bluuebunny/crossref_metadata_2025 huggingface dataset:
The Type IV Secretion System Component VirB5 Binds to the
trans
-Zeatin Biosynthetic Enzyme Tzs and Enables Its Translocation to the Cell Surface of
Agrobacterium tumefaciens
Authors: Aly Khaled Ahmed , Krall Lilian , Lottspeich Friedrich , Baron Christian
Published: December 2007

VirB5 is a minor component of the extracellular T pilus determined by the
Agrobacterium tumefaciens
type IV secretion system. To identify proteins that interact with VirB5 during the pilus assembly process, we purified VirB5 as a recombinant fusion protein and, by using a gel overlay assay, we detected a 26-kDa interacting protein in
Agrobacterium
cell lysates. The VirB5-binding protein was purified from
A. tumefaciens
and identified as the cytokinin biosynthetic enzyme Tzs. The VirB5-Tzs interaction was confirmed using pulldown assays with purified proteins and the yeast two-hybrid system. An analysis of the subcellular localization in
A. tumefaciens
showed that Tzs was present in the soluble as well as the membrane fraction. Tzs was extracted from the membranes with the mild detergent dodecyl-β-
d
-maltoside in complexes of different molecular masses, and this association was strongly reduced in the absence of VirB5. Using immunoelectron microscopy, we also detected Tzs on the
Agrobacterium
cell surface. A functional type IV secretion system was required for efficient translocation to the surface, but Tzs was not secreted into the cell supernatant. The fact that Tzs localizes on the cell surface suggests that it may contribute to the interaction of
Agrobacterium
with plants.
Nanoarchaeum equitansandIgnicoccus hospitalis: New Insights into a Unique, Intimate Association of Two Archaea
Authors: Jahn Ulrike , Gallenberger Martin , Paper Walter , Junglas Benjamin , Eisenreich Wolfgang , Stetter Karl O. , Rachel Reinhard , Huber Harald
Published: December 2007

Nanoarchaeum equitansandIgnicoccus hospitalisrepresent a unique, intimate association of two archaea. Both form a stable coculture which is mandatory forN.equitansbut not for the hostI.hospitalis. Here, we investigated interactions and mutual influence between these microorganisms. Fermentation studies revealed that during exponential growth only about 25% ofI.hospitaliscells are occupied byN.equitanscells (one to three cells). The latter strongly proliferate in the stationary phase ofI.hospitalis, until 80 to 90% of theI.hospitaliscells carry around 10N.equitanscells. Furthermore, the expulsion of H2S, the major metabolic end product ofI.hospitalis, by strong gas stripping yields huge amounts of freeN.equitanscells.N.equitanshad no influence on the doubling times, final cell concentrations, and growth temperature, pH, or salt concentration ranges or optima ofI.hospitalis. However, isolation studies using optical tweezers revealed that infection withN.equitansinhibited the proliferation of individualI.hospitaliscells. This inhibition might be caused by deprivation of the host of cell components like amino acids, as demonstrated by13C-labeling studies. The strong dependence ofN.equitansonI.hospitaliswas affirmed by live-dead staining and electron microscopic analyses, which indicated a tight physiological and structural connection between the two microorganisms. No alternative hosts, including otherIgnicoccusspecies, were accepted byN.equitans. In summary, the data show a highly specialized association ofN.equitansandI.hospitaliswhich so far cannot be assigned to a classical symbiosis, commensalism, or parasitism.
Novel Multiplex PCR Assay for Simultaneous Identification of Community-Associated Methicillin-Resistant
Staphylococcus aureus
Strains USA300 and USA400 and Detection of
mecA
and Panton-Valentine Leukocidin Genes, with Discrimination of
Staphylococcus aureus
from Coagulase-Negative Staphylococci
Authors: Zhang Kunyan , McClure Jo-Ann , Elsayed Sameer , Louie Thomas , Conly John M.
Published: December 2007

We developed a novel multiplex PCR assay for rapid identification and discrimination of the USA300 and USA400 strains and concomitant detection of Panton-Valentine leukocidin genes, with simultaneous discrimination of methicillin-resistant
Staphylococcus aureus
strains from methicillin-susceptible
S. aureus
strains,
S. aureus
strains from coagulase-negative staphylococci, and staphylococci from other bacteria.
Comparative Study of Virulence Traits of
Escherichia coli
Clinical Isolates Causing Early and Late Neonatal Sepsis
Authors: Soto S. M. , Bosch J. , Jimenez de Anta M. T. , Vila J.
Published: December 2007

Neonatal meningitis and septicemia caused by
Escherichia coli
are still major health problems in industrialized countries. Forty-seven
E. coli
strains causing neonatal sepsis were analyzed. Twenty-two and 25 strains caused early (detected from 0 to 3 days after birth) and late (detected from 4 to 28 days after birth) infections, respectively. Only the
ibeA
gene was significantly more prevalent in the strains causing early infections.
Contribution of the (1→3)-β-
d
-Glucan Assay for Diagnosis of Invasive Fungal Infections
Authors: Persat Florence , Ranque Stéphane , Derouin Francis , Michel-Nguyen Annie , Picot Stéphane , Sulahian Annie
Published: December 2007

