bluuebunny/crossref_metadata_2025 huggingface dataset:
Comparison and evaluation of experimental mediastinitis models: precolonized foreign body implants and bacterial suspension inoculation seems promising
Authors: Ersoz Gulden , Aytacoglu Barlas Naim , Sucu Nehir , Tamer Lulufer , Bayindir Ismet , Kose Necmi , Kaya Ali , Dikmengil Murat
Published: April 2006

Background

Post-sternotomy mediastinitis (PSM) is a devastating surgical complication affecting 1–3% of patients that undergo cardiac surgery. Staphylococcus aureus is one of the most commonly encountered bacterial pathogen cultured from mediastinal samples obtained from patients with PSM. A component of the membrane of the gram positive bacteria, lipoteichoic acid, stimulates the blood monocytes and macrophages to secrete cytokines, radicals and nitrogen species leading to oxido-inflammatory damage. This seems to be responsible for the high mortality rate in PSM. For the evaluation of the pathogenesis of infection or for the investigation of alternative treatment models in infection, no standard model of mediastinitis seems to be available. In this study, we evaluated four mediastinitis models in rats.

Methods

The rats were divided into four groups to form different infection models. Group A: A suspension of 1 × 107 colony-forming units Staphylococcus aureus in 0,5 mL was inoculated from the right second intercostal space into the mediastinum. Group B: A hole was created in the right second intercostal space and a piece of stainless-steel implant with a length of 0.5 cm was inserted into the mediastinum and a suspension of 1 × 107 cfu bacteria in 0,5 mL was administered via the tail vein. Group C: Precolonized stainless-steel implant was inserted into the mediastinum. Group D: Precolonized stainless-steel implant was inserted into the mediastinum and the bacteria suspension was also injected into the mediastinum. On the 10th day, rats were sacrificed and the extension of infection in the mediastenae was evaluated by quantitative cultures. Myeloperoxidase activity (MPO) and malondialdehyde (MDA) levels were determined in the sera to evaluate the neutrophil activation and assess the inflammatory oxidation.

Results

The degree of infection in group C and D were 83.3% and 100% respectively (P < 0.001). MDA levels were significantly higher in these two groups than the others (P < 0.001).

Conclusion

Infected implants and high bacterial concentration administration were the two important components that played a significant role in the outcome of a successful infection in mediastinum in a rat model.
Scaling up health interventions in resource-poor countries: what role does research in stated-preference framework play?
Authors: Pokhrel Subhash
Published: April 2006

Despite improved supply of health care services in low-income countries in the recent past, their uptake continues to be lower than anticipated. This has made it difficult to scale-up those interventions which are not only cost-effective from supply perspectives but that might have substantial impacts on improving the health status of these countries. Understanding demand-side barriers is therefore critically important. With the help of a case study from Nepal, this commentary argues that more research on demand-side barriers needs to be carried out and that the stated-preference (SP) approach to such research might be helpful. Since SP techniques place service users' preferences at the centre of the analysis, and because preferences reflect individual or social welfare, SP techniques are likely to be helpful in devising policies to increase social welfare (e.g. improved service coverage). Moreover, the SP data are collected in a controlled environment which allows straightforward identification of effects (e.g. that of process attributes of care) and large quantities of relevant data can be collected at moderate cost. In addition to providing insights into current preferences, SP data also provide insights into how preferences are likely to respond to a proposed change in resource allocation (e.g. changing service delivery strategy). Finally, the SP-based techniques have been used widely in resource-rich countries and their experience can be valuable in conducting scaling-up research in low-income countries.
Assembly of the phagocyte NADPH oxidase complex: chimeric constructs derived from the cytosolic components as tools for exploring structure-function relationships
Authors: Mizrahi Ariel , Berdichevsky Yevgeny , Ugolev Yelena , Molshanski-Mor Shahar , Nakash Yael , Dahan Iris , Alloul Nathalie , Gorzalczany Yara , Sarfstein Rive , Hirshberg Miriam , Pick Edgar
Published: April 2006

Phagocytes generate superoxide (O2.−) by an enzyme complex known as reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase. Its catalytic component, responsible for the NADPH-driven reduction of oxygen to O2.−, is flavocytochrome b559, located in the membrane and consisting of gp91phox and p22phox subunits. NADPH oxidase activation is initiated by the translocation to the membrane of the cytosolic components p47phox, p67phox, and the GTPase Rac. Cytochrome b559 is converted to an active form by the interaction of gp91phox with p67phox, leading to a conformational change in gp91phox and the induction of electron flow. We designed a new family of NADPH oxidase activators, represented by chimeras comprising various segments of p67phox and Rac1. The prototype chimera p67phox (1–212)-Rac1 (1–192) is a potent activator in a cell-free system, also containing membrane p47phox and an anionic amphiphile. Chimeras behave like bona fide GTPases and can be prenylated, and prenylated (p67phox-Rac1) chimeras activate the oxidase in the absence of p47phox and amphiphile. Experiments involving truncations, mutagenesis, and supplementation with Rac1 demonstrated that the presence of intrachimeric bonds between the p67phox and Rac1 moieties is an absolute requirement for the ability to activate the oxidase. The presence or absence of intrachimeric bonds has a major impact on the conformation of the chimeras, as demonstrated by fluorescence resonance energy transfer, small angle X-ray scattering, and gel filtration. Based on this, a “propagated wave” model of NADPH oxidase activation is proposed in which a conformational change initiated in Rac is propagated to p67phox and from p67phox to gp91phox.
A review of nasal trauma
Authors: Weller MD , Drake-Lee AB
Published: April 2006

