bluuebunny/crossref_metadata_2025 huggingface dataset:
Cytokine signalling in rat pulp interstitial fluid and transcapillary fluid exchange during lipopolysaccharide‐induced acute inflammation
Authors: Bletsa Athanasia , Berggreen Ellen , Fristad Inge , Tenstad Olav , Wiig Helge
Published: March 2006

The dental pulp consists of loose connective tissue encased in rigid dentinal walls. Because of its topography the tissue has low interstitial compliance and limited capacity to expand during fluid volume changes. Due to limitations regarding access to interstitial fluid, basic knowledge on transcapillary fluid transport parameters is lacking for this organ. The scope of this project was dual: first we aimed at establishing a method for isolation of pulp interstitial fluid (IF), and second we applied the method in rats subjected to lipopolysaccharide (LPS)‐induced endotoxaemia. The aim was to measure colloid osmotic pressure (COP) and pro‐inflammatory cytokines in the pulp IF during acute inflammation. Fluid volumes and pulpal blood flow (PBF) were measured to obtain more information about microcirculatory changes that take place in this pulpitis model. By centrifugation of incisor pulp at 239 g we were able to extract fluid representative for IF. Pulp IF had a relative high control COP (∼83% of plasma COP) and was similar to plasma COP 3 h after LPS challenge. The pulp exhibited a high content of IF (0.60 ± 0.03 ml (g wet weight)−1) and a vascular volume of 0.03 ± 0.01 ml (g w.w.)−1 No differences were observed in the distribution of fluid volumes after 1.5 and 3 h LPS exposure. PBF and systemic blood pressure dropped significantly after LPS administration. PBF remained low whereas systemic blood pressure was re‐established during the 3‐h period, implying organ dysfunction. There was a differential pattern of cytokine expression in pulp IF and serum with cytokines such as IL‐1α, IL‐1β and TNF‐α locally produced, whereas others such as IFN‐γ and IL‐6 were produced systemically and probably spilled over to the pulp IF after LPS exposure. Our findings show that pulp IF can be isolated by centrifugation and that this method is useful when studying fluid balance and extracellular signalling mechanisms in the dental pulp in normal and pathological conditions.
Dynamic behaviour of half‐sarcomeres during and after stretch in activated rabbit psoas myofibrils: sarcomere asymmetry but no ‘sarcomere popping’
Authors: Telley I. A. , Stehle R. , Ranatunga K. W. , Pfitzer G. , Stüssi E. , Denoth J.
Published: March 2006

We examined length changes of individual half‐sarcomeres during and after stretch in actively contracting, single rabbit psoas myofibrils containing 10–30 sarcomeres. The myofibrils were fluorescently immunostained so that both Z‐lines and M‐bands of sarcomeres could be monitored by video microscopy simultaneously with the force measurement. Half‐sarcomere lengths were determined by processing of video images and tracking the fluorescent Z‐line and M‐band signals. Upon Ca2+ activation, during the rise in force, active half‐sarcomeres predominantly shorten but to different extents so that an active myofibril consists of half‐sarcomeres of different lengths and thus asymmetric sarcomeres, i.e. shifted A‐bands, indicating different amounts of filament overlap in the two halves. When force reached a plateau, the myofibril was stretched by 15–20% resting length (L0) at a velocity of ∼0.2 L0 s−1. The myofibril force response to a ramp stretch is similar to that reported from muscle fibres. Despite the ∼2.5‐fold increase in force due to the stretch, the variability in half‐sarcomere length remained almost constant during the stretch and A‐band shifts did not progress further, independent of whether half‐sarcomeres shortened or lengthened during the initial Ca2+ activation. Moreover, albeit half‐sarcomeres lengthened to different extents during a stretch, rapid elongation of individual sarcomeres beyond filament overlap (‘popping’) was not observed. Thus, in contrast to predictions of the ‘popping sarcomere’ hypothesis, a stretch rather stabilizes the uniformity of half‐sarcomere lengths and sarcomere symmetry. In general, the half‐sarcomere length changes (dynamics) before and after stretch were slow and the dynamics after stretch were not readily predictable on the basis of the steady‐state force–sarcomere length relation.
Kinetic and functional analysis of transient, persistent and resurgent sodium currents in rat cerebellar granule cells in situ: an electrophysiological and modelling study
Authors: Magistretti Jacopo , Castelli Loretta , Forti Lia , D'Angelo Egidio
Published: March 2006

Cerebellar neurones show complex and differentiated mechanisms of action potential generation that have been proposed to depend on peculiar properties of their voltage‐dependent Na+ currents. In this study we analysed voltage‐dependent Na+ currents of rat cerebellar granule cells (GCs) by performing whole‐cell, patch‐clamp experiments in acute rat cerebellar slices. A transient Na+ current (INaT) was always present and had the properties of a typical fast‐activating/inactivating Na+ current. In addition to INaT, robust persistent (INaP) and resurgent (INaR) Na+ currents were observed. INaP peaked at ∼−40 mV, showed half‐maximal activation at ∼−55 mV, and its maximal amplitude was about 1.5% of that of INaT. INaR was elicited by repolarizing pulses applied following step depolarizations able to activate/inactivate INaT, and showed voltage‐ and time‐dependent activation and voltage‐dependent decay kinetics. The conductance underlying INaR showed a bell‐shaped voltage dependence, with peak at −35 mV. A significant correlation was found between GC INaR and INaT peak amplitudes; however, GCs expressing INaT of similar size showed marked variability in terms of INaR amplitude, and in a fraction of cells INaR was undetectable. INaT, INaP and INaR could be accounted for by a 13‐state kinetic scheme comprising closed, open, inactivated and blocked states. Current‐clamp experiments carried out to identify possible functional correlates of INaP and/or INaR revealed that in GCs single action potentials were followed by depolarizing afterpotentials (DAPs). In a majority of cells, DAPs showed properties consistent with INaR playing a role in their generation. Computer modelling showed that INaR promotes DAP generation and enhances high‐frequency firing, whereas INaP boosts near‐threshold firing activity. Our findings suggest that special properties of voltage‐dependent Na+ currents provides GCs with mechanisms suitable for shaping activity patterns, with potentially important consequences for cerebellar information transfer and computation.
Translocation of an endoproteolytically cleaved maxi‐K channel isoform: mechanisms to induce human myometrial cell repolarization
Authors: Korovkina Victoria P. , Brainard Adam M. , England Sarah K.
Published: March 2006