Diagnosis of invasive fungal infection (IFI) remains a challenge. A retrospective study was performed on 279 patients at three French university hospitals to evaluate the performance of the (1→3)-β-
d
-glucan assay (BG assay; Fungitell; Associates of Cape Cod, Inc.) for the diagnosis of IFI. The results of one serum per subject were analyzed for 117 patients who had probable or proven IFI according to the European Organization for Research and Treatment of Cancer criteria (70 invasive pulmonary aspergilloses [IPA], 27 fungal bloodstream infections, and 20
Pneumocystis jiroveci
pneumonias), 40 blood donors, and 122 patients who were hospitalized in hematology wards or intensive care units and were at risk for IFI but in whom IFI had not been diagnosed. For the overall IFI diagnosis, the BG assay had 77.8% sensitivity and specificities of 92.5 and 70.5% for blood donors and patients at risk, respectively. The assay was positive in 48 patients with IPA (68%), in 23 with bloodstream infections (85.2%), and in all who had
P. jiroveci
pneumonias (100%), and the false-positive rate varied depending on the controls used. It allowed a higher rate of detection among IPA patients compared to the galactomannan enzyme-linked immunosorbent assay (ELISA) (48 versus 39 patients, respectively) and among candidemia patients compared to the mannan ELISA (20 versus 11 patients, respectively). This assay therefore appears to be useful in the diagnosis of IFI, particularly for serum analysis of pneumocystosis pneumonia patients, but further studies are needed to evaluate false-positive rates and its future role in IFI diagnosis.
Novel Loop-Mediated Isothermal Amplification Method for Detection of the JP2 Clone of
Aggregatibacter actinomycetemcomitans
in Subgingival Plaque
Authors: Seki Mitsuko , Poulsen Knud , Haubek Dorte , Kilian Mogens
Published: December 2007

We developed a loop-mediated isothermal amplification method that detects the JP2 clone of
Aggregatibacter actinomycetemcomitans
, which induces aggressive periodontitis in adolescents of North and West African descents. Being independent of special equipment, this specific and sensitive method offers significant advantages for screening of patients on a population basis and in clinical settings.
Temporal Association of the Herpes Simplex Virus Genome with Histone Proteins during a Lytic Infection
Authors: Oh Jaewook , Fraser Nigel W.
Published: December 2007

Previous work has determined that there are nucleosomes on the herpes simplex virus (HSV) genome during a lytic infection but that they are not arranged in an equally spaced array like in cellular DNA. However, like in cellular DNA, the promoter regions of several viral genes have been shown to be associated with nucleosomes containing modified histone proteins that are generally found associated with actively transcribed genes. Furthermore, it has been found that the association of modified histones with the HSV genome can be detected at the earliest times postinfection (1 h postinfection) and increases up to 3 h postinfection. However from 3 h to 6 h postinfection (the late phase of the replication cycle), the association decreases. In this study we have examined histone association with promoter regions of all kinetic classes of genes. This was done over the time course of an infection in Sy5y cells using sucrose gradient sedimentation, bromodeoxyuridine labeling, chromatin immunoprecipitation assays, Western blot analysis, trypsin and DNase digestion, and quantitative real-time PCR. Because no histones were detected inside HSV type 1 capsids, the viral genome probably starts to associate with histones after being transported from infecting virions into the host nucleus. Promoter regions of all gene classes (immediate early, early, and late) bind with histone proteins at the start of viral gene expression. However, after viral DNA replication initiates, histones appear not to associate with newly synthesized viral genomes.
Human Immunodeficiency Virus Type 1 Vpr Induces Cell Cycle G
2
Arrest through Srk1/MK2-Mediated Phosphorylation of Cdc25
Authors: Huard Sylvain , Elder Robert T. , Liang Dong , Li Ge , Zhao Richard Y.
Published: December 2007

Human immunodeficiency virus type 1 (HIV-1) Vpr induces cell cycle G
2
arrest in fission yeast (
Schizosaccharomyces pombe
) and mammalian cells, suggesting the cellular pathway(s) targeted by Vpr is conserved among eukaryotes. Our previous studies in fission yeast demonstrated that Vpr induces G
2
arrest in part through inhibition of Cdc25, a Cdc2-specific phosphatase that promotes G
2
/M transition. The goal of this study was to further elucidate molecular mechanism underlying the inhibitory effect of Vpr on Cdc25. We show here that, similar to the DNA checkpoint controls, expression of
vpr
promotes subcellular relocalization of Cdc25 from nuclear to cytoplasm and thereby prevents activation of Cdc2 by Cdc25. Vpr-induced nuclear exclusion of Cdc25 appears to depend on the serine/threonine phosphorylation of Cdc25 and the presence of Rad24/14-3-3 protein, since amino acid substitutions of the nine possible phosphorylation sites of Cdc25 with Ala (9A) or deletion of the
rad24
gene abolished nuclear exclusion induced by Vpr. Interestingly, Vpr is still able to promote Cdc25 nuclear export in mutants defective in the checkpoints (
rad3
and
chk1/cds1
), the kinases that are normally required for Cdc25 phosphorylation and nuclear exclusion of Cdc25, suggesting that others kinase(s) might modulate phosphorylation of Cdc25 for the Vpr-induced G
2
arrest. We report here that this kinase is Srk1. Deletion of the
srk1
gene blocks the nuclear exclusion of Cdc25 caused by Vpr. Overexpression of
srk1
induces cell elongation, an indication of cell cycle G
2
delay, in a similar fashion to Vpr; however, no additive effect of cell elongation was observed when
srk1
and
vpr
were coexpressed, indicating Srk1 and Vpr are likely affecting the cell cycle G
2
/M transition through the same cellular pathway. Immunoprecipitation further shows that Vpr and Srk1 are part of the same protein complex. Consistent with our findings in fission yeast, depletion of the
MK2
gene, a human homologue of Srk1, either by small interfering RNA or an MK2 inhibitor suppresses Vpr-induced cell cycle G
2
arrest in mammalian cells. Collectively, our data suggest that Vpr induces cell cycle G
2
arrest at least in part through a Srk1/MK2-mediated mechanism.
Hepatitis C Virus Core Protein Upregulates Serine Phosphorylation of Insulin Receptor Substrate-1 and Impairs the Downstream Akt/Protein Kinase B Signaling Pathway for Insulin Resistance
Authors: Banerjee Sutapa , Saito Kousuke , Ait-Goughoulte Malika , Meyer Keith , Ray Ratna B. , Ray Ranjit
Published: December 2007

