bluuebunny/crossref_metadata_2025 huggingface dataset:
A Mutant Influenza Virus That Uses an N1 Neuraminidase as the Receptor-Binding Protein
Authors: Hooper Kathryn A. , Bloom Jesse D.
Published: September 2013

In the vast majority of influenza A viruses characterized to date, hemagglutinin (HA) is the receptor-binding and fusion protein, whereas neuraminidase (NA) is a receptor-cleaving protein that facilitates viral release but is expendable for entry. However, the NAs of some recent human H3N2 isolates have acquired receptor-binding activity via the mutation D151G, although these isolates also appear to retain the ability to bind receptors via HA. We report here the laboratory generation of a mutation (G147R) that enables an N1 NA to completely co-opt the receptor-binding function normally performed by HA. Viruses with this mutant NA grow to high titers even in the presence of extensive mutations to conserved residues in HA's receptor-binding pocket. When the receptor-binding NA is paired with this binding-deficient HA, viral infectivity and red blood cell agglutination are blocked by NA inhibitors. Furthermore, virus-like particles expressing only the receptor-binding NA agglutinate red blood cells in an NA-dependent manner. Although the G147R NA receptor-binding mutant virus that we characterize is a laboratory creation, this same mutation is found in several natural clusters of H1N1 and H5N1 viruses. Our results demonstrate that, at least in tissue culture, influenza virus receptor-binding activity can be entirely shifted from HA to NA.
Middle East Respiratory Syndrome Coronavirus Infection Mediated by the Transmembrane Serine Protease TMPRSS2
Authors: Shirato Kazuya , Kawase Miyuki , Matsuyama Shutoku
Published: September 2013

The Middle East respiratory syndrome coronavirus (MERS-CoV) utilizes host proteases for virus entry into lung cells. In the current study, Vero cells constitutively expressing type II transmembrane serine protease (Vero-TMPRSS2 cells) showed larger syncytia at 18 h after infection with MERS-CoV than after infection with other coronaviruses. Furthermore, the susceptibility of Vero-TMPRSS2 cells to MERS-CoV was 100-fold higher than that of non-TMPRSS2-expressing parental Vero cells. The serine protease inhibitor camostat, which inhibits TMPRSS2 activity, completely blocked syncytium formation but only partially blocked virus entry into Vero-TMPRSS2 cells. Importantly, the coronavirus is thought to enter cells via two distinct pathways, one mediated by TMPRSS2 at the cell surface and the other mediated by cathepsin L in the endosome. Simultaneous treatment with inhibitors of cathepsin L and TMPRSS2 completely blocked virus entry into Vero-TMPRSS2 cells, indicating that MERS-CoV employs both the cell surface and the endosomal pathway to infect Vero-TMPRSS2 cells. In contrast, a single camostat treatment suppressed MERS-CoV entry into human bronchial submucosal gland-derived Calu-3 cells by 10-fold and virus growth by 270-fold, although treatment with both camostat and (23,25)-
trans
-epoxysuccinyl-
l
-leucylamindo-3-methylbutane ethyl ester, a cathepsin inhibitor, or treatment with leupeptin, an inhibitor of cysteine, serine, and threonine peptidases, was no more efficacious than treatment with camostat alone. Further, these inhibitors were not efficacious against MERS-CoV infection of MRC-5 and WI-38 cells, which were derived from lung, but these characters differed from those of mature pneumocytes. These results suggest that a single treatment with camostat is sufficient to block MERS-CoV entry into a well-differentiated lung-derived cell line.
Characterization of a Novel G3P[3] Rotavirus Isolated from a Lesser Horseshoe Bat: a Distant Relative of Feline/Canine Rotaviruses
Authors: He Biao , Yang Fanli , Yang Weihong , Zhang Yuzhen , Feng Yun , Zhou Jihua , Xie Jinxin , Feng Ye , Bao Xiaolei , Guo Huancheng , Li Yingying , Xia Lele , Li Nan , Matthijnssens Jelle , Zhang Hailin , Tu Changchun
Published: September 2013

Bats are considered important animal reservoirs for many viruses pathogenic to humans. An approach based on viral metagenomics was used to study gut specimens from 78 insectivorous bats in Yunnan Province, China. Seventy-four reads were found to be related to group A rotavirus (RVA). Further reverse transcription-PCR screening and viral isolation on cell cultures confirmed the presence of a novel RVA strain, named RVA/Bat-tc/MSLH14/2012/G3P[3], in 1 (6%) of 16 lesser horseshoe bats. Full genomic sequencing analyses showed that MSLH14 possessed the genotype constellation G3-P[3]-I8-R3-C3-M3-A9-N3-T3-E3-H6, which is akin to human and animal rotaviruses believed to be of feline/canine origin. Phylogenetic analysis indicated that VP7 was most closely related to bovine RVA strains from India, whereas VP4 was most closely related to an unusual human RVA strain, CMH222, with animal characteristics isolated in Thailand. The remaining gene segments were only distantly related to a range of animal RVA strains, most of which are believed to be related to feline/canine RVAs. Experimental infection showed that bat RVA strain MSLH14 was highly pathogenic to suckling mice, causing 100% mortality when they were inoculated orally with a titer as low as 5 × 10
2
50% tissue culture infective doses. As this virus is not closely related to any known RVA strain, it is tempting to speculate that it is a true bat RVA strain rather than a virus transmitted between species. However, further screening of bat populations, preferably juvenile animals, will be crucial in determining whether or not this virus is widely distributed in the bat population.
Chimeric Exchange of Coronavirus nsp5 Proteases (3CLpro) Identifies Common and Divergent Regulatory Determinants of Protease Activity
Authors: Stobart Christopher C. , Sexton Nicole R. , Munjal Havisha , Lu Xiaotao , Molland Katrina L. , Tomar Sakshi , Mesecar Andrew D. , Denison Mark R.
Published: September 2013

