bluuebunny/crossref_metadata_2025 huggingface dataset:
Development of a Mechanical Damage Classification System for Use in the Post Construction Code for Nonmetallic Repair of Piping and Pressure Vessels
Authors: French Jesse J. , Henshaw John M.
Published: July 2010

ASME PCC-2, Repair of Pressure Equipment and Piping, Article 4.1, addresses the repair of high-pressure systems with nonmetallic composite systems and generally permits the repair of dents and gouges in paragraph 1.2(b) with certain substrate preparation requirements. Research done at The University of Tulsa along with meta-analysis of the work preformed over the last three decades at PRCI, GRI and Battelle has shown that classification of mechanical damage must be further refined and delineated to adequately address all applicable facets of non-metallic repair. This paper covers the author’s work with the Nonmetallic Repair Subgroup (SG NMR) to classify types of mechanical damage and the associated permissible repair methods. These damage classifications are set to be included in future editions of the PCC-2. The types of mechanical damage that were considered for this research were dents, cracks, gouges and certain repairable combinations of the three. Specific definitions are laid forth for the damage types and their subclasses along with geometric nomenclature in an attempt to standardize classification across the differences in the international standards. Other forms of mechanical damage and associated amplifying factors exist and are issues that pipeline and refinery assurance managers are concerned with. Additional examples of mechanical damage that are often considered for non-metallic repair are wrinkles and wrinkle bends, creases, buckling points and seismic folds. A common amplifying factor for mechanical damage is that increased corrosion rates often appear at the site of damage due to breeching of protective coatings or creation of extreme geometries that facilitate crevice corrosion, or the development of residual stresses.
Buckling of Cylindrical, Thin Wall, Trailer Truck Tanks and ASME Section XII
Authors: Karcher Guido G. , Ward Monte , Spoelstra Gary
Published: July 2010

New hazmat (hazardous material) transport tanks will be constructed to the rules of the ASME Code Section XII. The criteria and basis to design these tanks have been questioned and a full scale buckling test has been performed to evaluate maximum allowable over-the-road loadings and required design details. This paper summarizes the results of this testing and provides comparisons with classical Code buckling criteria, Code Case 2286 and other methods for such evaluations. The objective is to provide the ASME Section XII committee with a basis for establishing buckling design criteria and shell stiffening details for both hazmat and nonhazmat transport tanks.
Effects of Surface Finish and Loading Conditions on the Low Cycle Fatigue Behavior of Austenitic Stainless Steel in PWR Environment for Various Strain Amplitude Levels
Authors: Le Duff Jean Alain , Lefranc¸ois Andre´ , Vernot Jean Philippe
Published: July 2010

In February/March 2007, The NRC issued Regulatory Guide “RG1.207” and Argonne National Laboratory issued NUREG/CR-6909 that is now applicable in the US for evaluations of PWR environmental effects in fatigue analyses of new reactor components. In order to assess the conservativeness of the application of this NUREG report, Low Cycle Fatigue (LCF) tests were performed by AREVA NP on austenitic stainless steel specimens in a PWR environment. The selected material exhibits in air environment a fatigue behavior consistent with the ANL reference “air” mean curve, as published in NUREG/CR-6909. LCF tests in a PWR environment were performed at various strain amplitude levels (± 0.6% or ± 0.3%) for two loading conditions corresponding to a simple or to a complex strain rate history. The simple loading condition is a fully reverse triangle signal (for comparison purposes with tests performed by other laboratories with the same loading conditions) and the complex signal simulates the strain variation for an actual typical PWR thermal transient. In addition, two various surface finish conditions were tested: polished and ground. This paper presents the comparisons of penalty factors, as observed experimentally, with penalty factors evaluated using ANL formulations (considering the strain integral method for complex loading), and on the other, the comparison of the actual fatigue life of the specimen with the fatigue life predicted through the NUREG report application. For the two strain amplitudes of ± 0.6% and ± 0.3%, LCF tests results obtained on austenitic stainless steel specimens in PWR environment with triangle waveforms at constant low strain rates give “Fen” penalty factors close to those estimated using the ANL formulation (NUREG/6909). However, for the lower strain amplitude level and a triangle loading signal, the ANL formulation is pessimistic compared to the AREVA NP test results obtained for polished specimens. Finally, it was observed that constant amplitude LCF test results obtained on ground specimens under complex loading simulating an actual sequence of a cold and hot thermal shock exhibits lower combined environmental and surface finish effects when compared to the penalty factors estimated on the basis of the ANL formulations. It appears that the application of the NUREG/CR-6909 in conjunction with the Fen model proposed by ANL for austenitic stainless steel provides excessive margins, whereas the current ASME approach seems sufficient to cover significant environmental effects for representative loadings and surface finish conditions of reactor components.
Procedural Assistance Working Within ASME Code Case 2235-9
Authors: McLay Alex
Published: July 2010

