bluuebunny/crossref_metadata_2025 huggingface dataset:
Isolation, Characterization, and In Situ Detection of a Novel Chemolithoautotrophic Sulfur-Oxidizing Bacterium in Wastewater Biofilms Growing under Microaerophilic Conditions
Authors: Ito Tsukasa , Sugita Kenichi , Okabe Satoshi
Published: May 2004

We successfully isolated a novel aerobic chemolithotrophic sulfur-oxidizing bacterium, designated strain SO07, from wastewater biofilms growing under microaerophilic conditions. For isolation, the use of elemental sulfur (S
0
), which is the most abundant sulfur pool in the wastewater biofilms, as the electron donor was an effective measure to establish an enrichment culture of strain SO07 and further isolation. 16S rRNA gene sequence analysis revealed that newly isolated strain SO07 was affiliated with members of the genus
Halothiobacillus
, but it was only distantly related to previously isolated species (89% identity). Strain SO07 oxidized elemental sulfur, thiosulfate, and sulfide to sulfate under oxic conditions. Strain SO07 could not grow on nitrate. Organic carbons, including acetate, propionate, and formate, could not serve as carbon and energy sources. Unlike other aerobic sulfur-oxidizing bacteria, this bacterium was sensitive to NaCl; growth in medium containing more than 150 mM was negligible. In situ hybridization combined with confocal laser scanning microscopy revealed that a number of rod-shaped cells hybridized with a probe specific for strain SO07 were mainly present in the oxic biofilm strata (ca. 0 to 100 μm) and that they often coexisted with sulfate-reducing bacteria in this zone. These results demonstrated that strain SO07 was one of the important sulfur-oxidizing populations involved in the sulfur cycle occurring in the wastewater biofilm and was primarily responsible for the oxidation of H
2
S and S
0
to SO
4
2−
under oxic conditions.
Functional Expression of the Ectoine Hydroxylase Gene (
thpD
) from
Streptomyces chrysomallus
in
Halomonas elongata
Authors: Prabhu Julia , Schauwecker Florian , Grammel Nicolas , Keller Ullrich , Bernhard Michael
Published: May 2004

The formation of hydroxyectoine in the industrial ectoine producer
Halomonas elongata
was improved by the heterologous expression of the ectoine hydroxylase gene,
thpD
, from
Streptomyces chrysomallus.
The efficient conversion of ectoine to hydroxyectoine was achieved by the concerted regulation of
thpD
by the
H. elongata ectA
promoter.
Antimicrobial Resistance of
Enterococcus
Species Isolated from Produce
Authors: Johnston Lynette M. , Jaykus Lee-Ann
Published: May 2004

The purpose of this study was to characterize the antibiotic resistance profiles of
Enterococcus
species isolated from fresh produce harvested in the southwestern United States. Among the 185
Enterococcus
isolates obtained, 97 (52%) were
Enterococcus faecium
, 38 (21%) were
Enterococcus faecalis
, and 50 (27%) were other
Enterococcus
species. Of human clinical importance,
E. faecium
strains had a much higher prevalence of resistance to ciprofloxacin, tetracycline, and nitrofurantoin than
E. faecalis. E. faecalis
strains had a low prevalence of resistance to antibiotics used to treat
E. faecalis
infections of both clinical and of agricultural relevance, excluding its intrinsic resistance patterns. Thirty-four percent of the isolates had multiple-drug-resistance patterns, excluding intrinsic resistance. Data on the prevalence and types of antibiotic resistance in
Enterococcus
species isolated from fresh produce may be used to describe baseline antibiotic susceptibility profiles associated with
Enterococcus
spp. isolated from the environment. The data collected may also help elucidate the role of foods in the transmission of antibiotic-resistant strains to human populations.
pmoA
-Based Analysis of Methanotrophs in a Littoral Lake Sediment Reveals a Diverse and Stable Community in a Dynamic Environment
Authors: Pester Michael , Friedrich Michael W. , Schink Bernhard , Brune Andreas
Published: May 2004

Diversity and community structure of aerobic methane-oxidizing bacteria in the littoral sediment of Lake Constance was investigated by cloning analysis and terminal restriction fragment length polymorphism (T-RFLP) fingerprinting of the
pmoA
gene. Phylogenetic analysis revealed a high diversity of type I and type II methanotrophs in the oxygenated uppermost centimeter of the sediment. T-RFLP profiles indicated a high similarity between the active methanotrophic community in the oxic layer and the inactive community in an anoxic sediment layer at a 10-cm depth. There were also no major changes in community structure between littoral sediment cores sampled in summer and winter. By contrast, the fingerprint patterns showed substantial differences between the methanotrophic communities of littoral and profundal sediments.
Plastic Encapsulation of Stabilized
Escherichia coli
and
Pseudomonas putida
Authors: Manzanera M. , Vilchez S. , Tunnacliffe A.
Published: May 2004

Escherichia coli
and
Pseudomonas putida
dried in hydroxyectoine or trehalose are shown to be highly resistant to the organic solvents chloroform and acetone, and consequently, they can be encapsulated in a viable form in solid plastic materials. Bacteria are recovered by rehydration after physical disruption of the plastic.
P. putida
incorporated into a plastic coating of maize seeds was shown to colonize roots efficiently after germination.
Transformation of Folate-Consuming
Lactobacillus gasseri
into a Folate Producer
Authors: Wegkamp Arno , Starrenburg Marjo , de Vos Willem M. , Hugenholtz Jeroen , Sybesma Wilbert
Published: May 2004

