bluuebunny/crossref_metadata_2025 huggingface dataset:
Perspectives on Ecological and Evolutionary Systems Biology
Authors: Richards Christina L. , Hanzawa Yoshie , Katari Manpreet S. , Ehrenreich Ian M. , Engelmann Kathleen E. , Purugganan Michael D.
Published: June 2017

Understanding the emergent properties inherent to genome function requires an integrated approach of data from all levels of biology. Molecular biology data alone does not describe the complex interacting functions of organisms, while studies at the level of ecological communities and ecosystems have provided little insight into the molecular underpinnings of adaptation. Merging ecology and evolution into systems biology allows researchers to exploit a wealth of genomic information by incorporating the natural phenotypic, genetic and epigenetic diversity of model systems as well as their diverse ecologies and evolutionary histories. Here, we suggest that systems biology could more fully address the question of how organisms respond to environment if studies incorporated real field settings or experimental manipulation of relevant environmental factors. In addition, although the application of genomic approaches to non‐model systems has been slow, we highlight some of the significant progress that has been made. Ecological and evolutionary systems biology will lead to a much more sophisticated understanding of the origins and functions of biological diversity, and serve as a critical component in deciphering how organisms respond to complex environments.
Effect of Nutrient Availability on Root System Development
Authors: Cruz‐Ramírez Alfredo , Calderón‐Vázquez Carlos , Herrera‐Estrella Luis
Published: June 2017

Modern‐day plant root systems have been modeled from distinct ancestors and by different evolutionary pathways influenced by environmental cues, leading to varied root system architectures. Availability of mineral nutrients has played a major role in evolution and development of the diverse root system architectures. In this chapter, we review the importance of the study of mineral nutrition on root development and describe the physiological, morphological and molecular responses of plant roots to diverse nutrient deficiencies and how the different hormone signaling pathways participate in these responses.
Biosynthesis of Plant Cell Wall and Related Polysaccharides by Enzymes of theGT2andGT48Families
Authors: Stone Bruce A. , Jacobs Andrew K. , Hrmova Maria , Burton Rachel A. , Fincher Geoffrey B.
Published: June 2017

In this chapter we outline the evolution of our understanding of the biological functions, genetics and regulation of enzymes of the GT2 and GT48 families of glycosyl transferases. The GT2 family is very large and includes enzymes encoded by the cellulose synthase gene superfamily, together with many other transferases with diverse substrate specificities that are distributed from the Archaea to humans. On the other hand, there are relatively few known members of the GT48 family, in which activities are limited to putative (1,3)‐β‐d‐glucan synthases of embryophytes, fungi and yeasts. The review is focused on a number of individual case studies, which have been chosen on the basis of their biological and economical importance in plant biology. The history of our knowledge of (1,4)‐β‐d‐glucan (cellulose) synthases and (1,3;1,4)‐β‐d‐glucan synthases in plants will be reviewed as representatives of the GT2 family, while the (1,3)‐β‐d‐glucan (or callose) synthases will be compared with these, as representatives of the GT48 family. These synthases have been extensively studied using biochemical techniques, but they are associated with membranes and are not easily purified. Emerging functional genomics technologies have been used to identify the genes encoding the cellulose synthases of the GT2 family, while new proteomics procedures have provided amino acid sequence that has in turn been applied to the identification of the GT48 (1,3)‐β‐d‐glucan synthases of plants. The enzymes are involved in the synthesis of structurally similar polysaccharides, but appear to have evolved independently.
Adaptive backstepping neural control of state‐delayed nonlinear systems with full‐state constraints and unmodeled dynamics
Authors: Zhang Tianping , Xia Meizhen , Zhu Jiaming
Published: June 2017

In this paper, the problem of adaptive neural control is discussed for a class of strict‐feedback time‐varying delays nonlinear systems with full‐state constraints and unmodeled dynamics, as well as distributed time‐varying delays. The considered nonlinear system with full‐state constraints is transformed into a nonlinear system without state constraints by introducing a one‐to‐one asymmetric nonlinear mapping. Based on modified backstepping design and using radial basis function neural networks to approximate the unknown smooth nonlinear function and using a dynamic signal to handle dynamic uncertainties, a novel adaptive backstepping control is developed for the transformed system without state constraints. The uncertain terms produced by state time delays and distributed time delays are compensated for by constructing appropriate Lyapunov‐Krasovskii functionals. All signals in the closed‐loop system are proved to be semiglobally uniformly ultimately bounded. A numerical example is provided to illustrate the effectiveness of the proposed design scheme.
Improving Mechanical Properties of cp Titanium by Heat Treatment Optimization
Authors: Wagner Francis , Ouarem Abdelouahab , Richeton Thiebaud , Toth Laszlo S.
Published: June 2017

The yield stress and the ductility are very important mechanical quantities for materials selection. The paper deals with the question: how far is it possible to increase the yield stress without a significant loss of ductility by optimizing the final heat treatment in the elaboration stage. Commercially pure titanium sheets are subjected to different thermo‐mechanical treatments to produce seven metallurgical states. The textures and the microstructures of the samples are studied by Electron Back Scattering Diffraction measurements, the mechanical behavior by tensile testing along the previous rolling, and the transverse directions of the sheet. The obtained microstructures display different grain sizes and varying fractions of recrystallized grains, together with slightly dissimilar textures. The yield stress increases with the decreasing grain size and obeys the classical Hall–Petch law. The grain size reduction results in a small decrease of ductility for extension along the rolling direction when the recrystallized volume fraction is higher than 80%. For extension along the transverse direction, however, the homogeneous deformation strongly decreases as soon as the material contains a small fraction of non‐recrystallized grains. A good compromise between high yield stress and ductility is identified in a metallurgical state close to the end of primary recrystallization. This material state insures a relatively small grain size with all grains being in a recrystallized state.
Heterogeneous Catalysis in Zeolites, Mesoporous Silica, and Metal–Organic Frameworks
Authors: Liang Jie , Liang Zibin , Zou Ruqiang , Zhao Yanli
Published: June 2017

