bluuebunny/crossref_metadata_2025 huggingface dataset:
Electrochemical Nitric Oxide Sensors for Biological Samples – Principle, Selected Examples and Applications
Authors: Bedioui Fethi , Villeneuve Nicole
Published: February 2003

The discoveries made in the 1980s that NO could be synthesized by mammalian cells and could act as physiological messenger and cytotoxic agent had elevated the importance of its detection. The numerous properties of NO, that enable it to carry out its diverse functions, also present considerable problems when attempting its detection and quantification in biological systems. Indeed, its total free concentration in physiological conditions has been established to be in nanomolar range. Thus, detection of nitric oxide remains a challenge, pointing out the difficult dual requirements for specificity and sensitivity. Exception made for the electrochemical techniques, most of the approaches (namely UV‐visible spectroscopy, fluorescence, electron paramagnetic resonance spectroscopy) use indirect methods for estimating endogenous NO, relying on measurements of secondary species such as nitrite and nitrate or NO‐adducts. They also suffer from allowing only ex situ measurements. So, the only strategies that allow a direct and in vivo detection of NO are those based on the use of ultramicroelectrodes. The reality is that surface electrode modification is needed to make the ultramicroelectrode material selective for NO. Therefore, the design of modified electrode surfaces using organized layers is very attractive and provides the ideal strategy. This review addresses a global description of the various approaches that have involved chemically modified microelectrodes specially designed for the electrochemical detection of NO in biological media. Selected significant examples of applications in biological tissues are also reported in order to highlight the importance of this approach in having new insights into the modulatory role of NO in physiology and pathophysiology.
Background and operating parameters in microemulsion electrokinetic chromatography
Authors: Altria Kevin D. , Mahuzier Pierre‐Etienne , Clark Brian J.
Published: February 2003

Microemulsion electrokinetic chromatography (MEEKC) is an electrodriven separation technique. Separations are generally achieved using microemulsions consisting of surfactant‐coated nanometer‐sized oil droplets suspended in aqueous buffer. A cosurfactant such as a short‐chain alcohol is generally used to stabilize the microemulsion. This review summarizes the various microemulsion types and compositions that have been used in MEEKC. The effects of key‐operating variables such as surfactant type and concentration, cosurfactant type and concentration, buffer pH and type, oil type and concentration, use of organic solvent and cyclodextrin additions, and temperature are described. Specific examples of water‐in‐oil microemulsions and chirally selective separations are also covered.
Analysis of selected withanolides in plant extract by capillary electrochromatography and microemulsion electrokinetic chromatography
Authors: Cherkaoui Samir , Cahours Xavier , Veuthey Jean‐Luc
Published: February 2003

Microemulsion electrokinetic chromatography (MEEKC) coupled with a diode‐array detector was developed for the simultaneous analysis of natural steroidal compounds, withanolides including withaferin A, withacnistin and iochromolide. Optimal resolution was obtained with a microemulsion consisting of 70 mM octane, 800 mM 1‐butanol, 100 mM sodium dodecyl sulfate (SDS), and 10 mM phosphate‐borate buffer (pH 7) using a fused‐silica capillary at 25 kV and 40°C. Since this technique is not compatible with mass spectrometry detection, a capillary electrochromatographic method was developed to separate the investigated withanolides. The effects of mobile phase composition and pH were systematically investigated. Complete separation was obtained with a capillary electrochromatography (CEC) Hypersil C18 bonded silica column (packed length, 25 cm×100 μm ID and 375 μm OD), packed with 3 μm particles. The mobile phase consisted of formic acid‐ammonia, pH 8 / acetonitrile (40/60 v/v); the voltage was set at 25 kV and the temperature at 20°C. Under these conditions, resolution of these closely related compounds, including the critical pair withacnistin and iochromolide, was achieved in less than 5 min. The separations by MEEKC and CEC were compared with that obtained by reversed‐phase liquid chromatography and showed similar retention order, indicating the analogy of the retention mechanism of these techniques. To further improve specificity and sensitivity, the developed CEC method was interfaced with electrospray ionization mass spectrometry using a Teflon connection between the CEC column and a void fused‐silica capillary. Finally, the described methods were applied to the qualitative analysis of withanolides in Iochroma gesnerioides plant extract.
Influence of methanol as a buffer additive on the mobilities of organic cations in capillary electrophoresis
Authors: Roy Kimberly I. , Lucy Charles A.
Published: February 2003

The mobilities of a series of aromatic ammonium ions, ranging in charge from +1 to + 3, were investigated by capillary electrophoresis using buffers consisting of 0–75% v/v methanol. This is an extension of our previous studies involving anion mobility in methanol‐water media [1]. Absolute mobilities were determined by extrapolation of the effective mobilities to zero ionic strength according to the Pitts' equation. For all of the buffer compositions studied, the ionic strength effect increased with increasing cation charge, and varied as a function of solvent 1/ηε1/2 as predicted by the electrophoretic term within the Pitts' equation. In the presence of methanol, the ionic strength effects became more dramatic. The absolute mobilities of the cations were altered by the addition of methanol to the electrophoretic media. For example, at 75% MeOH, a migration order reversal was observed between the + 2 and + 3 ammonium ions. These solvent‐induced selectivity changes are attributed to dielectric friction. As predicted by the Hubbard‐Onsager dielectric friction model, dielectric friction increased with increasing methanol content and with increasing analyte charge. Further, the changes in cation mobility correlated to the changes in solvent relaxation time (τ), ε and η. Although not predicted by the Hubbard‐Onsager theory, the + 3 ammonium ion experienced more dielectric friction than the − 3 sulfonate and − 3 carboxylate investigated previously [1]. This apparent failure of the Hubbard‐Onsager model results from its continuum nature, whereby ion‐solvent interactions are not taken into account.
Control of electroosmotic flow in nonaqueous capillary electrophoresis by polymer capillary coatings
Authors: Steiner Frank , Hassel Marc
Published: February 2003