Five genes essential for folate biosynthesis in
Lactococcus lactis
were cloned on a broad-host-range lactococcal vector and were transferred to the folate auxotroph
Lactobacillus gasseri
. As a result
L. gasseri
changed from a folate consumer to a folate producer. This principle can be used to increase folate levels in many fermented food products.
Characterization of
Nocardia asteroides
Isolates from Different Ecological Habitats on the Basis of Repetitive Extragenic Palindromic-PCR Fingerprinting
Authors: Yamamura Hideki , Hayakawa Masayuki , Nakagawa Youji , Iimura Yuzuru
Published: May 2004

Thirteen isolates of
Nocardia asteroides
from both soils and aquatic samples (lake and moat sediments, as well as scum from activated sludge), together with a type strain and two known clinical isolates of this species, were characterized by repetitive extragenic palindromic-PCR fingerprinting with the BOX-A1R primer. The resulting DNA fingerprint patterns proved to be strain specific, and cluster analysis distinguished the soil isolates, the aquatic isolates, and the known strains as being in separate groups.
Salinity Decreases Nitrite Reductase Gene Diversity in Denitrifying Bacteria of Wastewater Treatment Systems
Authors: Yoshie Sachiko , Noda Naohiro , Tsuneda Satoshi , Hirata Akira , Inamori Yuhei
Published: May 2004

Investigation of the diversity of
nirK
and
nirS
in denitrifying bacteria revealed that salinity decreased the diversity in a nitrate-containing saline wastewater treatment system. The predominant
nirS
clone was related to
nirS
derived from marine bacteria, and the predominant
nirK
clone was related to
nirK
of the genus
Alcaligenes
.
Bacteriophage-Based Genetic System for Selection of Nonsplicing Inteins
Authors: Cann Isaac K. O. , Amaya Kensey R. , Southworth Maurice W. , Perler Francine B.
Published: May 2004

A genetic selection system that detects splicing and nonsplicing activities of inteins was developed based on the ability to rescue a T4 phage strain with a conditionally inactive DNA polymerase. This phage defect can be complemented by expression of plasmid-encoded phage RB69 DNA polymerase. Insertion of an intein gene into the active site of the RB69 DNA polymerase gene renders polymerase activity and phage viability dependent on protein splicing. The effectiveness of the system was tested by screening for thermosensitive splicing mutants. Development of genetic systems with the potential of identifying protein splicing inhibitors is a first step towards controlling proliferation of pathogenic microbes harboring inteins in essential proteins.
Degradation of Polycyclic Aromatic Hydrocarbons by a Newly Discovered Enteric Bacterium,
Leclercia adecarboxylata
Authors: Sarma Priyangshu Manab , Bhattacharya Dhruva , Krishnan S. , Lal Banwari
Published: May 2004

A bacterial strain, PS4040, capable of degrading polycyclic aromatic hydrocarbons for use as the sole carbon source was isolated from oily-sludge-contaminated soil. The 16S rRNA gene showed 98.8% homology to that of
Leclercia adecarboxylata
. Comparative molecular typing with the clinical strain of
L. adecarboxylata
revealed that there were few comigrating and few distinct amplimers among them.
Albusin B, a Bacteriocin from the Ruminal Bacterium
Ruminococcus albus
7 That Inhibits Growth of
Ruminococcus flavefaciens
Authors: Chen Junqin , Stevenson David M. , Weimer Paul J.
Published: May 2004

An ∼32-kDa protein (albusin B) that inhibited growth of
Ruminococcus flavefaciens
FD-1 was isolated from culture supernatants of
Ruminococcus albus
7. Traditional cloning and gene-walking PCR techniques revealed an open reading frame (
albB
) encoding a protein with a predicted molecular mass of 32,168 Da. A BLAST search revealed two homologs of AlbB from the unfinished genome of
R. albus
8 and moderate similarity to LlpA, a recently described 30-kDa bacteriocin from
Pseudomonas
sp. strain BW11M1.
Multiplex PCR with 16S rRNA Gene-Targeted Primers of
Bifidobacterium
spp. To Identify Sources of Fecal Pollution
Authors: Bonjoch X. , Ballesté E. , Blanch A. R.
Published: May 2004

Bifidobacteria are one of the most common bacterial types found in the intestines of humans and other animals and may be used as indicators of human fecal pollution. The presence of nine human-related
Bifidobacterium
species was analyzed in human and animal wastewater samples of different origins by using species-specific primers based on 16S rRNA sequences. Only
B. adolescentis
and
B. dentium
were found exclusively in human sewage. A multiplex PCR approach with strain-specific primers was developed. The method showed a sensitivity threshold of 10 cells/ml. This new molecular method could provide useful information for the characterization of fecal pollution sources.
Osmotic Stress Leads to Decreased Intracellular pH of
Listeria monocytogenes
as Determined by Fluorescence Ratio-Imaging Microscopy
Authors: Fang Weihuan , Siegumfeldt Henrik , Budde Birgitte Bjørn , Jakobsen Mogens
Published: May 2004

Intracellular pH (pH
i
) of
Listeria monocytogenes
was determined after exposure to NaCl or sorbitol in liquid and solid media (agar). Both compounds decreased pH
i
, and recovery on solid medium was impaired compared to that in liquid medium.
N
,
N
′-dicyclohexylcarbodiimide abolished pH
i
recovery, and lowering a
w
with glycerol showed no effect on pH
i
.
Specific PCR Assay for a Tannin-Tolerant
Selenomonas ruminantium
Isolate, Derived from Helicase Coding Sequences
Authors: Bishop Richard , Obura Moses , Odongo David , Odenyo Agnes
Published: May 2004

Sequences from a tannin-tolerant
Selenomonas ruminantium
isolate (EAT2) that hydrolyzes gallic acid were identified. Two exhibited identity to helicases with a wide phylogenetic distribution. PCR amplification by using primers from one helicase gene detected 2,000 to 5,000 EAT2 genome equivalents but did not amplify total gastrointestinal microbial DNA of nine other ungulate species.
Multiplex PCR Assay for Detection of Bacterial Pathogens Associated with Warm-Water Streptococcosis in Fish
Authors: Mata A. I. , Gibello A. , Casamayor A. , Blanco M. M. , Domínguez L. , Fernández-Garayzábal J. F.
Published: May 2004

