bluuebunny/crossref_metadata_2025_split huggingface dataset:
Standardized Assay Medium To Measure Lactococcus lactis Enzyme Activities while Mimicking Intracellular Conditions
Authors: Goel Anisha , Santos Filipe , de Vos Willem M. , Teusink Bas , Molenaar Douwe 
Published: October 2011

ABSTRACT

Knowledge of how the activity of enzymes is affected under
in vivo
conditions is essential for analyzing their regulation and constructing models that yield an integrated understanding of cell behavior. Current kinetic parameters for
Lactococcus lactis
are scattered through different studies and performed under different assay conditions. Furthermore, assay conditions often diverge from conditions prevailing in the intracellular environment. To establish uniform assay conditions that resemble intracellular conditions, we analyzed the intracellular composition of anaerobic glucose-limited chemostat cultures of
L. lactis
subsp.
cremoris
MG 1363. Based on this, we designed a new assay medium for enzyme activity measurements of growing cells of
L. lactis
, mimicking as closely as practically possible its intracellular environment. Procedures were optimized to be carried out in 96-well plates, and the reproducibility and dynamic range were checked for all enzyme activity measurements. The effects of freezing and the carryover of ammonium sulfate from the addition of coupling enzymes were also established. Activities of all 10 glycolytic and 4 fermentative enzymes were measured. Remarkably, most
in vivo
-like activities were lower than previously published data. Yet, the ratios of
V
max
over measured
in vivo
fluxes were above 1. With this work, we have developed and extensively validated standard protocols for enzyme activity measurements for
L. lactis
.
Expression Vectors for Acinetobacter baylyi ADP1
Authors: Murin Charles Daniel , Segal Kristy , Bryksin Anton , Matsumura Ichiro 
Published: October 2011

ABSTRACT

Acinetobacter baylyi
ADP1 is naturally competent and proficient at homologous recombination, so it can be transformed without restriction digests or ligation reactions. Expression vectors for this system, however, are not yet widely available. Here we describe the construction and characterization of inducible expression vectors that replicate as plasmids in
A. baylyi
or integrate into a nonessential part of its chromosome. These tools will facilitate the engineering of genes and genomes in this promising model organism.
Growth Inhibition of Various Enterobacteriaceae Species by the Yeast Hansenula anomala during Storage of Moist Cereal Grain
Authors: Olstorpe Matilda , Schnürer Johan , Passoth Volkmar 
Published: October 2011

ABSTRACT

Eleven of 13
Enterobacteriaceae
species tested grew in moist stored wheat, highlighting a potential risk of this energy-saving airtight storage method. When
Hansenula anomala
was coinoculated, all
Enterobacteriaceae
species were significantly inhibited after 2 months of storage, six of them to below the detection limit.
Acaricide Treatment Affects Viral Dynamics in Varroa destructor-Infested Honey Bee Colonies via both Host Physiology and Mite Control
Authors: Locke Barbara , Forsgren Eva , Fries Ingemar , de Miranda Joachim R. 
Published: October 2011

ABSTRACT

Honey bee (
Apis mellifera
) colonies are declining, and a number of stressors have been identified that affect, alone or in combination, the health of honey bees. The ectoparasitic mite
Varroa destructor
, honey bee viruses that are often closely associated with the mite, and pesticides used to control the mite population form a complex system of stressors that may affect honey bee health in different ways. During an acaricide treatment using Apistan (plastic strips coated with tau-fluvalinate), we analyzed the infection dynamics of deformed wing virus (DWV), sacbrood virus (SBV), and black queen cell virus (BQCV) in adult bees, mite-infested pupae, their associated
Varroa
mites, and uninfested pupae, comparing these to similar samples from untreated control colonies. Titers of DWV increased initially with the onset of the acaricide application and then slightly decreased progressively coinciding with the removal of the
Varroa
mite infestation. This initial increase in DWV titers suggests a physiological effect of tau-fluvalinate on the host's susceptibility to viral infection. DWV titers in adult bees and uninfested pupae remained higher in treated colonies than in untreated colonies. The titers of SBV and BQCV did not show any direct relationship with mite infestation and showed a variety of possible effects of the acaricide treatment. The results indicate that other factors besides
Varroa
mite infestation may be important to the development and maintenance of damaging DWV titers in colonies. Possible biochemical explanations for the observed synergistic effects between tau-fluvalinate and virus infections are discussed.
Functional Analysis of Lactobacillus rhamnosus GG Pili in Relation to Adhesion and Immunomodulatory Interactions with Intestinal Epithelial Cells
Authors: Lebeer Sarah , Claes Ingmar , Tytgat Hanne L. P. , Verhoeven Tine L. A. , Marien Eyra , von Ossowski Ingemar , Reunanen Justus , Palva Airi , de Vos Willem M. , De Keersmaecker Sigrid C. J. , Vanderleyden Jos 
Published: October 2011