The authors have undertaken a review of the diagnosis and management of nasal trauma. It deals with the normal anatomy of the nose, injury patterns to the nose, treatment of nasal trauma and some of the complications of nasal trauma.
Automated External Defibrillator Program Does Not Impair Cardiopulmonary Resuscitation Initiation in the Public Access Defibrillation Trial
Authors: Hedges Jerris R. , Sehra Ruchir , Van Zile Jonathan W. , Anton Andrew R. , Bosken Lois A. , O'Connor Robert E. , Moore Richard , Powell Judy L. , McBurnie Mary Ann
Published: April 2006

Objectives:
To evaluate whether automated external defibrillator (AED) training and AED availability affected the response of volunteer rescuers and performance of cardiopulmonary resuscitation (CPR) in presumed out‐of‐hospital cardiac arrest (OOH‐CA) during the multicenter Public Access Defibrillation Trial.

Methods:
The Public Access Defibrillation Trial recruited 1,260 facilities in 24 North American regional sites to participate in a trial addressing survival from OOH‐CA when AED training and availability were added to a volunteer‐based emergency response team. Volunteers at each facility were trained to perform either CPR alone (CPR) or CPR in conjunction with AED use (CPR+AED) according to randomized assignments. This study reports the frequency of response and initiation of CPR actions (chest compressions and/or ventilations) by volunteers in the CPR and CPR+AED study groups.

Results:
A total of 314 presumed OOH‐CA episodes occurred in CPR facilities, and 308 occurred in CPR+AED facilities. The volunteers were matched well for age, gender, and other features. Overall, ventilations (23.1% vs. 13.1%), chest compressions (24.4% vs. 12.1%), and both actions (19.8% vs. 10.5%; all p < 0.05) were more commonly performed in OOH‐CA cases in the CPR+AED group. However, when only OOH‐CA cases with volunteers responding were analyzed, the rates of CPR actions were similar. In the subgroup of CPR+AED cases with a responding volunteer, the AED was turned on for only 47% of cases. Volunteers initiated a CPR action more commonly when the AED was turned on (60.7% vs. 39.3%; p = 0.003).

Conclusions:
In the Public Access Defibrillation Trial, rates of CPR actions for presumed OOH‐CA victims were low but similar for CPR and CPR+AED responding volunteer rescuers. Factors associated with volunteer response, CPR action initiation, and AED activation warrant further investigation.
Reliability of a Core Competency Checklist Assessment in the Emergency Department: The Standardized Direct Observation Assessment Tool
Authors: Shayne Philip , Gallahue Fiona , Rinnert Stephan , Anderson Craig L. , Hern Gene , Katz Eric
Published: April 2006

Objectives:
A Council of Emergency Medicine Residency Directors task force developed the Standardized Direct Observation Assessment Tool (SDOT), a 26‐item checklist assessment tool to evaluate Accreditation Council for Graduate Medical Education resident core competencies by direct observation. Each of the checklist items is assigned to one or more of five core competencies. The objective of this study was to test the interrater measurement properties of the SDOT instrument.

Methods:
Two videos of simulated patient–resident–attending physician encounters were produced. Academic emergency medicine faculty members not involved in the development of the form viewed the two encounters and completed the SDOT for each. Faculty demographic data were collected. Data were collected from 82 faculty members at 16 emergency medicine residency programs. The checklist items were used to generate a composite score for each core competency of patient care, medical knowledge, interpersonal and communication skills, professionalism, and systems‐based practice.

Results:
Univariate analysis demonstrated a high degree of agreement between evaluators in evaluating residents for both videos. Multivariate analysis found no differences in rating by faculty when examined by experience, academic title, site, or previous use of the SDOT.

Conclusions:
Faculty from 16 emergency medicine residency programs had a high interrater agreement when using the SDOT to evaluate resident core competency performance. This study did not test the validity of the tool. This data analysis is mainly descriptive, and scripted video scenarios may not approximate direct observation in the emergency department.
Comprehensive Analysis ofUGT1APolymorphisms Predictive for Pharmacokinetics and Treatment Outcome in Patients With Non–Small-Cell Lung Cancer Treated With Irinotecan and Cisplatin
Authors: Han Ji-Youn , Lim Hyeong-Seok , Shin Eun Soon , Yoo Yeon-Kyeong , Park Yong Hoon , Lee Jong-Eun , Jang In-Jin , Ho Lee Dae , Soo Lee Jin
Published: April 2006

Purpose

To determine whether uridine diphosphate-glucuronosyltransferase 1A1, UGT1A7, and UGT1A9 polymorphisms affect the pharmacokinetics (PK) of irinotecan and treatment outcome of Korean patients with advanced non–small-cell lung cancer (NSCLC).

Methods

Eighty-one patients with advanced NSCLC were treated with irinotecan (80 mg/m2) on day 1 and 8 and cisplatin (60 mg/m2) on day 1 intravenously of each 3-week cycle. Genomic DNA was extracted from peripheral blood and genotyped using direct sequencing. We analyzed the association of UGT1A genotypes with irinotecan PK and clinical outcomes. All statistical tests were two-sided.