In aqueous capillary electrophoresis the electroosmotic flow (EOF) can be strongly suppressed or eliminated by coating the capillary surface silanols either by buffer additive adsorption or chemical modification. Hydrophilic coatings, e.g., polyvinyl alcohol (PVA) proved to be most efficient for EOF control in applications like DNA analysis. In nonaqueous capillary electrophoresis (NACE), however, the EOF cannot be totally suppressed with these capillaries and coating efficiency turned out to be solvent‐depending. In this paper, fused‐silica capillaries with monomeric and polymeric coatings differing in hydrophobicity and chemical properties (vinyl, vinyl acetate, vinyl alcohol and acrylates with different alkyl chain length) were investigated. Besides studying the EOF characteristics with different organic solvents and water, gas chromatography (GC) measurements were carried out to probe the silanol reduction via ether retention and the surface hydrophobicity by retention of nonane. Good correlations between GC results and EOF magnitude could be found. It could be demonstrated that the polymeric coating has to be solvatized by the buffer solvent to reduce the EOF. The PVA coating was optimal for aqueous systems but not effective for some nonaqueous buffers. On the other hand, polyvinyl acetate and polyethyl acrylate as polymeric coatings proved to be optimal to reduce the EOF in NACE.
Anomalously fast migration of triplet‐repeat DNA in capillary electrophoresis with linear polymer solution
Authors: Kiba Yuriko , Zhang Lihua , Baba Yoshinobu
Published: February 2003

We investigated the capillary electrophoretic behavior of single‐stranded DNA fragments in methylcellulose solution, and found that triplet‐repeat DNA showed anomalously faster mobilities than DNA markers with random sequence. Through the further study on the electrophoretic data, reptation model is proven appropriate to describe the migration of DNA under our experimental conditions. Accordingly, with the equations based on reptation theory, we could obtain the persistence length of DNA fragments and find that these values of triplet‐repeat DNAs are larger than that of DNA markers with random sequence, which means the former DNAs are less flexible than the latter ones when they migrate in the electric field. This phenomenon is supposed to result from the characteristic higher‐order structure formed by GC base pairs within triplet‐repeat DNA, which is further proven by the resumed migration order in accordance with DNA size when the denaturant is added into the sieving matrix.
Stacking phenomena in electromigration: From basic principles to practical procedures
Authors: Urbánek Marek , Křivánková Ludmila , Boček Petr
Published: February 2003

The review describes principles of procedures and techniques used in capillary zone electrophoresis for enhancement of sensitivity that are based on increasing analyte mass in its zone during the electromigration process, for which the term stacking is generally used. Attention is paid to intrinsic stacking in samples with low conductivity, transient isotachophoretic stacking applied in samples with high conductivity, and sweeping in micellar electrokinetic chromatography. Principles of these stacking schemes are explained, new procedures and instrumental arrangements are discussed, and all contributions involving stacking principles that have been published since the year 2000 are surveyed.
High‐sensitive capillary zone electrophoresis analysis by electrokinetic injection with transient isotachophoretic preconcentration: Electrokinetic supercharging
Authors: Hirokawa Takeshi , Okamoto Hikaru , Gaš Bohuslav
Published: February 2003

The principle of an on‐line preconcentration method for capillary zone electrophoresis (CZE) named electrokinetic supercharging (EKS), is described and based on computer simulation the preconcentration behavior of the method is discussed. EKS is an electrokinetic injection method with transient isotachophoretic process, is a powerful preconcentration technique for the analysis of dilute samples. After filling the separation capillary with supporting electrolyte, an appropriate amount of a leading electrolyte was filled and the electrokinetic injection was started. After a while, terminating electrolyte was filled subsequently and migration current was applied. This procedure enabled the introduction of a large amount of sample components from a dilute sample without deteriorating separation. Computer simulation of the electrokinetic injection revealed that EKS was effective for the preconcentration of analytes with wide mobility ranges by proper choice of transient isotachophoresis (ITP) system and electroosmotic flow (EOF) should be suppressed to increase injectable amount of analytes under constant voltage mode. A test mixture of rare‐earth chlorides was used to demonstrate the uses of EKS‐CZE. When a 100 μL sample was used, the low limit of detectable concentration was 0.3 μg/L (1.8 nM for Er), which was comparable or even better than that of ion chromatography and inductively coupled plasma‐atomic emission spectrometry (ICP‐AES).
Chloride present in biological samples as a tool for enhancement of sensitivity in capillary zone electrophoretic analysis of anionic trace analytes
Authors: Křivánková Ludmila , Pantůčková Pavla , Gebauer Petr , Boček Petr , Caslavska Jitka , Thormann Wolfgang
Published: February 2003

Effects originating from the variability of the sample matrix can be efficiently eliminated when the separation conditions are selected so that compounds of like charge with high concentration referred to as macrocomponents are embodied into the system of transient isotachophoresis. For stacking and separation of anionic trace analytes in biological samples, the presence of chloride is shown to be important to balance out effects of other macrocomponents that act against isotachophoretic stacking. Having acetoacetate, malate, citrate, and some drug metabolites in untreated human serum samples, the stacking mechanism of these compounds in an electrolyte system comprising 5 mM mandelic acid and ε‐aminocaproic acid, pH 3.8, is explained. Analytes are monitored by indirect UV‐absorption detection. Attention is paid to the minimum chloride concentration required with respect to the concentration ratio of phosphate (stacker) and lactate (destacker) present in the sample so as to ensure both stacking and separation of trace analytes. Insight into the separation process is given both with computer simulations and experiments. For selected analytes, the effect of chloride concentration on quantitative evaluation, sensitivity and limit of detection is demonstrated as well. Moreover, the applicability of the mobility window between phosphate and lactate for an additional group of metabolites is sketched.
The preparation of background electrolytes in capillary zone electrophoresis: Golden rules and pitfalls
Authors: Beckers Jozef L. , Boček Petr
Published: February 2003

