bluuebunny/crossref_metadata_2025 huggingface dataset:
Influence of Extraction Processing on Rheological Properties of Rapeseed Oils
Authors: Liu Changsheng , Yang Mei , Huang Fenghong
Published: July 2011

Rheological behavior of six crude rapeseed oils with different extraction methods including hot‐pressing, solvent‐extraction and cold‐pressing were studied. Viscosities of the oils were measured with shear rates ranging from 0.1 to 200 s−1 at three different temperatures. The Casson model was used to fit the experimental data and the Arrhenius equation was applied to estimate the energy of activation for viscosity (Ea). The extraction methods affected the total tocopherol, total phytosterols, total phenols, phosphorus and fatty acid composition. The hot‐pressed medium‐erucic rapeseed oil (HMRO) had the greatest viscosity, and the cold‐pressed low‐erucic rapeseed oil (CLRO) had the lowest viscosity among all the oils with shear rates >5 s−1 at 10 °C. The crude rapeseed oils exhibited Newtonian behavior at higher shear rates. The significant difference of viscosity of the six oils was reduced with increasing temperature, and there was no significant change (P > 0.01) among the oils with a shear rate of 100 s−1 at 50 °C. According to the values of Ea, the following order of a change in viscosity was obtained as follows: CMRO > SMRO > HMRO > CLRO > SLRO > HLRO(C, cold pressed; S, solvent extracted; H, hot pressed; M, medium erucic; L, low erucic; RO, rapeseed oil). Minor components may be the contributing factors for the values of Ea of rapeseed oils. The higher shear limiting viscosity (ηc) values calculated by the Casson model decreased as the temperature increased, but no significant change (P > 0.01) was observed for ηc by using different extraction methods at 50 °C.
Diagnosis and treatment of developmental dysplasia of the hip in the Netherlands: National questionnaire of paediatric orthopaedic surgeons on current practice in children less than 1 year old
Authors: Heeres Rick H. M. , Witbreuk M. M. E. H. , van der Sluijs J. A.
Published: July 2011

Purpose

There is no consensus in the literature regarding the diagnosis and treatment of developmental dysplasia of the hip (DDH). We designed a national questionnaire to assess the various opinions and current practice of paediatric orthopaedic surgeons in the Netherlands regarding the diagnosis and treatment of DDH in children less than 1 year old.

Methods

The questionnaire was sent to all members of the Dutch Paediatric Orthopaedic Society (DPOS). It discusses different methods and criteria used in the diagnosis of DDH, the use of different therapies and the use of different imaging techniques to evaluate the result of treatment.

Results

With 38 responders, the overall response rate to the survey was 67%. Most surgeons use clinical, radiographic and/or ultrasound examination for the diagnosis. The starting point of treatment is usually on the mild part of the DDH spectrum. The Pavlik harness is most popular in the treatment of dislocated hips, whereas in dysplastic hips, most surgeons use a rigid splint. The duration of treatment has a wide range and evaluation of the effect of treatment is predominantly done by radiography.

Conclusions

The diagnosis and treatment of DDH in the Netherlands has as much diversity as the literature has recommendations about this subject. The lack of consensus on many aspects of DDH diagnosis and treatment should form the basis for a discussion among Dutch paediatric orthopaedic surgeons. Using the available evidence, it should be possible to formulate a more uniform protocol for the diagnosis and treatment of DDH.
Präsentismus: Arbeiten mit eingeschränkter Gesundheit
Authors: Steinke Mika
Published: July 2011

Einleitung

Nicht jeder, der bei der Arbeit fehlt, ist krank. Und nicht jeder, der am Arbeitsplatz anwesend ist, ist gesund. Angesichts des demographischen Wandels und der damit einhergehenden Zunahme chronischer Erkrankungen wird insbesondere der letztere Fall, Krankheit bei der Arbeit, zunehmend ein Thema – und die hierfür bestehende Bezeichnung Präsentismus erlangt ein stetig wachsendes Interesse.
Sildenafil and Bosentan Improve Arterial Oxygenation During Acute Hypoxic Exercise: A Controlled Laboratory Trial
Authors: Olfert I. Mark , Loeckinger Alexander , Treml Benedikt , Faulhaber Martin , Flatz Markus , Burtscher Martin , Truebsbach Susanne , Kleinsasser Axel
Published: July 2011

Objectives

Sildenafil and, recently, bosentan have been reported to increase arterial saturation and exercise capacity at altitude. The mechanisms behind this are still poorly defined but may be related to attenuation of hypoxic pulmonary vasoconstriction (HPV) and improved gas exchange. This study was designed to examine and compare the effect of sildenafil and bosentan on pulmonary gas exchange during acute hypoxic exercise in a controlled laboratory setting.

Methods

Sixteen athletic university students (8 males, 8 females) were examined during exercise in a hypoxic chamber (11% oxygen) before and after the administration of either sildenafil (n=10) or bosentan (n=6). Respiratory and metabolic measurements were taken at rest and during increasing exercise intensity (up to 90% of their individual maximal oxygen uptake [V̇O2max]) in concert with arterial blood gas sampling.

Results

Both drugs resulted in small, but significant increases in arterial PO2 (2–3 Torr) and O2 saturation (3–4%) at rest and during hypoxic exercise, in both men and women. No significant changes in arterial PCO2 or ventilation were seen at rest or during exercise in hypoxia; however, heart rate (both at rest and during exercise) was increased with both sildenafil and bosentan in both men and women.

