bluuebunny/crossref_metadata_2025 huggingface dataset:
Gene expression profiling to identify markers associated with deregulated hTERT in HPV‐transformed keratinocytes and cervical cancer
Authors: de Wilde Jillian , Wilting Saskia M. , Meijer Chris J.L.M. , van de Wiel Mark A. , Ylstra Bauke , Snijders Peter J.F. , Steenbergen Renske D.M.
Published: October 2007

Although high‐risk human papillomavirus (HPV) infection plays a major role in the development of cervical cancer, additive oncogenic events are involved as well. One key event involves increased activity of telomerase resulting from a deregulated expression of its catalytic subunit hTERT. Our previous microcell‐mediated chromosome transfer studies revealed that introduction of human chromosome 6 in the HPV16‐immortalized keratinocyte cell line FK16A and in the HPV16‐containing cervical cancer cell line SiHa induced growth arrest, resulting from a repression of hTERT mRNA expression and telomerase activity. Here, this model was used to analyze expression profiles associated with hTERT deregulation in HPV‐transformed cells. Microarray expression analysis of 12 FK16A/chromosome 6 hybrids, 4 of which were negative for endogenous hTERT and 8 of which were positive for endogenous hTERT, resulted in the identification of 164 differentially expressed genes. Differential expression of a selection of 5 genes was verified by real‐time RT‐PCR. Of these 164 genes, 32 were also differentially expressed in other HPV transformed cells with deregulated hTERT. For 2 of these genes, encoding AQP3 and MGP, altered expression in hTERT positive cervical carcinomas was confirmed by real‐time RT‐PCR and immunohistochemistry, respectively. Moreover, increased MGP protein expression was significantly more frequent in high‐grade cervical premalignant lesions with elevated hTERT mRNA expression compared to those without. In summary, we identified 32 candidate biomarkers for deregulated hTERT mRNA expression, which may enable the identification of cervical premalignant lesions that are at highest risk to progress to invasive cancer. © 2007 Wiley‐Liss, Inc.
p68 (Ddx5) interacts with Runx2 and regulates osteoblast differentiation
Authors: Jensen Eric D. , Niu Lingling , Caretti Giuseppina , Nicol Samantha M. , Teplyuk Nadiya , Stein Gary S. , Sartorelli Vittorio , van Wijnen Andre J. , Fuller‐Pace Frances V. , Westendorf Jennifer J.
Published: October 2007

Runx2 is an essential transcription factor for osteoblast development from mesenchymal progenitors. Runx2 regulates gene expression by interacting with numerous transcription factors and co‐activators to integrate signaling events within the nucleus. In this study we used affinity purification and proteomic techniques to identify novel Runx2 interacting proteins. One of these proteins is the DEAD box RNA helicase, p68 (Ddx5). p68 regulates many aspects of RNA expression, including transcription and splicing. p68 co‐localized with Runx2 in punctate foci within the nucleus. In transcription assays, p68 functioned as a co‐activator of Runx2, but its helicase activity was not essential for co‐activation. In accordance, Runx2 transcriptional activity was muted in p68‐suppressed cells. Surprisingly, osteoblast differentiation of the multipotent progenitor C2C12 cell line was accelerated by p68 suppression and Runx2 suppressed p68 expression in calvarial progenitor cells. Together these data demonstrate that p68 is a novel co‐activator for Runx2, but it inhibits osteogenic differentiation of progenitor cells. Moreover Runx2 has an active role in regulating p68 levels in osteoblast precursors. Thus, crosstalk between Runx2 and p68 controls osteoblast specification and maturation at multiple levels. J. Cell. Biochem. 103: 1438–1451, 2008. © 2007 Wiley‐Liss, Inc.
Chondroitin sulfate prevents platelet derived growth factor‐mediated phosphorylation of PDGF‐Rβ in normal human fibroblasts severely impairing mitogenic responses
Authors: Fthenou E. , Zafiropoulos A. , Katonis P. , Tsatsakis A. , Karamanos N.K. , Tzanakakis G.N.
Published: October 2007

Platelet‐derived growth factor (PDGF) is a major polypeptide mitogen for cells of mesenchymal origin such as fibroblasts. Chondroitin sulfate chains (CS), which are abundant in the extracellular matrix have been shown to physically interact with PDGF‐BB modulating its biological function. The aim of the present study was to examine the involvement of CS on PDGF‐BB induced proliferative responses and receptor activation in human lung fibroblasts. The addition of exogenous free CS chains caused a significant downregulation of the PDGF‐BB mediated mitogenic and chemotactic responses. Similar results were obtained by the increase of endogenous CS biosynthesis after β‐D‐xyloside treatment. Furthermore, removal of the membrane‐bound CS chains by selective enzymatic treatment significantly increased the proliferative capacity of human fibroblasts. Analysis of PDGF‐R phosphorylation in the presence of CS or β‐D‐xyloside, revealed a reduction of PDGF‐Rβ phosphorylation in the tyrosine residue 1021. These results demonstrate, for the first time, that CS either soluble or surface bound downregulates the mitogenic responses of PDGF‐BB in normal human lung fibroblasts through the reduction of PDGF‐Rβ phosphorylation. J. Cell. Biochem. 103: 1866–1876, 2007. © 2007 Wiley‐Liss, Inc.
Flow‐dependent regulation of angiopoietin‐2
Authors: Goettsch Winfried , Gryczka Corina , Korff Thomas , Ernst Evelyn , Goettsch Claudia , Seebach Jochen , Schnittler Hans‐Joachim , Augustin Hellmut G. , Morawietz Henning
Published: October 2007

