bluuebunny/crossref_metadata_2025 huggingface dataset:
Truncated forms of viral VP2 proteins fused to EGFP assemble into fluorescent parvovirus-like particles
Authors: Gilbert Leona , Toivola Jouni , Välilehto Outi , Saloniemi Taija , Cunningham Claire , White Daniel , Mäkelä Anna R , Korhonen Eila , Vuento Matti , Oker-Blom Christian
Published: December 2006

Fluorescence correlation spectroscopy (FCS) monitors random movements of fluorescent molecules in solution, giving information about the number and the size of for example nano-particles. The canine parvovirus VP2 structural protein as well as N-terminal deletion mutants of VP2 (-14, -23, and -40 amino acids) were fused to the C-terminus of the enhanced green fluorescent protein (EGFP). The proteins were produced in insect cells, purified, and analyzed by western blotting, confocal and electron microscopy as well as FCS. The non-truncated form, EGFP-VP2, diffused with a hydrodynamic radius of 17 nm, whereas the fluorescent mutants truncated by 14, 23 and 40 amino acids showed hydrodynamic radii of 7, 20 and 14 nm, respectively. These results show that the non-truncated EGFP-VP2 fusion protein and the EGFP-VP2 constructs truncated by 23 and by as much as 40 amino acids were able to form virus-like particles (VLPs). The fluorescent VLP, harbouring VP2 truncated by 23 amino acids, showed a somewhat larger hydrodynamic radius compared to the non-truncated EGFP-VP2. In contrast, the construct containing EGFP-VP2 truncated by 14 amino acids was not able to assemble into VLP-resembling structures. Formation of capsid structures was confirmed by confocal and electron microscopy. The number of fluorescent fusion protein molecules present within the different VLPs was determined by FCS. In conclusion, FCS provides a novel strategy to analyze virus assembly and gives valuable structural information for strategic development of parvovirus-like particles.
Is mammalian chromosomal evolution driven by regions of genome fragility?
Authors: Ruiz-Herrera Aurora , Castresana Jose , Robinson Terence J
Published: December 2006

Background

A fundamental question in comparative genomics concerns the identification of mechanisms that underpin chromosomal change. In an attempt to shed light on the dynamics of mammalian genome evolution, we analyzed the distribution of syntenic blocks, evolutionary breakpoint regions, and evolutionary breakpoints taken from public databases available for seven eutherian species (mouse, rat, cattle, dog, pig, cat, and horse) and the chicken, and examined these for correspondence with human fragile sites and tandem repeats.

Results

Our results confirm previous investigations that showed the presence of chromosomal regions in the human genome that have been repeatedly used as illustrated by a high breakpoint accumulation in certain chromosomes and chromosomal bands. We show, however, that there is a striking correspondence between fragile site location, the positions of evolutionary breakpoints, and the distribution of tandem repeats throughout the human genome, which similarly reflect a non-uniform pattern of occurrence.

Conclusion

These observations provide further evidence that certain chromosomal regions in the human genome have been repeatedly used in the evolutionary process. As a consequence, the genome is a composite of fragile regions prone to reorganization that have been conserved in different lineages, and genomic tracts that do not exhibit the same levels of evolutionary plasticity.
Generation of medaka gene knockout models by target-selected mutagenesis
Authors: Taniguchi Yoshihito , Takeda Shunichi , Furutani-Seiki Makoto , Kamei Yasuhiro , Todo Takeshi , Sasado Takao , Deguchi Tomonori , Kondoh Hisato , Mudde Josine , Yamazoe Mitsuyoshi , Hidaka Masayuki , Mitani Hiroshi , Toyoda Atsushi , Sakaki Yoshiyuki , Plasterk Ronald HA , Cuppen Edwin
Published: December 2006

We have established a reverse genetics approach for the routine generation of medaka (Oryzias latipes) gene knockouts. A cryopreserved library of N-ethyl-N-nitrosourea (ENU) mutagenized fish was screened by high-throughput resequencing for induced point mutations. Nonsense and splice site mutations were retrieved for the Blm, Sirt1, Parkin and p53 genes and functional characterization of p53 mutants indicated a complete knockout of p53 function. The current cryopreserved resource is expected to contain knockouts for most medaka genes.
Controversies in the Management of Clinical Stage I Testis Cancer
Authors: de Wit Ronald , Fizazi Karim
Published: December 2006

During the last two decades, definitive primary treatments and surveillance with definitive treatment deferred until relapse have demonstrated 98% to 99% cure rates in patients with stage I testis cancer, and these options have obtained firm positions in standard management. The development of optimal management strategies in various countries were at least partly guided by available surgical expertise in retroperitoneal lymph node dissection in the United States, and easy access to reference hospitals in densely populated countries in Western Europe that facilitated close surveillance programs; hence, treatment preferences differ on the two sides of the Atlantic. The success of both approaches is highly dependent on the skills of the practitioner, particularly of surgery and of scrutinized surveillance. As a result, local expertise and familiarity with a chosen modality has strengthened over the years, and investigators have been reluctant to embark on randomized trials designed to compare one modality with another. Such expertise with one particular technique, with the other approach being less familiar territory, has created controversy, because both physicians and patients seek evidence-based data coming from randomized clinical trials on which to make management decisions. Moreover, the reduced risk of relapse resulting from the use of radiotherapy or carboplatin in stage I seminoma and of cisplatin-based chemotherapy in stage I nonseminoma must be balanced against the potential long-term adverse effects in this population of patients with a normal life expectancy. The purpose of this review is to present the currently available data and discuss the merits and the disadvantages of the various approaches, yielding to the possible conclusion that all options appear to be equal in terms of efficacy, but that modality-associated adverse effects differ.
Late Relapses of Germ Cell Malignancies: Incidence, Management, and Prognosis
Authors: Oldenburg Jan , Martin Jarad M. , Fosså Sophie D.
Published: December 2006