Results

In genotype-PK association analysis, UGT1A1*6/*6 (n = 6), UGT1A7*3/*3 (n = 6), and UGT1A9-118(dT)9/9(n = 11) were associated with significantly lower area under the time-concentration curve (AUC) SN-38G to SN-38 (AUCSN-38G/AUCSN-38) ratio (P = .002, P = .009, and P = .001, respectively). In linkage disequilibrium analysis, the UGT1A7 variants were highly linked with the UGT1A1*6 (D′ = 0.85, r2= 0.63) and UGT1A9*22 (D′ = 0.95, r2= 0.88), which was substantiated in haplotype analysis. Patients with UGT1A1*6/*6 had lower tumor response and higher incidence of severe neutropenia. UGT1A9-118(dT)9/9also showed a trend for high incidence of severe diarrhea, but not tumor response. In survival analysis, patients with UGT1A1*6/*6 had significantly shorter progression-free survival (P = .001) and overall survival (P = .017).

Conclusion

These findings suggest that UGT1A1*6 and UGT1A9*22 genotypes may be important for SN-38 glucuronidation and associate with irinotecan-related severe toxicity. Specifically, UGT1A1*6 might be useful for predicting tumor response and survival outcome of Korean patients with NSCLC treated with irinotecan-based chemotherapy.
Epigenetic Modification of the X Chromosome Influences Susceptibility to Polycystic Ovary Syndrome
Authors: Hickey T. E. , Legro R. S. , Norman R. J.
Published: April 2006

Context: The cause of polycystic ovary syndrome (PCOS) is unknown, although genetic and environmental influences are clearly implicated. Some genetic studies have suggested the involvement of X-linked genes in PCOS, but the influence of X chromosome inactivation (XCI) on manifestation of this disorder has not previously been examined.

Objective: The objective of the study was to test the null hypothesis that XCI has no influence on clinical presentation of PCOS.

Design: We examined patterns of XCI between sister pairs with the same genotype at a polymorphic locus on the X chromosome in families with PCOS.

Setting: The study was conducted at a private practice.

Participants: PCOS was defined as hyperandrogenemia with chronic anovulation. Forty families were studied in which DNA was obtained from at least one parent, the proband, and one sister that could be accurately diagnosed as being affected or unaffected.

Main Outcome Measure(s): Relative expression of two X-linked alleles was determined, and the ratio of one to the other represented the pattern of XCI.

Results: The statistical odds on a different clinical presentation between sisters was approximately 29 times higher in sister pairs with different patterns of XCI, compared with sister pairs with the same pattern of XCI (odds ratio 28.9; 95% confidence interval 4.0–206; P = 0.0008).

Conclusions: This study provides evidence to refute the null hypothesis and propose a closer inspection of X-linked genes in PCOS, one in which both genotype and epigenotype are considered. Environmental determinants of PCOS may alter clinical presentation via epigenetic modifications, which currently remain undetected in traditional genetic analyses.
Surgical Remission of Pituitary Adenomas Confined to the Neurohypophysis in Cushing’s Disease
Authors: Weil Robert J. , Vortmeyer Alexander O. , Nieman Lynnette K. , DeVroom Hetty L. , Wanebo John , Oldfield Edward H.
Published: April 2006

Context: Partial or total removal of the pituitary cures 60–80% of patients with Cushing’s disease (CD) in whom an adenoma cannot be identified at surgery. Many patients who fail complete or partial hypophysectomy are cured by sellar and parasellar irradiation.

Design/Patients: As part of a series of prospective studies of CD, we identified 12 patients (34.5 ± 19.9 yr; 11 females; four children) with tumors located completely within the neurohypophysis among 730 patients undergoing surgery for CD.

Setting: The study was conducted at a tertiary referral center at a clinical research hospital.

Results: All 12 patients had clinical and biochemically defined CD. Tumor was visible at surgery in 11 patients; all 12 tumors were positive for ACTH by immunohistochemistry. Two tumors were excised at repeat surgery because of persistent hypercortisolism within 14 d of negative exploration of the adenohypophysis. There were no long-term complications. At follow-up of 71.9 ± 34.2 months (range, 30–138 months), all patients are in remission of CD. Adult patients have had significant improvement in weight and body mass indices, with restoration of normal menses in all women. In the four pediatric patients, height, weight, and body mass indices have been restored toward normal by surgical remission of CD. Hypopituitarism or long-term neurohypophysial dysfunction has not occurred.

Conclusion: We report a new subset of patients with CD, ACTH-secreting adenomas that arise wholly within the posterior lobe of the pituitary gland. In cases of CD in which an adenoma is not identified in the adenohypophysis and in patients with persistent hypercortisolism after complete or partial excision of the anterior lobe, tumor within the neurohypophysis should be considered; selective adenomectomy of a neurohypophyseal, ACTH-secreting tumor can produce long-term remission.
Myosin light chain phosphorylation regulates barrier function by remodeling tight junction structure
Authors: Shen Le , Black Eric D. , Witkowski Edwina D. , Lencer Wayne I. , Guerriero Vince , Schneeberger Eveline E. , Turner Jerrold R.
Published: April 2006

Epithelial tight junctions form a barrier against passive paracellular flux. This barrier is regulated by complex physiologic and pathophysiologic signals that acutely fine-tune tight junction permeability. Although actomyosin contraction and myosin light chain phosphorylation are clearly involved in some forms of tight junction regulation, the contributions of other signaling events and the role of myosin light chain phosphorylation in this response are poorly understood. Here we ask if activation of myosin light chain kinase alone is sufficient to induce downstream tight junction regulation. We use a confluent polarized intestinal epithelial cell model system in which constitutively active myosin light chain kinase, tMLCK, is expressed using an inducible promoter. tMLCK expression increases myosin light chain phosphorylation, reorganizes perijunctional F-actin, and increases tight junction permeability. TJ proteins ZO-1 and occludin are markedly redistributed, morphologically and biochemically, but effects on claudin-1 and claudin-2 are limited. tMLCK inhibition prevents changes in barrier function and tight junction organization induced by tMLCK expression, suggesting that these events both require myosin light chain phosphorylation. We conclude that myosin light chain phosphorylation alone is sufficient to induce tight junction regulation and provide new insights into the molecular mechanisms that mediate this regulation.
Toxoplasma gondiitriggers Gi-dependent PI 3-kinase signaling required for inhibition of host cell apoptosis
Authors: Kim Leesun , Denkers Eric Y.
Published: April 2006