Endothelial cells are constantly exposed to high or low shear stress in arteries and veins by the flowing blood. Angiopoietin‐2 (Ang‐2) is acting as a critical regulator of vessel maturation and endothelial cell quiescence. In this study, flow‐dependent regulation of Ang‐2 was analyzed in vitro and in vivo. Ang‐2 mRNA, protein expression and release was upregulated by 24 h of low (1 dyne/cm2), but downregulated by high flow (30 dyne/cm2) in human endothelial cells. Increased endothelial NO synthase expression and NO formation was not affecting regulation of Ang‐2 by low or high flow. Low and high flow increased VEGF‐A expression. Inhibition of VEGFR‐2 prevented upregulation of Ang‐2 by low flow, but not downregulation of Ang‐2 by high flow. Furthermore, upregulation of Ang‐2 by VEGF was reduced by application of high flow. Forkhead box O (FOXO) transcription factor FOXO1 has been shown to regulate Ang‐2 expression in endothelial cells. FOXO1 binding activity was reduced by high flow. Nuclear localization of transcription factor FOXO1 was not changed by low flow, but reduced by high flow. In vivo, Ang‐2 was higher expressed in veins compared to arteries. Arterial ligation augmented Ang‐2 expression in distal arterial low flow areas. Our results support a VEGF‐dependent induction of Ang‐2 in low flow areas, and FOXO1‐dependent downregulation of Ang‐2 in high flow areas. These data suggest a new mechanism of flow‐dependent regulation of vessel stability and differentiation. J. Cell. Physiol. 214: 491–503, 2008. © 2007 Wiley‐Liss, Inc.
Functional characterization and cloning of amino acid transporter B0,+ (ATB0,+) in primary cultured rat pneumocytes
Authors: Uchiyama Tomomi , Fujita Takuya , Gukasyan Hovhannes J. , Kim Kwang‐Jin , Borok Zea , Crandall Edward D. , Lee Vincent H.L.
Published: October 2007

Cationic amino acid transport in primary cultured rat pneumocytes exhibiting characteristics of alveolar epithelial type I‐like cells are described. Asymmetry and activator ion dependency of 3H‐L‐arginine uptake were characterized from the apical or basolateral fluid of pneumocytes grown on permeable support. Substrate specificity of transport was evaluated as a function of 3H‐L‐arginine uptake inhibition in the presence of other amino acids. Transepithelial transport studies estimated 3H‐L‐arginine flux in the apical‐to‐basolateral and basolateral‐to‐apical directions. Full length cDNA of rat amino acid transporter B0,+ (rATB0,+) was cloned and its relative expression level studied. Results indicate that uptake of 3H‐L‐arginine from apical fluid is dependent on Na+ and Cl−. Zwitterionic and cationic amino acids (excluding L‐proline and anionic amino acids) inhibited uptake of 3H‐L‐arginine from apical, but not basolateral incubation fluid. Apical‐to‐basolateral transepithelial flux of 3H‐L‐arginine was 20× higher than basolateral‐to‐apical transport. Kinetic studies of 3H‐L‐arginine uptake from apical fluid revealed maximal velocity (Vmax) and Michaelis–Menten constants (Kt) of 33.32 ± 2.12 pmol/mg protein/15 min and 0.50 ± 0.11 mM, respectively, in a cooperative process having a coupling ratio of 1.18 ± 0.16 with Na+ and 1.11 ± 0.13 with Cl−. Expression of rATB0,+ mRNA was identified by RT‐PCR and Northern analysis. Corresponding cloned 3.2 kb rATB0,+ cDNA sequence exhibits pronounced homology in deduced amino acid sequence to mouse (95% identity and 97% similarity) and human (89% identity and 95% similarity) ATB0,+ homologues. We conclude that rat pneumocytes express ATB0,+, which may partly contribute towards recovering cationic and neutral amino acids from alveolar luminal fluid. J. Cell. Physiol. 214: 645–654, 2008. © 2007 Wiley‐Liss, Inc.
xRic‐8 is a GEF for Gsα and participates in maintaining meiotic arrest in Xenopus laevis oocytes
Authors: Romo Ximena , Pastén Pamela , Martínez Silvana , Soto Ximena , Lara Pablo , de Arellano Antonieta Ramírez , Torrejón Marcela , Montecino Martín , Hinrichs María Victoria , Olate Juan
Published: October 2007

Immature stage VI Xenopus oocytes are arrested at the G2/M border of meiosis I until exposed to progesterone, which induces meiotic resumption through a non‐genomic mechanism. One of the earliest events produced by this hormone is inhibition of the plasma membrane enzyme adenylyl cyclase (AC), with the concomitant drop in intracellular cAMP levels and reinitiation of the cell cycle. Recently Gsα and Gβγ have been shown to play an important role as positive regulators of Xenopus oocyte AC, maintaining the oocyte in the arrested state. However, a question that still remains unanswered, is how the activated state of Gsα and Gβγ is achieved in the immature oocyte, since no receptor or ligand have been found to be required. Here we provide evidence that xRic‐8 can act in vitro and in vivo as a GEF for Gsα. Overexpression of xRic‐8, through mRNA injection, greatly inhibits progesterone induced oocyte maturation and endogenous xRic‐8 mRNA depletion, through siRNA microinjection, induces spontaneous oocyte maturation. These results suggest that xRic‐8 is participating in the immature oocyte by keeping Gsα‐Gβγ‐AC signaling complex in an activated state and therefore maintaining G2 arrest. J. Cell. Physiol. 214: 673–680, 2008. © 2007 Wiley‐Liss, Inc.
Epidermal growth factor upregulates motility of Mat‐LyLu rat prostate cancer cells partially via voltage‐gated Na+ channel activity
Authors: Ding Yanning , Brackenbury William J. , Onganer Pinar U. , Montano Ximena , Porter Louise M. , Bates Lucy F. , Djamgoz Mustafa B.A.
Published: October 2007

