bluuebunny/crossref_metadata_2025 huggingface dataset:
Protein switches identified from diverse insertion libraries created using S1 nuclease digestion of supercoiled‐form plasmid DNA
Authors: Tullman Jennifer , Guntas Gurkan , Dumont Matthew , Ostermeier Marc
Published: May 2011

We demonstrate that S1 nuclease converts supercoiled plasmid DNA to unit‐length, linear dsDNA through the creation of a single, double‐stranded break in a plasmid molecule. These double‐stranded breaks occur not only in the origin of replication near inverted repeats but also at a wide variety of locations throughout the plasmid. S1 nuclease exhibits this activity under conditions typically employed for the nuclease's single‐stranded nuclease activity. Thus, S1 nuclease digestion of plasmid DNA, unlike analogous digestion with DNaseI, effectively halts after the first double‐stranded break. This property makes easier the construction of large domain insertion libraries in which the goal is to insert linear DNA at a variety of locations throughout a plasmid. We used this property to create a library in which a circularly permuted TEM1 β‐lactamase gene was inserted throughout a plasmid containing the gene encoding Escherichia coli ribose binding protein. Gene fusions that encode allosteric switch proteins in which ribose modulates β‐lactamase catalytic activity were isolated from this library using a combination of a genetic selection and a screen. Biotechnol. Bioeng. 2011;108: 2535–2543. © 2011 Wiley Periodicals, Inc.
Trends and outcomes after surgical lower limb revascularization in England
Authors: Moxey P W , Hofman D , Hinchliffe R J , Jones K , Thompson M M , Holt P J E
Published: May 2011

Background

The aim was to analyse contemporary data on the number of surgical revascularization procedures performed each year in England, and their outcome.

Methods

Hospital Episode Statistics and Office for National Statistics data were used to quantify numbers and identify factors associated with outcome after all femoropopliteal and femorodistal bypass procedures performed between 2002 and 2006. Outcome measures were repeat bypass, major amputation, death and a composite measure. Single-level multivariable logistic regression modelling was used to quantify the effect of these variables on outcome.

Results

A total of 21 675 femoropopliteal and 3458 femorodistal bypass procedures were performed. Mean in-hospital mortality rates were 6·7 and 8·0 per cent respectively. One-year survival rates were 82·8 and 79·1 per cent; both increased over the study interval. The mean 1-year major amputation rate after femoropopliteal bypass was 10·4 per cent, which decreased significantly over the 5 years (P < 0·001); after distal bypass the rate of 20·8 per cent remained unchanged (P = 0·456). Diabetes mellitus and chronic kidney disease were significant predictors of adverse outcome for both procedures: odds ratio (OR) at 1 year 1·56 (95 per cent confidence interval 1·46 to 1·67; P < 0·001) and 2·15 (1·88 to 2·45; P < 0·001) respectively for femoropopliteal bypass. Previous femoral angioplasty was associated with an increased rate of major amputation 1 year after proximal bypass (OR 1·18, 1·05 to 1·33; P = 0·004).

Conclusion

Although all mortality rates are improving, the major amputation rate remains high after femorodistal bypass. Adverse events occurred after 37·6 per cent of femoropopliteal and 49·7 per cent of femorodistal bypasses; diabetes and chronic renal failure were the main predictors of poor outcome.
Roimatacene: An Antibiotic against Gram‐Negative Bacteria Isolated from Cystobacter ferrugineus Cb G35 (Myxobacteria)
Authors: Zander Wiebke , Gerth Klaus , Mohr Kathrin I. , Kessler Wolfgang , Jansen Rolf , Müller Rolf
Published: May 2011

Roimatacene (1) was isolated from the myxobacterium Cystobacter ferrugineus strain Cb G35 in a bioactivity‐guided process, by following the antimicrobial activity against Escherichia coli. Since 1 was extremely sensitive to oxygen, a protective isolation and handling protocol was developed, by utilizing the free radical scavenger 4‐ethoxyphenol. The structure of 1 was determined by HRMS, 1D and 2D NMR spectroscopy and chemical derivatization to acetonides and Mosher esters to finally establish the absolute configuration. Methionine and acetate were identified as building blocks in the biosynthesis of 1 by feeding experiments with differently 13C‐labeled precursors. The antimicrobial activity of 1 was determined in a broad screening revealing 1 to inhibit several Gram‐negative bacteria.
Hydrothermal Synthesis of Na0.5La0.5TiO3–LaCrO3 Solid‐Solution Single‐Crystal Nanocubes for Visible‐Light‐Driven Photocatalytic H2 Evolution
Authors: Shi Jinwen , Ye Jinhua , Zhou Zhaohui , Li Mingtao , Guo Liejin
Published: May 2011

Cubic perovskite structure photocatalysts of Na0.5La0.5TiO3 and (Na0.5La0.5TiO3)1.00(LaCrO3)0.08 solid solution that consisted of well‐defined single‐crystal nanocubes were successfully prepared by means of facile and surfactant‐free hydrothermal reactions for the first time. The results from different instrumental characterizations and theoretical calculations consistently confirmed the formation of nanocubic single‐crystal solid solution of (Na0.5La0.5TiO3)1.00(LaCrO3)0.08, and clearly revealed the modification of its physicochemical properties compared with those of Na0.5La0.5TiO3. In particular, the effective narrowing of the bandgap (from 3.19 to 2.25 eV) by Cr3+ in the solid solution made it possible to utilize visible light. The solid‐solution configuration maintained the charge balance to preserve the valence of Cr3+ rather than Cr6+, and accommodated Cr3+ with high content to form new energy bands instead of localized impurity levels. The hydrothermal preparation strategy ensured the formation of single crystals with high purity, few defects, and regulated morphology; it also guaranteed the valences of Ti4+ and Cr3+ in the solid solution. Consequently, the recombination of photogenerated carriers could be effectively suppressed to benefit photocatalytic H2 evolution. (Na0.5La0.5TiO3)1.00(LaCrO3)0.08 nanocubic single‐crystal solid solution showed stable photocatalytic activity, and thus was proved to be a promising candidate for visible‐light‐driven photocatalytic H2 evolution.
Push–Pull Amino Succinimidyl Ester Thiophene‐Based Fluorescent Dyes: Synthesis and Optical Characterization
Authors: Sotgiu Giovanna , Galeotti Matteo , Samorí Cristian , Bongini Alessandro , Mazzanti Andrea
Published: May 2011