Large conductance Ca2+‐ and voltage‐activated K+ (maxi‐K) channels modulate human myometrial smooth muscle cell (hMSMC) excitability; however, the role of individual alternatively spliced isoforms remains unclear. We have previously shown that the transcript of a human maxi‐K channel isoform (mK44) is expressed predominantly in myometrial and aortic smooth muscle and forms a functional channel in heterologous expression systems. The mK44 isoform contains unique consensus motifs for both endoproteolytic cleavage and N‐myristoylation, although the function of these post‐translational modifications is unknown. The goal of these studies was to determine the role of post‐translational modifications in regulating mK44 channel function in hMSMCs. An mK44‐specific antibody indicated that this channel is localized intracellularly in hMSMCs and translocates to the cell membrane in response to increases in intracellular Ca2+. Immunological analyses using an N‐terminally myc‐tagged mK44 construct demonstrated endoproteolytical cleavage of mK44 in hMSMCs resulting in membrane localization of the mK44 N‐termini and intracellular retention of the pore‐forming C‐termini. Caffeine‐induced Ca2+ release from intracellular stores resulted in translocation of the C‐termini of mK44 to the cell membrane and co‐localization with its N‐termini. Translocation of mK44 channels to the cell membrane was concomitant with repolarization of the hMSMCs. Endoproteolytic digest of mK44 did not occur in HEK293 cells or mouse fibroblasts. MK44 truncated at a putative N‐myristoylation site did not produce current when expressed alone, but formed a functional channel when co‐expressed with the N‐terminus. These findings provide novel insight into cell‐specific regulation of maxi‐K channel function.
The human adult subtype ACh receptor channel has high Ca2+ permeability and predisposes to endplate Ca2+ overloading
Authors: Fucile Sergio , Sucapane Antonietta , Grassi Francesca , Eusebi Fabrizio , Engel Andrew G.
Published: March 2006

Slow‐channel congenital myasthenic syndrome, caused by mutations in subunits of the endplate ACh receptor (AChR), results in prolonged synaptic currents and excitotoxic injury of the postsynaptic region by Ca2+ overloading. The Ca2+ overloading could be due entirely to the prolonged openings of the AChR channel or could be abetted by enhanced Ca2+ permeability of the mutant channels. We therefore measured the fractional Ca2+ current, defined as the percentage of the total ACh‐evoked current carried by Ca2+ ions (Pf), for AChRs harbouring the αG153S or the αV249F slow‐channel mutation, and for wild‐type human AChRs in which Pf has not yet been determined. Experiments were performed in transiently transfected GH4C1 cells and human myotubes with simultaneous recording of ACh‐evoked whole‐cell currents and fura‐2 fluorescence signals. We found that the Pf of the wild‐type human endplate AChR was unexpectedly high (Pf∼7%), but neither the αV249F nor the αG153S mutation altered Pf. Fetal human AChRs containing either the wild‐type or the mutated α subunit had a much lower Pf (2–3%). We conclude that the Ca2+ permeability of human endplate AChRs is higher than that reported for any other human nicotinic AChR, with the exception of α7‐containing AChRs (Pf > 10%); and that neither the αG153S nor the αV249F mutations affect the Pf of fetal or adult endplate AChRs. However, the intrinsically high Ca2+ permeability of human AChRs probably predisposes to development of the endplate myopathy when opening events of the AChR channel are prolonged by altered AChR‐channel kinetics.
Genome-Wide Detection of Polymorphisms at Nucleotide Resolution with a Single DNA Microarray
Authors: Gresham David , Ruderfer Douglas M. , Pratt Stephen C. , Schacherer Joseph , Dunham Maitreya J. , Botstein David , Kruglyak Leonid
Published: March 2006

A central challenge of genomics is to detect, simply and inexpensively, all differences in sequence among the genomes of individual members of a species. We devised a system to detect all single-nucleotide differences between genomes with the use of data from a single hybridization to a whole-genome DNA microarray. This allowed us to detect a variety of spontaneous single–base pair substitutions, insertions, and deletions, and most (>90%) of the ∼30,000 known single-nucleotide polymorphisms between two
Saccharomyces cerevisiae
strains. We applied this approach to elucidate the genetic basis of phenotypic variants and to identify the small number of single–base pair changes accumulated during experimental evolution of yeast.
Acetylated Tat Regulates Human Immunodeficiency Virus Type 1 Splicing through Its Interaction with the Splicing Regulator p32
Authors: Berro Reem , Kehn Kylene , de la Fuente Cynthia , Pumfery Anne , Adair Richard , Wade John , Colberg-Poley Anamaris M. , Hiscott John , Kashanchi Fatah
Published: March 2006