Late relapses of malignant germ cell tumors (MGCTs) are rare and occur, by definition, 2 years or later after successful treatment. They represent a major challenge of today's treatment of MGCTs. Because of the rarity and heterogeneity of late relapses, many aspects of their main characteristics remain obscure. We present relevant literature on relapsing MGCTs to highlight the following issues: incidence, impact of initial treatment on the subsequent risk of late relapse, treatment, and survival. In a pooled analysis, the incidence is 1.4% and 3.2% in seminoma and nonseminoma patients, respectively. The predominant site of relapse is the retroperitoneal space in both histologic types. The initial treatment appears to be important for the risk and localization of late relapses. The treatment of late relapses should be based on a representative presalvage biopsy and includes radical surgery and salvage chemotherapy in most cases. Five-year cancer-specific survival is above 50% in the recent large series and reaches 100% in case of single-site teratoma. Diagnosis and treatment of late-relapsing MGCT patients is challenging and should be performed in experienced centers only. Referral of late-relapsing patients to high-volume institutions ensures the best chances of cure and enables increasing understanding of tumor biology and the clinical course of these patients.
Molecular Pathways in Invasive Bladder Cancer: New Insights Into Mechanisms, Progression, and Target Identification
Authors: Mitra Anirban P. , Datar Ram H. , Cote Richard J.
Published: December 2006

Papillary and invasive cancers of the urinary bladder appear to evolve and progress through distinct molecular pathways. Invasion in bladder cancer forebodes a graver prognosis, and these tumors are generally characterized by alterations in the p53 and retinoblastoma (RB) pathways that normally regulate the cell cycle by interacting with the Ras–mitogen activated protein kinase signal transduction pathway. Tumor angiogenesis further contributes to the neoplastic growth by providing a constant supply of oxygen and nutrients. Distinct epigenetic and genetic events characterize the interplay between the molecules involved in these pathways, thus affording their use as indicators of prognosis. Efforts are now underway to construct molecular panels comprising multiple markers that can serve as more robust predictors of outcome. While clinical trials for targeted chemotherapy for bladder cancer have commenced, novel genetic and pharmacologic agents that can target pathway-specific molecules are currently under development. The next generation of clinical management for urothelial carcinoma will witness the use of multimarker panels for prognostic prediction and combination therapy directed at novel molecular targets for treatment.
Chemotherapy for Advanced Germ Cell Tumors
Authors: Kondagunta G. Varuni , Motzer Robert J.
Published: December 2006

Purpose

Germ cell tumors constitute the most curable of all cancers. Standard treatment of previously untreated and treated patients has evolved on the basis of prospective clinical trials and prognostic factors. This review summarizes the prognostic criteria on which treatment decisions may be based, and outlines the current treatment approaches.

Patients and Methods

Randomized and nonrandomized trials of first-line, salvage, and palliative therapy and the role of surgery after chemotherapy were reviewed. In the treatment of previously untreated patients, emphasis was placed on interpretation of data of trials according to the International Germ Cell Cancer Collaborative Group model, which has evolved into a universally accepted classification algorithm for determining appropriate risk-directed chemotherapy. This system permits treatment choices based on the balance between benefit and toxicity and allows comparison of results across multiple clinical trials.

Results

Standard therapy for good-risk patients is four cycles of etoposide plus cisplatin or three cycles of cisplatin, etoposide plus bleomycin (BEP × 3); both approaches cure approximately 90% of patients. After chemotherapy and normalization of markers, patients should generally undergo resection of residual masses. Approximately 75% of intermediate-risk and 45% of poor-risk patients group achieve a durable complete response with BEP × 4. Potentially curative options in the salvage setting include ifosfamide plus cisplatin-containing standard dose therapy and high-dose carboplatin plus stem-cell rescue. Surgery remains an essential component of care.

Conclusion

Curative therapy exists even in patients with resistant disease, and treatment choices can be based on established clinical criteria. Serum tumor markers and surgery after chemotherapy have essential roles in patient management.
Molecular Biology of Renal Cell Cancer and the Identification of Therapeutic Targets
Authors: Iliopoulos Othon
Published: December 2006