Conclusions

These data demonstrate that sildenafil and bosentan equally improve arterial oxygenation in acute hypoxia in both men and women, which could account for improved physical performance at altitude.
Wilderness Communications
Authors: Worley Gordon H.
Published: July 2011

When an emergency situation arises in a remote location, the ability to communicate with outside sources of assistance can prove very valuable. This article reviews the different types of communications technologies available to individuals in remote locations, including satellite telephones, personal locator beacons, satellite messengers, cellular telephones, and the different licensed and non-licensed 2-way radio services available for personal use. It also discusses basic radio communications techniques, emergency communication, requesting ground or air casualty evacuation, and selecting communications devices for different applications.
‘Gonzalo Oleas, Defensor’: Cultural Intermediation in Mid-Twentieth-Century Ecuador
Authors: BECKER MARC
Published: July 2011

Gonzalo Oleas Zambrano was a socialist lawyer from Quito who, from the 1930s to the 1970s, became deeply involved in assisting rural communities in Ecuador with their legal petitions. Intermediaries have a long and varied history in negotiating relationships between the city and the countryside, and one that is often not well understood. At various points in his career Oleas acted like atinterillo, a socialist and anindigenista. An examination of Oleas’ petitions quickly breaks down a simplistic characterisation of his actions and interpretation of his motivation. Rather, his ability to transcend existing categories helps explain why rural litigants so often turned to Oleas for assistance.
Do Exchange Rate Regimes Matter for Inflation and Exchange Rate Dynamics? The Case of Central America
Authors: CAPUTO G. RODRIGO , MAGENDZO IGAL
Published: July 2011

This paper makes an empirical contribution to discussion of the optimal exchange rate regime. Using a new dataset for Central American countries, we compare the dynamics of the real exchange rate (RER) between dollarised and non-dollarised countries. Our results show that the two dollarised countries in the region, El Salvador and Panama, are quite different in terms of RER dynamics. In El Salvador the RER spends more time away from its equilibrium level than in the non-dollarised countries in the region, while the contrary is true for Panama. We also find that inflation persistence is similar in El Salvador to the other countries, but smaller in Panama. This leads us to the conclusion that some degree of exchange rate flexibility helps countries to have a more aligned RER. Nevertheless, a long-lived, highly credible dollarised economy, like Panama, can reduce inflation persistence to such an extent that RER misalignments are in fact less frequent than in countries with more flexible exchange rate regimes.
The impact of scaffolding and overhearing on young children's use of the spatial termsbetweenandmiddle
Authors: FOSTER EMILY K. , HUND ALYCIA M.
Published: July 2011

The primary goal was to specify the impact of scaffolding and overhearing on young children's use of the spatial termsbetweenandmiddle. Four- and five-year-old children described the location of a mouse hidden between two furniture items in a dollhouse with assistance from a parent. Children's use ofbetweenandmiddleincreased significantly across trials, and in concert, parents' directive scaffolding involvingmiddledecreased across trials. In the second study, three common scaffolding types (Between Directive, Middle Directive, non-directive) were compared with a no prompt condition by having children receive prompts from a doll and with overhearing conditions in which children overheard conversations between two adult experimenters containingbetweenormiddle. Children's use ofbetweenandmiddlewas much more frequent following directive prompting than following non-directive prompting, no prompting, or overhearing. Moreover, children showed some evidence of usingbetweenandmiddlein response to non-directive prompting and overhearing.
FRAYED AT THE EDGES: COLLECTIVE MEMORY AND HISTORY ON THE BORDERS OF CLASSIC MAYA POLITIES
Authors: Golden Charles
Published: July 2011

This article explores social memory and history as they pertain particularly to secondary political centers on the edges of the Classic Maya kingdoms of Piedras Negras and Yaxchilan. Over the course of the Late Classic period (a.d. 600–900) the rulers of Maya polities in the Usumacinta River basin increasingly relied on the subordinate lords who governed these secondary centers to patrol and control the boundaries of their territories. For the rulers of any state, formulating an appropriate and coherent history to guide social memory is a critical political act for maintaining the cohesion of the political community. But as the Classic period progressed, client lords were increasingly permitted a formerly royal prerogative; they were accorded their own inscribed monuments. The monuments, together with associated ritual performances, were an integral part of the construction of history and collective memory in local communities and allowed secondary nobles to restructure social memory for their own interests. This trend, in turn, increased the potential for royal history and authority to be contested throughout the kingdom. Through several case studies this paper examines the ways that subordinate nobles could contest social memory and history sanctioned by primary rulers and the ways in which kings acted to maintain the reins of history and memory.
RITUAL REMAINS AND COLLECTIVE MEMORY: MAYA EXAMPLES FROM WEST CENTRAL BELIZE
Authors: Schwake Sonja A. , Iannone Gyles
Published: July 2011

The idea of collective or social memory is explored in this paper as a way to understand ancient Maya ritual behavior. The theoretical characteristics of collective memory are defined and a strategy to operationalize the theory of collective memory using archaeological remains is presented. Two archaeological examples from the sites of Minanha and Zubin in west central Belize are discussed in terms of how they fit this new model. Finally, the social motivations that underlie this behavior are examined.
SITES OF MEMORY IN THE MAKING: POLITICAL STRATEGIZING IN THE CONSTRUCTION AND DECONSTRUCTION OF PLACE IN LATE TO TERMINAL CLASSIC SOUTHEASTERN MESOAMERICA
Authors: Stockett Miranda
Published: July 2011

Archaeological spaces can be viewed as material manifestations of human drama—sites for the production, expression, and manipulation of social life, power, and history. By viewing such spaces as stages for the enactment of processes of social memory, we may further enrich considerations of the interplay of materiality and history. Here I address the insights archaeologists may gain from engaging with theories of social memory by exploring their application to the analysis of settlements occupied during the Late to Terminal Classic period transition (a.d. 650–900) in pre-Columbian southeastern Mesoamerica. I also consider their relevance to community initiatives engaged by archaeologists today. Ultimately, I argue that processes of making, altering, and remaking place are one among many ways that memory may have served as a tool for political strategies and discourses about power.
Transcatheter closure in two rare cases of left-to-right shunt with Cardio-O-Fix occluders
Authors: Bialkowski Jacek , Akdeniz Celal , Celebi Ahmet
Published: July 2011