Human coronaviruses (CoVs) such as severe acute respiratory syndrome CoV (SARS-CoV) and Middle East respiratory syndrome CoV (MERS-CoV) cause epidemics of severe human respiratory disease. A conserved step of CoV replication is the translation and processing of replicase polyproteins containing 16 nonstructural protein domains (nsp's 1 to 16). The CoV nsp5 protease (3CLpro; Mpro) processes nsp's at 11 cleavage sites and is essential for virus replication. CoV nsp5 has a conserved 3-domain structure and catalytic residues. However, the intra- and intermolecular determinants of nsp5 activity and their conservation across divergent CoVs are unknown, in part due to challenges in cultivating many human and zoonotic CoVs. To test for conservation of nsp5 structure-function determinants, we engineered chimeric betacoronavirus murine hepatitis virus (MHV) genomes encoding nsp5 proteases of human and bat alphacoronaviruses and betacoronaviruses. Exchange of nsp5 proteases from HCoV-HKU1 and HCoV-OC43, which share the same genogroup, genogroup 2a, with MHV, allowed for immediate viral recovery with efficient replication albeit with impaired fitness in direct competition with wild-type MHV. Introduction of MHV nsp5 temperature-sensitive mutations into chimeric HKU1 and OC43 nsp5 proteases resulted in clear differences in viability and temperature-sensitive phenotypes compared with MHV nsp5. These data indicate tight genetic linkage and coevolution between nsp5 protease and the genomic background and identify differences in intramolecular networks regulating nsp5 function. Our results also provide evidence that chimeric viruses within coronavirus genogroups can be used to test nsp5 determinants of function and inhibition in common isogenic backgrounds and cell types.
Involvement of the Rabies Virus Phosphoprotein Gene in Neuroinvasiveness
Authors: Yamaoka Satoko , Ito Naoto , Ohka Seii , Kaneda Shohei , Nakamura Hiroko , Agari Takahiro , Masatani Tatsunori , Nakagawa Keisuke , Okada Kazuma , Okadera Kota , Mitake Hiromichi , Fujii Teruo , Sugiyama Makoto
Published: September 2013

Rabies virus (RABV), which is transmitted via a bite wound caused by a rabid animal, infects peripheral nerves and then spreads to the central nervous system (CNS) before causing severe neurological symptoms and death in the infected individual. Despite the importance of this ability of the virus to spread from a peripheral site to the CNS (neuroinvasiveness) in the pathogenesis of rabies, little is known about the mechanism underlying the neuroinvasiveness of RABV. In this study, to obtain insights into the mechanism, we conducted comparative analysis of two fixed RABV strains, Nishigahara and the derivative strain Ni-CE, which cause lethal and asymptomatic infections, respectively, in mice after intramuscular inoculation. Examination of a series of chimeric viruses harboring the respective genes from Nishigahara in the genetic background of Ni-CE revealed that the Nishigahara phosphoprotein (P) gene plays a major role in the neuroinvasiveness by mediating infection of peripheral nerves. The results obtained from both
in vivo
and
in vitro
experiments strongly suggested that the Nishigahara P gene, but not the Ni-CE P gene, is important for stable viral replication in muscle cells. Further investigation based on the previous finding that RABV phosphoprotein counteracts the host interferon (IFN) system demonstrated that the Nishigahara P gene, but not the Ni-CE P gene, functions to suppress expression of the beta interferon (IFN-β) gene (
Ifn
-β) and IFN-stimulated genes in muscle cells. In conclusion, we provide the first data strongly suggesting that RABV phosphoprotein assists viral replication in muscle cells by counteracting the host IFN system and, consequently, enhances infection of peripheral nerves.
Stress-Induced Cellular Transcription Factors Expressed in Trigeminal Ganglionic Neurons Stimulate the Herpes Simplex Virus 1 ICP0 Promoter
Authors: Sinani Devis , Cordes Ethan , Workman Aspen , Thunuguntia Prasanth , Jones Clinton
Published: September 2013

Alphaherpesvirinaefamily members can reactivate from latency following stress. The synthetic corticosteroid dexamethasone induces certain cellular transcription factors in murine and bovine trigeminal ganglionic neurons. Three dexamethasone-induced transcription factors, Krüppel-like factor 15, Slug, and SPDEF, stimulated the herpes simplex virus type 1-infected cell protein 0 (ICP0) promoter more than 150-fold. Conversely, other viral promoters (VP16 and ICP4) were not strongly stimulated, suggesting that the ICP0 promoter is preferentially activated by dexamethasone-simulated stress.
Systematic review of the application of the plan–do–study–act method to improve quality in healthcare
Authors: Taylor Michael J , McNicholas Chris , Nicolay Chris , Darzi Ara , Bell Derek , Reed Julie E
Published: September 2013

Background

Plan–do–study–act (PDSA) cycles provide a structure for iterative testing of changes to improve quality of systems. The method is widely accepted in healthcare improvement; however there is little overarching evaluation of how the method is applied. This paper proposes a theoretical framework for assessing the quality of application of PDSA cycles and explores the consistency with which the method has been applied in peer-reviewed literature against this framework.

Methods

NHS Evidence and Cochrane databases were searched by three independent reviewers. Empirical studies were included that reported application of the PDSA method in healthcare. Application of PDSA cycles was assessed against key features of the method, including documentation characteristics, use of iterative cycles, prediction-based testing of change, initial small-scale testing and use of data over time.

