islam , Lundgren Anna , Harris Jason B. , Calderwood Stephen B. , Qadri Firdausi
Published: February 2009

Considerable effort is being made to understand the acute and memory antibody responses in natural cholera infection, while rather less is known about the roles of cellular immune responses involving T and B lymphocytes. We studied responses in adult patients hospitalized with cholera caused by
Vibrio cholerae
O1. Peripheral blood mononuclear cells from patients (
n
= 15) were analyzed by flow cytometry after stimulation with
V. cholerae
O1 membrane protein (MP) or toxin-coregulated pilus antigen (TcpA). The gamma interferon (IFN-γ) and interleukin-13 (IL-13) responses in stimulated-lymphocyte supernatants were studied. The responses were compared with those of healthy controls (
n
= 10). Patients responded with increased frequencies of gut-homing CD4
+
T cells (CD4
+
β7
+
), gut-homing CD8
+
T cells (CD8
+
β7
+
), and gut-homing B cells (CD19
+
β7
+
) at the early and/or late convalescent stages compared to the acute stage. After stimulation with MP or TcpA, proliferation of CD4
+
and CD8
+
T cells was increased at the acute stage and/or early convalescent stage compared to healthy controls. Increased IL-13 and IFN-γ responses were observed after antigenic stimulation at the acute and convalescent stages compared to healthy controls. Thus, increases in the levels of gut-homing T and B cells, as well as involvement of CD8 and CD4 Th1-mediated (IFN-γ) and CD4 Th2-mediated (IL-13) cytokine responses, take place in acute dehydrating disease caused by
V. cholerae
O1. Further studies are needed to determine if such responses are also stimulated after immunization with oral cholera vaccines and if these responses play a role in protection following exposure to cholera.