bluuebunny/crossref_metadata_2025 huggingface dataset:
CLONING AND CHARACTERIZATION OF TWO EcR ISOFORMS FROM JAPANESE PINE SAWYER,Monochamus alternates
Authors: Weng Hongbiao , Shen Weifeng , Liu Yan , He Lihua , Niu Baolong , Meng Zhiqi , Mu Jianjun
Published: August 2013

The ecdysone receptor (EcR) is the hormonal receptor of ecdysteroids, which regulates insect growth and development. In this study, we cloned and characterized two isoforms ofEcRinMonochamus alternatesnamedMaEcRA andMaEcR B. The cDNAs ofMaEcR AandMaEcR Bhave open repeating frames of 1,695 and 1,392 bp, respectively. The deduced proteins have the same C‐terminal sequence and varied in N‐terminal, and are consistent with reports on other insect species, particularly with the receptor of another coleopteran,Tribolium castaneum. The isoform‐specific developmental expression profile ofEcRin the epidermis and the midgut were analyzed with quantitative real‐time reverse‐transcriptase polymerase chain reaction in the pupal stage. RNA interference (RNAi) with common or isoform‐specific regions induced developmental stagnation. When treated in the later larval stage, RNAi with either the common sequence or anEcR Aspecific sequence caused more severe effects and most larvae died prior to adulthood. TheEcR Bspecific sequence caused less severe effects and about half of the treated larvae became adults, but some showed developmental defects. RNAi with both isoforms at early pupal stage attenuated the expression of 20E‐regulated genesE74,E75, andHR3. The study demonstrates the role of EcR in the transduction of ecdysteroid response inMonochamus alternatus.
Visualizing the history of evidence‐based medicine: A bibliometric analysis
Authors: Shen Jiantong , Yao Leye , Li Youping , Clarke Mike , Wang Li , Li Dan
Published: August 2013

The aim of this paper is to visualize the history of evidence‐based medicine (EBM) and to examine the characteristics of EBM development in China and the West. We searched the Web of Science and the Chinese National Knowledge Infrastructure database for papers related to EBM. We applied information visualization techniques, citation analysis, cocitation analysis, cocitation cluster analysis, and network analysis to construct historiographies, themes networks, and chronological theme maps regarding EBM in China and the West. EBM appeared to develop in 4 stages: incubation (1972–1992 in the West vs. 1982–1999 in China), initiation (1992–1993 vs. 1999–2000), rapid development (1993–2000 vs. 2000–2004), and stable distribution (2000 onwards vs. 2004 onwards). Although there was a lag in EBM initiation in China compared with the West, the pace of development appeared similar. Our study shows that important differences exist in research themes, domain structures, and development depth, and in the speed of adoption between China and the West. In the West, efforts in EBM have shifted from education to practice, and from the quality of evidence to its translation. In China, there was a similar shift from education to practice, and from production of evidence to its translation. In addition, this concept has diffused to other healthcare areas, leading to the development of evidence‐based traditional Chinese medicine, evidence‐based nursing, and evidence‐based policy making.
Robust Unknown Input Observer Design for Linear Uncertain Time Delay Systems with Application to Fault Detection
Authors: Ahmadizadeh Saeed , Zarei Jafar , Karimi Hamid Reza
Published: August 2013

In this paper, a novel approach is proposed to design a robust fault detection observer for uncertain linear time delay systems. The system is composed of both norm‐bounded uncertainties and exogenous signals (noise, disturbance, and fault) which are considered to be unknown. The main contribution of this paper is to present unknown input observer (UIO)‐based fault detection system which shows the maximum sensitivity to fault signals and the minimum sensitivity to other signals. Since the system contains uncertainty terms, an H∞ model‐matching approach is used in design procedure. The reference residual signal generator system is designed so that the fault signal has maximum sensitivity while the exogenous signals have minimum sensitivity on the residual signal. Then, the fault detection system is designed by minimizing the estimation error between the reference residual signal and the UIO residual signal in the sense of H∞ norm. A sufficient condition for the existence of such a filter is exploited in terms of certain linear matrix inequalities (LMIs). Application of the proposed method in a numerical example and an engineering process are simulated to demonstrate the effectiveness of the proposed algorithm. Simulation results show the validity of the proposed approach to detect the occurrence of faults in the presence of modeling errors, disturbances, and noise.
Die energetische Sanierung der Stadtbibliothek Nürnberg – Entwicklung des Energie‐ und Klimakonzepts mittels hygrothermischer Gebäudesimulation
Authors: Anlauft Eva , Meinhold Uwe , Wagner Moritz , Wenzel Urs
Published: August 2013

Das 1911 errichtete und nach weitgehender Zerstörung 1956 wieder aufgebaute Luitpoldhaus wurde im Rahmen der Zusammenlegung von drei bisher räumlich separaten Einrichtungen der Stadtbibliothek Nürnberg an einen Standort generalsaniert, umgebaut und erweitert. Mit der baulichen und anlagentechnischen Komplettsanierung verbindet sich eine städtebauliche Verbesserung des “soziokulturellen Zentrums südliche Altstadt“. Besonders anspruchsvoll ist die konservatorisch angemessene Unterbringung des für die Stadt Nürnberg überaus wertvollen Bestandes an mittelalterlichen Handschriften, Inkunabeln, alten Drucken und Karten. Mit der Sanierung wurde eine wesentliche Verbesserung der Energieeffizienz von mindestens 30 % unter dem Anforderungsniveau der Energieeinsparverordnung 2007 für Neubauten angestrebt. Die strengen konservatorischen Raumklimabedingungen sollen dabei mit weitgehend passiven baulichen Maßnahmen unter Einbindung innovativer Lösungen sowie mit minimierter anlagentechnischer Ausstattung gewährleistet werden. Für die Auswahl und Dimensionierung der Maßnahmen wurden umfangreiche thermische und hygrische Raumklima‐ und Strömungssimulationen in den Planungsprozess integriert.