Results

73 of 409 individual articles identified met the inclusion criteria. Of the 73 articles, 47 documented PDSA cycles in sufficient detail for full analysis against the whole framework. Many of these studies reported application of the PDSA method that failed to accord with primary features of the method. Less than 20% (14/73) fully documented the application of a sequence of iterative cycles. Furthermore, a lack of adherence to the notion of small-scale change is apparent and only 15% (7/47) reported the use of quantitative data at monthly or more frequent data intervals to inform progression of cycles.

Discussion

To progress the development of the science of improvement, a greater understanding of the use of improvement methods, including PDSA, is essential to draw reliable conclusions about their effectiveness. This would be supported by the development of systematic and rigorous standards for the application and reporting of PDSAs.
The challenges in developing a finite element injury model of the neck to predict the penetration of explosively propelled projectiles
Authors: Breeze Johno , Newbery T , Pope D , Midwinter M J
Published: September 2013

Introduction

Neck injuries sustained by UK service personnel serving on current operations from explosively propelled fragments result in significant mortality and long-term morbidity. Many of these injuries could potentially have been prevented had the soldiers been wearing their issued neck collars at the time of injury. The aim of this research is to develop an accurate method of predicting the resultant damage to cervical neurovascular structures from explosively propelled fragments.

Current status

A finite element numerical model has been developed based on an anatomically accurate, anthropometrically representative 3D mathematical mesh of cervical neurovascular structures. Currently, the model simulates the passage of a fragment simulating projectile through all anatomical components of the neck using material models based upon 20% ballistic gelatin on the simplification that all tissue types act like homogenous muscle.

Future research

The material models used to define the properties of each element within the model will be sequentially replaced by ones specific to each individual tissue within an anatomical structure. However, the cumulative effect of so many additional variables will necessitate experimental validation against both animal models and post-mortem human subjects to improve the credibility of any predictions made by the model. We believe this approach will in the future have the potential to enable objective comparisons between the mitigative effects of different body armour systems to be made with resultant time and financial savings.
Local insulin application on the carotid artery inhibits neointima formation
Authors: Chiang Simon , Breen Danna M. , Guo June , Mori Yusaku , Giacca Adria
Published: September 2013

Anti-mitogenic agents currently used to prevent restenosis in drug-eluting stents delay re-endothelialization. Delayed re-endothelialization is now considered as the main cause of late stent thrombosis with drug-eluting stents, which emphasizes the need for new treatments. We have shown that systemic insulin treatment decreases neointimal growth and accelerates re-endothelialization after arterial injury in a rat model of restenosis. However, systemic insulin treatment cannot be given to non-diabetic individuals because of the risk of hypoglycemia. Thus, we investigated whether local insulin treatment is also effective in reducing neointimal growth after arterial injury. Rats were given local vehicle or local insulin delivered via Pluronic gel applied around the carotid artery immediately following balloon injury. Plasma glucose and systemic insulin levels were not affected by local insulin treatment. Insulin decreased intimal area at 28 days (P < 0.05) and also inhibited vascular smooth muscle cell migration by 60% at 4 days (P < 0.05). NPH (a longer-lasting insulin) also decreased neointimal area. These results indicate that local insulin treatment can lead to decreased restenosis, suggesting a protective vascular effect of insulin in vivo and that local insulin treatment, possibly via insulin-eluting stents, may be clinically relevant.
Prolonged AMP-activated protein kinase induction impairs vascular functions
Authors: Turkseven Saadet , Ertuna Elif
Published: September 2013

AMP-activated protein kinase (AMPK) is a regulator of cellular metabolism and is involved in the pathogenesis of several diseases, including type 2 diabetes and cardiovascular diseases. Data showing the effects of AMPK on vasculature are controversial. Therefore, the aim of this study was to determine the impact of prolonged AMPK activation on vascular functions. For this purpose we have examined the role of AMPK in endothelium-dependent and -independent relaxation and vascular contractions. For this, we incubated thoracic aortic rings, from rats, with AMPK activator 5-aminoimidazole-4-carboxamide-1-4-ribofuranoside (AICAR, 500 μmol/L or 2 mmol/L) in the presence or absence of AMPK inhibitor compound C (10 μmol/L). Next, cumulative dose–response curves to acetylcholine (ACh) (10−9−10−4mol/L), nitroglycerine (NG) (10−9–3 × 10−5mol/L), and noradrenaline (NA) (10−9−10−4mol/L) were obtained. Endothelial nitric oxide synthase (eNOS) protein expression was determined. Our results show that endothelium-dependent relaxation was inhibited after AICAR treatment, and that this effect was reversed by AMPK inhibition. Moreover, AICAR enhanced the contractile response to NA and caused a decrease in eNOS protein expression. In conclusion, prolonged AMPK induction causes endothelial impairment, possibly via increased degradation and (or) reduced expression of eNOS.
EQUIDISTRIBUTION TOWARD THE GREEN CURRENT IN BIG COHOMOLOGY CLASSES
Authors: BAYRAKTAR TURGAY
Published: September 2013

We study limiting distribution of the sequence of pull-backs of smooth (1, 1) forms and positive closed currents by meromorphic endomorphisms of compact Kähler manifolds.
CHARGED LEPTON CORRECTION TO TRI-BIMAXIMAL LEPTON MIXING AND ITS IMPLICATIONS TO NEUTRINO PHENOMENOLOGY
Authors: GOLLU SRINU , DEEPTHI K. N. , MOHANTA R.
Published: September 2013

