bluuebunny/crossref_metadata_2025 huggingface dataset:
Self-interaction-free exchange-correlation functional for thermochemistry and kinetics
Authors: Mori-Sánchez Paula , Cohen Aron J. , Yang Weitao
Published: March 2006

We develop a self-interaction-free exchange-correlation functional which is very accurate for thermochemistry and kinetics. This is achieved by theoretical construction of the functional form and nonlinear fitting. We define a simple interpolation of the adiabatic connection that uses exact exchange, generalized gradient approximation (GGA) and meta-GGA functionals. The performance is optimized by fitting a small number of empirical parameters. Overall the new functional improves significantly upon hybrids and meta-GGAs while correctly describing one-electron systems. The mean absolute error on a large set of reaction barriers is reduced to 1.99kcal∕mol.
Hydrogen passivation of carbon Pb like centers at the 3C- and 4H-SiC∕SiO2 interfaces in oxidized porous SiC
Authors: Cantin J. L. , von Bardeleben H. J. , Ke Yue , Devaty R. P. , Choyke W. J.
Published: March 2006

The effect of forming gas and vacuum annealing on the concentration of carbon dangling bond (PbC) centers at 3C- and 4H-SiC∕SiO2 interfaces has been studied by electron paramagnetic resonance (EPR) spectroscopy. Our results show efficient passivation at 400°C and depassivation for temperatures above 850°C. A dissociation energy of (4.3±0.3)eV has been determined for both polytypes.
Effect of fly height and refractive index on the transmission efficiency of near-field optical transducers
Authors: Şendur Kürşat , Jones Paul
Published: March 2006

Heat-assisted magnetic recording is a potential remedy to extend the limits of magnetic recording. A high temperature with a steep gradient is used to reduce the local coercivity of the magnetic medium. To achieve such a thermal profile, an intense optical spot well below the diffraction limit is necessary. Transmission efficiency of a near-field optical transducer is affected by various factors at the head-medium interface. Effect of fly height and presence of high refractive index material at the head-medium interface is investigated. Favorable conditions are identified.
Fabrication of hetero-binary and honeycomb photonic crystals by one-step holographic lithography
Authors: Wu Lijun , Zhong Yongchun , Wong Kam Sing , Wang Guo Ping , Yuan Liang
Published: March 2006

We report the simulation and fabrication of two-dimensional hetero-binary and honeycomb photonic crystals by one-step holographic lithography. These structures are realized by introducing one or three auxiliary beams into the three basic beams forming a regular hexagonal structure. The size contrast between the center rod and its six neighbors in a hetero-binary structure can be tuned by adjusting the intensity contrast between the auxiliary beam and basic beams. The idea of creating heterogeneity into the interference pattern by auxiliary beams could be easily extended to three-dimensional case by adding more beams in the vertical direction.
Polarization characteristics of ZnO rib waveguide random lasers
Authors: Leong Eunice S. P. , Yu S. F. , Abiyasa A. P. , Lau S. P.
Published: March 2006

Ultraviolet ZnO rib waveguide random lasers with stripe width ranging from 1.7 to 6.5μm have been fabricated. It is found that the ZnO random lasers demonstrate strong TE lasing emission especially for rib waveguide with a narrower width. Rate equation analysis has shown that the dominant TE lasing emission is due to the corresponding large confinement factor and scattering strength inside the random media with rib waveguide. In addition, Fourier transform studies of the lasing spectra show that the width of rib waveguide constrains the formation of closed-loop paths of lights and, hence, reduces the number of lasing peaks.
Relationships between chlorophyll fluorescence parameters and photochemical reflectance index of tree species adapted to different temperature regimes
Authors: Weng Jen-Hsien , Chen Yaw-Nan , Liao Tien-Szu
Published: March 2006

Chlorophyll fluorescence parameters and spectral reflectance at leaf level were measured at both predawn and noon, under different temperatures and natural light conditions from autumn to winter. Predawn Fv / Fm of both mango (Mangifera indica L.), a tropical fruit tree, and Podocarpus nagi Zoll. et Moritz., a subtropical conifer, decreased with decreasing temperature, with the former to a greater extent than the latter. Yet, predawn Fv / Fm of Taiwan alder (Alnus formosana Makino), a broadleaf tree widely distributed from the lowlands to 3000 m above sea level in Taiwan, was less influenced by temperature. Nevertheless, taking all three species into consideration, predawn Fv / Fm showed a strong correlation with predawn photochemical reflectance index [(PRIp), PRI = (R531 − R570) / (R531 + R570), where R = reflectance]. For the data obtained at noon, ΔF / Fm′ showed a significant but weak correlation with PRI (PRIn). However, stronger correlation between ΔF / Fm′ and ΔPRI (PRIp − PRIn) was found. In addition, while a non-significant or weak correlation between non-photochemical quenching (NPQ) and PRIn was observed in species sensitive to low temperature, their NPQ was significantly correlated with ΔPRI. We conclude that PRIp can serve as an indicator of the seasonal variation of potential PSII efficiency; and ΔPRI reflects the actual photodissipation as well as actual PSII efficiency during illumination. For the three species in this study, the PRI provides a more consistent measure of the variation in predawn fluorescence values than for steady-state values measured under normal seasonally varying daylight illumination.
Structural basis for rodlet assembly in fungal hydrophobins
Authors: Kwan A. H. Y. , Winefield R. D. , Sunde M. , Matthews J. M. , Haverkamp R. G. , Templeton M. D. , Mackay J. P.
Published: March 2006

