bluuebunny/crossref_metadata_2025 huggingface dataset:
Long‐term stabilized blood samples as controls for flow cytometric HLA‐B27 screening: A feasibility study
Authors: Levering Wilfried H. B. M. , Wind Henk , Granger Vivian , Sintnicolaas Kees , Hooijkaas Herbert , Reilly John T. , Gratama Jan W. , Barnett David
Published: January 2008

Background:

Long‐term stabilized blood samples are potentially useful as positive or negative procedure controls for flow cytometric HLA‐B27 screening, and could serve as test samples in an external quality assessment (EQA) scheme. We evaluated long‐term stabilized whole blood specimens as prepared for the UK NEQAS for Leucocyte Immunophenotyping EQA scheme (Sheffield, UK).

Methods:

Peripheral blood samples were obtained from nine blood bank donors with known HLA‐B typing. Short‐term stabilization with Trans‐FIX™ was performed before shipment to Sheffield. Thereafter, long‐term stabilization was performed. Commercially available HLA‐B27 mAb were tested periodically between 1 week and 12 months on (i) fresh, (ii) short‐term stabilized, and (iii) long‐term stabilized blood samples using a stain, lyse, and wash technique. We compared the forward scatter (FSC), sideward scatter (SSC), and fluorescence signals of lymphocytes as a function of time. Furthermore, a pilot send‐out with stabilized blood samples of four blood bank donors was distributed among the participants to the Benelux EQA scheme for HLA‐B27 screening, and results were compared with historical EQA data obtained using nonstabilized blood samples from the same donors.

Results:

There were no major effects on FSC and SSC characteristics of lymphocytes. Background fluorescence of stabilized samples increased and specific fluorescence of stabilized HLA‐B27 positive samples decreased as compared with fresh samples. However, discrimination between the investigated HLA‐B27 positive and HLA‐B27 negative samples remained feasible poststabilization. In the pilot send‐out, the results obtained with stabilized samples were less concordant than with the corresponding fresh samples due to variable quality of the stabilized samples.

Conclusion:

Long‐term stabilized whole blood samples are potentially useful as true HLA‐B27 positive and true HLA‐B27 negative control cells for daily and longitudinal quality control of flow cytometric HLA‐B27 screening. In the same way, long‐term stabilized samples may be used for EQA purposes. However, these samples are currently not feasible for reagent validation purposes. Extensive quality control of long‐term stabilized samples is necessary before distribution in multicenter surveys. © 2008 Clinical Cytometry Society
A new case of acquired Glanzmann's thrombasthenia: Diagnostic value of flow cytometry
Authors: Giannini S. , Mezzasoma A. M. , Guglielmini G. , Rossi R. , Falcinelli E. , Gresele P.
Published: January 2008

Background:

Acquired Glanzmann's thrombasthenia (aGT) is a rare hemorrhagic disorder caused by autoantibodies, alloantibodies, or paraproteins directed against platelet GPIIb/IIIa. Its diagnosis requires several laboratory assays and mixing tests, which are complex and time consuming. We describe here a new case of aGT and compare different tests for the detection of GPIIb/IIIa‐blocking autoantibodies.

Methods:

A previously healthy 27‐year‐old male developed severe mucocutaneous bleeding, despite a normal platelet count, associated with non Hodgkin lymphoma.

Results:

Blood clotting tests were normal. Bleeding time and PFA‐100® were unmeasurable. Platelet aggregation was absent in response to all agonists except ristocetin. Platelet adhesion to collagen at high shear was impaired. Platelet granular content and release was normal. Flow cytometry showed normal binding of some anti‐GPIIb/IIIa antibodies (SZ21 and SAP), and decreased binding of others (P2, SZ22, A2A 9/6). Binding of PAC‐1, against activated GPIIb/IIIa, and of fibrinogen, was absent. In mixing tests, patient's serum inhibited aggregation, adhesion, and PAC‐1 and A2A9/6 binding to control platelets.

The patient's antibody, purified by affinity chromatography, recognized purified GPIIb by western blotting. Isolated patient's IgG inhibited platelet aggregation and A2A 9/6 binding by flow cytometry.

Conclusions:

Flow cytometry is especially useful for the diagnosis of aGT, being the only test able to characterize both the functional effect and the molecular target of the patient's autoantibody. © 2008 Clinical Cytometry Society
Determination of P‐glycoprotein, MDR‐related protein 1, breast cancer resistance protein, and lung‐resistance protein expression in leukemic stem cells of acute myeloid leukemia
Authors: de Figueiredo‐Pontes Lorena L. , Pintão Maria‐Carolina T. , Oliveira Luciana C. O. , Dalmazzo Leandro F. F. , Jácomo Rafael H. , Garcia Aglair B. , Falcão Roberto P. , Rego Eduardo M.
Published: January 2008

Background:

The most primitive leukemic precursor in acute myeloid leukemia (AML) is thought to be the leukemic stem cell (LSC), which retains the properties of self‐renewal and high proliferative capacity and quiescence of the hematopoietic stem cell. LSC seems to be immunophenotypically distinct and more resistant to chemotherapy than the more committed blasts. Considering that the multidrug resistance (MDR) constitutive expression may be a barrier to therapy in AML, we have investigated whether various MDR transporters were differentially expressed at the protein level by different leukemic subsets.