Renal cell cancer (RCC) is a heterogeneous disease consisting of different histologic types. Major advances have been accomplished during the last 15 years in our understanding of the genetic events that initiate RCC. These advances were greatly facilitated by meticulous clinical description and registration of patients with familial predisposition to RCC. The cloning of the susceptibility genes that underline familial predisposition to RCC has offered entry points into the signaling pathways that are also deregulated in sporadic RCC. Biochemical studies of these signaling pathways and target validation experiments have already culminated in the discovery and clinical application of small molecules with promising activity in RCC. In this article, we highlight the molecular genetic features of RCC that are more directly related to identification and validation of promising targets for molecular therapy.
Ceramide-Activated Protein Phosphatase Involvement in Insulin Resistance via Akt, Serine/Arginine-Rich Protein 40, and Ribonucleic Acid Splicing in L6 Skeletal Muscle Cells
Authors: Ghosh Nilanjan , Patel Niketa , Jiang Kun , Watson James E. , Cheng Jin , Chalfant Charles E. , Cooper Denise R.
Published: December 2006

Elevated TNFα levels are associated with insulin resistance, but the molecular mechanisms linking cytokine signaling to impaired insulin function remain elusive. We previously demonstrated a role for Akt in insulin regulation of protein kinase CβII alternative splicing through phosphorylation of serine/arginine-rich protein 40, a required mechanism for insulin-stimulated glucose uptake. We hypothesized that TNFα attenuated insulin signaling by dephosphorylating Akt and its targets via ceramide-activated protein phosphatase. Western blot analysis of L6 cell lysates demonstrated impaired insulin-stimulated phosphorylation of Akt, serine/arginine-rich protein 40, and glycogen synthase kinase 3β in response to TNFα and the short chain C6 ceramide analog. TNFα increased serine/threonine phosphatase activity of protein phosphatase 1 (PP1) in response to C6, but not insulin, suggesting a ceramide-specific effect. Myriocin, an inhibitor of de novo ceramide synthesis, blocked stimulation of the PP1 activity. Ceramide species measurement by liquid chromatography-mass spectrometry showed consistent increases in C24:1 and C16 ceramides. Effects of TNFα and C6 on insulin-stimulated phosphorylation of glycogen synthase kinase 3β were prevented by myriocin and tautomycin, a PP1 inhibitor, further implicating a de novo ceramide-PP1 pathway. Alternative splicing assays demonstrated that TNFα abolished insulin-mediated inclusion of the protein kinase CβII exon. Collectively, our work demonstrates a role for PP1-like ceramide-activated protein phosphatase in mediating TNFα effects blocking insulin phosphorylation cascades involved in glycogen metabolism and alternative splicing.
Postnatal Body Growth Is Dependent on the Transcription Factors Signal Transducers and Activators of Transcription 5a/b in Muscle: A Role for Autocrine/Paracrine Insulin-Like Growth Factor I
Authors: Klover Peter , Hennighausen Lothar
Published: December 2006

The transcription factors signal transducers and activators of transcription (STAT)5a and STAT5b (STAT5) are essential mediators of many actions of GH, including transcription of the IGF-I gene. Here, we present evidence that skeletal muscle STAT5 is important for postnatal growth and suggest that this is conveyed by the production of localized IGF-I. To investigate the role of STAT5 signaling in skeletal muscle, mice with a skeletal-muscle-specific deletion of the Stat5a and Stat5b genes (Stat5MKO mice) were used. IGF-I mRNA levels were reduced by 60% in muscle tissue of these mice. Despite only a 15% decrease in circulating IGF-I, 8-wk-old male Stat5MKO mice displayed approximately 20% reduction in body weight that was accounted for by a reduction in lean mass. The skeletons of Stat5MKO mice were found to be smaller than controls, indicating the growth defect was not restricted to skeletal muscle. These results demonstrate an as yet unreported critical role for STAT5 in skeletal muscle for local IGF-I production and postnatal growth and suggest the skeletal muscle as a major site of GH action.
Two-person red-and-black games with bet-dependent win probability functions
Authors: Chen May-Ru , Hsiau Shoou-Ren
Published: December 2006

In this paper a two-person red-and-black game is investigated. We suppose that, at every stage of the game, player I's win probability, f, is a function of the ratio of his bet to the sum of both players' bets. Two results are given: (i) if f is convex then a bold strategy is optimal for player I when player II plays timidly; and (ii) if f satisfies f(s)f(t) ≤ f(st) then a timid strategy is optimal for player II when player I plays boldly. These two results extend two formulations of red-and-black games proposed by Pontiggia (2005), and also provide a sufficient condition to ensure that the profile (bold, timid) is the unique Nash equilibrium for players I and II. Finally, we give a counterexample to Pontiggia's conjecture about a proportional N-person red-and-black game.
Central limit theorems for generalized Pólya urn models
Authors: Higueras I. , Moler J. , Plo F. , San Miguel M.
Published: December 2006

In this paper we obtain central limit theorems for generalized Pólya urn models with L ≥ 2 colors where one out of K different replacements (actions) is applied randomly at each step. Each possible action constitutes a row of the replacement matrix, which can be nonsquare and random. The actions are chosen following a probability distribution given by an arbitrary function of the proportions of the balls of different colors present in the urn. Moreover, under the same hypotheses it is proved that the covariance matrix of the asymptotic distribution is the solution of a Lyapunov equation, and a procedure is given to obtain the covariance matrix in an explicit form. Some applications of these results to random trees and adaptive designs in clinical trials are also presented.
Critical sizing of LRU caches with dependent requests
Authors: Jelenković Predrag R. , Radovanović Ana , Squillante Mark S.
Published: December 2006