Class I hydrophobins are a unique family of fungal proteins that form a polymeric, water-repellent monolayer on the surface of structures such as spores and fruiting bodies. Similar monolayers are being discovered on an increasing range of important microorganisms. Hydrophobin monolayers are amphipathic and particularly robust, and they reverse the wettability of the surface on which they are formed. There are also significant similarities between these polymers and amyloid-like fibrils. However, structural information on these proteins and the rodlets they form has been elusive. Here, we describe the three-dimensional structure of the monomeric form of the class I hydrophobin EAS. EAS forms a β-barrel structure punctuated by several disordered regions and displays a complete segregation of charged and hydrophobic residues on its surface. This structure is consistent with its ability to form an amphipathic polymer. By using this structure, together with data from mutagenesis and previous biophysical studies, we have been able to propose a model for the polymeric rodlet structure adopted by these proteins. X-ray fiber diffraction data from EAS rodlets are consistent with our model. Our data provide molecular insight into the nature of hydrophobin rodlet films and extend our understanding of the fibrillar β-structures that continue to be discovered in the protein world.
Conserved oligomeric Golgi complex subunit 1 deficiency reveals a previously uncharacterized congenital disorder of glycosylation type II
Authors: Foulquier François , Vasile Eliza , Schollen Els , Callewaert Nico , Raemaekers Tim , Quelhas Dulce , Jaeken Jaak , Mills Philippa , Winchester Bryan , Krieger Monty , Annaert Wim , Matthijs Gert
Published: March 2006

The conserved oligomeric Golgi (COG) complex is a heterooctameric complex that regulates intraGolgi trafficking and the integrity of the Golgi compartment in eukaryotic cells. Here, we describe a patient with a mild form of congenital disorder of glycosylation type II (CDG-II) that is caused by a deficiency in the Cog1 subunit of the complex. This patient has a defect in both N- and O-glycosylation. Mass spectrometric analysis of the structures of the N-linked glycans released from glycoproteins from the patient's serum revealed a reduction in sialic acid and galactose residues. Peanut agglutinin (PNA) lectin staining revealed a decrease in sialic acids on core 1 mucin type O-glycans, indicating a combined defect in N- and O-glycosylation. Sequence analysis of the
COG1
cDNA and gene identified a homozygous insertion of a single nucleotide (2659–2660insC), which is predicted to lead to a premature translation stop and truncation of the C terminus of the Cog1 protein by 80 amino acids. This mutation destabilizes several other COG subunits and alters their subcellular localization and hence the overall integrity of the COG complex. This results in reduced levels and/or altered Golgi localization of α-mannosidase II and β-1,4 galactosyltransferase I, which links it to the glycosylation deficiency. Transfection of primary fibroblasts of this patient with the full length hemagglutinin-tagged Cog1 indeed restored β-1,4 galactosyltransferase Golgi localization. We propose naming this disorder CDG-II/Cog1, or CDG-II caused by Cog1 deficiency.
Adenylyl cyclase type 5 (AC5) is an essential mediator of morphine action
Authors: Kim Kyoung-Shim , Lee Ko-Woon , Lee Kang-Woo , Im Joo-Young , Yoo Ji Yeoun , Kim Seung-Woo , Lee Ja-Kyeong , Nestler Eric J. , Han Pyung-Lim
Published: March 2006

Opioid drugs produce their pharmacological effects by activating inhibitory guanine nucleotide-binding regulatory protein-linked μ, δ, and κ opioid receptors. One major effector for these receptors is adenylyl cyclase, which is inhibited upon receptor activation. However, little is known about which of the ten known forms of adenylyl cyclase are involved in mediating opioid actions. Here we show that all of the major behavioral effects of morphine, including locomotor activation, analgesia, tolerance, reward, and physical dependence and withdrawal symptoms, are attenuated in mice lacking adenylyl cyclase type 5 (AC5), a form of adenylyl cyclase that is highly enriched in striatum. Furthermore, the behavioral effects of selective μ or δ opioid receptor agonists are lost in
AC5
−/−
mice, whereas the behavioral effects of selective κ opioid receptor agonists are unaffected. These behavioral data are consistent with the observation that the ability of a μ or δ opioid receptor agonist to suppress adenylyl cyclase activity was absent in striatum of
AC5
−/−
mice. Together, these results establish AC5 as an important component of μ and δ opioid receptor signal transduction mechanisms
in vivo
and provide further support for the importance of the cAMP pathway as a critical mediator of opioid action.
Functional mapping of disease susceptibility loci using cell biology
Authors: Antinozzi Peter A. , Garcia-Diaz Alejandro , Hu Chuan , Rothman James E.
Published: March 2006

In most genome-wide linkage studies, implication of a causative disease gene often requires years of expanding the study to more families and finer mapping of the initially described region. Even after such efforts, unobtainable sample sizes can be required to make statistically meaningful conclusions about a single gene. Here we demonstrate that by adding a layer of functional biology to statistical genetic results, this process can be accelerated. The diabetes susceptibility locus (chromosome 18p11) was systematically dissected by using a cell-based secretion assay and RNA interference, and we identified laminin α1 to have a role in pancreatic β cell secretion. The screen was extended to identify laminin receptor 1 as a functional partner in regards to β cell function. Our approach can potentially be widely used in the setting of high-throughput cellular screening of other loci to identify candidate genes.
High-amylose wheat generated by RNA interference improves indices of large-bowel health in rats
Authors: Regina Ahmed , Bird Anthony , Topping David , Bowden Sarah , Freeman Judy , Barsby Tina , Kosar-Hashemi Behjat , Li Zhongyi , Rahman Sadequr , Morell Matthew
Published: March 2006