The design and synthesis of new fluorescent dyes with emission range at 490–650 nm are described. Their structural and electronic properties have been characterized by both experimental techniques and quantum‐chemical calculations. The chromophores are donor–π‐bridge–acceptor push–pull compounds with a π bridge of phenyl and thiophene rings and their combination. Compared with previous thiophene fluorophores, these dyes show significant redshift in the absorption and emission spectra and offer compact, red‐emitting fluorophores. The dyes have amino succinimidyl active ester and can be readily conjugated to proteins, polymers and other amino‐group‐containing materials.
Synthesis of 5‐Acetyloxazoles and 1,2‐Diketones from β‐Alkoxy‐β‐ketoenamides and Their Subsequent Transformations
Authors: Lechel Tilman , Gerhard Markus , Trawny Daniel , Brusilowskij Boris , Schefzig Luise , Zimmer Reinhold , Rabe Jürgen P. , Lentz Dieter , Schalley Christoph A. , Reissig Hans‐Ulrich
Published: May 2011

Lithiated alkoxyallenes, nitriles, and carboxylic acids have been employed as precursors in a three‐component reaction leading to highly substituted β‐alkoxy‐β‐ketoenamides. Upon treatment with trifluoroacetic acid, these enamides could be easily cyclized to 5‐acetyloxazole derivatives. The synthesis is very flexible with respect to the substitution pattern at C‐2 and C‐4 of the oxazole core. A mechanistic suggestion for the oxazole formation is presented on the basis of 18O‐labeled compounds and their mass spectrometric analysis. In several cases, 1,2‐diketones are formed as side products or even as major components. The acetyl moiety at C‐5 of the oxazole derivatives can efficiently be converted into alkenyl or alkynyl moieties, which allows a multitude of subsequent reactions. Condensation reactions of the acetyl group provided the expected oxime or hydrazone. By applying a Fischer reaction, the phenylhydrazone could be transferred into an indole, which emphasizes the potential of 5‐acetyloxazoles for the preparation of highly substituted (poly)heterocyclic systems. The alkynyl group at C‐2 is prone to addition reactions, providing an enamine with interesting photophysical properties. Sonogashira couplings were performed with 5‐alkynyl‐substituted oxazoles, furnishing the expected aryl‐substituted products. This alkynyl unit was employed for the preparation of a new, star‐shaped trisoxazole derivative. The ability of this multivalent compound to form self‐assembled monolayers between the basal plane of highly oriented pyrolytic graphite and 1‐phenyloctane was demonstrated by scanning tunneling microscopy (STM). The star‐shaped compound seems to prefer the C3‐symmetric arrangement in this two‐dimensional crystal. Two 1,2‐diketones were smoothly converted into functionalized quinoxaline derivatives.
Frequency‐Domain Fourier‐Transform Terahertz Spectroscopy of the Single‐Molecule Magnet (NEt4)[Mn2(5‐Brsalen)2(MeOH)2Cr(CN)6]
Authors: Dreiser Jan , Schnegg Alexander , Holldack Karsten , Pedersen Kasper S. , Schau‐Magnussen Magnus , Nehrkorn Joscha , Tregenna‐Piggott Philip , Mutka Hannu , Weihe Høgni , Bendix Jesper , Waldmann Oliver
Published: May 2011

We have investigated the novel single‐molecule magnet (NEt4)[Mn2(5‐Brsalen)2(MeOH)2Cr(CN)6] (1; 5‐Brsalen=N,N′‐ethylenebis(5‐bromosalicylidene)iminato anion) using spectroscopic as well as magnetization and susceptibility measurements. Frequency‐domain Fourier‐transform terahertz electron paramagnetic resonance (FDFT THz‐EPR) based on the generation of THz radiation from a synchrotron in combination with inelastic neutron scattering (INS) allows for the discrimination between intermultiplet and intramultiplet transitions. Together with ac/dc magnetic susceptibility measurements the obtained set of data provides a complete characterization of the lowest energetic magnetic excitations. We find that the new compound 1 exhibits much weaker intermolecular interactions than found in the closely related compound: K[Mn2(5‐Brsalen)2(H2O)2Cr(CN)6] (2). Furthermore, two phonon lines in the vicinity of the magnetic excitations are detected.
A quantitative measure of chirality inside nucleic acid databank
Authors: Pietropaolo Adriana , Parrinello Michele
Published: May 2011

We show the capability of a chirality index (Pietropaolo et al., Proteins 2008;70:667–677) to investigate nucleic acid structures because of its high sensitivity to helical conformations. By analyzing selected structures of DNA and RNA, we have found that sequences rich in cytosine and guanine have a tendency to left‐handed chirality, in contrast to regions rich in adenine or thymine which show strong negative, right‐handed, chirality values. We also analyze RNA structures, where specific loops and hairpin motifs are characterized by a well‐defined chirality value. We find that in nucleosome the chirality is exalted, whereas in ribosome it is reduced. Our results illustrate the sensitivity of this descriptor for nucleic acid conformations. Chirality 2011. © 2011 Wiley‐Liss, Inc.
Conversions of Cellobiose and Inulin to Deoxyfructosazine in Aqueous Solutions
Authors: Wu Suxiang , Fan Honglei , Zhang Qian , Cheng Yan , Wang Qian , Yang Guanying , Han Buxing
Published: May 2011

