bluuebunny/crossref_metadata_2025 huggingface dataset:
Protein refolding using chemical refolding additives
Authors: Yamaguchi Satoshi , Yamamoto Etsushi , Mannen Teruhisa , Nagamune Teruyuki , Nagamune Teruyuki
Published: September 2012

In laboratories and manufacturing settings, a rapid and inexpensive method for the preparation of a target protein is crucial for promoting resesrach in protein science and engineering. Inclusion‐body‐based protein production is a promising method because high yields are achieved in the upstream process, although the refolding of solubilized, unfolded proteins in downstream processes often leads to significantly lower yields. The most challenging problem is that the effective condition for refolding is protein dependent and is therefore difficult to select in a rational manner. Accordingly, considerable time and expense using trial‐and‐error approaches are often needed to increase the final protein yield. Furthermore, for certain target proteins, finding suitable conditions to achieve an adequate yield cannot be obtained by existing methods. Therefore, to convert such a troublesome refolding process into a routine one, a wide array of methods based on novel technologies and materials have been developed. These methods select refolding conditions where productive refolding dominates over unproductive aggregation in competitive refolding reactions. This review focuses on synthetic refolding additives and describes the concepts underlying the development of reported chemical additives or chemical‐additive‐b
Monitoring and control of microbioreactors: An expert opinion on development needs
Authors: Gernaey Krist V. , Baganz Frank , Franco‐Lara Ezequiel , Kensy Frank , Krühne Ulrich , Luebberstedt Marc , Marx Uwe , Palmqvist Eva , Schmid Andreas , Schubert Frank , Mandenius Carl‐Fredrik
Published: September 2012

This perspective article is based on an expert panel review on microbioreactor applications in biochemical and biomedical engineering that was organized by the M3C (measurement, monitoring, modelling and control) Working Group of the European Section of Biochemical Engineering Science (ESBES) in the European Federation of Biotechnology (EFB). The aim of the panel was to provide an updated view on the present status of the subject and to identify critical needs and issues for furthering the successful development of microbioreactor monitoring and control. This will benefit future bioprocess development and in vitro toxicity testing. The article concludes with a set of recommendations for extended use and further development of microbioreactors.
Approach to Describe the Flow Conditions Relevant for Droplet Break Up During Emulsification of W/O Emulsions with Orifices
Authors: Wolf Frederik , Schuch Anna , Köhler Karsten , Schuchmann Heike P.
Published: September 2012

Eine indirekte Möglichkeit zur Einordnung der Strömung beim Emulgieren mit Lochblenden wird vorgestellt und der Einfluss der Strömungsform auf die Tropfengrößenverteilung der resultierenden W/O‐Emulsionen untersucht. Anhand verschiedener Bereiche im Verlauf des Durchflusskoeffizienten konnten unterschiedliche Strömungsformen identifiziert werden. In verschiedenen Strömungsbereichen wurden W/O‐Emulsionen zweier Stoffsysteme hergestellt und die Tropfengrößenverteilung der Emulsionen gemessen. Es konnte beobachtet werden, dass die resultierenden Tropfengrößenverteilungen der Emulsionen stark von der vorliegenden Strömungsform abhängen.
Resource Efficient Forming Technologies for Titanium and High Temperature Resistant Alloys
Authors: Erxleben Stefan , Fellmann Holger , Güther Volker , Janschek Peter , Lorenz Bernd
Published: September 2012

Halbzeuge von Nichteisenwerkstoffen wie Titan‐, Nickel‐ oder Kobaltlegierungen sind einerseits nur in begrenzten Abmaß‐Sortimenten verfügbar sowie anderseits schwer verarbeitbar. Das führt in der Regel zu einer erheblichen Ineffizienz verbunden mit hohen Materialverlusten insbesondere in der Klein‐ und Mittelserienfertigung. Die vorgestellten Arbeiten zeigen erste Ergebnisse der Entwicklung inkrementeller Umformverfahren, um neue Möglichkeiten der ressourcen‐, energie‐ und kosteneffizienten Verarbeitung dieser Werkstoffe zu eröffnen. Diese partiell wirkenden Verfahren zeichnen sich durch vorteilhafte Besonderheiten aus.
An Integrated Process for Innovative Extraction of Metals from Kupferschiefer Mine Dumps, Germany
Authors: Kamradt Andreas , Borg Gregor , Schaefer Juliane , Kruse Stephanie , Fiedler Marco , Romm Phillip , Schippers Axel , Gorny Ralf , Bois Michael du , Bieligk Christin , Liebetrau Nadine , Nell Sylvie , Friedrich Bernd , Morgenroth Henning , Wotruba Hermann , Merkel Charlotte
Published: September 2012

Dumps in the Central German Mansfeld mining district have been investigated by a multi‐disciplinary research team to develop a holistic concept for the recovery of base and trace metals from low‐grade Kupferschiefer ore. Geochemical and mineralogical data from representative sampling indicate that the low‐grade Kupferschiefer dumps contain significant amounts of Cu (0.6 %) and Ag (up to 800 ppm), besides exploitable grades of Pb and Zn. Additionally to flotation and hydrometallurgy, first shake flask bioleaching experiments on otherwise untreated Kupferschiefer ore yield Cu recovery rates of up to 95 %. A total of 900 t Cu and 5 t Ag contained in black copper can thus be produced from the pilot dump of 100 000 m3 low‐grade Kupferschiefer ore. Additionally, large volumes of limestone waste, separated from the dumps by sensor‐based sorting, can be utilized as aggregate in the construction industry.
Drosophila LSD1‐CoREST demethylase complex regulates DPP/TGFβ signaling during wing development
Authors: Curtis Brenda J. , Zraly Claudia B. , Dingwall Andrew K.
Published: September 2012