Foods high in resistant starch have the potential to improve human health and lower the risk of serious noninfectious diseases. RNA interference was used to down-regulate the two different isoforms of starch-branching enzyme (SBE) II (SBEIIa and SBEIIb) in wheat endosperm to raise its amylose content. Suppression of SBEIIb expression alone had no effect on amylose content; however, suppression of both SBEIIa and SBEIIb expression resulted in starch containing >70% amylose. When the >70% amylose wheat grain was fed to rats in a diet as a wholemeal, several indices of large-bowel function, including short-chain fatty acids, were improved relative to standard wholemeal wheat. These results indicate that this high-amylose wheat has a significant potential to improve human health through its resistant starch content.
A small interfering RNA screen for modulators of tumor cell motility identifies MAP4K4 as a promigratory kinase
Authors: Collins Cynthia S. , Hong Jiyong , Sapinoso Lisa , Zhou Yingyao , Liu Zheng , Micklash Kenneth , Schultz Peter G. , Hampton Garret M.
Published: March 2006

Cell motility is a complex biological process, involved in development, inflammation, homeostasis, and pathological processes such as the invasion and metastatic spread of cancer. Here, we describe a genomic screen designed to identify inhibitors of cell migration. A library of 10,996 small interfering RNAs (targeting 5,234 human genes) was screened for their ability to block the migration of a highly motile ovarian carcinoma cell line, SKOV-3, by using a 384-well wound-healing assay coupled with automated microscopy and wound quantification. Two or more small interfering RNAs against four genes,
CDK7
,
DYRK1B
,
MAP4K4
(
NIK
/
HGK
) (
MAP4K4
, mitogen-activated protein 4 kinase 4), and
SCCA-1
(SerpinB3), potently blocked the migration of SKOV-3 cells, concordant with reduced transcript levels. Further studies of the promigratory role of
MAP4K4
showed that the knockdown of this transcript inhibited the migration of multiple carcinoma cell lines, indicating a broad role in cell motility and potently suppressed the invasion of SKOV-3 cells
in vitro
. The effect of
MAP4K4
on cellular migration was found to be mediated through c-Jun N-terminal kinase, independent of AP1 activation and downstream transcription. Accordingly, small molecule inhibition of c-Jun N-terminal kinase suppressed SKOV-3 cell migration, underscoring the potential therapeutic utility of mitogen-activated protein kinase pathway inhibition in cancer progression.
Structural characterization of autoinhibited c-Met kinase produced by coexpression in bacteria with phosphatase
Authors: Wang Weiru , Marimuthu Adhirai , Tsai James , Kumar Abhinav , Krupka Heike I. , Zhang Chao , Powell Ben , Suzuki Yoshihisa , Nguyen Hoa , Tabrizizad Maryam , Luu Catherine , West Brian L.
Published: March 2006

Protein kinases are a large family of cell signaling mediators undergoing intensive research to identify inhibitors or modulators useful for medicine. As one strategy, small-molecule compounds that bind the active site with high affinity can be used to inhibit the enzyme activity. X-ray crystallography is a powerful method to reveal the structures of the kinase active sites, and thus aid in the design of high-affinity, selective inhibitors. However, a limitation still exists in the ability to produce purified kinases in amounts sufficient for crystallography. Furthermore, kinases exist in different conformation states as part of their normal regulation, and the ability to prepare crystals of kinases in these various states also remains a limitation. In this study, the c-Abl, c-Src, and c-Met kinases are produced in high yields in
Escherichia coli
by using a bicistronic vector encoding the PTP1B tyrosine phosphatase. A 100-fold lower dose of the inhibitor, Imatinib, was observed to inhibit the unphosphorylated form of c-Abl kinase prepared by using this vector, compared to the phosphorylated form produced without PTP1B, consistent with the known selectivity of this inhibitor for the unactivated conformation of the enzyme. Unphosphorylated c-Met kinase produced with this vector was used to obtain the crystal structure, at 2.15-Å resolution, of the autoinhibited form of the kinase domain, revealing an intricate network of interactions involving c-Met residues documented previously to cause dysregulation when mutated in several cancers.
Reduced c-Myc signaling triggers telomere-independent senescence by regulating Bmi-1 and p16
INK4a
Authors: Guney Isil , Wu Shirley , Sedivy John M.
Published: March 2006

Increased mitogenic signaling by positive effectors such as Ras or Myc can trigger senescence in normal cells, a response believed to function as a tumor-suppressive mechanism. We report here the existence of a checkpoint that monitors hypoproliferative signaling imbalances. Normal human fibroblasts with one copy of the
c-myc
gene inactivated by targeted homologous recombination switched with an increased frequency to a telomere-independent senescent state mediated by the cyclin-dependent kinase inhibitor p16
INK4a
. p16
INK4a
expression was regulated by the Polycomb group repressor Bmi-1, which we show is a direct transcriptional target of c-Myc. The Myc–Bmi circuit provides a mechanism for the conversion of environmental inputs that converge on c-Myc into discrete cell-fate decisions coupled to cell-cycle recruitment. A mechanism for limiting the proliferation of damaged or otherwise physiologically compromised cells would be expected to have important consequences on the generation of replicatively senescent cells during organismal aging.
Neutrophil polarization: Spatiotemporal dynamics of RhoA activity support a self-organizing mechanism
Authors: Wong Kit , Pertz Olivier , Hahn Klaus , Bourne Henry
Published: March 2006