The recent results from Daya Bay and RENO reactor neutrino experiments have firmly established that the smallest reactor mixing angle θ13 is nonvanishing at the 5 σ level, with a relatively large value, i.e. θ13 ≈ 9°. Using the fact that the neutrino mixing matrix can be represented as [Formula: see text], where Ul and Uν result from the diagonalization of the charged lepton and neutrino mass matrices and Pν is a diagonal matrix containing the Majorana phases and assuming the tri-bimaximal (TBM) form for Uν, we investigate the possibility of accounting for the large reactor mixing angle due to the corrections of the charged lepton mixing matrix. The form of Ul is assumed to be that of CKM mixing matrix of the quark sector. We find that with this modification it is possible to accommodate the large observed reactor mixing angle θ13. We also study the implications of such corrections on the other phenomenological observables.
THE INVESTIGATION ON STABILITY AND STRUCTURAL PROPERTIES OF COHESION-BASED NANOSTRUCTURES MATERIAL
Authors: BAHARI ALI , GHOLIPUR REZA , DERAKHSHI MARYAM
Published: September 2013

Styrene-doped ZrLaO y nanostructures were obtained by sol–gel method low-temperature synthesis. The nanostructures were characterized by X-ray diffraction (XRD), scanning electron microscope (SEM), atomic force microscope (AFM) and transmission electron microscopy (TEM) techniques. The observation using SEM and TEM revealed that the ring-shaped nanostructures were very uniform. Further characterization using XRD disclosed that the cohesion of the samples was controllable with annealing temperatures in the range of 800–1500°C. Cohesion property was investigated for the samples. The cohesion increased when increasing the annealing temperature. This was linked to the reinforcement of the oxygen bound on the ZrLaO y nanostructures The shape of nanostructures showed a transformation from a ring-shaped growth mode to a hole-surfaced growth mode with increasing annealing temperature. The styrene-doped ZrLaO y nanostructures with controllable crystallinity will have great potential for various applications in fuel, cells, batteries, electronics devices and chemical sensors.
PROCESSING–STRUCTURE–PROPERTY CORRELATION IN DC SPUTTERED MOLYBDENUM THIN FILMS
Authors: KHAN MAJID , ISLAM MOHAMMAD , AKRAM AFTAB , MANZOOR UMAIR
Published: September 2013

Molybdenum thin films were sputter deposited under different conditions of DC power and chamber pressure. The structure and topography of the films were investigated using AFM, SEM and XRD techniques. Van der Pauw method and tape test were employed to determine electrical resistivity and interfacial strength to the substrate, respectively. All the films are of sub-micron thickness with maximum growth rate of 78 nm/min and crystallite size in the range of 4 to 21 nm. The films produced at high power and low pressure exhibit compressive residual strains, low electrical resistivity and poor adhesion to the glass substrate, whereas the converse is true for films produced at high pressure.
PLASMA NITRIDING FOR IMPROVING WEAR RESISTANCE OF CABLE BOLT
Authors: NIU SHAOQING , YANG SHUANGSUO , LI YI
Published: September 2013

In order to improve the wear resistance of the cable bolt and increase its life-time during operation, plasma nitriding was employed to obtain a protective nitriding layer on its surface. The microstructure, phase constitution, microhardness and wear resistance of the nitriding layer were investigated. It was shown that continuous and dense nitriding layers were formed on the surface of the samples. The microhardness of the nitrided sample was enhanced by the formation of nitriding layer, which mainly consisted of Fe 4 N and Fe 3 N . The mass losses of the nitrided samples were much smaller and the wear rates were almost hundred times lesser than that of the substrate. Plasma nitriding treatment can effectively enhance the wear resistance of cable bolt.
A comparison of compositional schedulability analysis techniques for hierarchical real-time systems
Authors: Anand Madhukar , Fischmeister Sebastian , Lee Insup
Published: September 2013

Schedulability analysis of hierarchical real-time embedded systems involves defining interfaces that represent the underlying system faithfully and then compositionally analyzing those interfaces. Whereas commonly used abstractions, such as periodic and sporadic tasks and their interfaces, are simple and well studied, results for more complex and expressive abstractions and interfaces based on task graphs and automata are limited. One contributory factor may be the hardness of compositional schedulability analysis with task graphs and automata. Recently, conditional task models, such as the recurring branching task model, have been introduced with the goal of reaching a middle ground in the trade-off between expressivity and ease of analysis. Consequently, techniques for compositional analysis with conditional models have also been proposed, and each offer different advantages. In this work, we revisit those techniques, compare their advantages using an automotive case study, and identify limitations that would need to be addressed before adopting these techniques for use with real-world problems.
Wall Teichoic Acids of Gram-Positive Bacteria
Authors: Brown Stephanie , Santa Maria John P. , Walker Suzanne
Published: September 2013

The peptidoglycan layers of many gram-positive bacteria are densely functionalized with anionic glycopolymers known as wall teichoic acids (WTAs). These polymers play crucial roles in cell shape determination, regulation of cell division, and other fundamental aspects of gram-positive bacterial physiology. Additionally, WTAs are important in pathogenesis and play key roles in antibiotic resistance. We provide an overview of WTA structure and biosynthesis, review recent studies on the biological roles of these polymers, and highlight remaining questions. We also discuss prospects for exploiting WTA biosynthesis as a target for new therapies to overcome resistant infections.
Histone deacetylase 6-mediated deacetylation of α-tubulin coordinates cytoskeletal and signaling events during platelet activation
Authors: Aslan Joseph E. , Phillips Kevin G. , Healy Laura D. , Itakura Asako , Pang Jiaqing , McCarty Owen J. T.
Published: September 2013