The human immunodeficiency virus type 1 (HIV-1) potent transactivator Tat protein mediates pleiotropic effects on various cell functions. Posttranslational modification of Tat affects its activity during viral transcription. Tat binds to TAR and subsequently becomes acetylated on lysine residues by histone acetyltransferases. Novel protein-protein interaction domains on acetylated Tat are then established, which are necessary for both sustained transcriptional activation of the HIV-1 promoter and viral transcription elongation. In this study, we investigated the identity of proteins that preferentially bound acetylated Tat. Using a proteomic approach, we identified a number of proteins that preferentially bound AcTat, among which p32, a cofactor of splicing factor ASF/SF-2, was identified. We found that p32 was recruited to the HIV-1 genome, suggesting a mechanism by which acetylation of Tat may inhibit HIV-1 splicing needed for the production of full-length transcripts. Using Tat from different clades, harboring a different number of acetylation sites, as well as Tat mutated at lysine residues, we demonstrated that Tat acetylation affected splicing in vivo. Finally, using confocal microscopy, we found that p32 and Tat colocalize in vivo in HIV-1-infected cells.
Functional Characterization of Heptad Repeat 1 and 2 Mutants of the Spike Protein of Severe Acute Respiratory Syndrome Coronavirus
Authors: Chan Woan-Eng , Chuang Chin-Kai , Yeh Shiou-Hwei , Chang Mau-Sun , Chen Steve S.-L.
Published: March 2006

To understand the roles of heptad repeat 1(HR1) and HR2 of the spike (S) protein of the severe acute respiratory syndrome coronavirus (SARS-CoV) in virus-cell interactions, the conserved Leu or Ile residues located at positions 913, 927, 941, and 955 in HR1 and 1151, 1165, and 1179 in HR2 were individually replaced with an α-helix-breaker Pro residue. The 913P mutant was expressed mainly as a faster-migrating, lower-molecular-weight S
L
form, while the wild type and all other mutants produced similar levels of both the S
L
form and the slower-migrating, higher-molecular-weight S
H
form. The wild-type S
L
form was processed to the S
H
form, whereas the S
L
form of the 913P mutant was inefficiently converted to the S
H
form after biosynthesis. None of these mutations affected cell surface expression or binding to its cognate ACE2 receptor. In a human immunodeficiency virus type 1/SARS S coexpression study, all mutants except the 913P mutant incorporated the S
H
form into the virions as effectively as did the wild-type S
H
form. The mutation at Ile-1151 did not affect membrane fusion or viral entry. The impaired viral entry of the 927P, 941P, 955P, and 1165P mutants was due to their inability to mediate membrane fusion, whereas the defect in viral entry of the 1179P mutant occurred not at the stage of membrane fusion but rather at a postfusion stage. Our study demonstrates the functional importance of HR1 and HR2 of the SARS-CoV spike protein in membrane fusion and viral entry.
Characterization of Two Novel Polyomaviruses of Birds by Using Multiply Primed Rolling-Circle Amplification of Their Genomes
Authors: Johne Reimar , Wittig Walter , Fernández-de-Luco Daniel , Höfle Ursula , Müller Hermann
Published: March 2006

Polyomaviruses are small nonenveloped particles with a circular double-stranded genome, approximately 5 kbp in size. The mammalian polyomaviruses mainly cause persistent subclinical infections in their natural nonimmunocompromised hosts. In contrast, the polyomaviruses of birds—avian polyomavirus (APV) and goose hemorrhagic polyomavirus (GHPV)—are the primary agents of acute and chronic disease with high mortality rates in young birds. Screening of field samples of diseased birds by consensus PCR revealed the presence of two novel polyomaviruses in the liver of an Eurasian bullfinch (
Pyrrhula pyrrhula griseiventris
) and in the spleen of a Eurasian jackdaw (
Corvus monedula
), tentatively designated as finch polyomavirus (FPyV) and crow polyomavirus (CPyV), respectively. The genomes of the viruses were amplified by using multiply primed rolling-circle amplification and cloned. Analysis of the FPyV and CPyV genome sequences revealed a close relationship to APV and GHPV, indicating the existence of a distinct avian group among the polyomaviruses. The main characteristics of this group are (i) involvement in fatal disease, (ii) the existence of an additional open reading frame in the 5′ region of the late mRNAs, and (iii) a different manner of DNA binding of the large tumor antigen compared to that of the mammalian polyomaviruses.
Enhanced Local Symmetry Interactions Globally Stabilize a Mutant Virus Capsid That Maintains Infectivity and Capsid Dynamics
Authors: Speir Jeffrey A. , Bothner Brian , Qu Chunxu , Willits Deborah A. , Young Mark J. , Johnson John E.
Published: March 2006

Structural transitions in viral capsids play a critical role in the virus life cycle, including assembly, disassembly, and release of the packaged nucleic acid.
Cowpea chlorotic mottle virus
(CCMV) undergoes a well-studied reversible structural expansion in vitro in which the capsid expands by 10%. The swollen form of the particle can be completely disassembled by increasing the salt concentration to 1 M. Remarkably, a single-residue mutant of the CCMV N-terminal arm, K42R, is not susceptible to dissociation in high salt (salt-stable CCMV [SS-CCMV]) and retains 70% of wild-type infectivity. We present the combined structural and biophysical basis for the chemical stability and viability of the SS-CCMV particles. A 2.7-Å resolution crystal structure of the SS-CCMV capsid shows an addition of 660 new intersubunit interactions per particle at the center of the 20 hexameric capsomeres, which are a direct result of the K42R mutation. Protease-based mapping experiments of intact particles demonstrate that both the swollen and closed forms of the wild-type and SS-CCMV particles have highly dynamic N-terminal regions, yet the SS-CCMV particles are more resistant to degradation. Thus, the increase in SS-CCMV particle stability is a result of concentrated tethering of subunits at a local symmetry interface (i.e., quasi-sixfold axes) that does not interfere with the function of other key symmetry interfaces (i.e., fivefold, twofold, quasi-threefold axes). The result is a particle that is still dynamic but insensitive to high salt due to a new series of bonds that are resistant to high ionic strength and preserve the overall particle structure.
Genetic and Functional Analysis of R5X4 Human Immunodeficiency Virus Type 1 Envelope Glycoproteins Derived from Two Individuals Homozygous for the CCR5Δ32 Allele
Authors: Gray Lachlan , Churchill Melissa J. , Keane Niamh , Sterjovski Jasminka , Ellett Anne M. , Purcell Damian F. J. , Poumbourios Pantelis , Kol Chenda , Wang Bin , Saksena Nitin K. , Wesselingh Steven L. , Price Patricia , French Martyn , Gabuzda Dana , Gorry Paul R.
Published: March 2006