Chemoattractants like fMet-Leu-Phe (fMLP) induce neutrophils to polarize with phosphatidylinositol 3,4,5-trisphosphate (PIP3) and protrusive F-actin at the front and actomyosin contraction at the sides and back. RhoA and its downstream effector, myosin II, mediate the “backness” response, which locally inhibits the “frontness” response and constrains its location to one part of the cell. In living HL-60 cells, we used a fluorescent PIP3 probe or a single-chain FRET biosensor for RhoA-GTP to assess spatial distribution of frontness or backness responses, respectively, during the first 3 min after exposure to a uniform concentration of fMLP. Increased PIP3 signal or RhoA activity initially localized randomly about the cell’s periphery but progressively redistributed to the front or to the back and sides, respectively. Cells rendered unable to mount the frontness response (by inhibiting actin polymerization or Gi, a trimeric G protein) responded to a micropipette source of attractant by localizing RhoA activity at the up-gradient edge. We infer that protrusive F-actin, induced by the frontness response, constrains the spatial distribution of backness by locally reducing activation of RhoA, thereby reducing its active form at the front. Mutual incompatibility of frontness and backness is responsible for self-organization of neutrophil polarity.
Cofactor-containing antibodies: Crystal structure of the original yellow antibody
Authors: Zhu Xueyong , Wentworth Paul , Kyle Robert A. , Lerner Richard A. , Wilson Ian A.
Published: March 2006

Antibodies are generally thought to be a class of proteins that function without the use of cofactors. However, it is not widely appreciated that antibodies are believed to be the major carrier protein in human circulation for the important riboflavin cofactor that is involved in a host of biological phenomena. A further link between riboflavin and antibodies was discovered 30 years ago when a bright-yellow antibody, IgG
GAR
, was purified from a patient with multiple myeloma who had turned yellow during the course of her disease. It was subsequently shown that the yellow color of this antibody was due to riboflavin binding. However, it was not known how and where riboflavin was bound to this antibody. We now report the crystal structure of this historically important IgG
GAR
Fab at 3.0-Å resolution. The riboflavin is located in the antigen-combining site with its isoalloxazine ring stacked between the parallel aromatic moieties of TyrH33, PheH58, and TyrH100A. Together with additional hydrogen bonds, these interactions reveal the structural basis for high-affinity riboflavin binding. The ligand specificity of IgG
GAR
is compared with another riboflavin-binding antibody, IgG
DOT
, which was purified from a second patient with multiple myeloma. The crystal structure of IgG
GAR
provides a starting point for attempts to understand the physiological relevance and chemical functions of cofactor-containing antibodies.
Imaging Methods for Morphological and Functional Phenotyping of the Rodent Heart
Authors: Badea Cristian T. , Bucholz Elizabeth , Hedlund Laurence W. , Rockman Howard A. , Johnson G. Allan
Published: March 2006

Small animal imaging has a critical role in phenotyping, drug discovery, and in providing a basic understanding of mechanisms of disease. Translating imaging methods from humans to small animals is not an easy task. The purpose of this work is to compare two cardiac imaging modalities, i.e., magnetic resonance microscopy (MRM) and microcomputed tomography (CT) for preclinical studies on rodents. We present the two technologies, the parameters that they can measure, the types of alterations that they can detect, and show how these imaging methods compare to techniques available in clinical medicine. While this paper does not refer per se to the cardiac risk assessment for drug or chemical development, we hope that the information will effectively address how MRM and micro-CT might be exploited to measure biomarkers critical for safety assessment.
Utility of hERG Assays as Surrogate Markers of Delayed Cardiac Repolarization and QT Safety
Authors: Gintant Gary A. , Su Zhi , Martin Ruth L. , Cox Bryan F.
Published: March 2006

HERG (human-ether-a-go-go-related gene) encodes for a cardiac potassium channel that plays a critical role in defining ventricular repolarization. Noncardiovascular drugs associated with a rare but potentially lethal ventricular arrhythmia (Torsades de Pointes) have been linked to delayed cardiac repolarization and block of hERG current. This brief overview will discuss the role of hERG current in cardiac electrophysiology, its involvement in drug-induced delayed repolarization, and approaches used to define drug effects on hERG current. In addition, examples of hERG blocking drugs acting differently (i.e., overt and covert hERG blockade due to multichannel block) together with the utility and limitations of hERG assays as tools to predict the risk of delayed repolarization and proarrhythmia are discussed.
Preclinical Restenosis Models: Challenges and Successes
Authors: Touchard Arturo G. , Schwartz Robert S.
Published: March 2006

Coronary artery disease remains a major problem for Western societies. The advent of percutaneous interventions, including stents has brought clinical care to a new level of efficacy, yet problems remain. Restenosis following stenting in human coronary arteries appears at last to be yielding to therapeutic strategies, especially drug eluting stents. Because therapeutic percutaneous coronary intervention is widely dominated by the intracoronary stent, restenosis therapies must include the stented coronary artery. Animal models and in particular the porcine coronary model seem to represent the human coronary artery reaction to stenting. It mimics several clinical conditions including thrombosis and neointimal formation. A key question in the era of intravascular technologies is how well this and other models can predict clinical events. This paper discusses the models and their application.
Angiogenesis: Bench to Bedside, Have We Learned Anything?
Authors: Lee Seung Uk , Wykrzykowska Joanna J. , Laham Roger J.
Published: March 2006