The conversion of the biomass into high value‐added chemicals is an attractive topic in green chemistry. In this work, we studied the synthesis of 2,6‐deoxyfructosazine (2,6‐DOF) and 2,5‐deoxyfructosazine (2,5‐DOF) from cellobiose and inulin, respectively, in aqueous solution by an one‐pot two‐step route. In this method, cellobiose or inulin was first hydrolyzed into glucose or fructose and then reacted directly with ammonium formate to form 2,6‐DOF or 2,5‐DOF. The effects of reaction temperature, reaction time, pH value on the yields of the products were studied. It was demonstrated that the cellobiose and inulin could be converted to 2,6‐DOF and 2,5‐DOF effectively at optimized condition.
Inputs to the midbrain dopaminergic complex in the rat, with emphasis on extended amygdala‐recipient sectors
Authors: Zahm Daniel S. , Cheng Anita Y. , Lee Tristan J. , Ghobadi Comeron W. , Schwartz Zachary M. , Geisler Stefanie , Parsely Kenneth P. , Gruber Clemens , Veh Ruediger W.
Published: May 2011

The midbrain dopaminergic neuronal groups A8, A9, A10, and A10dc occupy, respectively, the retrorubral field (RRF), substantia nigra compacta (SNc), ventral tegmental area (VTA), and ventrolateral periaqueductal gray (PAGvl). Collectively, these structures give rise to a mixed dopaminergic and nondopaminergic projection system that essentially permits adaptive behavior. However, knowledge is incomplete regarding how the afferents of these structures are organized. Although the VTA is known to receive numerous afferents from cortex, basal forebrain, and brainstem and the SNc is widely perceived as receiving inputs mainly from the striatum, the afferents of the RRF and PAGvl have yet to be assessed comprehensively. This study was performed to provide an account of those connections and to seek a better understanding of how afferents might contribute to the functional interrelatedness of the VTA, SNc, RRF, and PAGvl. Ventral midbrain structures received injections of retrograde tracer, and the resulting retrogradely labeled structures were targeted with injections of anterogradely transported Phaseolus vulgaris leucoagglutinin. Whereas all injections of retrograde tracer into the VTA, SNc, RRF, or PAGvl produced labeling in many structures extending from the cortex to caudal brainstem, pronounced labeling of structures making up the central division of the extended amygdala occurred following injections that involved the RRF and PAGvl. The anterograde tracing supported this finding, and the combination of retrograde and anterograde labeling data also confirmed reports from other groups indicating that the SNc receives robust input from many of the same structures that innervate the VTA, RRF, and PAGvl. J. Comp. Neurol. 519:3159–3188, 2011. © 2011 Wiley‐Liss, Inc.
Assembly properties of lamprey neurofilament subunits and their expression after spinal cord transection
Authors: Zhang Guixin , Jin Liqing , Selzer Michael E.
Published: May 2011

In mammals neurofilaments (NF) are formed by coassembly of three subunits: NFL, NFM, and NFH (light, medium, and heavy). It had been believed that lampreys have only one subunit, NF180. However, a previous study showed that NF180 could not self‐assemble but could coassemble with rat NFL, suggesting the existence of additional NF subunits in lamprey. More recently, we cloned three additional NF subunits. These new subunits and NF180 have now been transfected in combinations into SW13cl.2Vim− cells, which lack endogenous cytoplasmic intermediate filaments. None of the subunits could self‐assemble. No combination of NF subunits could form filaments in the absence of lamprey NFL (L‐NFL). Assembly occurred at 28°C, but not at 37°C. L‐NFL could form thick NF bundles with NF180 but not with NF132 and NF95, which formed only fine filamentous arrays. To determine which parts of the NF subunits are required for filament or bundle formation, we constructed deletion mutants of NF180 and cotransfected them with L‐NFL. As with mammalian NF, only constructs with intact head and core domains could form filaments with L‐NFL. However, the full length of NF180 was required to form NF bundles. As with NF180, in situ hybridization indicated that mRNA for L‐NFL and NF132 was downregulated in identified reticulospinal neurons by 5 weeks after spinal cord transection, but was reexpressed at 10 weeks selectively in those neurons whose axons have a high probability of regenerating. This is consistent with a possible role of NFs in the mechanism of axon regeneration. J. Comp. Neurol. 519:3657–3671, 2011. © 2011 Wiley‐Liss, Inc.
Subcellular localization of patched and smoothened, the receptors for sonic hedgehog signaling, in the hippocampal neuron
Authors: Petralia Ronald S. , Schwartz Catherine M. , Wang Ya‐Xian , Mattson Mark P. , Yao Pamela J.
Published: May 2011

Cumulative evidence suggests that, aside from patterning the embryonic neural tube, Sonic hedgehog (Shh) signaling plays important roles in the mature nervous system. In this study, we investigate the expression and localization of the Shh signaling receptors, Patched (Ptch) and Smoothened (Smo), in the hippocampal neurons of young and mature rats. Reverse transcriptase‐polymerase chain reaction and immunoblotting analyses show that the expression of Ptch and Smo remains at a moderate level in young postnatal and adult brains. By using immunofluorescence light microscopy and immunoelectron microscopy, we examine the spatial distribution of Ptch and Smo within the hippocampal neurons. In young developing neurons, Ptch and Smo are present in the processes and are clustered at their growth cones. In mature neurons, Ptch and Smo are concentrated in dendrites, spines, and postsynaptic sites. Synaptic Ptch and Smo often co‐exist with unusual structures—synaptic spinules and autophagosomes. Our results reveal the anatomical organization of the Shh receptors within both the young and the mature hippocampal neurons. J. Comp. Neurol. 519:3684‐3699, 2011. ©2011 Wiley‐Liss, Inc.
Redox‐Responsive Rhodocenium [O,O]‐, [N,O]‐, [N,N]‐, and [N,C,N]‐Metalloligands
Authors: Eisenstecken Daniela , Enk Barbara , Kopacka Holger , Wurst Klaus , Müller Thomas , Pevny Florian , Winter Rainer F. , Bildstein Benno
Published: May 2011