We characterized human immunodeficiency virus type 1 (HIV-1) envelope glycoproteins (Env) isolated from two HIV-1-infected CCR5Δ32 homozygotes. Envs from both subjects used CCR5 and CXCR4 for entry into transfected cells. Most R5X4 Envs were lymphocyte-tropic and used CXCR4 exclusively for entry into peripheral blood mononuclear cells (PBMC), but a subset was dually lymphocyte- and macrophage-tropic and used either CCR5 or CXCR4 for entry into PBMC and monocyte-derived macrophages. The persistence of CCR5-using HIV-1 in two CCR5Δ32 homozygotes suggests the conserved CCR5 binding domain of Env is highly stable and provides new mechanistic insights important for HIV-1 transmission and persistence.
A rapid, microwave-assisted, one-pot cyclopropanation reaction to synthesize (4-ferrocenyl)benzoylcyclopropane in the solid state
Authors: Yin Juxing , Li Xiaolong , Wu Xiaoli , Chen Baohua , Liu Zenglu , Song Qingbao
Published: March 2006

Some new (4-ferrocenyl)benzoylcyclopropanes were prepared rapidly via a microwave-assisted, one-pot cyclopropanation reaction in the solid state, directly using 4-ferrocenylacetophenone, aldehyde, and sulfur ylide as the starting materials. The main advantages of this procedure are convenient operation, shorter reaction time, lower environmental pollution, and no need to isolate the intermediate product (chalcone) in a separate step.Key words: 4-ferrocenylacetophenone, sulfur ylide, microwave-assisted, one-pot, solid state.
EFFECTIVE FIELD THEORY APPROACH TO THE NUCLEON–NUCLEON INTERACTION REVISITED
Authors: GEGELIA JAMBUL , SCHERER STEFAN
Published: March 2006

It is argued that Weinberg's approach to the nucleon–nucleon (NN) interaction problem within effective field theory provides a consistent power counting for renormalized diagrams. Within this scheme the NN potential is organized as an expansion in terms of small quantities like small external momenta and the pion mass (divided by the characteristic large scale of the effective theory). Physical observables to any given order in these small quantities are calculated from the solutions of the Lippmann–Schwinger (or Schrödinger) equation.
Pharmacological chaperone corrects lysosomal storage in Fabry disease caused by trafficking-incompetent variants
Authors: Yam Gary Hin-Fai , Bosshard Nils , Zuber Christian , Steinmann Beat , Roth Jürgen
Published: March 2006

Fabry disease is a lysosomal storage disorder caused by deficiency of α-galactosidase A (α-Gal A) resulting in lysosomal accumulation of glycosphingolipid globotriosylceramide Gb3. Misfolded α-Gal A variants can have residual enzyme activity but are unstable. Their lysosomal trafficking is impaired because they are retained in the endoplasmic reticulum (ER) by quality control. Subinhibitory doses of the competitive inhibitor of α-Gal A, 1-deoxygalactonojirimycin (DGJ), stabilize mutant α-Gal A in vitro and correct the trafficking defect. We showed by immunolabeling that the chaperone-like action of DGJ significantly reduces the lysosomal Gb3 storage in human Fabry fibroblasts harboring the novel mutations T194I and V390fsX8. The specificity of the DGJ effect was proven by RNA interference. Electron microscopic morphometry demonstrated a reduction of large-size, disease-associated lysosomes and loss of characteristic multilamellar lysosomal inclusions on DGJ treatment. In addition, the pre-Golgi intermediates were decreased. However, the rough ER was not different between DGJ-treated and untreated cells. Pulse-chase experiments revealed that DGJ treatment resulted in maturation and stabilization of mutant α-Gal A. Genes involved in cell stress signaling, heat shock response, unfolded protein response, and ER-associated degradation show no apparent difference in expression between untreated and DGJ-treated fibroblasts. The DGJ treatment has no apparent cytotoxic effects. Thus our data show the usefulness of a pharmacological chaperone for correction of the lysosomal storage in Fabry fibroblasts harboring different mutations with residual enzyme activity. Pharmacological chaperones acting on misfolded, unstable mutant proteins that exhibit residual biological activity offer a convenient and cost-efficient therapeutic strategy.
Myocardial β1-adrenergic receptor polymorphisms affect functional recovery after ischemic injury
Authors: Akhter Shahab A. , D'Souza Karen M. , Petrashevskaya Natalia N. , Mialet-Perez Jeanne , Liggett Stephen B.
Published: March 2006