A multiplex PCR-based method was designed for the simultaneous detection of the main pathogens involved in warm-water streptococcosis in fish (
Streptococcus iniae
,
Streptococcus difficilis
,
Streptococcus parauberis
, and
Lactococcus garvieae
). Each of the four pairs of oligonucleotide primers exclusively amplified the targeted gene of the specific microorganism. The sensitivity of the multiplex PCR using purified DNA was 25 pg for
S. iniae
, 12.5 pg for
S. difficilis
, 50 pg for
S. parauberis
, and 30 pg for
L. garvieae
. The multiplex PCR assay was useful for the specific detection of the four species of bacteria not only in pure culture but also in inoculated fish tissue homogenates and naturally infected fish. Therefore, this method could be a useful alternative to the culture-based method for the routine diagnosis of warm-water streptococcal infections in fish.
Detection Rate and Antigenic Specificities of Antineutrophil Cytoplasmic Antibodies in Chinese Patients with Clinically Suspected Vasculitis
Authors: Xin Gang , Zhao Ming-hui , Wang Hai-yan
Published: May 2004

The detection rate of antineutrophil cytoplasmic antibodies (ANCA) in Chinese patients with clinically suspected small vessel vasculitis was investigated, and their antigen specificity and demographic features were analyzed. A number of sera (
n
= 5,604) sent to our referral laboratory for ANCA screening were tested by indirect immunofluorescence (IIF), enzyme-linked immunosorbent assays (ELISAs) for myeloperoxidase (MPO)- and proteinase 3 (PR3)-ANCA. Then the IIF-ANCA-positive sera that were negative for MPO- and PR3-ANCA were further tested by antigen-specific ELISA by using other five highly purified known ANCA antigens as solid-phase ligands. The known antigens included bactericidal/permeability-increasing protein (BPI), human leukocyte elastase (HLE), lactoferrin, cathepsin G, and azurocidins. Of the 5,604 sera, 267 (4.76%) sera were IIF-ANCA positive and 390 (7%) were antinuclear antibody (ANA) positive in the IIF assay. Of the IIF-positive samples, 213 were anti-MPO positive, 32 were anti-PR3 positive, and five cases were positive for both. Of the 48 sera positive for IIF-ANCA but negative for MPO- and PR3-ANCA, 13 sera (27%) recognized other target antigens, 7 sera recognized BPI, 5 recognized HLE, 1 recognize cathepsin G, and 1 recognized azurocidin. None of the sera recognized lactoferrin, and one serum sample recognized both BPI and HLE. The majority of ANCA-positive patients presented in summer or winter. There was no difference in gender (male/female ratio, 1:1.12) in ANCA-positive patients with a mean age of 53.1 years. The male/female ratio was 1.17:1 for patients over 60 years of age; however, it was 1:4 for patients under 20 years of age. We conclude that ANCA-related diseases are not rare in China, and the major antigens are MPO and PR3. When the IIF technique is used to detect ANCA, ANA should be carefully distinguished.
Comparison of Different Criteria for Interpretation of Immunoglobulin G Immunoblotting Results for Diagnosis of
Helicobacter pylori
Infection
Authors: Lepper Philipp M. , Möricke Angelika , Vogt Konstanze , Bode Günter , Trautmann Matthias
Published: May 2004

Gastric infection with
Helicobacter pylori
is one of the most common chronic infections in humans, causing substantial morbidity and mortality. The diagnosis of
H. pylori
infection usually involves upper endoscopy with biopsy since the only noninvasive method of comparable accuracy, the [
13
C]urea breath test, requires technical equipment that is not available in most gastroenterological units. Serological methods for detection of
H. pylori
infection have reached sufficient accuracy to be used as screening tests before endoscopy or for seroepidemiological surveys. In the present study we evaluated different interpretation criteria for use with immunoglobulin G immunoblotting for the diagnosis of
H. pylori
infection. We applied five different sets of interpretation criteria, four of which had been published previously, to the Western blot results of 294 patients with different gastrointestinal symptoms. Since it is known that less than 2% of patients who are infected with
H. pylori
fail to seroconvert, an optimally sensitive Western blotting system should be able to detect approximately 98% of active infections. When the different criteria were applied to our patient population, it became apparent that the abilities of the systems to detect active
H. pylori
infection were quite varied. The results for the sensitivity and specificity, according to the different applied criteria, ranged from 62.8 to 95.9% and from 85.7 to 100.0%, respectively. Positive predictive values and negative predictive values, according to the published criteria, ranged from 97.2 to 100.0% and from 37.7 to 82.4%, respectively. Recommendations for the optimal use of the different interpretation criteria are discussed.
Enhanced Surfactant Protein and Defensin mRNA Levels and Reduced Viral Replication during Parainfluenza Virus Type 3 Pneumonia in Neonatal Lambs
Authors: Grubor Branka , Gallup Jack M. , Meyerholz David K. , Crouch Erika C. , Evans Richard B. , Brogden Kim A. , Lehmkuhl Howard D. , Ackermann Mark R.
Published: May 2004