The Time of Flight Diffraction Technique (TOFD) is becoming increasingly used as an alternative to Radiography within both the construction industry and the in service inspection of pressure vessels and pipe work. TOFD, like any other technique requires to be applied in a concise manner in order to achieve the theoretical targets of high probability of detection (POD), a low false call rate and proof of coverage. The introduction of ASME and European standards has brought some coherence to the application of TOFD for fabrication inspection. However, a difficulty lies in the determination of actual coverage by TOFD on different geometries. Although the ASME specifically requires documented evidence of the coverage achieved, common practice is that at best a table is included in the procedure indicating the setups for the different weld geometries involved. To produce a documented examination strategy showing component coverage is cumbersome without software aids. Even with software tools such as professional drawing packages, producing such scan plans is not a simple task if many different geometries are involved. Sonovation felt there was a need to assist industry in simplifying this requirement by producing a simple to use software program that will assist in visualizing the coverage achieved for any setup. This paper will describe the logic behind this software package and demonstrate how it can assist companies in not only preparing procedures in accordance with ASME Rules but acting as a validation tool then on to its use within organisations QA and Data Base systems.
Prevention of Weld Metal Reheat Cracking During Cr-Mo-V Heavy Reactors Fabrication
Authors: Chauvy Ce´dric , Pillot Sylvain
Published: July 2010

CrMoV heavy reactors fabrication has undergone some weld metal reheat cracking issues during the end 2007 / beginning 2008. This paper addresses this particular problem and explains the methodology used to solve it. Usual techniques, such as chemical factors and required NDE, have shown their limits, underlining the fact that this problem was unpredictable. Special mechanical testing as well as very accurate chemical analyses have allowed the authors to find the root cause. Very small amounts of particular impurities were responsible for the cracking and a criterion is then proposed to ensure that the problem will not come out again.
Effective mobility characteristics of platinum-silicided p-type Schottky barrier metal-oxide-semiconductor field-effect transistor
Authors: Jang Moongyu , Jun Myungsim , Zyung Taehyoung
Published: July 2010

A 20 μm long channel platinum-silicided p-type Schottky barrier metal-oxide-semiconductor field-effect transistor (SB-MOSFET) is manufactured. The manufactured p-type SB-MOSFET shows 60 mV/decade subthreshold swing characteristic with leakage current less than 10−7 μA/μm and on/off current ratio larger than 106. Using this platinum-silicided p-type SB-MOSFET, the effective mobility of hole is extracted for the first time. The extracted effective hole mobility has slightly lower value compared to universal hole mobility. The reason for this is due to the existence of Schottky barrier between platinum-silicided and silicon.
Exposure to Bioaerosols during the Growth Season of Tomatoes in an Organic Greenhouse Using Supresivit (
Trichoderma harzianum
) and Mycostop (
Streptomyces griseoviridis
)
Authors: Hansen Vinni Mona , Winding Anne , Madsen Anne Mette
Published: July 2010

In working environments, especially in confined spaces like greenhouses, elevated concentrations of airborne microorganisms may become a problem for workers' health. Additionally, the use of microbial pest control agents (MPCAs) may increase exposure to microorganisms. The aim of this study was to investigate tomato growers' exposure to naturally occurring bioaerosol components [dust, bacteria, fungi, actinomycetes, (1→3)-β-
d
-glucans, and endotoxin] and MPCAs applied by drip irrigation. Airborne dust was collected with filter samplers and analyzed for microorganisms by plate counts and total counts using a microscope. Analysis of (1→3)-β-
d
-glucan and endotoxin content was performed by kinetic, chromatic
Limulus
amoebocyte lysate tests. The fungal strain (
Trichoderma harzianum
) from the biocontrol product Supresivit was identified by PCR analysis. Measurements were performed on the day of drip irrigation and 1 week, 1 month, and 3 months after the irrigation.
T. harzianum
from Supresivit could be detected only on the day of treatment.
Streptomyces griseoviridis
, an applied MPCA, was not detected in the air during this investigation. We found that bioaerosol exposure increases during the growth season and that exposure to fungi, bacteria, and endotoxin can reach levels during the harvest period that may cause respiratory symptoms in growers. The collected data indicate that MPCAs applied by drip irrigation do not become airborne later in the season.
Exploring the Role of Microorganisms in the Disease-Like Syndrome Affecting the Sponge
Ianthella basta
Authors: Luter Heidi M. , Whalan Steve , Webster Nicole S.
Published: July 2010

A disease-like syndrome is currently affecting a large percentage of the
Ianthella basta
populations from the Great Barrier Reef and central Torres Strait. Symptoms of the syndrome include discolored, necrotic spots leading to tissue degradation, exposure of the skeletal fibers, and disruption of the choanocyte chambers. To ascertain the role of microbes in the disease process, a comprehensive comparison of bacteria, viruses, fungi, and other eukaryotes was performed in healthy and diseased sponges using multiple techniques. A low diversity of microbes was observed in both healthy and diseased sponge communities, with all sponges dominated by an
Alphaproteobacteria
, a
Gammaproteobacteria
, and a group I crenarchaeota. Bacterial cultivation, community analysis by denaturing gradient gel electrophoresis (
Bacteria
and
Eukarya
), sequencing of 16S rRNA clone libraries (
Bacteria
and
Archaea
), and direct visual assessment by electron microscopy failed to reveal any putative pathogens. In addition, infection assays could not establish the syndrome in healthy sponges even after direct physical contact with affected tissue. These results suggest that microbes are not responsible for the formation of brown spot lesions and necrosis in
I. basta
.
Rapid Multiplex Detection and Differentiation of
Listeria
Cells by Use of Fluorescent Phage Endolysin Cell Wall Binding Domains
Authors: Schmelcher Mathias , Shabarova Tatiana , Eugster Marcel R. , Eichenseher Fritz , Tchang Vincent S. , Banz Manuel , Loessner Martin J.
Published: July 2010