Association studies suggest β1-adrenergic receptor (β1-AR) polymorphisms are disease modifiers in heart failure. The Arg389 variant has increased coupling to Gsin transfected cells and evokes enhanced ventricular function in transgenic mice. Here, we assessed the differential effects of the human Gly389 and Arg389 β1-AR polymorphisms on myocardial recovery after ischemic injury. Function was studied in transgenic mice with cardiac-specific expression of either human Gly389 or Arg389 β1-AR at baseline and after 20 min of ex vivo ischemia and reperfusion (I/R). In 3-mo-old mice of either genotype, there was poor recovery after I/R (∼38% vs. ∼68% for nontransgenic). Paradoxically, at 6 mo of age, functional recovery remained severely depressed in Gly389 hearts (∼32%) but was similar to nontransgenic for Arg389 hearts (∼60%). In Arg389 hearts, agonist-promoted adenylyl cyclase activities were depressed by ∼35% at 6 mo of age, and G protein-coupled receptor kinase (GRK) activity was increased by approximately twofold compared with Gly389. Furthermore, I/R evoked an approximately threefold increase in ERK2 phosphorylation in Arg389 but an approximately twofold decrease in Gly389 hearts. Individually, these changes have been shown to mitigate I/R injury; thus the Arg389-β1-AR uniquely evokes specialized pathways that act to protect against I/R injury. The improved recovery of function after I/R in Arg389 hearts relative to Gly389 appears to be due to an adaptive multimechanism program with allele-specific alterations in receptor signaling, GRK activity, and ERK2. Thus genetic variation of the human β1-AR may play a role in cardiac functional recovery after ischemic injury.
Review – 3D Micro CT Imaging of Renal Micro-Structural Changes
Authors: Gössl Mario , Bentley Michael D. , Lerman Lilach O.
Published: March 2006

Appropriate nephron function is dependent on the detailed spatial interrelationship of blood vesselswith the tubular components. However, because of methodological limitations,the three-dimensional anatomical complexity of the renal vasculatureand its geometrical relationship with specific tubular segmentsalong the nephron has been difficult to study in a quantitativemanner. Three-dimensional microcomputed tomography (3D micro CT) offers the unique opportunity to image kidney sample volumes with a high spatial resolution (of up to 5 µm cubic voxel size) without physically sectioning them, thereby allowing accurate calculations of vessel tortuosity and density, as indices of neovascularization, as well as volume and distribution of various kidney structures. In conjunction with molecular biology techniques, valuable associations between renal microstructures and activation of local molecular pathways can be drawn to elucidate mechanisms of renal disease and design therapeutic approaches. For example, recent studies in animal models of renal hypertension, hypercholesterolemia, and diabetes mellitus have shown that medical intervention to decrease oxidative stress and micro-inflammation may preserve renal microstructures as well as renal function in these chronic renal diseases. Future developments will be needed to establish the place of 3D micro CT in developing, directing, and monitoring the treatment of chronic kidney diseases.
Assessment of Morphological and Immunohistological Alterations in Long-Term Keloid Skin Explants
Authors: Duong Hai S. , Zhang Qunzhou , Kobi Abraham , Le Anh , Messadi Diana V.
Published: March 2006

One of the major impediments in keloid research is the lack of a keloid animal model that can mimic human keloid. This imposes investigative constraints on studying cellular interactions and biochemical processes that normally occur in vivo.Our main objective is to establish an in vitro model for maintaining long-term viable keloid dermal explants as a tool for investigating the pathogenesis of keloid scar formation. Explants of adult keloid scars were cultured in vitro by embedding them in enriched collagen gel matrix and maintaining them for up to 6 weeks, whereupon changes in tissue morphology and cellular differentiation were examined. The effects of medium enrichment, air versus liquid submersion, and different substrates on the explants were examined. Our results indicated that keloid explants embedded in a collagen gel matrix were morphologically better preserved than explants placed on a plastic substrate. Explants with epidermis at the air-liquid interface had better morphology than collagen-submerged explants, and there were no differences between serum-free and serum-supplemented explant cultures. Immunohistochemical and apoptotic analyses were performed to assess cellular viability and differentiation. In situ hybridization confirmed that keloid fibroblasts had sustained collagen type I gene expression throughout the 6 weeks in culture, thus validating the integrity of a long-term keloid culture system. In conclusion, the collagen-embedded skin explant system demonstrates that keloid tissues could be maintained for up to 6 weeks for long-term in vitro studies.
Nuclear Medicine Procedures for the Diagnosis of Acute and Chronic Renal Failure
Authors: Haufe Sabine E. , Riedmüller K. , Haberkorn U.
Published: March 2006

The focus of this review is on the current role of nuclear imaging studies in the clinical evaluation of patients with acute and chronic renal failure. In this setting nuclear imaging has two roles: diagnostic and prognostic, indicating that these methods are an essential component in the evaluation of renal diseases. The functional assessment of the kidney by nuclear medicine procedures is based on the use of radioisotopes bound to non-metabolized molecules with known pharmacokinetics. Renal scintigraphy is usually applied for the assessment of renal function expressed as glomerular filtration rate, effective renal plasma flow or more generally kidney perfusion. Newer methods rely on positron emission tomography, which allows the generation of images with higher resolution and absolute quantitation of biological processes such as transport activities, enzyme activities or angiotensin receptors.
Damage to Human Hair Caused by Repeated Bleaching Combined with Daily Weathering during Daily Life Activities
Authors: Masukawa Y. , Tsujimura Hisashi , Tanamachi H. , Narita H. , Imokawa Genji
Published: March 2006