The choice and timing of specific developmental pathways in organogenesis are determined by tissue‐specific temporal and spatial cues that are acted upon to impart unique cellular and compartmental identities. A consequence of cellular signaling is the rapid transcriptional reprogramming of a wide variety of target genes. To overcome intrinsic epigenetic chromatin barriers to transcription modulation, histone modifying and remodeling complexes are employed. The deposition or erasure of specific covalent histone modifications, including acetylation, methylation, and ubiquitination are essential features of gene activation and repression. We have found that the activity of a specific class of histone demethylation enzymes is required for the specification of vein cell fates during Drosophila wing development. Genetic tests revealed that the Drosophila LSD1‐CoREST complex is required for proper cell specification through regulation of the DPP/TGFβ pathway. An important finding from this analysis is that LSD1‐CoREST functions through control of rhomboid expression in an EGFR‐independent pathway. genesis
51:16–31, 2013. © 2012 Wiley Periodicals, Inc.
The origins of Atlantic salmon (Salmo salar L.) recolonizing the River Mersey in northwest England
Authors: Ikediashi Charles , Billington Sam , Stevens Jamie R.
Published: September 2012

By the 1950s, pollution had extirpated Atlantic salmon in the river Mersey in northwest England. During the 1970s, an extensive restoration program began and in 2001, an adult salmon was caught ascending the river. Subsequently, a fish trap was installed and additional adults are now routinely sampled. In this study, we have genotyped 138 adults and one juvenile salmon at 14 microsatellite loci from across this time period (2001–2011). We have used assignment analysis with a recently compiled pan‐European microsatellite baseline to identify their most probable region of origin. Fish entering the Mersey appear to originate from multiple sources, with the greatest proportion (45–60%, dependent on methodology) assigning to rivers in the geographical region just north of the Mersey, which includes Northwest England and the Solway Firth. Substantial numbers also appear to originate from rivers in western Scotland, and from rivers in Wales and Southwest England; nonetheless, the number of fish originating from proximal rivers to the west of the Mersey was lower than expected. Our results suggest that the majority of salmon sampled in the Mersey are straying in a southerly direction, in accordance with the predominantly clockwise gyre present in the eastern Irish Sea. Our findings highlight the complementary roles of improving water quality and in‐river navigability in restoring salmon to a river and underlines further the potential benefits of restoration over stocking as a long‐term solution to declining fish stocks.
Production of interleukin‐27 by human neutrophils regulates their function during bacterial infection
Authors: Rinchai Darawan , Khaenam Prasong , Kewcharoenwong Chidchamai , Buddhisa Surachat , Pankla Rungnapa , Chaussabel Damien , Bancroft Gregory J. , Lertmemongkolchai Ganjana
Published: September 2012

Septicemia is the most severe form of melioidosis caused by the Gram‐negative bacterium, Burkholderia pseudomallei. Here, we show that levels of IL‐27p28 transcript and protein were both significantly elevated in patients with sepsis, particularly melioidosis and in patients with unfavorable disease outcome. Moreover, human monocytes/macrophages and neutrophils were the major source of IL‐27 during infection. The addition of exogenous IL‐27 in vitro resulted in significantly increased bacterial survival, reduced B. pseudomallei‐induced oxidative burst, and enhanced IL‐1β and TNF‐α production by purified neutrophils from healthy subjects. Finally, blockade of endogenous IL‐27 in neutrophils using soluble IL‐27 receptor antagonist prior to infection led to significantly reduced survival of bacteria and decreased IL‐1β, but not TNF‐α production. These results indicate a potential role for IL‐27 in the suppression of anti‐bacterial defense mechanisms that might contribute to disease severity in sepsis. The targeting of this cytokine may be beneficial in the management of human sepsis.
Avidity of influenza‐specific memory CD8+T‐cell populations decays over time compromising antiviral immunity
Authors: Humphreys Ian R. , Clement Mathew , Marsden Morgan , Ladell Kristin , McLaren James E. , Smart Kathryn , Hindley James P. , Bridgeman Hayley M. , van den Berg Hugo A. , Price David A. , Ager Ann , Wooldridge Linda , Godkin Andrew , Gallimore Awen M.
Published: September 2012

Decline of cell‐mediated immunity is often attributed to decaying T‐cell numbers and their distribution in peripheral organs. This study examined the hypothesis that qualitative as well as quantitative changes contribute to the declining efficacy of CD8+ T‐cell memory. Using a model of influenza virus infection, where loss of protective CD8+ T‐cell immunity was observed 6 months postinfection, we found no decline in antigen‐specific T‐cell numbers or migration to the site of secondary infection. There was, however, a large reduction in antigen‐specific CD8+ T‐cell degranulation, cytokine secretion, and polyfunctionality. A profound loss of high‐avidity T cells over time indicated that failure to confer protective immunity resulted from the inferior functional capacity of remaining low avidity cells. These data imply that high‐avidity central memory T cells wane with declining antigen levels, leaving lower avidity T cells with reduced functional capabilities.
CD73 is a phenotypic marker of effector memory Th17 cells in inflammatory bowel disease
Authors: Doherty Glen A. , Bai Aiping , Hanidziar Dusan , Longhi Maria S. , Lawlor Garrett O. , Putheti Prabhakar , Csizmadia Eva , Nowak Martina , Cheifetz Adam S. , Moss Alan C. , Robson Simon C.
Published: September 2012