The main aim of this investigation was to determine whether a functional relationship existed between epidermal growth factor (EGF) and voltage‐gated sodium channel (VGSC) upregulation, both associated with strongly metastatic prostate cancer cells. Incubation with EGF for 24 h more than doubled VGSC current density. Similar treatment with EGF significantly and dose‐dependently enhanced the cells' migration through Transwell filters. Both the patch clamp recordings and the migration assay suggested that endogenous EGF played a similar role. Importantly, co‐application of EGF and tetrodotoxin, a highly selective VGSC blocker, abolished 65% of the potentiating effect of EGF. It is suggested that a significant portion of the EGF‐induced enhancement of migration occurred via VGSC activity. J. Cell. Physiol. 215: 77–81, 2008. © 2007 Wiley‐Liss, Inc.
Protein kinase A and mitogen‐activated protein kinase pathways mediate cAMP induction of α‐Epithelial Na+ channels (α‐ENaC)
Authors: Mustafa Shamimunisa B. , Castro Robert , Falck Alison J. , Petershack Jean A. , Henson Barbara M. , Mendoza Yvonne M. , Choudary Ahsan , Seidner Steven R.
Published: October 2007

A major mechanism for Na+ transport across epithelia occurs through epithelial Na+ channels (ENaC). ENaC is a multimeric channel consisting of three subunits (α, β, and γ). The α‐subunit is critical for ENaC function. In specific culture conditions, the rat submandibular gland epithelial cell line (SMG‐C6) demonstrates minimal Na+ transport properties and exposure to dibutyryl cAMP (DbcAMP) for up to 48 h caused an elevation of α‐ENaC mRNA and protein expression and amiloride‐sensitive short‐circuit current (ISC). Here we examined the early signaling pathways evoked by DbcAMP which contribute to the eventual increase in Na+ transport is present. Treatment with either of the protein kinase A (PKA) inhibitors KT5720 or H‐89 followed by exposure to 1 mM DbcAMP for 24 h markedly attenuated DbcAMP‐induced α‐ENaC protein formation and ISC. Exposure of SMG‐C6 cells to 1 mM DbcAMP induced a rapid, transient phosphorylation of the cAMP response element binding protein (CREB). This response was attenuated in the presence of either KT5720 or H‐89. Dominant‐negative CREB decreased DbcAMP‐induced α‐ENaC expression. Suppression of the extracellular signal‐regulated protein kinase (ERK 1,2) with PD98059 or the p38 mitogen‐activated protein kinase (MAPK) pathway with SB203580 reduced DbcAMP‐induced α‐ENaC protein levels in SMG‐C6 cells. DbcAMP‐induced phosphorylation of CREB was markedly attenuated by PD98059 or SB203580. DbcAMP‐induced activation of the either the p38 or the ERK 1,2 MAPK pathways was abolished by either of the PKA inhibitors, H‐89 or KT5720. Cross talk between these signaling pathways induced by DbcAMP via the activation of CREB appears to contribute to increased levels of α‐ENaC observed after 24 h of treatment in SMG‐C6 epithelial cells. J. Cell. Physiol. 215: 101–110, 2008. © 2007 Wiley‐Liss, Inc.
Centrosome duplication proceeds during mimosine‐induced G1 cell cycle arrest
Authors: Durcan Thomas M. , Halpin Elizabeth S. , Casaletti Luciana , Vaughan Kevin T. , Pierson Maggie R. , Woods Shane , Hinchcliffe Edward H.
Published: October 2007

Centrosome duplication must remain coordinated with cell cycle progression to ensure the formation of a strictly bipolar mitotic spindle, but the mechanisms that regulate this coordination are poorly understood. Previous work has shown that prolonged S‐phase is permissive for centrosome duplication, but prolonging either G2 or M‐phase cannot support duplication. To examine whether G1 is permissive for centrosome duplication, we release serum‐starved G0 cells into mimosine, which delays the cell cycle in G1. We find that in mimosine, centrosome duplication does occur, albeit slowly compared with cells that progress into S‐phase; centrosome duplication in mimosine‐treated cells also proceeds in the absence of a rise in Cdk2 kinase activity normally associated with the G1/S transition. CHO cells arrested with mimosine can also assemble more than four centrioles (termed “centrosome amplification”), but the extent of centrosome amplification during prolonged G1 is decreased compared to cells that enter S‐phase and activate the Cdk2‐cyclin complex. Together, our results suggest a model, which predicts that entry into S‐phase and the rise in Cdk2 activity associated with this transition are not absolutely required to initiate centrosome duplication, but rather, serve to entrain the centrosome reproduction cycle with cell cycle progression. J. Cell. Physiol. 215: 182–191, 2008. © 2007 Wiley‐Liss, Inc.
Systemically transplanted bone marrow stromal cells contributing to bone tissue regeneration
Authors: Li S. , Tu Q. , Zhang J. , Stein G. , Lian J. , Yang P.S. , Chen J.
Published: October 2007