Background: Although it is well known that chemical oxidation with alkaline hydrogen peroxide damages human hair, little is known about the effects of bleaching during daily life activities which may alter hair components. Objective: To determine the effects of bleaching combined with daily weathering on lipids and amino acids in hair. Methods: Levels of lipids and amino acids were analyzed by chromatography in scalp hair fibers collected from three different females and in experimentally treated hair. Results: Bleaching combined with daily weathering induced the loss of 18-methyleicosanoic acid (18-MEA) and the conversion of half-cystine (H-CYS) to cysteic acid (CYS-A), which were the most remarkable changes among all lipids and amino acids tested. Although a single bleaching had no effect, repeated bleaching combined with daily weathering elicited significant decreases in other intrinsic endogenous lipids and alterations in amino acids that are characteristic for the cuticle. Conclusion: Hair damage, eventually leading to the worst damage such as splitting, is caused by bleaching during daily life activities and is mainly attributable to the loss of 18-MEA as well as the conversion of H-CYS to CYS-A.
Recharacterization of the Nonlesional Dry Skin in Atopic Dermatitis through Disrupted Barrier Function
Authors: Matsuki Hayato , Kiyokane Kimihiro , Matsuki Takahiro , Sato Sayuri , Imokawa Genji
Published: March 2006

Background: The etiology of the nonlesional dry and barrier-disrupted skin of patients with atopic dermatitis (AD) is still unclear. Objective: To determine whether disrupted barrier function in the nonlesional skin is associated with inflammatory or postinflammatory events, which are relevant to the severity of AD or local dry skin properties, respectively. Methods: We evaluated the barrier function and the water content of nonlesional forearm skin and compared these with the severity of AD and the intensity of dryness/scaling/itchiness at the same skin sites. Results: The transepidermal water loss (TEWL) significantly increased in proportion to the severity of AD with a markedly high correlation coefficient (r = 0.834, p < 0.0001, n = 106), while the capacitance decreased in proportion to the severity of AD with a relatively lower correlation coefficient (r = –0.720, p < 0.0001, n = 106) compared with TEWL. Relationship between TEWL and capacitance values in association with the AD severity revealed that the two parameters are well distributed, corresponding to the severity of AD, and that the elevated TEWL more adequately reflects the difference between healthy control and the mild group of AD compared with the reduced capacitance. Comparison with dry skin properties revealed that while the capacitance values were highly correlated with dryness (r = –0.752, p < 0.0001, n = 106) and with scaling (r = –0.697, p < 0.0001, n = 106), the TEWL was also related to dryness (r = 0.788, p < 0.0001, n = 106) with a higher correlation coefficient compared with capacitance and to scaling (r = 0.697, p < 0.0001, n = 106). Conclusion: Our results indicate that the barrier disruption in the nonlesional skin is well suited to reflect the severity of AD as well as the dry skin properties, providing a useful insight into understandings of diagnosis and clinical improvement during therapy.
Reevaluation of the Importance of Barrier Dysfunction in the Nonlesional Dry Skin of Atopic Dermatitis Patients through the Use of Two Barrier Creams
Authors: Matsuki Hayato , Kiyokane Kimihiro , Matsuki Takahiro , Sato Sayuri , Imokawa Genji
Published: March 2006

Background: Atopic dermatitis (AD) can be considered a barrier disease in which antigens and irritants that can easily penetrate clinically normal, nonlesional skin due to its defective barrier function trigger and worsen the dermatitis. Thus, replenishing the barrier function in clinically normal, nonlesional skin of patients with AD seems to be a key for preventing the refractory nature of the dermatitis. Objective: To determine whether the disrupted barrier function of AD nonlesional skin can be repaired by topical application of a synthetic ceramide known to induce barrier recovery and to subsequently evaluate the relationship between enhanced barrier function and improved dry skin conditions. Methods: We applied topically a synthetic ceramide (CER) or hirudoid (HIRU)-containing cream to the nonlesional skin of AD patients for 4 weeks and evaluated their efficacy by measuring transepidermal water loss (TEWL) and capacitance values as well as clinical scoring for scaling/dryness/itchiness. Results: Treatment for 4 weeks with the CER cream significantly reduced dryness/scaling/itchiness which was accompanied by significant decreases in TEWL and increases in capacitance values at 2 and 4 weeks. In contrast, treatment for 4 weeks with the HIRU cream elicited a similar but lesser reduction in dryness/scaling/itchiness which was accompanied by significant but lesser decreases and increases in TEWL and capacitance values, respectively, at 2 and 4 weeks. Comparison of TEWL and capacitance values during the 4 weeks of treatment with CER or HIRU creams revealed that while the two parameters of CER cream-treated skin were generally similar to healthy control skin, those of the HIRU cream-treated skin remained similar to mild or moderate AD skin. Conclusion: It is likely that the recovery of barrier function reflects the improvement in clinically evaluated dry skin conditions of the nonlesional skin to a greater extent than that in water deficiency, which suggests that the barrier-replenishing effect is a more important factor for treatment of AD nonlesional skin than is the improvement of water deficiency.
Hypoglycemia in Newborn Infants: Features Associated with Adverse Outcomes
Authors: Rozance Paul J. , Hay William W.
Published: March 2006