Methods:

The relative expression of the drug‐efflux pumps P‐gp, MRP, LRP, and BCRP was evaluated by mean fluorescence index (MFI) and the Kolmogorov‐Smirnov analysis (D values) in five leukemic subpopulations: CD34+CD38−CD123+ (LSCs), CD34+CD38+CD123−, CD34+CD38+CD123+, CD34+CD38+CD123−, and CD34− mature cells in 26 bone marrow samples of CD34+ AML cases.

Results:

The comparison between the two more immature subsets (LSC versus CD34+CD38−CD123− cells) revealed a higher P‐gp, MRP, and LRP expression in LSCs. The comparative analysis between LSCs and subsets of intermediate maturation (CD34+CD38+) demonstrated the higher BCRP expression in the LSCs. In addition, P‐gp expression was also significantly higher in the LSC compared to CD34+CD38+CD123− subpopulation. Finally, the comparative analysis between LSC and the most mature subset (CD34−) revealed higher MRP and LRP and lower P‐gp expression in the LSCs.

Conclusions:

Considering the cellular heterogeneity of AML, the higher MDR transporters expression at the most immature, self‐renewable, and quiescent LSC population reinforces that MDR is one of the mechanisms responsible for treatment failure. © 2008 Clinical Cytometry Society.
MHC‐restricted T cell receptor signaling is required for αβ TCR replacement of the pre T cell receptor
Authors: Croxford Andrew L. , Akilli‐Ozturk Özlem , Rieux‐Laucat Frederic , Förster Irmgard , Waisman Ari , Buch Thorsten
Published: January 2008

A developmental block is imposed on CD25+CD44– thymocytes at the β‐selection checkpoint in the absence of the pre T cell receptor (preTCR) α‐chain, pTα. Early surface expression of a transgenic αβ TCR has been shown to partially circumvent this block, such that thymocytes progress to the CD4+CD8+ double‐positive stage. We wanted to analyze whether a restricting MHC element is required for αβ TCR‐expressing double‐negative (DN) thymocytes to overcome the developmental block in pTα‐deficient animals. We used the HY‐I knock‐in model that endows thymocytes with αβ TCR expression in the DN compartment but has the advantage of physiological expression levels, in contrast to conventional TCR transgenes. On a pTα‐deficient background, this HY‐I TCR transgene ‘rescued’ CD25+CD44– thymocytes from apoptosis and enabled progression to later differentiation stages. On a non‐selecting MHC background, however, pTα‐deficient HY‐I mice presented a pronounced reduction in numbers of splenocytes and thymocytes when compared to animals of selecting MHC genotype, showing that MHC restriction is necessary to drive HY‐TCR‐mediated rescue of pTα‐deficient thymocytes.
Opposing regulation of T cell function by Egr‐1/NAB2 and Egr‐2/Egr‐3
Authors: Collins Sam , Lutz Michael A. , Zarek Paul E. , Anders Robert A. , Kersh Gilbert J. , Powell Jonathan D.
Published: January 2008

TCR‐induced NF‐AT activation leads to the up‐regulation of multiple genes involved in T cell anergy. Since NF‐AT is also involved in T cell activation, we have endeavored to dissect TCR‐induced activating and inhibitory genetic programs. This approach revealed roles for the early growth response (Egr) family of transcription factors and the Egr coactivator/corepressor NGFI‐A‐binding protein (NAB)2 in regulating T cell function. TCR‐induced Egr‐1 and NAB2 enhance T cell function, while Egr‐2 and Egr‐3 inhibit T cell function. In this report, we demonstrate that Egr‐2 and Egr‐3 are induced by NF‐AT in the absence of AP‐1, while Egr‐1 and NAB2 both require AP‐1‐mediated transcription. Our data suggest that Egr‐3 is upstream of Egr‐2, and that mechanistically Egr‐2 and Egr‐3 suppress Egr‐1 and NAB2 expression. Functionally, T cells from Egr‐2 and Egr‐3 null mice are hyperresponsive while T cells from Egr‐3 transgenic, overexpressing mice are hyporesponsive. Furthermore, an in vivo model of autoimmune pneumonitis reveals that T cells from Egr‐3 null mice hasten death while Egr‐3‐overexpressing T cells cause less disease. Overall, our data suggest that just as the Egr/NAB network of genes control cell fate in other systems, TCR‐induced Egr‐1, 2, 3 and NAB2 control the fate of antigen recognition in T cells.
Unexpected regulatory roles of TLR4 and TLR9 in experimental autoimmune encephalomyelitis
Authors: Marta Monica , Andersson Åsa , Isaksson Magnus , Kämpe Olle , Lobell Anna
Published: January 2008