Bone marrow stromal cells (BMSCs) are a rich source of osteogenic progenitor cells. A fundamental question is whether systemically transplanted BMSCs participate in bone regeneration. Luciferase and GFP double‐labeled BMSCs were transplanted into irradiated mice. Five weeks after transplantation, artificial bone wounds were created in the mandibles and calvaria of the recipients. Animals were sacrificed at weeks 2, 4, and 6 after surgery and the expressions of luciferase and GFP were determined using Xenogen IVIS Imaging System, immunohistochemical staining and RT‐PCR. The results demonstrated that transplanted BMSCs can be detected in wound sites as early as 2 weeks and lasted the whole experimental period. Luciferase expression peaked at 2 weeks after surgery and decreased thereafter, exhibiting a similar expression pattern as that of BSP, while GFP expression was relatively stable during the experimental period. In conclusion, BMSCs can migrate to bone wound sites and participate in bone regeneration in orocraniofacial region. J. Cell. Physiol. 215: 204–209, 2008. © 2007 Wiley‐Liss, Inc.
Regulation of collagen gene expression in the Tsk2 mouse
Authors: Barisic‐Dujmovic Tatjana , Boban Ivana , Clark Stephen H.
Published: October 2007

The tight skin 2 (Tsk2) mutation is an ENU induced dominant mutation localized on mouse chromosome 1. While the molecular defect is unknown, Tsk2/+ mice display cutaneous thickening associated with excessive matrix production and are used as a model of scleroderma. The purpose of this study was to examine the cellular mechanisms associated with the excessive synthesis of matrix macromolecules using a collagen promoter GFP reporter transgene (pOBCol3.6GFP) as a marker of Col1a1 expression. This analysis of pOBCol3.6GFP expression in Tsk2/+ skin showed an increase in transgene activity compared to wild‐type (+/+) samples. In addition, an increased area of “high” GFP fluorescence in Tsk2/+ dermis in both 1‐ and 4‐month‐old mice was observed that was also associated with an increased number of dermal fibroblasts per unit area of dermis. These data collectively suggest an important mechanism of Tsk2/+ skin fibrosis; an increased number of collagen expressing cells as well as elevated collagen expression on a per cell basis. During this study it was noted that Tsk2/+ mice appeared consistently smaller than wild‐type (+/+) siblings and measurements of body length revealed a decrease (5–10%) in 1‐ and 2‐month‐old Tsk2/+ mice as well as a decrease in body weight in both age groups as compared to wild‐type (+/+) control mice. Femur length was also decreased (2–9%) in Tsk2/+ mice. Finally, in contrast to Tsk/+ mice that display an emphysema‐like lung pathology, histological sections of lungs from Tsk2/+ mice were normal and indistinguishable from wild‐type (+/+) controls. J. Cell. Physiol. 215: 464–471, 2008. © 2007 Wiley‐Liss, Inc.
Microscopic observation and laser‐controlled microoptothermal drive mechanism
Authors: Zhang Dongxian , Zhang Haijun , Liu Chao , Jiang Jianzhong
Published: October 2007

The mechanism of novel optothermal microactuators (OTA) was analyzed with a microscope system, a charge‐coupled device‐combined light microscope, and a computer system. One OTA, two optothermal microswitches (OTS) with different length and shape were machined by an excimer laser micromatching system using single layer material. They all had two thin expansion arms with different widths. The mechanism of the OTA/OTS is that the different optothermal expansion controlled by asymmetric topology and shape of the arms causes a magnified lateral deflection or vibration. A red laser diode (650 nm) with maximum power output of 30 mW was employed as the external power source to drive the OTA/OTS. Experiments were carried out with the microscope system, and serial images and videos were acquired. The results indicate that the structure of the OTA/OTS is simple and easy to be manufactured. They can practically generate an obvious lateral deflection or vibration. When the location of the laser spot on the OTA changed, the direction of its deflection is also changed. When the value of laser power that irradiated on the OTS increased, the lateral deflection of the OTS enlarged, the maximum could be larger than 30 μm. This kind of novel microactuator has the advantages of remote wireless controlling, large displacement, simple structure, easy to be machined, and therefore will be quite useful for the practical applications in the fields of micro/nanotechnology. Microsc. Res. Tech., 2008. © 2007 Wiley‐Liss, Inc.
Enhanced contraceptive response by co‐immunization of DNA and protein vaccines encoding the mouse zona pellucida 3 with minimal oophoritis in mouse ovary
Authors: Li Jinyao , Jin Huali , Zhang Ailian , Li Yijie , Wang Bin , Zhang Fuchun
Published: October 2007

Zona pellucida 3 (ZP3) acts as the primary sperm receptor, induces autoantibody that can prevent oocyte fertilization and has been proposed as a vaccine candidate for contraception in humans. Due to the elicited autoreactive T cell inflammation that causes ovarian destruction, ZP3‐based vaccine with removed T epitopes from the ZP3 is considered as a preferred approach. We present here a new strategy to eliminate the T cell inflammation while retaining a high level of antibody by co‐immunization of mZP3 DNA and protein vaccines, which resulted in a higher reduction rate of fertility in this group. Histological analysis showed that there were normal follicular developments of infertile mice in the co‐immunized group; while other vaccine groups of the most infertile mice lacked mature follicles. There was a significant correlation between normal follicular development and the inhibition of T cell response in co‐immunized mice. At the same time, co‐immunization reduced the production of inflammatory cytokine, IFN‐γ, and increased the productions of IL‐10 and FoxP3 in CD4 T cells, suggesting the anti‐inflammation may be via a T regulatory function. The results indicate that co‐immunization of mZP3 DNA‐ and protein‐based vaccines can reduce fertility without interfering with the normal follicular development and present a novel strategy to develop a contraceptive vaccine in humans. Copyright © 2007 John Wiley & Sons, Ltd.
Proteomic analysis of the cortisol‐mediated stress response in THP‐1 monocytes using DIGE technology
Authors: Billing Anja M. , Fack Fred , Renaut Jenny , Olinger Christophe M. , Schote Andrea B. , Turner Jonathan D. , Muller Claude P.
Published: October 2007

