bluuebunny/crossref_metadata_2025 huggingface dataset:
Tissue‐specific up‐regulation of the proteasome subunit β5i (LMP7) in Sjögren's syndrome
Authors: Egerer Thomas , Martinez‐Gamboa Lorena , Dankof Anja , Stuhlmüller Bruno , Dörner Thomas , Krenn Veit , Egerer Karl , Rudolph Paul E. , Burmester Gerd‐R. , Feist Eugen
Published: April 2006

Objective

Sjögren's syndrome (SS) is characterized by autoimmune infiltration and focal accumulation of lymphocytes in the exocrine glands, with a predominance of CD4‐positive T cells. Since these histologic findings are nonspecific, determination of clinical and serologic abnormalities contribute to the diagnosis. The aim of this study was to identify a novel, disease‐specific, immunologically relevant marker for SS.

Methods

To analyze disease‐related and tissue‐specific expression of candidate markers, we examined biopsied minor salivary glands and peripheral blood mononuclear cells from patients with primary and secondary SS (n = 26) as well as from patients with sicca symptoms without autoimmune sialadenitis (n = 15). Expression of the Th1/Th2‐related chemokines CCL3 (macrophage inflammatory protein 1α) and CCL2 (monocyte chemoattractant protein 1), CXCL7 (neutrophil‐activating peptide 2 [NAP‐2]), interleukin‐1β, inducible costimulator, and the proteasome subunits α3 (C9) and β5i (LMP7) was analyzed at the messenger RNA (mRNA) level using real‐time polymerase chain reaction techniques. Immunohistochemical analysis was used to identify the β5i (LMP7)–expressing cell populations in minor salivary glands.

Results

The expression profiles revealed a significant up‐regulation of β5i (LMP7) exclusively in the salivary glands of SS patients. Immunohistochemistry confirmed expression of the immunoproteasome subunit β5i (LMP7) within the acinar and ductal epithelial cells. No significant difference in the distinct histologic focus scores was evident for the expression of the markers investigated. In the peripheral blood compartment, the expression of CXCL7 was up‐regulated both in primary and in secondary SS.

Conclusion

Tissue‐specific up‐regulation of β5i (LMP7) mRNA was shown to be characteristic of SS, indicating a disease‐specific modulation of the proteasome system. Expression of β5i (LMP7) represents an independent parameter that can be used in addition to the focus score to distinguish SS in biopsied labial salivary glands.
A comparison of response criteria to evaluate therapeutic response in patients with juvenile idiopathic arthritis treated with methotrexate and/or anti–tumor necrosis factor α agents
Authors: Lurati A. , Pontikaki I. , Teruzzi B. , Desiati F. , Gerloni V. , Gattinara M. , Cimaz R. , Fantini F.
Published: April 2006

Objective

There are no validated criteria to evaluate clinical response in juvenile idiopathic arthritis (JIA). The purpose of this study was to compare 4 sets of criteria (2 from the American College of Rheumatology [ACR] and 2 from the European League Against Rheumatism [EULAR]) for clinical response evaluation in JIA patients treated with methotrexate and/or anti–tumor necrosis factor α drugs.

Methods

Seventy‐five patients with JIA were evaluated at baseline and after 6 months of therapy with second‐line drugs. Mean age at study onset was 12.8 years (range 2–32.9 years). Diagnoses were systemic JIA (n = 16), rheumatoid factor–positive JIA (n = 5), rheumatoid factor–negative JIA (n = 9), persistent oligoarticular JIA (n = 10), extended oligoarticular JIA (n = 33), and psoriatic arthritis (n = 2). Clinical response was evaluated with the ACR Pediatric 30 criteria and the ACR 20% response criteria (ACR20), and with the EULAR Disease Activity Score (DAS) and 28‐joint DAS (DAS28). Patients with EULAR criteria responses of “good” or “moderate” were classified as responders. Responders and nonresponders according to the different criteria were then compared.

Results

For patients younger than 16 years, Cohen's kappa varied between 0.51 and 0.72, with a good‐to‐excellent reproducibility index for all comparisons, except for the DAS28/ACR20 comparison. The best agreement was obtained by comparing the DAS and the ACR Pediatric 30. For patients older than 16 years, the reproducibility index was good or excellent in only 2 cases, i.e., comparing the DAS and the ACR Pediatric 30 and comparing the DAS and the DAS28 (as expected).

Conclusion

Our study shows a good agreement overall for the different criteria tested. The highest concordance was observed between the DAS and the ACR Pediatric 30, the lowest between the DAS28 and the ACR20. Our data suggest that the ACR Pediatric 30 criteria can be used also in adult patients affected by JIA, and that the original DAS can be an alternative to the ACR Pediatric 30 in both children and young adults with JIA.
Major reduction in spinal inflammation in patients with ankylosing spondylitis after treatment with infliximab: Results of a multicenter, randomized, double‐blind, placebo‐controlled magnetic resonance imaging study
Authors: Braun Jürgen , Landewé Robert , Hermann Kay‐Geert A. , Han John , Yan Songkai , Williamson Paul , van der Heijde Désirée
Published: April 2006

Objective

To determine whether the effects of anti–tumor necrosis factor α (TNFα) in reducing the signs and symptoms of ankylosing spondylitis (AS) coincide with a reduction in spinal inflammation as detected by magnetic resonance imaging (MRI).

Methods

Pre‐ and postgadolinium T1 and STIR MR images of the spine were acquired at baseline and at week 24 in patients with AS who participated in a multicenter, randomized, double‐blind, placebo‐controlled study. Patients were randomly assigned at an 8:3 ratio to receive infusions of infliximab (5 mg/kg) or placebo at weeks 0, 2, and 6 and then every 6 weeks thereafter. MR images were obtained and evaluated independently by 2 readers who were blinded to the treatment allocation and time sequence of the images.