Defensins and surfactant protein A (SP-A) and SP-D are antimicrobial components of the pulmonary innate immune system. The purpose of this study was to determine the extent to which parainfluenza type 3 virus infection in neonatal lambs alters expression of sheep beta-defensin 1 (SBD-1), SP-A, and SP-D, all of which are constitutively transcribed by respiratory epithelia. Parainfluenza type 3 viral antigen was detected by immunohistochemistry (IHC) in the bronchioles of all infected lambs 3 days postinoculation and at diminished levels 6 days postinoculation, but it was absent 17 days postinoculation. At all times postinoculation, lung homogenates from parainfluenza type 3 virus-inoculated animals had increased SBD-1, SP-A, and SP-D mRNA levels as detected by fluorogenic real-time reverse transcriptase PCR. Protein levels of SP-A in lung homogenates detected by quantitative-competitive enzyme-linked immunosorbent assay and protein antigen of SP-A detected by IHC were not altered. These studies demonstrate that parainfluenza type 3 virus infection results in enhanced expression of constitutively transcribed innate immune factors expressed by respiratory epithelia and that this increased expression occurs concurrently with decreased viral replication.
Caspase Inhibitors Attenuate Superantigen-Induced Inflammatory Cytokines, Chemokines, and T-Cell Proliferation
Authors: Krakauer Teresa
Published: May 2004

Proinflammatory cytokines mediate the toxic effect of superantigenic staphylococcal exotoxins (SE). A pan-caspase inhibitor suppressed SE-stimulated T-cell proliferation and the production of cytokines and chemokines by human peripheral blood mononuclear cells. These data suggest that caspase inhibitors may represent a novel therapeutic modality for treating SE-induced toxic shock.
SPECTROSCOPY OF Bc MESON IN A SEMI-RELATIVISTIC QUARK MODEL USING THE SHIFTED LARGE-N EXPANSION METHOD
Authors: IKHDAIR SAMEER M. , SEVER RAMAZAN
Published: May 2004

We calculate the [Formula: see text] mass spectrum, the splitting values and some other properties in the framework of the semirelativistic equation by applying the shifted large-N expansion technique. We use seven different central potentials together with an improved QCD-motivated interquark potentials calculated to two loops in the modified minimal-subtraction [Formula: see text] scheme. The parameters of these potentials are fitted to generate the semirelativistic bound states of [Formula: see text] quarkonium system in close conformity with the experimental and the present available calculated center-of-gravity (c.o.g.) data. Calculations of the energy bound states are carried out up to third order. Our results are in excellent fit with the results of the other works.
COLLECTIVE STRING FIELD THEORY OF MATRIX MODELS IN THE BMN LIMIT
Authors: DE MELLO KOCH ROBERT , JEVICKI ANTAL , RODRIGUES JOÃO P.
Published: May 2004

We develop a systematic procedure for deriving canonical string field theory from large N matrix models in the Berenstein–Maldacena–Nastase limit. The approach, based on collective field theory, provides a generalization of the standard string field theory.
DUAL PROJECTION IN A TWO-DIMENSIONAL CURVED EXPANDING UNIVERSE
Authors: ABREU EVERTON M. C.
Published: May 2004

It is a well-known result that the scalar field is composed of two chiral particles (Floreanini–Jackiw particles) of opposite chiralities. Also, that a Siegel particle spectrum is formed by a nonmover field (a Hull's noton) and a FJ particle. In this work, we show that in a scalar field spectrum, in a curved expanding universe scenario, we find a different result, two dynamical chiral fields.
PROTON DECAY, FERMION MASSES AND TEXTURE FROM EXTRA DIMENSIONS IN SUSY GUT's
Authors: KAKIZAKI MITSURU , YAMAGUCHI MASAHIRO
Published: May 2004

In supersymmetry, there are gauge invariant dimension 5 proton decay operators which must be suppressed by a mass scale much larger than the Planck mass. It is natural to expect that this suppression should be explained by a mechanism that explains the hierarchical structure of the fermion mass matrices. We apply this argument to the case where wave functions of chiral multiplets are localized under a kink background along an extra spatial dimension and the Yukawa couplings as well as the coefficients of the proton decay operators are determined by the overlap of the relevant wave functions. A configuration is found in the context of SU(5) supersymmetric grand unified theory that yields realistic values of quark masses, mixing angles, CP phase and charged lepton masses and sufficiently small genuine dimension 5 proton decay operators. Inclusion of SU(5) breaking effects is essential in order to obtain non-vanishing CP phase as well as correct lepton masses. The resulting mass matrix has a texture structure in which texture zeros are a consequence of extremely small overlap of the wave functions. Our approach requires explicit breaking of supersymmetry in the extra dimension, which can be realized in (de)constructing extra dimension.
A NOTE ON GAUSSIAN INTEGRALS OVER PARA-GRASSMANN VARIABLES
Authors: CUGLIANDOLO LETICIA F. , LOZANO G. S. , MORENO E. F. , SCHAPOSNIK F. A.
Published: May 2004

We discuss the generalization of the connection between the determinant of an operator entering a quadratic form and the associated Gaussian path-integral valid for Grassmann variables to the para-Grassmann case [θp+1=0 with p=1(p>1) for Grassmann (para-Grassmann) variables]. We show that the q-deformed commutation relations of the para-Grassmann variables lead naturally to consider q-deformed quadratic forms related to multiparametric deformations of GL (n) and their corresponding q-determinants. We suggest a possible application to the study of disordered systems.
SUPERSTRUCTURE FORMATION AND X-RAY PHOTOEMISSION PROPERTIES OF THE TlTiOPO4 SURFACE
Authors: ATUCHIN V. V. , POKROVSKY L. D. , KESLER V. G. , YU. MAKLAKOVA N. , VORONKOVA V. I. , YANOVSKII V. K.
Published: May 2004