ABSTRACT

Lactobacillus rhamnosus
GG, a probiotic with good survival capacity in the human gut, has well-documented adhesion properties and health effects. Recently,
spaCBA
-encoded pili that bind to human intestinal mucus were identified on its cell surface. Here, we report on the phenotypic analysis of a
spaCBA
pilus knockout mutant in comparison with the wild type and other adhesin mutants. The SpaCBA pilus of
L. rhamnosus
GG showed to be key for efficient adherence to the Caco-2 intestinal epithelial cell (IEC) line and biofilm formation. Moreover, the
spaCBA
mutant induces an elevated level of interleukin-8 (IL-8) mRNA in Caco-2 cells compared to the wild type, possibly involving an interaction of lipoteichoic acid with Toll-like receptor 2. In contrast, an
L. rhamnosus
GG mutant without exopolysaccharides but with an increased exposure of pili leads to the reduced expression of IL-8. Using Transwells to partition bacteria from Caco-2 cells, IL-8 induction is blocked completely regardless of whether wild-type or mutant
L. rhamnosus
GG cells are used. Taken together, our data suggest that
L. rhamnosus
GG SpaCBA pili, while promoting strong adhesive interactions with IECs, have a functional role in balancing IL-8 mRNA expression induced by surface molecules such as lipoteichoic acid.
Identification and Characterization of the Rhizobium sp. Strain GIN611 Glycoside Oxidoreductase Resulting in the Deglycosylation of Ginsenosides
Authors: Kim Eun-Mi , Kim Juhan , Seo Joo-Hyun , Park Jun-Seong , Kim Duck-Hee , Kim Byung-Gee 
Published: October 2011

ABSTRACT

Using enrichment culture,
Rhizobium
sp. strain GIN611 was isolated as having activity for deglycosylation of a ginsenoside, compound K (CK). The purified heterodimeric protein complex from
Rhizobium
sp. GIN611 consisted of two subunits with molecular masses of 63.5 kDa and 17.5 kDa. In the genome, the coding sequence for the small subunit was located right after the sequence for the large subunit, with one nucleotide overlapping. The large subunit showed CK oxidation activity, and the deglycosylation of compound K was performed via oxidation of ginsenoside glucose by glycoside oxidoreductase. Coexpression of the small subunit helped soluble expression of the large subunit in recombinant
Escherichia coli
. The purified large subunit also showed oxidation activity against other ginsenoside compounds, such as Rb1, Rb2, Rb3, Rc, F2, CK, Rh2, Re, F1, and the isoflavone daidzin, but at a much lower rate. When oxidized CK was extracted and incubated in phosphate buffer with or without enzyme, (
S
)-protopanaxadiol [PPD(S)] was detected in both cases, which suggests that deglycosylation of oxidized glucose is spontaneous.
A Highly Organized Structure Mediating Nuclear Localization of a Myb2 Transcription Factor in the Protozoan Parasite Trichomonas vaginalis
Authors: Chu Chien-Hsin , Chang Lung-Chun , Hsu Hong-Ming , Wei Shu-Yi , Liu Hsing-Wei , Lee Yu , Kuo Chung-Chi , Indra Dharmu , Chen Chinpan , Ong Shiou-Jeng , Tai Jung-Hsiang 
Published: October 2011

ABSTRACT

Nuclear proteins usually contain specific peptide sequences, referred to as nuclear localization signals (NLSs), for nuclear import. These signals remain unexplored in the protozoan pathogen,
Trichomonas vaginalis
. The nuclear import of a Myb2 transcription factor was studied here using immunodetection of a hemagglutinin-tagged Myb2 overexpressed in the parasite. The tagged Myb2 was localized to the nucleus as punctate signals. With mutations of its polybasic sequences, 48KKQK51 and 61KR62, Myb2 was localized to the nucleus, but the signal was diffusive. When fused to a C-terminal non-nuclear protein, the Myb2 sequence spanning amino acid (aa) residues 48 to 143, which is embedded within the R2R3 DNA-binding domain (aa 40 to 156), was essential and sufficient for efficient nuclear import of a bacterial tetracycline repressor (TetR), and yet the transport efficiency was reduced with an additional fusion of a firefly luciferase to TetR, while classical NLSs from the simian virus 40 T-antigen had no function in this assay system. Myb2 nuclear import and DNA-binding activity were substantially perturbed with mutation of a conserved isoleucine (I74) in helix 2 to proline that altered secondary structure and ternary folding of the R2R3 domain. Disruption of DNA-binding activity alone by point mutation of a lysine residue, K51, preceding the structural domain had little effect on Myb2 nuclear localization, suggesting that nuclear translocation of Myb2, which requires an ordered structural domain, is independent of its DNA binding activity. These findings provide useful information for testing whether myriad Mybs in the parasite use a common module to regulate nuclear import.
Identification of Tissue Cyst Wall Components by Transcriptome Analysis of
In Vivo
and In Vitro Toxoplasma gondii Bradyzoites
Authors: Buchholz Kerry R. , Fritz Heather M. , Chen Xiucui , Durbin-Johnson Blythe , Rocke David M. , Ferguson David J. , Conrad Patricia A. , Boothroyd John C. 
Published: October 2011