We present two procedures of transcatheter closure: that of an aorto-pulmonary window in a 12-month-old infant with a body weight of 7 kilograms, and that of ruptured sinus of Valsalva aneurysm into the right atrium in an adult patient. In the first case, we applied the muscular ventricular septal defect Cardio-O-Fix, while in the other we applied the patent ductus arteriosus Cardio-O-Fix occluder. The procedures were successful in both patients, and we achieved complete closure of the unwanted shunts.
Konversionsstörungen: Störungsbild, Diagnostik, Psychoedukation und Intervention
Authors: Noeker Meinolf
Published: July 2011

Konversionsstörungen umfassen nichtepileptische Anfälle sowie sensorische oder motorische Funktionsausfälle (Gangstörungen, Kraftverlust und Lähmung, Sprechstörungen) ohne organmedizinische Ursache. Sie nehmen in klassifikatorischer, ätiologischer und therapeutischer Hinsicht eine Zwischenstellung zwischen den somatoformen und dissoziativen Störungen ein. Bei hohen Raten psychopathologischer Vulnerabilität und Komorbidität können traumatische Erfahrungen, akute Belastungsfaktoren sowie aktuell überfordernde Entwicklungsaufgaben als Auslöser wirken. Besonders typisch ist ein emotional invalidierendes Familienklima, das den authentischen und direkten Ausdruck von Bedürfnissen und Befindlichkeiten des Kindes hemmt. Die Manifestation der Konversionssymptome erfüllt einerseits eine entlastende und verstärkende Funktion, erzeugt andererseits aber neue Belastungsfaktoren wie funktionelle Behinderung, Einschränkungen der sozialen und schulischen Teilhabe, hypochondrische Sorge vor einer unerkannten Grunderkrankung sowie Angst vor Stigmatisierung als Simulant oder psychisch Kranker. Ein Störungsmodell wird vorgestellt, das auch als Grundlage für einen Leitfaden zur klinisch-psychologischen Exploration, Anamneseerhebung, Fallkonzeption sowie das das Procedere der Diagnosemitteilung dient. Patientenorientierte Metaphern werden im Rahmen der Psychoedukation eingesetzt, um die Entstehung der Symptomatik trotz intakter neuronaler Reizleitung für Patient und Eltern nachvollziehbar zu gestalten. Die psychotherapeutische Intervention startet mit einer schrittweisen Wiederaufnahme altersgerechter Aktivitäten und Rollen. Physiotherapie zum Training der geschwächten Muskulatur wird initiiert, um dem Patienten ein Ritual anzubieten, das eine Wiederherstellung der Funktionsausfälle ohne Gesichtsverlust ermöglicht. Psychotherapeutische Strategien im engeren Sinne greifen die vorgegebene Dissoziation des Bewusstseins von der körperlichen Symptomatik auf. Altersgerecht wird dem Kind ein Persönlichkeitsanteil beschrieben, der die Symptomatik überwinden will, ein anderer Persönlichkeitsanteil wird gegenübergestellt, der die Konversionssymptomatik und dessen Anliegen repräsentiert. Der Therapeut entwickelt und moderiert mit dem Kind Lösungen, die geeignet sind, beide Persönlichkeitsanteile wieder neu zu integrieren und damit die Dissoziation zwischen Bewusstsein und körperlicher Symptomatik aufzulösen.
Kognitive Leistungsprofile motorisch- und sprachentwicklungsverzögerter Vorschulkinder
Authors: Kastner Julia , Lipsius Maike , Hecking Mascha , Petermann Franz , Petermann Ulrike , Mayer Hermann , Springer Stephan
Published: July 2011

Unterschiedliche Studien belegen, dass Kinder mit umschriebenen motorischen Entwicklungsstörungen häufig auch komorbide Sprachentwicklungsstörungen aufweisen. Vermutlich liegt der Kombination beider Störungen eine gemeinsame Ursache zugrunde. Für die klinische Praxis ist es von Interesse zu untersuchen, ob Kinder mit motorischen und sprachlichen Entwicklungsstörungen ein spezifisches kognitives Leistungsprofil aufweisen, um frühzeitig umfassende Interventionen einleiten zu können. Dazu wurden die WPPSI-III-Resultate von insgesamt 53 Kindern im Alter von vier bis sechs Jahren mit isolierten motorischen Entwicklungsstörungen, isolierten expressiven Sprachstörungen und Kindern mit kombinierten motorischen und sprachlichen Entwicklungsstörungen untereinander, aber auch mit einer normativ entwickelten Referenzgruppe verglichen. Die Kinder mit isolierten expressiven Sprachstörungen weisen ausschließlich gravierende Defizite in den sprachlichen Subtests auf; Kinder mit motorischen Entwicklungsstörungen schneiden in allen Kerntests geringfügig schlechter als unauffällige Kinder ab, wobei signifikante Differenzen für die Untertests Mosaik-Test, Matrizen-Test, Allgemeines Wissen und Wortschatz-Test auffallen. Kinder mit motorischen und sprachlichen Entwicklungsstörungen zeigen im Vergleich zu ausschließlich sprachauffälligen Kindern identische Defizite in den verbalen Untertests der WPPSI-III. Der Vergleich mit den ausschließlich motorisch auffälligen Kindern lässt erkennen, dass sich die Testresultate in den handlungsbezogenen Subtests jedoch verschlechtern. Die erhobenen Befunde unterstützen die Argumente eines eigenen Störungskonzeptes; das kombinierte Auftreten motorischer und sprachlicher Entwicklungsstörungen unterstreicht die Notwendigkeit den kognitiven Leistungsstand umfassend zu überprüfen.
Enhancement of mDia2 activity by Rho-kinase-dependent phosphorylation of the diaphanous autoregulatory domain
Authors: Staus Dean P. , Taylor Joan M. , Mack Christopher P.
Published: July 2011