The genus
Listeria
comprises food-borne pathogens associated with severe infections and a high mortality rate. Endolysins from bacteriophages infecting
Listeria
are promising tools for both their detection and control. These proteins feature a modular organization, consisting of an N-terminal enzymatically active domain (EAD), which contributes lytic activity, and a C-terminal cell wall binding domain (CBD), which targets the lysin to its substrate. Sequence comparison among 12 different endolysins revealed high diversity among the enzyme's functional domains and allowed classification of their CBDs into two major groups and five subclasses. This diversity is reflected in various binding properties, as determined by cell wall binding assays using CBDs fused to fluorescent marker proteins. Although some proteins exhibited a broad binding range and recognize
Listeria
strains representing all serovars, others target specific serovars only. The CBDs also differed with respect to the number and distribution of ligands recognized on the cells, as well as their binding affinities. Surface plasmon resonance analysis revealed equilibrium affinities in the pico- to nanomolar ranges for all proteins except CBD006, which is due to an internal truncation. Rapid multiplexed detection and differentiation of
Listeria
strains in mixed bacterial cultures was possible by combining CBDs of different binding specificities with fluorescent markers of various colors. In addition, cells of different
Listeria
strains could be recovered from artificially contaminated milk or cheese by CBD-based magnetic separation by using broad-range CBDP40 and subsequently identified after incubation with two differently colored CBD fusion proteins of higher specificity.
Diversity of Glycosyl Hydrolases from Cellulose-Depleting Communities Enriched from Casts of Two Earthworm Species
Authors: Beloqui Ana , Nechitaylo Taras Y. , López-Cortés Nieves , Ghazi Azam , Guazzaroni María-Eugenia , Polaina Julio , Strittmatter Axel W. , Reva Oleg , Waliczek Agnes , Yakimov Michail M. , Golyshina Olga V. , Ferrer Manuel , Golyshin Peter N.
Published: July 2010

The guts and casts of earthworms contain microbial assemblages that process large amounts of organic polymeric substrates from plant litter and soil; however, the enzymatic potential of these microbial communities remains largely unexplored. In the present work, we retrieved carbohydrate-modifying enzymes through the activity screening of metagenomic fosmid libraries from cellulose-depleting microbial communities established with the fresh casts of two earthworm species,Aporrectodea caliginosaandLumbricus terrestris, as inocula. Eight glycosyl hydrolases (GHs) from theA. caliginosa-derived community were multidomain endo-β-glucanases, β-glucosidases, β-cellobiohydrolases, β-galactosidase, and β-xylosidases of known GH families. In contrast, two GHs derived from theL. terrestrismicrobiome had no similarity to any known GHs and represented two novel families of β-galactosidases/α-arabinopyranosidases. Members of these families were annotated in public databases as conserved hypothetical proteins, with one being structurally related to isomerases/dehydratases. This study provides insight into their biochemistry, domain structures, and active-site architecture. The two communities were similar in bacterial composition but significantly different with regard to their eukaryotic inhabitants. Further sequence analysis of fosmids and plasmids bearing the GH-encoding genes, along with oligonucleotide usage pattern analysis, suggested that those apparently originated fromGammaproteobacteria(pseudomonads andCellvibrio-like organisms),Betaproteobacteria(Comamonadaceae), andAlphaproteobacteria(Rhizobiales).
In Vivo
Biotinylation of Bacterial Magnetic Particles by a Truncated Form of
Escherichia coli
Biotin Ligase and Biotin Acceptor Peptide
Authors: Maeda Yoshiaki , Yoshino Tomoko , Matsunaga Tadashi
Published: July 2010

Escherichia coli
biotin ligase can attach biotin molecules to a lysine residue of biotin acceptor peptide (BAP), and biotinylation of particular BAP-fused proteins in cells was carried out by coexpression of
E. coli
biotin ligase (
in vivo
biotinylation). This
in vivo
biotinylation technology has been applied for protein purification, analysis of protein localization, and protein-protein interaction in eukaryotic cells, while such studies have not been reported in bacterial cells. In this study,
in vivo
biotinylation of bacterial magnetic particles (BacMPs) synthesized by
Magnetospirillum magneticum
AMB-1 was attempted by heterologous expression of
E. coli
biotin ligase. To biotinylate BacMPs
in vivo
, BAP was fused to a BacMP surface protein, Mms13, and
E. coli
biotin ligase was simultaneously expressed in the truncated form lacking the DNA-binding domain. This truncation-based approach permitted the growth of AMB-1 transformants when biotin ligase was heterologously expressed.
In vivo
biotinylation of BAP on BacMPs was confirmed using an alkaline phosphatase-conjugated antibiotin antibody. The biotinylated BAP-displaying BacMPs were then exposed to streptavidin by simple mixing. The streptavidin-binding capacity of BacMPs biotinylated
in vivo
was 35-fold greater than that of BacMPs biotinylated
in vitro
, where BAP-displaying BacMPs purified from bacterial cells were biotinylated by being mixed with
E. coli
biotin ligase. This study describes not only a simple method to produce biotinylated nanomagnetic particles but also a possible expansion of
in vivo
biotinylation technology for bacterial investigation.
Effective Gene Silencing in a Microsporidian Parasite Associated with Honeybee (
Apis mellifera
) Colony Declines
Authors: Paldi Nitzan , Glick Eitan , Oliva Maayan , Zilberberg Yaron , Aubin Lucie , Pettis Jeffery , Chen Yanping , Evans Jay D.
Published: July 2010