It was recently proved by Jelenković and Radovanović (2004) that the least-recently-used (LRU) caching policy, in the presence of semi-Markov-modulated requests that have a generalized Zipf's law popularity distribution, is asymptotically insensitive to the correlation in the request process. However, since the previous result is asymptotic, it remains unclear how small the cache size can become while still retaining the preceding insensitivity property. In this paper, assuming that requests are generated by a nearly completely decomposable Markov-modulated process, we characterize the critical cache size below which the dependency of requests dominates the cache performance. This critical cache size is small relative to the dynamics of the modulating process, and in fact is sublinear with respect to the sojourn times of the modulated chain that determines the dependence structure.
Reversibility and entropy production of inhomogeneous Markov chains
Authors: Ge Hao , Jiang Da-Quan , Qian Min
Published: December 2006

In this paper we introduce the concepts of instantaneous reversibility and instantaneous entropy production rate for inhomogeneous Markov chains with denumerable state spaces. The following statements are proved to be equivalent: the inhomogeneous Markov chain is instantaneously reversible; it is in detailed balance; its entropy production rate vanishes. In particular, for a time-periodic birth-death chain, which can be regarded as a simple version of a physical model (Brownian motors), we prove that its rotation number is 0 when it is instantaneously reversible or periodically reversible. Hence, in our model of Markov chains, the directed transport phenomenon of Brownian motors can occur only in nonequilibrium and irreversible systems.
On the total reward variance for continuous-time Markov reward chains
Authors: Van Dijk Nico M. , Sladký Karel
Published: December 2006

As an extension of the discrete-time case, this note investigates the variance of the total cumulative reward for continuous-time Markov reward chains with finite state spaces. The results correspond to discrete-time results. In particular, the variance growth rate is shown to be asymptotically linear in time. Expressions are provided to compute this growth rate.
Exact distributions for reward functions on semi-Markov and Markov additive processes
Authors: Stefanov Valeri T.
Published: December 2006

The distribution theory for reward functions on semi-Markov processes has been of interest since the early 1960s. The relevant asymptotic distribution theory has been satisfactorily developed. On the other hand, it has been noticed that it is difficult to find exact distribution results which lead to the effective computation of such distributions. Note that there is no satisfactory exact distribution result for rewards accumulated over deterministic time intervals [0, t], even in the special case of continuous-time Markov chains. The present paper provides neat general results which lead to explicit closed-form expressions for the relevant Laplace transforms of general reward functions on semi-Markov and Markov additive processes.
On the asymptotic distribution of the discrete scan statistic
Authors: Boutsikas Michael V. , Koutras Markos V.
Published: December 2006

The discrete scan statistic in a binary (0-1) sequence of n trials is defined as the maximum number of successes within any k consecutive trials (n and k, n ≥ k, being two positive integers). It has been used in many areas of science (quality control, molecular biology, psychology, etc.) to test the null hypothesis of uniformity against a clustering alternative. In this article we provide a compound Poisson approximation and subsequently use it to establish asymptotic results for the distribution of the discrete scan statistic as n, k → ∞ and the success probability of the trials is kept fixed. An extreme value theorem is also provided for the celebrated Erdős-Rényi statistic.
Strategic equilibrium versus global optimum for a pair of competing servers
Authors: Avi-Itzhak Benjamin , Golany Boaz , Rothblum Uriel G.
Published: December 2006

Christ and Avi-Itzhak (2002) analyzed a queueing system with two competing servers who determine their service rates so as to optimize their individual utilities. The system is formulated as a two-person game; Christ and Avi-Itzhak proved the existence of a unique Nash equilibrium which is symmetric. In this paper, we explore globally optimal solutions. We prove that the unique Nash equilibrium is generally strictly inferior to a globally optimal solution and that optimal solutions are symmetric and require the servers to adopt service rates that are smaller than those occurring in equilibrium. Furthermore, given a symmetric globally optimal solution, we show how to impose linear penalties on the service rates so that the given optimal solution becomes a unique Nash equilibrium. When service rates are not observable, we show how the same effect is achieved by imposing linear penalties on a corresponding signal.
A relation between positive dependence of signal and the variability of conditional expectation given signal
Authors: Mizuno Toshihide
Published: December 2006

Let S1 and S2 be two signals of a random variable X, where G1(s1 ∣ x) and G2(s2 ∣ x) are their conditional distributions given X = x. If, for all s1 and s2, G1(s1 ∣ x) - G2(s2 ∣ x) changes sign at most once from negative to positive as x increases, then the conditional expectation of X given S1 is greater than the conditional expectation of X given S2 in the convex order, provided that both conditional expectations are increasing. The stochastic order of the sufficient condition is equivalent to the more stochastically increasing order when S1 and S2 have the same marginal distribution and, when S1 and S2 are sums of X and independent noises, it is equivalent to the dispersive order of the noises.
Shapes of stationary autocovariances
Authors: Lund Robert , Zhao Ying , Kiessler Peter C.
Published: December 2006

This note introduces shape orderings for stationary time series autocorrelation and partial autocorrelation functions and explores some of their convergence rate ramifications. The shapes explored include decreasing hazard rate and new better than used, orderings that are familiar from stochastic processes settings. Time series models where these shapes arise are presented. The shapes are used to obtain explicit geometric convergence rates for mean squared errors of one-step-ahead forecasts.
The Role of Private Decentralized Institutions in Sustaining Industry Self-Regulation
Authors: Lenox Michael J.
Published: December 2006