Infection with the intracellular parasite Toxoplasma gondii renders cells resistant to multiple pro-apoptotic signals, but underlying mechanisms have not been delineated. The phosphoinositide 3-kinase (PI 3-kinase) pathway and the immediate downstream effector protein kinase B (PKB/Akt) play important roles in cell survival and apoptosis inhibition. Here, we show that Toxoplasma infection of mouse macrophages activates PKB/Akt in vivo and in vitro. In a mixed population of infected and non-infected macrophages, activation is only observed in parasite-infected cells. The PI 3-kinase inhibitors wortmannin and LY294002 block parasite-induced PKB phosphorylation. PKB activation occurs independently of Toll-like receptor adaptor protein MyD88 but uncoupling of Gi-protein-mediated signaling with pertussis toxin prevents PKB phosphorylation. Moreover, in the presence of PI 3-kinase inhibitors or pertussis toxin, not only PKB activation but also ERK1/2 activation during T. gondii infection is defective. Most importantly, the parasite's ability to induce macrophage resistance to pro-apoptotic signaling is prevented by incubation with PI 3-kinase inhibitors. This study demonstrates that T. gondii exploits host Gi-protein-dependent PI 3-kinase signaling to prevent induction of apoptosis in infected macrophages.
Lgl mediates apical domain disassembly by suppressing the PAR-3-aPKC-PAR-6 complex to orient apical membrane polarity
Authors: Yamanaka Tomoyuki , Horikoshi Yosuke , Izumi Natsuko , Suzuki Atsushi , Mizuno Keiko , Ohno Shigeo
Published: April 2006

The basolateral tumor suppressor protein Lgl is important for the regulation of epithelial cell polarity and tissue morphology. Recent studies have shown a physical and functional interaction of Lgl with another polarity-regulating protein machinery, the apical PAR-3-aPKC-PAR-6 complex, in epithelial cells. However, the mechanism of Lgl-mediated regulation of epithelial cell polarity remains obscure. By an siRNA method, we here show that endogenous Lgl is required for the disassembly of apical membrane domains in depolarizing MDCK cells induced by Ca2+ depletion. Importantly, this Lgl function is mediated by the suppression of the apical PAR-3-aPKC-PAR-6 complex activity. Analysis using 2D- or 3D-cultured cells in collagen gel suggests the importance of this suppressive regulation of Lgl on the collagen-mediated re-establishment of apical membrane domains and lumen formation. These results indicate that basolateral Lgl plays a crucial role in the disassembly of apical membrane domains to induce the orientation of apical membrane polarity, which is mediated by the suppression of apical PAR-3-aPKC-PAR-6 complex activity.
Elucidation of megalin/LRP2-dependent endocytic transport processes in the larval zebrafish pronephros
Authors: Anzenberger Uwe , Bit-Avragim Nana , Rohr Stefan , Rudolph Franziska , Dehmel Bastian , Willnow Thomas E. , Abdelilah-Seyfried Salim
Published: April 2006

Megalin/LRP2 is an endocytic receptor in the proximal tubules of the mammalian kidney that plays a central role in the clearance of metabolites from the glomerular filtrate. To establish a genetic model system for elucidation of molecular components of this retrieval pathway, we characterized orthologous transport processes in the zebrafish. We show that expression of megalin/LRP2 and its co-receptor cubilin is conserved in the larval zebrafish pronephros and demarcates a segment of the pronephric duct that is active in clearance of tracer from the ultrafiltrate. Knock-down of megalin/LRP2 causes lack of Rab4-positive endosomes in the proximal pronephric duct epithelium and abrogates apical endocytosis. Similarly, knock-down of the megalin/LRP2 adaptor Disabled 2 also blocks renal clearance processes. These results demonstrate the conservation of the megalin/LRP2 retrieval pathway between the larval zebrafish pronephros and the mammalian kidney and set the stage for dissection of the renal endocytic machinery in a simple model organism. Using this model system, we provide first genetic evidence that renal tubular endocytosis and formation of endosomes is a ligand-induced process that crucially depends on megalin/LRP2 activity.
Poly(N-isopropylacrylamide) Microparticle Formation in Water by Infrared Laser-Induced Photo-Thermal Phase Transition
Authors: Ishikawa Masazumi , Misawa Hiroaki , Kitamura Noboru , Masuhara Hiroshi
Published: April 2006

Irradiation of a focused 1064 nm laser beam to an aqueous poly(N-isopropylacrylamide) solution resulted in the reversible microparticle (diameter ≈ 7 μm) formation through local heating of water by the laser beam and subsequent phase transition of the polymer solution. The microparticle produced was simultaneously manipulated in three-dimensional space by the incident 1064 nm laser beam.
Giant Metal-Insulator-Metal Transition Induced by Selective Deuteration of the Molecular Conductor, (DMe-DCNQI)2Cu (DMe-DCNQI = 2,5-dimethyl-N,N′-dicyanoquinonediimine)
Authors: Aonuma Shuji , Sawa Hiroshi , Kato Reizo , Kobayashi Hayao
Published: April 2006