The purpose of this review article is to document from the literature values of blood/plasma glucose concentration and associated clinical signs and conditions in newborn infants (both term and preterm) that indicate a reasonable clinical probability that hypoglycemia is a proximate cause of acute and/or sustained neurological injury and to review the physiological and pathophysiological responses to hypoglycemia that may influence the ultimate outcome of newborns with low blood glucose. Our overall conclusion is that there is inadequate information in the literature to define any one value of glucose below which irreparable hypoglycemic injury to the central nervous system occurs, at any one time or for any defined period of time, in a population of infants or in any given infant. Clinical signs of prolonged and severe neurological disturbance (coma, seizures), extremely and persistently low plasma/blood glucose concentrations (0 to <1.0 mmol/l [0 to <18–20 mg/dl] for more than 1–2 h), and the absence of other obvious central nervous system (CNS) pathology (hypoxia-ischemia, intracranial hemorrhage, infection, etc.) are important for the diagnosis of injury due to glucose deficiency. Specific conditions, such as persistent hyperinsulinemia with severe hypoglycemic episodes that include seizures, also contribute to the diagnosis of hypoglycemic injury. Such lack of definitive measures of injury specific to glucose deficiency indicates that clinicians should be on the alert for infants at risk of hypoglycemia and for clinical signs and conditions that might herald severe hypoglycemia; they should have a low threshold for investigating and diagnosing ‘hypoglycemia’ with frequent measurements of plasma/blood glucose concentration; and they should treat low glucose concentrations promptly and maintain them in a safe range. Because there is no conclusive evidence or consensus in the literature that defines an absolute value or duration of ‘hypoglycemia’ that must occur, with our without related clinical complications, to produce neurological injury, clinicians should consider the information currently available, determine a ‘target’ plasma or blood glucose concentration that is acceptable, and treat infants with glucose concentrations below this value accordingly. Our intent in this review article is to highlight the studies relevant to this issue and help clinicians formulate a safe and, hopefully, effective strategy for the diagnosis and treatment of hypoglycemia.
Contrast-Enhanced 3-D MRA in Decision Making for Carotid Endarterectomy: A 6-Year Experience
Authors: Barth Alain , Arnold Marcel , Mattle Heinrich P. , Schroth Gerhard , Remonda Luca
Published: March 2006

Background: Systematic need for angiography in diagnosis of carotid artery stenosis and indication of surgical therapy is still debated. Noninvasive imaging techniques such as MR angiography (MRA) or CT angiography (CTA) offer an alternative to digital subtraction angiography (DSA) and are increasingly used in clinical practice. In this study, we present the radiological characteristics and clinical results of a series of patients operated on the basis of combined ultrasonography (US)/MRA. Methods: This observational study included all the patients consecutively operated for a carotid stenosis in our Department from October 1998 to December 2004. The applied MRA protocol had previously been established in a large correlation study with DSA. DSA was used only in case of discordance between US and MRA. The preoperative radiological information furnished by MRA was compared with intraoperative findings. The outcome of the operation was assessed according to ECST criteria. Results: Among 327 patients, preoperative MRA was performed in 278 (85%), DSA in 44 (13.5%) and CT angiography in 5 (1.5%). Most of DSA studies were performed as emergency for preparation of endovascular therapy or for reasons other than carotid stenosis. Eleven additional DSA (3.3%) complemented US/MRA, mostly because diverging diagnosis of subocclusion of ICA. No direct morbidity or intraoperative difficulty was related to preoperative MRA. Combined mortality/major morbidity rate was 0.9% (3 patients) and minor morbidity rate 5.5% (18 patients). Conclusions: This observational study describes a well-established practice of carotid surgery and supports the exclusive use of non invasive diagnostic imaging for indicating and deciding the operation.
Does Thrombolysis Benefit Patients with Lacunar Syndrome?
Authors: Cocho Dolores , Belvís Roberto , Martí-Fàbregas Joan , Bravo Yolanda , Aleu Aitziber , Pagonabarraga Javier , Molina-Porcel Laura , Díaz-Manera Jorge , San Roman Luis , Martínez-Lage María , Martínez Alejandro , Moreno Mireia , Martí-Vilalta Josep-Lluis
Published: March 2006

The efficacy of thrombolysis in clinical stroke subtypes is unclear. We compared the benefit of intravenous rt-PA in 11 patients with lacunar syndrome with that in 33 patients with a non-lacunar syndrome. Patients were matched by NIHSS score and time to treatment. Although no statistically significant differences were detected in outcome, the benefit was greater in the non- lacunar syndrome group.
Progressive Overlap Syndrome due to Small Cell Lung Cancer as a Paraneoplastic Syndrome
Authors: Yano Shuichi , Kobayashi Kanako , Kato Kazuhiro
Published: March 2006

The association between rheumatic diseases and malignancy has been documented in many studies. Polymyositis and especially dermatomyositis are associated with a higher risk of malignant disease. Furthermore, lung cancer usually develops in patients with a prolonged history of systemic sclerosis accompanied by pulmonary fibrosis. Though overlap syndrome is rarely reported in the associated cancer, this is the first report describing progressive overlap syndrome in small cell lung cancer as a paraneoplastic syndrome.
Onset and Duration of Action of Formoterol and Tiotropium in Patients with Moderate to Severe COPD
Authors: Richter Kai , Stenglein Stephan , Mücke Marion , Sieder Christian , Schmidtmann Sören , Harnest Ulf , Weidinger Gottfried , Magnussen Helgo
Published: March 2006