The energy optimised renovation of Nuremberg City Library: Designing for energy and climate using hygrothermal building simulation. The Luitpoldhaus was built in 1911, largely destroyed in 1945, then rebuilt in 1956. Three previously separate built elements of a Nuremberg City Library site were recently combined, refurbished, converted and expanded. This thoroughgoing renovation involving both construction and systems engineering was combined with urban planning improvements to the southern part of the old town, a socio‐cultural focal point in the city. One particular challenge faced was how to house the city‘s treasured collection of mediaeval manuscripts and incunabula, old prints and maps in an environment suited to their conservation. The renovation has significantly improved energy efficiency at the site: the target was at least 30 % below the level required of new buildings under the 2007 German Building Energy Conservation Ordinance. The stringent conservation requirements on indoor climatic conditions will be maintained with the extensive use of passive construction techniques, involving innovative solutions and a minimum of systems engineering equipment. Comprehensive thermal and hygrothermal climatic and flow simulations were incorporated into the planning process to help select and dimension the relevant measures.
Untersuchung zur Dämmwirkung von Wärmedämm‐Materialien mit Infrarot reflektierenden Oberflächen
Authors: Al‐Sibai Faruk , Hillemacher Bernd , Burghold Michael , Kneer Reinhold
Published: August 2013

Für Wärmedämm‐Materialien mit Infrarot reflektierenden Oberflächen gibt es bisher keine Möglichkeiten, die wesentlichen Merkmale der IR‐Folien zu beschreiben und eine Möglichkeit zu etablieren, den IR‐Dämmprodukten verständliche, produktspezifische und wärmetechnische Eigenschaften zuzuweisen. Für Lufträume, deren Begrenzungsflächen niedrige Emissionsgrade aufweisen, sind die Näherungsverfahren der DIN EN ISO 6946 nur eingeschränkt geeignet, da die dominierenden, konvektiven Prozesse in den Lufträumen nicht genau genug berechnet werden können. Es wird daher ein universelles Verfahren für die Berechnung des Wärmedurchlasswiderstandes durch ein Bauteil vorgestellt, das im Besonderen die Wärmeübergangsprozesse durch Leitung, Konvektion und Strahlung in den Lufträumen detailliert mit Hilfe dimensionsloser Wärmeübergangskoeffizienten für unterschiedlichste Rayleigh‐Zahl‐Bereiche simuliert und quantitative Voraussagen über den Bemessungswert des Wärmedurchlasswiderstandes R des Dämmsystems und somit auch über den Wärmedurchgangskoeffizienten U durch die gesamte Baukonstruktion erlaubt.

Analysis of the insulation effect of infrared reflective products. Up to now there are no adequate methods to quantify the thermal resistance of IR‐reflective insulation products. For air filled rooms with low‐emissivity bounding surfaces, the approximation procedure of the DIN EN ISO 6946 is not suitable, because the dominating convective heat transfer processes cannot be calculated precisely enough. The Institute of Heat and Mass Transfer at RWTH Aachen University has developed a simulation procedure for the calculation of the simultaneous heat transfer by conduction, convection and radiation in a fluid layer in detail for various Rayleigh number ranges with the help of Nusselt correlations to obtain quantitative predictions for the thermal resistance of different roof systems.
Teacher perspectives on factors facilitating implementation of whole school approaches for resolving conflict
Authors: Freeman Elizabeth , Wertheim Eleanor H. , Trinder Margot
Published: August 2013

While social–emotional learning programmes in schools often have positive outcomes, many such initiatives are not well implemented and maintained. This paper reports on teacher reflections on the process of planning and implementing a whole school social–emotional learning (SEL) programme with a conflict resolution focus, called Enhancing Relationships in School Communities (ERIS). Ten primary schools participated in an 18‐month professional development programme offered by the researchers to assist schools to develop a whole school approach to handling interpersonal issues using a cooperative problem‐solving approach. Implementation (core) teams of three to five members, including principal or assistant principal and relevant teachers, attended workshops and managed the change process. In individual interviews, 29 core team members described the most significant changes that had taken place, what factors facilitated those changes and what assisted in overcoming barriers. A synthesised model of facilitators of implementation was developed on the basis of these interviews. The major components of the model included ensuring a whole school vision and process; pre‐programme engagement confirming commitment and alignment of researcher and teachers' visions; facilitative programme structure and processes such as linking the current programme to existing programmes and processes in the school; leadership and support for staff in the change process, for example, through the implementation team; the nature of the programme content; and monitoring and feedback processes to sustain motivation and inform on needs and outcomes.
‘Digital natives’: Honour and respect in computerized encounters between Israeli Jewish and Arab children and adult learners
Authors: Gamliel Tova , Hazan Haim
Published: August 2013

In Israel's Multigenerational Connection Program (MCP), children instruct adults in computer and Internet use. Taking children's advantage in digital literacy as a given, the study examines their generational status in computerized encounters that MCP creates in two schools, one Jewish and one Arab. The data were gathered by means of qualitative participant observation. The results suggest that the family–community‐based habitus is reflected in the interpretation of the program by the computer teacher at each school, who, by ‘importing’ it to MCP, encouraged relations of respect or honour between children and adults. The significance of these relations, rooted in ethno‐national relations between the groups, transcends the global discourse about the uniformity of the generational digital divide. While the Arab group's ambivalence toward MCP weakened the children's status, the Jewish children attained empowerment as young teachers. The conclusions focus on the implications for intergenerational relations and children's self‐image of the preservation of the Arab honour‐contingent habitus in a Western educational context—a self‐chosen strategy that may widen the digital divide between Arab and Jewish children.
Polyploidie: Mehr ist nicht immer besser
Authors: Kronberg Inge
Published: August 2013