The glucocorticoid (GC) cortisol, the main mediator of the hypothalamic‐pituitary‐adrenal axis has many implications in metabolism, stress response and the immune system. Its function is mediated via binding to the glucocorticoid receptor (GR), a member of the superfamily of ligand‐activated nuclear hormone receptors. The activity of the ligated GR results from its binding as a transcription factor to glucocorticoid response elements (GREs). Two‐dimensional gel electrophoresis with DIGE (fluorescence difference gel electrophoresis) technology was applied to study the effects of cortisol on the human THP‐1 monocytic cell line. A total of 28 cortisol‐modulated proteins were identified belonging to five functional groups: cytoskeleton (8), chaperones (9), immune response (4), metabolism (3) and transcription/translation (4). Their corresponding genes were screened for putative GREs in their + 10 kb/− 0.2 kb promoter regions including all alternative promoters available within the Database for Transcription Start Sites (DBTSS). FKBP51, known to be induced by cortisol, was identified as the strongest differentially expressed protein, and contains the highest number of strict GREs. Genomic analysis of five alternativeFKBP5promoter regions suggests GC inducibility of all transcripts. Additionally, proteomics (2D DIGE and 2D immunoblotting) revealed the existence of several FKBP51 isoforms, which were not previously described. To our knowledge this is the first proteomic study that addresses the effects of cortisol on immune cells. FKBP51 isoforms found on the gel map were linked to alternative promoter usage on the genetic level, successfully correlating both the specific proteomic and genomic findings. Copyright © 2007 John Wiley & Sons, Ltd.
Application of two‐dimensional electrophoresis and matrix‐assisted laser desorption/ionization time‐of‐flight mass spectrometry for proteomic analysis of the sexually transmitted parasite Trichomonas vaginalis
Authors: De Jesus Jose Batista , Cuervo Patricia , Junqueira Magno , Britto Constança , Silva‐Filho Fernando Costa , Sabóia‐Vahia Leonardo , González Luis Javier , Barbosa Domont Gilberto
Published: October 2007

Trichomonas vaginalis is a sexually transmitted protozoan parasite that infects the human urogenital tract causing trichomoniasis, a worldwide disease. In this work, a fresh clinical isolate of T. vaginalis was used for study of the protein expression in this species. Two‐dimensional gel electrophoresis (2‐DE) and MALDI‐TOF/TOF mass spectrometry (MS) were employed to create a reference map of soluble proteins in the pH range 4–7. A set of 116 proteins belonging to functional classes expressed in high and low abundance was identified by peptide mass fingerprinting and tandem MS. These identifications corresponded to 67 different proteins, suggesting that post‐translational modifications are common phenomena in T. vaginalis. Identified proteins were classified into 16 groups according to biological processes. Among detected proteins we identified the major enzymes involved in both cytosolic and hydrogenosomal metabolic pathways, as well as putative protein targets for new drug design. In addition, this analysis allows validation of previous gene predictions confirming the expression of 15 hypothetical proteins. Finally, the findings here reported represent the first reference proteome map of T. vaginalis and the first steps towards the description of a comprehensive proteome map of this parasite. Copyright © 2007 John Wiley & Sons, Ltd.
LC‐MSMS identification of Arabidopsis thaliana heat‐stable seed proteins: Enriching for LEA‐type proteins by acid treatment
Authors: Oliveira E. , Amara I. , Bellido D. , Odena M. A. , Domínguez E. , Pagès M. , Goday A.
Published: October 2007

Protein identification in systems containing very highly abundant proteins is not always efficient and usually requires previous enrichment or fractionation steps in order to uncover minor proteins. In plant seeds, identification of late embryogenesis abundant (LEA) proteins is often masked by the presence of the large family of storage proteins. LEA‐proteins are predicted to play a role in plant stress tolerance. They are highly hydrophilic proteins, generally heat‐stable, and correlate with dehydration in seeds or vegetative tissues. In the present work, we analyze the protein composition of heat‐stable Arabidopsis thaliana seed extracts after treatment with trichloroacetic acid (TCA). The composition of the proteins that precipitate and those that remain in solution in 3% TCA was analyzed by two different approaches: 1D SDS‐PAGE coupled to LC‐ESI‐MSMS analysis and a gel‐free protocol associated with LC‐MALDI‐MSMS.

Our results indicate that treating total heat‐soluble extracts with 3% TCA is an effective procedure to remove storage proteins by selective precipitation and this fractionation step provides a soluble fraction highly enriched in Lea‐type proteins. The analysis and determination of protein identities in this acid‐soluble fraction by MS technology is a suitable system for large‐scale identification of Lea‐proteins present in seeds. Copyright © 2007 John Wiley & Sons, Ltd.
Systematic determination of ion score cutoffs based on calculated false positive rates: application for identifying ubiquitinated proteins by tandem mass spectrometry
Authors: Vasilescu Julian , Smith Jeffrey C. , Zweitzig Daniel R. , Denis Nicholas J. , Haines Dale S. , Figeys Daniel
Published: October 2007

We report a simple approach for determining ion score cutoffs that permit the confident identification of ubiquitinated proteins by tandem mass spectrometry (MS/MS). Initial experiments involving the analysis of gel bands containing multi‐Ubiquitin chains with quadrupole time‐of‐flight and quadrupole ion trap mass spectrometers revealed that standard ion score cutoffs used for database searching were not sufficiently stringent. We also found that false positive and false negative rates (FPR and FNR) varied significantly depending on the cutoff scores used and that appropriate cutoffs could only be determined following a systematic evaluation of false positive rates. When standard cutoff scores were used for the analysis of complex mixtures of ubiquitinated proteins, unacceptably high FPR were observed. Finally, we found that FPR for ubiquitinated proteins are affected by the size of the protein database that is searched. These observations may be applicable for the study of other post‐translational modifications. Copyright © 2007 John Wiley & Sons, Ltd.
Local injection of thrombin‐related peptide (TP508) in PPF/PLGA microparticles–enhanced bone formation during distraction osteogenesis
Authors: Wang Yan , Wan Chao , Szöke George , Ryaby James T. , Li Gang
Published: October 2007