The tubulin cytoskeleton plays a key role in maintaining the characteristic quiescent discoid shape of resting platelets. Upon activation, platelets undergo a dramatic change in shape; however, little is known of how the microtubule system contributes to regulating platelet shape and function. Here we investigated the role of the covalent modification of α-tubulin by acetylation in the regulation of platelet physiology during activation. Superresolution microscopy analysis of the platelet tubulin cytoskeleton showed that the marginal band together with an interconnected web of finer tubulin structures collapsed upon platelet activation with the glycoprotein VI (GPVI)-agonist collagen-related peptide (CRP). Western blot analysis revealed that α-tubulin was acetylated in resting platelets and deacetylated during platelet activation. Tubacin, a specific inhibitor of the tubulin deacetylase HDAC6, prevented tubulin deacetylation upon platelet activation with CRP. Inhibition of HDAC6 upregulated tubulin acetylation and disrupted the organization of the platelet microtubule marginal band without significantly affecting platelet volume changes in response to CRP stimulation. HDAC6 inhibitors also inhibited platelet aggregation in response to CRP and blocked platelet signaling events upstream of platelet Rho GTPase activation. Together, these findings support a role for acetylation signaling in controlling the resting structure of the platelet tubulin marginal band as well as in the coordination of signaling systems that drive platelet cytoskeletal changes and aggregation.
Heart failure-induced changes of voltage-gated Ca2+ channels and cell excitability in rat cardiac postganglionic neurons
Authors: Tu Huiyin , Liu Jinxu , Zhang Dongze , Zheng Hong , Patel Kaushik P. , Cornish Kurtis G. , Wang Wei-Zhong , Muelleman Robert L. , Li Yu-Long
Published: September 2013

Chronic heart failure (CHF) is characterized by decreased cardiac parasympathetic and increased cardiac sympathetic nerve activity. This autonomic imbalance increases the risk of arrhythmias and sudden death in patients with CHF. We hypothesized that the molecular and cellular alterations of cardiac postganglionic parasympathetic (CPP) neurons located in the intracardiac ganglia and sympathetic (CPS) neurons located in the stellate ganglia (SG) possibly link to the cardiac autonomic imbalance in CHF. Rat CHF was induced by left coronary artery ligation. Single-cell real-time PCR and immunofluorescent data showed that L (Cav1.2 and Cav1.3), P/Q (Cav2.1), N (Cav2.2), and R (Cav2.3) types of Ca2+ channels were expressed in CPP and CPS neurons, but CHF decreased the mRNA and protein expression of only the N-type Ca2+ channels in CPP neurons, and it did not affect mRNA and protein expression of all Ca2+ channel subtypes in the CPS neurons. Patch-clamp recording confirmed that CHF reduced N-type Ca2+ currents and cell excitability in the CPP neurons and enhanced N-type Ca2+ currents and cell excitability in the CPS neurons. N-type Ca2+ channel blocker (1 μM ω-conotoxin GVIA) lowered Ca2+ currents and cell excitability in the CPP and CPS neurons from sham-operated and CHF rats. These results suggest that CHF reduces the N-type Ca2+ channel currents and cell excitability in the CPP neurons and enhances the N-type Ca2+ currents and cell excitability in the CPS neurons, which may contribute to the cardiac autonomic imbalance in CHF.
Evolution and Cell Physiology. 4. Why invent yet another protein complex to build junctions in epithelial cells?
Authors: Bivic André Le
Published: September 2013

The formation of the first epithelium was an essential step for animal evolution, since it has allowed coordination of the behavior of a cell layer and creation of a selective barrier between the internal medium and the outside world. The possibility of coupling the cells in a single layer has allowed morphogenetic events, such as tube formation, or gastrulation, to form more complex animal morphologies. The invention of sealed junctions between cells has allowed, on the other hand, creation of an asymmetry of nutrients or salts between the apical and the basal side of the epithelial layer. Creation of an internal medium has led to homeostasis, allowing the evolution of more complex physiological functions and the emergence of sophisticated animal shapes. During evolution, the origins of the first animals coincided with the invention of several protein complexes, including true cadherins and the polarity protein complexes. How these complexes regulate formation of the apicolateral border and the adherens junctions is still not fully understood. This review focuses on the role of these apical polarity complexes and, in particular, the Crumbs complex, which is essential for proper organization of epithelial layers from Drosophila to humans.
Mineralocorticoid receptor antagonizes Dot1a-Af9 complex to increaseαENaCtranscription
Authors: Zhang Xi , Zhou Qiaoling , Chen Lihe , Berger Stefan , Wu Hongyu , Xiao Zhou , Pearce David , Zhou Xiaodong , Zhang Wenzheng
Published: September 2013

Aldosterone is a major regulator of Na+absorption and acts by activating the mineralocorticoid receptor (MR) to stimulate the epithelial Na+channel (ENaC). MR−/−mice exhibited pseudohypoaldosteronism type 1 (hyponatremia, hyperkalemia, salt wasting, and high levels of aldosterone) and died around postnatal day 10. However, if and how MR regulates ENaC transcription remain incompletely understood. Our earlier work demonstrated that aldosterone activates αENaC transcription by reducing expression of Dot1a and Af9 and by impairing Dot1a-Af9 interaction. Most recently, we reported identification of a major Af9 binding site in the αENaC promoter and upregulation of αENaC mRNA expression in mouse kidneys lacking Dot1a. Despite these findings, the putative antagonism between the MR/aldosterone and Dot1a-Af9 complexes has never been addressed. The molecular defects leading to PHA-1 in MR−/−mice remain elusive. Here, we report that MR competes with Dot1a to bind Af9. MR/aldosterone and Dot1a-Af9 complexes mutually counterbalance ENaC mRNA expression in inner medullary collecting duct 3 (IMCD3) cells. Real-time RT-quantitative PCR revealed that 5-day-old MR−/−vs. MR+/+mice had significantly lower αENaC mRNA levels. This change was associated with an increased Af9 binding and H3 K79 hypermethylation in the αENaC promoter. Therefore, this study identified MR as a novel binding partner and regulator of Af9 and a novel mechanism coupling MR-mediated activation with relief of Dot1a-Af9-mediated repression via MR-Af9 interaction. Impaired ENaC expression due to failure to inhibit Dot1a-Af9 may play an important role in the early stages of PHA-1 (before postnatal day 8) in MR−/−mice.
Development of enzyme-linked immunosorbent assays for urinary thiazide-sensitive Na-Cl cotransporter measurement
Authors: Isobe Kiyoshi , Mori Takayasu , Asano Takako , Kawaguchi Hiroyuki , Nonoyama Shigeaki , Kumagai Naonori , Kamada Fumiaki , Morimoto Tetsuji , Hayashi Matsuhiko , Sohara Eisei , Rai Tatemitsu , Sasaki Sei , Uchida Shinichi
Published: September 2013