Polyploidie ist vor allem im Pflanzenreich eine verbreitete Genommutation. Die Vervielfachung des Genoms verändert den Phänotyp: Aus kleinen wildwachsenden Holzäpfeln werden große, saftige Kulturäpfel, Baumwollfäden werden lang und spinnbar, Weizen ertragreich und dreschfähig. Die Polyploidisierung ist aber nicht nur das Ergebnis von Züchtung, sondern kommt natürlicherweise bei den meisten heutigen Pflanzen vor. Sie hat Impulse in der Evolution gegeben, aber auch in Sackgassen geführt. Was sich auf genetischer und epigenetischer Ebene dahinter verbirgt, wird trotz neuer Labormethoden erst allmählich klarer.
In jedem Winkel ein Netz: Spinnen‐Vielfalt rund ums Haus
Authors: Petrischak Hannes
Published: August 2013

Spinnen lieben die vielfältigen Strukturen menschlicher Bauwerke: In Winkeln und Spalten finden sie ihre Verstecke, und viele Insekten, angelockt vom Licht, gehen ihnen hier ins Netz. Auch wer sich vielleicht zunächst vor diesen Tieren gruselt, wird bei genauerer Betrachtung schnell von ihnen fasziniert sein.
Mycophenolic acid regulates spleen tyrosine kinase to repress tumour necrosis factor‐alpha‐induced monocyte chemotatic protein‐1 production in cultured human aortic endothelial cells
Authors: Koo Tai Yeon , Kim Yoon Ji , Yang Won Seok , Park Jung Sik , Han Nam Jeong , Lee Joo Mi , Park Su Kil
Published: August 2013

Atherosclerosis develops from cascades of inflammatory processes. Spleen tyrosine kinase (Syk) and monocyte chemotatic protein‐1 (MCP‐1) play important roles in the pathogenesis of atherosclerosis. Mycophenolic acid (MPA) has an anti‐inflammatory effect. We have investigated whether MPA regulates Syk to repress tumour necrosis factor‐α (TNF‐α)‐induced MCP‐1 production in cultured human aortic endothelial cells. Expression of MCP‐1 mRNA and its protein were measured by real time RT‐PCR and ELISA, respectively. Reactive oxygen species (ROS) production were measured using 2′7′‐dichlorofluorescein diacetate. Activation of AP‐1 and NF‐κB were assessed by electrophoretic mobility shift assay. Tyrosine phosphorylation of Syk was examined by Western blot analysis. TNF‐α increased MCP‐1 at both mRNA and protein levels. TNF‐α‐induced MCP‐1 mRNA expression was inhibited by N‐acetylcysteine (NAC), Syk inhibitor, Syk‐siRNA and MPA. TNF‐α‐induced MCP‐1 protein production was also inhibited by Syk inhibitor and MPA. TNF‐α increased DNA binding activity of AP‐1 and NF‐κB, whereas both AP‐1 and NF‐κB decoy oligodeoxynucleotides downregulated TNF‐α‐induced MCP‐1 mRNA expression. TNF‐α increased ROS generation, which was inhibited by NAC and MPA, but not by Syk inhibitor. TNF‐α increased tyrosine phosphorylation of Syk, which was attenuated by NAC and MPA. MPA and Syk inhibitor attenuated TNF‐α‐induced DNA binding activity of NF‐κB and AP‐1. TNF‐α induced MCP‐1 expression via activation of AP‐1 and NF‐κB. AP‐1 and NF‐κB were mediated through ROS, followed by Syk. MPA exerts anti‐inflammatory effect by inhibiting MCP‐1 expression via suppression of ROS and Syk.
All‐trans retinoic acid displays multiple effects on the growth, lipogenesis and adipokine gene expression of AML‐I preadipocyte cell line
Authors: Morikawa Keiko , Hanada Haruka , Hirota Kaori , Nonaka Mitsuko , Ikeda Chiharu
Published: August 2013

Adipose tissue is a potential site of retinoic acid (RA) action, but its physiological significance remains to be clarified. We have examined the effect of all‐trans retinoic acid (ATRA) on growth and differentiation of preadipocytes, and on adipokine gene expression in mature adipocytes using human preadipocyte cell model, AML‐I. Both ATRA and 9‐cis RA induced growth arrest in AML‐I preadipocyte at between 50 and 100 µM, which was accompanied by apoptosis. Western blotting showed a loss of NF‐κB, Bcl‐2 and p‐Akt, and the accumulation of Bad and Akt in cytoplasm of ATRA‐treated AML‐I preadipocytes. Exposure of AML‐I to ATRA or 9‐cis RA increased intracellular lipid accumulation in a time‐dependent manner compared to vehicle‐treated cells. Expression of fatty acid synthase (FAS) and peroxisome proliferator‐activated receptor‐γ (PPAR‐γ) proteins was increased in ATRA‐treated cells. Thus, both ATRA and 9‐cis RA promoted differentiation, inhibited proliferation and induced apoptosis in AML‐I preadipocytes. ATRA also modulated adipokine expression by increasing the mRNA level of adipocytokines (adiponectin, leptin and LPL), and by inhibiting PAI‐1 mRNA expression in mature AML‐I adipocytes. The data suggest that ATRA exerts a wide range of effects—growth arrest, apoptosis, lipogenesis and modulation of adipokine gene expression—during the maturation of preadipocytes into adipocytes.
Role of 14‐3‐3σ in resistance to cisplatin in non‐small cell lung cancer cells
Authors: Cetintas Vildan Bozok , Tetik Aslı , Cok Gursel , Kucukaslan Ali Sahin , Kosova Buket , Gunduz Cumhur , Veral Ali , Eroglu Zuhal
Published: August 2013