Results

A total of 194 patients in the infliximab group and 72 patients in the placebo group had evaluable images at baseline and week 24. About 80% of the patients had at least 1 active spinal lesion at baseline, as assessed by MRI. The improvement in the MRI Activity Score after 6 months was significantly greater in the patients who received infliximab (mean 5.02, median 2.72) than in those who received placebo (mean 0.60, median 0.0) (P < 0.001). Almost complete resolution of spinal inflammation was seen in most patients who received infliximab, irrespective of baseline activity.

Conclusion

Patients with AS who received infliximab therapy showed a decrease in spinal inflammation as detected by MRI, whereas those who received placebo showed persistent inflammatory spondylitis.
Linkage analysis of rheumatoid arthritis in US and UK families reveals interactions between HLA–DRB1 and loci on chromosomes 6q and 16p
Authors: John Sally , Amos Christopher , Shephard Neil , Chen Wei , Butterworth Adam , Etzel Carol , Jawaheer Damini , Seldin Michael , Silman Alan , Gregersen Peter , Worthington Jane
Published: April 2006

Objective

HLA is the most strongly associated locus in rheumatoid arthritis (RA), accounting for up to one‐third of the genetic contribution. Conditioning on the effect of true disease loci such as HLA can lead to increased power to detect effects at other loci and, in addition, allows investigation of the underlying disease models, including interactions. The aim of this study was to detect susceptibility loci for RA by conditioning on HLA in a large sample of affected sibling pairs (ASPs) and to test for evidence of interaction between novel loci and HLA.

Methods

Genotype data from 3 whole‐genome linkage scans for RA in a US population and a UK population were pooled, resulting in a combined data set of 886 ASPs. This pooling of data increased the power to detect loci showing low levels of heterogeneity. Nonparametric linkage analysis was performed to identify regions of interest. Joint 2‐locus analysis was then performed for HLA and each of the loci that demonstrated evidence of linkage in the 886 ASPs.

Results

Evidence for linkage was most significant at HLA (P = 4 × 10−16), with 7 non‐HLA loci showing some evidence for linkage (P = 0.05–0.003). Joint modeling of these loci with HLA provided evidence for linkage at a genome‐wide significance level for loci on 6q (P = 2.7 × 10−6) and 16p (P = 2 × 10−4).

Conclusion

These data provide the most convincing evidence to date that 6q and 16p harbor susceptibility genes. In addition, these loci may interact with HLA, facilitating the search for candidate genes within this region.
Importance of genetic influences on chronic widespread pain
Authors: Kato Kenji , Sullivan Patrick F. , Evengård Birgitta , Pedersen Nancy L.
Published: April 2006

Objective

To estimate the relative importance of genetic and environmental factors in chronic widespread pain, and to assess whether there are sex differences in the type or magnitude of these influences.

Methods

Data were collected from a national sample of twins ≥42 years of age, all of whom were participants in the Swedish Twin Registry. The presence of chronic widespread pain was assessed via computer‐assisted telephone interviews, which were conducted between 1998 and 2002, using the American College of Rheumatology criteria for fibromyalgia. No clinical examinations were performed. In preliminary analyses, probandwise concordance rates and tetrachoric correlations were calculated. Structural equation modeling was then performed to estimate additive genetic, shared environmental, and nonshared environmental sources of variability in susceptibility for the development of chronic widespread pain.

Results

Of 61,355 eligible twins, 44,897 individuals (73.2%) responded to the interview. Both members of 15,950 pairs responded to the items regarding pain symptoms; of these pairs, 4,170 were monozygotic, 5,881 were same‐sex dizygotic, and 5,755 were opposite‐sex dizygotic. The prevalence of chronic widespread pain was 4.1%, and the ratio of women to men was 3.3 to 1. Probandwise concordance rates and tetrachoric correlations suggested modest genetic influences for both women and men. Genetic and shared environmental influences explained approximately half of the total variance, with no indication of sex differences in either the type or magnitude of these influences.

Conclusion

Individual differences in the likelihood of developing chronic widespread pain reflect modest genetic influences. There are no significant sex differences in the type or expression of the genes responsible for chronic widespread pain or in the magnitude of the relative importance of these influences on chronic widespread pain.
Performance of an automated computer‐based scoring method to assess joint space narrowing in rheumatoid arthritis: A longitudinal study
Authors: Finckh Axel , de Pablo Paola , Katz Jeffrey N. , Neumann Gesa , Lu Ying , Wolfe Frederick , Duryea Jeffrey
Published: April 2006

Objective

To compare the diagnostic performance of a computer‐based method for measuring joint space width with the Sharp joint space narrowing (JSN) scoring method in patients with rheumatoid arthritis (RA).

Methods

A random sample of patients with early RA, for whom sequential hand radiographs and Sharp scores were available, was selected from the National Data Bank for Rheumatic Diseases. Hand joint space width was measured using an automated, computer‐based method in random order and with blinding for clinical information. We constructed a receiver operating characteristic curve and compared the diagnostic performance of the computer‐based and Sharp methods based on the areas under the curve.

Results

One hundred twenty‐nine patients with early RA who underwent serial radiography were included. Changes in the computer‐based and Sharp methods were highly correlated (r = 0.75, P < 0.001). The computer‐based method was significantly more discriminant than the Sharp JSN subscale. The area under the curve of the computer‐based method was 0.96 (95% confidence interval [95% CI] 0.94, 0.99) compared with 0.93 (95% CI 0.89, 0.96) for the Sharp subscale (P = 0.024). At the most discriminant cutoff, specificity of the computer‐based method was 88.4% compared with 81.4% for the Sharp subscale (P = 0.11); sensitivity was 87.6% for the computer‐based method compared with 82.2% for Sharp subscale (P = 0.19). The signal‐to‐noise ratio for the computer‐based method was 83% compared with 70% for the Sharp subscale (P = 0.013).