Chronic hepatitis C virus (HCV) infection has a significantly increased prevalence of type 2 diabetes mellitus (T2DM). Insulin resistance is a critical component of T2DM pathogenesis. Several mechanisms are likely to be involved in the pathogenesis of HCV-related insulin resistance. Since we and others have previously observed that HCV core protein activates c-Jun N-terminal kinase (JNK) and mitogen-activated protein kinase, we examined the contribution of these pathways to insulin resistance in hepatocytes. Our experimental findings suggest that HCV core protein alone or in the presence of other viral proteins increases Ser
312
phosphorylation of the insulin receptor substrate-1 (IRS-1). Hepatocytes infected with cell culture-grown HCV genotype 1a or 2a displayed a significant increase in the Ser
473
phosphorylation status of the Ser/Thr kinase protein kinase B (Akt/PKB), while Thr
308
phosphorylation was not significantly altered. HCV core protein-mediated Ser
312
phosphorylation of IRS-1 was inhibited by JNK (SP600125) and phosphatidylinositol-3 kinase (LY294002) inhibitors. A functional assay also suggested that hepatocytes expressing HCV core protein alone or infected with cell culture-grown HCV exhibited a suppression of 2-deoxy-
d
-[
3
H]glucose uptake. Inhibition of the JNK signaling pathway significantly restored glucose uptake despite HCV core expression in hepatocytes. Taken together, our results demonstrated that HCV core protein increases IRS-1 phosphorylation at Ser
312
which may contribute in part to the mechanism of insulin resistance.
Genetic Determinants of Sindbis Virus Mosquito Infection Are Associated with a Highly Conserved Alphavirus and Flavivirus Envelope Sequence
Authors: Pierro Dennis J. , Powers Erik L. , Olson Ken E.
Published: December 2007

Wild-type Sindbis virus (SINV) strain MRE16 efficiently infects
Aedes aegypti
midgut epithelial cells (MEC), but laboratory-derived neurovirulent SINV strain TE/5′2J infects MEC poorly. SINV determinants for MEC infection have been localized to the E2 glycoprotein. The E2 amino acid sequences of MRE16 and TE/5′2J differ at 60 residue sites. To identify the genetic determinants of MEC infection of MRE16, the TE/5′2J virus genome was altered to contain either domain chimeras or more focused nucleotide substitutions of MRE16. The growth patterns of derived viruses in cell culture were determined, as were the midgut infection rates (MIR) in
A. aegypti
mosquitoes. The results showed that substitutions of MRE16 E2 aa 95 to 96 and 116 to 119 into the TE/5′2J virus increased MIR both independently and in combination with each other. In addition, a unique PPF/.GDS amino acid motif was located between these two sites that was found to be a highly conserved sequence among alphaviruses and flaviviruses but not other arboviruses.
Replication of ICP0-Null Mutant Herpes Simplex Virus Type 1 Is Restricted by both PML and Sp100
Authors: Everett Roger D. , Parada Carlos , Gripon Philippe , Sirma Hüseyin , Orr Anne
Published: December 2007

Herpes simplex virus type 1 (HSV-1) mutants that fail to express the viral immediate-early protein ICP0 have a pronounced defect in viral gene expression and plaque formation in limited-passage human fibroblasts. ICP0 is a RING finger E3 ubiquitin ligase that induces the degradation of several cellular proteins. PML, the organizer of cellular nuclear substructures known as PML nuclear bodies or ND10, is one of the most notable proteins that is targeted by ICP0. Depletion of PML from human fibroblasts increases ICP0-null mutant HSV-1 gene expression, but not to wild-type levels. In this study, we report that depletion of Sp100, another major ND10 protein, results in a similar increase in ICP0-null mutant gene expression and that simultaneous depletion of both proteins complements the mutant virus to a greater degree. Although chromatin assembly and modification undoubtedly play major roles in the regulation of HSV-1 infection, we found that inhibition of histone deacetylase activity with trichostatin A was unable to complement the defect of ICP0-null mutant HSV-1 in either normal or PML-depleted human fibroblasts. These data lend further weight to the hypothesis that ND10 play an important role in the regulation of HSV-1 gene expression.
EXCITON IN ZnO FILM
Authors: ZHAO HUA , XIONG WEN , ZHU MENG-ZAO
Published: December 2007

The present study variationally calculates the ground state energy and the first excited energy of an exciton in an ZnO film in effective mass approximation. Change of the ground state energy, the first excited energy of an exciton, and radius of the exciton with film thickness are studied, as well as the correction due to the quantum tunneling of the exciton wave function through the film.
HIGH HARMONIC GENERATION IN ATOMS, MOLECULES AND NANOSTRUCTURES
Authors: ZHANG G. P.
Published: December 2007

High harmonic generation (HHG) has attracted tremendous attention over two decades. This paper presents a brief review of high harmonic generation in atoms, molecules and nanostructures. Through a simple classical model, we first review a few important concepts in HHG. We then discuss how the electric field initial phase affects the harmonic yield, and explain the reason behind the cutoff law. We focus on HHG in nanostructures. As an example, a detailed calculation in C 60 is presented.
EFFECTS OF Ge DOPING THROUGH ION IMPLANTATION ON THE STRUCTURAL AND OPTICAL PROPERTIES OF ZnO THIN FILMS PREPARED BY SOL-GEL TECHNIQUE
Authors: XUE S. W. , ZU X. T. , XIANG X. , CHEN M. Y. , ZHENG W. G.
Published: December 2007