The Cu salts of selectively deuterated DMe-DCNQI’s were prepared, and the electrical conductivities were examined. By deuteration of two hydrogens of the methyl groups, a drastic metal-insulator-metal transition was induced for the first time in a non-alloy system at ambient pressure. On the other hand, deuteration of two ring hydrogens or one methyl hydrogen did not affect the stability of a metallic state down to 4.2 K. These phenomena are discussed in terms of the “chemical pressure” effect.
Effect of Substituents on the Equilibrium between Trans and Perpendicular Conformers in the Triplet Excited State of Substituted Styrylpyrenes
Authors: Kikuchi Yoshihiko , Okamoto Hiroaki , Arai Tatsuo , Tokumaru Katsumi
Published: April 2006

In the equilibrium between trans (3t*) and perpendicular (3p*) conformers in the triplet excited state of 1-styrylpyrenes, substitution by electron-accepting groups at the para-position of styryl group more effectively stabilizes 3t* than 3p*, therefore affording very high composition of their trans isomers at the photostationary state.
Efficient Electrochemical Reduction of N2 to NH3 Catalyzed by Lithium
Authors: Tsuneto Akira , Kudo Akihiko , Sakata Tadayoshi
Published: April 2006

A novel electrochemical N2 reduction system is reported. Ammonia was formed with a significant current efficiency (up to 8%) by a simple electrolysis, which was conducted under N2 now (1 atm) in a solution of LiClO4 (0.2 M) in tetrahydrofuran/ethanol (99:1 v/v) on some metal electrodes. Higher current efficiency (48.7%) for NH3 formation was achieved when the electrolysis was conducted under high pressure of N2 (50 atm). Lithium seemed to act as a catalyst (mediator) in this electrochemical system.
The Strange Attractor of Chaotic Voltage Oscillation in Water-Oil-Water Liquid Membrane Containing Cationic Surfactant
Authors: Takahashi Koichi , Yasukawa Hideaki , Sugimura Takaaki
Published: April 2006

A strange attractor of chaotic voltage oscillation caused by micelle formation of cationic surfactant in the water-oil-water liquid membrane was found. The topology of the strange attractor is the mirror image of the Rössler’s attractor. The correlation dimension of the attractor is 2.4.
Precursor of Carthamin, A Constituent of Safflower
Authors: Kumazawa Toshihiro , Sato Shingo , Kanenari Daisuke , Kunimatsu Akira , Hirose Ryoji , Matsuba Shigeru , Obara Heitaro , Suzuki Masanobu , Sato Masaya , Onodera Jun-ichi
Published: April 2006

Precursor of Carthamin was isolated from the flowers of Carthamin tinctorius L. (Safflower) and its structure was characterized as 1 on the basis of spectroscopic analysis and by comparing its properties with those of synthetic model compound.
Dehydroepiandrosterone sulphate interferes with the Abbott Architect direct immunoassay for testosterone
Authors: Warner M H , Kane J W , Atkin S L , Kilpatrick E S
Published: April 2006

Background: There are known difficulties in measuring testosterone in women using 'direct' (non-extraction) immunoassays. The positive bias encountered in many of these assays has been attributed to an unidentified interfering substance. We have evaluated the potential for dehydroepiandrosterone sulphate (DHEA-S) to cross-react in the Abbott Architect testosterone assay.

Methods: Samples ( n = 81) from female patients who had DHEA-S, direct and extracted testosterone measured were examined retrospectively. Possible cross-reactivity was evaluated by spiking both testosterone free assay diluent and serum from a woman with exogenous DHEA-S.

Results: The retrospective patient analysis of the data demonstrated a relationship between interference and DHEA-S ( r = 0.78, P - 0.0001). Spiking studies revealed a linear relationship between added DHEA-S and the measured testosterone concentration. Consistent with the retrospective study, for every 1 μmol/L increase in DHEA-S, 0.23 nmol/L of testosterone was measured in the diluent. Spiking DHEA-S into serum gave only a 0.12 nmol/L increase in measured testosterone for every 1 μmol/L rise in DHEA-S.

Conclusion: DHEA-S cross-reacts in the Abbott Architect direct immunoassay for testosterone to a clinically significant effect. This cross-reactivity is likely to be responsible for much of the interference observed when measuring testosterone in women by this method.
Effect of plantain on therapeutic drug monitoring of digoxin and thirteen other common drugs
Authors: Dasgupta Amitava , Davis Bonnette , Wells Alice
Published: April 2006

Introduction: Plantain, a herbal remedy, has been reported to interfere with therapeutic drug monitoring of digoxin. We evaluated three commercially available plantain products for potential interference with therapeutic drug monitoring of digoxin and 13 other common drugs.

Method: Dry content of plantain capsule or plantain leaf was extracted with either methanol or ethanol:water (60:40 by volume), added to drug-free serum and apparent digoxin was measured by both fluorescence polarization immunoassay and microparticle enzyme immunoassay. Using immunoassays, we also measured apparent concentrations of 13 other drugs (tobramycin, procainamide, tricyclic antidepressants, quinidine, carbamazepine, phenytoin, theophylline, valproic acid, amikacin, gentamycin, phenobarbital, salicylate and acetaminophen [paracetamol]) due to the presence of plantain. In separate experiments, a serum pool prepared from patients receiving digoxin was further supplemented with plantain and observed digoxin values were compared with original digoxin concentration. The presence of any cardiac glycoside in plantain was also investigated using thin layer chromatography (TLC).