End-stage ischemic cardiomyopathy patients are an ever-increasing group of coronary artery disease patients, often with no options in our current treatment armamentarium. Angiogenesis therapy pre-clinical and phase I clinical trials showed great promise, however, the benefits of single growth factor treatments have not been borne out in the larger phase II randomized trials. The complexity of angiogenesis process and the challenges in creating animal models to replicate and study this process in ischemic adult human myocardium have been major limitations to progress in this field. In addition failure to control for the powerful placebo effect in the clinical trials and inadequate methods of outcomes measures assessment have created difficult to overcome road blocks in establishing the efficacy of angiogenic strategies. Herein we review the challenges of angiogenesis research and development of treatment strategies. We also propose a structured model for further investigations of angiogenic therapies. The adherence to such a regimented approach as proposed here is, in our opinion, the only way to achieve success in angiogenesis approach development to treatment of patients with end-stage cardiac ischemia refractory to other established therapies.
Sudden Cardiac Death not Related to Coronary Atherosclerosis
Authors: Ladich Elena , Virmani Renu , Burke Allen
Published: March 2006

Sudden cardiac death (SCD) accounts for approximately 300,000 cardiac events in the United States each year, representing an overall incidence of 0.1–0.2% per year. Although the vast majority of these may be attributed to coronary atherosclerosis, a wide variety of nonatherosclerotic-related cardiac diseases have been associated with SCD. This review highlights three general categories of cardiac disease not related to atherosclerosis: the cardiomyopathies, inflammatory myocardial diseases, and ion channel disorders. The important role played by genetics in some of these cardiovascular diseases is presented as well as toxic and drug-related etiologies.
Diffusion magnetic resonance imaging may provide prognostic information in osmotic demyelination syndrome: report of a case
Authors: Dervisoglu E. , Yegenaga I. , Anik Y. , Sengul E. , Turgut T.
Published: March 2006

Hyponatremia and its rapid correction may cause osmotic demyelination syndrome (ODS) with damage to the pontine and extrapontine areas of the brain. The damage may become persistent or may regress and disappear during follow-up. We describe the case of a 35-year-old woman with chronic renal failure who was admitted to the emergency department with profound hyponatremia which was corrected rapidly after hemodialysis treatment. During follow-up, she developed quadriparesis and dysartria. Magnetic resonance imaging (MRI) demonstrated abnormalities characteristic of ODS in the pons as well as the basal ganglia with increased signal intensity on T2 and diffusion-weighted (DW) MRI and low apparent diffusion coefficient (ADC) values. After the sixth day, her clinical status improved progressively. Control MRI revealed rapid normalization of the ADC values during the first week and month parallel to the clinical improvement. However, the hyperintensities on T2-weighted images persisted. Four months later the MRI findings were completely normal. The close relationship between the ADC abnormality and the clinical status suggests that DW-MRI may be useful in predicting the prognosis of ODS.
Behavioral Responses to Threatened Ethnic Identity in a Sports Setting
Authors: Dru Vincent
Published: March 2006

Boxing competitions with two rounds between French and Arab students (N = 58) were used to investigate intergroup relations and social identity. Behavioral measures were observed in two rounds of the competition to assess possible changes depending on threat conditions and strength of ethnic identification. Analyses showed that behavioral effort in round 2 was not influenced by the degree of ethnic identification or threat conditions. However, behavioral competitiveness (the number of initiatives and ripostes attempted relative to the opponent's) in round 2 was greater in the higher threat condition for high ethnic identifiers. Behavioral competitiveness decreased slightly from the no-threat to the threat condition for low ethnic identifiers. The results are discussed in terms of the social identity processes that might influence behavioral responses in naturalistic intergroup settings.
Insulin Glulisine—A Comprehensive Preclinical Evaluation
Authors: Stammberger Ingo , Seipke Gerhard , Bartels Thomas
Published: March 2006

Receptor binding and signaling and the mitogenic potential of insulin glulisine (glulisine), regular human insulin (RHI), and Asp(B10) were compared in vivo and in vitro. Insulin and insulin-like growth factor 1 (IGF-1) receptor binding was studied with human insulin receptors (293HEK cells) and the human osteosarcoma-derived cell line B10. Insulin receptor–mediated signaling was assessed in rat-1 fibroblasts overexpressing insulin receptors. Activation of insulin receptor substrates 1 and 2 (IRS-1/IRS-2) was studied in rat and human myoblasts and rat cardiomyocytes. DNA synthesis induction was assessed by [3H] thymidine incorporation in the human epithelial breast cell line MCF10. Interaction with the IGF-1 receptor, DNA synthesis, and intracellular signal transduction were assessed in cardiac K6 myoblasts. Immunohistochemical examination of Sprague-Dawley rat tissue treated with glulisine for 6 months ( n = 40), and glulisine and RHI for 12 months ( n = 60), was performed. Steady-state insulin receptor binding affinity was slightly lower for glulisine versus RHI (~0.70). IGF-1 receptor binding affinity was lower (four-to fivefold) for glulisine, but significantly higher (four-fold) for Asp(B10) versus RHI. Glulisine, Asp(B10), and RHI showed similar insulin receptor–association kinetics; however, Asp(B10) revealed increased insulin receptor affinity. Glulisine and RHI showed similar insulin receptor–mediated phosphorylation and IRS-2 activation. Activation of IRS-1 was 6- to 10-fold lower with glulisine; glulisine was less potent and Asp(B10) slightly more potent in stimulating DNA synthesis versus RHI. Stimulation of DNA synthesis was comparable for glulisine and RHI in K6 myoblasts. At 12 months, there was no significant difference between glulisine and RHI in proliferative activity. This preclinical evaluation suggests that structural changes in glulisine versus RHI are not associated with any safety issues.
Involvement of Calcium in 7β-Hydroxycholesterol and Cholesterol-5β,6β-Epoxide-Induced Apoptosis
Authors: Ryan L. , O’Callaghan Y. C. , O’Brien N. M.
Published: March 2006