ZnO thin films were first prepared by the sol–gel process, and then Ge ions were implanted into the ZnO films. The effects of ion implantation on the structural and optical properties of the ZnO films were investigated by X-ray diffraction, photoluminescence (PL), and optical transmittance measurements. Measurement results showed that the intensity of the (002) diffraction peak was decreased and the full width at half maximum was narrowed. PL emission was greatly extinguished after Ge ion implantation. Both the near band edge (NBE) excitonic UV emission at 391 nm and the defect related deep level emission centered at 470 nm in the visible region were decreased after Ge ion implantation. NBE peak and the absorption edge were observed to have a blueshift toward higher energy.
HIERARCHICAL QUANTUM SEARCH
Authors: KOREPIN VLADIMIR E. , XU YING
Published: December 2007

Database search has wide applications and is used as a subroutine in many important algorithms. In this paper, we will consider a database with a single target item. Quantum algorithm (Grover) locates the target item faster than any classical algorithm. In addition to a full (Grover) search, it frequently occurs that one is looking for a group of items (a block) containing the target item, rather than the target item itself. This problem is known as partial search. As a generalization of the full search, partial search is of particular importance in practice. Partial search trades accuracy for speed, i.e., it works faster than a full search. There exists different versions of partial search. We will study the optimized version of the algorithm discovered by Grover and Radhakrishnan and call it GRK. GRK can be applied successively (in a sequence). First, the database is partitioned into blocks and GRK is applied to find the target block. This target block is then partitioned into subblocks and GRK is used again to find the target subblock. This procedure can be repeated if the database is large enough. (This sequence of GRK's is called a hierarchy.) Another possibility is to partition the database into subblocks directly and use GRK to find the target subblock once. In this paper, we will prove that the latter is faster (makes less queries to the Oracle).
CHAOS SYNCHRONIZATION IN MULTIPLE TIME DELAY ELECTROOPTICAL SEMICONDUCTOR LASERS
Authors: SHAHVERDIEV E. M. , SHORE K. A.
Published: December 2007

We make the first report of the chaos synchronization regimes in the unidirectionally-coupled multiple time delay electrooptical semiconductor lasers. We derive the existence and sufficient stability conditions for the synchronization regimes. Analytical results are fully supported by numerical simulations.
STABLE DETERMINATION OF A BOUNDARY COEFFICIENT IN AN ELLIPTIC EQUATION
Authors: CHENG JIN , CHOULLI MOURAD , LIN JUNSHAN
Published: December 2007

We prove a logarithmic stability estimate for a Cauchy problem associated with a second order elliptic operator. Our proof is essentially based on a Carleman estimate by A. L. Bukhgeim. This result is applied to establish a stability estimate for the inverse problem of determining a boundary coefficient (or a boundary function) by a single boundary measurement. This kind of inverse problems is motivated by the corrosion detection problem.
MOMENTS AND COMMUTATORS OF PROBABILITY MEASURES
Authors: ACCARDI LUIGI , KUO HUI-HSIUNG , STAN AUREL
Published: December 2007

Let a0, a- and a+ be the preservation, annihilation, and creation operators of a probability measure μ on ℝd, respectively. The operators a0 and [a-, a+] are proven to uniquely determine the moments of μ. We discuss the question: "What conditions must two families of operators satisfy, in order to ensure the existence of a probability measure, having finite moments of any order, so that, its associated preservation operators and commutators between the annihilation and creation operators are the given families of operators?" For the case d = 1, a satisfactory answer to this question is obtained as a simple condition in terms of the Szegö–Jacobi parameters. For the multidimensional case, we give some necessary conditions for the answer to this question. We also give a table with the associated preservation and commutator between the annihilation and creation operators, for some of the classic probability measures on ℝ.
POISSON–MASLOV TYPE FORMULAS FOR SCHRÖDINGER EQUATIONS WITH MATRIX-VALUED POTENTIALS
Authors: SHAMAROV N. N.
Published: December 2007

Cauchy problems for Schrödinger equations with matrix-valued potentials are explicitly solved under following assumptions:.

— equations are written in momentum form;.

— the potentials are Fourier transformed matrix-valued measures with, in general, noncommuting values;.

— initial Cauchy data are good enough.

The solutions at time t are presented in form of integrals over some spaces of piecewise continuous mappings of the segment [0, t] to a finite-dimensional momentum space, and measures of the integration are countably additive but matrix-valued (resulting in matrices of ordinary Lebesgue integrals).

Known results gave solutions either when the measure had commuting (complex) values, or when the integration over infinite dimensional spaces was quite symbolic, or when such integrating was of chronological type and hence more complicated.

The method used below is based on technique of matrix-valued transition amplitudes.
A REMARK ON THE GENERATOR OF A RIGHT-CONTINUOUS MARKOV PROCESS
Authors: RÖCKNER MICHAEL , TRUTNAU GERALD
Published: December 2007

Given a right-continuous Markov process (Xt)t ≥ 0 on a second countable metrizable space E with transition semigroup (pt)t ≥ 0, we prove that there exists a σ-finite Borel measure μ with full support on E, and a closed and densely defined linear operator [Formula: see text] generating (pt)t ≥ 0 on Lp (E; μ). In particular, we solve the corresponding Cauchy problem in Lp (E; μ) for any initial condition [Formula: see text]. Furthermore, for any real β > 0 we show that there exists a generalized Dirichlet form which is associated to (e-βt pt)t ≥ 0. If the β-subprocess of (Xt)t ≥ 0 corresponding to (e-βt pt)t ≥ 0, β > 0, is μ-special standard then all results from generalized Dirichlet form theory become available, and Fukushima's decomposition holds for [Formula: see text]. If (Xt)t ≥ 0 is transient, then β can be chosen to be zero.
IDENTIFYING GENE-SPECIFIC VARIATIONS IN BIOMEDICAL TEXT
Authors: KLINGER ROMAN , FRIEDRICH CHRISTOPH M. , MEVISSEN HEINZ THEODOR , FLUCK JULIANE , HOFMANN-APITIUS MARTIN , FURLONG LAURA I. , SANZ FERRAN
Published: December 2007