An emerging body of institutional scholarship emphasizes the role played by private decentralized institutions in facilitating collective action among organizations. However, these institutions often suffer from free riding and opportunism. This may lead firms to exit the institution and eventually cause its collapse. In this paper, we explore how private decentralized institutions may be sustained despite these problems. We focus on one form of private decentralized institution—trade association–sponsored industry self-regulatory programs. We advance two alternative hypotheses to explain the sustained existence of industry self-regulatory institutions: (a) Firms participate to gain a participation-contingent benefit, and (b) firms participate to maintain a generally beneficial institution. Using a 10-year panel of data from the chemical industry, we find evidence consistent with the latter hypothesis for at least one prominent example of self-regulation. Our findings have implications for both specific models of industry self-regulation and general theories of collective action.
Stages of the Recruitment Process and the Referrer’s Performance Effect
Authors: Yakubovich Valery , Lup Daniela
Published: December 2006

Although the existing theory predicts that a referral’s chances of being hired increase with the job performance of the referrer, no empirical evidence is available to support this claim. To address this discrepancy, we decompose the recruitment process into objective selection, subjective selection, and self-selection and theorize that the likelihood of passing a particular recruitment stage increases with the performance of the referrer under objective selection and self-selection, but remains undetermined at a stage of subjective selection. Our analysis of unique comprehensive data on online recruitment of sales agents in a virtual call center supports these arguments. The effectiveness of personnel as a recruitment channel varies with the type of the recruitment stage and performance of the referrer. When the firm evaluates candidates by an objective criterion, the advantage of a referral increases with the performance of his or her referrer; those referred by relatively high-performing workers are significantly better than the applicants who learned about the job from Internet ads. When job candidates self-select into the next stage of the online application process, the referral of any agent is more likely to continue than a nonreferral, and this likelihood increases with the performance of the referrer. On a subjective stage, the outcome is contingent on the intricacies of the recruitment process. In our case, an applicant’s chances of being hired increase with the performance of his or her referrer because the firm rejects the referrals of low-performing workers at a higher rate than it does nonreferrals, while it treats equally the referrals of high-performing workers and nonreferrals. The study’s contributions to the literature on social networks in labor markets are discussed.
Organizational Identity and Firm Performance: What Happens When Leaders Disagree About “Who We Are?”
Authors: Voss Zannie Giraud , Cable Daniel M. , Voss Glenn B.
Published: December 2006

This study examines how the organizational success of 113 nonprofit professional theatres was affected when two top leaders responsible for different parts of the organization held divergent views about organizational identity. Focusing on five values that differentiate theatres, we find that leaders’ disagreement about organizational identity was related to lower ticket revenues and lower net income, and that organizational performance was lowest when disagreement about identity was extreme. Although some findings suggest that minor identity disagreement between leaders may not hurt organizations, results generally support the perspective that leaders should actively promote a single identity.
Increased Plasma Concentrations of Antiprothrombin Antibodies in Women with Recurrent Spontaneous Abortions
Authors: Sabatini Laura , Torricelli Michela , Scaccia Valentina , Fineschi Daniela , Pescaglini Monica , Gasparri Laura , Florio Pasquale , Petraglia Felice
Published: December 2006

Background: Antiphospholipid antibodies are associated with recurrent fetal loss, but the clinical relevance of antiprothrombin (aPT) antibodies remains controversial. This study was designed to evaluate the relationship of plasma concentrations of aPT antibodies (IgG, IgM, and IgA isotypes) and recurrent spontaneous abortion (RSA) not associated with antiphospholipid-antibody syndrome.

Methods: In this retrospective case–control study, we measured plasma aPT antibodies in 100 pregnant women at 8–12 weeks of gestation who had histories of recurrent abortion not associated with antiphospholipid-antibody syndrome. The controls were 200 healthy gestational-age–matched women with uncomplicated gestations.

Results: The mean (SD) plasma aPT concentrations were significantly (P <0.001) higher in women with histories of recurrent abortion than in healthy controls [7.97 (0.79) and 2.08 (0.07) kU/L]. Similarly, the concentrations of IgM aPT were significantly (P <0.001) higher in patients than in controls [5.73 (0.85) and 1.83 (0.05) kU/L]. No differences were found for IgA aPT (P = 0.358).

Conclusions: High concentrations of aPT antibodies (IgG and IgM isotypes) are associated with pregnancy loss in women with RSA. We suggest that the antibodies may have a relevant role in the etiology and pathogenesis of the condition.
Drug Screening of Preserved Oral Fluid by Liquid Chromatography–Tandem Mass Spectrometry
Authors: Øiestad Elisabeth Leere , Johansen Unni , Christophersen Asbjorg Solberg
Published: December 2006

Background: Oral fluid is an alternative matrix with potential applications in road-side drug screening, work-place testing, drug treatment programs, and epidemiological surveys. Development of methods for extensive drug screening in oral fluid is warranted.