Crystalline porous materials are important in the development of catalytic systems with high scientific and industrial impact. Zeolites, ordered mesoporous silica, and metal–organic frameworks (MOFs) are three types of porous materials that can be used as heterogeneous catalysts. This review focuses on a comparison of the catalytic activities of zeolites, mesoporous silica, and MOFs. In the first part of the review, the distinctive properties of these porous materials relevant to catalysis are discussed, and the corresponding catalytic reactions are highlighted. In the second part, the catalytic behaviors of zeolites, mesoporous silica, and MOFs in four types of general organic reactions (acid, base, oxidation, and hydrogenation) are compared. The advantages and disadvantages of each porous material for catalytic reactions are summarized. Conclusions and prospects for future development of these porous materials in this field are provided in the last section. This review aims to highlight recent research advancements in zeolites, ordered mesoporous silica, and MOFs for heterogeneous catalysis, and inspire further studies in this rapidly developing field.
Conductive Carbon Nitride for Excellent Energy Storage
Authors: Xu Jijian , Xu Feng , Qian Meng , Xu Fangfang , Hong Zhanglian , Huang Fuqiang
Published: June 2017

Conductive carbon nitride, as a hypothetical carbon material demonstrating high nitrogen doping, high electrical conductivity, and high surface area, has not been fabricated. A major challenge towards its fabrication is that high conductivity requires high temperature synthesis, but the high temperature eliminates nitrogen from carbon. Different from conventional methods, a facile preparation of conductive carbon nitride from novel thermal decomposition of nickel hydrogencyanamide in a confined space is reported. New developed nickel hydrogencyanamide is a unique precursor which provides self‐grown fragments of ⋅NCN⋅ or NCCN and conductive carbon (C‐sp2) catalyst of Ni metal during the decomposition. The final product is a tubular structure of rich mesoporous and microporous few‐layer carbon with extraordinarily high N doping level (≈15 at%) and high extent of sp2 carbon (≈65%) favoring a high conductivity (>2 S cm−1); the ultrahigh contents of nongraphitic nitrogen, redox active pyridinic N (9 at%), and pyrrolic N (5 at%), are stabilized by forming NiN bonds. The conductive carbon nitride harvests a large capacitance of 372 F g−1 with >90% initial capacitance after 10 000 cycles as a supercapacitor electrode, far exceeding the activated carbon electrodes that have <250 F g−1.
Making Copper(0) Nanoparticles in Glycerol: A Straightforward Synthesis for a Multipurpose Catalyst
Authors: Dang‐Bao Trung , Pradel Christian , Favier Isabelle , Gómez Montserrat
Published: June 2017

Small zero‐valent copper nanoparticles (CuNPs) have been straightforwardly prepared from Cu(I) and Cu(II) precursors in glycerol and in the presence of polyvinylpyrrolidone as stabilizer. Thanks to the negligible vapor pressure of the solvent, these original nano‐systems could be directly characterized in glycerol as well as in the solid state, exhibiting relevantly homogeneous colloidal dispersions, also even after catalysis. CuNPs coming from the well‐defined coordination complex di‐μ‐hydroxobis[(N,N,N′,N′‐tetramethylethylenediamine)copper(II)] chloride {[Cu(κ2‐N,N‐TMEDA)(μ‐OH)]2Cl2} have been highly efficient in C–C and C–heteroatom bond formation processes. This new catalytic system has proved its performance in C–N couplings and in the synthesis of differently substituted propargylic amines through cross‐dehydrogenative couplings, multi‐component reactions such as A3 (aldehyde‐alkyne‐amine) and KA2 (ketone‐alkyne‐amine) couplings, as well as in the formation of heterocycles such as benzofurans, indolizines, and quinolines under smooth conditions. No significant copper amount was detected in the extracted organic compounds from the catalytic phase by inductively coupled plasma‐atomic emission spectroscopic (ICP‐AES) analyses, proving a highly efficient immobilization of copper nanoparticles in glycerol. From a mechanistic point of view, spectroscopic data (infrared and ultraviolet‐visible spectra) agree with a surface‐like catalytic reactivity.

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Comparative performance of deciduous and permanent dental morphology in detecting biological relatives
Authors: Paul Kathleen S. , Stojanowski Christopher M.
Published: June 2017

Objectives

Dental morphology plays a key role in reconstructing population history and evolutionary relationships at global, regional, and intracemetery scales. At the inter‐individual level, it is assumed that close biological kin exhibit greater phenotypic similarity than non‐relatives. Heritability estimates provide one measure of phenotypic resemblance but are not easily incorporated into analyses of archaeological samples. In this study we evaluate the assumption that relatives are more similar phenotypically than non‐relatives. We compare results for permanent dental morphology to those obtained using deciduous dental morphology in a matched dataset (Paul & Stojanowski,).