In this article the methodology of the design of suitable background electrolytes (BGEs) in capillary zone electrophoresis (CZE) is described. The principal aspects of the role of a BGE in CZE are discussed with respect to an appropiate migration behavior of analytes, including the transport of the electric current, the buffering of pH, the Joule heat, the electro‐endosmotic flow (EOF) and the principal migration and detection modes. The impact of the composition of the BGE upon migration and detection is discussed. It is shown that the total concentration of the BGE is a principal factor and the adjustment of migrating analyte zones according to the Kohlrausch regulating function (KRF) is the principal effect in most of the sample stacking techniques. The number of co‐ions and their properties are of key importance for peak shapes of the analyte peaks and for the existence of system zones. The detection of UV‐transparent analytes may advanteously be done in the indirect UV mode, by using UV‐absorbing co‐ions, however, both peaks and dips may be expected in the UV trace in case of multiple co‐ionic BGEs. Properties of BGEs can be predicted applying mathematical models and it is shown that with SystCharts, predictions can be given concerning the existence of system zones, detection modes and the peak shapes of analytes for a given BGE. Practical examples of methodological considerations are given in the design of suitable BGEs for four principal combinations of migration and detection modes. The properties of the BGEs selected are exemplified with experimental results. Golden rules are summarized for the preparation of suitable BGEs in CZE.
Ampholytes as background electrolytes in capillary zone electrophoresis: Sense or nonsense? Histidine as a model ampholyte
Authors: Beckers Jozef L.
Published: February 2003

A lot of phenomena, occuring in capillary zone electrophoresis (CZE), are linked with the ionic concentration of the background electrolyte (BGE). If weak bases and acids are used as BGEs in CZE, at a pH where they are scarcely ionized, the ionic concentration of the BGE is very low and this brings a strong peak broadening, limited sample stacking and low sample load. Because the electromigration dispersion increases extremely, moreover, the existence of low‐conductivity BGEs in CZE is a contradiction in terms. The behavior of ampholytes as BGE in CZE is examined, by means of histidine as a model ampholyte. For BGEs consisting of histidine, important parameters, including the ionic concentrations, buffer capacity, transfer ratio, and the indicator for electromigration dispersion E1m1/E2m2, are calculated at various pH. Although the transfer ratio is fairly constant over the whole pH traject, the ionic concentration and buffer capacity decrease whereas the electromigration dispersion strongly increases near the pI of histidine. I.e., that ampholytes can be applied as BGEs in CZE, however, just not at pH near their pI value, except as the difference between the pK values of the basic and acidic group, the δpK value, is very small. For ampholytes with a low δpK value or at high concentrations, all the before‐mentioned effects are less fatal, but in that case we can not speak of a real low‐conductivity BGE. If ampholytes are used at pH near their pK values, the use of ampholytes as BGE is not advantageously compared with simple weak bases and acids. This has been confirmed by calculations and experiments.
Sensitive indirect photometric detection of inorganic and small organic anions by capillary electrophoresis using Orange G as a probe ion
Authors: Johns Cameron , Shaw Matthew J. , Macke Miroslav , Haddad Paul R.
Published: February 2003

This study addresses the two major problems in the use of dyes as highly absorbing probes for indirect photometric detection in capillary electrophoresis (CE). First, effective electroosmotic flow (EOF) modification or suppression to allow separation and detection of a wide mobility range of analytes is not straightforward when electrolytes containing increased dye concentrations are used. The suppression of EOF to less than + 5×10−9 m2V−1s−1 was achieved with a combination of a poly(ethylenimine) (PEI)‐coated capillary and the addition of the neutral polymer hydroxypropylmethylcellulose (HPMC) to the background electrolyte. Second, the deterioration of baselines due to adsorption of the dye probe to the capillary wall is generally a problem. In this work, baseline quality at higher probe concentrations was significantly improved by a rather unusual but highly effective combination of a simultaneous application of a slight overpressure (25 mbar) at the injection end during the separation, and the use of a relatively narrow capillary of 50 μm inner diameter. Both measures would appear to be counterproductive. Optimisation of the probe concentration with regard to signal‐to‐noise ratio resulted in an electrolyte of 4 mM Orange G, 0.05% HPMC buffered at pH 7.7 by the addition of 10.0 mM histidine isoelectric buffer. Very high separation efficiencies of 128 000–297 000 plates were made possible by the relatively high probe concentration. Combined with excellent detection sensitivity, even with the introduction of hydrodynamic flow and a reduced optical path length, these measures resulted in limits of detection ranging from 0.216 to 0.912 μM with a deuterium lamp light source (248 nm) and from 0.147 to 0.834 μM with a 476 nm blue light‐emitting diode (LED) light source. Reproducibility over 30 consecutive runs without changing the electrolyte was excellent, with relative standard deviation (RSD) values of 0.14–0.80% for migration time, 1.27–3.36% for peak area and 0.88–5.12% for peak heights. The optimised electrolyte was used for the analysis of inorganic anions in air filter samples, providing good agreement with results obtained by ion chromatography.
Indirect laser‐induced fluorescence detection for capillary electrophoresis using a frequency‐doubled diode laser
Authors: Ragozina Natalia , Pütz Michael , Faubel Werner , Pyell Ute
Published: February 2003