Methods: We developed a liquid chromatography– tandem mass spectrometry (LC-MS/MS) method for drug screening of preserved oral fluid collected with the Intercept® collection device. Samples were prepared by liquid–liquid extraction with ethylacetate/heptane (4:1). LC-separation was achieved with an Atlantis dC18-column (2.1 × 50 mm, 3 μm particle). Mass detection was performed by positive ion mode electrospray LC-MS/MS and included the following drugs/metabolites: morphine, 6-monoacetylmorphine, codeine, buprenorphine, methadone, amphetamine, methamphetamine, 3,4-methylenedioxymethamphetamine, 3,4-methylenedioxyamphetamine, 3,4-methylenedioxyethylamphetamine, cocaine, benzoylecgonine, Δ-9-tetrahydrocannabinol, lysergic acid diethylamide, alprazolam, bromazepam, clonazepam, 7-aminoclonazepam, diazepam, N-desmethyldiazepam, 3-OH-diazepam, fenazepam, flunitrazepam, 7-aminoflunitrazepam, lorazepam, nitrazepam, 7-aminonitrazepam, oxazepam, zopiclone, zolpidem, carisoprodol, and meprobamat.

Results: Screening of 32 drugs was performed with a run time of 14 min. Within- and between-day relative CVs varied from 2.0% to 31.8% and from 3.6% to 39.1%, respectively. Extraction recoveries were >50% except for morphine (30%) and benzoylecgonine (0.2%). The concentrations of the lowest calibrator were 1 nmol/L (0.28 μg/L) to 500 nmol/L (68 μg/L), depending on the drug.

Conclusion: The method allowed rapid and sensitive oral fluid screening for the most commonly abused drugs in Norway and will be used for a road-side survey of drug use in normal traffic.
Mitochondrial DNA Mutation Detection by Electrospray Mass Spectrometry
Authors: Jiang Yun , Hall Thomas A , Hofstadler Steven A , Naviaux Robert K
Published: December 2006

Background: Mitochondrial DNA (mtDNA) mutations cause a large spectrum of clinically important neurodegenerative, neuromuscular, cardiovascular, and endocrine disorders. We describe the novel application of electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry (ESI-FTICR MS) to the rapid and accurate identification of pathogenic mtDNA variants.

Methods: In a blinded study, we used ESI-FTICR MS to analyze 24 unrelated samples of total cellular DNA containing 12 mtDNA variants and compared the results with those obtained by conventional PCR-restriction fragment length polymorphism (PCR-RFLP) analysis and gel electrophoresis.

Results: From the 24-sample blinded panel, we correctly identified 12 of the samples as bearing an mtDNA variant and found the remaining 12 samples to have no pathogenic variants. The correlation coefficient between the 2 methods for mtDNA variant detection was 1.0; there were no false positives or false negatives in this sample set. In addition, the ESI-FTICR method identified 4 single-nucleotide polymorphisms (SNP) that had previously been missed by standard PCR-RFLP analysis.

Conclusions: ESI-FTICR MS is a rapid, sensitive, and accurate method for the identification and quantification of mtDNA mutations and SNPs.
Experimental Study on Fracture Behavior of a Silty Clay
Authors: Wang Jun-Jie , Zhu Jun-Gao , Chiu C. F. , Chai He-Jun
Published: December 2006

Fracture toughness KIC and KIIC are two important parameters in fracture mechanics. In this paper, a device was designed to perform improved three-point and four-point unsymmetrical bending tests on soil beams. The device can easily remove the influence of specimen self-weight on the test results. Using the improved methods and the device, KIC, KIIC, and mixed mode I-II stress intensity factors KI and KII at fracture failure of a compacted silty clay were investigated. Test results indicated that the fracture of the silty clay exhibits linear and elastic behavior. The influences of dry density, water content, and preconsolidation pressure of specimens on parameter KIC were investigated. It was found that the classical mixed mode I-II fracture failure theories such as the strain energy density factor theory, maximum circumferential stress theory, and energy release rate theory cannot adequately predict the fracture failure behavior of the specimens, but a circle agrees well with the test results. The directions of crack growing in the specimens under mode II and mixed mode I-II loading conditions were expressed by a linear function of KI/KII.
Intensifiers of adjectives in German
Authors: Claudi Ulrike
Published: December 2006

Work on grammaticalization that has bee carried out in the course of the last decades has shown that there are some domains of grammatical structure that are more conservative while others are more innovative. The present paper deals with one domain that appears to belong to the most innovative parts of grammar, namely that of adjectival intensifiers. Looking in more detail into the nature and the use of such intensifiers in one particular language, which is German, the paper aims to show that intensifiers in particular and hyperbolic language in general are a culture-dependent phenomenon, and that their study can offer new insights into the mechanism underlying the nature of grammatical change.
Constructions of equative comparison
Authors: Henkelmann Peter
Published: December 2006

The morphosyntactic structures of equative constructions, which represent one class of comparison constructions (alongside positive, comparative, and superlative con-structions), are cross-linguistically quite diverse. In the present investigation, which is based on a convenience sample consisting of 25 languages, constructions of this kind are exemplified, analyzed, and classified. Underlying these different morphosyntactic structures are various semantic structures, which are classified and illustrated, too.
Smoking and susceptibility to thyroid cancer: an inverse association with CYP1A1 allelic variants
Authors: Bufalo Natassia E , Leite Janaina L , Guilhen Ana C T , Morari Elaine C , Granja Fabiana , Assumpcao Ligia V M , Ward Laura S
Published: December 2006