Results: We observed no apparent digoxin in the presence of plantain in serum. Moreover, when aliquots of digoxin serum pool were supplemented with various amounts of plantain, the observed digoxin concentrations in the presence of plantain compared well with original digoxin concentration. TLC analysis did not show the presence of either digoxin or digitoxin in plantain products studied. Moreover, plantain did not affect immunoassay results of the 13 other drugs studied.

Conclusions: The plantain products studied did not interfere with therapeutic drug monitoring of digoxin as well as 13 other commonly monitored drugs.
Categorical effects in children's colour search: A cross‐linguistic comparison
Authors: Daoutis Christine A. , Franklin Anna , Riddett Amy , Clifford Alexandra , Davies Ian R. L.
Published: April 2006

In adults, visual search for a colour target is facilitated if the target and distractors fall in different colour categories (e.g. Daoutis, Pilling, & Davies, in press). The present study explored category effects in children's colour search. The relationship between linguistic colour categories and perceptual categories was addressed by comparing native speakers of languages differing in the number of colour terms. Experiment 1 compared English and Kwanyama (Namibian) children aged 4 to 7 years on a visual search task, using target‐distractor pairs (blue‐green, blue‐purple, red‐pink) for which the Kwanyama did not have distinct names. The presence of a category advantage in the English, but not in the Kwanyama, suggested that linguistic boundaries may affect search performance. Experiment 2 examined visual search performance in the green‐yellow and the blue‐green region, in English and Himba (Namibian) 6‐year‐olds. The number of distractors was varied to assess search efficiency. Cross‐category search was more efficient than within‐category search in the English group, but this advantage was absent in the Himba. Increasing the number of distractors affected search speed in the English group, but not in the Himba. Overall, these findings suggest cross‐language differences in categorical effects on colour search, but also in the way the children performed the search. The nature of the category effect in search is discussed with respect to these findings.
Residues Determining the Binding Specificity of Uncompetitive Inhibitors to Tissue-Nonspecific Alkaline Phosphatase
Authors: Kozlenkov Alexey , Le Du Marie Helene , Cuniasse Philippe , Ny Tor , Hoylaerts Marc F , Millán José Luis
Published: April 2006

Recent data have pointed to TNALP as a therapeutic target for soft-tissue ossification abnormalities. Here, we used mutagenesis, kinetic analysis, and computer modeling to identify the residues important for the binding of known ALP inhibitors to the TNALP active site. These data will enable drug design efforts aimed at developing improved specific TNALP inhibitors for therapeutic use.

Introduction: We have shown previously that the genetic ablation of tissue-nonspecific alkaline phosphatase (TNALP) function leads to amelioration of soft-tissue ossification in mouse models of osteoarthritis and ankylosis (i.e., Enpp1−/− and ank/ank mutant mice). We surmise that the pharmacologic inhibition of TNALP activity represents a viable therapeutic approach for these diseases. As a first step toward developing suitable TNALP therapeutics, we have now clarified the residues involved in binding well-known uncompetitive inhibitors to the TNALP active site.

Materials and Methods: We compared the modeled 3D structure of TNALP with the 3D structure of human placental alkaline phosphatase (PLALP) and identified the residues that differ between these isozymes within a 12 Å radius of the active site, because these isozymes differ significantly in inhibitor specificity. We then used site-directed mutagenesis to substitute TNALP residues to their respective homolog in PLALP. In addition, we mutagenized most of these residues in TNALP to Ala and the corresponding residues in PLALP to their TNALP homolog. All mutants were characterized for their sensitivity toward the uncompetitive inhibitors l-homoarginine (L-hArg), levamisole, theophylline, and L-phenylalanine.

Results and Conclusions: We found that the identity of residue 108 in TNALP largely determines the specificity of inhibition by L-hArg. The conserved Tyr-371 is also necessary for binding of L-hArg. In contrast, the binding of levamisole to TNALP is mostly dependent on His-434 and Tyr-371, but not on residues 108 or 109. The main determinant of sensitivity to theophylline is His-434. Thus, we have clarified the location of the binding sites for all three TNALP inhibitors, and we have also been able to exchange inhibitor specificities between TNALP and PLALP. These data will enable drug design efforts aimed at developing improved, selective, and drug-like TNALP inhibitors for therapeutic use.
Secreted Frizzled-Related Protein-1 Inhibits RANKL-Dependent Osteoclast Formation
Authors: Häusler Karl D , Horwood Nicole J , Chuman Yoshiro , Fisher Jane L , Ellis Jennifer , Martin T John , Rubin Jeffrey S , Gillespie Matthew T
Published: April 2006

We determined that sFRP-1 mRNA was differentially expressed by osteoblast/stromal cell lines and that sFRP-1 neutralizing antibodies and siRNA complementary to sFRP-1 coding sequence enhanced, while recombinant sFRP-1 inhibited, osteoclast formation. In studying the mechanism of action for sFRP-1, we found that sFRP-1 could bind recombinant RANKL. These results suggest potential cross-talk between Wnt and RANKL pathways.

Introduction: Osteoclast formation in normal bone remodeling requires the presence of osteoblast lineage cells that express RANKL and macrophage-colony-stimulating factor (M-CSF), which interact with their cognate receptors on the osteoclast precursor. We identified secreted Frizzled-related protein-1 (sFRP-1), which is known to bind to Wnt and inhibit the Wnt signaling pathway, as an osteoblast-derived factor that impinges on osteoclast formation and activity.