Conclusion

The computer‐based method for measuring joint space width is more discriminant than the semiquantitative Sharp JSN subscale.
Hyaluronan molecular reflection by synovial lining is concentration dependent and reduced in dilute effusions in a rabbit model
Authors: Sabaratnam S. , Mason R. M. , Levick J. R.
Published: April 2006

Objective

Hyaluronan (HA) has a major role in regulating synovial fluid volume. This role depends on the synovium functioning as an ultrafilter that reflects HA during trans–synovial fluid drainage. Reflection boosts the HA concentration on the membrane surface, leading to osmotic retention of synovial fluid (“buffering”). In arthritic effusions, however, HA concentration and osmotic buffering are greatly reduced. We tested the hypothesis that reflection is reduced (escape increased) when the HA concentration falls below the molecular entanglement concentration (C*).

Methods

HA at 0.2 mg/ml (C*) was infused continuously into rabbit knee joints to set up a steady trans‐synovial filtration. Joint‐derived lymph was sampled over 3 hours, and subsynovial fluid was sampled at the end of the 3‐hour period. HA was quantified by high‐performance liquid chromatography to evaluate the reflected fraction. C* was determined by viscometry.

Results

Viscometry showed that 0.2 mg/ml HA was below C* and 1.5 mg/ml was above it. At 0.2 mg/ml, the mean ± SEM HA reflected fraction was 0.66 ± 0.04 (n = 7). At 1.5 mg/ml the reflection increased to 0.88 ± 0.04 (n = 5) (P < 0.005). HA permeation increased almost 3‐fold, from 12% to 34%, at the lower concentration.

Conclusion

Chain–chain interaction at >C* increases effective molecular domain size and hence HA reflection, promoting effective conservation of synovial fluid in normal joints. HA can fall below C* (∼1 mg/ml) in arthritic effusions, promoting loss of HA. The attendant failure of outflow buffering facilitates fluid escape and periarticular edema.
Recovery from deafness in a patient with Muckle‐Wells syndrome treated with anakinra
Authors: Mirault Tristan , Launay David , Cuisset Laurence , Hachulla Eric , Lambert Marc , Queyrel Viviane , Quemeneur Thomas , Morell‐Dubois Sandrine , Hatron Pierre‐Yves
Published: April 2006

Muckle‐Wells syndrome (MWS) is a dominantly inherited autoinflammatory disease characterized by rashes, fever, arthralgia, sensorineural deafness, and the possible development of systemic AA amyloidosis. We used anakinra to treat a 22‐year‐old patient with MWS who had deafness and a high serum level of C‐reactive protein (CRP). Following treatment with anakinra, the patient's CRP level normalized, and she recovered from deafness. The fact that this occurrence has never been previously reported strengthens the role of anakinra in MWS but also raises new questions about the physiopathology of such deafness.
Followup radiographic data on patients with rheumatoid arthritis who participated in a two‐year trial of prednisone therapy or placebo
Authors: Jacobs Johannes W. G. , Everdingen Amalia A. Van , Verstappen Suzan M. M. , Bijlsma Johannes W. J.
Published: April 2006

Objective

In a previous clinical trial of patients with early rheumatoid arthritis (RA), it was determined that patients who received 10 mg of prednisone per day for 2 years had less radiographic joint damage compared with those who received placebo. Our goal was to investigate whether this beneficial effect persisted after the end of the trial.

Methods

A blinded assessment of radiographic joint damage was performed ∼3 years after the end of the original 2‐year study. Twenty‐four patients from the original prednisone group (60%) and 28 patients from the original placebo group (68%) participated in this followup study. At the end of the original trial, prednisone dosages were tapered down in the prednisone group and stopped, if possible. Patients from the original prednisone group took prednisone during 35% of the followup period (∼1 year) at a mean daily dose of ∼5 mg. Two patients from the original placebo group started taking prednisone during followup. Radiographs of the hands and feet were scored according to the van der Heijde modification of the Sharp method.

Results

During 3 additional years of followup, radiographic scores showed significantly less progression in the original prednisone group than in the original placebo group. Radiographic damage in the original prednisone group did not show an accelerated rate of progression during the followup period.

Conclusion

The inhibition of radiographic joint damage in patients with early active RA treated with 10 mg of prednisone per day for 2 years seems to persist after the end of prednisone therapy.
Vascular endothelial growth factor gene haplotypes in Kawasaki disease
Authors: Breunis W. B. , Biezeveld M. H. , Geissler J. , Ottenkamp J. , Kuipers I. M. , Lam J. , Hutchinson A. , Welch R. , Chanock S. J , Kuijpers T. W.
Published: April 2006

Objective

To investigate whether common genetic variants in the vascular endothelial growth factor (VEGF) gene are associated with Kawasaki disease (KD) and the subsequent development of coronary artery lesions.

Methods

Common genetic variants in the VEGF gene were analyzed in an association study in a Dutch cohort of 170 KD patients and 300 healthy Dutch Caucasian controls. Genotyping was done with 5′‐nuclease TaqMan assays and 3′‐hybridization–triggered fluorescence minor groove binder Eclipse assays.

Results

An association with susceptibility to KD was observed with 2 of the 6 single‐nucleotide polymorphisms analyzed in VEGF: –2594 A>C (rs699947) and the 236 bp 3′ of STP C>T (rs3025039). Also for an 18‐bp deletion in the promoter of VEGF a significant difference in the genotype and allele frequencies was observed between the KD patients and the controls. The haplotype CGCC (based on rs699947, rs2010963, rs25648, and rs3025039) was significantly associated with the development of KD (hap score 3.8; P = 0.0002). VEGF plasma levels were significantly higher in patients with the early phase of KD than in the healthy controls, and there was a trend toward higher VEGF plasma levels in KD patients with the –2594 CC and 236 bp 3′ of STP CC genotypes.