We have previously demonstrated that injections of the thrombin‐related peptide, TP508, into the lengthening gap have significantly enhanced bone consolidation in a rabbit model of distraction osteogenesis. This study was to further test the effect of a single TP508 injection in slow release preparation on bone formation during distraction osteogenesis. Rabbits had left tibiae lengthened unilateral lengthener at rate of 1.4 mm/day for 6 days. TP508 was injected into as the following: Group 1, TP508 in saline; Group 2, in PPF/PLGA [poly(propylene fumarate)/poly(D,L‐lactic‐co‐glycolic acid)] microparticles; and Group 3, dextran gel only. All the animals were killed 2 weeks after lengthening. On radiographies, more bone was formed in the two TP508‐treated groups at first and secnd week postlengthening than that of the control Group 3. Microcomputed tomography (microCT) at 2 weeks indicated that the most advanced bone formation and remodeling was seen in Group 2. The mean volumetric BMD of the regenerates was significantly higher in the TP508 treated groups compared to the control group (p < 0.05). Histological evaluations supported the radiographic and the microCT results. In conclusion, we have demonstrated that a single injection of small amount of TP508 (300 µg) at the end of lengthening phases has significantly enhanced bone consolidation process in a rabbit model of distraction osteogenesis. The delivery of TP508 in PPF/PLGA microparticles appears to lead to a better quality bone formation over the saline delivery, further examinations are needed to confirm if PPF/PLGA microparticles may be desirable drug delivery form in augmenting bone formation. © 2007 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 26:539–546, 2008
Prediction of final tumor response to preoperative chemotherapy by Tc‐99m MIBI imaging at the middle of chemotherapy in malignant bone and soft tissue tumors: Comparison with Tl‐201 imaging
Authors: Taki Junichi , Higuchi Takahiro , Sumiya Hisashi , Tsuchiya Hiroyuki , Minato Hiroshi , Tomita Katsuro , Tonami Norihisa
Published: October 2007

The aim of this study was to assess the predictive power of 99mTc‐MIBI scintigraphy performed in the middle of chemotherapy, for the final tumor response to chemotherapy, and compare it to that of 201Tl in malignant bone and soft tissue tumors (MBST). Sixty‐eight patients with MBST underwent 99mTc‐MIBI and 201Tl scintigraphies at 15 min after tracer injection before the first, and after the third, chemotherapy cycles. After five cycles of chemotherapy and tumor resection, therapeutic effect was assessed by histopathology. Less than 90% and ≥90% necrosis were judged as poor and good response to chemotherapy, respectively. Tracers uptake ratios were calculated by dividing the lesion count density by that of the background. 99mTc‐MIBI perfusion index was also calculated. The % reduction of the perfusion index (ΔPI) and uptake ratios (ΔUR) calculated by 100 × [(prechemotherapy value − postchemotherapy value)/prechemotherapy value] were compared with histologic response. The sensitivity, specificity, and accuracy for the prediction of effective chemotherapy in 99mTc‐MIBI imaging were 80%, 95%, 88% in ΔUR, and 74%, 74%, 74% in ΔPI, respectively. The area under the receiver operator characteristic curve (Az) of the 99mTc‐MIBI‐ΔUR (0.923) was significantly higher than that of ΔPI (0.809, p = 0.025) but only marginally higher than that of the 201Tl‐ΔUR (0.865, p = 0.079). Az in 201Tl (0.865) was not significantly different from that of ΔPI (0.809, p = 0.35). 99mTc‐MIBI imaging performed in the middle of chemotherapy well predicts the final tumor response to chemotherapy in patients with malignant bone and soft tissue tumors. © 2007 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 26:411–418, 2008
An Overview of Important Microstructural Distributions for Polyolefin Analysis
Authors: Soares João B.P.
Published: October 2007

Summary: Polyolefins with complex microstructures are becoming increasingly common in academic and industrial applications. Polyolefin analytical techniques are evolving to provide a more detailed picture of these microstructures, with the development and improvement of hyphenated‐techniques and cross‐fractionation methods. These modern analytical techniques provide a wealth of information on polyolefin microstructure and, despite being extremely useful, they can also be hard to interpret without the help of mathematical models that link polymerization kinetics to chain microstructure and polymer characterization results. In this paper we review some of the most important distributions for polyolefin microstructure and derive a few new expressions that help understand the results obtained with several polyolefin characterization techniques.
Characterization of Ethylene‐1‐Hexene Copolymers Made with Supported Metallocene Catalysts: Influence of Support Type
Authors: Paredes Beatriz , Soares João B.P. , van Grieken Rafael , Carrero Alicia , Suarez Inmaculada
Published: October 2007