A blue (452 nm) frequency‐doubled diode laser with a quasi‐cw optical output power of 10 μW is used for indirect laser‐induced fluorescence detection in combination with the capillary electrophoretic separation of inorganic anions. As fluorescing probe ion the anion of 8‐hydroxypyrene‐1,3,6‐trisulfonic acid (HPTS) was selected having an absorption maximum of 454 nm in alkaline medium. Employing a capillary coated with linear acrylamide, baseline separation of eight inorganic anions was possible within 5 min. With a separation buffer containing 50 μmol·L‐1 HPTS and 10 mmol·L‐1 lysine the limits of detection for sulfate, nitrite, nitrate, azide, thiocyanate, and chlorate were between 0.9 and 4.7 μmol·L‐1. Separation of chloride and sulfate was achieved by adding 0.25 mmol·L‐1 calcium hydroxide to the separation buffer. Inorganic anions in several mineral and tap water samples have been determined with the technique developed and results are compared to data obtained by ion chromatography in combination with conductivity detection after conductivity suppression.
Cutting the Wastewater Peaks for Cyclic Batch Production Plants
Authors: Hui C.‐W. , Li B. , Smith R.
Published: February 2003

This paper proposes systematic approaches for cutting the wastewater peaks generated in cyclic batch production plants in which wastewater generation rates and concentration are varied with time. To reduce the investment for the wastewater treatment unit or not to violate the discharge limits, the fluctuations of both flow rate and concentration of the wastewater generated need to be controlled or equalized. Buffer tanks (or basin) are commonly used to regulate the wastewater flow and concentration before the wastewater is sent to treatment units or discharge. The size of the buffer tank needs to be designed properly so that its size is minimized and still has enough capacity to handle various operating conditions. In this paper, conceptual approaches are presented to minimize the size of the buffer tank for either controlling wastewater flow rate or concentration. Mathematical models are proposed for controlling both flow rate and concentration.
New Cells for the Human Retina
Authors: Thumann Gabriele
Published: February 2003

Age‐dependent damage to the retina can lead to blindness. It may be possible to restore vision by transplanting cells from the iris onto the retina
When Squinting becomes a Problem
Authors: Wassill Klaus‐Heiko , Kaufmann Herbert
Published: February 2003

The distortion of vision in a prism is normally pleasurable. However, for people who squint, these undesirable double images can quickly become a nuisance
Psychiatric Penal Sentencing
Authors: Rasch Wilfried
Published: February 2003

Fresh concepts are required for preventive detention. Current regulations result in unjustified commitments to clinics
The Man at the Piano or how Sound came to Image
Authors: Mungen Anno
Published: February 2003

By the mid‐19th century artists were already beginning to enhance the emotional effect of images and moving image sequences through musical expression. Their compositions were the precursors of modern film music
When Crystals grow in the Computer
Authors: Dzuik Gerhard , Schmidt Alfred
Published: February 2003

For many materials in use, the transition from liquid to solid influences internal structures. Novel computer programmes show how micro‐structures grow during solidification and impair their quality
Cell adhesion and matrix remodeling genes identified by co‐expression analysis
Authors: Walker Michael G. , Volkmuth Wayne
Published: February 2003

Cell adhesion and matrix remodeling are elements in many diseases, ranging from atherosclerosis and fibrosis to metastatic cancer. However, many genes that participate in these processes have not yet been identified. To find such genes, we looked for previously uncharacterized genes that are co‐expressed with known cell adhesion and matrix remodeling genes. The known genes in this study included MMP2, TIMP3, BM‐40, chondroitin, connective tissue growth factor, fibromodulin, IGFBP5, laminin, MGP, myosin light chain kinase, several collagens, and other matrix and adhesion proteins. We found eight previously uncharacterized genes, here named MXRA1 through MXRA8, that were strongly co‐expressed with these known adhesion and matrix genes. Five of the MXRA genes have a significant similarity to uncharacterized cDNA sequences or predicted proteins listed in the Genbank database, but otherwise show distant or no sequence similarity to genes with known function. Subsequent to our entry of the MXRA gene sequences in the Genbank, three of the eight genes have been independently described by other researchers: MXRA2 is α‐parvin, a cell‐matrix adhesion protein, MXRA4 is a C1 complement component receptor involved in cell adhesion, and MXRA5 is adlican, an adhesion proteoglycan. The analysis described here provides further evidence for the role of these genes in adhesion and matrix remodeling.
Structural variation in a novel zinc finger protein and investigation of its role in Hirschsprung disease
Authors: Andrew Scott D. , Kuiper Michael J. , Wride Michael A. , Mansergh Fiona C. , Rancourt Derrick E. , Mulligan Lois M.
Published: February 2003