As we have recently shown, the doubly substituted [1,2‐O,O]H, [1,2‐O,N]H, [1,2‐N,N]H, and [1,3‐N,N]H pentafulvenes are convenient N/O‐functionalized cyclopentadienide (Cp) precursors. Deprotonation of the pentafulvenes by potassium hydride followed by reaction with [Cp*RhCl2]2 or [Cp*IrCl2]2 gave access to the first functionalized rhodocenium and iridocenium salts that contain two acyl and/or imidoyl substituents. These air‐stable compounds represent interesting bis(acyl/imidoyl) or mixed acyl/imidoyl metalloligand systems that combine an axially shielding, electrochemically active metallocene moiety with directly attached, conjugating oxygen and/or nitrogen donor sites. The structural properties of these novel metallocene metalloligands in solution and in the solid state were studied by NMR spectroscopy (1H, 13C, 103Rh) and by single‐crystal structure analysis. The electrochemical investigations on complexes 5a, 6b, and 7a showed the effect of the appended functional groups on the potential and on the reversibility of the rhodocenium/rhodocene reduction and provided evidence for the dimerization that followed the reduction. Further redox processes that were due to the heteroatom functions included the stepwise reduction of the 1,2‐diketo entity of 5a and the oxidation of the 1,2‐diimino moiety of 7a.
Electrochemical Studies of Ochratoxin A Mycotoxin at Gold Electrodes Modified with Cysteamine Self‐Assembled Monolayers. Its Ultrasensitive Quantification in Red Wine Samples
Authors: Perrotta Patricio René , Vettorazzi Nelio Roberto , Arévalo Fernando Javier , Granero Adrián Marcelo , Chulze Sofía Noemí , Zón María Alicia , Fernández Héctor
Published: May 2011

The quantification of ochratoxin A is studied at cysteamine self‐assembled monolayer modified gold electrodes in red wine samples by square wave voltammetry. Detection and quantification limits of 0.004 µg L−1 and 0.012 µg L−1, respectively, were determined. The recovery percentages were in the range from 146 % to 94.0 % at spiking levels ranging from 0.02 to 5 µg L−1. The variation coefficients for within‐laboratory repeatability varied from 31.4 to 11.5 % for spiked level from 0.02 to 2.0 µg L−1. The developed electrochemical method is efficient, reproducible, and ultrasensitive for the quantification of OTA in red wine samples.
Alcohol diagnoses among older Tennessee Medicare beneficiaries: race and gender differences
Authors: Kilbourne Barbara Jean , Cummings Sherry M. , Levine Robert
Published: May 2011

Background

These analyses bolster a sparse body of research focusing on the rate of alcohol disorders among older adults, particularly race and gender subgroups.

Methods

We based the study on cross‐sectional data from all Medicare billed physician/patient encounters. Analyses of these data included cross‐tabulations, difference of means tests, and difference of proportions tests, logistic regression and multinomial logistic regression. These analyses were based Medicare billing records from physician/patient encounters in Tennessee. Data included Tennessee Medicare billings beneficiaries enrolled in Medicare Part B, who saw a physician at least once in 2000. Patients with billings containing ICD‐9 codes: 303 (alcohol abuse), 305 (alcohol dependence), 291 (alcohol psychosis), or 571.1‐571.3 (alcohol‐related liver disease including cirrhosis of the liver) as to primary diagnosis were considered alcohol‐disordered.

Results

Analyses reveal the overall rate of alcohol disorders, subgroup variation in rates and differences in pattern of specific disorders. Merely 0.04% of Tennessee Medicare beneficiaries were diagnosed with any type of alcoholism, a rate much lower than those reported in previous studies. Rates of alcohol disorders varied across groups, with significantly higher rates for Black men. The type alcohol disorder also varied across groups.

Conclusions

Many encounters with the medical system result in missed opportunities to identify and treat alcohol disorders, a significant risk factor among older adults. Alcoholism both triggers and exacerbates many chronic conditions among older adults. The earlier in the disease trajectory the more of these conditions could be prevented or more efficiently managed, resulting in substantial savings in health care costs. Copyright © 2011 John Wiley & Sons, Ltd.
White matter integrity and behavioral activation in healthy subjects
Authors: Xu Jiansong , Kober Hedy , Carroll Kathleen M. , Rounsaville Bruce J. , Pearlson Godfrey D. , Potenza Marc N.
Published: May 2011

Individual differences in behavioral inhibition and behavioral activation may place certain people at greater risk for neuropsychiatric disorders and engagement in risky behaviors. Therefore, studying the neural correlates of behavioral inhibition and activation may help us understand neural mechanisms underlying risk behaviors in both clinical and non‐clinical populations. To investigate, we assessed the relationships between white matter integrity and measures of behavioral inhibition and behavioral activation in 51 healthy participants using diffusion tensor imaging (DTI) and the Behavioral Inhibition System/Behavioral Activation System (BIS/BAS) scale. Scores on the Fun‐Seeking subscale of the BAS positively correlated with DTI fractional anisotropy in the left corona radiata and adjacent superior longitudinal fasciculus, and with mean diffusivity in the left inferior longitudinal fasciculus and inferior fronto‐occipital fasciculus after controlling for age, gender, and education. These findings suggest that the integrity of white matter connecting extensive brain regions implicated in self‐control and the processing of rewards and emotions are associated with individual differences in the motivation for seeking and participating in fun and novel experiences. Hum Brain Mapp , 2012. © 2011 Wiley Periodicals, Inc.
sLORETA allows reliable distributed source reconstruction based on subdural strip and grid recordings
Authors: Dümpelmann Matthias , Ball Tonio , Schulze‐Bonhage Andreas
Published: May 2011