Purinergic signaling and associated ectonucleotidases, such as CD39 and CD73, have been implicated in the pathogenesis of inflammatory bowel disease (IBD). CD39 is known to be a Treg memory cell marker, and here we determine the phenotype and function of CD73+CD4+ T lymphocytes in patients with IBD. We describe elevated levels of CD73+CD4+ T cells in the peripheral blood and intestinal lamina propria of patients with active IBD. The functional phenotype of these CD73+CD4+ T cells was further determined by gene expression, ecto‐enzymatic activity, and suppressive assays. Increased numbers of CD73+CD4+ T cells in the periphery and lamina propria were noted during active inflammation, which returned to baseline levels following anti‐TNF treatment. Peripheral CD73+CD4+ T cells predominantly expressed CD45RO, and were enriched with IL‐17A+ cells. The CD73+CD4+ cell population expressed higher levels of RORC, IL‐17A, and TNF, and lower levels of FOXP3 and/or CD25, than CD73−CD4+ T cells. Expression of CD73 by peripheral CD4+ T cells was increased by TNF, and decreased by an anti‐TNF monoclonal antibody (infliximab). In vitro, these peripheral CD73+CD4+ T cells did not suppress proliferation of CD25− effector cells, and expressed higher levels of pro‐inflammatory markers. We conclude that the CD73+CD4+ T‐cell population in patients with active IBD are enriched with cells with a T‐helper type 17 phenotype, and could be used to monitor disease activity during treatment.
Influence of congenital human cytomegalovirus infection and the NKG2C genotype on NK‐cell subset distribution in children
Authors: Noyola Daniel E. , Fortuny Claudia , Muntasell Aura , Noguera‐Julian Antoni , Muñoz‐Almagro Carmen , Alarcón Ana , Juncosa Teresa , Moraru Manuela , Vilches Carlos , López‐Botet Miguel
Published: September 2012

Human cytomegalovirus (HCMV) has been reported to reshape the NK‐cell receptor (NKR) distribution, promoting an expansion of CD94/NKG2C+ NK and T cells. The role of NK cells in congenital HCMV infection is ill‐defined. Here we studied the expression of NKR (i.e., NKG2C, NKG2A, LILRB1, CD161) and the frequency of the NKG2C gene deletion in children with past congenital infection, both symptomatic (n = 15) and asymptomatic (n = 11), including as controls children with postnatal infection (n = 11) and noninfected (n = 20). The expansion of NKG2C+ NK cells in HCMV‐infected individuals appeared particularly marked and was associated with an increased number of LILRB1+ NK cells in cases with symptomatic congenital infection. Increased numbers of NKG2C+, NKG2A+, and CD161+ T cells were also associated to HCMV infection. The NKG2C deletion frequency was comparable in children with congenital HCMV infection and controls. Remarkably, the homozygous NKG2C+/+ genotype appeared associated with increased absolute numbers of NKG2C+ NK cells. Moreover, HCMV‐infected NKG2C+/+ children displayed higher absolute numbers of NKG2A+ and total NK cells than NKG2C+/− individuals. Our study provides novel insights on the impact of HCMV infection on the homeostasis of the NK‐cell compartment in children, revealing a modulatory influence of NKG2C copy number.
The Simple Synthesis of Ferrocene Ligands from a Practitioner's Perspective
Authors: Butler Ian R.
Published: September 2012

This review is a personal account of the research carried out in our laboratories on the design and synthesis of ferrocene compounds used in catalysis, primarily as ligands but also in material science applications. The emphasis of the research has always been to find the simplest possible synthetic routes towards these ferrocene compounds, which may be used as convenient starting materials. This is why lithiation and quench methodology has been used extensively and successfully in most applications. It has taken many years to achieve the convenient synthesis of some very simple compounds such as 1,2‐dibromoferrocene and 1,2,3,4,5‐pentabromoferrocene, and the reasons for this are highlighted. The review concentrates on the synthesis of ferrocenophanes, ferrocenylphosphanes, ferrocenylalkylamines and their precursor compounds, describing some of their early coordination chemistry. Subsequently, the successful use of ferrocene‐based ligands in the Lucite Alpha process is discussed. The coordination chemistry of a selection of the ligands prepared is discussed, and the rationale behind the specific ligand design strategies is given. Included is some research material, previously unpublished, which serves to fill in gaps in the overview. Finally, some derivatives prepared for applications in material science are described from a synthetic standpoint.
3‐(2‐Furyl)‐2‐sulfanylpropenoic Acid as a Chelating Agent for Lead(II) and Diorganonolead(IV) Compounds – Chemical Behaviour and In Vitro Protective Effect against Dimethyllead(IV) Toxicity
Authors: Casas José S. , Castaño M. Victoria , Couce Maria D. , Sánchez Agustín , Sordo José , Torres M. D. , Vázquez López Ezequiel M.
Published: September 2012

The reaction of the title acid (H2fspa) with lead(II) acetate and dimethyl‐ or diphenyllead(IV) acetate in alcoholic medium gives the 1:1 complexes [Pb(fspa)] and [PbR2(fspa)] (R = Me, Ph). When the reaction was carried out in the presence of diisopropylamine (Q), the [HQ]2[Pb(fspa)2] and [HQ]2[PbPh2(fspa)2] derivatives formed in low yield. Additionally, the compounds Na[PbMe2(OAc)3], [PbPh2(fspa)(DMSO)] and [PbPh2(fspa)(DMSO)]·DMSO were also isolated as crystalline solids in subsequent crystallization processes. All these compounds were characterized by IR and NMR (1H and 13C) spectroscopy and, in some cases, by X‐ray diffractometry and 207Pb NMR spectroscopy. Pig renal proximal tubule LLC‐PK1 culture cells were used to determine in vitro the nephroprotective effect of H2fspa, [HQ]2[Zn(fspa)2], vitamin B1 and vitamin B6 against PbMe2(NO3)2 toxicity. The beneficial effects of the acid, the zinc(II) complex and/or their combinations with vitamin B1 (thiamine nitrate) are described.
Arene–Ruthenium Complexes with Phosphanylferrocenecarboxamides Bearing Polar Hydroxyalkyl Groups – Synthesis, Molecular Structure, and Catalytic Use in Redox Isomerizations of Allylic Alcohols to Carbonyl Compounds
Authors: Schulz Jiří , Císařová Ivana , Štěpnička Petr
Published: September 2012