The influence of genetic variations on diseases or cellular processes is the main focus of many investigations, and results of biomedical studies are often only accessible through scientific publications. Automatic extraction of this information requires recognition of the gene names and the accompanying allelic variant information. In a previous work, the OSIRIS system for the detection of allelic variation in text based on a query expansion approach was communicated. Challenges associated with this system are the relatively low recall for variation mentions and gene name recognition. To tackle this challenge, we integrate the ProMiner system developed for the recognition and normalization of gene and protein names with a conditional random field (CRF)-based recognition of variation terms in biomedical text. Following the newly developed normalization of variation entities, we can link textual entities to Single Nucleotide Polymorphism database (dbSNP) entries. The performance of this novel approach is evaluated, and improved results in comparison to state-of-the-art systems are reported.
RETRIEVING MUTATION-SPECIFIC INFORMATION FOR HUMAN PROTEINS IN UNIPROT/SWISS-PROT KNOWLEDGEBASE
Authors: YIP YUM LINA , LACHENAL NATHALIE , PILLET VIOLAINE , VEUTHEY ANNE-LISE
Published: December 2007

The UniProt/Swiss-Prot Knowledgebase records about 30,500 variants in 5,664 proteins (Release 52.2). Most of these variants are manually curated single amino acid polymorphisms (SAPs) with references to the literature. In order to keep the list of published documents related to SAPs up to date, an automatic information retrieval method is developed to recover texts mentioning SAPs. The method is based on the use of regular expressions (patterns) and rules for the detection and validation of mutations. When evaluated using a corpus of 9,820 PubMed references, the precision of the retrieval was determined to be 89.5% over all variants. It was also found that the use of nonstandard mutation nomenclature and sequence positional correction is necessary to retrieve a significant number of relevant articles. The method was applied to the 5,664 proteins with variants. This was performed by first submitting a PubMed query to retrieve articles using gene or protein names and a list of mutation-related keywords; the SAP detection procedure was then used to recover relevant documents. The method was found to be efficient in retrieving new references on known polymorphisms. New references on known SAPs will be rendered accessible to the public via the Swiss-Prot variant pages.
Role of PKC-δ on substance P-induced chemokine synthesis in pancreatic acinar cells
Authors: Ramnath Raina Devi , Sun Jia , Adhikari Sharmila , Zhi Liang , Bhatia Madhav
Published: December 2007

Interaction of the neuropeptide substance P (SP) with its high-affinity neurokinin-1 receptor (NK1R) plays an important role in the pathophysiology of acute pancreatitis. SP is known to stimulate the production of chemokines monocyte chemoattractant protein-1 (MCP-1), macrophage inflammatory protein (MIP)-1α, and MIP-2 in pancreatic acinar cells via the activation of NF-κB. However, the signaling mechanisms by which the SP-NK1R interaction induces NF-κB activation and chemokine production remain unclear. To that end, in the present study, we investigated the participation of PKC in SP-induced chemokine production in pancreatic acinar cells. In this study, we showed that SP stimulated an early phosphorylation of PKC isoform PKC-δ followed by increased activation of MAPKKK MEKK1 and MAPK ERK and JNK as well as transcription factor NF-κB and activator protein-1 driven chemokine production. Depletion of PKC-δ with its inhibitor rottlerin or the specific PKC-δ translocation inhibitor peptide dose dependently decreased SP-induced PKC-δ, MEKK1, ERK, JNK, NF-κB, and AP-1 activation. Moreover, rottlerin as well as PKC-δ translocation inhibitor inhibited SP-induced chemokine production in a concentration-dependent manner. We also demonstrated that PKC-δ activation was attenuated by CP96345, a selective NK1R antagonist, thus showing that PKC-δ activation was indeed mediated by SP in pancreatic acinar cells. These results show that PKC-δ is an important proinflammatory signal transducer for SP-NK1R-induced chemokine production in pancreatic acinar cells.
Acetoacetate and β-hydroxybutyrate in combination with other metabolites release insulin from INS-1 cells and provide clues about pathways in insulin secretion
Authors: MacDonald Michael J. , Longacre Melissa J. , Stoker Scott W. , Brown Laura J. , Hasan Noaman M. , Kendrick Mindy A.
Published: December 2007

Mitochondrial anaplerosis is important for insulin secretion, but only some of the products of anaplerosis are known. We discovered novel effects of mitochondrial metabolites on insulin release in INS-1 832/13 cells that suggested pathways to some of these products. Acetoacetate, β-hydroxybutyrate, α-ketoisocaproate (KIC), and monomethyl succinate (MMS) alone did not stimulate insulin release. Lactate released very little insulin. When acetoacetate, β-hydroxybutyrate, or KIC were combined with MMS, or either ketone body was combined with lactate, insulin release was stimulated 10-fold to 20-fold the controls (almost as much as with glucose). Pyruvate was a potent stimulus of insulin release. In rat pancreatic islets, β-hydroxybutyrate potentiated MMS- and glucose-induced insulin release. The pathways of their metabolism suggest that, in addition to producing ATP, the ketone bodies and KIC supply the acetate component and MMS supplies the oxaloacetate component of citrate. In line with this, citrate was increased by β-hydroxybutyrate plus MMS in INS-1 cells and by β-hydroxybutyrate plus succinate in mitochondria. The two ketone bodies and KIC can also be metabolized to acetoacetyl-CoA and acetyl-CoA, which are precursors of other short-chain acyl-CoAs (SC-CoAs). Measurements of SC-CoAs by LC-MS/MS in INS-1 cells confirmed that KIC, β-hydroxybutyrate, glucose, and pyruvate increased the levels of acetyl-CoA, acetoacetyl-CoA, succinyl-CoA, hydroxymethylglutaryl-CoA, and malonyl-CoA. MMS increased incorporation of 14C from β-hydroxybutyrate into citrate, acid-precipitable material, and lipids, suggesting that the two molecules complement one another to increase anaplerosis. The results suggest that, besides citrate, some of the products of anaplerosis are SC-CoAs, which may be precursors of molecules involved in insulin secretion.
Acylation-stimulating protein deficiency and altered adipose tissue in alternative complement pathway knockout mice
Authors: Paglialunga Sabina , Fisette Alexandre , Yan Yafeng , Deshaies Yves , Brouillette Jean-Francois , Pekna Marcela , Cianflone Katherine
Published: December 2007