Conclusion

Our results suggest that polymorphisms of the VEGF gene may play a role in the pathogenesis of KD.
Antibodies to ribosomal P proteins: A potential serologic marker for lupus membranous glomerulonephritis
Authors: do Nascimento Ana Patrícia , dos Santos Trindade Viana Vilma , de Abreu Testagrossa Leonardo , Leon Elaine Pires , Borba Eduardo Ferreria , Barros Rui Toledo , Bonfá Eloísa
Published: April 2006

Objective

To evaluate the relevance of antibodies to ribosomal P proteins (anti‐P antibodies) in discriminating histopathologic patterns of lupus nephritis.

Methods

The study group comprised 81 consecutive patients with systemic lupus erythematosus who underwent renal biopsy and for whom frozen serum was available at the time of biopsy. All biopsy specimens were reviewed in a blinded manner, according to the 2004 criteria of the International Society of Nephrology and the Renal Pathology Society. Anti‐P antibodies were detected by enzyme‐linked immunosorbent assay (ELISA)/immunoblot analysis, and anti–double‐stranded DNA (anti‐dsDNA) was detected by indirect immunofluorescence/ELISA.

Results

Anti‐P antibodies were detected in 18 patients (22%). The demographic and clinical features of patients with and those without anti‐P antibodies were similar. Remarkably, analyses of biopsy specimens revealed that the frequency of anti‐P antibodies in patients with class V lupus nephritis was higher than the frequency among patients with other classes of renal disease (72% versus 28%; P = 0.005). Accordingly, anti‐P antibody–positive patients had a higher mean (±SD) proteinuria level compared with anti‐P antibody–negative patients (6.4 ± 4.8 versus 4.7 ± 3.9 gm/dl; P = 0.046). Renal function was preserved in 6 of 7 patients who had both isolated anti‐P antibodies and class V lupus nephritis. In contrast, anti‐dsDNA was associated with proliferative‐class lupus nephritis (P = 0.050) and higher creatinine levels (P = 0.014). Furthermore, 7 of 9 patients with isolated anti‐P antibodies had class V lupus nephritis, and, more importantly, 5 of these 7 patients (71%) displayed a pure membranous pattern. Conversely, a tendency toward the predominance of class V lupus nephritis (67%) with concomitant proliferative lesions was observed when anti‐P antibody was associated with anti‐dsDNA.

Conclusion

Our data introduce anti‐P antibody as a novel serologic marker for membranous lupus nephritis and support the notion that the presence of isolated anti‐P antibodies may discriminate patients with pure class V lupus nephritis, whereas the simultaneous presence of anti‐dsDNA antibodies suggests class V disease with concomitant proliferative lesions.
Cardiovascular outcomes in new users of coxibs and nonsteroidal antiinflammatory drugs: High‐risk subgroups and time course of risk
Authors: Solomon Daniel H. , Avorn Jerry , Stürmer Til , Glynn Robert J. , Mogun Helen , Schneeweiss Sebastian
Published: April 2006

Objective

Controversy persists regarding the cardiovascular risks of treatment with selective cyclooxygenase 2 inhibitors (coxibs) and nonselective nonsteroidal antiinflammatory drugs (NSAIDs). This study was undertaken to examine, in a large group of new users of coxibs and NSAIDs, the rate of cardiovascular events, their time course, and whether baseline cardiovascular risk modified the rate ratios (RRs) for future events.

Methods

This cohort study included Medicare beneficiaries who enrolled in a state‐run prescription drug plan that fully covered NSAIDs and coxibs without restriction. All study patients started use of a coxib or NSAID after January 1, 1999. The primary composite end point was a hospital admission for either myocardial infarction or ischemic stroke. Predefined exposure groups included the 3 coxibs available in the US during the study period (celecoxib, rofecoxib, and valdecoxib), as well as oral formulations of diclofenac, ibuprofen, naproxen, and a composite of all other NSAIDs. We compared the rate of cardiovascular events associated with each of these agents with that in a reference group of patients who did not use NSAIDs or coxibs, but started other medications unrelated to cardiovascular risk. Daily exposure to all study drugs was assessed based on filled prescription data. A Cox proportional hazards model stratified on calendar year that included other baseline cardiovascular risk factors constituted the primary analysis.

Results

We identified 74,838 users of NSAIDs or coxibs, and 23,532 comparable users of other drugs comprised the reference group. Adjusted models demonstrated a significant elevation in the event rate for rofecoxib (RR 1.15, 95% confidence interval [95% CI] 1.06–1.25) and a significant reduction in the rate for naproxen (RR 0.75, 95% CI 0.62–0.92). No other coxib or NSAID was associated with a significant increase or decrease in cardiovascular event rate. The increased rate associated with rofecoxib was seen in the first 60 days of use (adjusted RR 1.14, 95% CI 1.01–1.29) and thereafter (adjusted RR 1.14, 95% CI 1.02–1.28). Kaplan‐Meier event curves showed a similar pattern of risk (early and persistent separation of the event curves) among long‐term rofecoxib users at low or high baseline cardiovascular risk.

Conclusion

We found an increased cardiovascular event rate among users of rofecoxib, and a decreased rate with naproxen use. Other coxibs and NSAIDs did not appear to be associated with a difference in event rate compared with users of other drugs. The increase in rate associated with rofecoxib was seen within the first 60 days and persisted. There was no important modification of the event rate based on the patient's baseline cardiovascular risk.
On‐line preparation of peroxymonocarbonate and its application for the study of energy transfer chemiluminescence to lanthanide inorganic coordinate complexes
Authors: Liu Meilin , Cheng Xianglei , Zhao Lixia , Lin Jin‐Ming
Published: April 2006