Source localization based on invasive recordings by subdural strip and grid electrodes is a topic of increasing interest. This simulation study addresses the question, which factors are relevant for reliable source reconstruction based on sLORETA. MRI and electrode positions of a patient undergoing invasive presurgical epilepsy diagnostics were the basis of sLORETA simulations. A boundary element head model derived from the MRI was used for the simulation of electrical potentials and source reconstruction. Focal dipolar sources distributed on a regular three‐dimensional lattice and spatiotemporal distributed patches served as input for simulation. In addition to the distance between original and reconstructed source maxima, the activation volume of the reconstruction and the correlation of time courses between the original and reconstructed sources were investigated. Simulations were supplemented by the localization of the patient's spike activity. For noise‐free simulated data, sLORETA achieved results with zero localization error. Added noise diminished the percentage of reliable source localizations with a localization error ≤15 mm to 67.8%. Only for source positions close to the electrode contacts the activation volume correctly represented focal generators. Time‐courses of original and reconstructed sources were significantly correlated. The case study results showed accurate localization. sLORETA is a distributed source model, which can be applied for reliable grid and strip based source localization. For distant source positions, overestimation of the extent of the generator has to be taken into account. sLORETA‐based source reconstruction has the potential to improve the localization of distributed generators in presurgical epilepsy diagnostics and cognitive neuroscience. Hum Brain Mapp , 2011. © 2011 Wiley‐Liss, Inc.
Overnight deprivation from smoking disrupts amygdala responses to fear
Authors: Onur Oezguer A. , Patin Alexandra , Mihov Yoan , Buecher Boris , Stoffel‐Wagner Birgit , Schlaepfer Thomas E. , Walter Henrik , Maier Wolfgang , Hurlemann René
Published: May 2011

Cigarette smoking, a major, yet avoidable, cause of disability and premature death, is the most prevalent form of nicotine addiction. An emerging theme in the neurobiology of nicotine addiction is the integrity of the amygdala. Using functional MRI, amygdala responses during a face perception task were compared between 28 chronic smokers [14 females, 14 males; age, 26.3 (2.8) years; age at onset of smoking, 15.8 (2.6) years; years smoked, 9.1 (2.1); cigarettes per day, 17.1 (3.7); Fagerström test for nicotine dependence score, 4.1 (1.9); exhaled carbon‐monoxide level, 17.8 (9.5) ppm] and 28 age‐ and education‐matched nonsmokers [14 females, 14 males; age, 26.9 (2.4) years]. Subjects underwent imaging on two separate occasions 1 week apart: smoking satiety versus overnight smoking deprivation, in a randomized counterbalanced order. Our results show no difference in amygdala responses to faces between nonsmokers and satiated smokers. However, overnight deprivation from smoking was associated with a significantly lowered amygdala response to fear, an effect that was probabilistically mapped to the basolateral amygdala. We suggest that aberrant amygdala reactivity in overnight‐deprived smokers may reflect a pre‐existing vulnerability to smoking and/or increase the risk of smoking relapse after a cessation attempt. Hum Brain Mapp, 2011. © 2011 Wiley‐Liss, Inc.
Acid/base‐catalyzed cyclization of O‐alkynylphenylphosphonic acid monoesters and (O‐hydroxyphenyl)ethynylphosphinates
Authors: Peng Ai‐Yun , Du Yidan , Wei Yingling , Xie Liang , Ding Yi‐Xiang
Published: May 2011

In this work, we found that acids (i.e., HCl and NaHSO3) rather than bases could catalyze the cyclization of o‐alkynylphenylphosphonic acid monoesters at slow rates and give phosphaisocoumarins in low to medium yields, whereas the cyclization of (o‐hydroxyphenyl)ethynylphosphinates proceeded very smoothly under basic conditions rather than acidic conditions, and a series of phosphachromones could be prepared in excellent yields at room temperature using K2CO3 as catalyst. This is the first example of the synthesis of phosphorus heterocycles via acid/base‐catalyzed intramolecular cyclization of alkynes. Possible mechanisms were discussed. © 2011 Wiley Periodicals, Inc. Heteroatom Chem 22:649–652, 2011; View this article online at wileyonlinelibrary.com. DOI 10.1002/hc.20728
Impact of viral amino acid substitutions and host interleukin-28b polymorphism on replication and susceptibility to interferon of hepatitis C virus
Authors: Hiraga Nobuhiko , Abe Hiromi , Imamura Michio , Tsuge Masataka , Takahashi Shoichi , Hayes Nelson C. , Ochi Hidenori , Tateno Chise , Yoshizato Katsutoshi , Nakamura Yusuke , Kamatani Naoyuki , Chayama Kazuaki
Published: May 2011

Amino acid (aa) substitutions of core 70 and 91 and in the NS5A (nonstructural protein 5A) interferon sensitivity determining region (ISDR) as well as genetic polymorphisms in the host interleukin-28B (IL28B) locus affect the outcome of interferon (IFN)-based therapies for patients with chronic hepatitis C. The combination of these factors and the quasispecies nature of the virus complicate understanding of the underlying mechanism. Using infectious hepatitis C virus (HCV) genotype 1b clone HCV-KT9, we introduced substitutions at both core aa70 (Arg to Gln) and aa91 (Leu to Met). We also introduced four and nine ISDR aa substitutions into core mutant HCV-KT9. Using human hepatocyte chimeric mice with different IL28B genotypes, we examined the infectivity, replication ability, and susceptibility to IFN of these clones. Although aa substitutions in the ISDR significantly impaired infectivity and replication ability of the virus, core aa70 and 91 substitutions did not. The effect of IFN treatment was similar in core wild-type and mutant viruses. Interestingly, virus titer was significantly higher in mice with the favorable IL28B allele (rs8099917 TT and rs12979860 CC) in the transplanted hepatocytes than in mice with hepatocytes from rs8099917 TG and rs12979860 TT donors

(P

< 0.001). However, the effect of IFN was significantly greater, and intrahepatic expression levels of IFN-stimulated genes were significantly higher in mice with the favorable IL28B allele.