Summary: It is known that the nature of the support, as well as the technique used to anchor the metallocene onto it, play important roles on catalytic activity and on the properties of the polymers produced with supported metallocenes. In the present work, the effect of different support types on the microstructure of ethylene/1‐hexene copolymers made with supported metallocene catalysts has been investigated through the analysis of molecular weight and chemical composition distributions using high temperature gel permeation chromatography (GPC) and crystallization analysis fractionation (Crystaf). The copolymer samples obtained using commercial carriers (silica and silica‐alumina) had unimodal chemical composition distributions and were used to create a linear calibration curve relating the peak crystallization temperature from Crystaf and the comonomer content as determined by 13C NMR. This calibration curve is useful to determine the 1‐hexene fractions for each peak in the resins showing bimodal chemical composition distributions, such as those obtained with catalysts supported on MCM‐41 and SBA‐15 materials. The structure and chemistry of the support used had a large influence on comonomer incorporation and the shape of the chemical composition distribution of the polymer, which suggests that the supporting process creates different types of active sites.
Application of Fractionation Techniques to the Study of Olefin Polymerization Kinetics and Polymer Degradation
Authors: Nakatani Hisayuki , Matsuoka Hitoshi , Suzuki Shoutarou , Taniike Toshiaki , Boping Liu , Terano Minoru
Published: October 2007

Summary: Temperature rising elution fractionation (TREF) has been regarded as a powerful technique for study of semicrystalline polymers. In this paper, two examples of unique applications of TREF were introduced. One was the study on the influence of extraction of internal donor on the variation of isospecific active sites of a MgCl2‐ supported Ziegler catalyst, and the other was the estimation of the relationship between polymer micro‐tacticity and degradation rate of isotactic polypropylene (iPP). The former example revealed the conversion from high to low isospecific site by the extraction of internal donors, whereas the latter showed a negative correlation between the level of isotacticity and the degradation rate. These results demonstrated that TREF was useful in these research applications.
Synthesis and Characterization of Ethylene/Propylene Copolymers in the Whole Composition Range
Authors: Caballero Ma. Joaquina , Suarez Inmaculada , Coto Baudilio , Van Grieken Rafael , Monrabal Benjamín
Published: October 2007

Summary: The incorporation of comonomer molecules in the backbone of a homopolymer can influence the final properties of the material, decreasing its crystallinity and the melting and glass transition temperatures, and increasing its impact resistance and transparency. In the present work, ten ethylene/propylene copolymers have been synthesized using a supported metallocene catalytic system covering the whole composition range. Any desired composition was obtained by controlling the feed composition during the reaction. These synthesized copolymers have been characterized by different techniques in order to study the effect of the comonomer incorporation onto their final properties. When the comonomer content is low, the behaviour of the copolymer is similar to that of the corresponding homopolymer. Nevertheless, if the comonomer content increases, the copolymer becomes more amorphous (low crystallization temperature and soft XRD signals) and easily deformable, reaching a behaviour close to that corresponding to an elastomeric material. In order to corroborate these results the samples have been characterized by TREF and GPC‐MALS. TREF analysis showed that copolymers containing less than 10% and more than 80% of ethylene are semicrystalline, with elution temperatures typical of this kind of polymers. Molecular weights are higher for homopolymers and they decrease as the comonomer concentration increases, whereas the polydispersity index keeps almost constant at the expected value for this kind of samples.
Characterization of Polyethylene Nascent Powders Synthesized with TpTiCl2(OR) Catalysts
Authors: Casas Emilio , Karam Arquímedes , Díaz‐Barrios Antonio , Albano Carmen , Sánchez Yanixia , Méndez Bernardo
Published: October 2007

Summary: Different kinds of polyethylene and ethylene‐1‐hexene copolymers were synthesized with TpTiCl2(OR) (Tp = hydrotris(pyrazolyl)borate; R = Et, i‐Pr, n‐Bu) catalysts with and without H2. The polymers were characterized by 13C NMR, capillary viscosimetry or GPC, and DSC. The homopolymers showed properties characteristic of ultra‐high molecular weight polyethylenes (UHMWPE) with linear structure and high density polyethylenes (HDPE) with molecular weights in the range of commercial grades under hydrogen atmosphere. The copolymers showed a 1‐hexene incorporation up to 6 mol‐%. Important differences in the thermal properties were observed between the first DSC (nascent powders) and the second DSC heatings (melt‐crystallized samples), which evidenced the molecular weights influence on the melt‐crystallized samples.
Characterization of LDPE grafted with Diethylmaleate by Gamma Radiation: Application of FTIR, GPC and SSA Techniques
Authors: Sánchez Y. , Albano C. , Perera R. , Karam A. , Silva P.
Published: October 2007

Summary: The grafting of diethyl maleate onto low‐density polyethylene using gamma irradiation as initiator has been evaluated. The grafting degree was estimated by FTIR and with the use of a calibration curve. The functionalized polymers were characterized by DSC, MFI, SSA and GPC. An increase in the grafting degree with the radiation dose and the concentration of the functional monomer was found. Branching seemed to be the preferential reaction induced by radiation, causing an increase in average molecular weights with the consequently reduction of melt flow index values. The molecular segregation induced by the calorimetric treatment showed that grafting occurs preferentially through secondary carbon sequences.
Boundary integral method for Stokes flow past a porous body
Authors: Kohr Mirela , Raja Sekhar G. P. , Wendland Wolfgang L.
Published: October 2007

In this paper we obtain an indirect boundary integral method in order to prove existence and uniqueness of the classical solution to a boundary value problem for the Stokes–Brinkman‐coupled system, which describes an unbounded Stokes flow past a porous body in terms of Brinkman's model. Therefore, one assumes that the flow inside the body is governed by the continuity and Brinkman equations. Some asymptotic results in both cases of large and, respectively, of low permeability are also obtained. Copyright © 2007 John Wiley & Sons, Ltd.
Comparing treatment effects after adjustment with multivariable Cox proportional hazards regression and propensity score methods
Authors: Martens Edwin P. , de Boer Anthonius , Pestman Wiebe R. , Belitser Svetlana V. , Stricker Bruno H. Ch. , Klungel Olaf H.
Published: October 2007

Purpose

To compare adjusted effects of drug treatment for hypertension on the risk of stroke from propensity score (PS) methods with a multivariable Cox proportional hazards (Cox PH) regression in an observational study with censored data.