It has been shown that peroxymonocarbonate ion (HCO4−) is a potent oxidant. In this study, a flow‐injection system was developed in order to prepare on‐line HCO4− ion and the optimum conditions for the on‐line preparation of HCO4− were studied in detail. We used 99% 13C‐enriched NaHCO3 to examine peroxymonocarbonate by 13C‐NMR at 25°C. An ultra‐weak chemiluminescence (CL) was observed after mixing H2O2 and sodium bicarbonate in an organic co‐solvent that can accelerate the formation of HCO4− ion. When lanthanide inorganic coordinate complex, Eu(II)–EDTA, was added into this HCO4− system, the CL intensity was significantly enhanced. The CL mechanism was investigated by various methods. The experimental results indicate that peroxymonocarbonate oxidizes Eu(II) to Eu(III) and produces singlet oxygen; meanwhile, the energy originating from dimers of singlet oxygen is accepted by the Eu(III)–EDTA− complex. The excited Eu(III) ions undergo radiative deactivation and emit CL. Copyright © 2006 John Wiley & Sons, Ltd.
Firefly Luciferase Produces Hydrogen Peroxide as a Coproduct in Dehydroluciferyl Adenylate Formation
Authors: Fraga Hugo , Fernandes Diogo , Novotny Jiri , Fontes Rui , Esteves da Silva Joaquim C. G.
Published: April 2006

Firefly luciferase catalyzes the synthesis of H2O2 from the same substrates as the bioluminescence reaction: ATP and luciferin (d‐LH2). About 80 % of the enzyme‐bound intermediate d‐luciferyl adenylate (d‐LH2‐AMP) is oxidized into oxyluciferin, and a photon is emitted during this reaction. The enzyme pathway responsible for the generation of H2O2 is a side reaction in which d‐LH2‐AMP is oxidized into dehydroluciferyl adenylate (L‐AMP). Like the bioluminescence reaction, the luciferase‐catalyzed synthesis of H2O2 and L‐AMP is a stereospecific process, involving only the natural D enantiomer. However, the intramolecular electron transfer postulated as essential to the light emission process is not involved in this side reaction.
Reaction of Cytidine Nucleotides with Cyanoacetylene: Support for the Intermediacy of Nucleoside‐2′,3′‐cyclic Phosphates in the Prebiotic Synthesis of RNA
Authors: Crowe Michael A. , Sutherland John D.
Published: April 2006

A robust and prebiotically plausible synthesis of RNA is a key requirement of the “RNA World” hypothesis, but, to date, no such synthesis has been demonstrated. Monomer synthesis strategies involving attachment of preformed nucleobases to sugars have failed, and, even if activated 5′‐nucleotides could be made, the hydrolysis of these intermediates in water makes their efficient oligomerisation appear unlikely. We recently reported a synthesis of cytidine‐2′,3′‐cyclic phosphate 1 (C>p) in which the nucleobase was assembled in stages on a sugar‐phosphate template. However, 2′,3′‐cyclic nucleotides (N>p's) also undergo hydrolysis, in this case giving a mixture of the 2′‐ and 3′‐monophosphates. This hydrolysis has previously been seen as making the, otherwise promising, oligomerisation of N>p's seem as unlikely as that of the 5′‐activated nucleotides. We now find that cyanoacetylene, the reagent used for the second stage of nucleobase assembly in the synthesis of C>p, also reverses the effect of the hydrolysis by driving efficient cyclisation of C2′p and C3′p back to C>p. Excess cyanoacetylene also derivatises the nucleobase, but this modification is reversible at neutral pH. These findings significantly strengthen the case for N>p's in a prebiotic synthesis of RNA.
Characterization of BlsM, a Nucleotide Hydrolase Involved in Cytosine Production for the Biosynthesis of Blasticidin S
Authors: Grochowski Laura L. , Zabriskie T. Mark
Published: April 2006

Biosynthesis of the antifungal agent blasticidin S in Streptomyces griseochromogenes requires the formation of free cytosine. The blsM gene in the blasticidin S gene cluster is predicted to encode a protein that has sequence homology with several nucleoside transferases. In vitro analysis of recombinant BlsM revealed that the enzyme functions as a nucleotide hydrolase and catalyzes the formation of free cytosine by using cytidine 5′‐monophosphate (CMP) as the preferred substrate. Cytosine production was significantly lower with CDP, CTP, and dCMP as alternate substrates. BlsM was also observed to have low‐level cytidine deaminase activity, converting cytidine and deoxycytidine to uridine and deoxyuridine, respectively. Point mutations were introduced in blsM at putative catalytic residues to generate three mutant enzymes, BlsM Ser98Asp, Glu104Ala, and Glu104Asp. All three mutants lost CMP hydrolysis activity, but the Ser98Asp mutant showed a modest increase in cytidine deaminase activity.
Modulation of Infectivity in Phage Display as a Tool to Determine the Substrate Specificity of Proteases
Authors: Chaparro‐Riggers Javier F. , Breves Roland , Maurer Karl‐Heinz , Bornscheuer Uwe
Published: April 2006

Proteases play an important role in human and animal diseases. Rapid determination of substrate specificity is possible through the use of substrate phage display; however, current methods possess several drawbacks. They require phage‐immobilization and cannot be used for infectivity‐destroying or affinity tag‐destroying proteases; this can make entire libraries useless. To overcome these limitations, here we introduce infectivity‐modulated phage display (IMOP). IMOP uses a protease‐resistant and infectivity‐reducing tag fused to substrate‐displaying polyvalent phages, and the specific cleavage of the substrate increases the infectivity of the phages by releasing the infectivity‐reducing tag. The resulting phages were first tested with the infectivity‐destroying detergent protease subtilisin; this resulted in a highly specific substrate at a 200‐fold enrichment. In a second example, the protease ompT was used and led to an enrichment of the known double‐arginine motif. The IMOP system thus substantially improves and simplifies previous systems.
Heteropolynuclear Palladium Complexes with Pyrazolate and Its 3‐tert‐Butyl Derivatives: The Effect of Heterometal Ions on the Rate of Isomerization
Authors: Umakoshi Keisuke , Kojima Takashi , Arikawa Yasuhiro , Onishi Masayoshi
Published: April 2006