The efficacies of chemotherapeutic agents are often limited by side effects and acquired drug resistance. We have investigated whether the differential expression pattern of 14‐3‐3σ affects cisplatin response in non‐small cell lung cancer cell lines. Two pairs of parental/cisplatin resistant cell lines (A549/CRA549 and Calu1/CR‐Calu1) and clinical lung cancer biopsy samples were analysed for 14‐3‐3σ expression. Cell viability was assessed by WST assay; and 14‐3‐3σ expression was suppressed by siRNA transfection. 14‐3‐3σ mRNA expression increased in CR‐A549 and CR‐Calu1 compared with their cisplatin‐sensitive parental A549 and Calu1 cell lines. But when 14‐3‐3σ expression was suppressed, elevated cisplatin response was seen in A549 and CR‐Calu1 cell lines. Increased 14‐3‐3σ expression might also account for reduced cisplatin response in vivo, since, 14‐3‐3σ expression in clinical biopsy samples obtained from lung cancer patients undergoing cisplatin‐based chemotherapy significantly higher in the non‐responder compared with the responder group. We therefore propose that increased 14‐3‐3σ expression is correlated with cisplatin response in non‐small cell lung cancer cells; monitoring its expression might become useful in the future in predicting poor outcome to cisplatin treatment and/or the verification of acquired cisplatin resistance in lung cancer patients.
Protective effects of ADAM8 against cisplatin‐mediated apoptosis in non‐small‐cell lung cancer
Authors: Zhang Weifeng , Wan Minghui , Ma Lunchao , Liu Xiang , He Jianxing
Published: August 2013

A disintegrin, metalloproteinase 8 (ADAM8), is overexpressed in the vast majority of lung cancers and can be a diagnostic marker of lung cancer. We have investigated the effect of ADAM8 on the cisplatin resistance in non‐small‐cell lung cancer (NSCLC) cell lines. Stable cell lines overexpressing ADAM8 in A549 and H460 cells were generated, both of which have low endogenous ADAM8. Ectopic expression of ADAM8 rendered cells more resistant to cisplatin‐induced toxicity, increasing the half maximal inhibitory concentration (IC50) values by 1.85‐fold in A549 cells and 3.91‐fold in H460 cells relative to mock‐transfected cells. Moreover, silencing of ADAM8 in H647 cells with high endogenous level of ADAM8 sensitised them to cisplatin‐induced toxicity, with a lower IC50 value of 11.2 µM relative to an IC50 of 25.3 µM in mock‐transfected cells. Moreover, knockdown of ADAM8 caused a significant increase in cisplatin‐induced apoptosis assessed by annexin‐V/propidium iodide double staining, accompanying with enhanced cleavage of caspase‐3 and poly(ADP‐ribose) polymerase. Western blot analysis showed that a greater amount of phosphorylated signal transducer and activator of transcription 3 (STAT3) in ADAM8‐overexpressing A549 cells compared to parental or mock‐transfected cells. STAT3 silencing increased the susceptibility of ADAM8‐overexpressing A549 cells to cisplatin. Both Bcl‐2 and Mcl‐1 in ADAM8‐overexpressing A549 cells were profoundly diminished by STAT3 knockdown. Thus, ADAM8 is implicated in cisplatin resistance of NSCLC cells through activation of the STAT3 signalling pathway, and thus represents a potential therapeutic target in this malignancy.
Flavonoid hesperidin protects neural crest cells from death caused by aflatoxin B1
Authors: Nones Jader , Nones Janaína , Trentin Andrea Gonçalves
Published: August 2013

The neural crest (NC) corresponds to a collection of multipotent and oligopotent progenitors endowed with both neural and mesenchymal potentials. The derivatives of the NC at trunk level include neurons and glial cells of the peripheral nervous system. Despite the well‐known influence of aflatoxins on the development of cancer, the issue of whether they also influence NC cells has not been yet addressed. In the present work, we have investigated the effects of aflatoxin B1 on quail NC cells and the concomitant effects of the flavonoid hesperidin associated with this mycotoxin. We show for the first time that aflatoxin B1 decreases the viability and the total number of glial and neuronal cells/field, although their proportions in relation to the total number of cells were not altered. Therefore, aflatoxin has no effect on NC differentiation. However, this compound was able to reduce NC proliferation and NC survival. Furthermore, the co‐administration of hesperidin, a well‐known polyphenolic protector of cell death, partially prevented the effect of aflatoxin B1. Taken together, our results demonstrate that aflatoxin B1 is toxic to NC cells, an effect partially prevented by the flavonoid hesperidin. This study may contribute to the understanding of the effects of these compounds during early embryonic development and offer potentially more assertive diets and treatments for pregnant animals.
Association of nucleus and centrosome: magnet or velcro?
Authors: Burakov Anton V. , Nadezhdina Elena S.
Published: August 2013

A structural link between cell's nucleus and centrosome was proposed years ago. Such a link was suggested to maintain nucleus–centrosome axis, determine polarity of interphase cells and ensure spindle assembly in mitotic cells. The idea of structural link is supported by the facts that centrosomes are usually located in close proximity to the nuclei and remain attached to the nuclei in mildly homogenated cells. However, juxtaposed location can result rather from the tendency of both organelles to occupy central position in cell than from the existence of a specific structural link. Moreover, the nucleus was shown to be transported towards the centrosome along microtubules by dynein bound to nuclear envelope; inhibition of dynein results in the increase of nucleus–centrosome distance. The interaction of both organelles is disturbed in response to actin depolymerisation, although the exact role of actin filaments in this process remains unknown. The link between the nucleus and the centrosome can support simultaneous migration of nuclei and centrosomes in large cells and in syncytia, but its existence in interphase fibroblast‐like and epithelia‐like cells was not confirmed yet. Further studies include direct visualisation of a specific link between centrosome and nucleus and elucidation of actin role in its formation.
Measurement of entropy production in living cells under an alternating electric field
Authors: Ding Changjiang , Luo Liaofu
Published: August 2013