Phosphanylferrocenecarboxamide Ph2P–fc–CONHCH2CH2OH (1, fc = ferrocene‐1,1′‐diyl) and its newly synthesized congeners, Ph2P–fc–CONHCH(CH2OH)2 (2) and Ph2P–fc–CONHC(CH2OH)3 (3), were converted to a series of (η6‐arene)ruthenium complexes [(η6‐arene)RuCl2(L‐κP)] 5–7, where arene is benzene, p‐cymene, and hexamethylbenzene and L = 1–3. All compounds were characterized by multinuclear NMR and IR spectroscopy, by mass spectrometry, and by elemental analysis. The molecular structures of 2, 3, 3O (a phosphane oxide resulting from the oxidation of 3), 5c·CH2Cl2, and 6c·Et2O were determined by single‐crystal X‐ray diffraction analysis. The ruthenium complexes were further evaluated as catalysts in the redox isomerization of allyl alcohols to carbonyl compounds. Complex [(η6‐p‐cymene)RuCl2(1‐κP)] (5b) proved to be a particularly attractive catalyst, being both readily available and catalytically active. Substrates with unsubstituted double bonds were cleanly isomerized with this catalyst in 1,2‐dichloroethane (0.5 mol‐% Ru, 80 °C), whereas for those bearing substituents at the double bond (particularly in the position closer to the OH group) lower conversions and selectivities were achieved. A similar trend was noted when pure water was used as the solvent, except that the best results (complete conversion with 2 mol‐% Ru) were seen for 1,3‐diphenylallyl alcohol, the most hydrophobic substrate.
Click Synthesis and Redox Chemistry of Mono‐ and Heterobimetallic Triazolyl and Triazolium‐Ferrocene and Cobalticinium Complexes
Authors: Rapakousiou Amalia , Mouche Claire , Duttine Mathieu , Ruiz Jaime , Astruc Didier
Published: September 2012

Mono‐ and heterobimetallic triazolylferrocene and cobalticinium complexes and triazolium derivatives were synthesized by click reactions between the ethynylmetallocenes and benzyl azide or (azidomethyl)ferrocene followed by methylation reactions, respectively. Cyclic voltammetry data shed light on the electron‐withdrawing character of the 1,2,3‐triazolyl and triazolium substituents of the metallocenes, and cathodic reduction of the triazolium group was found to be irreversible even at –50 °C. Chemical reduction of orange triazolylcobalticinium hexafluorophosphate using NaBH4 yielded a mixture of red isomeric triazolyl η4‐cyclopentadiene–cobalt–η5‐cyclopentadienyl complexes, whereas reduction by the single‐electron reductant [FeICp(η6‐C6Me6)] yielded the brown 19‐electron triazolylcobaltocene.
A Stereoselective Route to Aza‐C‐aryl Glycosides from Arynes and Chiral Nitrones
Authors: Khangarot Rama Kanwar , Kaliappan Krishna P.
Published: September 2012

Arynes are regarded as potential and versatile intermediates in organic synthesis. These highly‐reactive, strained and kinetically unstable species have numerous applications in synthetic chemistry. In this article, a highly‐diasteroselective and efficient 1,3‐dipolar cycloaddition reaction between a range of arynes and sugar‐derived cyclic nitrones leading to an interesting class of sugar‐based benzo[d]isoxazolines is described. These benzo[d]isoxazolines upon selective N–O bond cleavage provide various substituted aza‐C‐aryl glycosides in good yield. These substituted pyrrolidine derivatives are chiral aminophenols and could be potential chiral ligands and organocatalysts in asymmetric synthesis.
Colorimetric Probe for the Detection of Thiols: The Dihydroazulene/Vinylheptafulvene System
Authors: Cacciarini Martina , Della Pia Eduardo A. , Nielsen Mogens B.
Published: September 2012

A new application of the dihydroazulene/vinylheptafulvene (DHA/VHF) system as a chemodosimeter for thiols is reported herein. A color change visible to the naked eye can be detected in acetonitrile/water (4:1) in the presence of cysteine and alkanethiols but not with other amino acids. The higher reactivity of DHA towards cysteine compared with VHF has been demonstrated. The easy synthesis of the main core and its versatility in terms of structural and physicochemical changes are essential elements for developing a new family of thiol‐specific probes with higher selectivity and sensitivity than the reported prototype.
Phosphane‐Mediated Domino Synthesis of Tetrasubstituted Furans from Simple Terminal Activated Olefins
Authors: Wang Jianfang , Zhou Rong , He Zheng‐Rong , He Zhengjie
Published: September 2012

Convenient and highly efficient syntheses of tetrasubstituted furans with flexible substituent patterns from simple and readily available starting materials have been developed. Under very mild conditions, with the mediation of stoichiometric nBu3P, simple terminal activated olefins and acyl chlorides or anhydrides smoothly furnish tetrasubstituted furans in modest to excellent yields. This synthetic strategy features a flexible selection of substituent pattern and a C‐acylation/O‐acylation/C‐acylation/intramolecular Wittig reaction multiple domino assembly sequence.
Palladium‐Catalyzed Mono‐α‐arylation of Carbonyl‐Containing Compounds with Aryl Halides using DalPhos Ligands
Authors: Crawford Sarah M. , Alsabeh Pamela G. , Stradiotto Mark
Published: September 2012

We report the extension and optimization of the [Pd(cinnamyl)Cl]2/DalPhos catalyst system, previously found effective for the mono‐α‐arylation of acetone, to the mono‐α‐arylation of a variety of carbonyl‐containing compounds with aryl halides and heteroaryl halides. Aryl methyl ketones, heteroaryl methyl ketones, propiophenones, malonates, and methoxyacetone can be α‐arylated under relatively mild conditions and in good yields. We also report the limitations of the ligand/catalyst system towards other classes of carbonyl‐containing compounds.
Searching for the Hidden Hydrides: The Competition between Alkene Isomerization and Metathesis with Grubbs Catalysts
Authors: Ashworth Ian W. , Hillier Ian H. , Nelson David J. , Percy Jonathan M. , Vincent Mark A.
Published: September 2012