The heteropolynuclear complexes [Pd2M′2(μ‐pz)6] (M′=Ag (1), Au (2); pzH=pyrazole), HT‐[Pd2M′2(μ‐3‐tBupz)6] (M′=Ag (3 a), Au (4 a); 3‐tBupzH=3‐tert‐butylpyrazole), and HH‐[Pd2Au2(μ‐3‐tBupz)6] (4 b) have been prepared and some of them were structurally characterized. When 3‐tert‐butylpyrazolate was employed as a bridging ligand, two linkage isomers (head‐to‐tail (HT) and head‐to‐head (HH)) arise from the difference in orientation of the substituent groups on the pyrazolate bridges between the two Pd atoms. 1H NMR spectroscopy has been used to identify and to follow the reversible stereochemical rearrangement of the HH isomer of [Pd2Ag2(μ‐3‐tBupz)6] (3 b) to form the HT isomer 3 a in CDCl3 and the HT isomer of [Pd2Au2(μ‐3‐tBupz)6] (4 a) to form the HH isomer 4 b in C6D6. Kinetic studies of the reaction have established the rate law to be −d(HH)/dt=d(HT)/dt=k2[HH]−k1[HT] for 3 b and −d(HT)/dt=d(HH)/dt=k1[HT]−k2[HH] for 4 a, where k1 and k2 denote the rate of isomerization from the HT to the HH isomer and that from the HH to the HT isomer, respectively. For typical runs at 50 °C in C6D6, k1=13.8×10−5 s−1, k2=18.6×10−5 s−1, and Keq=k2/k1=1.24 for 3 b, and k1=1.26×10−5 s−1, k2=3.52×10−5 s−1, and Keq=k1/k2=0.36 for 4 a. Temperature‐dependent rate measurements reveal ΔH≠ and ΔS≠ to be 100(1) kJ mol−1 and 0(3) J mol−1 K−1 for 3 b and 112(5) kJ mol−1 and 20(17) J mol−1 K−1 for 4 a, respectively. The rate of isomerization is essentially unaffected by the concentration of the complex or by the presence of neutral bridging ligands. These data and observations imply that the isomerization involves an intramolecular exchange process.
trans–cis Photoisomerization of the Styrylpyridine Ligand in [Re(CO)3(2,2′‐bipyridine)(t‐4‐styrylpyridine)]+: Role of the Metal‐to‐Ligand Charge‐Transfer Excited States
Authors: Bossert Julien , Daniel Chantal
Published: April 2006

The trans‐cis isomerization of the styrylpyridine carbon–carbon double bond induced by visible light irradiation in fac‐[Re(CO)3(bpy)(stpy)]+ (bpy = 2,2′‐bipyridine; stpy = t‐4‐styrylpyridine) has been investigated by means of quantum‐chemical methods. The structures of the various cis and trans conformers of [Re(CO)3(bpy)(stpy)]+ have been optimized at the density functional theory (DFT) level. Three rotational conformers for the most stable trans isomer lie within 2.3 kJ mol−1 each other. The energy difference between the cis and trans isomers is 27.0 kJ mol−1. The electronic spectroscopy of the most stable conformers has been investigated by time‐dependent DFT (TD‐DFT) and complete active space self‐consistent field/CAS second order perturbation theory (CASSCF/CASPT2) calculations. The lowest absorption bands are dominated by metal‐to‐ligand charge‐transfer (MLCT, dRe→π*bpy) transitions calculated at about 25 000 cm−1 and by a strong intraligand 1IL (πstpy→π*stpy) transition in the near UV region. On the basis of CASSCF potential energy curves (PECs) calculated as a function of the torsion angle of the CC bond of the styrylpyridine ligand, it is shown that the role of the low‐lying MLCT states is important in the photoisomerization mechanism. In contrast to the free organic ligand, in which the singlet mechanism is operational via the 1IL (S1) and electronic ground (S0) states, coordination to the rhenium steers the isomerization to the triplet PEC corresponding to the 3IL state. From the 3ILt (t = trans) the system evolves to the perpendicular intermediate 3ILp (p = perpendicular) following a 90° rotation around the styrylpyridine CC bond. The metal center acts as a photosensitizer because of the presence of photoactive MLCT states under visible irradiation. The position of the crossing between the 3IL and electronic ground state PEC determines the quantum yield of the isomerization process.
High‐Spin‐ and Low‐Spin‐State Structures of [Fe(chloroethyltetrazole)6](ClO4)2 from Synchrotron Powder Diffraction Data
Authors: Dova Eva , Peschar René , Sakata Makoto , Kato Kenichi , Schenk Henk
Published: April 2006

The spin‐crossover complex [Fe(teec)6](ClO4)2 (teec = chloroethyltetrazole) exhibits a 50 % incomplete spin crossover in the temperature range 300–30 K. Time‐resolved synchrotron powder diffraction experiments have been carried out to elucidate its structural behavior. We report crystal structure models of this material at 300 K (high spin) and 90 K (low spin), as solved from synchrotron powder diffraction data by using Genetic Algorithm and Parallel Tempering techniques and refined with Rietveld refinement. During short synchrotron powder diffraction experiments (five minutes duration) two distinguishable lattices were observed the quantities of which vary with temperature. The implication of this phenomenon, that is interpreted as a structural phase transition associated with the high‐to‐low spin crossover, and the structural characteristics of the high‐spin and low‐spin models are discussed in relation to other compounds showing a similar type of spin‐crossover behavior.
Exploring the Hydration of Pb2+: Ab Initio Studies and First‐Principles Molecular Dynamics
Authors: Gourlaouen Christophe , Gérard Hélène , Parisel Olivier
Published: April 2006