Conclusion:

Our data suggest that HCV replication levels and response to IFN are affected by human hepatocyte IL28B single-nucleotide polymorphism genotype and mutations in the ISDR. The mechanism underlying the clinically observed association of wild-type core protein in eradication-favorable host cells should be investigated further. (HEPATOLOGY 2011;)
Molecular imaging of hepatic stellate cell activity by visualization of hepatic integrin αvβ3 expression with SPECT in rat
Authors: Li Feng , Song Zhengji , Li Qinghua , Wu Jian , Wang Jiyao , Xie Cao , Tu Chuantao , Wang Jian , Huang Xiaowei , Lu Weiyue
Published: May 2011

The key factors in the pathogenesis of liver fibrosis are the activation and proliferation of hepatic stellate cells (HSCs), which express integrin αvβ3 after activation. This study aimed to explore the potential of 99mTc-labeled cyclic arginine-glycine-aspartic acid pentapeptide (cRGD) as a single photon emission computed tomography (SPECT) radiotracer to image hepatic integrin αvβ3 expression to reflect HSC activity in fibrotic livers. Rat models of liver fibrosis caused by thioacetamide or carbon tetrachloride (CCl4) treatment were employed to examine the expression and distribution of integrin αvβ3 during fibrotic progression or regression. The binding activity of radiolabeled cRGD to integrin αvβ3 was assessed in liver sections. SPECT was performed to determine hepatic integrin αvβ3 expression in rats with different stages of liver fibrosis. Protein and messenger RNA (mRNA) levels of integrin αv and β3 subunits were increased with the progression of liver fibrosis and reduced with its regression. The cell type that expressed the majority of integrin αvβ3 in fibrotic livers was found to be activated HSCs. The cRGD binding to activated HSCs displayed a high receptor-coupling affinity and an abundant receptor capacity. Iodine-125 (125I)-labeled cRGD bound to fibrotic liver sections and the binding activity was the highest in advanced fibrosis. Intravenously administered carboxyfluorescein-labeled cRGD was accumulated in fibrotic liver, and the accumulation amount was increased with the progression and reduced with the regression of fibrosis. A SPECT imaging study with 99mTc-labeled cRGD as a tracer demonstrated that the radioactivity ratio of liver to heart increased progressively along with severity of hepatic fibrosis. Conclusion: Hepatic integrin αvβ3 expression in fibrotic liver reflects HSC activity and its imaging using 99mTc-labeled cRGD as a SPECT radiotracer may distinguish different stages of liver fibrosis in rats. (HEPATOLOGY 2011;)
Temporal factors control hippocampal contributions to fear renewal after extinction
Authors: Zelikowsky Moriel , Pham Daniel L. , Fanselow Michael S.
Published: May 2011

Fear can be extinguished by repeated exposure to a cue that signals threat. However, extinction does not erase fear, as an extinguished cue presented in a context distinct from that of extinction results in renewed fear of that cue. The hippocampus, which is involved in the formation of contextual representations, is a natural candidate structure for investigations into the neural circuitry underlying fear renewal. Thus far, studies examining the necessity of the hippocampus for fear renewal have produced mixed results. We isolated the conditions under which the hippocampus may be required for renewal. Rats received lesions of the dorsal hippocampus either prior to tone fear conditioning or following extinction. Fear renewal was measured using discrete tone presentations or a long, continuous tone. The topography of fear responding at test was assessed by comparing “early” and “sustained” renewal, where early fear was determined by freezing to the first discrete tone or the equivalent initial segment of a continuous tone and sustained fear was determined by freezing averaged across all discrete tones or the entire continuous tone. We found that following pretraining damage of the hippocampus, early renewal remained intact regardless of lesion condition. However, sustained renewal only persisted in discrete, but not continuous, tone‐tested animals. A more extensive analysis of the topography of fear responding revealed that the disruption of renewal was generated when the tone duration at test began to violate that used during extinction, suggesting that the hippocampus is sensitive to mismatches in CS‐duration. Postextinction lesions resulted in an overall reduction of fear renewal. This pattern of results is consistent with those observed for contextual fear conditioning, wherein animals display a resistance to anterograde amnesia despite the presence of a strong retrograde amnesia for the same contextual information. Furthermore, the data support a role for the hippocampus in sustaining renewal when the CS duration at test does not match that used during extinction. © 2011 Wiley Periodicals, Inc.
Proteins linked to spatial memory formation of CD1 mice in the multiple T‐maze
Authors: Patil Sudarshan S. , Li Kongzhao , Heo Seok , Höger Harald , Lubec Gert
Published: May 2011

In own previous work CD1 mice were tested in the Multiple T‐maze (MTM), a robust land maze allowing determination of latency to reach the goal box with food reward and to evaluate correct decisions made on the way to the goal box. Herein, hippocampi of these animals were used for the current study with the aim to investigate differences in protein levels between trained and yoked mice and, moreover, to determine differences in protein levels between trained and yoked mice with and without memory formation in the MTM. Three training sessions were carried out for four training days each, followed by probe trials on Days 5 and 12. Good and no‐performers in the MTM were separated based on means and median of latency to reach the goal box on probe trial Day 12. Six hours following the probe trial on Day 12, animals were sacrificed and hippocampi were taken. Proteins were extracted and run on two‐dimensional gel electrophoresis, spots were quantified and differentially expressed proteins were identified by mass spectrometry using an ion trap. Levels of 17 proteins were significantly different in trained vs. yoked mice. Seven proteins were differentially expressed comparing trained vs. yoked mice from good and no‐performers. A series of proteins were significantly correlated with latency and may link these proteins to spatial memory formation. Differential protein expression in trained vs. yoked mice and in good and no‐performers may allow insight into spatial memory formation as well as represent tentative pharmacological targets. © 2011 Wiley Periodicals, Inc.
Inhibition of VEGF induces cellular senescence in colorectal cancer cells
Authors: Hasan Mohammad R. , Ho Shirley H. Y. , Owen David A. , Tai Isabella T.
Published: May 2011