Innate immune mechanisms essential for priming encephalitogenic T cells in autoimmune neuroinflammation are poorly understood. Experimental autoimmune encephalomyelitis (EAE) is a IL‐17‐producing Th (Th17) cell‐mediated autoimmune disease and an animal model of multiple sclerosis. To investigate how upstream TLR signals influence autoimmune T cell responses, we studied the role of individual TLR and MyD88, the common TLR adaptor molecule, in the initiation of innate and adaptive immune responses in EAE. Wild type (WT) C57BL/6, TLR‐deficient and MyD88‐deficient mice were immunized with myelin oligodendrocyte glycoprotein (MOG) in CFA. MyD88–/– mice were completely EAE resistant. Purified splenic myeloid DC (mDC) from MyD88–/– mice expressed much less IL‐6 and IL‐23, and serum and T cell IL‐17 were absent. TLR4–/– and TLR9–/– mice surprisingly exhibited more severe EAE symptoms than WT mice. IL‐6 and IL‐23 expression by mDC and Th17 responses were higher in TLR4–/– mice, suggesting a regulatory role of TLR4 in priming Th17 cells. IL‐6 expression by splenocytes was higher in TLR9–/– mice. Our data suggest that MyD88 mediates the induction of mDC IL‐6 and IL‐23 responses after MOG immunization, which in turn drives IL‐17‐producing encephalitogenic Th17 cell activation. Importantly, we demonstrate that TLR4 and TLR9 regulate disease severity in MOG‐induced EAE.
Mannose‐capped lipoarabinomannan‐ and prostaglandin E2‐dependent expansion of regulatory T cells in human Mycobacterium tuberculosis infection
Authors: Garg Ankita , Barnes Peter F. , Roy Sugata , Quiroga María F. , Wu Shiping , García Verónica E. , Krutzik Stephan R. , Weis Steven E. , Vankayalapati Ramakrishna
Published: January 2008

We evaluated the role of regulatory T cells (CD4+ CD25+ Foxp3+ cells, Tregs) in human Mycobacterium tuberculosis infection. Tregs were expanded in response to M. tuberculosis in healthy tuberculin reactors, but not in tuberculin‐negative individuals. The M. tuberculosis mannose‐capped lipoarabinomannan (ManLAM) resulted in regulatory T cell expansion, whereas the M. tuberculosis 19‐kDa protein and heat shock protein 65 had no effect. Anti‐IL‐10 and anti‐TGF‐β alone or in combination, did not reduce expansion of Tregs. In contrast, the cyclooxygenase enzyme‐2 inhibitor NS398 significantly inhibited expansion of Tregs, indicating that prostaglandin E2 (PGE2) contributes to Treg expansion. Monocytes produced PGE2 upon culturing with heat‐killed M. tuberculosis or ManLAM, and T cells from healthy tuberculin reactors enhanced PGE2 production by monocytes. Expanded Tregs produced significant amounts of TGF‐β and IL‐10 and depletion of Tregs from PBMC of these individuals increased the frequency of M. tuberculosis‐responsive CD4+ IFN‐γ cells. Culturing M. tuberculosis‐expanded Tregs with autologous CD8+ cells decreased the frequency of IFN‐γ+cells. Freshly isolated PBMC from tuberculosis patients had increased percentages of Tregs, compared to healthy tuberculin reactors. These findings demonstrate that Tregs expand in response to M. tuberculosis through mechanisms that depend on ManLAM and PGE2.
Delivery of antigen using a novel mannosylated dendrimer potentiates immunogenicity in vitro and in vivo
Authors: Sheng Kuo‐Ching , Kalkanidis Martha , Pouniotis Dodie S. , Esparon Sandra , Tang Choon Kit , Apostolopoulos Vasso , Pietersz Geoffrey A.
Published: January 2008

Antigen mannosylation has been shown to be an effective approach to potentiate antigen immunogenicity, due to the enhanced antigen uptake and presentation by APC. To overcome disadvantages associated with conventional methods used to mannosylate antigens, we have developed a novel mannose‐based antigen delivery system that utilizes a polyamidoamine (PAMAM) dendrimer. It is demonstrated that mannosylated dendrimer ovalbumin (MDO) is a potent immune inducer. With a strong binding avidity to DC, MDO potently induced OVA‐specific T cell response in vitro. It was found that the immunogenicity of MDO was due not only to enhanced antigen presentation, but also to induction of DC maturation. Mice immunized with MDO generated strong OVA‐specific CD4+/CD8+ T cell and antibody responses. MDO also targeted lymph node DC to cross‐present OVA, leading to OTI CD8+ T cell proliferation. Moreover, upon challenge with B16‐OVA tumor cells, tumors in mice pre‐immunized with MDO either did not grow or displayed a much more delayed onset, and had slower kinetics of growth than those of OVA‐immunized mice. This mannose‐based antigen delivery system was applied here for the first time to the immunization study. With several advantages and exceptional adjuvanticity, we propose mannosylated dendrimer as a potential vaccine carrier.
IL‐23‐mediated regulation of IL‐17 production in Helicobacter pylori‐infected gastric mucosa
Authors: Caruso Roberta , Fina Daniele , Paoluzi Omero Alessandro , Del Vecchio Blanco Giovanna , Stolfi Carmine , Rizzo Angelamaria , Caprioli Flavio , Sarra Massimiliano , Andrei Fabio , Fantini Massimo Claudio , MacDonald Thomas T. , Pallone Francesco , Monteleone Giovanni
Published: January 2008