Even though lead is a well‐known toxicant widely scattered throughout the world since antiquity, its chemistry is poorly documented at the molecular level. Here we investigate the hydration of the Pb2+ ion by means of first‐principles molecular dynamics (Car–Parrinello molecular dynamics, CPMD). We found that the hydrated cation is heptacoordinated in a dynamically holodirected arrangement roughly corresponding to a fluxional distorted pentagonal bipyramid. The time‐averaged PbO bond length is especially large and amounts to 2.70 Å with an associated root‐mean‐square deviation of 0.26 Å. This results from a dynamic exchange between short (<2.6 Å), intermediate (2.6–3.0 Å) and long (>3.0 Å) PbO bonds. The latter very long PbO distance implies that the determination of the coordination number nc from experimental work may not necessarily yield values directly comparable to the theoretical value of nc=7, since not all experimental techniques would recognize such a long distance as a bond to the metal cation. Pronounced disorders are evidenced in the second shell, characteristic of a chaotropic cation, and exchanges between the first and second shells cannot be excluded on a timescale of a few tens of picoseconds.
A DNA‐Binding Copper(I) Metallosupramolecular Cylinder that Acts as an Artificial Nuclease
Authors: Childs Laura J. , Malina Jaroslav , Rolfsnes Britt Elin , Pascu Mirela , Prieto Maria J. , Broome Mark J. , Rodger P. Mark , Sletten Einar , Moreno Virtudes , Rodger Alison , Hannon Michael J.
Published: April 2006

The DNA binding of a dicationic pyridylimine‐based dicopper(I) metallosupramolecular cylinder is reported together with its ability to act as an artificial nuclease. The cylinder binds strongly to DNA; more strongly than the spherical dication [Ru(phen)3]2+(phen=1,10‐phenanthroline), but more weakly than the corresponding tetracationic cylinders. DNA coiling effects are not observed with this dication, in contrast to the situation with the previously reported tetracationic cylinder involving a similar ligand. Linear dichroism (LD) data suggests that the dicopper cylinder binds in a different orientation from that of the tetracationic iron cylinder. Furthermore, the dicopper cylinder shows DNA‐cleavage activity in the presence of peroxide. Of particular note is that the cylinder displays a marked and unusual ability to cleave both DNA strands at the same site, probably reflecting its dinuclear nature and possibly its mode of binding to the DNA.
Anticancer Cyclometalated [AuIIIm(C∧N∧C)mL]n+ Compounds: Synthesis and Cytotoxic Properties
Authors: Li Carrie Ka‐Lei , Sun Raymond Wai‐Yin , Kui Steven Chi‐Fai , Zhu Nianyong , Che Chi‐Ming
Published: April 2006

A series of cyclometalated gold(III) compounds [Aum(C∧N∧C)mL]n+ (m=1–3; n=0–3; HC∧N∧CH=2,6‐diphenylpyridine) was prepared by ligand substitution reaction of L with N‐donor or phosphine ligands. The [Aum(C∧N∧C)mL]n+ compounds are stable in solution in the presence of glutathione. Crystal structures of the gold(III) compounds containing bridging bi‐ and tridentate phosphino ligands reveal the presence of weak intramolecular π⋅⋅⋅π stacking between the [Au(C∧N∧C)]+ units. Results of MTT assays demonstrated that the [Aum(C∧N∧C)mL]n+ compounds containing nontoxic N‐donor auxiliary ligands (2) exert anticancer potency comparable to that of cisplatin, with IC50 values ranging from 1.5 to 84 μM. The use of [Au(C∧N∧C)(1‐methylimidazole)]+ (2 a) as a model compound revealed that the gold(III)‐induced cytotoxicity occurs through an apoptotic cell‐death pathway. The cell‐free interaction of 2 a with double‐stranded DNA was also examined. Absorption titration showed that 2 a binds to calf‐thymus DNA (ctDNA) with a binding constant of 4.5×105 dm3 mol−1 at 298 K. Evidence from gel‐mobility‐shift assays and viscosity measurements supports an intercalating binding mode for the 2 a–DNA interaction. Cell‐cycle analysis revealed that 2 a causes S‐phase cell arrest after incubation for 24 and 48 hours. The cytotoxicity of 3 b–g toward cancer cells (IC50=0.04–4.3 μM) correlates to that of the metal‐free phosphine ligands (IC50=0.1–38.0 μM), with [Au2(C∧N∧C)2(μ‐dppp)]2+ (3 d) and dppp (dppp=1,2‐bis(diphenylphosphino)propane) being the most cytotoxic gold(III) and metal‐free compounds, respectively. Compound 3 d shows a cytotoxicity at least ten‐fold higher than the other gold(III) analogues; in vitro cellular‐uptake experiments reveal similar absorptions for all the gold(III) compounds into nasopharyngeal carcinoma cells (SUNE1) (1.18–3.81 ng/cell; c.f., 3 d=2.04 ng/cell), suggesting the presence of non‐gold‐mediated cytotoxicity. Unlike 2 a, both gold(III) compounds [Au(C∧N∧C)(PPh3)]+ (3 a) (PPh3=triphenylphosphine) and [Au2(C∧N∧C)2(μ‐dppp)]2+ (3 d) interact only weakly with ctDNA and do not arrest the cell cycle.
Lewis Base Stabilized Phosphanylborane
Authors: Schwan Karl‐Christian , Timoskin Alexey Y. , Zabel Manfred , Scheer Manfred
Published: April 2006

The abstraction of the Lewis acid from [W(CO)5(PH2BH2⋅NMe3)] (1) by an excess of P(OMe3)3 leads to the quantitative formation of the first Lewis base stabilized monomeric parent compound of phosphanylborane [H2PBH2⋅NMe3] 2. Density functional theory (DFT) calculations have shown a low energetic difference between the crystallographically determined antiperiplanar arrangement of the lone pair and the trimethylamine group relative to the PB core and the synperiplanar conformation. Subsequent reactions with the main‐group Lewis acid BH3 as well as with an [Fe(CO)4] unit as a transition‐metal Lewis acid led to the formation of [(BH3)PH2BH2⋅NMe3] (3), containing a central H3BPH2BH2 unit, and [Fe(CO)4(PH2BH2⋅NMe3)] (4), respectively. In oxidation processes with O2, Me3NO, elemental sulfur, and selenium, the boranylphosphine chalcogenides [H2P(Q)BH2⋅NMe3] (Q = S 5 b; Se 5 c) as well as the novel boranyl phosphonic acid [(HO)2P(O)BH2⋅NMe3] (6 a) are formed. All products have been characterized by spectroscopic as well as by single‐crystal X‐ray structure analysis.
Neutral Helium Compounds: Theoretical Evidence for a Large Class of Polynuclear Complexes
Authors: Borocci Stefano , Bronzolino Nicoletta , Grandinetti Felice
Published: April 2006