X-ray photoemission spectroscopy (XPS) measurements have been executed for TlTiOPO 4 to elucidate the general features in the electronic structure of the KTiOPO 4 family compounds. The peculiarities of the valence band structure have been discussed for the crystals. The persistence of core level binding energy differences O 1s–P 2p and O 1s–Ti 2p 3/2 has been detected in TlTiOPO 4 and KTiOPO 4, which relates well with the constancy of averaged P – O and Ti – O chemical bond lengths in this crystal family. The superstructure ordering of the TlTiOPO 4 surface subjected to polishing and annealing has been detected by reflectance high energy electron diffraction (RHEED). From comparison of surface crystallographic properties of TlTiOPO 4 and KTiOPO 4, the most typical superstructure indices have been revealed.
TWO DIRECT METHODS FOR OBTAINING SOLUTIONS TO VARIATIONAL PROBLEMS
Authors: CARLSON DEAN A.
Published: May 2004

In this paper we compare two methods for obtaining solutions for free problems in the Calculus of Variations. The first appears at least as early as 1935 and is due to Carathéodory while the second, due to Leitmann, first appears in 1967. Both methods introduce the notion of equivalent variational problems. Using either approach, an auxiliary problem is obtained for which the solution is more easily obtained. We compare both approaches by using each to solve the same class of examples. We conclude our discussion by unifying the two approaches into one and illustrating this new method's potential through the use of an elementary example.
AN EXTENSION OF LEITMANN'S DIRECT METHOD TO INEQUALITY CONSTRAINTS
Authors: CARTIGNY PIERRE , DEISSENBERG CHRISTOPHE
Published: May 2004

George Leitmann's direct method permits to find the global minimum of a variational problem by use of a change of variables. In this article, we extend this method to a class of scalar, inequality constrained problems. An application example is given.
DIFFERENTIAL GAMES WITH SEMI-UNCERTAIN DYNAMICS
Authors: CLEMHOUT SIMONE , WAN HENRY
Published: May 2004

We introduce competitive markets into an extension of the differential game model. The non-uniqueness of market-clearing prices implies the constant presence of uninsurable risks for the players. In a class of examples, some characteristics of the system as well as part of the players' rational response remain to be perfectly predictable. This offers a distinct view regarding the recent financial crises and new insight concerning institutional reform.
The 1.4-MDa apoptosome is a critical intermediate in apoptosome maturation
Authors: Beem Elaine , Holliday L. Shannon , Segal Mark S.
Published: May 2004

Previously, we demonstrated that both 150 mM KCl and alkaline pH inhibit cytochrome c-mediated activation of procaspase-3 in a unique manner. To determine the mechanism of inhibition, we analyzed the effect of KCl and alkaline pH on the formation of apoptosomes (a large complex consisting of cytochrome c, Apaf-1, and procaspase-9/caspase-9) in vitro. Our results suggest that an initial ∼700-kDa apoptosome matures through a 1.4-MDa intermediate before a ∼700-kDa apoptosome is reformed and procaspase-3 is activated. We further demonstrate that 150 mM KCl interferes with the conversion of the initial ∼700-kDa apoptosome to the 1.4-MDa intermediate, while alkaline pH “traps” the apoptosome in the 1.4-MDa intermediate. Analysis of the cleaved state of procaspase-9 and procaspase-3 suggests that the 1.4-MDa intermediate may be required for cleavage of procaspase-9. Consistent with these results, in vivo data suggest that blocking acidification during the induction of apoptosis inhibits activation of procaspase-3. On the basis of these results, we propose a model of apoptosome maturation.
Neutral endopeptidase 24.11 is important for the degradation of both endogenous and exogenous glucagon in anesthetized pigs
Authors: Trebbien Ramona , Klarskov Letty , Olesen Mette , Holst Jens J. , Carr Richard D. , Deacon Carolyn F.
Published: May 2004

Glucagon has a short plasma t1/2in vivo, with renal extraction playing a major role in its elimination. Glucagon is degraded by neutral endopeptidase (NEP) 24.11 in vitro, but the physiological relevance of NEP 24.11 in glucagon metabolism is unknown. Therefore, the influence of candoxatril, a selective NEP inhibitor, on plasma levels of endogenous and exogenous glucagon was examined in anesthetized pigs. Candoxatril increased endogenous glucagon concentrations, from 6.3 ± 2.5 to 20.7 ± 6.3 pmol/l [COOH-terminal (C)-RIA, P < 0.05]. During glucagon infusion, candoxatril increased the t1/2determined by C-RIA (from 3.0 ± 0.5 to 17.0 ± 2.5 min, P < 0.005) and midregion (M)-RIA (2.8 ± 0.5 to 17.0 ± 3.0 min, P < 0.01) and reduced metabolic clearance rates (MCR; 19.1 ± 3.2 to 9.4 ± 2.0 ml·kg−1·min−1, P < 0.02, C-RIA; 19.2 ± 4.8 to 9.0 ± 2.3 ml·kg−1·min−1, P < 0.05, M-RIA). However, neither t1/2nor MCR determined by NH2-terminal (N)-RIA were significantly affected ( t1/2, 2.7 ± 0.4 to 4.5 ± 1.6 min; MCR, 30.3 ± 6.4 to 28.5 ± 9.0 ml·kg−1·min−1), suggesting that candoxatril had no effect on NH2-terminal degradation but leads to the accumulation of NH2-terminally truncated forms of glucagon. Determination of arteriovenous glucagon concentration differences revealed that renal glucagon extraction was reduced (but not eliminated) by candoxatril (from 40.4 ± 3.8 to 18.6 ± 4.1%, P < 0.02, C-RIA; 29.2 ± 3.1 to 14.7 ± 2.2%, P < 0.02, M-RIA; 26.5 ± 4.0 to 19.7 ± 3.5%, P < 0.06, N-RIA). Femoral extraction was reduced by candoxatril when determined by C-RIA (from 22.7 ± 2.4 to 8.0 ± 5.1%, P < 0.05) but was not changed significantly when determined using M- or N-RIAs (10.0 ± 2.8 to 4.7 ± 3.7%, M-RIA; 10.5 ± 2.5 to 7.8 ± 4.2%, N-RIA). This study provides evidence that NEP 24.11 is an important mediator of the degradation of both endogenous and exogenous glucagon in vivo.
Low myo-inositol and high glutamine levels in brain are associated with neuropsychological deterioration after induced hyperammonemia
Authors: Shawcross D. L. , Balata S. , Olde Damink S. W. M. , Hayes P. C. , Wardlaw J. , Marshall I. , Deutz N. E. P. , Williams R. , Jalan R.
Published: May 2004