Honeybee colonies are vulnerable to parasites and pathogens ranging from viruses to vertebrates. An increasingly prevalent disease of managed honeybees is caused by the microsporidian
Nosema ceranae
. Microsporidia are basal fungi and obligate parasites with much-reduced genomic and cellular components. A recent genome-sequencing effort for
N. ceranae
indicated the presence of machinery for RNA silencing in this species, suggesting that RNA interference (RNAi) might be exploited to regulate
Nosema
gene expression within bee hosts. Here we used controlled laboratory experiments to show that double-stranded RNA homologous to specific
N. ceranae
ADP/ATP transporter genes can specifically and differentially silence transcripts encoding these proteins. This inhibition also affects
Nosema
levels and host physiology. Gene silencing could be mediated solely by
Nosema
or in concert with known systemic RNAi mechanisms in their bee hosts. These results are novel for the microsporidia and provide a possible avenue for controlling a disease agent implicated in severe honeybee colony losses. Moreover, since microsporidia are pathogenic in several known veterinary and human diseases, this advance may have broader applications in the future for disease control.
Identification and Characterization of a Phosphodiesterase That Inversely Regulates Motility and Biofilm Formation in
Vibrio cholerae
Authors: Liu Xianxian , Beyhan Sinem , Lim Bentley , Linington Roger G. , Yildiz Fitnat H.
Published: July 2010

Vibrio cholerae
switches between free-living motile and surface-attached sessile lifestyles. Cyclic diguanylate (c-di-GMP) is a signaling molecule controlling such lifestyle changes. C-di-GMP is synthesized by diguanylate cyclases (DGCs) that contain a GGDEF domain and is degraded by phosphodiesterases (PDEs) that contain an EAL or HD-GYP domain. We constructed in-frame deletions of all
V. cholerae
genes encoding proteins with GGDEF and/or EAL domains and screened mutants for altered motility phenotypes. Of 52 mutants tested, four mutants exhibited an increase in motility, while three mutants exhibited a decrease in motility. We further characterized one mutant lacking VC0137 (
cdgJ
), which encodes an EAL domain protein. Cellular c-di-GMP quantifications and
in vitro
enzymatic activity assays revealed that CdgJ functions as a PDE. The
cdgJ
mutant had reduced motility and exhibited a small decrease in
flaA
expression; however, it was able to produce a flagellum. This mutant had enhanced biofilm formation and
vps
gene expression compared to that of the wild type, indicating that CdgJ inversely regulates motility and biofilm formation. Genetic interaction analysis revealed that at least four DGCs, together with CdgJ, control motility in
V. cholerae
.
Localization and Cellular Amounts of the WalRKJ (VicRKX) Two-Component Regulatory System Proteins in Serotype 2Streptococcus pneumoniae
Authors: Wayne Kyle J. , Sham Lok-To , Tsui Ho-Ching T. , Gutu Alina D. , Barendt Skye M. , Keen Susan K. , Winkler Malcolm E.
Published: July 2010

The WalRK two-component regulatory system coordinates gene expression that maintains cell wall homeostasis and responds to antibiotic stress in low-GC Gram-positive bacteria. Phosphorylated WalR (VicR) of the major human respiratory pathogenStreptococcus pneumoniae(WalRSpn) positively regulates transcription of several surface virulence genes and, most critically,pcsB, which encodes an essential cell division protein. Despite numerous studies of several species, little is known about the signals sensed by the WalK histidine kinase or the function of the WalJ ancillary protein encoded in thewalRKSpnoperon. To better understand the functions of the WalRKJSpnproteins inS. pneumoniae, we performed experiments to determine their cellular localization and amounts. In contrast to WalK fromBacillus subtilis(WalKBsu), which is localized at division septa, immunofluorescence microscopy showed that WalKSpnis distributed throughout the cell periphery. WalJSpnis also localized to the cell surface periphery, whereas WalRSpnwas found to be localized in the cytoplasm around the nucleoid. In fractionation experiments, WalRSpnwas recovered from the cytoplasmic fraction, while WalKSpnand the majority of WalJSpnwere recovered from the cell membrane fraction. This fractionation is consistent with the localization patterns observed. Lastly, we determined the cellular amounts of WalRKJSpnby quantitative Western blotting. The WalRSpnresponse regulator is relatively abundant and present at levels of ≈6,200 monomers per cell, which are ≈14-fold greater than the amount of the WalKSpnhistidine kinase, which is present at ≈460 dimers (920 monomers) per cell. We detected ≈1,200 monomers per cell of WalJSpnancillary protein, similar to the amount of WalKSpn.
REALIZATION OF SQUARE ROOT DOMAIN TOW–THOMAS BIQUADRATIC FILTER
Authors: ÖLMEZ SİNEM , ÇAM UĞUR
Published: July 2010