Materials and Methods

Permanent trait expression was scored from dental casts representing 69 sibling pairs, curated as part of the longitudinal Burlington Growth Study. Simulating a biodistance approach, 22 morphological traits of permanent tooth crowns were used to generate 69 inter‐relative and 2,076 non‐relative Euclidean distances. Following distance ordination, family‐specific dispersion values were calculated from multidimensional scaling coordinates. Output was compared to that of a previous study that focused on deciduous crown variation in the same set of individuals (Paul & Stojanowski,). Mantel tests were used to evaluate the correlation of a proxy genetic distance matrix to both the permanent and deciduous dental distance matrices.

Results

On average, inter‐relative distances generated from morphological traits of permanent tooth crowns were smaller than expected by chance based on resampling (p < 0.001). While family‐specific dispersion varied greatly for both permanent and deciduous datasets, over 75% of the families exhibited greater dispersion with permanent traits. This suggests that morphological traits of the permanent dentition provide a less faithful reflection of biological relatedness than morphological traits of the deciduous dentition. Mantel tests indicate that both the deciduous and permanent distance matrices are significantly correlated with a matrix of genetic relatedness coefficients; however, the magnitude of the correlations was low.

Discussion

Overall, morphological traits of permanent tooth crowns perform moderately well in distinguishing relatives from non‐relatives, but deciduous crown variations may provide a more direct reflection of the underlying genetic structure of intra‐site or intra‐cemetery samples. These findings have implications for bioarchaeological research and biodistance practices. In particular, morphological traits of the deciduous dentition should be incorporated into standard data collection protocols because of their stronger signal of relatedness.
Gene therapy decreases seizures in a model of Incontinentia pigmenti
Authors: Dogbevia Godwin K. , Töllner Kathrin , Körbelin Jakob , Bröer Sonja , Ridder Dirk A. , Grasshoff Hanna , Brandt Claudia , Wenzel Jan , Straub Beate K. , Trepel Martin , Löscher Wolfgang , Schwaninger Markus
Published: June 2017

Objective

Incontinentia pigmenti (IP) is a genetic disease leading to severe neurological symptoms, such as epileptic seizures, but no specific treatment is available. IP is caused by pathogenic variants that inactivate the Nemo gene. Replacing Nemo through gene therapy might provide therapeutic benefits.

Methods

In a mouse model of IP, we administered a single intravenous dose of the adeno‐associated virus (AAV) vector, AAV‐BR1‐CAG‐NEMO, delivering the Nemo gene to the brain endothelium. Spontaneous epileptic seizures and the integrity of the blood–brain barrier (BBB) were monitored.

Results

The endothelium‐targeted gene therapy improved the integrity of the BBB. In parallel, it reduced the incidence of seizures and delayed their occurrence. Neonate mice intravenously injected with the AAV‐BR1‐CAG‐NEMO vector developed no hepatocellular carcinoma or other major adverse effects 11 months after vector injection, demonstrating that the vector has a favorable safety profile.

Interpretation

The data show that the BBB is a target of antiepileptic treatment and, more specifically, provide evidence for the therapeutic benefit of a brain endothelial‐targeted gene therapy in IP. Ann Neurol 2017;82:93–104
A novel cause of chronic viral meningoencephalitis: Cache Valley virus
Authors: Wilson Michael R. , Suan Dan , Duggins Andrew , Schubert Ryan D. , Khan Lillian M. , Sample Hannah A. , Zorn Kelsey C. , Rodrigues Hoffman Aline , Blick Anna , Shingde Meena , DeRisi Joseph L.
Published: June 2017

Objective

Immunodeficient patients are particularly vulnerable to neuroinvasive infections that can be challenging to diagnose. Metagenomic next generation sequencing can identify unusual or novel microbes and is therefore well suited for investigating the etiology of chronic meningoencephalitis in immunodeficient patients.

Methods

We present the case of a 34‐year‐old man with X‐linked agammaglobulinemia from Australia suffering from 3 years of meningoencephalitis that defied an etiologic diagnosis despite extensive conventional testing, including a brain biopsy. Metagenomic next generation sequencing of his cerebrospinal fluid and brain biopsy tissue was performed to identify a causative pathogen.

Results

Sequences aligning to multiple Cache Valley virus genes were identified via metagenomic next generation sequencing. Reverse transcription polymerase chain reaction and immunohistochemistry subsequently confirmed the presence of Cache Valley virus in the brain biopsy tissue.

Interpretation

Cache Valley virus, a mosquito‐borne orthobunyavirus, has only been identified in 3 immunocompetent North American patients with acute neuroinvasive disease. The reported severity ranges from a self‐limiting meningitis to a rapidly fatal meningoencephalitis with multiorgan failure. The virus has never been known to cause a chronic systemic or neurologic infection in humans. Cache Valley virus has also never previously been detected on the Australian continent. Our research subject traveled to North and South Carolina and Michigan in the weeks prior to the onset of his illness. This report demonstrates that metagenomic next generation sequencing allows for unbiased pathogen identification, the early detection of emerging viruses as they spread to new locales, and the discovery of novel disease phenotypes. Ann Neurol 2017;82:105–114
Optoelektrochemische In‐situ‐Beobachtung der kathodischen Abscheidung einzelner Cobaltnanopartikel
Authors: Brasiliense Vitor , Clausmeyer Jan , Dauphin Alice L. , Noël Jean‐Marc , Berto Pascal , Tessier Gilles , Schuhmann Wolfgang , Kanoufi Fréderic
Published: June 2017