Vascular endothelial growth factor (VEGF) inhibitors, such as bevacizumab, have improved outcomes in metastatic colorectal cancer (CRC). Recent studies have suggested that VEGF can delay the onset of cellular senescence in human endothelial cells. As VEGF receptors are known to be upregulated in CRC, we hypothesized that VEGF inhibition may directly influence cellular senescence in this disease. In our study, we observed that treatment with bevacizumab caused a significant increase (p < 0.05) in cellular senescence in vitro in several CRC cells, such as MIP101, RKO, SW620 and SW480 cells, compared to untreated or human IgG‐treated control cells. Similar results were also obtained from cells treated with a VEGFR2 kinase inhibitor Ki8751. In vivo, cellular senescence was detected in MIP101 tumor xenografts from 75% of mice treated with bevacizumab, while cellular senescence was undetectable in xenografts from mice treated with saline or human IgG (p < 0.05). Interestingly, we also observed that the proportion of senescent cells in colon cancer tissues obtained from patients treated with bevacizumab was 4.4‐fold higher (p < 0.01) than those of untreated patients. To understand how VEGF inhibitors may regulate cellular senescence, we noted that among the two important regulators of senescent growth arrest of tumor cells, bevacizumab‐associated increase in cellular senescence coincided with an upregulation of p16 but appeared to be independent of p53. siRNA silencing of p16 gene in MIP101 cells suppressed bevacizumab‐induced cellular senescence, while silencing of p53 had no effect. These findings demonstrate a novel antitumor activity of VEGF inhibitors in CRC, involving p16.
Cetuximab ± chemotherapy enhances dendritic cell‐mediated phagocytosis of colon cancer cells and ignites a highly efficient colon cancer antigen‐specific cytotoxic T‐cell response in vitro
Authors: Correale P. , Botta C. , Cusi M.G. , Del Vecchio M.T. , De Santi M.M. , Gori Savellini G. , Bestoso E. , Apollinari S. , Mannucci S. , Marra M. , Abbruzzese A. , Aquino A. , Turriziani M. , Bonmassar L. , Caraglia M. , Tagliaferri P.
Published: May 2011

Cetuximab is a human/mouse chimeric IgG1 monoclonal antibody (mAb) to epidermal growth factor receptor, approved for colorectal carcinoma treatment in combination with chemotherapy. The immune‐mediated effects elicited by its human fraction of crystallization moiety might critically contribute to the overall anti‐tumor effectiveness of the antibody. We therefore investigated cetuximab ability to promote colon cancer cell opsonization and phagocytosis by human dendritic cells (DCs) that are subsequently engaged in antigen‐cross presentation to cytotoxic T‐lymphocyte (CTL) precursors. Human colon cancer cell lines were evaluated for susceptibility to DC‐mediated phagocytosis before and after treatment with chemotherapy ± cetuximab in vitro. Human DCs loaded with control or drug‐treated cetuximab‐coated colon cancer cells were used to in vitro generate cytotoxic T cell clones from peripheral blood mononuclear cells of human leucocyte antigen‐A(*)02.01+ donors. T‐cell cultures were characterized for immune‐phenotype and tumor‐antigen specific CTL activity. The results confirmed that treatment of tumor cells with irinotecan + L‐folinate + 5‐flurouracil (ILF) or with gemcitabine + ILF increased tumor antigen expression. Moreover, malignant cells exposed to chemotherapy and cetuximab were highly susceptible to phagocytosis by human DCs and were able to promote their activation. The consequent DC‐mediated cross‐priming of antigens derived from mAb‐covered/drug‐treated cancer cells elicited a robust CTL anti‐tumor response. On the basis of our data, we suggest a possible involvement of CTL‐dependent immunity in cetuximab anti‐cancer effects.
MUC4: A novel prognostic factor of oral squamous cell carcinoma
Authors: Hamada Tomofumi , Wakamatsu Tsunenobu , Miyahara Mayumi , Nagata Satoshi , Nomura Masahiro , Kamikawa Yoshiaki , Yamada Norishige , Batra Surinder K. , Yonezawa Suguru , Sugihara Kazumasa
Published: May 2011

MUC4 mucin is now known to be expressed in various normal and cancer tissues. We have previously reported that MUC4 expression is a novel prognostic factor in several malignant tumors; however, it has not been investigated in oral squamous cell carcinoma (OSCC). The aim of our study is to evaluate the prognostic significance of MUC4 expression in OSCC. We examined the expression profile of MUC4 in OSCC tissues from 150 patients using immunohistochemistry. Its prognostic significance in OSCC was statistically analyzed. MUC4 was expressed in 61 of the 150 patients with OSCC. MUC4 expression was significantly correlated with higher T classification (p = 0.0004), positive nodal metastasis (p = 0.049), advanced tumor stage (p = 0.002), diffuse invasion of cancer cells (p = 0.004) and patient's death (p = 0.004) in OSCC. Multivariate analysis showed that MUC4 expression (p = 0.011), tumor location (p = 0.032) and diffuse invasion (p = 0.009) were statistically significant risk factors. Backward stepwise multivariate analysis demonstrated MUC4 expression (p = 0.0015) and diffuse invasion (p = 0.018) to be statistically significant independent risk factors of poor survival in OSCC. The disease‐free and overall survival of patients with MUC4 expression was significantly worse than those without MUC4 expression (p < 0.0001 and p = 0.0001). In addition, the MUC4 expression was a significant risk factor for local recurrence and subsequent nodal metastasis in OSCC (p = 0.017 and p = 0.0001). We first report MUC4 overexpression is an independent factor for poor prognosis of patients with OSCC; therefore, patients with OSCC showing positive MUC4 expression should be followed up carefully.
Expression and pharmacological inhibition of thymidylate synthase and Src kinase in nonsmall cell lung cancer
Authors: Ceppi Paolo , Rapa Ida , Lo Iacono Marco , Righi Luisella , Giorcelli Jessica , Pautasso Marisa , Billè Andrea , Ardissone Francesco , Papotti Mauro , Scagliotti Giorgio V.
Published: May 2011