It is clear that RhoA activates the DRF (diaphanous-related formin) mDia2 by disrupting the molecular interaction between the DAD (diaphanous autoregulatory domain) and the DID (diaphanous inhibitory domain). Previous studies indicate that a basic motif within the DAD contributes to mDia2 auto-inhibition, and results shown in the present study suggest these residues bind a conserved acidic region within the DID. Furthermore, we demonstrate that mDia2 is phosphorylated by ROCK (Rho-kinase) at two conserved residues (Thr1061 and Ser1070) just C-terminal to the DAD basic region. Phosphomimetic mutations to these residues in the context of the full-length molecule enhanced mDia2 activity as measured by increased actin polymerization, SRF (serum response factor)-dependent smooth muscle-specific gene transcription, and nuclear localization of myocardin-related transcription factor B. Biochemical and functional data indicate that the T1061E/S1070E mutation significantly inhibited the ability of DAD to interact with DID and enhanced mDia2 activation by RhoA. Taken together, the results of the present study indicate that ROCK-dependent phosphorylation of the mDia2 DAD is an important determinant of mDia2 activity and that this signalling mechanism affects actin polymerization and smooth muscle cell-specific gene expression.
Cholesterol-induced conformational changes in the oxytocin receptor
Authors: Muth Sabine , Fries Anja , Gimpl Gerald
Published: July 2011

Recent studies suggest that cholesterol binding is widespread among GPCRs (G-protein-coupled receptors). In the present study, we analysed putative cholesterol-induced changes in the OTR [OT (oxytocin) receptor], a prototype of cholesterol-interacting GPCRs. For this purpose, we have created recombinant OTRs that are able to bind two small-sized fluorescence-labelled ligands simultaneously. An OTR antagonist was chosen as one of the ligands. To create a second ligand-binding site, a small-sized α-BTB (bungarotoxin binding) site was inserted at the N-terminus or within the third extracellular loop of the OTR. All receptor constructs were functionally active and bound both ligands with high affinity in the nanomolar range. Measurements of the quenching behaviour, fluorescence anisotropy and energy transfer of both receptor-bound ligands were performed to monitor receptor states at various cholesterol concentrations. The quenching studies suggested no major changes in the molecular environment of the fluorophores in response to cholesterol. The fluorescence anisotropy data indicated that cholesterol affects the dynamics or orientation of the antagonist. The energy transfer efficiency between both ligands clearly increased with increasing cholesterol. Overall, cholesterol induced both a changed orientation and a decreased distance of the receptor-bound ligands, suggesting a more compact receptor state in association with cholesterol.
Mechanistic studies on the enzymatic processing of fluorinated methionine analogues by Trichomonas vaginalis methionine γ-lyase
Authors: Moya Ignace A. , Westrop Gareth D. , Coombs Graham H. , Honek John F.
Published: July 2011

TFM (L-trifluoromethionine), a potential prodrug, was reported to be toxic towards human pathogens that express MGL (L-methionine γ-lyase; EC 4.4.1.11), a pyridoxal phosphate-containing enzyme that converts L-methionine into α-oxobutyrate, ammonia and methyl mercaptan. It has been hypothesized that the extremely reactive thiocarbonyl difluoride is produced when the enzyme acts upon TFM, resulting in cellular toxicity. The potential application of the fluorinated thiomethyl group in other areas of biochemistry and medicinal chemistry requires additional studies. Therefore a detailed investigation of the theoretical and experimental chemistry and biochemistry of these fluorinated groups (CF3S− and CF2HS−) has been undertaken to trap and identify chemical intermediates produced by enzyme processing of molecules containing these fluorinated moieties. TvMGL (MGL from Trichomonas vaginalis) and a chemical model system of the reaction were utilized in order to investigate the cofactor-dependent activation of TFM and previously uninvestigated DFM (L-difluoromethionine). The differences in toxicity between TFM and DFM were evaluated against Escherichia coli expressing TvMGL1, as well as the intact human pathogen T. vaginalis. The relationship between the chemical structure of the reactive intermediates produced from the enzymatic processing of these analogues and their cellular toxicity are discussed.
Farnesoid X receptor protects human and murine gastric epithelial cells against inflammation-induced damage
Authors: Lian Fan , Xing Xiangbin , Yuan Gang , Schäfer Claus , Rauser Sandra , Walch Axel , Röcken Christoph , Ebeling Martin , Wright Matthew B. , Schmid Roland M. , Ebert Matthias P. A. , Burgermeister Elke
Published: July 2011