Die Elektrochemie einzelner Partikel auf einer Nanoelektrode wird mittels optischer Dunkelfeldmikroskopie untersucht. Die Betrachtung des gestreuten Lichts macht es möglich, die elektrochemische Abscheidung einzelner Cobaltnanopartikel mit Radien bis hinunter zu 65 nm in situ zu verfolgen. Die Dimensionen von größeren Submikropartikeln können durch Superlokalisierung der Partikelgrenzen bestimmt werden. Zusätzlich enthält das gestreute Licht ergänzende Informationen über die komplexe Redoxchemie. Mit der optoelektrochemischen Technik werden wertvolle Einblicke in elektrokatalytische Prozesse während der Abscheidung gewonnen, die durch elektrochemische Messungen an einzelnen Partikeln allein nicht zugänglich wären.
Protonierung von Nitraminen: Bildung des O‐ oder N‐protonierten Kations
Authors: Saal Thomas , Rahm Martin , Christe Karl O. , Haiges Ralf
Published: June 2017

Die Reaktionen von Nitramin, N‐Methylnitramin und N,N‐Dimethylnitramin mit wasserfreiem HF und den binären supersauren Systemen HF/MF5 (M=As, Sb) wurden bei Temperaturen unterhalb von −40 °C untersucht. Mittels multinuklearer NMR‐Spektroskopie konnte in Lösung ausschließlich die O‐Protonierung beobachtet werden, wobei aus reiner HF‐Lösung selbst bei −78 °C kein Feststoff isoliert werden konnte. Im HF/MF5‐System konnte jedoch protoniertes Nitramin zum ersten Mal in Form feuchtigkeitsempfindlicher MF6−‐Salze isoliert werden, die sich bei Temperaturen oberhalb von −40 °C zersetzen. Im Feststoff resultiert abhängig vom Gegenion ein O‐protoniertes oder N‐protoniertes Kation. Dies ist in Einklang mit Rechnungen, die einen nur minimalen Energieunterschied zwischen O‐Protonierung und N‐Protonierung aufzeigen. Die Salze [H2N‐NO2H][AsF6], [H3N‐NO2][SbF6], [MeHNNO2H][SbF6] und [Me2NNO2H][SbF6] wurden durch ihre Röntgenkristallstrukturen charakterisiert.
From Lithium‐Ion to Sodium‐Ion Batteries: Advantages, Challenges, and Surprises
Authors: Nayak Prasant Kumar , Yang Liangtao , Brehm Wolfgang , Adelhelm Philipp
Published: June 2017

Mobile and stationary energy storage by rechargeable batteries is a topic of broad societal and economical relevance. Lithium‐ion battery (LIB) technology is at the forefront of the development, but a massively growing market will likely put severe pressure on resources and supply chains. Recently, sodium‐ion batteries (SIBs) have been reconsidered with the aim of providing a lower‐cost alternative that is less susceptible to resource and supply risks. On paper, the replacement of lithium by sodium in a battery seems straightforward at first, but unpredictable surprises are often found in practice. What happens when replacing lithium by sodium in electrode reactions? This review provides a state‐of‐the art overview on the redox behavior of materials when used as electrodes in lithium‐ion and sodium‐ion batteries, respectively. Advantages and challenges related to the use of sodium instead of lithium are discussed.
γ‐Functionalizations of Amines through Visible‐Light‐Mediated, Redox‐Neutral C−C Bond Cleavage
Authors: Shu Wei , Genoux Alexandre , Li Zhaodong , Nevado Cristina
Published: June 2017

Cleavage of unstrained C−C bonds under mild, redox‐neutral conditions represents a challenging endeavor which is accomplished here in the context of a flexible, visible‐light‐mediated, γ‐functionalization of amines. In situ generated C‐centered radicals are harvested in the presence of Michael acceptors, thiols and alkyl halides to efficiently form new C(sp3)−C(sp3), C(sp3)−H and C(sp3)−Br bonds, respectively.
A statistical approach to develop biocomposites from epoxy resin, poly(furfuryl alcohol), poly(propylene carbonate), and biochar
Authors: Mashouf Roudsari Ghodsieh , Mohanty Amar K. , Misra Manjusri
Published: June 2017

Epoxy composites are typically petroleum based and prone to fracture. Increasing concerns about climate change have motivated scientists to find green alternatives. To address both drawbacks, effect of addition of poly(propylene carbonate) polyol (PPC), a polyol derived from carbon dioxide, and biochar, a byproduct of pyrolysis in an epoxy/poly(furfuryl alcohol) (PFA) network was studied. It is hypothesized that addition of PPC will increase impact strength while addition of inexpensive biochar will offset cost of final product. In addition, incorporation of PFA, PPC, and biochar can increase biobased content of thermoset. A statistical approach was used to find a systematic correlation between constituent contents and mechanical properties of biocomposites. Mixture design of experiment and backward elimination regression were used to model mechanical properties of biocomposites. The fitted models showed a great ability to predict mechanical properties of new formulations. Addition of 10% biochar increased tensile strength and toughness by 13% and 34%, respectively. Biochar also increased modulus while it had adverse effect on impact strength. Promising effect of PPC on toughening of matrix was proved and it was found that addition of 30% of PPC increased impact strength by fivefolds. © 2017 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2017, 134, 45307.
Cranial Indicators Identified for Peak Incidence of Otitis Media
Authors: Pagano Anthony S. , Wang Eugene , Yuan Derek , Fischer Daniel , Bluestone Charles , Marquez Samuel , Laitman Jeffrey
Published: June 2017