ABSTRACT

The
Toxoplasma gondii
bradyzoite is essential to establish persistent infection, yet little is known about what factors this developmental form secretes to establish the cyst or interact with its host cell. To identify candidate bradyzoite-secreted effectors, the transcriptomes of
in vitro
tachyzoites 2 days postinfection,
in vitro
bradyzoites 4 days postinfection, and
in vivo
bradyzoites 21 days postinfection were interrogated by microarray, and the program SignalP was used to identify signal peptides indicating secretion. One hundred two putative bradyzoite-secreted effectors were identified by this approach. Two candidates, bradyzoite pseudokinase 1 and microneme adhesive repeat domain-containing protein 4, were chosen for further investigation and confirmed to be induced and secreted by bradyzoites
in vitro
and
in vivo
. Thus, we report the first analysis of the transcriptomes of
in vitro
and
in vivo
bradyzoites and identify two new protein components of the
Toxoplasma
tissue cyst wall.
Zygotic Expression of the Double-Stranded RNA Binding Motif Protein Drb2p Is Required for DNA Elimination in the Ciliate Tetrahymena thermophila
Authors: Motl Jason A. , Chalker Douglas L. 
Published: October 2011

ABSTRACT

Double-stranded RNA binding motif (DSRM)-containing proteins play many roles in the regulation of gene transcription and translation, including some with tandem DSRMs that act in small RNA biogenesis. We report the characterization of the genes for double-stranded RNA binding proteins 1 and 2 (
DRB1
and
DRB2
), two genes encoding nuclear proteins with tandem DSRMs in the ciliate
Tetrahymena thermophila.
Both proteins are expressed throughout growth and development but exhibit distinct peaks of expression, suggesting different biological roles. In support of this, we show that expression of
DRB2
is essential for vegetative growth while
DRB1
expression is not. During conjugation, Drb1p and Drb2p localize to distinct nuclear foci. Cells lacking all
DRB1
copies are able to produce viable progeny, although at a reduced rate relative to wild-type cells. In contrast, cells lacking germ line
DRB2
copies, which thus cannot express Drb2p zygotically, fail to produce progeny, arresting late into conjugation. This arrest phenotype is accompanied by a failure to organize the essential DNA rearrangement protein Pdd1p into DNA elimination bodies and execute DNA elimination and chromosome breakage. These results implicate zygotically expressed Drb2p in the maturation of these nuclear structures, which are necessary for reorganization of the somatic genome.
Abdominal symptoms during physical exercise and the role of gastrointestinal ischaemia: a study in 12 symptomatic athletes
Authors: ter Steege Rinze WF , Geelkerken Robert H , Huisman Ad B , Kolkman Jeroen J 
Published: October 2011

BackgroundGastrointestinal (GI) symptoms during exercise may be caused by GI ischaemia. The authors report their experience with the diagnostic protocol and management of athletes with symptomatic exercise-induced GI ischaemia. The value of prolonged exercise tonometry in the diagnostic protocol of these patients was evaluated.MethodsPatients referred for GI symptoms during physical exercise underwent a standardised diagnostic protocol, including prolonged exercise tonometry. Indicators of GI ischaemia, as measured by tonometry, were related to the presence of symptoms during the exercise test (S+ and S− tests) and exercise intensity.Results12 athletes were specifically referred for GI symptoms during exercise (five males and seven females; median age 29 years (range 15–46 years)). Type of sport was cycling, long-distance running and triathlon. Median duration of symptoms was 32 months (range 7–240 months). Splanchnic artery stenosis was found in one athlete. GI ischaemia was found in six athletes during submaximal exercise. All athletes had gastric and jejunal ischaemia during maximum intensity exercise. No significant difference was found in gastric and jejunal Pco2or gradients between S+ and S− tests during any phase of the exercise protocol. In S+ tests, but not in S− tests, a significant correlation between lactate and gastric gradient was found. In S+ tests, the regression coefficients of gradients were higher than those in S− tests. Treatment advice aimed at limiting GI ischaemia were successful in reducing complaints in the majority of the athletes.ConclusionGI ischaemia was present in all athletes during maximum intensity exercise and in 50% during submaximal exercise. Athletes with GI symptoms had higher gastric gradients per mmol/l increase in lactate, suggesting an increased susceptibility for the development of ischaemia during exercise. Treatment advice aimed at limiting GI ischaemia helped the majority of the referred athletes to reduce their complaints. Our results suggest an important role for GI ischaemia in the pathophysiology of their complaints.
Feasibility of precompetition medical assessment at FIFA World Cups for female youth players
Authors: Dvorak Jiri , Grimm Katharina , Schmied Christian , Junge Astrid 
Published: October 2011