Acylation-stimulating protein (C3adesArg/ASP) is an adipokine that acts on its receptor C5L2 to stimulate triglyceride (TG) synthesis in adipose tissue. The present study investigated ASP levels in mouse models of obesity and leanness and the effect of ASP deficiency in C3 knockout (C3KO) mice on adipose tissue morphology. Plasma ASP levels in wild-type (WT) mice correlated positively with plasma nonesterified fatty acids (NEFA) ( R = 0.664, P < 0.001) and total cholesterol ( R = 0.515, P < 0.001). Plasma ASP was increased by 85% in obese ob/ob leptin-deficient mice and decreased in lean diacylglycerol acyltransferase 1 (DGAT1) KO mice (−54%) and C/EBPαβ/βtransgenic mice (−70%) compared with WT. Mice lacking alternative complement factor B or adipsin (FBKO or ADKO), required for ASP production, were also ASP deficient. Both FBKO and C3KO mice had delayed postprandial TG and NEFA clearance on low-fat (LF) and high-fat (HF) diets, suggesting that lack of ASP, not C3, drives the metabolic phenotype. Adipocyte size distribution in C3KO mice was polarized (increased number of both small and large cells), with decreased adipsin expression (−33% gonadal HF), DGAT1 expression (−31% to −50%) and DGAT activity (−41%). Overall, a reduction/deficiency in ASP is associated with an antiadipogenic state and ASP may provide a target for controlling fat storage.
Adverse effects of pneumoperitoneum on renal function: involvement of the endothelin and nitric oxide systems
Authors: Abassi Zaid , Bishara Bishara , Karram Tony , Khatib Samer , Winaver Joseph , Hoffman Aaron
Published: December 2007

Increased intra-abdominal pressure (IAP) during laparoscopy adversely affects kidney function. The mechanism underlying this phenomenon is largely unknown. This study was designed to investigate the involvement of endothelin (ET)-1 and nitric oxide (NO) systems in IAP-induced renal dysfunction. Rats were subjected to IAP of 14 mmHg for 1 h, followed by a deflation for 60 min (recovery). Four additional groups were pretreated with 1) ABT-627, an ETAantagonist; 2) A-192621, an ETBantagonist; 3) nitroglycerine; and 4) NG-nitro-l-arginine methyl ester, a NO synthase inhibitor, before IAP. Urine flow rate (V), absolute Na+excretion (UNaV), glomerular filtration rate (GFR), and renal plasma flow (RPF) were determined. Significant reductions in kidney function and hemodynamics were observed when IAP was applied. V decreased from 8.1 ± 1.0 to 5.8 ± 0.5 μl/min, UNaV from 1.08 ± 0.31 to 0.43 ± 0.10 μeq/min, GFR from 1.84 ± 0.12 to 1.05 ± 0.06 ml/min (−46.9 ± 2.7% from baseline), and RPF from 8.62 ± 0.87 to 3.82 ± 0.16 ml/min (−54 ± 3.5% from baseline). When the animals were pretreated with either ABT-627 or A-192621, given alone or combined, the adverse effects of IAP on GFR, RPF, V, and UNaV were significantly augmented. When the animals were pretreated with nitroglycerine, the adverse effects of pneumoperitoneum on GFR and RPF were substantially improved. In contrast, pretreatment with NG-nitro-l-arginine methyl ester remarkably aggravated pneumoperitoneum-induced renal dysfunction. In conclusion, decreased renal excretory function and hypofiltration are induced by increased IAP. These effects are related to impairment of renal hemodynamics and could be partially ameliorated by pretreatment with nitroglycerine and aggravated by NO and ET blockade.
Bradykinin modulates focal adhesions and induces stress fiber remodeling in renal papillary collecting duct cells
Authors: Márquez María Gabriela , del Carmen Fernández-Tome María , Favale Nicolás Octavio , Pescio Lucila Gisele , Sterin-Speziale Norma Beatriz
Published: December 2007

Focal adhesions (FAs) are specialized regions of cell attachment to the extracellular matrix. Previous works have suggested that bradykinin (BK) can modulate cell-matrix interaction. In the present study, we used a physiological cellular model to evaluate the potential role of BK in modulating FAs and stress fibers. We performed a quantitative morphometric analysis of FAs in primary cultured rat renal papillary collecting duct cells, which included size, axial ratio (shape), and average length. After 1, 5, or 10 min of incubation with BK, cultured cells were immunostained and analyzed by confocal microscopy. Although the shape of FAs was not altered, BK induced a decrease in the number of vinculin-stained FAs per cell, and a decrease in both their size and their average length, but not in talin-containing FAs, thus suggesting that BK could be inducing a restructuring of FAs. BK also induced a remodeling of the actin filament assemblies rather than their dissipation. Since we have previously demonstrated that BK stimulates activation of PLCβ in rat renal papillae, we attempted to determine whether BK can modulate FA restructuring by this mechanism, by pretreating cultured cells with the PLCβ inhibitor U73122. The present study, performed under physiological conditions with cells that were not genetically manipulated, provides new experimental evidence supporting the notion that the intrarenal hormone BK modulates FAs and actin cytoskeleton organization through a mechanism that involves the activation of PLCβ. We propose this finding as a novel mechanism for BK modulation of tubular collecting duct function.
Amelioration of oxidative mitochondrial DNA damage and deletion after renal ischemic injury by the KATP channel opener diazoxide
Authors: Sun Zhaoli , Zhang Xiuying , Ito Kazushige , Li Yulin , Montgomery Robert A. , Tachibana Shingo , Williams George Melville
Published: December 2007