Acute otitis media (AOM) is one of the most common pediatric conditions worldwide. Peak age of occurrence for AOM has been identified within the first postnatal year and it remains frequent until approximately six postnatal years. Morphological differences between adults and infants in the cartilaginous Eustachian tube (CET) and associated structures may be responsible for development of this disease yet few have investigated normal growth trajectories. We tested hypotheses on coincidence of skeletal growth changes and known ages of peak AOM occurrence. Growth was divided into five dental eruption stages ranging from edentulous neonates (Stage 1) to adults with erupted third maxillary molars (Stage 5). A total of 32 three‐dimensional landmarks were used and Generalized Procrustes Analysis was performed. Next, we performed principal components analysis and calculated univariate measures. It was found that growth change in Stage 1 was the most rapid and comprised the largest amount of overall growth in upper respiratory tract proportions (where time is represented by the natural logarithmic transformation of centroid size). The analysis of univariate measures showed that Stage 1 humans did indeed possess the relatively shortest and most horizontally oriented CET's with the greatest amount of growth change occurring at the transition to Stage 2 (eruption of deciduous dentition at five postnatal months, commencing peak AOM incidence) and ceasing by Stage 3 (approximately six postnatal years). Skeletal indicators appear related to peak ages of AOM incidence and may contribute to understanding of a nearly ubiquitous human disease. Anat Rec, 2017. © 2017 Wiley Periodicals, Inc. Anat Rec, 300:1721–1740, 2017. © 2017 Wiley Periodicals, Inc.
Multivariate binary classification of imbalanced datasets—A case study based on high‐dimensional multiplex autoimmune assay data
Authors: Schlieker Laura , Telaar Anna , Lueking Angelika , Schulz‐Knappe Peter , Theek Carmen , Ickstadt Katja
Published: June 2017

The classification of a population by a specific trait is a major task in medicine, for example when in a diagnostic setting groups of patients with specific diseases are identified, but also when in predictive medicine a group of patients is classified into specific disease severity classes that might profit from different treatments. When the sizes of those subgroups become small, for example in rare diseases, imbalances between the classes are more the rule than the exception and make statistical classification problematic when the error rate of the minority class is high. Many observations are classified as belonging to the majority class, while the error rate of the majority class is low. This case study aims to investigate class imbalance for Random Forests and Powered Partial Least Squares Discriminant Analysis (PPLS‐DA) and to evaluate the performance of these classifiers when they are combined with methods to compensate imbalance (sampling methods, cost‐sensitive learning approaches). We evaluate all approaches with a scoring system taking the classification results into consideration. This case study is based on one high‐dimensional multiplex autoimmune assay dataset describing immune response to antigens and consisting of two classes of patients: Rheumatoid Arthritis (RA) and Systemic Lupus Erythemathodes (SLE). Datasets with varying degrees of imbalance are created by successively reducing the class of RA patients. Our results indicate possible benefit of cost‐sensitive learning approaches for Random Forests. Although further research is needed to verify our findings by investigating other datasets or large‐scale simulation studies, we claim that this work has the potential to increase awareness of practitioners to this problem of class imbalance and stresses the importance of considering methods to compensate class imbalance.
Role of cytoskeletal mechanics and cell membrane fluidity in the intracellular delivery of molecules mediated by laser‐activated carbon nanoparticles
Authors: Holguin Stefany Y. , Anderson Caleb F. , Thadhani Naresh N. , Prausnitz Mark R.
Published: June 2017

Exposure of cells and nanoparticles to near‐infrared nanosecond pulsed laser light can lead to efficient intracellular delivery of molecules while maintaining high cell viability by a photoacoustic phenomenon known as transient nanoparticle energy transduction (TNET). Here, we examined the influence of cytoskeletal mechanics and plasma membrane fluidity on intracellular uptake of molecules and loss of cell viability due to TNET. We found that destabilization of actin filaments using latrunculin A led to greater uptake of molecules and less viability loss caused by TNET. Stabilization of actin filaments using jasplakinolide had no significant effect on uptake or viability loss caused by TNET. To study the role of plasma membrane fluidity, we increased fluidity by depletion of membrane cholesterol using methyl‐β‐cyclodextrin and decreased fluidity by enrichment of the membrane with cholesterol using water‐soluble cholesterol. Neither of these membrane fluidity changes significantly altered cellular uptake or viability loss caused by TNET. We conclude that weakening mechanical integrity of the cytoskeleton can increase intracellular uptake and decrease loss of cell viability, while plasma membrane fluidity does not appear to play a significant role in uptake or viability loss caused by TNET. The positive effects of cytoskeletal weakening may be due to an enhanced ability of the cell to recover from the effects of TNET and maintain viability. Biotechnol. Bioeng. 2017;114: 2390–2399. © 2017 Wiley Periodicals, Inc.
Use of a protein engineering strategy to overcome limitations in the production of “Difficult to Express” recombinant proteins
Authors: Hussain Hirra , Fisher David I. , Abbott W. Mark , Roth Robert G. , Dickson Alan J.
Published: June 2017