BackgroundAlthough most experts agree that preparticipation screening is important to prevent sudden cardiac death in sport, only a few reports have been published on the feasibility of its practical implementation.MethodsThe football associations participating in the U-17 and U-20 Women's World Cups 2010 were asked to perform a standardised precompetition medical assessment (PCMA) of their players (in total 672).ResultsCompliance with the requirement for performing the PCMA was high among all teams, particularly from African, Asian and Central/South American countries. No relevant abnormal findings in personal history and clinical cardiological examination were reported. Athletic ECG patterns were frequent, but very few findings were considered to require further investigation. All players were declared as eligible to play.ConclusionsBased on the demonstrated feasibility of performing a comprehensive PCMA in elite female youth players, the Fédération Internationale de Football Association (FIFA) Executive Committee decided to make the PCMA a compulsory requirement for all FIFA competitions.
Procedure competence versus number performed: a survey of graduate emergency medicine specialists in a developing nation: Table 1
Authors: Wen Leana S , Nagurney J Tobias , Geduld Heike I , Wen Angela P , Wallis Lee A 
Published: October 2011

BackgroundEmergency medicine (EM) training programmes are being conducted around the world but no study has assessed the procedural competence of developing nations' EM trainees.ObjectivesTo quantify the number of core procedures and resuscitations performed and describe the perceived procedural competency of graduates of Africa's first EM registrarship at the University of Cape Town/Stellenbosch University (UCT/SUN) in Cape Town, South Africa.MethodsAll 30 graduates from the first four classes in the UCT/SUN EM programme (2007–10) were asked to complete a written, self-administered survey on the number of procedures needed for competency, the number of procedures performed during registrarship and the perceived competence in each procedure ranked on a five-point Likert scale. The procedures selected were the 10 core procedures and four types of resuscitations as defined by the US-based Residency Review Committee. Results were compiled and analysed using descriptive statistics.ResultsTwenty-seven (90%) completed surveys. For most core procedures and all resuscitations, the number of procedures reported by respondents far exceeded the Residency Review Committee minimum. The three procedures not meeting the minimum were internal cardiac pacing, cricothyrotomy and periocardiocentesis. Respondents reported perceived competence in most procedures and all resuscitations.ConclusionsEM trainees in a South Africa registrarship report a high number of procedures performed for most procedures and all resuscitations. As medical education moves to the era of direct observation and other methods of assessment, more studies are needed to define and ensure procedural competence in trainees of nascent EM programmes.
MULTIPLE LOCALIZED GIANT CELL TUMOR OF THE TENDON SHEATH (GCTTS) AFFECTING A SINGLE TENDON: A VERY RARE CASE REPORT AND REVIEW OF RECENT CASES
Authors: Singh Talvinder , Noor Saqib , Simons Adrian W. 
Published: October 2011

Introduction Giant cell tumors of the tendon sheath (GCTTS) are very common. More recently, a small number of case reports have identified the presence of multifocal GCTTS in the hand. These case reports have identified the presence of a maximum of two simultaneous lesions of a giant cell tumor affecting the same tendon sheath. We present an exceptionally rare case of simultaneous multiple localized GCTTS in which five lesions were identified on a single tendon simultaneously. This number of lesions on a single tendon has never been previously reported. Case: A 37-year-old tree surgeon initially complained of pain in the region of the base of the ring and little fingers. A month later, he developed multiple soft tissue swellings at these sites and a soft tissue mass in the center of the palm relating to the left ring finger. A magnetic resonance imaging (MRI) scan suggested multiple GCTTS. These masses were excised completely without MRI evidence of a recurrence. Multiple GCTTS should be a differential diagnosis of multiple soft tissue swellings in the hand with an MRI scan and complete excision being the appropriate imaging and treatment modality respectively.
In VivoPositron Emission Tomography Imaging of Protease Activity by Generation of a Hydrophobic Product from a Noninhibitory Protease Substrate
Authors: Chuang Chih-Hung , Chuang Kuo-Hsiang , Wang Hsin-Ell , Roffler Steve R. , Shiea Jen-taie , Tzou Shey-Cherng , Cheng Ta-Chun , Kao Chien-Han , Wu Shih-Yen , Tseng Wei-Lung , Cheng Chiu-Min , Hou Ming-Feng , Wang Ju-Ming , Cheng Tian-Lu 
Published: October 2011