The combination of cytotoxic chemotherapy with signaling pathway inhibitors represents a potential strategy to improve the treatment of nonsmall cell lung cancer (NSCLC). Thymidylate synthase (TS) is an enzyme essential for DNA synthesis, and its overexpression has been associated with the reduced sensitivity to antifolate agents. Src is a tyrosine kinase that modulates the cytotoxicity of cancer cells after drug treatment, and in vitro data indicate that its inhibition could revert the resistance to TS‐inhibiting drugs. Our study investigated the significance of TS and Src expression in NSCLC tissues, and the effects of their pharmacological inhibition in cell lines. In tumor and normal tissues from 94 resected NSCLC patients, TS and Src transcript levels were found positively correlated (RS = 0.66), associated with patients smoking history and overall survival. At multivariate analysis, TS gene expression was an independent prognostic factor (relative risk (RR) = 1.78, from 1.16 to 2.72; p < 0.01). Immunohistochemical detection in tumor specimens confirmed that Src kinase activation, evaluated by phospho‐specific antibody, was associated to a higher TS expression. In cell lines, dasatinib, a Src‐inhibiting agent, synergistically enhanced pemetrexed‐cytotoxicity of A549 cells, as evaluated by MTT and apoptosis assays. The biological explanation for this interaction was based on the upregulation of TS messenger RNA and protein levels induced by pemetrexed, which was significantly prevented by dasatinib cotreatment. The data of our study suggest that TS and Src may belong to a common pathway that bears prognostic significance in NSCLC, and that Src represents a potential target to improve the efficacy of TS‐inhibiting agents.
Dark cell change of the cerebellar Purkinje cells induced by terbutaline under transient disruption of the blood–brain barrier in adult rats: morphological evaluation
Authors: Yamada Naoaki , Sasaki Satoshi , Ishii Hiroyuki , Sato Junko , Kanno Takeshi , Wako Yumi , Tsuchitani Minoru
Published: May 2011

This study aimed to establish a cerebellar degeneration animal model and to characterize the dark cell change of Purkinje cells. We hypothesized that terbutaline, a β2‐adrenoceptor agonist, induces cerebellar degeneration not only in neonatal rats, but also in adult rats. Nine‐week‐old adult male Sprague–Dawley rats were anesthetized and infused with 25% mannitol via the left common carotid artery. Thirty seconds later, terbutaline was infused via the same artery. Dark‐stained Purkinje cells were observed in the entire cerebellum on day 3. Prominent Bergmann glial cells accompanied by swelling of the glial processes were present, and were closely associated with the dark‐stained Purkinje cells. These findings were found continuously throughout day 30. Ultrastructurally, dilated Golgi vesicles and/or endoplasmic reticulum and large lamella bodies were present in both severely changed and slightly changed Purkinje cells. Bergmann glial cells in the area of synaptic contacts of the severely changed Purkinje cells showed swelling. The Bergmann glial process in close contact with the slightly changed Purkinje cell dendrite in molecular layer showed slight swelling, and large lamella bodies in the dendrite were observed close to the dendritic spines. These findings may suggest that terbutaline induced a failure of Bergmann glial cell and resulted in dark cell degeneration of the Purkinje cells due to glutamate excitotoxicity. Copyright © 2011 John Wiley & Sons, Ltd.
Hepatocyte growth factor preferentially activates the anti‐inflammatory arm of NF‐κB signaling to induce A20 and protect renal proximal tubular epithelial cells from inflammation
Authors: da Silva Cleide G. , Maccariello Elizabeth R. , Wilson Szuhuei Wu , Putheti Prabhakar , Daniel Soizic , Damrauer Scott M. , Peterson Clayton R. , Siracuse Jeffrey J. , Kaczmarek Elzbieta , Ferran Christiane
Published: May 2011

Inflammation induces the NF‐κB dependent protein A20 in human renal proximal tubular epithelial cells (RPTEC), which secondarily contains inflammation by shutting down NF‐κB activation. We surmised that inducing A20 without engaging the pro‐inflammatory arm of NF‐κB could improve outcomes in kidney disease. We showed that hepatocyte growth factor (HGF) increases A20 mRNA and protein levels in RPTEC without causing inflammation. Upregulation of A20 by HGF was NF‐κB/RelA dependent as it was abolished by overexpressing IκBα or silencing p65/RelA. Unlike TNFα, HGF caused minimal IκBα and p65/RelA phosphorylation, with moderate IκBα degradation. Upstream, HGF led to robust and sustained AKT activation, which was required for p65 phosphorylation and A20 upregulation. While HGF treatment of RPTEC significantly increased A20 mRNA, it failed to induce NF‐κB dependent, pro‐inflammatory MCP‐1, VCAM‐1, and ICAM‐1 mRNA. This indicates that HGF preferentially upregulates protective (A20) over pro‐inflammatory NF‐κB dependent genes. Upregulation of A20 supported the anti‐inflammatory effects of HGF in RPTEC. HGF pretreatment significantly attenuated TNFα‐mediated increase of ICAM‐1, a finding partially reversed by silencing A20. In conclusion, this is the first demonstration that HGF activates an AKT‐p65/RelA pathway to preferentially induce A20 but not inflammatory molecules. This could be highly desirable in acute and chronic renal injury where A20‐based anti‐inflammatory therapies are beneficial. J. Cell. Physiol. 227: 1382–1390, 2012. © 2011 Wiley Periodicals, Inc.