In this paper, a Tow–Thomas biquadratic filter designed in square root domain is proposed. The presented filter is constructed with a lossy integrator, a lossless integrator, and a summer block. To the best knowledge of the authors, the filter is the first square root domain Tow–Thomas filter in the literature. The state space synthesis method is used to design the biquadratic filter. The filter operated at 2.5 V supply voltage is simulated by using SPICE simulation program with 0.25 μm TSMC CMOS model parameters. Simulation results are in good agreement with theoretical results that the cut-off frequency and quality factor of the filter are tunable electronically.
Biomechanics of Pressure Ulcer in Body Tissues Interacting with External Forces during Locomotion
Authors: Mak Arthur F.T. , Zhang Ming , Tam Eric W.C.
Published: July 2010

Forces acting on the body via various external surfaces during locomotion are needed to support the body under gravity, control posture, and overcome inertia. Examples include the forces acting on the body via the seating surfaces during wheelchair propulsion, the forces acting on the plantar foot tissues via the insole during gait, and the forces acting on the residual-limb tissues via the prosthetic socket during various movement activities. Excessive exposure to unwarranted stresses at the body-support interfaces could lead to tissue breakdowns commonly known as pressure ulcers, often presented as deep-tissue injuries around bony prominences or as surface damage on the skin. In this article, we review the literature that describes how the involved tissues respond to epidermal loading, taking into account both experimental and computational findings from in vivo and in vitro studies. In particular, we discuss related literature about internal tissue deformation and stresses, microcirculatory responses, and histological, cellular, and molecular observations.
Image-Guided Interventions: Technology Review and Clinical Applications
Authors: Cleary Kevin , Peters Terry M.
Published: July 2010

Image-guided interventions are medical procedures that use computer-based systems to provide virtual image overlays to help the physician precisely visualize and target the surgical site. This field has been greatly expanded by the advances in medical imaging and computing power over the past 20 years. This review begins with a historical overview and then describes the component technologies of tracking, registration, visualization, and software. Clinical applications in neurosurgery, orthopedics, and the cardiac and thoracoabdominal areas are discussed, together with a description of an evolving technology named Natural Orifice Transluminal Endoscopic Surgery (NOTES). As the trend toward minimally invasive procedures continues, image-guided interventions will play an important role in enabling new procedures, while improving the accuracy and success of existing approaches. Despite this promise, the role of image-guided systems must be validated by clinical trials facilitated by partnerships between scientists and physicians if this field is to reach its full potential.
Mechanisms of Defibrillation
Authors: Dosdall Derek J. , Fast Vladimir G. , Ideker Raymond E.
Published: July 2010

Electrical shock has been the one effective treatment for ventricular fibrillation for several decades. With the advancement of electrical and optical mapping techniques, histology, and computer modeling, the mechanisms responsible for defibrillation are now coming to light. In this review, we discuss recent work that demonstrates the various mechanisms responsible for defibrillation. On the cellular level, membrane depolarization and electroporation affect defibrillation outcome. Cell bundles and collagenous septae are secondary sources and cause virtual electrodes at sites far from shocking electrodes. On the whole-heart level, shock field gradient and critical points determine whether a shock is successful or whether reentry causes initiation and continuation of fibrillation.
microRNA: A Master Regulator of Cellular Processes for Bioengineering Systems
Authors: Sun Wei , Julie Li Yi-Shuan , Huang Hsien-Da , Shyy John Y-J. , Chien Shu
Published: July 2010

microRNAs (miRNAs) are small RNAs 18 to 24 nucleotides in length that serve the pivotal function of regulating gene expression. Instead of being translated into proteins, the mature single-stranded miRNA binds to messenger RNAs (mRNAs) to interfere with the translational process. It is estimated that whereas only 1% of the genomic transcripts in mammalian cells encode miRNA, nearly one-third of the encoded genes are regulated by miRNA. Various bioinformatics databases, tools, and algorithms have been developed to predict the sequences of miRNAs and their target genes. In combination with the in silico approaches in systems biology, experimental studies on miRNA provide a new bioengineering approach for understanding the mechanism of fine-tuning gene regulation. This review aims to provide state-of-the-art information on this important mechanism of gene regulation for researchers working in biomedical engineering and related fields. Particular emphases are placed on summarizing the current tools and strategies for miRNA study from a bioengineering perspective and the possible applications of miRNAs (such as antagomirs and miRNA sponges) in biomedical engineering research.
Protein Engineering in the Development of Functional Hydrogels
Authors: Banta Scott , Wheeldon Ian R. , Blenner Mark
Published: July 2010

Proteins, which are natural heteropolymers, have evolved to exhibit a staggering array of functions and capabilities. As scientists and engineers strive to tackle important challenges in medicine, novel biomaterials continue to be devised, designed, and implemented to help to address critical needs. This review aims to cover the present advances in the use of protein engineering to create new protein and peptide domains that enable the formation of advanced functional hydrogels. Three types of domains are covered in this review: (a) the leucine zipper coiled-coil domains, (b) the EF-hand domains, and (c) the elastin-like polypeptides. In each case, the functionality of these domains is discussed as well as recent advancements in the use of these domains to create novel hydrogel-based biomaterials. As protein engineering is used to both create and improve protein domains, these advances will lead to exciting new biomaterials for use in a variety of applications.
Tetracyclines: a pleitropic family of compounds with promising therapeutic properties. Review of the literature
Authors: Griffin Michael O. , Fricovsky Eduardo , Ceballos Guillermo , Villarreal Francisco
Published: July 2010