Helicobacter pylori (Hp) infection is associated with a marked infiltration of the gastric mucosa by inflammatory cells. The molecular pathways that control Hp‐associated inflammatory reaction are complex, but locally induced cytokines seem to contribute to maintaining the ongoing inflammation. We have previously shown that IL‐17 is over‐produced in Hp‐infected gastric mucosa, and that IL‐17 stimulates the synthesis of IL‐8, the major neutrophil chemoattractant. Factors/mechanisms that regulate IL‐17 expression remain, however, unknown. In this study, we initially expanded our previous data, showing that CD4+ and CD8+ T cells are a source of IL‐17 in Hp‐infected samples. Since IL‐23 enhances T cell‐derived IL‐17 during bacterial infections, we then assessed the role of IL‐23 in controlling IL‐17 expression in Hp‐colonized stomach. Using real‐time PCR and ELISA, IL‐23 was detected in all gastric biopsies, but its expression was more pronounced in Hp‐infected samples in comparison to controls. Treatment of normal gastric lamina propria mononuclear cells (LPMC) with IL‐23 enhanced Stat3 activation and IL‐17 secretion, and pharmacological inhibition of Stat3 prevented IL‐23‐driven IL‐17 synthesis. Consistently, blockade of IL‐23 in cultures of LPMC from Hp‐infected patients reduced Stat3 activation and IL‐17 production. Data show that IL‐23 is overexpressed in Hp‐infected gastric mucosa where it could contribute to sustaining IL‐17 production.
Nippostrongylus brasiliensis infection leads to the development of emphysema associated with the induction of alternatively activated macrophages
Authors: Marsland Benjamin J. , Kurrer Michael , Reissmann Regina , Harris Nicola L. , Kopf Manfred
Published: January 2008

Chronic obstructive pulmonary disease (COPD) is the 5th most prevalent disease worldwide leading to severe morbidity and mortality in developed countries. The disease is strongly associated with smoking, and can be characterized by progressive and irreversible deterioration in lung function and destruction of the lung parenchyma. We show here that infection with the hookworm Nippostrongylus brasiliensis results in deterioration in lung function, destruction of alveoli and long‐term airways hyperresponsiveness, consistent with COPD and emphysema. N. brasiliensis infection leads to chronic low level hemorrhaging in the lung and the presence of hemosiderin‐laden macrophages in the absence of an overt inflammatory infiltrate. Microarray analysis of gene expression in diseased lungs and quantitative RT‐PCR analysis of purified macrophages revealed a state of prolonged tissue injury and the presence of alternatively activated macrophages producing MMP‐12. Taken together, these data show that lung tissue damage caused by hookworm infection can result in the development of COPD and emphysema.
Slamming the DOR on chemokine receptor signaling: Heterodimerization silences ligand‐occupied CXCR4 and δ‐opioid receptors
Authors: Hereld Dale , Jin Tian
Published: January 2008

Dimerization has emerged as a common mechanism for regulating the function of G protein‐coupled receptors (GPCR). Among these are chemokine receptors, which detect various chemokines and regulate a range of physiological process, including immune cell trafficking, cancer cell migration, and neuronal patterning. Homo‐ and heterodimerization in response to chemokine binding has been shown to be required for the initiation or alteration of signaling by a number of chemokine receptors. In this issue of the European Journal of Immunology, a new study indicates that the formation of heterodimers of chemokine receptor CXCR4 and the δ‐opioid receptor (DOR) prevents each of them from actively signaling, suggesting a novel mechanism for silencing GPCR function.

See accompanying article: http://dx.doi.org/10.1002/eji200737630
Benzylamines as Versatile Agents for the One‐Pot Synthesis and Highly Ordered Stacking of Anatase Nanoplatelets
Authors: Garnweitner Georg , Tsedev Ninjbadgar , Dierke Hanno , Niederberger Markus
Published: January 2008

The simple reaction of titanium isopropoxide in benzylamine is shown to lead to remarkably complex, highly ordered hybrid structures. These structures consist of anatase nanoplatelets that were stacked in a lamellar fashion with a small organic layer in between. By careful characterization of these structures, we show that indeed solely the benzylamine solvent is present in the organic moiety between the nanocrystals, which thereby provides both shape control and alignment of the inorganic crystals. The solvent also plays a central role during the anatase formation itself; hence, it enacts control on the forming materials on a multitude of levels.(© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2008)
Carbamates of Polyamines – Versatile Building Blocks for the Construction of Polynuclear Metal Complexes
Authors: Neis Christian , Weyhermüller Thomas , Bill Eckhard , Stucky Stefan , Hegetschweiler Kaspar
Published: January 2008

A new concept for the design of polynuclear metal complexes is described: Partial aeration of a polyamine solution with CO2 generates a system where both, a chelating agent (the unreacted polyamine) and a bridging agent (the polycarbamate) are present. Together with a metal cation, these components constitute an ideal set of building blocks for the construction of extended molecular architectures. These considerations are exemplified in the reaction of the well‐known triamine 6‐methyl‐1,4‐diazepan‐6‐amine (L) with FeII under aerobic conditions. After partial conversion of L into its dicarbamate derivative H–2L(CO2)22–, the components spontaneously assemble to the tetranuclear complex [(μ‐O)2(FeIIIL)4{μ‐H–2L(CO2)2}2]4+ (14+). 1(ClO4)4 was characterized by single‐crystal X‐ray analysis, Mössbauer spectroscopy, and magnetic susceptibility measurements.(© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2008)
Synthesis of Highly Substituted 2H‐Azirine‐2‐carboxylates via 3‐Azido‐4‐oxobut‐2‐enoates
Authors: Patonay Tamás , Jekő József , Juhász‐Tóth Éva
Published: January 2008