Although Grubbs metathesis catalysts have enabled syntheses of a range of molecules, alkene isomerization, a known and problematic side reaction, is poorly understood. Several mechanisms which have been advanced to account for isomerization were studied by using electronic structure calculations. The pathway catalyzed by a ruthenium hydride emerged as the most facile process by a significant margin. For the first time, we have obtained experimental evidence for the presence of this species in a metathesis‐active system.
DNA polymerase kappa microsatellite synthesis: Two distinct mechanisms of slippage‐mediated errors
Authors: Baptiste Beverly A. , Eckert Kristin A.
Published: September 2012

Microsatellite tandem repeats are frequent sites of strand slippage mutagenesis in the human genome. Microsatellite mutations often occur as insertion/deletion of a repeat motif (unit‐based indels), and increase in frequency with increasing repeat length after a threshold is reached. We recently demonstrated that DNA polymerase κ (Pol κ) produces fewer unit‐based indel errors within dinucleotide microsatellites than does polymerase δ. Here, we examined human Pol κ's error profile within microsatellite alleles of varying sequence composition and length, using an in vitro HSV‐tk gap‐filling assay. We observed that Pol κ displays relatively accurate synthesis for unit‐based indels, using di‐ and tetranucleotide repeat templates longer than the threshold length. We observed an abrupt increase in the unit‐based indel frequency when the total microsatellite length exceeds 28 nucleotides, suggesting that extended Pol κ protein–DNA interactions enhance fidelity of the enzyme when synthesizing these microsatellite alleles. In contrast, Pol κ is error‐prone within the HSV‐tk coding sequence, producing frequent single‐base errors in a manner that is highly biased with regard to sequence context. Single‐nucleotide errors are also created by Pol κ within di‐ and tetranucleotide repeats, independently of the microsatellite allele length and at a frequency per nucleotide similar to the frequency of single base errors within the coding sequence. These single‐base errors represent the mutational signature of Pol κ, and we propose them a mechanism independent of homology‐stabilized slippage. Pol κ's dual fidelity nature provides a unique research tool to explore the distinct mechanisms of slippage‐mediated mutagenesis.Environ. Mol. Mutagen., 2012. © 2012 Wiley Periodicals, Inc.
DNA polymerase zeta generates clustered mutations during bypass of endogenous DNA lesions inSaccharomyces cerevisiae
Authors: Stone Jana E. , Lujan Scott A. , Kunkel Thomas A.
Published: September 2012

Multiple sequence changes that are simultaneously introduced in a single DNA transaction have a higher probability of altering gene function than do single base substitutions. DNA polymerase zeta (Pol ζ) has been shown to introduce such clustered mutations under specific selective and/or DNA damage‐producing conditions. In this study, a forward mutation assay was used to determine the specificity of spontaneous mutations generated inSaccharomyces cerevisiaewhen either wild‐type Pol ζ or a mutator Pol ζ variant (rev3‐L979F) bypasses endogenous lesions. Mutagenesis in strains proficient for nucleotide excision repair (NER) was compared to mutagenesis in NER‐deficient strains that retain unrepaired endogenous DNA lesions in the genome. Compared to NER‐proficient strains, NER‐deficientrad14Δstrains have elevated mutation rates that depend on Pol ζ. Rates are most strongly elevated for tandem base pair substitutions and clusters of multiple, closely spaced mutations. Both types of mutations depend on Pol ζ, but not on Pol η. Rates of each are further elevated in yeast strains bearing therev3‐979Fallele. The results indicate that when Pol ζ performs mutagenic bypass of endogenous, helix‐distorting lesions, it catalyzes a short track of processive, error‐prone synthesis. We discuss the implications of this unique catalytic property of Pol ζ to its evolutionary conservation and possibly to multistage carcinogenesis.Environ. Mol. Mutagen., 2012. © 2012 Wiley Periodicals, Inc.
PtCo/Polypyrrole‐Multiwalled Carbon Nanotube Complex Cathode Catalyst Containing Two Types of Oxygen Reduction Active Sites Used in Direct Methanol Fuel Cells
Authors: Zhao H. , Pan L. , Jin J. , Li L. , Xu J.
Published: September 2012

Low oxygen reduction reaction (ORR) activity and high cost of noble metal catalysts are two major challenges in direct methanol fuel cells (DMFCs). Pt‐based catalysts are considered as an ideal alternative to deal with these two problems. While the second component metals play only the role of synergy effect with Pt, they themselves are inert towards activity towards ORR. It is necessary to design a new route to ultilize the second component metal by forming CoNx ORR active site on the base of PtM catalyst. In this paper, PtCo/polypyrrole‐multiwalled carbon nanotubes (PtCo/PPy‐MWCNTs) catalyst containing two types of ORR active site (Pt and CoNx) was synthesized by one pot synthesis route. The effect and dynamic mechanism of the named CoNx active site towards ORR was discussed by X‐ray photoelectron sprectroscopy and linear sweep voltammetry. PtCo/PPy‐MWCNTs cathode catalyst showed improved activity towards ORR and great potential in DMFCs.
Microstructural Analysis of Interfaces between Lanthanum Contained Glass and Two Different Electrolytes for SOFC Applications
Authors: Kaur G. , Pandey O. P. , Singh K.
Published: September 2012

The diffusion couples of lanthanum based strontium borosilicate glass with two different (high and low temperature) electrolytes have been heat‐treated at 850 and 800 °C, respectively, for 5, 100, and 750 h to understand the mechanism of interface formation and growth. These prepared diffusion couples have been characterized using various techniques like X‐ray diffraction (XRD), scanning electron microscope (SEM), X‐ray dot mapping, and electron probe microanalysis (EPMA). XRD revealed no detrimental phase formation. Other thermodynamic parameters like frequency factor, crystallization constants, free volume, and bulk thermal expansion coefficients have been calculated to understand the behavior of glass under the influence of temperature. These theoretical parameters will enable to understand the interfacial growth observed in heat‐treated couple for its end application in SOFC. The observed results indicate that lanthanum containing glass exhibit stable and smooth interface with yttria‐stabilized zirconia (high‐temperature electrolyte) as compared to doped bismuth vanadate (low temperature electrolyte).
t(11;14)(q23;q32) involving IGH and DDX6 in nodal marginal zone lymphoma
Authors: Stary Susanne , Vinatzer Ursula , Müllauer Leonhard , Raderer Markus , Birner Peter , Streubel Berthold
Published: September 2012