Zinc finger transcription factors play essential roles in neural crest cell development. Even subtle disruptions of the function of these genes could contribute significantly to complex developmental phenotypes such as the multigenic disorder Hirschsprung disease (HSCR), a congenital failure of enteric neurogenesis. Although germline mutations in the RET proto‐oncogene are the most common cause of familial disease, at least 8 genes including transcription factors have been implicated in sporadic HSCR to date. Thus, a further group of candidate genes are those involved in regulating expression of known HSCR genes. In this study, we have characterized the ZNF358 zinc finger gene, a human orthologue of the mouse gene zfend, which is expressed in early developmental stages in the gut and in the neural folds at the time of neural crest differentiation. ZNF358 represents a putative transcription factor with DNA binding activity confered by 9 Cys2His2 zinc fingers. Although we did not detect disease associated mutations in a panel of HSCR patients, we did identify a novel variable sequence within the coding region of ZNF358 that would result in a deletion of nine amino acids from a polyalanine domain. Our molecular model suggests that this variant could alter protein tertiary structure and the ability of ZNF358 to regulate transcription. Together, our data suggest that, while ZNF358 may be involved in the regulation of genes such as those necessary for development or differentiation of neural crest cells, it does not play an obvious role in HSCR.
Single Nucleotide Polymorphism analysis of the human adenosine A2B receptor gene: prevalence of SNPs in asthmatics and normal subjects
Authors: van Heeke Gino , Seamons Rachael , Metais Jean‐Yves , Fozard John R. , Goff Stephen , Wheatley Amanda , Dewar Jane , Hall Ian P.
Published: February 2003

The adenosine A2B receptor is found on human lung mast cells and is believed to mediate the bronchoconstriction in response to adenosine characteristic of asthmatics. As such it represents an attractive therapeutic target for asthma and allergic rhinitis. As genetic variability in drug targets may affect an individual's response to treatment, the adenosine A2B receptor gene was analyzed for polymorphisms and their prevalence defined in a Caucasian population. The coding region of the A2B receptor gene as well as the intron‐exon boundaries of the gene were found to be free of genetic variation. Three single nucleotide polymorphisms were identified in the promoter region of the gene, one of which created a BamHI restriction fragment length polymorphism. The prevalence of the three single nucleotide polymorphisms in an unselected population was <5%. The prevalence of the BamHI polymorphism was investigated in a British population of 40 asthmatics and 40 non‐asthmatics: no excess of the BamHI variant was observed in asthmatic subjects. The data indicates that any variability in drug response would likely not be due to direct structural variation of the A2B receptor protein. Further, polymorphism at the A2B receptor genomic locus is unlikely to contribute to the population risk of developing asthma.
Quantitation of inflammatory und proliferative genes as disease markers in laser‐microdissected, formalin‐fixed and paraffinized glomeruli from human renal biopsies
Authors: Fries Jochen Walter Ulrich , Pakula Alexandra , Roth Tanja , Dienes Hans‐Peter , Odenthal Margarete
Published: February 2003

Organ biopsies from kidney, liver, intestine or heart are presently only evaluated for pathologic lesions using formalin‐fixation and paraffin‐embedding. Changes in pattern and levels of gene expression underlying and preceding these lesions have been disregarded, since a quantitative assay system using minute paraffinized tissue samples with minimal amounts of RNA is currently unavailable. This paper describes such an assay using formalin‐fixed, paraffinized biopsies from human glomeruli after laser‐microdissection. Choosing proliferative glomerular lesions expected to express platelet‐derived growth factor‐β receptor (PDGF‐βR), we investigated whether vascular cell adhesion molecule‐1 (VCAM‐1), a marker of the inflammatory signal pathway, was coexpressed, indicating the potential to progress an already existing lesion. RNA extraction procedures including DNAse pretreatment were pretested on microdissected mouse glomeruli. β‐actin, not upregulated in cell culture by TNF (tumor necrosis factor) or PDGF stimulation, served as housekeeping gene. By qualitative real‐time (RT)‐PCR, transcription of both genes was detectable using only one microdissected glomerular section. Quantitative PCR for VCAM‐1 revealed unsuspected increased levels in the same cases positive by qualitative RT‐PCR, not detectable by morphologic analysis or immunohistochemistry. Thus, quantitative gene expression analysis is possible in paraffinized biopsies from minute tissue samples and will provide such important information as unexpected gene expression causing a potential progression of an existing lesions.
Molecular Recognition of an Amino Acid by a Novel Macrocyclic Monoxo‐tetraamine and Its ZnII Complex
Authors: Gao Jian , Martell Arthur E. , Reibenspies Joseph
Published: February 2003

The macrocycle monoxo‐tetraamine, 1,4,8,11‐tetraazacyclotridecane‐5‐one (mota), was found to be extremely efficient in the recognition of the amino acid, glycine (Gl), in aqueous solution. The single‐crystal structure of a five‐coordinated mota‐ZnII complex is reported. This ZnII complex acts as host for Gl to form a stable ternary complex. The recognition constants and the recognition mechanism for the binary and ternary complexes are based on potentiometric data.
Rapid Cleavage of Oligodeoxyribonucleotides from cis‐Diol‐Bearing Universal Polymer Support
Authors: Kumar Pradeep , Gupta Kailash C.
Published: February 2003

Two sets of conditions have been proposed for the cleavage of oligodeoxyribonucleotides from cis‐diol‐bearing polymer supports and their complete deprotection. The first condition involves the use of spermine in the presence of Zn2+ ions in aqueous NH3 at 60° for 1 h, while the second one employs the use of 0.5M LiOH in conjunction with 3.5M Et3N/MeOH at 75° for 40–60 min, or under microwaves for 5–8 min, depending on the protection employed for nucleic bases.
Synthesis, Enantiomeric Conformations, and Stereodynamics of Aromatic ortho‐Substituted Disulfones
Authors: Lacour Jérôme , Monchaud David , Mareda Jiri , Favarger France , Bernardinelli Gérald
Published: February 2003