Renal ischemia was induced in the rat by constriction of the renal artery for 45 min, and the ability of the ATP-sensitive K+ (KATP) channel opener diazoxide (DZ) to ameliorate renal ischemia-reperfusion (I/R) injury was evaluated. In this model, blood urea nitrogen and creatinine were elevated 2 days after I/R injury but returned closer to normal levels by 7 days after reperfusion. Histological staining for reactive oxygen species (ROS) was clearly positive and oxidized DNA, detected by the presence of the stable adduct 8-hydroxy-2′-deoxyguanosine, was clearly present in the cytoplasm of tubular cells after 1 h of reperfusion and declined 7 days after reperfusion. This finding was confirmed by ELISA, which detected 8-hydroxy-2′-deoxyguanosine in the mitochondrial fraction of kidney homogenates. Despite evidence of improved function measured by blood urea nitrogen and creatinine 7 days after reperfusion, the early changes in tubules were alarming. Mitochondrial DNA showed the common deletion, and the number of TdT-mediated dUTP nick-end label-positive tubular cells increased. Activation of caspase-3 continued, and abnormal levels of ROS were found in the mitochondrial fraction of cellular homogenates. Treatment with DZ before ischemia reduced or prevented the acute and subacute deleterious effects associated with renal I/R injury. We conclude that excess production of ROS by mitochondria on reperfusion is a major upstream event in renal reperfusion injury and that DZ functioned by preventing ROS accumulation in the mitochondria after I/R injury, thereby reducing oxidative stress as measured by the presence of oxidized mitochondrial DNA and features of apoptosis.
Inhibition of proinflammatory genes in anti-GBM glomerulonephritis by targeted dexamethasone-loaded AbEsel liposomes
Authors: Ásgeirsdóttir Sigrídur A. , Zwiers Peter J. , Morselt Henriëtte W. , Moorlag Hendrik E. , Bakker Hester I. , Heeringa Peter , Kok Jan Willem , Kallenberg Cees G. M. , Molema Grietje , Kamps Jan A. A. M.
Published: December 2007

E-selectin-directed targeted drug delivery was analyzed in anti-glomerular basement membrane glomerulonephritis. Liposomes conjugated with anti-E-selectin antibodies (AbEsel liposomes) were internalized by activated endothelial cells in vitro through E-selectin-mediated endocytosis. At the onset of glomerulonephritis in mice, E-selectin was expressed on glomerular endothelial cells, which resulted in homing of AbEsel liposomes to glomeruli after intravenous administration. Accumulation of AbEsel liposomes in the kidney was 3.6 times higher than nontargeted IgG liposomes, whereas the accumulation of both liposomes in the clearance organs liver and spleen and in heart and lungs was comparable. In glomeruli, the AbEsel liposomes colocalized with the endothelial cell marker CD31. Quantitative RT-PCR analysis of laser-microdissected arterioles, glomeruli, and postcapillary venules demonstrated that targeted delivery of dexamethasone by AbEsel liposomes reduced glomerular endothelial expression of P-selectin, E-selectin, and vascular cell adhesion molecule-1 by 60–70%. The expression of these genes was not modulated in endothelial cells in nontargeted renal microvasculatures. Decrease of glomerular endothelial activation at disease onset was followed by reduced albuminuria at day 7. This study demonstrates the potential of vascular bed-specific drug delivery aimed at disease-induced epitopes on the microvascular endothelial cells as a therapeutic strategy for glomerulonephritis.
Albumin-stimulated DNA synthesis is mediated by Ca2+/PKC as well as EGF receptor-dependent p44/42 MAPK and NF-κB signal pathways in renal proximal tubule cells
Authors: Lee Yu Jin , Han Ho Jae
Published: December 2007

It is now recognized that significant tubular reabsorption of albumin occurs under physiological conditions that may play an important role in maintaining proximal tubular integrity and function. Therefore, this study examined the effect of bovine serum albumin (BSA) on DNA synthesis and its related signal molecules in primary cultured rabbit renal proximal tubule cells (PTCs). BSA increased the level of [3H]thymidine incorporation in a dose (≥3 mg/ml)- and time (≥3 h)-dependent manner, intracellular Ca2+concentration, and the level of protein kinase C (PKC) phosphorylation and stimulated the phosphorylation of the epidermal growth factor receptor (EGFR), which was inhibited by EGTA (extracellular Ca2+chelator), 1,2-bis(2-aminophenoxy)ethane- N,N,N′,N′-tetraacetic acid acetoxymethyl ester (BAPTA-AM, intracellular Ca2+chelator), or PKC inhibitors (staurosporine or bisindolylmaleimide I). In addition, the PKC inhibitors or an EGFR inhibitor (AG-1478) blocked the BSA-induced phosphorylation of p44/42 mitogen-activated protein kinases (MAPKs). BSA also increased the level of nuclear factor-κB (NF-κB) and inhibitor of NF-κB (IκB) phosphorylation, which was blocked by staurosporine, AG-1478, or PD-98059 (p44/42 MAPK inhibitor). Inhibition of Ca2+, PKC, EGFR, p44/42 MAPK, or NF-κB signal pathways blocked the BSA-induced incorporation of [3H]thymidine. Consequently, the inhibition of Ca2+, PKC, EGFR, p44/42 MAPKs, or NF-κB blocked the BSA-induced increases in cyclin D1, cyclin-dependent kinase (CDK)4, cyclin E, or CDK2 and restored the BSA-induced inhibition of p21WAF/Cip1and p27Kip1expression. In conclusion, BSA stimulates DNA synthesis that is mediated by Ca2+/PKC as well as the EGFR-dependent p44/42 MAPK and NF-κB signal pathways in PTCs.
Sex-specific influence of aging on exercising leg blood flow
Authors: Parker Beth A. , Smithmyer Sandra L. , Pelberg Justin A. , Mishkin Aaron D. , Proctor David N.
Published: December 2007