Bile acids from duodenogastric reflux promote inflammation and increase the risk for gastro-oesophageal cancers. FXR (farnesoid X receptor/NR1H4) is a transcription factor regulated by bile acids such as CDCA (chenodeoxycholic acid). FXR protects the liver and the intestinal tract against bile acid overload; however, a functional role for FXR in the stomach has not been described. We detected FXR expression in the normal human stomach and in GC (gastric cancer). FXR mRNA and protein were also present in the human GC cell lines MKN45 and SNU5, but not in the AGS cell line. Transfection of FXR into AGS cells protected against TNFα (tumour necrosis factor α)-induced cell damage. We identified K13 (keratin 13), an anti-apoptotic protein of desmosomes, as a novel CDCA-regulated FXR-target gene. FXR bound to a conserved regulatory element in the proximal human K13 promoter. Gastric expression of K13 mRNA was increased in an FXR-dependent manner by a chow diet enriched with 1% (w/w) CDCA and by indomethacin (35 mg/kg of body weight intraperitoneal) in C57BL/6 mice. FXR-deficient mice were more susceptible to indomethacin-induced gastric ulceration than their WT (wild-type) littermates. These results suggest that FXR increases the resistance of human and murine gastric epithelial cells to inflammation-mediated damage and may thus participate in the development of GC.
Targeting the autolysis loop of urokinase-type plasminogen activator with conformation-specific monoclonal antibodies
Authors: Botkjaer Kenneth A. , Fogh Sarah , Bekes Erin C. , Chen Zhuo , Blouse Grant E. , Jensen Janni M. , Mortensen Kim K. , Huang Mingdong , Deryugina Elena , Quigley James P. , Declerck Paul J. , Andreasen Peter A.
Published: July 2011

Tight regulation of serine proteases is essential for their physiological function, and unbalanced states of protease activity have been implicated in a variety of human diseases. One key example is the presence of uPA (urokinase-type plasminogen activator) in different human cancer types, with high levels correlating with a poor prognosis. This observation has stimulated efforts into finding new principles for intervening with uPA's activity. In the present study we characterize the so-called autolysis loop in the catalytic domain of uPA as a potential inhibitory target. This loop was found to harbour the epitopes for three conformation-specific monoclonal antibodies, two with a preference for the zymogen form pro-uPA, and one with a preference for active uPA. All three antibodies were shown to have overlapping epitopes, with three common residues being crucial for all three antibodies, demonstrating a direct link between conformational changes of the autolysis loop and the creation of a catalytically mature active site. All three antibodies are potent inhibitors of uPA activity, the two pro-uPA-specific ones by inhibiting conversion of pro-uPA to active uPA and the active uPA-specific antibody by shielding the access of plasminogen to the active site. Furthermore, using immunofluorescence, the conformation-specific antibodies mAb-112 and mAb-12E6B10 enabled us to selectively stain pro-uPA or active uPA on the surface of cultured cells. Moreover, in various independent model systems, the antibodies inhibited tumour cell invasion and dissemination, providing evidence for the feasibility of pharmaceutical intervention with serine protease activity by targeting surface loops that undergo conformational changes during zymogen activation.
Structural changes in the BH3 domain of SOUL protein upon interaction with the anti-apoptotic protein Bcl-xL
Authors: Ambrosi Emmanuele , Capaldi Stefano , Bovi Michele , Saccomani Gianmaria , Perduca Massimiliano , Monaco Hugo L.
Published: July 2011

The SOUL protein is known to induce apoptosis by provoking the mitochondrial permeability transition, and a sequence homologous with the BH3 (Bcl-2 homology 3) domains has recently been identified in the protein, thus making it a potential new member of the BH3-only protein family. In the present study, we provide NMR, SPR (surface plasmon resonance) and crystallographic evidence that a peptide spanning residues 147–172 in SOUL interacts with the anti-apoptotic protein Bcl-xL. We have crystallized SOUL alone and the complex of its BH3 domain peptide with Bcl-xL, and solved their three-dimensional structures. The SOUL monomer is a single domain organized as a distorted β-barrel with eight anti-parallel strands and two α-helices. The BH3 domain extends across 15 residues at the end of the second helix and eight amino acids in the chain following it. There are important structural differences in the BH3 domain in the intact SOUL molecule and the same sequence bound to Bcl-xL.
Autotaxin induces lung epithelial cell migration through lysoPLD activity-dependent and -independent pathways
Authors: Zhao Jing , He Donghong , Berdyshev Evgeny , Zhong Mintao , Salgia Ravi , Morris Andrew J. , Smyth Susan S. , Natarajan Viswanathan , Zhao Yutong
Published: July 2011

Lung cell migration is a crucial step for re-epithelialization that in turn is essential for remodelling and repair after lung injury. In the present paper we hypothesize that secreted ATX (autotaxin), which exhibits lysoPLD (lysophospholipase D) activity, stimulates lung epithelial cell migration through LPA (lysophosphatidic acid) generation-dependent and -independent pathways. Release of endogenous ATX protein and activity was detected in lung epithelial cell culture medium. ATX with V5 tag overexpressed conditional medium had higher LPA levels compared with control medium and stimulated cell migration through Gαi-coupled LPA receptors, cytoskeleton rearrangement, phosphorylation of PKC (protein kinase C) δ and cortactin at the leading edge of migrating cells. Inhibition of PKCδ attenuated ATX–V5 overexpressed conditional medium-mediated phosphorylation of cortactin. In addition, a recombinant ATX mutant, lacking lysoPLD activity, or heat-inactived ATX also induced lung epithelial cell migration. Extracelluar ATX bound to the LPA receptor and integrin β4 complex on A549 cell surface. Finally, intratracheal administration of LPS (lipopolysaccharide) into the mouse airway induced ATX release and LPA production in BAL (bronchoalveolar lavage) fluid. These results suggested a significant role for ATX in lung epithelial cell migration and remodelling through its ability to induce LPA production-mediated phosphorylation of PKCδ and cortactin. In addition we also demonstrated assocation of ATX with the epithelial cell-surface LPA receptor and integrin β4.
Mutations in the transmembrane and juxtamembrane domains enhance IL27R transforming activity
Authors: Lambert Que T. , Pradhan Anuradha , Roll J. Devon , Reuther Gary W.
Published: July 2011