The Na-Cl cotransporter (NCC) in the distal convoluted tubules in kidney is known to be excreted in urine. However, its clinical significance has not been established because of the lack of quantitative data on urinary NCC. We developed highly sensitive enzyme-linked immunosorbent assays (ELISAs) for urinary total NCC (tNCC) and its active form, phosphorylated NCC (pNCC). We first measured the excretion of tNCC and pT55-NCC in urinary exosomes in pseudohypoaldosteronism type II (PHAII) patients since PHAII is caused by NCC activation. Highly increased excretion of tNCC and pNCC was observed in PHAII patients. In contrast, the levels of tNCC and pNCC in the urine of patients with Gitelman's syndrome were not detectable or very low, indicating that both assays could specifically detect the changes in urinary NCC excretion caused by the changes of NCC activity in the kidney. Then, to test whether these assays could be feasible for a more general patient population, we measured tNCC and pNCC in the urine of outpatients with different clinical backgrounds. Although urinary protein levels >30 mg/dl interfered with our ELISA, we could measure urinary pNCC in all patients without proteinuria. Thus we established highly sensitive and quantitative assays for urinary NCC, which could be valuable tools for estimating NCC activity in vivo.
Neurophysiological correlates of aging-related muscle weakness
Authors: Plow Ela B. , Cunningham David A. , Bonnett Corin , Gohar Dina , Bayram Mehmed , Wyant Alexandria , Varnerin Nicole , Mamone Bernadett , Siemionow Vlodek , Hou Juliet , Machado Andre , Yue Guang H.
Published: September 2013

Muscle weakness associated with aging implicates central neural degeneration. However, role of the primary motor cortex (M1) is poorly understood, despite evidence that gains in strength in younger adults are associated with its adaptations. We investigated whether weakness of biceps brachii in aging analogously relates to processes in M1. We enrolled 20 young (22.6 ± 0.87 yr) and 28 old (74.79 ± 1.37 yr) right-handed participants. Using transcranial magnetic stimulation, representation of biceps in M1 was identified. We examined the effect of age and sex on strength of left elbow flexion, voluntary activation of biceps, corticospinal excitability and output, and short-interval intracortical and interhemispheric inhibition. Interhemispheric inhibition was significantly exaggerated in the old ( P = 0.047), while strength tended to be lower ( P = 0.075). Overall, women were weaker ( P < 0.001). Processes of M1 related to strength or voluntary activation of biceps, but only in older adults. Corticospinal excitability was lower in weaker individuals ( r = 0.38), and corticospinal output, intracortical inhibition and interhemispheric inhibition were reduced too in individuals who poorly activated biceps ( r = 0.43, 0.54 and 0.38). Lower intracortical inhibition may reflect compensation for reduced corticospinal excitability, allowing weaker older adults to spread activity in M1 to recruit synergists and attempt to sustain motor output. Exaggerated interhemispheric inhibition, however, conflicts with previous evidence, potentially related to greater callosal damage in our older sample, our choice of proximal vs. distal muscle and differing influence of measurement of inhibition in rest vs. active states of muscle. Overall, age-specific relation of M1 to strength and muscle activation emphasizes that its adaptations only emerge when necessitated, as in a weakening neuromuscular system in aging.
Vascular endothelial growth factor-A signaling in bone marrow-derived endothelial progenitor cells exposed to hypoxic stress
Authors: Hoffmann Brian R. , Wagner Jordan R. , Prisco Anthony R. , Janiak Agnieszka , Greene Andrew S.
Published: September 2013

Bone marrow-derived endothelial progenitor cells (BM-EPCs) are stimulated by vascular endothelial growth factor-A (VEGF-A) and other potent proangiogenic factors. During angiogenesis, an increase in VEGF-A expression stimulates BM-EPCs to enhance endothelial tube formation and contribute to an increase in microvessel density. Hypoxia is known to produce an enhanced angiogenic response and heightened levels of VEGF-A have been seen in oxygen deprived epithelial and endothelial cells, yet the pathways for VEGF-A signaling in BM-EPCs have not been described. This study explores the influence of hypoxia on VEGF-A signaling in rat BM-EPCs utilizing a novel proteomic strategy to directly identify interacting downstream components of the combined VEGF receptor(s) signaling pathways, gene expression analysis, and functional phenotyping. VEGF-A signaling network analysis following liquid chromatographic separation and tandem mass spectrometry revealed proteins related to inositol/calcium signaling, nitric oxide signaling, cell survival, cell migration, and inflammatory responses. Alterations in BM-EPC expression of common angiogenic genes and tube formation in response to VEGF-A during hypoxia were measured and combined with the proteomic analysis to enhance and support the signaling pathways detected. BM-EPC tube formation assays in response to VEGF-A exhibited little tube formation; however, a cell projection/migratory phenotype supported the signaling data. Additionally, a novel assay measuring BM-EPC incorporation into preformed endothelial cell tubes indicated a significant increase of incorporated BM-EPCs after pretreatment with VEGF-A during hypoxia. This study verifies known VEGF-A pathway components and reveals several unidentified mechanisms of VEGF-A signaling in BM-EPCs during hypoxia that may be important for migration to sites of vascular regeneration.
Somatostatin Receptor-Based Molecular Imaging and Therapy for Neuroendocrine Tumors
Authors: Wang Ling , Tang Kun , Zhang Qi , Li Huanbin , Wen Zhengwei , Zhang Hongzheng , Zhang Hong
Published: September 2013