Ab initio calculations at the MP2 and CCSD(T) levels of theory disclose the conceivable existence of neutral complexes containing up to four helium atoms. These species are formally obtained by replacing the hydrogen atoms of parent molecules such as CH4, SiH4, NH3, PH3, H2O, H2S, C2H2, C2H4, and C6H6with NBeHe moieties, which behave as monovalent functional groups containing helium. The geometries and vibrational frequencies of these M(NBeHe)n(n>1; M=central moiety) polyhelium complexes have been investigated at the MP2(full)/6‐31G(d) level of theory, and their stability with respect to the loss of helium atom(s) has been evaluated by means of single‐point calculations at the CCSD(T)/6‐311G(d,p) level of theory. Molecules such as HnC(NBeHe)4−nand HnSi(NBeHe)4−n(n=0–3), C2(NBeHe)2, andortho‐,meta‐, andpara‐C6H4(NBeHe)2were invariably characterized as energy minima, and were found to be stable with respect to the loss of helium atom(s) by approximately 4–5 kcal mol−1. On the other hand, species such as C2(NBeHe)4and C6(NBeHe)6were characterized as high‐order saddle points on the potential‐energy surface, and were unstable with respect to helium atom(s) loss owing to the bending motion of the NBeHe groups. The molecules containing N, P, O, or S as the central atom also showed a variable topology and include second‐order saddle points such as S(NBeHe)2, third‐order saddle points such as HN(NBeHe)2, but also minimum‐energy structures such as O(NBeHe)2and HP(NBeHe)2, which are also stable by approximately 5 kcal mol−1with respect to the helium atom(s) loss. These results suggest the conceivable existence of an, in principle, very large class of M(NBeHe)n(n>1) polyhelium complexes, whose stability may be substantially affected by the nature and the size of the central moiety M. Atoms‐in‐Molecules (AIM) calculations on selected species invariably suggest that, in our investigated M(NBeHe)n(n>1) compounds, the beryllium–helium interaction is essentially electrostatic.
Nonlinear recursive method to solid deformable structure dynamic problems
Authors: Hwang Yunn‐Lin
Published: April 2006

The recursive projection schemes used in most existing recursive methods for solid deformable structure dynamic problems lead to dense coefficient matrices in the acceleration equations and consequently there is a strong dynamic coupling between the joint and elastic coordinates. When the number of elastic degrees of freedom increases, the size of the coefficient matrix in the acceleration equations becomes large and consequently the use of these recursive methods for solving for the joint and elastic accelerations becomes less efficient. This paper discusses the problems associated with the recursive projection schemes used in the existing recursive methods, and it is shown that decoupling the joint and elastic accelerations using the nonlinear recursive method requires the factorization of nonlinear matrices whose dimensions are independent of the number of elastic degrees of freedom of the open‐loop and closed‐loop system. An amalgamated formulation that can be used to decouple the elastic and joint accelerations is then proposed. The use of the nonlinear recursive method developed in this paper is demonstrated using an open‐loop free‐falling deformable double pendulum system and a closed‐loop four‐bar mechanism. Copyright © 2006 John Wiley & Sons, Ltd.
Numerical determination of the solvent diffusion coefficient in a concentrated polymer solution
Authors: Kojić Nikola , Kojić Aleksandar , Kojić Milos̆
Published: April 2006

In chemical engineering applications involving solvent removal from a concentrated polymer solution where diffusion is the governing process (e.g. dry spinning of synthetic silk‐like fibres), the determination of the concentration‐dependent diffusion coefficient is essential. Diffusion of solvent through such a polymer solution can be described by Fick's law. We propose a numerical procedure to obtain the dependence of the diffusion coefficient on solvent concentration. The procedure employs the finite element method to model a pan weighing experiment, in which a small amount of the polymer solution is placed in a pan and allowed to evaporate into air. In the computational model, the changing depth of the polymer solution is taken into account and the varying diffusion coefficient is determined so as to match the experimentally recorded mass loss. Copyright © 2006 John Wiley & Sons, Ltd.
Flow cytometry analysis of gap junction‐mediated cell–cell communication: Advantages and pitfalls
Authors: Fonseca Paula Candida , Nihei Oscar Kenji , Savino Wilson , Spray David C. , Alves Luiz Anastacio
Published: April 2006

Background:

Since the first morphological description of the gap junctions use electron microscopy, a considerable number of techniques has been introduced to evaluate gap junction channel functionality, many of which use dye transfer techniques, such as dye injection and fluorescent dye transfer, analyzed by flow cytometry.

Methods:

To analyze dye transfer, generally one population of cells is incubated with calcein‐AM (0.5 μM) for 30 min at 37°C, and the other population was incubated with the lipophilic dye DiIC18 (3) (10 μM) for 1 h at 37°C; after incubation, these cells were washed five times with PBS and cocultured for different times, and then the dye transfer was analyzed by flow cytometry.

Results:

In this short overview, we focus on some advantages and disadvantages of flow cytometry as a technique to investigate gap junction‐mediated intercellular communication (GJIC). In addition, we point out some technical pitfalls that we have encountered when applying this technique to study gap junctions in immune system cells.

Conclusions:

Analysis of fluorescent dye transfer by flow cytometry is a useful tool to investigate GJIC. However, some points must be taken into consideration before using this methodology, which are discussed herein. © 2006 International Society for Analytical Cytology