Cytokines and their receptors regulate haemopoiesis by controlling cellular growth, survival and differentiation. Thus it is not surprising that mutations of cytokine receptors contribute to the formation of haemopoietic disorders, including cancer. We recently identified transforming properties of IL27R, the ligand-binding component of the receptor for interleukin-27. Although wild-type IL27R exhibits transforming properties in haemopoietic cells, in the present study we set out to determine if the transforming activity of IL27R could be enhanced by mutation. We identified three mutations of IL27R that enhance its transforming activity. One of these mutations is a phenylalanine to cysteine mutation at residue 523 (F523C) in the transmembrane domain of the receptor. The two other mutations identified involve deletions of amino acids in the cytoplasmic juxtamembrane region of the receptor. Expression of each of these mutant IL27R proteins led to rapid cytokine-independent transformation in haemopoietic cells. Moreover, the rate of transformation induced by these mutants was significantly greater than that induced by wild-type IL27R. Expression of these IL27R mutants also induced enhanced activation of JAK (Janus kinase)/STAT (signal transducer and activator of transcription) signalling compared with wild-type. An activating deletion mutation of IL27R enhanced homodimerization of the receptor by a mechanism that may involve disulfide bonding. These transforming IL27R mutants displayed equal or greater transforming activity than bona fide haemopoietic oncogenes such as BCR–ABL (breakpoint cluster region–Abelson murine leukaemia viral oncogene homologue) and JAK2-V617F. Since IL27R is expressed on haemopoietic stem cells, lymphoid cells and myeloid cells, including acute myeloid leukaemia blast cells, mutation of this receptor has the potential to contribute to a variety of haemopoietic neoplasms.
Liver-specific deletion of protein tyrosine phosphatase (PTP) 1B improves obesity- and pharmacologically induced endoplasmic reticulum stress
Authors: Agouni Abdelali , Mody Nimesh , Owen Carl , Czopek Alicja , Zimmer Derek , Bentires-Alj Mohamed , Bence Kendra K. , Delibegović Mirela
Published: July 2011

Obesity is associated with induction of the ER (endoplasmic reticulum)-stress response signalling and insulin resistance. PTP1B (protein tyrosine phosphatase 1B) is a major regulator of adiposity and insulin sensitivity. The aim of the present study was to investigate the role of L-PTP1B (liver-specific PTP1B) in chronically HFD (high-fat diet) and pharmacologically induced (tunicamycin and thapsigargin) ER-stress response signalling in vitro and in vivo. We assessed the effects of ER-stress response induction on hepatic PTP1B expression, and consequences of hepatic-PTP1B deficiency, in cells and mouse liver, on components of ER-stress response signalling. We found that PTP1B protein and mRNA expression levels were up-regulated in response to acute and/or chronic ER stress, in vitro and in vivo. Silencing PTP1B in hepatic cell lines or mouse liver (L-PTP1B−/−) protected against induction of pharmacologically induced and/or obesity-induced ER stress. The HFD-induced increase in CHOP (CCAAT/enhancer-binding protein homologous protein) and BIP (binding immunoglobulin protein) mRNA levels were partially inhibited, whereas ATF4 (activated transcription factor 4), GADD34 (growth-arrest and DNA-damage-inducible protein 34), GRP94 (glucose-regulated protein 94), ERDJ4 (ER-localized DnaJ homologue) mRNAs and ATF6 protein cleavage were completely suppressed in L-PTP1B−/− mice relative to control littermates. L-PTP1B−/− mice also had increased nuclear translocation of spliced XBP-1 (X box-binding protein-1) via increased p85α binding. We demonstrate that the ER-stress response and L-PTP1B expression are interlinked in obesity- and pharmacologically induced ER stress and this may be one of the mechanisms behind improved insulin sensitivity and lower lipid accumulation in L-PTP1B−/− mice.
OEP61 is a chaperone receptor at the plastid outer envelope
Authors: von Loeffelholz Ottilie , Kriechbaumer Verena , Ewan Richard A. , Jonczyk Rafal , Lehmann Susann , Young Jason C. , Abell Ben M.
Published: July 2011

Chloroplast precursor proteins encoded in the nucleus depend on their targeting sequences for delivery to chloroplasts. There exist different routes to the chloroplast outer envelope, but a common theme is the involvement of molecular chaperones. Hsp90 (heat-shock protein 90) delivers precursors via its receptor Toc64, which transfers precursors to the core translocase in the outer envelope. In the present paper, we identify an uncharacterized protein in Arabidopsis thaliana OEP61 which shares common features with Toc64, and potentially provides an alternative route to the chloroplasts. Sequence analysis indicates that OEP61 possesses a clamp-type TPR (tetratricopeptide repeat) domain capable of binding molecular chaperones, and a C-terminal TMD (transmembrane domain). Phylogenetic comparisons show sequence similarities between the TPR domain of OEP61 and those of the Toc64 family. Expression of mRNA and protein was detected in all plant tissues, and localization at the chloroplast outer envelope was demonstrated by a combination of microscopy and in vitro import assays. Binding assays show that OEP61 interacts specifically with Hsp70 (heat-shock protein 70) via its TPR clamp domain. Furthermore, OEP61 selectively recognizes chloroplast precursors via their targeting sequences, and a soluble form of OEP61 inhibits chloroplast targeting. We therefore propose that OEP61 is a novel chaperone receptor at the chloroplast outer envelope, mediating Hsp70-dependent protein targeting to chloroplasts.
A drug targeting only p110α can block phosphoinositide 3-kinase signalling and tumour growth in certain cell types
Authors: Jamieson Stephen , Flanagan Jack U. , Kolekar Sharada , Buchanan Christina , Kendall Jackie D. , Lee Woo-Jeong , Rewcastle Gordon W. , Denny William A. , Singh Ripudaman , Dickson James , Baguley Bruce C. , Shepherd Peter R.
Published: July 2011