There must be something unique about a class of drugs (discovered and developed in the mid-1940s) where there are more than 130 ongoing clinical trials currently listed. Tetracyclines were developed as a result of the screening of soil samples for antibiotic organisms. The first of these compounds chlortetracycline was introduced in 1948. Soon after their development tetracyclines were found to be highly effective against various pathogens including rickettsiae, Gram-positive, and Gram-negative bacteria, thus, becoming a class of broad-spectrum antibiotics. The mechanism of action of tetracyclines is thought to be related to the inhibition of protein synthesis by binding to the 30S bacterial ribosome. Tetracyclines are also an effective anti-malarial drug. Over time, many other “protective” actions have been described for tetracyclines. Minocycline, which can readily cross cell membranes, is known to be a potent anti-apoptotic agent. Its mechanism of action appears to relate to specific effects exerted on apoptosis signaling pathways. Another tetracycline, doxycycline is known to exert antiprotease activities. Doxycycline can inhibit matrix metalloproteinases, which contribute to tissue destruction activities in diseases such as gingivitis. A large body of literature has provided additional evidence for the “beneficial” actions of tetracyclines, including their ability to act as oxygen radical scavengers and anti-inflammatory agents. This increasing volume of published work and ongoing clinical trials supports the notion that a more systematic examination of their possible therapeutic uses is warranted. This review provides a summary of tetracycline's multiple mechanisms of action and while using the effects on the heart as an example, this review also notes their potential to benefit patients suffering from various pathologies such as cancer, Rosacea, and Parkinson's disease.
Phospholipase C-dependent Ca2+-sensing pathways leading to endocytosis and inhibition of the plasma membrane vacuolar H+-ATPase in osteoclasts
Authors: Sakai Hiromu , Moriura Yoshie , Notomi Takuya , Kawawaki Junko , Ohnishi Keiko , Kuno Miyuki
Published: July 2010

In osteoclasts, elevation of extracellular Ca2+ is an endogenous signal that inhibits bone resorption. We recently found that an elevation of extracellular Ca2+ decreased proton extrusion through the plasma membrane vacuolar H+-ATPase (V-ATPase) rapidly. In this study we investigated mechanisms underlying this early Ca2+-sensing response, particularly in reference to the activity of the plasma membrane V-ATPase and to membrane retrieval. Whole cell clamp recordings allowed us to measure the V-ATPase currents and the cell capacitance ( Cm) simultaneously. Cm is a measure of cell surface. Extracellular Ca2+ (2.5–40 mM) decreased Cm and the V-ATPase current simultaneously. The decreased Cm, together with the enhanced uptake of a lipophilic dye (FM1–43), indicated that Ca2+ facilitated endocytosis. The endocytosis was blocked by dynamin inhibitors (dynasore and dynamin-inhibitory peptide), by small interfering RNA (siRNA) targeting for dynanmin-2 and also by bafilomycin A1, a blocker of V-ATPases. The extracellular Ca2+-induced endocytosis and inhibition of the V-ATPase current were diminished by a phospholipase C inhibitor (U73122) and siRNA targeting for phospholipase C γ2 subunit. Holding the cytosolic Ca2+ at either high (0.5–5 μM) or low levels or inhibiting calmodulin by an inhibitor (W7) or an antibody (anti-CaM) decreased the stimulated endocytosis and the inhibition of the V-ATPase current. These data suggest that extracellular Ca2+ facilitated dynamin- and V-ATPase-dependent endocytosis in association with an inhibition of the plasma membrane V-ATPase. Phospholipase C, cytosolic Ca2+, and calmodulin were involved in the signaling pathways. Membrane retrieval and the plasma membrane V-ATPase activity may cooperate during the early phase of Ca2+-sensing response in osteoclasts.
Optical molecular imaging and its emerging role in colorectal cancer
Authors: Sheth Rahul A. , Mahmood Umar
Published: July 2010

Colorectal cancer remains a major cause of morbidity and mortality in the United States. The advent of molecular therapies targeted against specific, stereotyped cellular mutations that occur in this disease has ushered in new hope for treatment options. However, key questions regarding optimal dosing schedules, dosing duration, and patient selection remain unanswered. In this review, we describe how recent advances in molecular imaging, specifically optical molecular imaging with fluorescent probes, offer potential solutions to these questions. We begin with an overview of optical molecular imaging, including discussions on the various methods of design for fluorescent probes and the clinically relevant imaging systems that have been built to image them. We then focus on the relevance of optical molecular imaging to colorectal cancer. We review the most recent data on how this imaging modality has been applied to the measurement of treatment efficacy for currently available as well as developmental molecularly targeted therapies. We then conclude with a discussion on how this imaging approach has already begun to be translated clinically for human use.
Insulin- and leptin-regulated fatty acid uptake plays a key causal role in hepatic steatosis in mice with intact leptin signaling but not inob/obordb/dbmice
Authors: Ge Fengxia , Zhou Shengli , Hu Chunguang , Lobdell Harrison , Berk Paul D.
Published: July 2010