The neuropsychological effect of hyperammonemia is variable. This study tests the hypothesis that the effect of ammonia on the neuropsychological function in patients with cirrhosis is determined by the ability of the brain to buffer ammonia-induced increase in glutamine within the astrocyte by losing osmolytes like myo-inositol (mI) and not by the magnitude of the induced hyperammonemia. Fourteen cirrhotic patients with no evidence of overt hepatic encephalopathy were given a 75-g amino acid (aa) solution mimicking the hemoglobin molecule to induce hyperammonemia. Measurement of a battery of neuropsychological function tests including immediate memory, ammonia, aa, and short-echo time proton magnetic resonance spectroscopy were performed before and 4 h after administration of the aa solution. Eight patients showed deterioration in the Immediate Memory Test at 4 h. Demographic factors, severity of liver disease, change in plasma ammonia, and aa profiles after the aa solution were similar in those that showed a deterioration compared with those who did not. In patients who showed deterioration in the memory test, the mI-to-creatine ratio (mI/Cr) was significantly lower at baseline than those that did not deteriorate. In contrast, the glutamate/glutamine-to-Cr ratio was significantly greater in the patients that deteriorated. The observation that deterioration in the memory test scores was greater in those with lower mI/Cr supports the hypothesis that the neuropsychological effects of induced hyperammonemia is determined by the capacity of the brain to handle ammonia-induced increase in glutamine.
High-resolution imaging reveals a limit in spatial resolution of blood flow measurements by microspheres
Authors: Decking Ulrich K. M. , Pai Vinay M. , Bennett Eric , Taylor Joni L. , Fingas Christian D. , Zanger Klaus , Wen Han , Balaban Robert S.
Published: May 2004

Density of 15-μm microspheres after left atrial application is the standard measure of regional perfusion. In the heart, substantial differences in microsphere density are seen at spatial resolutions <5 ml, implying perfusion heterogeneity. Microsphere deposition imaging permits a superior evaluation of the distribution pattern. Therefore, fluorescent microspheres (FMS) were applied, FMS deposition in the canine heart was imaged by epifluorescence microscopy in vitro, and the patterns were observed compared with MR images of iron oxide microspheres (IMS) obtained in vivo and in vitro. FMS deposition in myocardial slices revealed the following: 1) a nonrandom distribution, with sequentially applied FMS of different color stacked within the same vessel, 2) general FMS clustering, and 3) rather large areas devoid of FMS ( n = 3). This pattern was also seen in reconstructed three-dimensional images (<1 nl resolution) of FMS distribution ( n = 4). Surprisingly, the deposition pattern of sequentially applied FMS remained virtually identical over 3 days. Augmenting flow by intracoronary adenosine (>2 μM) enhanced local microsphere density, but did not alter the deposition pattern ( n = 3). The nonrandom, temporally stable pattern was quantitatively confirmed by a three-dimensional intermicrosphere distance analysis of sequentially applied FMS. T2-weighted short-axis MR images (2-μl resolution) of IMS revealed similar patterns in vivo and in vitro ( n = 6), as seen with FMS. The observed temporally stable microsphere patterns are not consistent with the notion that microsphere deposition is solely governed by blood flow. We propose that at high spatial resolution (<2 μl) structural aspects of the vascular network dominate microsphere distribution, resulting in the organized patterns observed.
Differential expression of α2D-adrenoceptor and eNOS in aortas from early and later stages of diabetes in Goto-Kakizaki rats
Authors: Kobayashi Tsuneo , Matsumoto Takayuki , Ooishi Kazuyuki , Kamata Katsuo
Published: May 2004

The aim of the present study was to compare vascular dysfunction between the early (12 wk old) and later (36 wk old) stages of spontaneous diabetes in Goto-Kakizaki (GK) rats. We also evaluated the aortic expression of the α2D-adrenoceptor and endothelial nitric oxide synthase (eNOS). Vascular reactivity was assessed in thoracic aortas from age-matched control rats and 12- and 36-wk GK rats. Using RT-PCR and immunoblots, we also examined the changes in expression of the α2D-adrenoceptor and eNOS. In aortas from GK rats (vs. those from age-matched control rats): 1) the relaxation response to ACh was enhanced at 12 wk but decreased at 36 wk; 2) the relaxation response to sodium nitroprusside was decreased at both 12 and 36 wk, 3) norepinephrine (NE)-induced contractility was decreased at 12 wk but not at 36 wk, 4) the expressions of α1B- and α1D-adrenoceptors were unaffected, whereas those of α2D-adrenoceptor and eNOS mRNAs were increased at both 12 and 36 wk; and 5) NE- and ACh-stimulated NOx (nitrite and nitrate) levels were increased at 12 wk, although at 36 wk ACh-stimulated NOx was lower, whereas NE-stimulated NOx showed no change. These results clearly demonstrate that enhanced ACh-induced relaxation and impaired NE-induced contraction, due to NO overproduction via eNOS and increased α2D-adrenoceptor expression, occur in early-stage GK rats and that the impaired ACh-induced relaxation in later-stage GK rats is due to reductions in both NO production and NO responsiveness (but not in eNOS expression).
Chronic mevastatin modulates receptor-dependent vascular contraction in eNOS-deficient mice
Authors: Budzyn Klaudia , Marley Philip D. , Sobey Christopher G.
Published: May 2004