Aromatic ortho‐disulfone derivatives are readily accessible from diiodide precursors by CuI‐mediated reaction with sodium sulfinate salts (DMF, 110°). The sulfonyl substituents adopt in solution and in the solid state two enantiomeric conformations (λ and δ) as evidenced by 31P‐ and 1H‐NMR data of the chiral D3‐symmetric tris{4,5‐bis[(4‐methylphenyl)sulfonyl]benzene‐1,2‐diolato(2−)‐κO,κO′}phosphate(v) anion (3a) and 1,2‐bis(camphor‐10‐sulfonyl)‐4,5‐dimethoxybenzene ((=1,2‐bis{{[(1S,4R)‐7,7‐dimethyl‐2‐oxobicyclo[2.2.1]hept‐1‐yl]methyl}sulfonyl}‐4,5‐dimethoxybenzene; 6c). X‐Ray structure analysis of 1,2‐dimethoxy‐4,5‐bis(methylsulfonyl)benzene (6a) and 1,2‐dimethoxy‐4,5‐bis(4‐methylphenyl)sulfonyl]benzene (6b) confirmed in the solid state the preferred chiral orientation of the sulfonyl groups. Dynamic conformational isomerism was detected for 6c in its 1H‐NMR in the temperature range of 110°, the corresponding free energy being 19.8 kcal⋅mol−1.
Novel Enantioselective Synthesis of Both Enantiomers of Furan‐2‐yl Amines and Amino Acids
Authors: Demir Ayhan S. , Sesenoglu Özge , Ülkü Dinçer , Arıcı Cengiz
Published: February 2003

A new enantioselective synthesis of furan‐2‐yl amines and amino acids is described, in which the key step is the oxazaborolidine‐catalyzed enantioselective reduction of O‐benzyl (E)‐ and (Z)‐furan‐2‐yl ketone oximes to the corresponding chiral amines. The chirality of the furan‐2‐yl amines is fully controlled by the appropriate choice of the geometrical isomer of the O‐benzyl oxime. Oxidation of the furan ring furnished amino acids in high yields.
Breast cancer cell line proliferation blocked by the Src‐related Rak tyrosine kinase
Authors: Meyer Tanya , Xu LiHui , Chang Jinli , Liu Edison T. , Craven Rolf J. , Cance William G.
Published: February 2003

Rak is a 54 kDa protein tyrosine kinase originally isolated from breast cancer cells and expressed in epithelial cells. It resembles the protooncogene Src structurally but lacks an amino‐terminal myristylation site and localizes to the nuclear and perinuclear regions of the cell. We report here that expression of Rak in 2 different breast cancer cell lines inhibits growth and causes G1 arrest of the cell cycle. This growth inhibition is kinase‐dependent but does not require the Rak SH2 or SH3 domain. Rak also binds to the pRb tumor‐suppressor protein but inhibits growth even in cells that lack pRb. These results suggest that Rak regulates cell growth by phosphorylating perinuclear proteins and has a function that is distinct from the Src‐related kinase family. © 2002 Wiley‐Liss, Inc.
Reduced expression of transforming growth factor‐β receptors is an unfavorable prognostic factor in human esophageal squamous cell carcinoma
Authors: Fukai Yasuyuki , Fukuchi Minoru , Masuda Norihiro , Osawa Hidenobu , Kato Hiroyuki , Nakajima Takashi , Kuwano Hiroyuki
Published: February 2003

Transforming growth factor‐β (TGF‐β) inhibits epithelial cell proliferation. Inactivation of the TGF‐β signaling pathway is thought to play a role in tumorigenesis. Our purpose was to clarify the correlation between TGF‐β receptors or TGF‐β1 expression and the clinicopathologic characteristics of patients with esophageal squamous cell carcinoma (SCC). Immunohistochemical staining for TGF‐β type I receptor (TGF‐βR‐I), TGF‐βR‐II and TGF‐β1 was performed on surgical specimens obtained from 80 patients with esophageal SCC. Preoperative plasma TGF‐β1 levels were measured and correlated with pathologic features and clinical outcomes. Expression of TGF‐βR‐I and TGF‐βR‐II was reduced in 43 (53.8%) and 23 (28.8%) specimens, respectively. TGF‐β1 was overexpressed in 29 (36.3%). Reduced expression of TGF‐βR‐I and TGF‐βR‐II showed a significant association with depth of invasion (p = 0.0015 and p = 0.0012), lymph node metastasis (p = 0.0309 and p = 0.0059) and pathologic stage (p = 0.0103 and p = 0.0401). Overexpression of TGF‐β1 had a significant association with depth of invasion only (p = 0.0335). Reduced expression of TGF‐βR‐I and TGF‐βR‐II was correlated with cancer‐specific survival (p = 0.0324 and p = 0.0243). The mean preoperative plasma TGF‐β1 level was 10.5 ± 0.8 ng/ml in patients with esophageal carcinoma and was significantly higher compared to healthy controls (p < 0.01). We demonstrate that reduced expression of TGF‐β receptors in esophageal SCC appears to be correlated with depth of invasion, lymph node metastasis, pathologic stage and poor prognosis. TGF‐β receptor expression may play a key role in the progression of this cancer. © 2003 Wiley‐Liss, Inc.
Inhibition of heparanase activity and heparanase‐induced angiogenesis by suramin analogues
Authors: Marchetti Dario , Reiland Jane , Erwin Brad , Roy Madhuchhanda
Published: February 2003