Nodal marginal zone lymphoma (NMZL) is a primary nodal B‐cell lymphoma that shares morphological and immunophenotypic characteristics with extranodal and splenic marginal zone lymphoma. Data on altered genes and signaling pathways are scarce in this rare tumor entity. To gain further insights into the genetic background of NMZL, seven cases were investigated by microarray analysis, G‐banding, and FISH. Chromosomal imbalances were observed in 3/7 cases (43%) with gains of chromosome arms 1q, 8q, and 12q being the most frequent findings. Furthermore, we identified a translocation t(11;14)(q23;q32) involving IGH and DDX6. Chromosomal walking, expression analysis, siRNA‐mediated gene knockdown and a yeast two hybrid screen were performed for further characterization of the translocation in vitro. In siRNA experiments, DDX6 appeared not to be involved in NF‐κB activation as frequently observed for genes promoting lymphomagenesis but was found to interfere with the expression of BCL6 and BCL2 in an NF‐κB independent manner. In conclusion, we identified several unbalanced aberrations and a t(11;14) involving IGH and DDX6 providing evidence for a contribution of DDX6 to lymphomagenesis by deregulation of BCL6 in NMZL. © 2012 Wiley Periodicals, Inc.
Expression of the MAP kinase phosphatase DUSP4 is associated with microsatellite instability in colorectal cancer (CRC) and causes increased cell proliferation
Authors: Gröschl Benedikt , Bettstetter Marcus , Giedl Christian , Woenckhaus Matthias , Edmonston Tina , Hofstädter Ferdinand , Dietmaier Wolfgang
Published: September 2012

DUSP4 (MKP‐2), a member of the mitogen‐activated protein kinase phosphatase (MKP) family and potential tumor suppressor, negatively regulates the MAPKs (mitogen‐activated protein kinases) ERK, p38 and JNK. MAPKs play a crucial role in cancer development and progression. Previously, using microarray analyses we found a conspicuously frequent overexpression of DUSP4 in colorectal cancer (CRC) with high frequent microsatellite instability (MSI‐H) compared to microsatellite stable (MSS) CRC. Here we studied DUSP4 expression on mRNA level in 38 CRC (19 MSI‐H and 19 MSS) compared to matched normal tissue as well as in CRC cell lines by RT‐qPCR. DUSP4 was overexpressed in all 19 MSI‐H tumors and in 14 MSS tumors. Median expression levels in MSI‐H tumors were significantly higher than in MSS‐tumors (p < 0.001). Consistently, MSI‐H CRC cell lines showed 6.8‐fold higher DUSP4 mRNA levels than MSS cell lines. DUSP4 expression was not regulated by promoter methylation since no methylation was found by quantitative methylation analysis of DUSP4 promoter in CRC cell lines neither in tumor samples. Furthermore, no DUSP4 mutation was found on genomic DNA level in four CRC cell lines. DUSP4 overexpression in CRC cell lines through DUSP4 transfection caused upregulated expression of MAPK targets CDC25A, CCND1, EGR1, FOS, MYC and CDKN1A in HCT116 as well as downregulation of mismatch repair gene MSH2 in SW480. Furthermore, DUSP4 overexpression led to increased proliferation in CRC cell lines. Our findings suggest that DUSP4 acts as an important regulator of cell growth within the MAPK pathway and causes enhanced cell growth in MSI‐H CRC.
Genetic variation in the lactase gene, dairy product intake and risk for prostate cancer in the European prospective investigation into cancer and nutrition
Authors: Travis Ruth C. , Appleby Paul N. , Siddiq Afshan , Allen Naomi E. , Kaaks Rudolf , Canzian Federico , Feller Silke , Tjønneland Anne , Føns Johnsen Nina , Overvad Kim , Ramón Quirós J. , González Carlos A. , Sánchez Maria‐José , Larrañaga Nerea , Chirlaque Maria‐Dolores , Barricarte Aurelio , Khaw Kay‐Tee , Wareham Nick , Trichopoulou Antonia , Valanou Elisavet , Oustoglou Erifili , Palli Domenico , Sieri Sabina , Tumino Rosario , Sacerdote Carlotta , Bueno‐de‐Mesquita H. B(as). , Stattin Pär , Ferrari Pietro , Johansson Mattias , Norat Teresa , Riboli Elio , Key Timothy J
Published: September 2012

High dairy protein intake has been found to be associated with increased prostate cancer risk in the European Prospective Investigation into Cancer and Nutrition (EPIC). To further examine this possible relationship, we investigated the hypothesis that a genetic polymorphism in the lactase (LCT) gene might be associated with elevated dairy product intake and increased prostate cancer risk in a case–control study nested in EPIC. The C/T‐13910 lactase variant (rs4988235) was genotyped in 630 men with prostate cancer and 873 matched control participants. Dairy product consumption was assessed by diet questionnaire. Odds ratios (ORs) for prostate cancer in relation to lactase genotype were estimated by conditional logistic regression. Lactase genotype frequency varied significantly between countries, with frequencies of the T (lactase persistence) allele ranging from 7% in Greece to 79% in Denmark. Intake of milk and total dairy products varied significantly by lactase genotype after adjustment for recruitment center; adjusted mean intakes of milk were 44.4, 69.8 and 82.3 g/day among men with CC, CT and TT genotypes, respectively. The lactase variant was not significantly associated with prostate cancer risk, both in our data (adjusted OR for TT vs. CC homozygotes: 1.10, 95% CI: 0.76–1.59) and in a meta‐analysis of all the published data (combined OR for T allele carriers vs. CC homozygotes: 1.12, 0.96–1.32). These findings show that while variation in the lactase gene is associated with milk intake in men, the lactase polymorphism does not have a large effect on prostate cancer risk.
Pharmacological validation of early and late phase of rat mono‐iodoacetate model using the Tekscan system
Authors: Rashid M.H. , Theberge Y. , Elmes S.J. , Perkins M.N. , McIntosh F.
Published: September 2012

Background

Previous pharmacological validations of the rat mono‐iodoacetate (MIA)‐induced chronic joint pain model were mostly performed by measuring weight‐bearing (WB) deficit with an incapacitance tester. However, conventional incapacitance testers have several drawbacks including restrain stress on animal and sole use of hind limbs WB.