Genetic alterations in PI3K (phosphoinositide 3-kinase) signalling are common in cancer and include deletions in PTEN (phosphatase and tensin homologue deleted on chromosome 10), amplifications of PIK3CA and mutations in two distinct regions of the PIK3CA gene. This suggests drugs targeting PI3K, and p110α in particular, might be useful in treating cancers. Broad-spectrum inhibition of PI3K is effective in preventing growth factor signalling and tumour growth, but suitable inhibitors of p110α have not been available to study the effects of inhibiting this isoform alone. In the present study we characterize a novel small molecule, A66, showing the S-enantiomer to be a highly specific and selective p110α inhibitor. Using molecular modelling and biochemical studies, we explain the basis of this selectivity. Using a panel of isoform-selective inhibitors, we show that insulin signalling to Akt/PKB (protein kinase B) is attenuated by the additive effects of inhibiting p110α/p110β/p110δ in all cell lines tested. However, inhibition of p110α alone was sufficient to block insulin signalling to Akt/PKB in certain cell lines. The responsive cell lines all harboured H1047R mutations in PIK3CA and have high levels of p110α and class-Ia PI3K activity. This may explain the increased sensitivity of these cells to p110α inhibitors. We assessed the activation of Akt/PKB and tumour growth in xenograft models and found that tumours derived from two of the responsive cell lines were also responsive to A66 in vivo. These results show that inhibition of p110α alone has the potential to block growth factor signalling and reduce growth in a subset of tumours.
Crystal structure of a novel type of odorant-binding protein from Anopheles gambiae, belonging to the C-plus class
Authors: Lagarde Amandine , Spinelli Silvia , Qiao Huili , Tegoni Mariella , Pelosi Paolo , Cambillau Christian
Published: July 2011

Agam (Anopheles gambiae) relies on its olfactory system to target human prey, leading eventually to the injection of Plasmodium falciparum, the malaria vector. OBPs (odorant-binding proteins) are the first line of proteins involved in odorant recognition. They interact with olfactory receptors and thus constitute an interesting target for insect control. In the present study, we undertook a large-scale analysis of proteins belonging to the olfactory system of Agam with the aim of preventing insect bites by designing strong olfactory repellents. We determined the three-dimensional structures of several Agam OBPs, either alone or in complex with model compounds. In the present paper, we report the first three-dimensional structure of a member of the C-plus class of OBPs, AgamOBP47, which has a longer sequence than classical OBPs and contains six disulfide bridges. AgamOBP47 possesses a core of six α-helices and three disulfide bridges, similar to the classical OBP fold. Two extra loops and the N- and C-terminal extra segments contain two additional α-helices and are held in conformation by three disulfide bridges. They are located either side of the classical OBP core domain. The binding site of OBP47 is located between the core and the additional domains. Two crevices are observed on opposite sides of OBP47, which are joined together by a shallow channel of sufficient size to accommodate a model of the best-tested ligand. The binding sites of C-plus class OBPs therefore exhibit different characteristics, as compared with classical OBPs, which should lead to markedly diverse functional implications.
A combined action of pulmonary surfactant proteins SP-B and SP-C modulates permeability and dynamics of phospholipid membranes
Authors: Parra Elisa , Moleiro Lara H. , López-Montero Ivan , Cruz Antonio , Monroy Francisco , Pérez-Gil Jesús
Published: July 2011

Proteins SP-B and SP-C are essential to promote formation of surface-active films at the respiratory interface, but their mechanism of action is still under investigation. In the present study we have analysed the effect of the proteins on the accessibility of native, quasi-native and model surfactant membranes to incorporation of the fluorescent probes Nile Red (permeable) and FM 1-43 (impermeable) into membranes. We have also analysed the effect of single or combined proteins on membrane permeation using the soluble fluorescent dye calcein. The fluorescence of FM 1-43 was always higher in membranes containing SP-B and/or SP-C than in protein-depleted membranes, in contrast with Nile Red which was very similar in all of the materials tested. SP-B and SP-C promoted probe partition with markedly different kinetics. On the other hand, physiological proportions of SP-B and SP-C caused giant oligolamellar vesicles to incorporate FM 1-43 from the external medium into apparently most of the membranes instantaneously. In contrast, oligolamellar pure lipid vesicles appeared to be mainly labelled in the outermost membrane layer. Pure lipidic vesicles were impermeable to calcein, whereas it permeated through membranes containing SP-B and/or SP-C. Vesicles containing only SP-B were stable, but prone to vesicle–vesicle interactions, whereas those containing only SP-C were extremely dynamic, undergoing frequent fluctuations and ruptures. Differential structural effects of proteins on vesicles were confirmed by electron microscopy. These results suggest that SP-B and SP-C have different contributions to inter- and intra-membrane lipid dynamics, and that their combined action could provide unique effects to modulate structure and dynamics of pulmonary surfactant membranes and films.
Intrinsic membrane association of the cytoplasmic tail of influenza virus M2 protein and lateral membrane sorting regulated by cholesterol binding and palmitoylation
Authors: Thaa Bastian , Levental Ilya , Herrmann Andreas , Veit Michael
Published: July 2011