Entropy is a thermodynamic property toward equilibrium based on the dissipation of energy. Cells constitute such a thermodynamic system, in which entropy production is both inevitable and highly significant. Although the experimental measurement of entropy production in a cell is very difficult, a new method to accomplish this in living cells is reported herein. Through heating the sample by alternating electric fields and recording the heat flow from cells, the entropy production in two normal cell lines, MCF10A and HL‐7702, and two cancerous cell lines, MDA‐MB‐231 and SMMC‐7721, was measured and compared. The scaled electroinduced entropy production rate (SEEP) of cancer cells monotonically increases with electric field strength at 5–40 V/cm, while that of normal cells changes nonmonotonically with electric field strength, reaching a peak at 5–30 V/cm. For all cell lines, the cancerous‐to‐normal ratio of field‐induced entropy production is clearly <1 in a large range of field strength from 5 to 25 V/cm. Therefore, this work presents an easy and effective strategy for experimentally investigating the thermodynamic properties of the cell, and gives deeper insight into the physical differences between normal and cancerous cells exposed to electric fields.
Mucins 1‐shRNA inhibit the proliferation and HIF‐1alpha protein expression on human cholangiocarcinoma cells
Authors: Li Zhuo‐Ri , Wei Jia‐Li , Li Zhen‐Zhen , Li Shu‐Fang
Published: August 2013

To explore the effects of Mucins (MUC)1‐shRNA on the proliferation and hypoxia inducible factor (HIF)‐1alpha expression of human cholangiocarcinoma (CCA) QBC939 cells in vitro. MUC1‐shRNA was constructed and transfected with Lipofectamine™ 2000 into cultured CCA cells. MUC1 mRNA and protein expression levels were determined by RT‐PCR and Western blot, respectively. The cellular proliferation and HIF‐1alpha expression of QBC939 cells were evaluated by the MTT assay and Western blot, respectively. After transfection, the expression levels of MUC1 mRNA and protein in the experimental group decreased significantly in QBC939 (P < 0.01). The proliferation of MUC1 shRNA‐transfected group was 0.30 ± 0.05, 38.32 ± 1.43%, 15.18 ± 1.32%, and there were remarkable differences when compared with the control groups (P < 0.05). Significant inhibition of HIF‐1alpha protein expression in MUC1 shRNA‐transfected group was also discovered (P < 0.05). MUC1‐shRNA could inhibit proliferation and significantly weaken HIF‐1alpha protein expression of QBC939 cells, suggesting its potential as a therapeutic target of CCA.
Phenotypic and functional comparison of optimum culture conditions for upscaling of dental pulp stem cells
Authors: Kanafi Mohammad Mahboob , Pal Rajarshi , Gupta Pawan Kumar
Published: August 2013

Advances in dental pulp stem cell (DPSC) biology and behaviour have promised much in the field of regenerative medicine. Their recent use in clinical trials for bone repair enforces the notion that DPSCs can be used successfully in patients; however they display diverse characteristics under different culture conditions. Since the success of any stem cell culture is regulated by its own micro‐environment, it is imperative to optimise the growth conditions and establish a generic protocol for maintenance and scale‐up. This study focused on optimisation of long‐term culture conditions of human exfoliated deciduous teeth (SHED) in comparison with DPSCs, employing three commonly used basal media – knockout Dulbecco's modified Eagle's medium (KO‐DMEM), α‐MEM and DMEM/F12. Based on their characterisation with respect to morphology, growth kinetics, cell surface marker expression, differentiation capacity and plating density, our findings suggest that cells can be expanded with the highest efficiency in KO‐DMEM medium supplemented with 10% FBS. Additionally, under our standardised xeno‐free (10% human plasma) growth conditions, DPSCs displayed and retained their multipotent attributes until late passages. The differences in the growth and differentiation characteristics between SHED and DPSCs are shown, and certify SHED can be a key element in tissue engineering.
Possible role of the ‘IDO‐AhR axis’ in maternal‐foetal tolerance
Authors: Hao Kehong , Zhou Qian , Chen Wen , Jia Wenwen , Zheng Jing , Kang Jiuhong , Wang Kai , Duan Tony
Published: August 2013

The induction and maintenance of immunologic tolerance at the feto‐maternal interface is necessary for a successful pregnancy. The most accepted hypothesis for the mechanism underlying this tolerance is that pregnancy‐induced foetal antigen‐specific maternal T regulatory (Treg) cells mediate maternal tolerance to the foetus. The aryl hydrocarbon receptor (AhR), which is highly expressed in the placenta, is widely studied in female reproductive biology and immunology. Activation of AhR can promote immune tolerance by controlling the differentiation of Treg cells in some autoimmune disorders. However, the specific mechanisms underlying tolerance are poorly understood. Indoleamine 2,3‐dioxygenase (IDO) is the initial and rate‐limiting enzyme of tryptophan catabolism in human placental trophoblasts. IDO produces kynurenine, an endogenous AhR ligand that directly activates AhR and is proposed to be central to the establishment and maintenance of immunologic tolerance at the maternal‐foetal interface. We propose that kynurenine activates AhR, leading to the AhR‐dependent Treg cells generation, which in turn critically regulates immunological tolerance at the feto‐maternal interface. This hypothesis must be tested and the proof of this hypothesis may provide a potential therapeutic target for the treatment of infertility and other adverse pregnancy outcomes resulted from inadequate immunological tolerance at the feto‐maternal interface.
Mesenchymal stem cells of human placenta and umbilical cord suppress T‐cell proliferation at G0 phase of cell cycle
Authors: Vellasamy Shalini , Sandrasaigaran Pratheep , Vidyadaran Sharmili , Abdullah Maha , George Elizabeth , Ramasamy Rajesh
Published: August 2013

Mesenchymal stem cells (MSC) generated from human umbilical cord (UC‐MSC) and placenta (PLC‐MSC) were assessed and compared for their immunomodulatory function on T cells proliferation by analysis of the cell cycle. Mitogen stimulated or resting T cells were co‐cultured in the presence or absence of MSC. T‐cell proliferation was assessed by tritiated thymidine (3H‐TdR) assay and the mechanism of inhibition was examined bycell cycle and apoptosis assay. Both UC‐MSC and PLC‐MSC profoundly inhibited the proliferation of T‐cell, mainly via cell‐to‐cell contact. MSC‐mediated anti‐proliferation does not lead to apoptosis,but prevented T cells from entering S phase and they therefore accumulated in the G0/G1 phases. The anti‐proliferative activity of MSC was related to this cell cycle arrest of T‐cell. UC‐MSC produced a greater inhibition than PLC‐MSC in all measured parameters.
Preparation of Kupffer cell enriched non‐parenchymal liver cells with high yield and reduced damage of surface markers by a modified method for flow cytometry
Authors: Xu Fan , Zhen Peng , Zheng Yu , LIjuan Feng , Aiting Yang , Min Cong , Hong You , Jidong Jia
Published: August 2013

The aim of this study was to optimise a collagenase perfusion protocol for the isolation of a liver non‐parenchymal cell (NPC) suspension enriched for Kupffer cells that reduced damage to F4/80 antigen cell surface expression to allow analysis by flow cytometry.