Objectives

The aim of the present study was to compare pharmacological sensitivity of the early (up to 1 week after MIA) versus late (between 2 and 4 weeks after MIA) phase of the rat MIA model using a highly sensitive tactile pressure measurement system (Tekscan®), which can measure weight borne by all four limbs and the tail in a non‐restrained animal.

Methods

The Tekscan® WB measurement system was used in MIA rats to examine the acute and chronic dosing effects of drugs that targeted different mechanisms. Electrophysiological recordings from joint afferents and biochemical analysis of synovial fluid were also performed.

Results

Dexamethasone, duloxetine and morphine significantly alleviated WB deficits in the Tekscan® system during both early and late phase of the MIA model while celecoxib and naproxen alleviated WB deficit only during the early phase. Similarly, naproxen was able to inhibit spontaneous neuronal activity from MIA joint afferents only during the early phase. Finally, concentrations of prostaglandin E2 in synovial fluid were elevated only during the early phase of the rat MIA model.

Conclusions

Our pharmacological validation studies using the Tekscan® system along with electrophysiological and biochemical results suggest different mechanisms for early and late phase of MIA‐induced chronic joint pain in rat.
Diethyl phthalate enhances expression of SIRT1 and DNMT3a during apoptosis in PC12 cells
Authors: Sun Yongkun , Guo Zhikun , Iku Shouhei , Saito Takeshi , Kurasaki Masaaki
Published: September 2012

Diethyl phthalate (DEP) works as a phthalate plasticizer and is ubiquitously used in personal care products, cosmetics, medical equipment and pharmaceutical coating. DEP is considered a potential endocrine disruptor. Previously we found DEP‐enhanced apoptosis induced by serum deprivation in PC12 cells. However, the relationship between DEP and longevity‐related factors, sirtuins and epigenetic factors (e.g. DNA methyltransferases) remains unclear, because genome modification caused by chemical toxicity, sirtuins and epigenetic factors have played key roles in abnormal metabolism and development. Here, we investigate whether DEP affects sirtuins (SIRT1 and SIRT2) and methyltranferases (DNMT1 and DNMT3a) on the apoptosis of PC12 cells. We found that DNMT3a was significantly decreased by serum deprivation. However, DNMT3a, DNMT3b and SIRT1 were significantly increased under the enhancement of apoptosis induced by serum deprivation. These results suggest that SIRT1, DNMT3a and DNMT3b play multiple and complex roles in different apoptotic stages. Our results showed DEP triggered epigenetic factors on PC12 cells apoptosis under nutrition stress. Finally, our results suggest that monitoring epigenetic factors such as DNMT3a, DNMT3b and SIRT1 could be a useful tool for chemical toxicity risk assessment. Copyright © 2012 John Wiley & Sons, Ltd.
Phagocytic uptake and ROS‐mediated cytotoxicity in human hepatic cell line of amphiphilic polyphosphazene nanoparticles
Authors: Qiu Liyan , Chen Yue , Gao Menghua , Zheng Cheng , Zhao Qinghe
Published: September 2012

The pH‐responsive amphiphilic polyphosphazenes bearing N,N‐diisopropylethylenediamine (DPA) have been proven to be promising nanovehicles for drug antitumor therapy. To further modify these amphiphilic polyphosphazenes with fluorescent labeling agent or other biochemical functional groups, serine methyl ester containing active chemical group NH2 was chosen to be introduced to get a novel polymer [NP(PEG)0.24(DPA)0.5(SME)1.26]n (PDS‐NH2). Considering the possible toxic effect of NH2 group, the biocompatibility in bloodstream and nanotoxicity on human normal hepatic L‐02 cells was evaluated in this study. The polymer [NP(PEG)0.24(DPA)0.5(SME‐BOC)1.26]n (PDS‐BOC) linked with tert‐butyloxycarbonyl groups to protect and hide NH2 group was applied as the comparison. First, the bovine serum albumin (BSA) adsorption and phagocytic uptake behavior in human THP‐1 macrophages were performed. The results suggested that only a minor percentage of the nanoparticles were involved in BSA binding and phagocytic uptake as the result of PEGylation on the particulate surface. To determine the nanotoxicity on human normal hepatic L‐02 cells, we measured cell viability, apoptosis and necrosis, reactive oxygen species generation, the loss of mitochondrial membrane potential, and the levels of the apoptotic signaling proteins in L‐02 cells after the cells being exposed to nanoparticles of different concentrations (0.1, 0.2, and 0.5 mg/mL) for 24 h. Our data indicated that the two nanoparticles induced cytotoxicity in a dose‐dependent manner; PDS‐NH2 caused more cytotoxicity than PDS‐BOC as a result of NH2 exposure. The increased expression of caspase‐3 and caspase‐9 suggested that they triggered apoptosis through mitochondria‐dependent pathways in L‐02 cells. © 2012 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 101A:285–297, 2013.
Cell adhesion and osteogenic differentiation on three‐dimensional pillar surfaces
Authors: Kaivosoja Emilia , Suvanto Pia , Barreto Gonçalo , Aura Susanna , Soininen Antti , Franssila Sami , Konttinen Yrjö T.
Published: September 2012