Phenacyl azides have been transformed into ethyl 4‐aryl‐3‐azido‐2‐methyl‐4‐oxobut‐2‐enoates by base‐induced coupling of the starting α‐azido ketone with ethyl pyruvate followed by elimination of methanesulfonic acid from the mesylate generated in situ from the labile aldol‐type intermediates ethyl 4‐aryl‐3‐azido‐2‐hydroxy‐2‐methyl‐4‐oxobutanoates. Thermolysis of ethyl 4‐aryl‐3‐azido‐2‐methyl‐4‐oxobut‐2‐enoates resulted in the formation of the hitherto unknown 3‐aroyl‐2‐ethoxycarbonyl‐2‐methyl‐2H‐azirines.(© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2008)
Application of Some Room Temperature Ionic Liquids in the Development of Biosensors at Carbon Film Electrodes
Authors: Pauliukaite Rasa , Doherty Andrew P. , Murnaghan Kevin D. , Brett Christopher M. A.
Published: January 2008

Two room temperature ionic liquids, 1‐butyl‐3‐methylimidazolium bistriflimide and 1‐butyl‐3‐methylimidazolium nitrate, were employed for enzyme immobilization in a new sol‐gel matrix and, for the first time, were successfully applied as electrolyte carriers in a biosensing system. The new sol‐gel matrix, based on 3‐aminopropyltrimethoxysilane and 1‐butyl‐3‐methylimidazolium bistriflimide mixtures, did not crack even after several weeks when kept dry, and exhibited similar analytical properties to aqueous sol‐gel based glucose biosensors. The linear range was up to 1.1 mM of glucose, sensitivity was 62 nA mM−1 and the limit of detection was 28.8 μM. The optimum ionic liquid electrolyte carrier was found to be 1‐butyl‐3‐methylimidazolium nitrate, where the biosensor was made by electrodeposition of the redox mediator, poly(neutral red), and the enzyme was immobilized by cross‐linking with glutaraldehyde. The results showed that application of room temperature ionic liquids to biosensors is very promising and can be further exploited.
A Renewable Potentiometric Immunosensor Based on Fe3O4 Nanoparticles Immobilized Anti‐IgG
Authors: Li Jianping , Gao Huiling
Published: January 2008

A renewable potentiometric immunosensor for detection of immunoglobulin G (IgG) has been developed by magnetic force attraction of Fe3O4 nanoparticles immobilized goat‐anti‐human IgG antibody. For preparing sensitive film of the sensor, cysteine was bonded on the nano‐Fe3O4 particles surface. The cysteine functionalized magnetic nanoparticles was attracted on a solid paraffin carbon paste electrode surface to covalently immobilize of anti‐immunoglobulin G (anti‐IgG) by employing a conventional glutaraldehyde‐crosslinking method. The immunosensor showed a specific response to human immunoglobulin G in the range of 0.1–1.2 ng/mL with a detection limit of 0.023 ng/mL. The immunosensor based on the magnetic nanoparticles was made easily by this method. It can be used expediently, renewed easily and low‐cost relatively. The renewable potentiometric immunosensor with better stability and higher sensitivity can be employed extensively in clinical diagnosis, monitoring of disease and environmental studies and etc.
Identification of deletion and duplication genotypes of thePMP22gene using PCR‐RFLP, competitive multiplex PCR, and multiplex ligation‐dependent probe amplification: A comparison
Authors: Hung Chia‐Cheng , Lee Chien‐Nan , Lin Chia‐Yun , Cheng Wen‐Fang , Chen Chi‐An , Hsieh Sung‐Tsang , Yang Chih‐Chao , Jong Yuh‐Jyh , Su Yi‐Ning , Lin Win‐Li
Published: January 2008

We evaluated the efficacy of PCR‐RFLP, competitive multiplex PCR, and a commercially available system of multiplex ligation‐dependent probe amplification (MLPA) for the determination of deletion and duplication genotypes of thePMP22gene. We compared the methods for efficiency, sensitivity, and specificity. We determined the gene dosage of thePMP22geneviaPCR‐RFLP, competitive multiplex PCR, and MLPA. To demonstrate the sensitivity and accuracy of these three methods, a total of 185 samples from 42 patients with hereditary neuropathy with liability to pressure palsies (HNPP), 57 patients with Charcot–Marie–Tooth disease type 1A (CMT1A), and 86 unaffected individuals, were analyzed. Molecular diagnosis by PCR‐RFLP was performed on all 185 samples; 24 HNPP deletions and 33 CMT1A duplications were identified. In contrast, 25 HNPP deletions and 38 CMT1A duplications were identified correctly using competitive multiplex PCR and MLPA. Six samples were incorrectly identified by PCR‐RFLP (one HNPP deletion and five CMT1A duplications). Competitive multiplex PCR and MLPA demonstrated reliability and relative speed compared to PCR‐RFLP; they were superior to PCR‐RFLP for gene dosage quantification. Multiplex PCR and MLPA should be the methods of choice for detection of deletion and duplication genotypes in molecular genetic diagnoses.
An ellipsoidal mirror for detection of laser‐induced fluorescence in capillary electrophoresis system: Applications for labelled antibody analysis
Authors: Rodat Audrey , Kalck Fabien , Poinsot Véréna , Feurer Bernard , Couderc François
Published: January 2008