Certain recombinant proteins are deemed “difficult to express” in mammalian expression systems requiring significant cell and/or process engineering to abrogate expression bottlenecks. With increasing demand for the production of recombinant proteins in mammalian cells, low protein yields can have significant consequences for industrial processes. To investigate the molecular mechanisms that restrict expression of recombinant proteins, naturally secreted model proteins were analyzed from the tissue inhibitors of metalloproteinase (TIMP) protein family. In particular, TIMP‐2 and TIMP‐3 were subjected to detailed study. TIMP proteins share significant sequence homology (∼50% identity and ∼70% similarity in amino acid sequence). However, they show marked differences in secretion in mammalian expression systems despite this extensive sequence homology. Using these two proteins as models, this study characterized the molecular mechanisms responsible for poor recombinant protein production. Our results reveal that both TIMP‐2 and TIMP‐3 are detectable at mRNA and protein level within the cell but only TIMP‐2 is secreted effectively into the extracellular medium. Analysis of protein localization and the nature of intracellular protein suggest TIMP‐3 is severely limited in its post‐translational processing. To overcome this challenge, modification of the TIMP‐3 sequence to include a furin protease‐cleavable pro‐sequence resulted in secretion of the modified TIMP‐3 protein, however, incomplete processing was observed. Based on the TIMP‐3 data, the protein engineering approach was optimized and successfully applied in combination with cell engineering, the overexpression of furin, to another member of the TIMP protein family (the poorly expressed TIMP‐4). Use of the described protein engineering strategy resulted in successful secretion of poorly (TIMP‐4) and non‐secreted (TIMP‐3) targets, and presents a novel strategy to enhance the production of “difficult” recombinant targets. Biotechnol. Bioeng. 2017;114: 2348–2359. © 2017 Wiley Periodicals, Inc.
Proteomic analysis of host cell protein dynamics in the supernatant of Fc‐fusion protein‐producing CHO DG44 and DUKX‐B11 cell lines in batch and fed‐batch cultures
Authors: Park Jin Hyoung , Jin Jong Hwa , Ji In Jung , An Hyun Joo , Kim Jong Won , Lee Gyun Min
Published: June 2017

Chinese hamster ovary (CHO) cells are the most widely used host cell lines for the commercial production of therapeutic proteins including Fc‐fusion proteins. During the culture of recombinant CHO (rCHO) cells, host cell proteins (HCPs), secreted from viable cells and released from dead cells, accumulate extracellularly, potentially impairing product quality. In this study, the HCPs that accumulated extracellularly in batch and fed‐batch cultures of Fc‐fusion protein‐producing rCHO cell lines (DG‐Fc and DUKX‐Fc) were identified and quantified using nanoflow liquid chromatography–tandem mass spectrometry (LC–MS/MS), followed by gene ontology and functional analysis. When the proteome database of Cricetulus griseus was used as a reference to identify the HCPs, more HCPs were identified for DG‐Fc (1632 HCPs in batch culture and 1733 HCPs in fed‐batch culture) than for DUKX‐Fc (1114 HCPs in batch culture and 1002 HCPs in fed‐batch culture). Clustering analysis of HCPs, which were classified into four clusters according to their concentration profiles during culture, showed that the concentration profiles of HCPs affecting the quality of Fc‐fusion proteins correlated with changes in Fc‐fusion protein quality. Taken together, the dataset of HCPs obtained in this study using the two different rCHO cell lines provides insights into the determination of appropriate target proteins to be removed during the culture and purification steps so as to ensure good Fc‐fusion protein quality. Biotechnol. Bioeng. 2017;114: 2267–2278. © 2017 Wiley Periodicals, Inc.
Enzyme‐free cell detachment mediated by resonance vibration with temperature modulation
Authors: Kurashina Yuta , Hirano Makoto , Imashiro Chikahiro , Totani Kiichiro , Komotori Jun , Takemura Kenjiro
Published: June 2017

Cell detachment is an essential process in adherent cell culture. However, trypsinization, which is the most popular detachment technique used in culture, damages cellular membranes. Reducing cellular membrane damage during detachment should improve the quality of cell culture. In this article, we propose an enzyme‐free cell detachment method based on resonance vibration with temperature modulation. We developed a culture device that can excite a resonance vibration and control temperature. We then evaluated the cell detachment ratio and the growth response, observed the morphology, and analyzed the cellular protein of the collected cells—mouse myoblast cell line (C2C12). With the temperature of 10°C and the maximum vibration amplitude of 2 μm, 77.9% of cells in number were successfully detached compared with traditional trypsinization. The 72‐h proliferation ratio of the reseeded cells was similar to that with trypsinization, whereas the proliferation ratio of proposed method was 12.6% greater than that of trypsinization after freezing and thawing. Moreover, the cells can be collected relatively intact and both intracellular and cell surface proteins in the proposed method were less damaged than in trypsinization. These results show that this method has definite advantages over trypsinization, which indicates that it could be applied to subcultures of cells that are more susceptible to trypsin damage for mass culture of sustainable clinical use. Biotechnol. Bioeng. 2017;114: 2279–2288. © 2017 Wiley Periodicals, Inc.
Meta-analysis of determinants of survival following treatment of recurrent hepatocellular carcinoma
Authors: Erridge S , Pucher P H , Markar S R , Malietzis G , Athanasiou T , Darzi A , Sodergren M H , Jiao L R
Published: June 2017

Background

Intrahepatic recurrence of hepatocellular carcinoma (HCC) following resection is common. However, no current consensus guidelines exist to inform management decisions in these patients. Systematic review and meta-analysis of survival following different treatment modalities may allow improved treatment selection. This review aimed to identify the optimum treatment strategies for HCC recurrence.

Methods

A systematic review, up to September 2016, was conducted in accordance with MOOSE guidelines. The primary outcome was the hazard ratio for overall survival of different treatment modalities. Meta-analysis of different treatment modalities was carried out using a random-effects model, with further assessment of additional prognostic factors for survival.

Results

Nineteen cohort studies (2764 patients) were included in final data analysis. The median 5-year survival rates after repeat hepatectomy (525 patients), ablation (658) and transarterial chemoembolization (TACE) (855) were 35·2, 48·3 and 15·5 per cent respectively. Pooled analysis of ten studies demonstrated no significant difference between overall survival after ablation versus repeat hepatectomy (hazard ratio 1·03, 95 per cent c.i. 0·68 to 1·55; P = 0·897). Pooled analysis of seven studies comparing TACE with repeat hepatectomy showed no statistically significant difference in survival (hazard ratio 1·61, 0·99 to 2·63; P = 0·056).