The influenza virus transmembrane protein M2 is a proton channel, but also plays a role in the scission of nascent virus particles from the plasma membrane. An amphiphilic helix in the CT (cytoplasmic tail) of M2 is supposed to insert into the lipid bilayer, thereby inducing curvature. Palmitoylation of the helix and binding to cholesterol via putative CRAC (cholesterol recognition/interaction amino acid consensus) motifs are believed to target M2 to the edge of rafts, the viral-budding site. In the present study, we tested pre-conditions of this model, i.e. that the CT interacts with membranes, and that acylation and cholesterol binding affect targeting of M2. M2-CT, purified as a glutathione transferase fusion protein, associated with [3H]photocholesterol and with liposomes. Mutation of tyrosine residues in the CRAC motifs prevented [3H]photocholesterol labelling and reduced liposome binding. M2-CT fused to the yellow fluorescent protein localized to the Golgi in transfected cells; membrane targeting was dependent on CRAC and (to a lesser extent) on palmitoylation. Preparation of giant plasma membrane vesicles from cells expressing full-length M2–GFP (green fluorescent protein) showed that the protein is partly present in the raft domain. Raft targeting required palmitoylation, but not the CRAC motifs. Thus palmitoylation and cholesterol binding differentially affect the intrinsic membrane binding of the amphiphilic helix.
Dock/Nck facilitates PTP61F/PTP1B regulation of insulin signalling
Authors: Wu Chia-Lun , Buszard Bree , Teng Chun-Hung , Chen Wei-Lin , Warr Coral G. , Tiganis Tony , Meng Tzu-Ching
Published: July 2011

PTP1B (protein tyrosine phosphatase 1B) is a negative regulator of IR (insulin receptor) activation and glucose homoeostasis, but the precise molecular mechanisms governing PTP1B substrate selectivity and the regulation of insulin signalling remain unclear. In the present study we have taken advantage of Drosophila as a model organism to establish the role of the SH3 (Src homology 3)/SH2 adaptor protein Dock (Dreadlocks) and its mammalian counterpart Nck in IR regulation by PTPs. We demonstrate that the PTP1B orthologue PTP61F dephosphorylates the Drosophila IR in S2 cells in vitro and attenuates IR-induced eye overgrowth in vivo. Our studies indicate that Dock forms a stable complex with PTP61F and that Dock/PTP61F associate with the IR in response to insulin. We report that Dock is required for effective IR dephosphorylation and inactivation by PTP61F in vitro and in vivo. Furthermore, we demonstrate that Nck interacts with PTP1B and that the Nck/PTP1B complex inducibly associates with the IR for the attenuation of IR activation in mammalian cells. Our studies reveal for the first time that the adaptor protein Dock/Nck attenuates insulin signalling by recruiting PTP61F/PTP1B to its substrate, the IR.
Characterization and functional analysis of GhRDR6, a novel RDR6 gene from cotton (Gossypium hirsutum L.)
Authors: Wang Mian , Li Shanwei , Yang Haifang , Gao Zheng , Wu Changai , Guo Xingqi
Published: July 2011

RDR6 (RNA-dependent RNA polymerase 6) is not only involved in virus resistance but also plays an important role in natural plant development. In the present study, a novel RDR gene, named GhRDR6 (Gossypium hirsutum RDR6), was isolated from cotton (G. hirsutum L.). Alignment and evolutionary relationship analyses showed that GhRDR6 was more closely related to RDR6 than to other RDRs. Expression analysis indicated that this single-copy gene is constitutively expressed in the roots, stems and leaves. Semi-quantitative RT–PCR (reverse transcription–PCR) showed that GhRDR6 was up-regulated by the application of various phytohormones, including MeJA [methyl JA (jasmonate)], ABA (abscisic acid), JA, α-naphthylacetic acid, gibberellins and ET (ethylene). In addition, GhRDR6 expression increased in response to wounding, cold (4°C) and NaCl treatments, but not by drought. Furthermore, overexpression of GhRDR6 in transgenic Nicotiana benthamiana plants resulted in root lengths longer than the wide-type during the seeding stage. Interestingly, the GhRDR6-overexpressing plants displayed reduced tolerance to oxidative damage, resulting in reduced ABA-sensitivity, but they tolerated freezing. Moreover, resistance to potato virus Y was enhanced in transgenic N. benthamiana plants. These results suggest that GhRDR6 may play an important role in plant defence responses and a pivotal role in plant development.
Permissive effect of EGFR-activated pathways on RVI and their anti-apoptotic effect in hypertonicity-exposed mIMCD3 cells
Authors: Ruiz-Martínez Alejandro , Vázquez-Juárez Erika , Ramos-Mandujano Gerardo , Pasantes-Morales Herminia
Published: July 2011

Hypertonicity is a stressful stimulus leading to cell shrinkage and apoptotic cell death. Apoptosis can be prevented if cells are able to activate the mechanism of RVI (regulatory volume increase). This study in mIMCD3 cells presents evidence of a permissive role of the EGFR (epidermal growth factor receptor) on RVI, achieved for the most part through the two main EGFR-triggered signalling chains, the MAPK (mitogen-activated protein kinase)/ERK (extracellular-signal-regulated kinase) and the PI3K (phosphoinositide 3-kinase)/Akt (also known as protein kinase B) pathways. Hyperosmotic solutions (450 mosM) made by addition of NaCl, increased EGFR phosphorylation, which is prevented by GM6001 and AG1478, blockers respectively, of MMPs (matrix metalloproteinases) and EGFR. Inhibition of EGFR, ERK (PD98059) or PI3K/Akt (wortmannin) phosphorylation reduced RVI by 60, 48 and 58% respectively. The NHE (Na+/H+ exchanger) seems to be the essential mediator of this effect since (i) NHE is the main contributor to RVI, (ii) EGFR, ERK and PI3K/Akt blockers added together with the NHE blocker zoniporide reduce RVI by non-additive effects and (iii) All the blockers significantly lowered the NHE rate in cells challenged by an NH4Cl pulse. Besides reducing RVI, the inhibition of MMP, EGFR and PI3K/Akt had a strong pro-apoptotic effect increasing cell death by 2–3.7-fold. This effect was significantly lower when RVI inhibition did not involve the EGFR-PI3K/Akt pathway. These results provide evidence that Akt and its permissive effect on RVI have a predominant influence on cell survival under hypertonic conditions in IMCD3 cells. This role of Akt operates under the influence of EGFR activation, promoted by MMP.