Heparanase, a heparan sulfate‐specific endo‐β‐D‐glucuronidase, plays an important role in tumor cell metastasis through the degradation of extracellular matrix heparan sulfate proteoglycans (ECM HSPG). Heparanase activity correlates with the metastatic propensity of tumor cells. Suramin, a polysulfonated naphthylurea, is an inhibitor of heparanase with suramin analogues shown to possess antiangiogenic and antiproliferative properties. We investigated the effects of selected suramin analogues (NF 127, NF 145 and NF 171) on heparanase activity and heparanase‐driven angiogenesis. Studies of the ability of cellular extracts and purified heparanase from human, highly invasive and brain‐metastatic melanoma (70W) cells revealed that heparanase expressed by these cells was effectively inhibited by suramin analogues in a dose‐dependent manner. These analogues possessed more potent heparanase inhibitory activities than suramin: The concentrations required for 50% heparanase inhibition (IC50) were 20–30 μM, or at least 2 times lower than that for suramin. One hundred percent inhibition was observed at concentrations of 100 μM and higher. Of relevance, these compounds significantly decreased (i) the invasive capacity of human 70W cells by chemoinvasion assays performed with filters coated with purified HSPG or Matrigel™, and (ii) blood vessel formation by in vivo angiogenic assays, thus linking their antiangiogenic properties with impedance of heparanase‐induced angiogenesis. Specifically, inhibition of invasion by NF 127, NF 145 and NF 171 was found at 10 μM concentrations of compounds with a significant decrease of invasive values at concentrations as low as 1.5 μM. In addition, NF 127, NF 145 and NF 171 promoted nearly complete inhibition of heparanase‐induced angiogenesis at values ranging from 236 μM (for NF 145) to 362 μM (for NF 127). These results further emphasize the importance of heparanase in invasive and angiogenic mechanisms and the potential clinical application of heparanase inhibitors such as suramin analogues in cancers and angiogenesis‐dependent diseases. © 2003 Wiley‐Liss, Inc.
α‐tocopherol protects against cisplatin‐induced toxicity without interfering with antitumor efficacy
Authors: Leonetti Carlo , Biroccio Annamaria , Gabellini Chiara , Scarsella Marco , Maresca Vittoria , Flori Enrica , Bove Loredana , Pace Andrea , Stoppacciaro Antonella , Zupi Gabriella , Cognetti Francesco , Picardo Mauro
Published: February 2003

Our aim was 2‐fold: to investigate the role of α‐tocopherol supplementation on the antitumor activity of DDP and to evaluate the effect of α‐tocopherol on the survival and neurotoxicity of DDP‐treated mice. Experiments performed on the M14 human melanoma line demonstrated that α‐tocopherol supplementation did not influence the efficacy of DDP; the inhibition of cell survival and of the in vivo tumor growth after treatment with α‐tocopherol and DDP combination was similar to that observed after DDP alone. Conversely, α‐tocopherol was also able to increase survival of mice treated with a high dose of DDP. While DDP alone produced death in about 70% of mice, the combination reduced deaths to about 30%. Analysis of oxidative stress markers and peroxidative damage in organs indicated that the protective effect of α‐tocopherol was mainly related to its antioxidant activity. A significant increase in the concentration of TBARS and decreased PUFAs and catalase activity were observed after DDP treatment, while with α‐tocopherol the levels of these markers were comparable to those observed in untreated mice. Histologic analysis performed on peripheral nerve revealed that α‐tocopherol also protected mice from severe neurologic damage induced by DDP treatment. In conclusion, our results demonstrate that α‐tocopherol protects against the systemic toxicity and neurotoxicity induced by DDP without interfering with its antitumor activity and suggest that this combination is a promising strategy to improve the therapeutic index of DDP‐based chemotherapy. © 2003 Wiley‐Liss, Inc.
Human papillomavirus infection among women in South and North Vietnam
Authors: Anh Pham Thi Hoang , Hieu Nguyen Trong , Herrero Rolando , Vaccarella Salvatore , Smith Jennifer S. , Thuy Nguyen Thi , Nga Nguyen Hoai , Duc Nguyen Ba , Ashley Rhoda , Snijders Peter J.F. , Meijer Chris J.L.M. , Muñoz Nubia , Parkin D. Max , Franceschi Silvia
Published: February 2003

The incidence rate of invasive cervical carcinoma (ICC) is 4‐fold higher in Ho Chi Minh City, in the South of Vietnam, than in Hanoi, in the North. Thus, we explored the prevalence of and the risk factors for human papillomavirus (HPV) infection in these 2 areas. A population‐based random sample of married women aged 15–69 years were interviewed and had a gynaecological examination in the urban district of Ho Chi Minh City and in a peri‐urban district in Hanoi. HPV DNA detection was performed using a GP5+/6+ primer‐mediated PCR enzyme immunoassay. A total of 922 women from Ho Chi Minh and 994 from Hanoi, for whom a Pap smear and HPV‐status were available, were evaluated. HPV DNA was detected among 10.9% of women in Ho Chi Minh City and 2.0% in Hanoi (age standardized prevalence, world standard population: 10.6% and 2.3%, respectively). In the 2 areas combined, 30 different HPV types were found, the most common being HPV 16 (in 14 single and 18 multiple infections), followed by HPV 58, 18 and 56. A peak of HPV DNA detection in women younger than age 25 was found in Ho Chi Minh City (22.3%) but not in Hanoi. Major risk factors for HPV DNA detection were indicators of sexual habits, most notably the presence of HSV‐2 antibodies, nulliparity and the current use of oral contraceptives. Women in Hanoi showed the lowest HPV prevalence ever reported so far, suggesting that HPV has not spread widely in this population. As expected, HPV prevalence in a population seemed to be closely correlated with ICC incidence rates. © 2003 Wiley‐Liss, Inc.
Mucosal immunoglobulin‐A and ‐G responses to oncogenic human papilloma virus capsids
Authors: Sasagawa Toshiyuki , Rose Robert C. , Azar Khadijeh K. , Sakai Akemi , Inoue Masaki
Published: February 2003