Oxidized low-density lipoprotein (oxLDL) is believed to play a central role in the development of atherosclerosis. The induction of apoptosis in cells of the arterial wall is a critical event in the development of atheroma. 7β-Hydroxycholesterol (7 β-OH) and cholesterol-5 β,6 β-epoxide ( β-epoxide) are components of oxLDL and have previously been shown to be potent inducers of apoptosis. The exact mechanism through which these oxysterols induce apoptosis remains to be fully elucidated. A perturbation of intra-cellular calcium homeostasis has been found to trigger apoptosis in many experimental systems. The aim of the present study was to determine the involvement of calcium signaling in 7 β-OH and β-epoxide–induced apoptosis. To this end, the authors employed the calcium channel blockers verapamil and nifedipine and inhibitors of calpain activation, ALLM and ALLN. Verapamil protected against the decrease in viability induced by 7 β-OH whereas nifedipine had a protective effect in both 7 β-OH and β-epoxide–treated cells, though these compounds did not restore viability to control levels. Verapamil, nifedipine, and ALLM prevented apoptosis induced by β-epoxide. None of the compounds employed in the current study protected against 7 β-OH–induced apoptosis. Our results implicate calcium signaling in the apoptotic pathway induced by β-epoxide and also highlight differences between apoptosis induced by 7 β-OH and β-epoxide.
Genotoxic Effects of Benzo[a]pyrene and Dibenzo[a,l]pyrene in a Human Lung Cell Line
Authors: Mourón Silvana Andrea , Grillo Claudia Alejandra , Dulout Fernando Noel , Golijow Carlos Daniel
Published: March 2006

Several studies have shown that polycyclic aromatic hydrocarbons (PAHs) produce genotoxic effects in assays performed in vivo and in vitro. This study was undertaken to investigate the ability of benzo[ a]pyrene (BP) and dibenzo[ a,l]pyrene (DBP) to induce DNA damage in a human lung fibroblast cell line (MRC-5), using sister-chromatid exchanges test (SCEs), the comet assay, and evaluating point mutations in codon 12 of the K- ras protooncogene by polymerase chain reaction–single-strand conformation polymorphisms (PCR-SSCPs) and restriction fragment length polymorphisms (RFLP)-enriched PCR methods. Sister-chromatid exchanges frequencies were significantly increased in cells exposed to benzo[ a]pyrene and dibenzo[ a,l]pyrene in relation to controls ( p < .001). Using the standard alkaline comet assay, significant differences between groups were found for the variable comet moment (CM) when cells were exposed to BP ( p < .001) and DBP ( p < .001). Nevertheless, PCR-SSCP and RFLP-enriched PCR methods did not show any association between treatments with BP and DBP and K- ras point mutations. The data presented in this study indicated that BP and DBP induced both DNA strand breaks and sister-chromatid exchanges but not significant point mutations at codon 12 of K- ras gene in the MRC-5 cell line.
Neural Cell Adhesion Molecule Induces Intracellular Signaling via Multiple Mechanisms of Ca2+ Homeostasis
Authors: Kiryushko Darya , Korshunova Irina , Berezin Vladimir , Bock Elisabeth
Published: March 2006

The neural cell adhesion molecule (NCAM) plays a pivotal role in the development of the nervous system, promoting neuronal differentiation via homophilic (NCAM–NCAM) as well as heterophilic (NCAM-fibroblast growth factor receptor [FGFR]) interactions. NCAM-induced intracellular signaling has been shown to affect and be dependent on the cytoplasmic Ca2+ concentration ([Ca2+]i). However, the molecular basis of this remains unclear. In this study, we determined [Ca2+]i regulating mechanisms involved in intracellular signaling induced by NCAM. To mimic the effect of homophilic NCAM interaction on [Ca2+]i in vitro, we used a peptide derived from a homophilic binding site of NCAM, termed P2, which triggers signaling cascades similar to those activated by NCAM–NCAM interaction. We found that P2 increased [Ca2+]i in primary hippocampal neurons. This effect depended on two signaling pathways. The first pathway was associated with activation of FGFR, phospholipase Cγ, and production of diacylglycerol, and the second pathway involved Src-family kinases. Moreover, NCAM-mediated Ca2+ entry required activation of nonselective cation and T-type voltage-gated Ca2+ channels. These channels, together with the Src-family kinases, were also involved in neuritogenesis induced by physiological, homophilic NCAM interactions. Thus, unanticipated mechanisms of Ca2+ homeostasis are shown to be activated by NCAM and to contribute to neuronal differentiation.
The APC tumor suppressor counteracts β-catenin activation and H3K4 methylation at Wnt target genes
Authors: Sierra Jose , Yoshida Tomonori , Joazeiro Claudio A. , Jones Katherine A.
Published: March 2006