Kupffer cell‐enriched liver NPCs were isolated from C57BL/6 mice using different protocols. Flow cytometry was used to examine the effect of collagenase digestion on F4/80 expression on Kupffer cells, and results were represented by the percentage of F4/80 positive cells and by the F4/80 mean fluorescence intensity (MFI). The perfusion temperature, concentration of collagenase solution and total dosage of collagenase for liver perfusion influenced the effect of collagenase perfusion on the expression of F4/80 antigen on Kupffer cells. Collagenase perfusion at 28°C resulted in an increased percentage of F4/80 positive cells (P = 0.001) and MFI (P = 0.005) compared with 37°C. Perfusion with a total dose of 1.0 g/kg BW collagenase (using a 0.75 mg/mL solution) resulted in the highest percentage of F4/80 positive cells (P = 0.001) compared with 0.8 g/kg BW and 1.2 g/kg BW collagenase. Isolation of cells using the modified protocol resulted in a higher percentage of Kupffer cells (P < 0.001) and a higher MFI of F4/80 antigen (P < 0.001) compared with the common protocol.
Long‐term muscle‐derived cell culture: multipotency and susceptibility to cell death stimuli
Authors: Kalvelyte Audrone , Krestnikova Natalija , Stulpinas Aurimas , Bukelskiene Virginija , Bironaite Daiva , Baltriukiene Daiva , Imbrasaite Ausra
Published: August 2013

Improvement in the yield of adult organism stem cells, and the ability to manage their differentiation and survival potential are the major goals in their application in regenerative medicine and in the adult stem cell research. We have demonstrated that adult rabbit muscle‐derived cell lines with an unlimited proliferative potential in vitro can differentiate into myogenic, osteogenic, adipogenic and neurogenic lineages. Studies of cell survival in vitro showed that differentiated cells, except neurogenic ones, are more resistant to apoptosis inducers compared to proliferating cells. Resistance to death signals correlated with the level of protein kinase AKT phosphorylation. Skeletal muscle‐derived cell lines can be multipurpose tools in therapy. Enhanced resistance of differentiated cells to certain types of damage shows their potential for long‐term survival and maintenance in an organism.

This article was published online on 29 January 2013. An error was subsequently identified. This notice is included in the online and print versions to indicate that both have been corrected 6 March 2013.
Engraftment of genetically modified human amniotic fluid‐derived progenitor cells to produce coagulation factor IX after in utero transplantation in mice
Authors: Yang Chen‐Min , Gong Xiu‐Li , Qiu Jin , Tang Hui‐Xiang , Gong Zhi‐Juan , Huang Shu‐Zhen , Zeng Fanyi
Published: August 2013

Human amniotic fluid derived progenitor cells (hAFPCs) may be multipotent and can be considered a potential tool in the field of cell therapy for haemophilia B. Their capacity to express human coagulation factor IX (hFIX) after transduction and their fate after in utero transplantation is unknown. hAFPCs isolated from second trimester pregnancies were assessed for their phenotypic markers, multilineage capacity, and expression of hFIX after transduction. Their engraftment potential was analysed in a mouse model after in utero transplantation at embryonic day 12.5. Immunohistochemistry, fluorescence in situ, ELISA and PCR were used to assess post‐transplant chimeras. hAFPCs expressed several pluripotent markers, including NANOG, SOX2, SSEA4 and TRA‐1‐60, and could differentiate into adipocytes and osteocytes. In vitro, after transduction with hFIX and EGFP cDNAs, constitutive hFIX protein expression and clotting activity were found. Engraftment was achieved in various foetal tissues after in utero transplantation. Safe engraftment without oncogenesis was confirmed, with low donor cell levels, but persistent engraftment, into different organs (liver, heart and lung) through to 12 weeks of age. Transgenic expression of circulating hFIX was detected in recipient mice for up to 12 weeks. hAFPCs can be engrafted long‐term in immunocompetent mice after in utero transplantation. Thus, cell transplantation approaches using genetically engineered hAFPCs may prove valuable for the prenatal treatment for haemophilia B.
c‐Myc suppressed E‐cadherin through miR‐9 at the post‐transcriptional level
Authors: Liu Mei , Zhu Hongxia , Yang Shangbin , Wang Zaozao , Bai Jinfeng , Xu Ningzhi
Published: August 2013

c‐Myc oncoprotein is overexpressed in most human cancers and regulates different genes and pathways in different cell types. E‐cadherin expression is repressed by MYC through a post‐transcriptional mechanism, but the exact mechanism remains elusive. Since E‐cadherin is a direct target of miR‐9 and miR‐9 can be activated by MYC and MYCN, this suggests that c‐Myc negatively modulates E‐cadherin through a microRNA pathway. We have established a c‐Myc‐inducible expression system in which the protein level and transcriptional activity of c‐Myc is significantly upregulated upon doxycycline induction. Overexpressed c‐Myc led to an EMT‐like conversion in the T‐REx‐293 cells and resulted in a significant decrease in E‐cadherin and an increase in Vimentin. Stem‐loop RT‐PCR showed elevated expression of miR‐9 when c‐Myc was induced to be overexpressed. Regarding the relationship of c‐Myc, miR‐9 and E‐cadherin, the expression of miR‐9 was curtailed by using antagomir‐9 in induced overexpressing c‐Myc. Restoration of E‐cadherin expression became much stronger in the presence of c‐Myc. Thus c‐Myc represses E‐cadherin at the post‐transcriptional level through miR‐9.
PPAR‐γ agonist pioglitazone prevents apoptosis of endothelial progenitor cells from rat bone marrow
Authors: Zhang Hui‐Feng , Wang Li , Yuan Hui‐Juan , Ma Yue‐Hua , Wang Yan‐Fang , Hu Zi‐Ying , Su Yong , Zhao Zhi‐Gang
Published: August 2013