An LIF detector was integrated into a CE system which uses a ball lens to focus the laser beam on the CE capillary. The detector employs an ellipsoid that is glued on the capillary window, to permit the collection of the fluorescence in the capillary. This ‘trapped’ fluorescence stays in the capillary because the angle of the silica/air interface is greater than the critical angle. The performance of this new detector setup is found to be identical to the collinear setup using the same ball lens. An application to the analysis of FITC‐labeled IgG was optimized using a 14 cm effective length capillary. The LOD of an FITC‐labeled IgG2 at an excitation wavelength of 488 nm was 150 pg/mL, which was 10 times better than the LOD recorded with slab gel silver staining. Using a tetramethylrhodamine (TAMRA)‐labeled IgG2 and a 532 nm excitation wavelength the LOD is 50 pg/mL. The electropherograms of four different commercial FITC conjugates of IgG were studied. The presence of aggregates was observed in two samples while close kinetics of reduction was observed between free aggregates and high aggregates concentration samples. The integrated LIF detector provides an extremely powerful and convenient tool for antibody analysis and should be useful for therapeutic MAb control in pharmaceutical facilities.
Removal of high‐abundance proteins for nuclear subproteome studies in rice (Oryza sativa) endosperm
Authors: Li Guosheng , Nallamilli Babi Ramesh Reddy , Tan Feng , Peng Zhaohua
Published: January 2008

Endosperm is a highly specialized storage organ with three sets of genomes. It is one of the most economically important organs in plants. Endosperm development involves parental imprinting and endoreduplication. A thorough study of the endosperm proteome, particularly the nuclear proteome, may provide critical insight into the regulation of seed development. Unfortunately, endosperm is extremely rich in starch grains and protein bodies of different sizes, making proteome studies on nonstorage proteins, particularly the low‐abundance proteins, very challenging. Here we have developed a chromatographic method to remove large starch grains and an electrophoresis method to recover low‐abundance proteins, respectively. Using these methods, we have identified 468 proteins from the nuclear enriched fraction of rice endosperm, including transcription factors, histone modification proteins, kinetochore proteins, centromere/microtubule binding proteins, and transposon proteins. Among the 468 proteins, 208 (44%) are hypothetical proteins, indicating that the endosperm proteome is poorly explored. In addition, analyses of the MS/MS data using BioWorks 3.1 have identified 59 putative acetylated proteins and 40 putative methylated proteins. Our studies have developed a method to remove starch grains and recover low‐abundance proteins, respectively. The methods should be applicable to other organisms.
Determination of stability constants of valinomycin complexes with ammonium and alkali metal ions by capillary affinity electrophoresis
Authors: Ehala Sille , Kašička Václav , Makrlík Emanuel
Published: January 2008

Capillary affinity electrophoresis (CAE) has been employed to investigate quantitatively the interactions of valinomycin, macrocyclic depsipeptide antibiotic ionophore, with univalent cations, ammonium and alkali metal ions, K+, Cs+, Na+, and Li+, in methanol. The study involved measuring the change in effective electrophoretic mobility of valinomycin while the cation concentrations in the BGE were increased. The corresponding apparent stability (binding) constants of the valinomycin–univalent cation complexes were obtained from the dependence of valinomycin effective mobility on the cation concentration in BGE using a nonlinear regression analysis. The calculated apparent stability constants of the above‐mentioned complexes show the substantially higher selectivity of valinomycin for K+ and Cs+ ions over Li+, Na+, and NH4+ ions. CAE proved to be a suitable method for the investigation of both weak and strong interactions of valinomycin with small ions.
Least‐squares finite element processes in h, p, k mathematical and computational framework for a non‐linear conservation law
Authors: Surana K. S. , Allu S. , Reddy J. N. , Tenpas P. W.
Published: January 2008

This paper considers numerical simulation of time‐dependent non‐linear partial differential equation resulting from a single non‐linear conservation law in h, p, k mathematical and computational framework in which k=(k1, k2) are the orders of the approximation spaces in space and time yielding global differentiability of orders (k1−1) and (k2−1) in space and time (hence k‐version of finite element method) using space–time marching process. Time‐dependent viscous Burgers equation is used as a specific model problem that has physical mechanism for viscous dissipation and its theoretical solutions are analytic. The inviscid form, on the other hand, assumes zero viscosity and as a consequence its solutions are non‐analytic as well as non‐unique (Russ. Math. Surv. 1962; 17(3):145–146; Russ. Math. Surv. 1960; 15(6):53–111). In references (Russ. Math. Surv. 1962; 17(3):145–146; Russ. Math. Surv. 1960; 15(6):53–111) authors demonstrated that the solutions of inviscid Burgers equations can only be approached within a limiting process in which viscosity approaches zero. Many approaches based on artificial viscosity have been published to accomplish this including more recent work on H(Div) least‐squares approach (Commun. Pure Appl. Math. 1965; 18:697–715) in which artificial viscosity is a function of spatial discretization, which diminishes with progressively refined discretizations. The thrust of the present work is to point out that: (1) viscous form of the Burgers equation already has the essential mechanism of viscosity (which is physical), (2) with progressively increasing Reynolds (Re) number (thereby progressively reduced viscosity) the solutions approach that of the inviscid form, (3) it is possible to compute numerical solutions for any Re number (finite) within hpk framework and space–time least‐squares processes, (4) the space–time residual functional converges monotonically and that it is possible to achieve the desired accuracy, (5) space–time, time marching processes utilizing a single space–time strip are computationally efficient. It is shown that viscous form of the Burgers equation without linearizing provides a physical and viablemechanism for approaching the solutions of inviscid form with progressively increasing Re. Numerical studies are presented and the computed solutions are compared with published work. Copyright © 2008 John Wiley & Sons, Ltd.
Buoyancy‐corrected k–ε models and large eddy simulation applied to a large axisymmetric helium plume
Authors: Chung W. , Devaud C. B.
Published: January 2008