Hepatic steatosis results from several processes. To assess their relative roles, hepatocellular long-chain fatty acid (LCFA) uptake was assayed in hepatocytes from C57BL/6J control mice, mice with steatosis from a high-fat diet (HFD) or 10%, 14%, or 18% ethanol (EtOH) in drinking water [functioning leptin-signaling groups (FLSGs)], and ob/ob and db/db mice. Vmaxfor uptake was increased vs. controls ( P < 0.001) and correlated significantly with liver weight and triglycerides (TGs) in all FLSG mice but was minimally or not increased in ob/ob and db/db mice, in which liver weights and TGs greatly exceeded projections from regressions in FLSG animals. Coefficients of determination ( R2) for these FLSG regressions suggest that increased LCFA uptake accounts for ∼80% of the increase in hepatic TGs within these groups, but increased lipogenic gene expression data suggest that enhanced LCFA synthesis is the major contributor in ob/ob and db/db. Got2, Cd36, Slc27a2, and Slc27a5 gene expression ratios were significantly upregulated in the EtOH groups, correlating with sterol regulatory element binding protein 1c ( SREBP1c) and Vmax, but only Cd36 expression was increased in HFD, ob/ob, and db/db mice. Comparison of Vmaxwith serum insulin and leptin suggests that both hormones contribute to upregulation of uptake in the FLSG animals. Thus, increased LCFA uptake, reflecting SREBP1c-mediated upregulation of four distinct transporters, is the dominant cause of steatosis in EtOH-fed mice. In ob/ob and db/db mice, increased LCFA synthesis appears more important. In FLSG animals, insulin upregulates hepatocellular LCFA uptake. Leptin appears to upregulate LCFA uptake or to be essential for full expression of upregulation by insulin.
Adiponectin primes human monocytes into alternative anti-inflammatory M2 macrophages
Authors: Lovren Fina , Pan Yi , Quan Adrian , Szmitko Paul E. , Singh Krishna K. , Shukla Praphulla C. , Gupta Milan , Chan Lawrence , Al-Omran Mohammed , Teoh Hwee , Verma Subodh
Published: July 2010

Altered macrophage kinetics is a pivotal mechanism of visceral obesity-induced inflammation and cardiometabolic risk. Because monocytes can differentiate into either proatherogenic M1 macrophages or anti-inflammatory M2 macrophages, approaches that limit M1 while promoting M2 differentiation represent a unique therapeutic strategy. We hypothesized that adiponectin may prime human monocytes toward the M2 phenotype. Adiponectin promoted the alternative activation of human monocytes into anti-inflammatory M2 macrophages as opposed to the classically activated M1 phenotype. Adiponectin-treated cells displayed increased M2 markers, including the mannose receptor (MR) and alternative macrophage activation-associated CC chemokine-1. Incubation of M1 macrophages with adiponectin-treated M2-derived culture supernatant resulted in a pronounced inhibition of tumor necrosis factor-α and monocyte chemotactic protein-1 secretion. Activation of human monocytes into M2 macrophages by adiponectin was mediated, in addition to AMP-activated protein kinase and peroxisome proliferator-activated receptor (PPAR)-γ, via PPAR-α. Furthermore, macrophages isolated from adiponectin knockout mice demonstrated diminished levels of M2 markers such as MR, which were restored with adiponectin treatment. We report a novel immunoregulatory mechanism through which adiponectin primes human monocyte differentiation into anti-inflammatory M2 macrophages. Conditions associated with low adiponectin levels, such as visceral obesity and insulin resistance, may promote atherosclerosis, in part through aberrant macrophage kinetics.
Pannexin 1 is the conduit for low oxygen tension-induced ATP release from human erythrocytes
Authors: Sridharan Meera , Adderley Shaquria P. , Bowles Elizabeth A. , Egan Terrance M. , Stephenson Alan H. , Ellsworth Mary L. , Sprague Randy S.
Published: July 2010

Erythrocytes release ATP in response to exposure to the physiological stimulus of lowered oxygen (O2) tension as well as pharmacological activation of the prostacyclin receptor (IPR). ATP release in response to these stimuli requires activation of adenylyl cyclase, accumulation of cAMP, and activation of protein kinase A. The mechanism by which ATP, a highly charged anion, exits the erythrocyte in response to lowered O2 tension or receptor-mediated IPR activation by iloprost is unknown. It was demonstrated previously that inhibiting pannexin 1 with carbenoxolone inhibits hypotonically induced ATP release from human erythrocytes. Here we demonstrate that three structurally dissimilar compounds known to inhibit pannexin 1 prevent ATP release in response to lowered O2 tension but not to iloprost-induced ATP release. These results suggest that pannexin 1 is the conduit for ATP release from erythrocytes in response to lowered O2 tension. However, the identity of the conduit for iloprost-induced ATP release remains unknown.
Ovarian cancer G protein-coupled receptor 1-dependent and -independent vascular actions to acidic pH in human aortic smooth muscle cells
Authors: Liu Jin-Peng , Komachi Mayumi , Tomura Hideaki , Mogi Chihiro , Damirin Alatangaole , Tobo Masayuki , Takano Mutsumi , Nochi Hiromi , Tamoto Koichi , Sato Koichi , Okajima Fumikazu
Published: July 2010