Neuroendocrine tumors (NETs) are tumors originated from neuroendocrine cells in the body. The localization and the detection of the extent of NETs are important for diagnosis and treatment, which should be individualized according to the tumor type, burden, and symptoms. Molecular imaging of NETs with high sensitivity and specificity is achieved by nuclear medicine method using single photon-emitting and positron-emitting radiopharmaceuticals. Somatostatin receptor imaging (SRI) using SPECT or PET as a whole-body imaging technique has become a crucial part of the management of NETs. The radiotherapy with somatostatin analogues labeled with therapeutic beta emitters, such as lutetium-177 or yttrium-90, has been proved to be an option of therapy for patients with unresectable and metastasized NETs. Molecular imaging can deliver an important message to improve the outcome for patients with NETs by earlier diagnosis, better choice of the therapeutic method, and evaluation of the therapeutic response.
Heavy Ethanol Intoxication Increases Proinflammatory Cytokines and Aggravates Hemorrhagic Shock-Induced Organ Damage in Rats
Authors: Hu Tsung-Ming , Lee Ru-Ping , Lee Chung-Jen , Subeq Yi-Maun , Lin Nien-Tsung , Hsu Bang-Gee
Published: September 2013

Hemorrhagic shock (HS) following acute alcohol intoxication can increase proinflammatory cytokine production and induce marked immunosuppression. We investigated the effects of ethanol on physiopathology and cytokine levels following HS in acutely alcohol-intoxicated rats. Rats received an intravenous injection of 5 g/kg ethanol over 3 h followed by HS induced by withdrawal of 40% of total blood volume from a femoral arterial catheter over 30 min. Mean arterial pressure (MAP) and heart rate (HR) were monitored continuously for 48 h after the start of blood withdrawal. Biochemical parameters, including hemoglobin, ethanol, glutamic oxaloacetic transaminase (GOT), glutamic pyruvic transaminase (GPT), blood urea nitrogen (BUN), creatinine (Cre), lactic dehydrogenase (LDH), and creatine phosphokinase (CPK), were measured at 30 min before induction of HS and 0, 1, 3, 6, 9, 12, 18, 24, and 48 h after HS. Serum tumor necrosis factor-α(TNF-α) and interleukin-6 (IL-6) levels were measured at 1 and 12 h after HS. The liver, kidneys, and lungs were removed for pathology at 48 h later. HS significantly increased HR, blood GOT, GPT, BUN, Cre, LDH, CPK, TNF-α, and IL-6 levels and decreased hemoglobin and MAP in rats. Acute ethanol intoxication further increased serum levels of GOT, GPT, BUN, Cre, LDH, CPK, TNF-αand IL-6 elevation following HS. Acutely intoxicated rats exacerbated the histopathologic changes in the liver, kidneys, and lungs following HS.
Antioxidant Properties of Brazilian Tropical Fruits by Correlation between Different Assays
Authors: Gregoris Elena , Pereira Lima Giuseppina Pace , Fabris Sabrina , Bertelle Mariangela , Sicari Michela , Stevanato Roberto
Published: September 2013

Four different assays (the Folin-Ciocalteu, DPPH, enzymatic method, and inhibitory activity on lipid peroxidation) based on radically different physicochemical principles and normally used to determine the antioxidant activity of food have been confronted and utilized to investigate the antioxidant activity of fruits originated from Brazil, with particular attention to more exotic and less-studied species (jurubeba,Solanum paniculatum; pequi,Caryocar brasiliense; pitaya,Hylocereus undatus; siriguela,Spondias purpurea; umbu,Spondias tuberosa) in order to (i) verify the correlations between results obtained by the different assays, with the final purpose to obtain more reliable results avoiding possible measuring-method linked mistakes and (ii) individuate the more active fruit species. As expected, the different methods give different responses, depending on the specific assay reaction. Anyhow all results indicate high antioxidant properties for siriguela and jurubeba and poor values for pitaya, umbu, and pequi. Considering that no marked difference of ascorbic acid content has been detected among the different fruits, experimental data suggest that antioxidant activities of the investigated Brazilian fruits are poorly correlated with this molecule, principally depending on their total polyphenolic content.
Cloud Infrastructures forIn SilicoDrug Discovery: Economic and Practical Aspects
Authors: D'Agostino Daniele , Clematis Andrea , Quarati Alfonso , Cesini Daniele , Chiappori Federica , Milanesi Luciano , Merelli Ivan
Published: September 2013