The APC tumor suppressor controls the stability and nuclear export of β-catenin (β-cat), a transcriptional coactivator of LEF-1/TCF HMG proteins in the Wnt/Wg signaling pathway. We show here that β-cat and APC have opposing actions at Wnt target genes in vivo. The β-cat C-terminal activation domain associates with TRRAP/TIP60 and mixed-lineage-leukemia (MLL1/MLL2) SET1-type chromatin-modifying complexes in vitro, and we show that β-cat promotes H3K4 trimethylation at the c-Myc gene in vivo. H3K4 trimethylation in vivo requires prior ubiquitination of H2B, and we find that ubiquitin is necessary for transcription initiation on chromatin but not nonchromatin templates in vitro. Chromatin immunoprecipitation experiments reveal that β-cat recruits Pygopus, Bcl-9/Legless, and MLL/SET1-type complexes to the c-Myc enhancer together with the negative Wnt regulators, APC, and βTrCP. Interestingly, APC-mediated repression of c-Myc transcription in HT29-APC colorectal cancer cells is initiated by the transient binding of APC, βTrCP, and the CtBP corepressor to the c-Myc enhancer, followed by stable binding of the TLE-1 and HDAC1 corepressors. Moreover, nuclear CtBP physically associates with full-length APC, but not with mutant SW480 or HT29 APC proteins. We conclude that, in addition to regulating the stability of β-cat, APC facilitates CtBP-mediated repression of Wnt target genes in normal, but not in colorectal cancer cells.
Gene-specific requirement for P-TEFb activity and RNA polymerase II phosphorylation within the p53 transcriptional program
Authors: Gomes Nathan P. , Bjerke Glen , Llorente Briardo , Szostek Stephanie A. , Emerson Beverly M. , Espinosa Joaquin M.
Published: March 2006

Activation of the p53 pathway mediates cellular responses to diverse forms of stress. Here we report that the p53 target gene p21CIP1 is regulated by stress at post-initiation steps through conversion of paused RNA polymerase II (RNAP II) into an elongating form. High-resolution chromatin immunoprecipitation assays (ChIP) demonstrate that p53-dependent activation of p21CIP1 transcription after DNA damage occurs concomitantly with changes in RNAP II phosphorylation status and recruitment of the elongation factors DSIF (DRB Sensitivity-Inducing Factor), P-TEFb (Positive Transcription Elongation Factor b), TFIIH, TFIIF, and FACT (Facilitates Chromatin Transcription) to distinct regions of the p21CIP1 locus. Paradoxically, pharmacological inhibition of P-TEFb leads to global inhibition of mRNA synthesis but activation of the p53 pathway through p53 accumulation, expression of specific p53 target genes, and p53-dependent apoptosis. ChIP analyses of p21CIP1 activation in the absence of functional P-TEFb reveals the existence of two distinct kinases that phosphorylate Ser5 of the RNAP II C-terminal domain (CTD). Importantly, CTD phosphorylation at Ser2 is not required for p21CIP1 transcription, mRNA cleavage, or polyadenylation. Furthermore, recruitment of FACT requires CTD kinases, yet FACT is dispensable for p21CIP1 expression. Thus, select genes within the p53 pathway bypass the requirement for P-TEFb and RNAP II phosphorylation to trigger a cellular response to inhibition of global mRNA synthesis.
HIF-2α regulates Oct-4: effects of hypoxia on stem cell function, embryonic development, and tumor growth
Authors: Covello Kelly L. , Kehler James , Yu Hongwei , Gordan John D. , Arsham Andrew M. , Hu Cheng-Jun , Labosky Patricia A. , Simon M. Celeste , Keith Brian
Published: March 2006

The division, differentiation, and function of stem cells and multipotent progenitors are influenced by complex signals in the microenvironment, including oxygen availability. Using a genetic “knock-in” strategy, we demonstrate that targeted replacement of the oxygen-regulated transcription factor HIF-1α with HIF-2α results in expanded expression of HIF-2α-specific target genes including Oct-4, a transcription factor essential for maintaining stem cell pluripotency. We show that HIF-2α, but not HIF-1α, binds to theOct-4promoter and inducesOct-4expression and transcriptional activity, thereby contributing to impaired development in homozygousHif-2αKI/KIembryos, defective hematopoietic stem cell differentiation in embryoid bodies, and large embryonic stem cell (ES)-derived tumors characterized by altered cellular differentiation. Furthermore, loss of HIF-2α severely reduces the number of embryonic primordial germ cells, which requireOct-4expression for survival and/or maintenance. These results identifyOct-4as a HIF-2α-specific target gene and indicate that HIF-2α can regulate stem cell function and/or differentiation through activation ofOct-4, which in turn contributes to HIF-2α's tumor promoting activity.
The pathogenesis of immune thrombocytopaenic purpura
Authors: Cooper Nichola , Bussel James
Published: March 2006

Immune thrombocytopaenic purpura (ITP) is an autoimmune bleeding disease that is rarely fatal. However, in many adults treatment is unsatisfactory, with as much morbidity from the immunosuppressive effects of treatment as from bleeding. Identifying the underlying disease process should help us to identify more targeted therapies and improve not only the treatment but also the quality of life of patients with this disorder.
The neurophysiology of osseointegrated oral implants. A clinically underestimated aspect
Authors: ABARCA M. , VAN STEENBERGHE D. , MALEVEZ C. , JACOBS R.
Published: March 2006