The present numerical study is focused on testing two different modeling approaches to simulate a large turbulent buoyant helium plume, in particular the near‐field region. First, buoyancy‐corrected k–ε models are applied in Reynolds‐averaged Navier–Stokes (RANS) calculations, then large eddy simulation (LES) using a standard Smagorinsky model is examined. Good results are produced using the buoyancy‐corrected models, in particular, excellent agreement is achieved for the radial profiles of the streamwise velocity. However, the predictions are very sensitive to the choice of the buoyancy constant, C3ε, in the models. The present LES calculations show that the puffing frequency is accurately predicted. Predictions for the time‐averaged velocities are within experimental uncertainty at all locations. The predicted plume concentrations are in good agreement at the base of the plume, but the centerline values are overpredicted farther downstream. The higher‐order statistics are best predicted with the finest mesh. A sensitivity analysis on grid refinement, values of the Smagorinsky constant and the Schmidt number are included. Copyright © 2008 John Wiley & Sons, Ltd.
Detailed comparisons of front‐capturing methods for turbulent two‐phase flow simulations
Authors: Trontin Pierre , Vincent Stephane , Estivalezes Jean‐Luc , Caltagirone Jean‐Paul
Published: January 2008

In the framework of numerical study of multiphase flows, different front‐capturing methods are compared. Two different approaches dealing with the incompressibility constraint will be compared, too. Then, every technique is tested on several relevant test cases in order to make a comparison. The surface tension treatment is evaluated for each method and every front‐capturing method is tested on a case where interface deformations have something in common with turbulent behaviour. Finally, every method is studied through a phase inversion problem, which is an unsteady test case. Front‐capturing methods are tested on physical cases where Navier–Stokes and advection equations are coupled in a non‐linear way. Copyright © 2008 John Wiley & Sons, Ltd.
Accuracy of semiGLS stabilization of FEM for solving Navier–Stokes equations and a posteriori error estimates
Authors: Burda P. , Novotný J. , Šístek J.
Published: January 2008

Stabilization of the finite element method for flow problems at high Reynolds numbers is the main subject of presented research. The semiGLS method is recalled as a modification of the Galerkin least‐squares method. The presented work extends our previous paper on this method by its other important aspects. The main aim of this paper is to analyse and comment on the accuracy of the method. A posteriori error estimates for incompressible Navier–Stokes equations are used as the main tool for error analysis and some conclusions concerning accuracy are derived. Several numerical experiments are presented for both benchmark and practical problems. Copyright © 2008 John Wiley & Sons, Ltd.
Genome‐wide significance for dense SNP and resequencing data
Authors: Hoggart Clive J. , Clark Taane G. , De Iorio Maria , Whittaker John C. , Balding David J.
Published: January 2008

The problem of multiple testing is an important aspect of genome‐wide association studies, and will become more important as marker densities increase. The problem has been tackled with permutation and false discovery rate procedures and with Bayes factors, but each approach faces difficulties that we briefly review. In the current context of multiple studies on different genotyping platforms, we argue for the use of truly genome‐wide significance thresholds, based on all polymorphisms whether or not typed in the study. We approximate genome‐wide significance thresholds in contemporary West African, East Asian and European populations by simulating sequence data, based on all polymorphisms as well as for a range of single nucleotide polymorphism (SNP) selection criteria. Overall we find that significance thresholds vary by a factor of >20 over the SNP selection criteria and statistical tests that we consider and can be highly dependent on sample size. We compare our results for sequence data to those derived by the HapMap Consortium and find notable differences which may be due to the small sample sizes used in the HapMap estimate. Genet. Epidemiol. 32:179–185, 2008. © 2007 Wiley‐Liss, Inc.
Naturalistic outcomes of minor and subsyndromal depression in older primary care patients
Authors: Lyness Jeffrey M.
Published: January 2008

Objective

To review the literature regarding the naturalistic outcomes of minor and subsyndromal depression (‘Min/SSD’) in older primary care patients, synthesizing and critiquing findings and discussing avenues for future research.

Design

The author obtained relevant articles from repeated computer‐assisted literature searches over the past 15 years, and by reviewing the reference citations of the articles so obtained.

Results

A variety of relevant outcome domains were identified, as were important putative predictors, moderators, and mediators of outcome. In general, minor and subsyndromal depression each have comparable outcomes, outcomes that are clearly worse than non‐depressed subjects, with substantially elevated risk of worsening into major depression, albeit not as poor as those with major depression.