Background: Chronic obstructive pulmonary disease (COPD) management guidelines recommend regular treatment with one or more long-acting bronchodilators for patients with moderate to severe COPD. Objective: To compare the onset and duration of action of formoterol and tiotropium in patients with COPD. Methods: This randomized, multicentre, open-label crossover study in 38 patients with COPD (mean age 64 years; mean FEV1 55% predicted) assessed the effect of 7 days of treatment with formoterol (12 µg b.i.d. via Foradil® Aerolizer®) vs. tiotropium (18 µg o.d. via Spiriva® HandiHaler®) on lung function measured over a period of 12 h after the first dose on day 1 and the last dose on day 8. Results: The primary efficacy variable, FEV1-AUC during the first 2 h post-dose (FEV1-AUC10–120 min), was significantly higher for formoterol compared with tiotropium, with between-treatment differences of 124 ml (p = 0.016) after the first dose and 80 ml (p = 0.036) after 7 days’ treatment in favour of formoterol. FEV1 measured 12 h after inhalation did not differ statistically significantly between treatments. Adverse events occurred in 2 (5%) patients after treatment with formoterol and in 5 (12%) patients after treatment with tiotropium. Conclusion: This study demonstrates faster onset of action and greater bronchodilation of formoterol vs. tiotropium for bronchodilation within the first 2 h of inhalation (FEV1-AUC10–120 min) and comparable bronchodilation 12 h post-inhalation in patients with moderate to severe COPD.
Decreased Plasma Fibronectin Leads to Delayed Thrombus Growth in Injured Arterioles
Authors: Matuskova Jana , Chauhan Anil K. , Cambien Beatrice , Astrof Sophie , Dole Vandana S. , Piffath Crystal L. , Hynes Richard O. , Wagner Denisa D.
Published: March 2006

Objective—

Plasma fibronectin (FN) is decreased in several clinical conditions. We were interested to study the thrombotic and hemostatic consequences of the decrease in plasma FN (pFN), the role of FN splice variants in thrombosis, and to examine whether pFN incorporates into thrombi in vivo.

Methods and Results—

We compared the thrombotic response to a vessel injury in FN heterozygous (
FN
+/−) mice and corresponding
FN
+/+ mice. Although normal thrombosis in venules was observed, a decrease to half in the pFN concentration in
FN
+/− mice caused a delay in the appearance of thrombi in arterioles and consequently a delay in their occlusion. We were able to rescue the thrombotic defect in the
FN
+/− mice by infusion of rat pFN. Additionally, we could show intense incorporation of fluorescent pFN-coated microspheres into the developing thrombi. Moreover, we found that mice expressing FN without the EIIIA or EIIIB domains specific to cellular FN including platelet FN had no thrombotic defect.

Conclusions—

Mice heterozygous for
FN
have a striking defect in thrombus initiation and growth in arterioles attributable to the decrease of pFN. Our study is an example of haploid insufficiency for FN, and it emphasizes the fundamental role of this plasma protein in thrombosis in the arterial system.
The Elastic Lamellae of Devitalized Arteries Calcify When Incubated in Serum
Authors: Price Paul A. , Si Chan Wai , Jolson Dawn M. , Williamson Matthew K.
Published: March 2006

Objective—

To determine whether serum contains an activity that induces artery calcification.

Methods and Results—

The elastic lamellae of devitalized rat aortas calcify rapidly in rat or bovine serum, or in human serum provided [Pi] ≥2 mmol/L. This calcification is attributable to a potent serum calcification factor (SCF), one that causes devitalized aortas to calcify when incubated in DMEM containing as little as 1.5% serum but not in DMEM alone. The SCF that initiates medial elastin calcification has the same 50- to 150-kDa size and protease sensitivity as the SCF shown previously to initiate calcification of type I collagen. Our working hypothesis is that the same SCF initiates calcification of collagen and elastin, and that this SCF arises from sites of normal bone mineralization and, like alkaline phosphatase, is released into general circulation. The SCF does not initiate medial elastin calcification in living arteries, which suggests that vascular cells may prevent this calcification. This hypothesis is supported by the observations that living arteries secrete the calcification inhibitor matrix Gla protein (MGP); that inactivation of MGP with warfarin causes living arteries to calcify; and that addition of MGP to medium containing warfarin prevents this calcification.

Conclusion—

The elastic lamellae of devitalized aortas calcify rapidly in serum.
Cyclophilin A Is Secreted by a Vesicular Pathway in Vascular Smooth Muscle Cells
Authors: Suzuki Jun , Jin Zheng-Gen , Meoli David F. , Matoba Tetsuya , Berk Bradford C.
Published: March 2006

Reactive oxygen species (ROS) contribute to the pathogenesis of atherosclerosis in part by promoting vascular smooth muscle cell (VSMC) growth. Previously we demonstrated that cyclophilin A (CyPA) is a secreted oxidative stress-induced factor (SOXF) that promotes inflammation, VSMC growth, and endothelial cell apoptosis. However, the mechanisms that regulate CyPA secretion are unknown. In this study, we hypothesized that ROS-induced CyPA secretion from VSMC requires a highly regulated process of vesicle transport, docking, and fusion at the plasma membrane. Conditioned medium and plasma membrane sheets were prepared by exposing VSMC to 1 μmol/L LY83583, which generates intracellular superoxide. A vesicular transport mechanism was confirmed by colocalization at the plasma membrane with vesicle-associated membrane protein (VAMP). CyPA transport to the plasma membrane and secretion were significantly increased by LY83583. Reduction of VAMP-2 expression by small interfering RNA inhibited LY83583-induced CyPA secretion. Pretreatment with 3 μmol/L cytochalasin D, an actin depolymerizing agent, abrogated CyPA secretion. Infection with dominant-negative RhoA and Cdc42 adenovirus inhibited CyPA secretion by 72% and 63%, respectively, whereas dominant-negative Rac1 had a small effect (11%). Pretreatment with the Rho kinase inhibitor Y27632 (3 to 30 μmol/L) and myosin II inhibitor blebbistatin (1 to 10 μmol/L) inhibited CyPA secretion in a dose-dependent manner. Simvastatin (3 to 30 μmol/L) also dose-dependently inhibited LY83583-induced CyPA secretion likely via decreased isoprenylation of small GTPases. Our findings define a novel VSMC vesicular secretory pathway for CyPA that involves actin remodeling and myosin II activation via RhoA-, Cdc42-, and Rho kinase-dependent signaling events.