We tested the hypothesis that endothelial nitric oxide (NO) synthase (eNOS)-derived NO modulates rho-kinase-mediated vascular contraction. Because 3-hydroxy-3-methylglutaryl (HMG)-CoA-reductase inhibition can both upregulate eNOS expression and inhibit rhoA/rho-kinase function, a second hypothesis tested was that statin treatment modulates rho-kinase-mediated contraction and that this can occur independently of eNOS. Contractile responses to the receptor-dependent agonists serotonin and phenylephrine but not to the receptor-independent agent KCl were greater in aortic rings from eNOS-null (eNOS−/−) vs. wild-type (eNOS+/+) mice. Similarly enhanced responses were seen in eNOS+/+rings after acute NOS inhibition. The rho-kinase inhibitor Y-27632 abolished or profoundly attenuated responses to receptor agonists in both eNOS+/+and eNOS−/−rings, but responses in eNOS+/+were more sensitive to Y-27632. Mevastatin treatment (20 mg/kg sc per day, 14 days) reduced responses to serotonin and phenylephrine in female mice of both strains. KCl-induced contractions were slightly smaller in eNOS+/+-derived aortic rings only. Levels of plasma cholesterol, and aortic expression of rhoA and rho-kinase, did not differ between groups. Thus eNOS-derived NO suppresses rhoA/rho-kinase-mediated vascular contraction. Moreover, a similar suppressive effect on rho-kinase-mediated vasoconstriction by statin therapy occurs independently of effects on eNOS or plasma cholesterol.
Effect of intracerebroventricular angiotensin II on body weight and food intake in adult rats
Authors: Porter James P. , Potratz Kristen R.
Published: May 2004

We recently reported that intracerebroventricular infusions of ANG II decreased food intake and increased energy expenditure in young rats. The aim of the present study was to determine if intracerebroventricular ANG II has similar effects in adult rats. The time course of the effect was also investigated with the idea that at earlier time points, a potential role for increased hypothalamic expression of corticotropin-releasing hormone (CRH) in the anorexia could be established. Finally, the contribution of ANG II-induced water drinking to the decrease in food intake was directly investigated. Rats received intracerebroventricular saline or ANG II using osmotic minipumps. Food intake, water intake, and body weight were measured daily. Experiments were terminated 2, 5, or 11 days after the beginning of the infusions. ANG II (∼ 32 ng·kg−1·min−1) produced a transient decrease in food intake that lasted for 4–5 days although body weight continued to be decreased for the entire experiment most likely due to increased energy expenditure as evidenced by increased uncoupling protein-1 mRNA expression in brown adipose tissue. At 11 and 5 days, the expression of CRH mRNA was decreased. At 2 days, CRH expression was not suppressed even though body weight was decreased. The decrease in food intake and body weight was identical whether or not rats were allowed to increase water consumption. These data suggest that in adult rats ANG II acts within the brain to affect food intake and energy expenditure in a manner that is not related to water intake.
Influence of vehicle resistance on transdermal iontophoretic delivery of acetylcholine and sodium nitroprusside in humans
Authors: Khan Faisel , Newton David J. , Smyth Emily C. , Belch Jill J. F.
Published: May 2004

Iontophoresis is a valuable method of noninvasive drug delivery for assessment of skin microvascular function, but it is important to consider and minimize its potential nonspecific electrical effects on blood flow. The use of sodium chloride (NaCl) instead of water as the iontophoresis vehicle has been reported to reduce these effects because it has a lower electrical resistance. However, this argument may not be valid when an agonist is added to the vehicle because its resistance will be changed. The aim of our study was to determine whether there is a difference in resistance between water and NaCl when used as vehicles for iontophoresis of acetylcholine (ACh) and sodium nitroprusside (SNP). Four cumulative doses of each drug, dissolved in either water or NaCl, were delivered via iontophoresis to the forearm skin of 14 healthy volunteers. We measured the resulting blood flow responses by using laser-Doppler imaging and the voltage across the electrodes for each delivery as an index of resistance. For ACh and SNP, there were no significant differences between the voltages measured when either water or NaCl was used as the vehicle. However, the blood flow responses to both agonists were significantly lower with NaCl (ACh: 25% lower, P < 0.001; SNP: 15% lower, P = 0.019). The use of NaCl is therefore unlikely to decrease any nonspecific electrical effects, and it may in fact reduce the effective dose of drug delivered. Deionized water is a better iontophoresis vehicle for the assessment of microvascular function in skin when using ACh and SNP.
Effects of prior heavy-intensity exercise on pulmonary O2 uptake and muscle deoxygenation kinetics in young and older adult humans
Authors: DeLorey Darren S. , Kowalchuk John M. , Paterson Donald H.
Published: May 2004

Pulmonary O2 uptake (V̇o2p) and muscle deoxygenation kinetics were examined during moderate-intensity cycling (80% lactate threshold) without warm-up and after heavy-intensity warm-up exercise in young ( n = 6; 25 ± 3 yr) and older ( n = 5; 68 ± 3 yr) adults. We hypothesized that heavy warm-up would speed V̇o2p kinetics in older adults consequent to an improved intramuscular oxygenation. Subjects performed step transitions ( n = 4; 6 min) from 20 W to moderate-intensity exercise preceded by either no warm-up or heavy-intensity warm-up (6 min). V̇o2p was measured breath by breath. Oxy-, deoxy-(HHb), and total hemoglobin and myoglobin (Hbtot) of the vastus lateralis muscle were measured continuously by near-infrared spectroscopy (NIRS). V̇o2p (phase 2; τ) and HHb data were fit with a monoexponential model. After heavy-intensity warm-up, oxyhemoglobin (older subjects: 13 ± 9 μM; young subjects: 9 ± 8 μM) and Hbtot (older subjects: 12 ± 8 μM; young subjects: 14 ± 10 μM) were elevated ( P < 0.05) relative to the no warm-up pretransition baseline. In older adults, τV̇o2p adapted at a faster rate ( P < 0.05) after heavy warm-up (30 ± 7 s) than no warm-up (38 ± 5 s), whereas in young subjects, τV̇o2p was similar in no warm-up (26 ± 7 s) and heavy warm-up (25 ± 5 s). HHb adapted at a similar rate in older and young adults after no warm-up; however, in older adults after heavy warm-up, the adaptation of HHb was slower ( P < 0.01) compared with young and no warm-up. These data suggest that, in older adults, V̇o2p kinetics may be limited by a slow adaptation of muscle blood flow and O2 delivery.
Age- and fitness-related differences in limb venous compliance do not affect tolerance to maximal lower body negative pressure in men and women
Authors: Hernandez J. P. , Franke W. D.
Published: May 2004