We hypothesized that when compared with conventional two‐dimensional (2D) cultures, substrates containing 3D micropillars would allow cells to grow at levels, activating their cytoskeleton to promote osteogenesis. Fibroblasts, osteoblast‐like cells, and mesenchymal stem cells (MSCs) were studied. Planar substrates were compared with 200‐nm‐, 5‐μm‐, and 20‐μm‐high pillars of Ormocomp®, Si, diamond‐like carbon, or TiO2. Scanning electron microscopy and staining of actin cytoskeleton showed 7.5‐h adhesion to pillar edges and 5‐day stretching between adhesion contacts > 100‐μm distances of fibroblast and MSC in 3D networks, whereas SaOS‐2 cells adhered flatly and individually on horizontal and vertical surfaces. ERK and ROCK immunostaining at 14 and 21 days confirmed activation of the cytoskeleton. In contrast to expectations, success to induce osteogenesis was dominated by the cytocompatibility of the substrate over the 3D structure. This was shown using early alkaline phosphatase, intermediate osteopontin, and late mineralization markers, together with bone nodule formation, which were seen in planar substrates and low‐profile TiO2 pillars, but were poor in the 20‐μm landscape. The lack of intercellular contacts seems to halt the osteogenesis‐promoting effects of cytoskeletal organization and tension described earlier. © 2012 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 101A: 842–852, 2013.
Novel selenium‐doped hydroxyapatite coatings for biomedical applications
Authors: Rodríguez‐Valencia C. , López‐Álvarez M. , Cochón‐Cores B. , Pereiro I. , Serra J. , González P.
Published: September 2012

Nowadays there is a short‐term need of investigating in orthopedic implants with a greater functionality, including an improved osseointegration and also antibacterial properties. The coating of metallic implants with hydroxyapatite (HA) remains to be the main proposal, but superior quality HA coatings with compositions closer to natural bone apatites, including carbonates, trace elements are required. Selenium is an essential nutrient in biological tissues and, at the same time, it also presents antibacterial properties. A pioneering study on the fabrication of selenium‐doped carbonated hydroxyapatite (iHA:Se) coatings by Pulsed Laser Deposition (PLD) is presented. Different proportions of selenium were incorporated to obtain the iHA:Se coatings. Their physicochemical characterization, performed by SEM/EDS, FTIR, FT‐Raman, Interferometric Profilometry and XPS, revealed typical columnar growth of HA in globular aggregates and the efficient incorporation of selenium into the HA coatings by the, most probably, substitution of SeO groups in the CO sites. Biological evaluation illustrated the absence of cytotoxicity when an amount of 0.6 at.% of Se was added to the iHA:Se coatings and excellent proliferation of the MC3T3‐E1 preosteoblasts. Antibacterial properties were also proved with the inhibition of P. aeruginosa and S. aureus from establishing bacterial biofilms. © 2012 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 101A: 853–861, 2013.
Study of the effect of membrane thickness on microcapsule strength, permeability, and cell proliferation
Authors: Ma Ying , Zhang Ying , Wang Yu , Wang Qiuyan , Tan Mingqian , Liu Yang , Chen Li , Li Na , Yu Weiting , Ma Xiaojun
Published: September 2012

Cell microencapsulation is one of the promising strategies for in vitro production of proteins or in vivo delivery of therapeutic products. Membrane thickness controls microcapsule strength and permeability, which may in return affect cell growth and metabolism. In this study, the strength, permeability, and encapsulated Chinese hamster ovary cell proliferation and metabolism of four groups of microcapsules with different membrane thicknesses were investigated. It was found that increasing membrane thickness increases microcapsule strength, whereas decreases membrane permeability. During the first 6 days, cells within microcapsules with 10 μm thickness membrane proliferated fast and could reach a cell density of 1.9 × 107 cells/mL microcapsule with 92% cell density. A cell density of 5.5 × 107 cells/mL microcapsule with >85% cell density was achieved within microcapsules with 15 μm membrane thickness and these microcapsules kept over 88% integrity ratio after 11 days, which was much higher than that of microcapsules with 10 μm membrane thickness. Membrane with more than 20 μm thickness was not suited for encapsulated cell culture owing to low‐protein diffusion rate. These results indicated that cells survived shortly within the thinnest membrane thickness. There was a specific membrane thickness more suitable for cell growth for a long‐time culture. These findings will be useful for preparing microcapsules with the desired membrane thickness for microencapsulated cell culture dependent on various purposes. © 2012 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2013.
In vitro evaluation of hydroxyapatite–chitosan–gelatin composite membrane in guided tissue regeneration
Authors: Hunter Kimberly T. , Ma Teng
Published: September 2012

Resorbable biomaterials have been investigated as barrier membranes to compartmentalize the periodontal defects while selectively guiding osteoprogenitor cell proliferation and bone tissue expansion. Hydroxyapatite (H), chitosan (C), and gelatin (G) have chemical similarity to the structural components of natural bone and their composites have been tested as bone scaffolds. Human mesenchymal stem or stromal cells (hMSCs) are inducible osteoprogenitors and are responsible for bone tissue repair and regeneration. In this study, the dynamic interactions of hMSC with composite hydroxyapatite–chitosan–gelatin (HCG) membranes were investigated. The association of HCG formed a biodegradable membrane with ∼60 wt % water and an initial stiffness of ∼20 kPa. Preconditioning in serum‐containing media resulted in the formation nanopores in the HCG membranes and the increase of extracellular matrix (ECM) protein adsorption. Expression of integrin α2β1 and α5β1 coincided with ECM enrichment, suggesting the enhanced cell–ECM interactions. The elevated expression of bone marker proteins and genes in the HCG membranes suggests the progression of hMSC osteogenic differentiation in the absence of chemical induction. The results showed that the HCG membranes possess sufficient mechanical and structural properties to function as a barrier membrane, and that the adsorbed ECM proteins effectively functionalized the HCG membranes and promoted hMSC osteogenic differentiation. © 2012 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2013.