Materials and Methods: Differential display of mRNA from osteoblast lineage cell lines established sFRP-1 to be highly expressed in an osteoclast supporting cell line. sFRP-1 expression in bone was determined by in situ hybridization, and the effects of sFRP-1 on osteoclast formation were determined using a neutralizing antibody, siRNA, for sFRP-1 and recombinant protein.

Results: In situ hybridization revealed sFRP-1 mRNA expression in osteoblasts and chondrocytes in murine bone. sFRP-1 mRNA expression could be elevated in calvarial primary osteoblasts in response to prostaglandin E2 (PGE2) or interleukin (IL)-11, whereas many other osteotropic agents (e.g., IL-1, IL-6, calcitrol, parathyroid hormone) were without any effect. In vitro assays of osteoclast formation established sFRP-1 to be an inhibitor of osteoclast formation. Neutralizing antibodies against sFRP-1 enhanced TRACP+ mononuclear and multinuclear osteoclast formation (3- and 2-fold, respectively) in co-cultures of murine osteoblasts with spleen cells, whereas siRNA complementary to sFRP-1 coding sequence significantly enhanced osteoclast formation in co-cultures of KUSA O (osteoblast/stromal cell line) and bone marrow cells, cultured in the presence of PGE2 and 1,25(OH)2 vitamin D3. Recombinant sFRP-1 dose-dependently inhibited osteoclast formation in osteoblast/spleen co-cultures, RANKL + M-CSF-treated splenic cultures, and RANKL-treated RAW264.7 cell cultures, indicating a direct action of sFRP-1 on hematopoietic cells. Consistent with this, sFRP-1 was found to bind to RANKL in ELISAs.

Conclusion: sFRP-1 is expressed by osteoblasts and inhibits osteoclast formation. While sFRP-1 activity might involve the blocking of endogenous Wnt signaling, our results suggest that, alternatively, it could be because of direct binding to RANKL. This study describes a new mechanism whereby osteoblasts regulate osteoclastogenesis through the expression and release of sFRP-1.
Skeletal Unloading Alleviates the Anabolic Action of Intermittent PTH(1–34) in Mouse Tibia in Association With Inhibition of PTH-Induced Increase in c-fos mRNA in Bone Marrow Cells
Authors: Tanaka Shinya , Sakai Akinori , Tanaka Masahiro , Otomo Hajime , Okimoto Nobukazu , Sakata Takeshi , Nakamura Toshitaka
Published: April 2006

We analyzed the effect of unloading by tail suspension on the anabolic action of intermittent PTH in the tibia of growing mice. Unloading alleviated the PTH-induced increase of bone formation and accelerated bone resorption, consequently reducing bone mass. Reduction of the PTH-induced anabolic actions on bone was associated with unloading, which was apparently related to suppression of c-fos mRNA expression in bone marrow.

Introduction: The effects of intermittent parathyroid hormone (PTH) administration on unloading bone have not been well elucidated at the cellular and molecular levels. We tested the effects of PTH on unloaded tibias of tail-suspended mice.

Materials and Methods: Eighty male C57BL/6J mice, 8 weeks of age, were divided into four groups with loading or unloading and administration of PTH (40 μg/kg body weight) or vehicle five times per week. Mice were killed at 8 or 15 days, and both tibias were obtained. Bone histomorphometry of the trabecular bone in the proximal tibia, development of osteogenic cells, and mRNA expression of osteogenic molecules in bone marrow cells were assessed.

Results and Conclusions: At 15 days of unloading, bone volume decreased in PTH-treated mice. The increase in the bone formation rate by PTH was depressed, and the osteoclast surface was thoroughly increased. The increase in alkaline phosphatase-positive colony-forming units-fibroblastic (CFU-f) colonies induced by PTH was maintained and that of TRACP+ multinucleated cells enhanced. The PTH-induced increase in c-fos mRNA was depressed, but the increases in Osterix and RANKL mRNA were maintained. Unloading promoted the PTH-associated osteoclastogenesis and seemed to delay the progression of osteogenic differentiation in association with reduction of the PTH-dependent increase of c-fos mRNA in bone marrow cells.
Long-Term Sensitivity of Uterus and Hypothalamus/Pituitary Axis to 17β-Estradiol Is Higher Than That of Bone in Rats
Authors: Erben Reinhold G , Brunner Katrin S , Breig Bianca
Published: April 2006

We examined the long-term sensitivity of uterus and bone to low-dose 17β-estradiol in a 4-month experiment in OVX rats and found that a dose of estradiol that fully protected against uterine atrophy did not protect against bone loss. Our results suggest higher estrogen sensitivity of the uterus compared with bone.

Introduction: Estrogen is essential for the function of reproductive tissues and for the normal acquisition and maintenance of bone mass in females. This study was designed to examine the long-term sensitivity of the uterus and bone to low-dose estrogen.

Materials and Methods: In preliminary experiments, we determined the lowest subcutaneous dose of 17β-estradiol able to fully protect against uterine atrophy in ovariectomized (OVX) rats. This dose was found to be 1.5 μg/kg, given five times per week. Subsequently, groups of sham-operated (SHAM) or OVX 6-month-old rats (n = 8 each) were subcutaneously injected with vehicle or 1.5 μg/kg 17β-estradiol five times per week. All animals were killed 4 months after surgery. Serum osteocalcin and urinary deoxypyridinoline were measured as biochemical markers of bone turnover. Bones were analyzed by bone histomorphometry and pQCT.