Aging and chronic exercise training influence leg venous compliance. Venous compliance affects responses to an orthostatic stress; its effect on tolerance to maximal lower body negative pressure (LBNP) in the elderly is unknown. The purpose of this study was to determine the influence of age and fitness, a surrogate measure of exercise training, on calf venous compliance and tolerance to maximal LBNP in men and women. Forty participants, 10 young fit (YF; age = 22.6 ± 0.5 yr, peak oxygen uptake = 57.1 ± 2.0 ml·kg−1·min−1), 10 young unfit (YU; 23.1 ± 1.0 yr, 41.1 ± 2.0 ml·kg−1·min−1), 10 older fit (OF; 73.9 ± 2.0 yr, 39.0 ± 2.0 ml·kg−1·min−1), and 10 older unfit (OU; 70.9 ± 1.6 yr, 27.1 ± 2.0 ml·kg−1·min−1), underwent graded LBNP to presyncope or 4 min at −100 mmHg. By utilizing venous occlusion plethysmography, calf venous compliance was determined by using the first derivative of the pressure-volume relation during cuff pressure reduction. We found that the more fit groups had greater venous compliance than their unfit peers ( P < 0.05) as did the young groups compared with their older peers ( P < 0.05) such that OU < YU = OF < YF. LBNP tolerance did not differ between groups. In conclusion, these data suggest that aging reduces, and chronic exercise increases, venous compliance. However, these data do not support a significant influence of venous compliance on LBNP tolerance.
VR1 Receptor Activation Induces Glutamate Release and Postsynaptic Firing in the Paraventricular Nucleus
Authors: Li De-Pei , Chen Shao-Rui , Pan Hui-Lin
Published: May 2004

Neurons in the paraventricular nucleus (PVN) are important in regulating autonomic function through projections to the brain stem and spinal cord. Although the vanilloid receptors (VR1) are present in the PVN, their physiological function is scarcely known. In this study, we determined the role of VR1 receptors in the regulation of synaptic inputs and the excitability of spinally projecting PVN neurons. Whole cell patch-clamp recordings were performed on the PVN neurons labeled by a retrograde fluorescence tracer injected into the thoracic spinal cord of rats. Capsaicin significantly increased the frequency of glutamatergic miniature excitatory postsynaptic currents (mEPSCs) without changing the amplitude and decay time constant of mEPSCs. On the other hand, capsaicin had no effect on GABAergic miniature inhibitory postsynaptic currents (mIPSCs). The effect of capsaicin on mEPSCs was abolished by a specific VR1 antagonist, iodo-resiniferatoxin (iodo-RTX), or ruthenium red. Importantly, iodo-RTX per se significantly reduced the amplitude of evoked EPSCs and the frequency of mEPSCs. Removal of extracellular Ca2+, but not Cd2+ treatment, also eliminated the effect of capsaicin on mEPSCs. Furthermore, capsaicin caused a large increase in the firing rate of PVN neurons, and such an effect was abolished in the presence of ionotropic glutamate receptor antagonists. Additionally, the double-immunofluorescence labeling revealed that all of the VR1 immunoreactivity was colocalized with a presynaptic marker, synaptophysin, in the PVN. Thus this study provides the first evidence that activation of VR1 receptors excites preautonomic PVN neurons through selective potentiation of glutamatergic synaptic inputs. Presynaptic VR1 receptors and endogenous capsaicin-like substances in the PVN may represent a previously unidentified mechanism in hypothalamic regulation of the autonomic nervous system.
Interlimb Differences in Control of Movement Extent
Authors: Sainburg Robert L. , Schaefer Sydney Y.
Published: May 2004

The current study was designed to examine potential interlimb asymmetries in controlling movement extent. Subjects made repetitive single-joint elbow extension movements while the arm was supported on a horizontal, frictionless, air-jet system. Four targets of 10, 20, 35, and 45° excursions were randomly presented over the course of 150 trials. For both arms, peak tangential hand velocity scaled linearly with movement distance. There was no significant difference between either peak velocities or movement accuracies for the two arms. However, the mechanisms responsible for achieving these velocities and extents were quite distinct for each arm. For the dominant arm, peak tangential finger acceleration varied systematically with movement distance. In contrast, nondominant-arm peak tangential acceleration varied little across targets and, as such, was a poor predictor of movement distance. Instead the velocities of the nondominant arm were determined primarily by variation in the duration of the initial acceleration impulse, which corresponds to the time of peak velocity. These different strategies reflect previously identified mechanisms in controlling movement distance: pulse-height control and pulse-width control. The former is characterized by a variation in peak acceleration and has been associated with preplanning mechanisms. The latter occurs after peak acceleration and has been shown to depend on peripheral sensory feedback. Our findings indicate that the dominant-arm system controls movement extent largely through planning mechanisms that specify pulse-height control, whereas the nondominant system does so largely through feedback mediated pulse-width control.