Selective peroxisome proliferator‐activated receptor‐γ (PPAR‐γ) agonist affects the functions of endothelial progenitor cells (EPCs). This study explores the effect of selective PPAR‐γ agonist, pioglitazone, on EPC apoptosis. The cells were cultured and identified via the double staining method in a medium containing different concentrations of pioglitazone. EPC apoptosis was detected by flow cytometry. On Day 7, EPCs engulfed DiL‐ac‐LDL and FITC‐UEA‐1, and showed yellow fluorescence in a laser‐scanning confocal microscope. EPC apoptosis inhibition was maximal at 50 µmol/L. The ability of pioglitazone to prevent EPC apoptosis may be mediated by the PI3K/Akt signal pathway. The use of thiazolidine two ketone (TZD) to reduce EPC apoptosis may have some potential in treating vascular diseases.
Curcumin attenuates cyclooxygenase‐2 expression via inhibition of the NF‐κB pathway in lipopolysaccharide‐stimulated human gingival fibroblasts
Authors: Hu Ping , Huang Ping , Chen Min Wei
Published: August 2013

Porphyromonas gingivalis lipopolysaccharide (LPS) induces the expression of the cyclooxygenase‐2 (COX‐2), which contributes to the process of periodontitis. Curcumin, a constituent of turmeric, exhibits anti‐inflammatory properties. We have investigated the anti‐inflammatory effect of curcumin in human gingival fibroblasts (HGFs) stimulated by P. gingivalis LPS and its mechanism of action. HGFs pretreated with curcumin were stimulated by P. gingivalis LPS. COX‐2 mRNA and protein expressions were analysed by real‐time PCR and Western blot analysis. Activation of nuclear factor kappa B (NF‐κB) was analysed by the NF‐κB‐dependent luciferase activity and electrophoretic mobility‐shift assay (EMSA). Curcumin inhibited COX‐2 mRNA and protein synthesis in LPS‐stimulated HGFs in a dose‐dependent manner. P. gingivalis LPS activated NF‐κB‐dependent transcription in HGFs, which were also downregulated by pretreatment with curcumin. Therefore, curcumin can inhibit P. gingivalis LPS‐induced COX‐2 expression, which may be due to the inhibition of the NF‐κB pathway.
Functional compartmentalisation of NF‐κB‐associated proteins in A431 cells
Authors: Bolshakova Anastasia , Magnusson Karl‐Eric , Pinaev George , Petukhova Olga
Published: August 2013

NF‐κB proteins belong to a family of ubiquitous transcription factors involved in a number of cellular responses. While the pathways of NF‐κB activation and input into the regulation of gene activity have been comprehensively investigated, its cytoplasmic functions are poorly understood. In this study we addressed effects of the compartmentalisation of NF‐κB proteins RelA/p65 and p50 in relation to the inhibitor IκB‐α, using fibronectin (FN) and epidermal growth factor (EGF) for environmental stimulation of epidermoid carcinoma A431 cells. We thus assessed the presence of NF‐κB family proteins in the cytosol, membrane, nuclear and cytoskeletal fractions with a special attention to the cytoskeletal fraction to define whether NF‐κB was active or not. Sub‐cellular fractionation demonstrated that the proportion of RelA/p65 differed in diverse sub‐cellular fractions, and that the cytoskeleton harboured about 7% thereof. Neither the nuclear nor the cytoskeleton fraction did contain IκB‐α. The cytoskeleton binding of RelA/p65 and p50 was further confirmed by co‐localisation and electron microscopy data. During 30‐min EGF stimulation similar dynamics were found for RelA/p65 and IκB‐α in the cytosol, RelA/p65 and p50 in the nucleus and p50 and IκB‐α in the membrane. Furthermore, EGF stimulation for 30 min resulted in a threefold accumulation of RelA/p65 in cytoskeletal fraction. Our results suggest that nuclear‐, membrane‐ and cytoskeleton‐associated NF‐κB are dynamic and comprise active pools, whereas the cytoplasmic is more constant and likely non‐active due to the presence of IκB‐α. Moreover, we discovered the existence of a dynamic, IκB‐α‐free pool of RelA/p65 associated with cytoskeletal fraction, what argues for a special regulatory role of the cytoskeleton in NF‐κB stimulation.
Role of altholactone in inducing type II apoptosis signalling pathway and expression of cancer‐related genes in cervical carcinoma HeLa cell line
Authors: Uthaisang‐Tanechpongtamb Wanlaya , Sriyabhaya Promjit , Wilairat Prapon
Published: August 2013

Goniothalamus species (Annonaceae) is a shrub that grows in the rainforest of tropical Asia. Several compounds have been isolated and exhibit the potential use for cancer treatment. In this work, altholactone isolated from Goniothalamus macrophyllus was investigated for its cytotoxicity, apoptosis signalling and the expression of cancer‐related genes in the cervical carcinoma HeLa cells. Cytotoxicity was evaluated by MTT assay. Apoptotic characteristics were evaluated by morphological studies. Caspase‐3 activity was detected using a fluorogenic substrate. Cytochrome c release from mitochondria and protein Bid were determined by Western blotting and cancer‐related genes expression by RT‐PCR. The results demonstrated that altholactone was cytotoxic to HeLa (IC50 = 9.6 μg/mL), and apoptotic cell death was manifested by appearance of chromatin condensation and caspase‐3 activation, which was inhibited by specific inhibitors of both caspase‐8 and ‐9. Release into the cytosol of cytochrome and cleavage of Bid occurred. Altholactone also caused a decrease in bcl‐2 and an increase in p53 expression. These unique properties of altholactone suggest a potential for cancer chemotherapy.
Calpains are required for invasive and metastatic potentials of human HCC cells
Authors: Chen Bo , Tang Juan , Guo Yun‐Shan , Li Yong , Chen Zhi‐Nan , Jiang Jian‐Li
Published: August 2013