summary  Amputation of a limb or a tooth leads to the loss of a large number of exteroceptors. These play an important role in sensory perception and feedback, which tune the motor control. Even after rehabilitation with a prosthetic device, tactile function remains impaired. This can present a subsequent risk of overloading the prosthesis. The peri‐implant interface of oral osseointegrated implants is characterized by the absence of a periodontal ligament, a crucial difference towards the natural dentition, from a biomechanical but also from a neurophysiologic point of view. Patients rehabilitated with osseointegrated implants seem subjectively not much impaired in their masticatory and other oral functions. This observation might be linked to the presence of some peripheral feedback pathway to the sensory cortex. It may be attributed to the activation of receptors in the peri‐implant environment (either bone or periosteum). This phenomenon called osseoperception, when it relates to the consciousness of the applied stimuli, has been described for both oral and skeletal implants. In the present review, besides osseoperecption other neurophysiological aspects of oral implants, such their reflex function, will be outlined and their clinical meaning pointed out.
Evaluation of genetic damage in human peripheral lymphocytes exposed to mineral trioxide aggregate and Portland cements
Authors: BRAZ M. G. , CAMARGO E. A. , SALVADORI D. M. F. , MARQUES M. E. A. , RIBEIRO D. A.
Published: March 2006

summary  Mineral trioxide aggregate (MTA) and Portland cement are being used in dentistry as root‐end‐filling material for periapical surgery and for the sealing of communications between the root canal system and the surrounding tissues. However, genotoxicity tests for complete risk assessment of these compounds have not been conducted up to now. In the present study, the genotoxic effects of MTA and Portland cements were evaluated in peripheral lymphocytes from 10 volunteers by the alkaline single cell gel (comet) assay. The results pointed out that the single cell gel (comet) assay failed to detect the presence of DNA damage after a treatment of peripheral lymphocytes by MTA and Portland cements for concentrations up to 1000 μg mL−1. In summary, our results indicate that exposure to MTA or Portland cements may not be a factor that increases the level of DNA lesions in human peripheral lymphocytes as detected by single cell gel (comet) assay.
Comparison of digital and spectrophotometric measurements of colour shade guides
Authors: CAL E. , GÜNERI P. , KOSE T.
Published: March 2006

summary  Colour determination may be precisely performed with spectrophotometers, however this technique is time consuming and requires special devices. The aim of this study was to determine the correlation between the colour measurements performed on three shade guides of the same manufacturer, obtained with digital and spectrophotometric analyses. Sixty shade tabs of three shade guides were analysed to determine the L*, a*, b* values with a spectrophotometer. The same shade tabs were also imaged with a digital camera, and same colour parameters were determined on the digital images with the software. Three measurements for each shade tab were made with both methods. Statistical differences between the results of two methods were determined using anova (α = 0·05). Additionally, presence of any correlation between two methods within each guide was investigated by using correlation analysis. It was observed that the a* and b* values obtained by digital method were correlated with those of the spectrophotometer. Spectrophotometric analysis revealed no disparities among the L* measurements of the shade tabs of three shade guides (P > 0·05), but significant differences were observed between the L* values of the shade tabs with digital analysis method (P = 0·000). The digital method formed more subgroups than the spectrophotometer. For a* and b* values, both methods revealed significant differences among the shade tabs (P < 0·05). Additionally, both analyses revealed that colour characteristics of the first shade guide was different than the other two. The results obtained by digital method were correlated with those of the spectrophotometer, especially for a* and b* values.
Quantitative analysis of allergenic ingredients in eluate extracted from used denture base resin
Authors: MIKAI M. , KOIKE M. , FUJII H.
Published: March 2006

summary  Allergenic ingredients extracted from used denture base resin were quantitatively analysed using a gas chromatograph/mass spectrometer and high performance liquid chromatography. Methyl methacrylate (MMA), hydroquinone (HQ), formaldehyde (FMA), benzoyl peroxide (BPO), benzoic acid (BA) and methyl benzoate (MB) were detected in a eluate from all of the dentures, while ethyleneglycol dimethacrylate (DME) was detected in the eluate from 87 dentures in use for <15 years. MMA, HQ, FMA, BPO and MB showed a decrease in correlation to the period of denture use, but continuing to be evident even after 29 years of use. The MMA showed the highest level of elution and a relatively moderate decrease over time. The elution of BA, on the contrary, showed an increase with the period of denture use up to about 10 years and subsequently reached a plateau. Our results indicate that purported allergens exist in the resin base and can be eluted into the oral cavity, even in patients using an old denture for a period of nearly 30 years.
Craniofacial morphometric analysis of mandibular prognathism
Authors: CHANG H. P. , LIU P. H. , YANG Y. H. , LIN H. C. , CHANG C. H.
Published: March 2006

summary The purpose of this study was to provide more information about the morphological characteristics of the craniofacial complex in mandibular prognathism. Forty young adult males having mandibular prognathism were compared with 40 having normal occlusion. This was conducted to carry out geometric morphometric assessments to localize alterations, using Procrustes analysis and thin‐plate spline analysis, in addition to conventional cephalometric techniques. Procrustes analysis indicated that the mean craniofacial, midfacial and mandibular morphology was significantly different in prognathic subjects compared with normal controls. This finding was corroborated by the multivariate HotellingT2‐test of cephalometric variables. Mandibular prognathism demonstrated a shorter and slightly retropositioned maxilla, a greater total length and anterior positioning of the mandible. Thin‐plate spline analysis revealed a developmental diminution of the palatomaxillary region anteroposteriorly and a developmental elongation of the mandible anteroposteriorly, leading to the appearance of a prognathic mandibular profile. In conclusion, thin‐plate spline analysis seems to provide a valuable supplement for conventional cephalometric analysis because the complex patterns of craniofacial shape change are visualized suggestive by means of grid deformations.