AbstractPurpose: To develop an imaging technology for protease activities in patients that could help in prognosis prediction and in design of personalized, protease-based inhibitors and prodrugs for targeted therapy.Experimental Design: Polyethylene glycol (PEG) was covalently attached to the N-terminus of a hydrophilic peptide substrate (GPLGVR) for matrix metalloproteinase (MMP) to increase hydrophilicity. PEG-peptide was then linked to a hydrophobic tetramethylrhodamine (TMR) domain and labeled with 18F to form a PEG-peptide-18F-TMR probe. Specific cleavage of the probe by MMP2 was tested in vitro by matrix-assisted laser desorption/ionization–time-of-flight (MALDI-TOF). In vivo imaging of MMP2-expressing tumors was evaluated by micro-PET.Results: The hydrophobic TMR fragment (948 Da) was specifically generated by MMP2 enzymes and MMP-expressing HT1080 cells but not control MCF-7 cells. MMP-expressing HT1080 cells and tumors selectively accumulated the hydrolyzed, hydrophobic TMR fragment at sites of protease activity. Importantly, we found that 18F-labeled probe (18F-TMR) preferentially localized in HT1080 tumors but not control MCF-7 tumors as shown by micro-PET. Uptake of the probe in HT1080 tumors was 18.4 ± 1.9-fold greater than in the MCF-7 tumors 30 minutes after injection. These results suggest that the PEG-peptide-18F-TMR probe displays high selectivity for imaging MMP activity.Conclusions: This strategy successfully images MMP expression in vivo and may be extended to other proteases to predict patient prognosis and to design personalized, protease-based inhibitors and prodrug-targeted therapies. Clin Cancer Res; 18(1); 238–47. ©2011 AACR.
Stent Coated With Antibody Against Vascular Endothelial-Cadherin Captures Endothelial Progenitor Cells, Accelerates Re-Endothelialization, and Reduces Neointimal Formation
Authors: Lim Woo-Hyun , Seo Won-Woo , Choe WonSeok , Kang Chan-Koo , Park Jonghanne , Cho Hyun-Ju , Kyeong San , Hur Jin , Yang Han-Mo , Cho Hyun-Jai , Lee Yoon-Sik , Kim Hyo-Soo 
Published: October 2011

Objective—
In contrast to CD34, vascular endothelial-cadherin (VE-cadherin) is exclusively expressed on the late endothelial progenitor cells (EPC) whereas not on the early or myeloid EPC. Thus, VE-cadherin could be an ideal target surface molecule to capture circulating late EPC. In the present study, we evaluated whether anti-VE–cadherin antibody-coated stents (VE-cad stents) might accelerate endothelial recovery and reduce neointimal formation through the ability of capturing EPC.

Methods and Results—

The stainless steel stents were coated with rabbit polyclonal anti-human VE-cadherin antibodies and exposed to EPC for 30 minutes in vitro. The number of EPC that adhered to the surface of VE-cad stents was significantly higher than bare metal stents (BMS) in vitro, which was obliterated by pretreatment of VE-cad stent with soluble VE-cadherin proteins. We deployed VE-cad stents and BMS in the rabbit right and left iliac arteries, respectively. At 48 hours after stent deployment in vivo, CD-31–positive endothelial cells adhered to VE-cad stent significantly more than to BMS. At 3 days, scanning electron microscopy showed that over 90% surface of VE-cad stents was covered with endothelial cells, which was significantly different from BMS. At 42 days, neointimal area that was filled with smooth muscle cells positive for actin or calponin was significantly smaller in VE-cad stents than in BMS by histological analysis (0.95±0.22 versus 1.34±0.43 mm
2
, respectively,
P
=0.02). Immuno-histochemical analysis revealed that infiltration of inflammatory cells was not significantly different between 2 stents.

Conclusion—
VE-cad stents captured EPC successfully in vitro, accelerated endothelial recovery on stent, and eventually reduced neointimal formation in vivo.