In contrast to most human malignancies, epidemiologic studies have frequently reported a reduced risk of differentiated thyroid cancer in tobacco consumers. Cytochrome P4501A1 (CYP1A1) gene variants may be related to an increased capacity to activate polycyclic aromatic hydrocarbons, producing highly reactive electrophilic intermediates that might damage DNA. Hence, the germline inheritance of a wild-type CYP1A1 gene may decrease the susceptibility for thyroid cancer. The present study was designed to investigate CYP1A1 (m1 and m2) role in thyroid tumorigenesis and its connection with GSTM1, GSTT1, GSTP1, GSTO1, and codon 72 of p53 genotypes. A total of 248 patients with thyroid nodules, including 67 benign goiters, 13 follicular adenomas, 136 papillary carcinomas, and 32 follicular carcinomas, and 277 controls with similar ethnic backgrounds were interviewed on their lifetime dietary and occupational histories, smoking habit, previous diseases, and other anamnestic data. DNA was extracted from a blood sample and submitted to PCR-restriction fragment length polymorphism assays. The wild-type CYP1A1m1 genotype was more frequent among papillary carcinoma patients (74.26%) than in the control population (62.45%; P = 0.0147), reducing the risk for this type of cancer (odds ratio = 0.564; 95% confidence interval = 0.357–0.894). A multiple logistic regression analysis showed an inverse correlation between cigarette smoking (P = 0.0385) and CYP1A1 germline inheritance (P = 0.0237) with the susceptibility to papillary carcinomas. We were not able to find any correlation between smoking, clinical features, parameters of aggressiveness at diagnosis or during follow-up, and any of the GST or CYP genotypes considered separately or in different combinations. We suggest that CYP1A1 genotype might be associated with the reported reduced risk to papillary carcinomas among smokers.
Androgen receptor corepressors and prostate cancer
Authors: Burd Craig J , Morey Lisa M , Knudsen Karen E
Published: December 2006

The androgen receptor (AR) mediates the effects of male steroid hormones (androgens) and contributes to a wide variety of physiological and pathophysiological conditions. As such, the regulatory mechanisms governing AR activity are of high significance. Concerted effort has been placed on delineating the mechanisms that control AR activity in prostate cancer, as AR is required for survival and proliferation in this tumor type. Moreover, AR is the central therapeutic target for metastatic prostate cancers, and recurrent tumors evade therapy by restoring AR activity. It is increasingly apparent that AR cofactors which modulate receptor activity can contribute to prostate cancer growth or progression, and this has been particularly well established for AR coactivators. The present review is focused on the role of AR corepressors in governing androgen action, with a specific emphasis on their activities in prostate cancer.
Identification of novel genes that co-cluster with estrogen receptor alpha in breast tumor biopsy specimens, using a large-scale real-time reverse transcription-PCR approach
Authors: Tozlu S , Girault I , Vacher S , Vendrell J , Andrieu C , Spyratos F , Cohen P , Lidereau R , Bieche I
Published: December 2006

The estrogen receptor alpha (ERα) plays a critical role in the pathogenesis and clinical behavior of breast cancer. To obtain further insights into the molecular basis of estrogen-dependent forms of this malignancy, we used real-time quantitative reverse transcription (RT)-PCR to compare the mRNA expression of 560 selected genes in ERα-positive and ERα-negative breast tumors. Fifty-one (9.1%) of the 560 genes were significantly upregulated in ERα-positive breast tumors compared with ERα-negative breast tumors. In addition to well-known ERα-induced genes (PGR, TFF1/PS2, BCL2, ERBB4, CCND1, etc.) and genes recently identified by cDNA microarray-based approaches (GATA3, TFF3, MYB, STC2, HPN/HEPSIN, FOXA1, XBP1, SLC39A6/LIV-1, etc.), an appreciable number of novel genes were identified, many of, which were weakly expressed. This validates the use of large-scale real-time RT-PCR as a method complementary to cDNA microarrays for molecular tumor profiling. Most of the new genes identified here encoded secreted proteins (SEMA3B and CLU), growth factors (BDNF, FGF2 and EGF), growth factor receptors (IL6ST, PTPRT, RET, VEGFR1 and FGFR2) or metabolic enzymes (CYP2B6, CA12, ACADSB, NAT1, LRBA, SLC7A2 and SULT2B1). Importantly, we also identified a large number of genes encoding proteins with either pro-apoptotic (PUMA, NOXA and TATP73) or anti-apoptotic properties (BCL2, DNTP73 and TRAILR3). Surprisingly, only a small proportion of the 51 genes identified in breast tumor biopsy specimens were confirmed to be ERα-regulated and/or E2-regulated in vitro (cultured cell lines). Therefore, this study identified a limited number of genes and signaling pathways, which better delineate the role of ERα in breast cancer. Some of the genes identified here could be useful for diagnosis or for predicting endocrine responsiveness, and could form the basis for novel therapeutic strategies.
Predictive factors of efficacy of the somatostatin analogue octreotide as first line therapy for advanced pancreatic endocrine carcinoma
Authors: Butturini G , Bettini R , Missiaglia E , Mantovani W , Dalai I , Capelli P , Ferdeghini M , Pederzoli P , Scarpa A , Falconi M
Published: December 2006