Results and Conclusions: Our study clearly showed that a dose of estradiol that restores physiological estradiol serum levels, fully maintains uterine weight in OVX rats at the SHAM control level, and suppresses serum follicle-stimulating hormone (FSH) by 67% relative to OVX vehicle controls does not provide significant protection against OVX-induced bone loss at different cancellous and cortical bone sites. We conclude that the long-term sensitivity of the uterus and the hypothalamus/pituitary axis to 17β-estradiol is higher than that of bone in rats.
Dissociation Between Global Markers of Bone Formation and Direct Measurement of Spinal Bone Formation in Osteoporosis
Authors: Frost Michelle L , Fogelman Ignac , Blake Glen M , Marsden Paul K , Cook Gary JR
Published: April 2006

Regional bone metabolism measured using18F-fluoride PET was assessed in 72 postmenopausal women classified as normal, osteopenic, or osteoporotic. Lower values of regional bone formation activity at the lumbar spine were seen in osteoporotic women, whereas global markers of bone formation were significantly increased.

Introduction: Evaluations of global bone remodeling have revealed that increased bone turnover is a feature of patients with osteoporosis. The noninvasive functional imaging technique of18F-fluoride positron emission tomography (PET) allows the direct quantitative assessment of bone metabolism at specific sites in the skeleton, including the clinically important site of the lumbar spine.

Materials and Methods: The aim of this study was to compare regional skeletal kinetics in 72 postmenopausal women (mean age, 61 years) classified as normal, osteopenic, or osteoporotic according to their BMD T score at the lumbar spine. Each woman had a dynamic PET scan of the lumbar spine after injection of 90 MBq18F-fluoride ion and measurements of biochemical markers of bone formation and resorption. The arterial plasma input function was derived using aorta arterial activity from the PET image. Time-activity curves were obtained by placing regions of interest over the lumbar vertebrae. A three-compartmental model was used to calculate bone blood flow (K1) and the net plasma clearance of tracer to bone mineral (Ki), reflecting regional osteoblastic activity (ml/minutes/ml). Rate constants k2, k3, and k4, which describe transport between plasma, the extracellular fluid (ECF) compartment, and the bone mineral compartment, were also measured.

Results: The net uptake of fluoride to the bone mineral compartment (Ki) was significantly lower in the osteoporotic group compared with both the osteopenic and normal groups, with a mean difference of 0.005 ml/minutes/ml (16.7%). The fraction of the tracer in the extravascular tissue space that underwent specific binding to bone mineral (k3/k2 + k3) was also significantly reduced in the women classified as osteoporotic. In contrast, levels of bone-specific alkaline phosphatase (BSALP) were significantly higher in the osteoporotic group compared with the normal and osteopenic groups by 35% and 27%, respectively. A significant negative correlation (r = −0.41) was observed between levels of BSALP and the fraction of the tracer that underwent specific binding to bone mineral.

Conclusion: Lower values of Ki, a measurement of regional bone formation activity, were seen in women classified as osteoporotic, whereas levels of BSALP, a measure of global bone formation, were significantly increased. These findings are suggestive of increased global skeletal bone turnover in women with postmenopausal osteoporosis but with relatively reduced regional bone formation at the predominantly trabecular site of the lumbar spine.
Degradation of the Organic Phase of Bone by Osteoclasts: A Secondary Role for Lysosomal Acidification
Authors: Henriksen Kim , Sørensen Mette G , Nielsen Rasmus H , Gram Jeppe , Schaller Sophie , Dziegiel Morten H , Everts Vincent , Bollerslev Jens , Karsdal Morten A
Published: April 2006

Osteoclasts degrade bone matrix by secretion of hydrochloric acid and proteases. We studied the processes involved in the degradation of the organic matrix of bone in detail and found that lysosomal acidification is involved in this process and that MMPs are capable of degrading the organic matrix in the absence of cathepsin K.

Introduction: Osteoclasts resorb bone by secretion of acid by the vacuolar H+-adenosine triphosphatase (V-ATPase) and the chloride channel ClC-7, followed by degradation of the matrix, mainly collagen type I, by cathepsin K and possibly by matrix metalloproteinases (MMPs). However, the switch from acidification to proteolysis and the exact roles of both the ion transporters and the proteinases still remain to be studied.

Materials and Methods: We isolated CD14+ monocytes from human peripheral blood from either controls or patients with autosomal dominant osteopetrosis type II (ADOII) caused by defective ClC-7 function and cultured them in the presence of RANKL and macrophage-colony stimulating factor (M-CSF) to generate osteoclasts. We decalcified cortical bovine bone slices and studied the osteoclasts with respect to morphology, markers, and degradation of the decalcified matrix in the presence of various inhibitors of osteoclast acidification and proteolysis, using normal calcified bone as a reference.

Results: We found that ADOII osteoclasts not only have reduced resorption of the calcified matrix, but also 40% reduced degradation of the organic phase of bone. We found that both acidification inhibitors and cathepsin K inhibitors reduced degradation of the organic matrix by 40% in normal osteoclasts, but had no effect in the ADOII osteoclasts. Furthermore, we showed that inhibition of MMPs leads to a 70% reduction in the degradation of the organic bone matrix and that MMPs and cathepsin K have additive effects. Finally, we show that osteoclastic MMPs mediate release of the carboxyterminal telopeptide of type I collagen (ICTP) fragment in the absence of cathepsin K activity, and therefore, to some extent, are able to compensate for the loss of cathepsin K activity.

Conclusions: These data clearly show that osteoclastic acidification of the lysosomes plays a hitherto nonrecognized role in degradation of the organic matrix. Furthermore, these data shed light on the complicated interplay between acidification dependent and independent proteolytic processes, mediated by cathepsin K and the MMPs, respectively.