Atherosclerosis is a chronic inflammation disease characterized by acidic micromilieu and the accumulation of numerous bioactive lipid mediators, such as lysophosphatidic acid (LPA) and prostaglandins, in the atherosclerotic lesion. Chronic acidification induced various effects on vascular smooth muscle cells, but the molecular mechanisms underlying these effects remain unknown. In this study, we examine the role of proton-sensing ovarian cancer G protein-coupled receptor 1 (OGR1) in extracellular acidification-induced regulation of cyclooxygenase (COX)-2 induction, PGI2production, MAPK phosphatase (MKP)-1 expression, and plasminogen activator inhibitor (PAI)-1 expression and proliferation in human aortic smooth muscle cells (AoSMCs). Experiments with knockdown with small interfering RNA specific to OGR1 and specific inhibitors for G proteins showed that acidification-induced COX-2 expression, PGI2production, and MKP-1 expression, but not PAI-1 expression and inhibition of proliferation, were dependent on OGR1 and mainly mediated by Gq/11protein. LPA remarkably enhanced, through the LPA1receptor/Giprotein, the OGR1-mediated vascular actions to acidic pH. In conclusion, acidic pH-induced vascular actions of AoSMCs can be dissected to OGR1-dependent and -independent pathways: COX-2 expression, PGI2production, and MKP-1 expression are mediated by OGR1, but PAI-1 expression and inhibition of proliferation are not. LPA, which is usually thought to be a proatherogenic lipid mediator, may exert antiatherogenic actions under acidic micromilieu through cross-talk between LPA1/Giprotein and OGR1/Gq/11protein.
Absence of mitochondrial activation during levosimendan inotropic action in perfused paced guinea pig hearts as demonstrated by modular control analysis
Authors: Deschodt-Arsac Véronique , Calmettes Guillaume , Raffard Gérard , Massot Philippe , Franconi Jean-Michel , Pollesello Piero , Diolez Philippe
Published: July 2010

Levosimendan is a calcium sensitizer developed for the treatment of heart failure. It increases contractile force by enhancing the sensitivity of myofilaments to calcium. Besides this sensitizing effect, the drug has also been reported to show some inhibitory action on phosphodiesterase 3 (PDE3). The inotropic effects of levosimendan have been studied on guinea pig paced perfused hearts by using modular control analysis (MoCA) (Diolez P, Deschodt-Arsac V, Raffard G, Simon C, Santos PD, Thiaudiere E, Arsac L, Franconi JM. Am J Physiol Regul Integr Comp Physiol 293: R13–R19, 2007.), an integrative approach of heart energetics using noninvasive31P NMR. The aim was to evaluate quantitatively the respective effects of this drug on energy supply and demand modules. Under our experimental conditions, 0.7 μM levosimendan induced a 45% increase in paced heart output associated with a 7% decrease in phosphocreatine and a negligible increase in oxygen consumption. Because MoCA allows in situ study of the internal regulations in intact beating heart energetics, it was applied to describe quantitatively by which routes levosimendan exerts its inotropic action. MoCA demonstrated the absence of any significant effect of the drug on the supply module, which is responsible for the lower increase in oxygen consumption, compared with epinephrine, which increases the ratio between myocardial oxygen consumption and cardiac contraction. This result evidences that, under our conditions, a possible effect of levosimendan on PDE3 activity and/or intracellular calcium remains very low on mitochondrial activity and insignificant on integrated cardiac energetics. Thus, levosimendan inotropic effect on guinea pig heart depends almost entirely on the calcium-sensitizing properties leading to myofilament activation and the concomitant activation of energy supply by the decrease in PCr, therefore improving energetic efficiency of contraction.
Acute daily psychological stress causes increased atrophic gene expression and myostatin-dependent muscle atrophy
Authors: Allen David L. , McCall Gary E. , Loh Amanda S. , Madden Molly C. , Mehan Ryan S.
Published: July 2010

Psychological stress is known to attenuate body size and lean body mass. We tested the effects of 1, 3, or 7 days of two different models of psychological stress, 1 h of daily restraint stress (RS) or daily cage-switching stress (CS), on skeletal muscle size and atrophy-associated gene expression in mice. Thymus weights decreased in both RS and CS mice compared with unstressed controls, suggesting that both models activated the hypothalamic-pituitary-adrenal axis. Body mass was significantly decreased at all time points for both models of stress but was greater for RS than CS. Mass of the tibialis anterior (TA) and soleus (SOL) muscles was significantly decreased after 3 and 7 days of RS, but CS only significantly decreased SOL mass after 7 days. TA mRNA levels of the atrophy-associated genes myostatin (MSTN), atrogin-1, and the phosphatidylinositol 3-kinase inhibitory subunit p85α were all significantly increased relative to unstressed mice after 1 and 3 days of RS, and expression of MSTN and p85α mRNA remained elevated after 7 days of RS. Expression of muscle ring finger 1 was increased after 1 day of RS but returned to baseline at 3 and 7 days of RS. MSTN, atrogin-1, and p85α mRNA levels also significantly increased after 1 and 3 days of CS but atrogen-1 mRNA levels had resolved back to normal levels by 3 days and p85α with 7 days of CS. p21CIP mRNA levels were significantly decreased by 3 days of CS or RS. Finally, body mass was minimally affected, and muscle mass was completely unaffected by 3 days of RS in mice null for the MSTN gene, and MSTN inactivation attenuated the increase in atrogin-1 mRNA levels with 4 days of RS compared with wild-type mice. Together these data suggest that acute daily psychological stress induces atrophic gene expression and loss of muscle mass that appears to be MSTN dependent.