About 40% of nonfunctioning pancreatic endocrine carcinomas (NF-PEC) cannot be cured by surgery due to advanced stage disease. Somatostatin analogues have been proposed as first line therapy in these cases. We performed a prospective phase IV study to assess the efficacy of octreotide in advanced NF-PEC and identify factors predictive of response to therapy. Twenty-one consecutive patients with octreoscan-positive advanced-stage well-differentiated NF-PEC were treated with long-acting release octreotide 20 mg i.m. at diagnosis. The immunohistochemical expression of somatostatin receptor 2 (SSTR2) and the quantitative mRNA analysis of SSTR2 and SSTR5 were assessed in 12 tumours. The tumour proliferative fraction was assessed by immunohistochemistry for Ki-67. Eight patients (38%) had stable disease (SD) after a median follow-up of 49.5 months. Thirteen patients (62%) developed progression after a median of 18 months. Tumour progression correlated with a proliferative index ≥5% (P=0.016), weight loss (P=0.006) and absence of abdominal pain (P=0.003) at diagnosis. Other clinical (age, gender and primary tumour resection) or pathological parameters (site, size and liver metastasis) lacked significant correlation with tumour progression. No difference in the amount of SSTR2 mRNA and protein or SSTR5 mRNA was found between tumours that were stable (n=5) and seven tumours that progressed (n=7). Treatment with long-acting release octreotide was associated with stabilization of disease and a good quality of life in 38% of patients. A Ki-67 index ≥5% and/or the presence of weight loss may justify more aggressive therapy without waiting for radiologically proven progression of disease.
Natural course of small, asymptomatic neuroendocrine pancreatic tumours in multiple endocrine neoplasia type 1: an endoscopic ultrasound imaging study
Authors: Kann P H , Balakina E , Ivan D , Bartsch D K , Meyer S , Klose K-J , Behr Th , Langer P
Published: December 2006

Endoscopic ultrasound (EUS) enables detection and localization of pancreatic neuroendocrine tumours. Even small tumours down to a diameter of 1–2 mm can be visualized. Since such small tumours usually cannot be detected by computed tomography (ct), magnetic resonance imaging (mri) and somatostatin receptor scintigraphy (srs), and experience with EUS imaging is limited, there is no clear evidence for clinical management in multiple endocrine neoplasia type 1 (MEN1). Knowledge about the natural course of growth and metastatic distribution is mandatory to come to appropriate clinical decisions and guidelines. This prospective study was aimed to assess the natural course of small (<15 mm) neuroendocrine pancreatic tumours without clinical symptoms due to endocrine activity or mechanical problems and without clear indication for surgical therapy in MEN1 by EUS.

A total of 82 asymptomatic tumours <15 mm (5.9 ± 3.2 mm diameter at baseline) in 20 patients with MEN1-disease (8 female/12 male, 43 ± 13 years) were studied over a period of 20 ± 12 months (33.8 patient years, 106.7 tumour years) by EUS. Change in largest diameter of each tumour and annual tumour incidence rate in the patients’ cohort were calculated.

Increase of largest tumour diameter was found to be 1.3 ± 3.2% per month, annual tumour incidence rate 0.62 new tumours per patient year. In one patient, rapid progressive pancreatic manifestation of MEN1 was observed. There was no evidence in ct and/or srs and/or mri for metastatic disease in all patients. Only 4/84 (4.8%) pancreatic tumours could be visualized by computed tomography, 5/79 (6.3%) by somatostatin receptor imaging and 4/39 (10.3%) by magnetic resonance imaging.

Small asymptomatic neuroendocrine pancreatic tumours in MEN1 usually seem to grow slowly. Annual tumour incidence rate is low. However, faster growing tumours and patients with rapidly progressive disease can be observed. Risk for obvious metastatic disease from asymptomatic neuroendocrine pancreatic tumours <15 mm in MEN1 seems to be low.
Human melanoma cells express functional receptors for thyroid-stimulating hormone
Authors: Ellerhorst Julie A , Sendi-Naderi Aresu , Johnson Marilyn K , Cooke Carolyn P , Dang Shyam M , Diwan A Hafeez
Published: December 2006

We have reported a high prevalence of hypothyroidism in the cutaneous melanoma population, suggesting that the pathologic hormonal environment of hypothyroidism promotes melanoma growth. The objective of this study was to test the hypothesis that TSH, which circulates at elevated levels in hypothyroid individuals, stimulates the growth of melanoma cells. Our results show that TSH receptors (TSHR) are expressed by virtually all cutaneous melanocytic lesions, including benign nevi, dysplastic nevi, and melanomas, with higher expression found in malignant and pre-malignant lesions. The finding of TSHR expression by human tumors is confirmed in cultured melanoma cells and melanocytes, in which TSHR expression is demonstrated by immunofluorescent staining, western blotting, and reverse transcriptase-PCR. Melanoma TSHR are functional, as evidenced by the ability of TSH to induce the formation of cAMP and to activate the mitogen-activated protein kinase (MAPK) pathway. Cultured melanoma cells, but not melanocytes, are induced to proliferate at a physiologically relevant concentration of TSH. Taken together, these data support the hypothesis that TSH is a growth factor for human melanoma. Our findings have broad clinical implications for the prevention of melanoma and the management of established disease.