Conclusion

Based on these limited data, there does not appear to be a significant difference in survival between patients undergoing repeat hepatectomy or ablation for recurrent HCC. The results also identify important negative prognostic factors (short disease-free interval, multiple hepatic metastases and large hepatic metastases), which may influence choice of treatment.
Amplicon Competition Enables End‐Point Quantitation of Nucleic Acids Following Isothermal Amplification
Authors: Jiang Yu Sherry , Stacy Apollo , Whiteley Marvin , Ellington Andrew D. , Bhadra Sanchita
Published: June 2017

It is inherently difficult to quantitate nucleic acid analytes with most isothermal amplification assays. We developed loop‐mediated isothermal amplification (LAMP) reactions in which competition between defined numbers of “false” and “true” amplicons leads to order of magnitude quantitation by a single endpoint determination. These thresholded LAMP reactions were successfully used to directly and quantitatively estimate the numbers of nucleic acids in complex biospecimens, including directly from cells and in sewage, with the values obtained closely correlating with qPCR quantitations. Thresholded LAMP reactions are amenable to endpoint readout by cell phone, unlike other methods that require continuous monitoring, and should therefore prove extremely useful in developing one‐pot reactions for point‐of‐care diagnostics without needing sophisticated material or informatics infrastructure.
DFT Studies of Dimerization Reactions of Boroles
Authors: Wang Zheng , Zhou Yu , Lee Ka‐Ho , Lam Wai Han , Dewhurst Rian D. , Braunschweig Holger , Marder Todd B. , Lin Zhenyang
Published: June 2017

Boroles undergo dimerization reactions to give Diels–Alder (DA) dimers, bridged‐bicyclic (BB) dimers or spiro dimers (SD) depending on the substituents on the borole. We performed DFT calculations to investigate how different substituents at the carbon atoms of the butadiene backbone as well as at the boron atom influence the dimerization reaction pathways. The DFT results show that, in general, both the DA and BB dimers are easily accessible kinetically, and the DA dimers are thermodynamically more stable than the BB dimers. When the substituent–substituent repulsive steric interactions are alleviated to a certain extent, the BB dimers are more stable than the DA dimer, and become accessible. The SD dimers are generally kinetically difficult to obtain. However, we found that aryl substituents promote the formation of the SD dimers.
Synthesis of gem‐Dinitromethylated and Fluorodinitromethylated Derivatives of 5,5′‐Dinitro‐bis‐1,2,4‐triazole as Promising High‐Energy‐Density Materials
Authors: Huang Shi , Tian Junjun , Qi Xiujuan , Wang Kangcai , Zhang Qinghua
Published: June 2017

gem‐Dinitromethylated and fluorodinitromethylated 5,5′‐dinitro‐3,3′‐bis‐1,2,4‐triazole (DNBT) (2 and 3) along with seven ionic derivatives 4–9, were synthesized and characterized by NMR and IR spectroscopies, elemental analysis, single‐crystal X‐ray diffraction (XRD), and differential scanning calorimetry (DSC). XRD revealed that compounds 2 and 3 crystallized in the monoclinic P21/n space group and compound 5 crystallized in the monoclinic P21/c space group. The physicochemical properties of the as‐synthesized compounds 2–9 were investigated and the results indicated that compounds 3, 6, and 7 exhibited a good balance between high energy and low sensitivity, demonstrating their potential as new high‐energy‐density materials (HEDMs). The proposed synthetic strategy for introducing gem‐dinitromethyl and fluorodinitromethyl groups into the DNBT framework to prepare materials with high energy and low sensitivity suitable for HEDMs looks promising.
Synthesis and Characterization of Heterobenzenacyclo‐octaphanes Derived from Cyclotetrabenzoin
Authors: Hahn Sebastian , Alrayyani Maymounah , Sontheim Anna , Wang Xiqu , Rominger Frank , Miljanić Ognjen Š. , Bunz Uwe H. F.
Published: June 2017

Invited for the cover of this issue is the groups of Ognjen Š. Miljanić at the University of Houston (United States) and Uwe H. F. Bunz at the Ruprecht‐Karls‐University of Heidelberg (Germany). The image depicts the versatility of our system, demonstrated by single crystal structures of our target molecules with different substitution and functionalities. Read the full text of the article at 10.1002/chem.201701125.
Transition and Alkali Metal Complex Ternary Amides for Ammonia Synthesis and Decomposition
Authors: Cao Hujun , Guo Jianping , Chang Fei , Pistidda Claudio , Zhou Wei , Zhang Xilun , Santoru Antonio , Wu Hui , Schell Norbert , Niewa Rainer , Chen Ping , Klassen Thomas , Dornheim Martin
Published: June 2017