Oncogenic human papilloma virus (HPV) infection is the most important risk factor for developing cervical cancer. It is known that serum antibody responses against these viruses are associated with persistent infection. We conducted an epidemiological study of 627 women to detect cervical mucosal immunoglobulin (Ig)A and IgG responses to oncogenic HPV capsids. Antibody reactivity and cervical HPV infection genotypes were examined by enzyme‐linked immunosorbent assay (ELISA) using HPV types 16, 18, 31, and 45 virus‐like particles, and a polymerase chain reaction‐based method, respectively. HPV infection was defined as being positive for HPV DNA. Multivariate analysis revealed that a mucosal IgA response was associated with the HPV infection, whereas the IgG response was associated with high‐grade cervical squamous intraepithelial lesions (SIL)/squamous cell cancer (SCC) and subject age (40–49 years). IgA was positive in 72% of women with oncogenic HPV infections, whereas IgG was positive in 64% of women with high‐grade SIL/SCC. The longitudinal study demonstrated that the IgA response was elicited earlier than the IgG response, and the IgG response was barely induced in the preclinical HPV infection. However, once an IgG response was induced, it persisted longer after HPV clearance. The mucosal IgA response reflects current HPV infection, whereas an IgG response may be induced with the development of cervical lesions. © 2003 Wiley‐Liss, Inc.
Letrozole as a potent inhibitor of cell proliferation and expression of metalloproteinases (MMP‐2 and MMP‐9) by human epithelial breast cancer cells
Authors: Mitropoulou Theoni N. , Tzanakakis George N. , Kletsas Dimitris , Kalofonos Haralabos P. , Karamanos Nikas K.
Published: February 2003

P450 aromatase catalyzes the conversion of androgens to estrogens and plays a key role in the cell growth of hormone‐dependent breast cancer in postmenopausal women. On the other hand, matrix metalloproteinases (MMPs), which can degrade almost all components of the extracellular matrix, play a crucial role in tumor cell invasion and cancer metastasis. In the present study the effect of letrozole on cell proliferation of estrogen receptor (ER)‐positive MCF‐7 human epithelial breast cancer and MCF‐12A human mammary epithelial cells was studied. The effect of letrozole on the in vitro release of MMPs, particularly type IV collagenases (MMP‐2 and MMP‐9), by the ER‐positive MCF‐7 cells was also investigated, using a solid‐phase method of high sensitivity and accuracy. Using RNA isolates from cell lines MCF‐7 and MCF‐12A, reverse transcriptase‐polymerase chain reaction analysis revealed that only MCF‐7 cells express the P450 aromatase gene. Study of the effects of letrozole alone and the hormones 17‐β‐estradiol, testosterone and 4‐androstene‐3, 17‐dione in the presence and absence of letrozole on cell growth at the DNA synthesis level showed that letrozole significantly suppressed the endogenous aromatase‐induced proliferation of MCF‐7 cells. The majority of MMPs secreted by MCF‐7 cells were identified in their pro‐forms, which was in accordance with the low metastatic potential determined for these cells. After treatment of cells with letrozole (10 nM) for 24 and 48 h, significant inhibition of MMP levels was obtained. Furthermore, concurrent treatment of MCF‐7 cells with 17‐β‐estradiol in the presence of letrozole significantly suppressed the estradiol‐induced stimulation of MMP levels. The data obtained suggest that letrozole is a potent in vitro inhibitor of cell proliferation and of type IV collagenases expressed by ER‐positive MCF‐7 cells and may be of value for suppressing breast tumor growth and invasiveness. © 2003 Wiley‐Liss, Inc.
The product of the candidate prostate cancer susceptibility gene ELAC2 interacts with the γ‐tubulin complex
Authors: Korver Wouter , Guevara Claudia , Chen Yang , Neuteboom Saskia , Bookstein Rob , Tavtigian Sean , Lees Emma
Published: February 2003

ELAC2 is a novel candidate cancer susceptibility gene located on chromosome 17p: Carriers of mutations in ELAC2 display a higher risk of developing prostate cancer. Overexpression of ELAC2 in tumor cells causes a delay in G2‐M progression characterized by accumulation of cyclin B levels. Consistent with a function in mitosis, further biochemical analysis revealed that ELAC2 physically interacts with the γ‐tubulin complex. This is the first biologic insight into the function of this new putative cancer susceptibility gene, providing clues of how perturbation of ELAC2 might promote tumorigenesis through irregular cell division. © 2003 Wiley‐Liss, Inc.
Differentially expressed transcripts in neoplastic hepatic nodules and neonatal rat liver studied by cDNA microarray analysis
Authors: Tellgren Åsa , Wood Timothy J.J. , Flores‐Morales Amilcar , Torndal Ulla‐Britta , Eriksson Lennart , Norstedt Gunnar
Published: February 2003

The molecular mechanisms underlying hepatocarcinogenesis remain unclear. Wistar rats treated with 2‐AAF develop hepatocarcinoma in histologically well‐characterised stages. In our study, cDNA microarrays were used to measure the expression of 3,000 genes during the progression of liver carcinogenesis in persistent neoplastic nodules. Because tumours frequently revert into a more poorly differentiated phenotype, we also studied the expression of the same set of transcripts in neonatal rat liver. Approximately 2,000 transcripts gave a detectable signal in experiments comparing gene expression in nodules and control tissue. Approximately 8% of these were identified as differentially expressed in liver nodules. The differentially expressed genes fell into several categories with putative or demonstrated roles in signal transduction, metabolism, detoxification, cell‐structure and transport. Many of the differentially expressed genes in nodules were not previously known to be regulated during liver carcinogenesis. A universal transcript profile for gene expression in hepatic liver nodules and neonatal liver has been created. © 2003 Wiley‐Liss, Inc.