Our previous work suggests that healthy human aging is associated with sex-specific differences in leg vascular responses during large muscle mass exercise (2-legged cycling) (Proctor DN, Parker BA. Microcirculation 13: 315–327, 2006). The present study determined whether age × sex interactions in exercising leg hemodynamics persist during small muscle mass exercise that is not limited by cardiac output. Thirty-one young (20–30 yr; 15 men/16 women) and 31 older (60–79 yr; 13 men/18 women) healthy, normally active adults performed graded single-leg knee extensions to maximal exertion. Femoral artery blood velocity and diameter (Doppler ultrasound), heart rate (ECG), and beat-to-beat arterial blood pressure (mean arterial pressure, radial artery tonometry) were measured during each 3-min work rate (4.8 and 8 W/stage for women and men, respectively). The results (means ± SE) were as follows. Despite reduced resting leg blood flow and vascular conductance, older men exhibited relatively preserved exercising leg hemodynamic responses. Older women, by contrast, exhibited attenuated hyperemic (young: 52 ± 3 ml·min−1·W−1; vs. older: 40 ± 4 ml·min−1·W−1; P = 0.02) and vasodilatory responses (young: 0.56 ± 0.06 ml·min−1·mmHg−1·W−1 vs. older: 0.37 ± 0.04 ml·min−1·mmHg−1 W−1; P < 0.01) to exercise compared with young women. Relative (percentage of maximal) work rate comparisons of all groups combined also revealed attenuated vasodilator responses in older women ( P < 0.01 for age × sex × work rate interaction). These sex-specific age differences were not abolished by consideration of hemoglobin, quadriceps muscle, muscle recruitment, and mechanical influences on muscle perfusion. Collectively, these findings suggest that local factors contribute to the sex-specific effects of aging on exercising leg hemodynamics in healthy adults.
Bcl-xL Inhibitor ABT-737 Reveals a Dual Role for Bcl-xL in Synaptic Transmission
Authors: Hickman John A. , Hardwick J. Marie , Kaczmarek Leonard K. , Jonas Elizabeth A.
Published: December 2007

A role for BCL-xL in regulating neuronal activity is suggested by its dramatic effects on synaptic function and mitochondrial channel activity. When recombinant BCL-xL is injected into the giant presynaptic terminal of squid stellate ganglion or applied directly to mitochondrial outer membranes within the living terminal, it potentiates synaptic transmission acutely, and it produces mitochondrial channel activity. The squid, however, is a genetically intractable model, making it difficult to apply genetic tools in squid to explore the role of endogenous BCL-xL in synaptic function. Therefore the small molecule inhibitor ABT-737, a mimetic of the BH3-only protein BAD, binding to the BH3-binding domain pocket, was tested in squid, revealing a dual role for BCL-xL. ABT-737 slowed recovery of synaptic responses after repetitive synaptic activity, indicating that endogenous BCL-xL is necessary for timely recovery of rapidly firing synapses. Unexpectedly, however, ABT-737 also protected neurons from hypoxia-induced synaptic rundown and from increased permeability of the mitochondrial outer membrane during hypoxia. This implies that endogenous BCL-xL or a modified form of BCL-xL, such as the N-truncated, proteolytic, pro-apoptotic cleavage product, ΔN BCL-xL, contributes to injurious responses of the hypoxic synapse. To determine if ABT-737 is also an inhibitor of ΔN BCL-xL, recombinant ΔN BCL-xL protein was injected into the synapse. ABT-737 potently inhibited synaptic rundown induced by recombinant ΔN BCL-xL. These observations support the possibility that endogenous proteolysis or a functionally equivalent modification of BCL-xL is responsible for the deleterious effects of hypoxia on synaptic activity.
Inferring Spike Trains From Local Field Potentials
Authors: Rasch Malte J. , Gretton Arthur , Murayama Yusuke , Maass Wolfgang , Logothetis Nikos K.
Published: December 2007

We investigated whether it is possible to infer spike trains solely on the basis of the underlying local field potentials (LFPs). Using support vector machines and linear regression models, we found that in the primary visual cortex (V1) of monkeys, spikes can indeed be inferred from LFPs, at least with moderate success. Although there is a considerable degree of variation across electrodes, the low-frequency structure in spike trains (in the 100-ms range) can be inferred with reasonable accuracy, whereas exact spike positions are not reliably predicted. Two kinds of features of the LFP are exploited for prediction: the frequency power of bands in the high γ-range (40–90 Hz) and information contained in low-frequency oscillations (<10 Hz), where both phase and power modulations are informative. Information analysis revealed that both features code (mainly) independent aspects of the spike-to-LFP relationship, with the low-frequency LFP phase coding for temporally clustered spiking activity. Although both features and prediction quality are similar during seminatural movie stimuli and spontaneous activity, prediction performance during spontaneous activity degrades much more slowly with increasing electrode distance. The general trend of data obtained with anesthetized animals is qualitatively mirrored in that of a more limited data set recorded in V1 of non-anesthetized monkeys. In contrast to the cortical field potentials, thalamic LFPs (e.g., LFPs derived from recordings in the dorsal lateral geniculate nucleus) hold no useful information for predicting spiking activity.