Methods

From two prospective population‐based cohort studies in The Netherlands a selection of subjects was used who either received drug treatment for hypertension (n = 1293) or were untreated ‘candidates’ for treatment (n = 954). A multivariable Cox PH was performed on the risk of stroke using eight covariates along with three PS methods.

Results

In multivariable Cox PH regression the adjusted hazard ratio (HR) for treatment was 0.64 (CI95%: 0.42, 0.98). After stratification on the PS the HR was 0.58 (CI95%: 0.38, 0.89). Matching on the PS yielded a HR of 0.49 (CI95%: 0.27, 0.88), whereas adjustment with a continuous PS gave similar results as Cox regression. When more covariates were added (not possible in multivariable Cox model) a similar reduction in HR was reached by all PS methods. The inclusion of a simulated balanced covariate gave largest changes in HR using the multivariable Cox model and matching on the PS.

Conclusions

In PS methods in general a larger number of confounders can be used. In this data set matching on the PS is sensitive to small changes in the model, probably because of the small number of events. Stratification, and covariate adjustment, were less sensitive to the inclusion of a non‐confounder than multivariable Cox PH regression. Attention should be paid to PS model building and balance checking. Copyright © 2007 John Wiley & Sons, Ltd.
Positron annihilation study of PbWO4 crystal doped with Y2O3 at different concentration
Authors: Xie Jianjun , Geng Zhengsong , Yuan Hui , Xiong Wei , Chen Liang , Ye Chongzhi , Xu Guoqing , Hao Xiaopeng , Zhang Zhiming , Ma Hongliang , Gu Mu , Feng Xiqi , Liao Jingying
Published: October 2007

A study of vacancy‐related defects in yttrium‐doped PbWO4 with doping concentration of 0–1 mol% has been performed using X‐ray diffraction and positron annihilation lifetime spectroscopy. It was found that the samples have vacancy‐like defects that act as positron traps. Two main defect lifetime components were found in all samples, one at about 0.195 ns to 0.208 ns and the other about 0.408 ns to 0.735 ns. These defect trapping sites can be attributed to single Pb‐site vacancies and interstitial oxygen ions, respectively. (© 2008 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)
Construction of confidence limits about effect measures: A general approach
Authors: Zou G. Y. , Donner A.
Published: October 2007

It is widely accepted that confidence interval construction has important advantages over significance testing for the presentation of research results, as now facilitated by readily available software. However, for a number of effect measures, procedures are either not available or not satisfactory in samples of small to moderate size. In this paper, we describe a general approach for estimating a difference between effect measures, which can also be used to obtain confidence limits for a risk ratio and a lognormal mean. Numerical evaluation shows that this closed‐form procedure outperforms existing methods, including the bootstrap. Copyright © 2007 John Wiley & Sons, Ltd.
Sample size formulae for trials comparing group and individual treatments in a multilevel model
Authors: Moerbeek Mirjam , Wong Weng Kee
Published: October 2007

In disease screening and prevention trials, subjects in the experimental condition are frequently nested within therapy groups, whereas subjects in the control group receive individual or no therapy and are therefore not nested within groups. Outcomes of subjects within the same therapy group are expected to be more alike than outcomes of subjects within different therapy groups. Ignoring this dependency in the design stage may result in less powerful designs. This paper presents a multilevel model for analyzing such trials and sample size formulae for continuous and binary outcomes with unequal variances and costs across groups. The proposed optimal design ensures that there is adequate power to detect a treatment effect with either minimal cost or a minimal number of subjects. We apply our strategy and design an improved trial where all subjects with musculoskeletal pain received conventional therapy and subjects in the intervention arm participated in a group‐learning program. Copyright © 2007 John Wiley & Sons, Ltd.
Mechanism of action and effectiveness of ester thiols as thermal stabilizers for poly(vinyl chloride)
Authors: Ge Xianlong , Culyba Elizabeth K. , Grinnell Cole L. , Zestos Alexander G. , Starnes William H.
Published: October 2007

Organic thiols containing at least one carboxylate ester group (ester thiols) are excellent thermal stabilizers for both rigid and plasticized poly(vinyl chloride) (PVC). Their mechanism of action is shown to involve the deactivation of unstable structural defects by nucleophilic chloride displacement, the retardation and removal of coloration through thiol additions to polyene double bonds, and the prevention of autoacceleration during thermal dehydrochlorination through polyene shortening reactions and the scavenging of free radicals formed from polyenes and HCl. An unusually facile displacement of labile chloride that is favored by thiol acidity can account, at least in part, for the relatively high effectiveness of dipentaerythritol hexakis(mercaptoacetate) as a stabilizer. J. VINYL ADDIT. TECHNOL., 13:170–175, 2007. © 2007 Society of Plastics Engineers
Method for assessing impact of large‐scale wind power integration on reserves
Authors: Norheim Ian , Pudjianto Danny
Published: October 2007

Concerns have been raised due to the impact that large‐scale wind power integration will have on reserves and secure operation of power systems. Case studies on a simplified model of the UK system with large penetration of wind power in Scotland are presented for illustration. A method called stepwise power flow was used to simulate the impact of wind power generation fluctuation output on the system frequency, reserves and power flow. It was demonstrated that the supply–demand imbalance caused by forecast errors in wind power production and load, and fall out of wind power production would influence the system frequency, system bias and the use of reserves. This method can also be used to estimate the amount of wind power that can be integrated before the system experiences serious problems due to inadequacy of the available reserves. Copyright © 2007 John Wiley & Sons, Ltd.