A new complex ternary amide, Rb2[Mn(NH2)4], which simultaneously contains both transition and alkali metal catalytic sites, is developed. This is in line with the recently reported TM‐LiH composite catalysts, which have been shown to effectively break the scaling relations and achieve ammonia synthesis under mild conditions. Rb2[Mn(NH2)4] can be facilely synthesized by mechanochemical reaction at room temperature. It exhibits two temperature‐dependent polymorphs, that is, a low‐temperature orthorhombic and a high‐temperature monoclinic structure. Rb2[Mn(NH2)4] decomposes to N2, H2, NH3, Mn3N2, and RbNH2 under inert atmosphere; whereas it releases NH3 at a temperature as low as 80 °C under H2 atmosphere. Those unique behaviors enable Rb2[Mn(NH2)4], and its analogue K2[Mn(NH2)4], to be excellent catalytic materials for ammonia decomposition and synthesis. Experimental results show both ammonia decomposition onset temperatures and conversion rates over Rb2[Mn(NH2)4] and K2[Mn(NH2)4] are similar to those of noble metal Ru‐based catalysts. More importantly, these ternary amides exhibit superior capabilities in catalyzing NH3 synthesis, which are more than 3 orders of magnitude higher than that of Mn nitride and twice of that of Ru/MgO. The in situ SR‐PXD measurement shows that manganese nitride, synergistic with Rb/KH or Rb/K(NH2)xH1−x, are likely the active sites. The chemistry of Rb2/K2[Mn(NH2)x] and Rb/K(NH2)xH1−x with H2/N2 and NH3 correlates closely with the catalytic performance.
A Simple Route to Strong Carbon‐13 NMR Signals Detectable for Several Minutes
Authors: Roy Soumya S. , Norcott Philip , Rayner Peter J. , Green Gary G. R. , Duckett Simon B.
Published: June 2017

Nuclear magnetic resonance (NMR) and magnetic resonance imaging (MRI) suffer from low sensitivity and limited nuclear spin memory lifetimes. Although hyperpolarization techniques increase sensitivity, there is also a desire to increase relaxation times to expand the range of applications addressable by these methods. Here, we demonstrate a route to create hyperpolarized magnetization in 13C nuclear spin pairs that last much longer than normal lifetimes by storage in a singlet state. By combining molecular design and low‐field storage with para‐hydrogen derived hyperpolarization, we achieve more than three orders of signal amplification relative to equilibrium Zeeman polarization and an order of magnitude extension in state lifetime. These studies use a range of specifically synthesized pyridazine derivatives and dimethyl p‐tolyl phenyl pyridazine is the most successful, achieving a lifetime of about 190 s in low‐field, which leads to a 13C‐signal that is visible for 10 minutes.
Application of SWATH Proteomics to Mouse Biology
Authors: Wu Yibo , Williams Evan G. , Aebersold Ruedi
Published: June 2017

The quantitative measurement of the proteome has been shown to yield new insights into physiology and cell biology that cannot be determined from the genome and transcriptome because the quantitative relationship between transcriptome and proteome is complex. MS‐based proteomics techniques, such as SWATH‐MS, have recently advanced to the point at which they may be reliably applied by biologists who are not specialists in mass spectrometry. Here we provide standard protocols for preparation of tissue samples for input into the SWATH‐MS analytical pipeline. These protocols are designed for high‐throughput processing of tissues with ≥5 mg of sample available for analysis. Studies with extremely limited amounts of tissue should consider PCT‐SWATH. An experienced single user should be able to process 48 samples per day for injection into the mass spectrometer, or up to 144 samples a week. The machine time necessary for running these samples with SWATH is approximately 1.5 hr per sample. Data acquisition protocols are also provided. © 2017 by John Wiley & Sons, Inc.
A High‐Throughput Process for the Solid‐Phase Purification of Synthetic DNA Sequences
Authors: Grajkowski Andrzej , Cieślak Jacek , Beaucage Serge L.
Published: June 2017

An efficient process for the purification of synthetic phosphorothioate and native DNA sequences is presented. The process is based on the use of an aminopropylated silica gel support functionalized with aminooxyalkyl functions to enable capture of DNA sequences through an oximation reaction with the keto function of a linker conjugated to the 5′‐terminus of DNA sequences. Deoxyribonucleoside phosphoramidites carrying this linker, as a 5′‐hydroxyl protecting group, have been synthesized for incorporation into DNA sequences during the last coupling step of a standard solid‐phase synthesis protocol executed on a controlled pore glass (CPG) support. Solid‐phase capture of the nucleobase‐ and phosphate‐deprotected DNA sequences released from the CPG support is demonstrated to proceed near quantitatively. Shorter than full‐length DNA sequences are first washed away from the capture support; the solid‐phase purified DNA sequences are then released from this support upon reaction with tetra‐n‐butylammonium fluoride in dry dimethylsulfoxide (DMSO) and precipitated in tetrahydrofuran (THF). The purity of solid‐phase‐purified DNA sequences exceeds 98%. The simulated high‐throughput and scalability features of the solid‐phase purification process are demonstrated without sacrificing purity of the DNA sequences. © 2017 by John Wiley & Sons, Inc.
Label‐Free Dynamic Mass Redistribution and Bio‐Impedance Methods for Drug Discovery
Authors: Grundmann Manuel
Published: June 2017

Label‐free biosensors are increasingly employed in drug discovery. Cell‐based biosensors provide valuable insights into the biological consequences of exposing cells and tissues to chemical agents and the underlying molecular mechanisms associated with these effects. Optical biosensors based on the detection of dynamic mass redistribution (DMR) and impedance biosensors using cellular dielectric spectroscopy (CDS) capture changes of the cytoskeleton of living cells in real time. Because signal transduction correlates with changes in cell morphology, DMR and CDS biosensors are exquisitely suited for recording integrated cell responses in an unbiased, yet pathway‐specific manner without the use of labels that may interfere with cell function. Described in this unit are several experimental approaches utilizing optical label‐free system capturing dynamic mass redistribution (DMR) in living cells (Epic System) and an impedance‐based CDS technology (CellKey). In addition, potential pitfalls associated with these assays and alternative approaches for overcoming such technical challenges are discussed. © 2017 by John Wiley & Sons, Inc.