Cloud computing opens new perspectives for small-medium biotechnology laboratories that need to perform bioinformatics analysis in a flexible and effective way. This seems particularly true for hybrid clouds that couple the scalability offered by general-purpose public clouds with the greater control and ad hoc customizations supplied by the private ones. A hybrid cloud broker, acting as an intermediary between users and public providers, can support customers in the selection of the most suitable offers, optionally adding the provisioning of dedicated services with higher levels of quality. This paper analyses some economic and practical aspects of exploiting cloud computing in a real research scenario for thein silicodrug discovery in terms of requirements, costs, and computational load based on the number of expected users. In particular, our work is aimed at supporting both the researchers and the cloud broker delivering an IaaS cloud infrastructure for biotechnology laboratories exposing different levels of nonfunctional requirements.
Choroidal Metastasis of Non-Small Cell Lung Cancer That Responded to Gefitinib
Authors: Shimomura Iwao , Tada Yuji , Miura Gen , Suzuki Toshio , Matsumura Takuma , Tsushima Kenji , Terada Jiro , Kurimoto Ryota , Sakaida Emiko , Sekine Ikuo , Takiguchi Yuichi , Yamamoto Shuichi , Tatsumi Koichiro
Published: September 2013

A 52-year-old Japanese woman presented with optical symptoms, including left-sided myodesopsia, blurred vision, narrowed visual field, and diminished visual acuity. Ocular evaluation revealed a metastatic tumor in the choroid. Further examinations identified pulmonary adenocarcinoma as the primary tumor. Because an epidermal growth factor receptor gene (EGFR) mutation was detected in a biopsy specimen, gefitinib treatment was initiated. Dramatic responses were obtained in the primary tumor and metastatic foci. Optical symptoms improved and remained stable for 5 months during the treatment, until relapse. This report demonstrates that gefitinib is effective for choroidal metastasis of pulmonary adenocarcinoma harboring anEGFRmutation.
Improved Properties of Poly(3-hydroxybutyrate-co-3-hydroxyvalerate) Produced byComamonassp. EB172 Utilizing Volatile Fatty Acids by Regulating the Nitrogen Source
Authors: Zakaria Mohd Rafein , Ariffin Hidayah , Abd-Aziz Suraini , Hassan Mohd Ali , Shirai Yoshihito
Published: September 2013

This study presents the effect of carbon to nitrogen ratio (C/N) (mol/mol) on the cell growth and poly(3-hydroxybutyrate-co-3-hydroxyvalerate) accumulation byComamonassp. EB172 in 2 L fermenters using volatile fatty acids (VFA) as the carbon source. This VFA was supplemented with ammonium sulphate and yeast extract in the feeding solution to achieve C/N (mol/mol) 5, 15, 25, and 34.4, respectively. By extrapolating the C/N and the source of nitrogen, the properties of the polymers can be regulated. The number average molecular weight (Mn) of P(3HB-co-3HV) copolymer reached the highest at 838 × 103 Da with polydispersity index (PDI) value of 1.8, when the culture broth was supplemented with yeast extract (C/N 34.4). Tensile strength and Young’s modulus of the copolymer containing 6–8 mol% 3HV were in the ranges of 13–14.4 MPa and 0.26–0.34 GPa, respectively, comparable to those of polyethylene (PE). Thus,Comamonassp. EB172 has shown promising bacterial isolates producing polyhydroxyalkanoates from renewable carbon materials.
A Simple HPLC‐DAD Method for the Analysis of Melamine in Protein Supplements: Validation Using the Accuracy Profiles
Authors: Montesano Domenico , Gennari Oriella , Festa Carmen , Zollo Franco , Seccia Serenella , Albrizio Stefania
Published: September 2013

The study presents a fully validated simple high‐performance liquid chromatography method with diode array detection (HPLC‐DAD), able to accurately determine the melamine, fraudulently added, in protein supplements, commonly used from healthy adults to enhance exercise or sport performance. The validation strategy was intentionally oriented towards routine use and the reliability of the method rather than extreme performance. For this reason, validation by accuracy profile, including estimation of uncertainty, was chosen. This procedure, based on the concept of total error (bias + standard deviation), clearly showed that this method was able to determine melamine over the range of 0.05–3.0 mg Kg−1, selected by taking into account the maximum residue levels (MRLs) proposed by European legislation to distinguish between the unavoidable background presence of melamine and unacceptable adulteration. The accuracy profile procedure established that at least 95% of the future results obtained with the proposed method would be within the ±15% acceptance limits of the validated HPLC‐DAD method over the whole defined concentration range.
Phylogenetic Relationships ofPseudorasbora,Pseudopungtungia, andPungtungia(Teleostei; Cypriniformes; Gobioninae) Inferred from Multiple Nuclear Gene Sequences
Authors: Kim Keun-Yong , Ko Myeong-Hun , Liu Huanzhang , Tang Qiongying , Chen Xianglin , Miyazaki Jun-Ichi , Bang In-Chul
Published: September 2013

Gobionine species belonging to the generaPseudorasbora,Pseudopungtungia, andPungtungia(Teleostei; Cypriniformes; Cyprinidae) have been heavily studied because of problems on taxonomy, threats of extinction, invasion, and human health. Nucleotide sequences of three nuclear genes, that is, recombination activating protein gene 1 (rag1), recombination activating gene 2 (rag2), and early growth response 1 gene (egr1), fromPseudorasbora,Pseudopungtungia, andPungtungiaspecies residing in China, Japan, and Korea, were analyzed to elucidate their intergeneric and interspecific phylogenetic relationships. In the phylogenetic tree inferred from their multiple gene sequences,Pseudorasbora,PseudopungtungiaandPungtungiaspecies ramified into three phylogenetically distinct clades; the “tenuicorpa” clade composed ofPseudopungtungia tenuicorpa, the “parva” clade composed of allPseudorasboraspecies/subspecies, and the “herzi” clade composed ofPseudopungtungia nigra, andPungtungia herzi. The genusPseudorasborawas recovered as monophyletic, while the genusPseudopungtungiawas recovered as polyphyletic. Our phylogenetic result implies the unstable taxonomic status of the genusPseudopungtungia.