Calpains are a conserved family of calcium‐dependent cysteine proteinases involved in various cellular functions. Two ubiquitous isoforms, µ‐ and m‐calpain, are key members of the calpain family that play essential roles in regulating cell migration and invasion. However, it remains unclear whether they are involved in the progression of hepatocellular carcinoma (HCC). Here, we investigated the functions of µ‐ and m‐calpain in the invasive and metastatic processes of human hepatoma cells. Our results indicated that the expression levels of calpains were elevated in HCC cells compared with those in normal hepatic cells. Our results indicated that small interfering RNA (siRNA)‐mediated silencing of µ‐ and m‐calpain expressions significantly suppressed the adhesive, migrative and invasive potentials of human hepatoma cells. The matrix metalloproteinases (MMPs) are key regulators of malignant tumour invasion and metastasis. siRNA‐mediated down‐regulation of µ‐ and m‐calpain expressions also significantly attenuated MMP‐2 and MMP‐9 secretion. Thus µ‐ and m‐calpain may play important roles in the invasion and metastasis of human hepatoma cells, and calpains may be drug targets for preventing HCC metastasis.
Identification of radiation‐sensitive expressed genes in the ICR and AKR/J mouse thymus
Authors: Bong Jin Jong , Kang Yu Mi , Shin Suk Chul , Choi Seung Jin , Lee Kyung Mi , Kim Hee Sun
Published: August 2013

We have investigated radiation‐sensitive expressed genes (EGs), their signal pathways, and the effects of ionizing radiation in the thymus of ICR and AKR/J mice. Whole‐body and relative thymus weights were taken and microarray analyses were done on the thymuses of high‐dose‐rate (HDR, 137Cs, 0.8 Gy/min, a single dose of 4.5 Gy) and low‐dose‐rate (LDR, 137Cs, 0.7 mGy/h, a cumulative dose of 1.7 Gy) irradiated ICR and AKR/J mice. Gene expression patterns were validated by quantitative polymerase chain reaction (qPCR). The effect of ionizing radiation on thymus cell apoptosis was measured terminal deoxynucleotidyl‐transferase‐mediated dUTP‐end labeling (TUNEL). LDR‐irradiation increased the mean whole‐body weight, but decreased the relative thymus weight of AKR/J mice. Radiation‐sensitive EGs were found by comparing HDR‐ and LDR‐irradiated ICR and AKR/J mice. qPCR analysis showed that 12 EGs had dose and dose‐rate dependent expression patterns. Gene‐network analysis indicated that Ighg, Igh‐VJ558, Defb6, Reg3g, and Saa2 may be involved in the immune response, leukocyte migration, and apoptosis. Our data suggest that expression of the HDR (Glut1, Glut4, and PKLR) and LDR radiation‐response genes (Ighg and Igh‐VJ558) can be dose or dose‐rate dependent. There was an increased number of apoptotic cells in HDR‐irradiated ICR mice and LDR‐irradiated AKR/J mice. Thus, changes of the mean whole‐body weight and relative thymus weight, EGs, signal pathways, and the effects of ionizing radiation on the thymus of ICR and AKR/J mice are described.
Effect of monochromatic visible light on intracellular superoxide anion production and mitochondrial membrane potential of B16F1 and B16F10 murine melanoma cells
Authors: Sato Kazuomi , Minai Yuji , Watanabe Hiroyuki
Published: August 2013

We have investigated the effect of visible light on animal cells using light‐emitting diodes to emit monochromatic visible light (red, yellow, green and blue light). To explore the relevant mechanism of apoptosis, we assessed the intracellular superoxide anion production and mitochondrial membrane potential (ΔΨm) of B16F1 and B16F10 murine melanoma cells after monochromatic light irradiation. Blue light caused ΔΨm depolarization subsequent to elevation of intracellular superoxide production. However, red and yellow light had no affect on both cell lines. Green light induced ΔΨm collapse only in B16F1 melanoma cells. ΔΨm is a key indicator of mitochondrial function, therefore its disruption causes mitochondria‐dependent apoptosis. Thus, blue light causes mitochondrial dysfunction and subsequent cell death.
Nitric oxide protects endothelium from cadmium mediated leakiness
Authors: Nagarajan Shunmugam , Rajendran Saranya , Saran Uttara , Priya M. Krishna , Swaminathan Akila , Siamwala Jamila H. , Sinha Swaraj , Veeriah Vimal , Sonar Punam , Jadhav Vivek , Jaffar Ali B.M. , Chatterjee Suvro
Published: August 2013

Cadmium targets the vascular endothelium causing endothelial dysfunction and leakiness of endothelial barrier. Nitric oxide plays a major role in mediating endothelial functions including angiogenesis, migration and permeability. The present study investigates the nitric oxide effects on cadmium induced endothelial leakiness. Results of ex vivo and in vitro permeability assays showed that even a sub‐lethal dose of cadmium chloride (1 µM) was sufficient to induce leakiness of endothelial cells. Cadmium drastically altered the actin polymerisation pattern and membrane tension of these cells compared to controls. Addition of nitric oxide donor Spermine NONOate (SP) significantly blunted cadmium‐mediated effects and recover endothelial cells integrity. Cadmium‐induced cytoskeletal rearrangements and membrane leakiness are associated with the low nitric oxide availability and high reactive oxygen species generation. In brief, we show the protective role of nitric oxide against cadmium‐mediated endothelial leakiness.