Conclusions

Min/SSD is common and of real clinical importance in primary care seniors. Several definitions of SSD may be used, each with overlapping but distinguishable utility in identifying patients. While the evidence base has expanded greatly in the past decade, considerable work remains to be done. Naturalistic studies of several outcome domains are needed, focusing on the predictive, moderating, and mediating roles of a wide range of psychopathological, medical, functional, and psychosocial factors. Such work will complement interventions and biomarker research approaches. Copyright © 2008 John Wiley & Sons, Ltd.
A new P‐heterocyclic family: A variety of six‐membered and bridged P‐heterocycles with 1‐benzyl substituent
Authors: Novák Tibor , Deme János , Ludányi Krisztina , Keglevich György
Published: January 2008

The ring enlargement of 1‐benzyl‐2,5‐dihydro‐1H‐phosphole oxide (1) via the corresponding 2‐phosphabicyclo[3.1.0]hexane 2‐oxide (2) afforded, depending on the conditions, the double bond isomers (A and B) of 1,2‐dihydrophosphinine oxide 4 or that of 3‐substituted 1,2,3,6‐tetrahydrophosphinine oxides 5 and 6. Dihydrophosphinine oxides (4) were suitable starting materials for 1,2,3,4,5,6‐hexahydrophosphinine oxide 7 and 1,2,3,6‐tetrahydrophosphinine oxide 8 obtained by reductive approaches and for the double bond isomers (A and B) of 2‐phosphabicyclo[2.2.2]octadiene 2‐oxide 9 and phosphabicyclooctene oxide 10 prepared in Diels–Alder cycloaddition. Precursor 9 was utilized in the fragmentation‐related phosphorylation of alcohols. © 2008 Wiley Periodicals, Inc. Heteroatom Chem 19:28–34, 2008; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/hc.20363
Synthesis and biological activities of novel bis‐heterocyclic pyrrodiazole derivatives
Authors: He Feng‐Qi , Liu Xing‐Hai , Wang Bao‐Lei , Li Zheng‐Ming
Published: January 2008

Novel structures of bis‐heterocyclic pyrrodiazole derivatives containing pyrazole were designed and synthesized. The title compounds were characterized by 1H NMR, IR, MS, and elemental analysis. Biological activities of three intermediate compounds and 25 pyrrodiazole derivatives were tested in vivo and in vitro. Some of the title compounds exhibited certain herbicidal activities against barnyardgrass and rape. © 2008 Wiley Periodicals, Inc. Heteroatom Chem 19:21–27, 2008; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/hc.20369
Synthesis, structure, and biological activity of novel 4,5‐disubstituted thiazolyl urea derivatives
Authors: Ling Shao , Xin Zhou , Zhong Jin , Jian‐xin Fang
Published: January 2008

Novel 1‐(2,4‐dichlorophenyl)‐3‐[4‐aryl‐5‐(1H‐1,2,4‐triazol‐1‐yl)thiazol‐2‐yl] urea derivatives were synthesized by the reaction of 2‐amino‐4‐sustituted phentyl‐5‐(1H‐1,2,4‐triazol‐1‐yl) thiazoles with 2,4‐dichloro‐1‐isocyanatobenzene. Structures of the title compounds were confirmed by the elemental analysis, 1H NMR, and single crystal X‐ray diffraction analysis. Biological evaluation showed that some of them possess promising antitumor activities. © 2008 Wiley Periodicals, Inc. Heteroatom Chem 19:2–6, 2008; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/hc.20375
LiClO4 catalyzed mild and efficient method for the synthesis of thiiranes from oxiranes
Authors: Reddy Sanjeeva , Nagavani S.
Published: January 2008

Oxiranes are efficiently converted to the corresponding thiiranes by potassium thiocyanate in the presence of catalytic amounts of LiClO4 with excellent yields under mild and nonaqueous conditions. © 2008 Wiley Periodicals, Inc. Heteroatom Chem 19:97–99, 2008; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/hc.20376
Synthesis and conformational analysis of a small meta‐cyclophane, bis(5‐carbomethoxy‐1,3‐phenylene)‐14‐crown‐4
Authors: Gibson Harry W. , Wang Hong , Chng Chin P. , Glass Thomas E. , Schoonover Daniel , Zakharov Lev N. , Rheingold Arnold L.
Published: January 2008

A small cyclophane, bis(5‐carbometh‐oxy‐1,3‐phenylene)‐14‐crown‐4 (BCMP14C4, 3) and its diacid, bis(5‐carboxy‐1,3‐phenylene)‐14‐crown‐4 (4), were synthesized and characterized. The solid‐state molecular structures of 3 and 4 were determined by X‐ray crystallography as ladder or stepped conformations in which the two aromatic rings are antiparallel to each other without overlap and the ethylene tethers both take trans‐conformations. Diester 3 is formed in the lowest cyclization yield (under the same reaction conditions) and exhibits the highest melting point compared to its larger ring (20‐, 26‐ and 32‐membered) analogs. In CD2Cl2 solution, diester 3 exists predominantly as a nonplanar gauche–gauche structure as deduced by H NMR studies. © 2008 Wiley Periodicals, Inc. Heteroatom Chem 19:48–54, 2008; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/hc.20393
Synthesis of phosphonodipeptide conjugates of ursolic acid and their homologs
Authors: Deng Sheng‐Lou , Baglin Isabelle , Nour Mohammed , Cavé Christian
Published: January 2008

To prepare novel derivatives of naturally bioactive 3β‐hydroxy‐urs‐12‐en‐28‐oic acid (ursolic acid) with unusual properties and broad spectrum of activities, a number of chemical reactions were conducted. First, a variety of α‐aminophosphonates were prepared by a series of reactions involving the three‐component Mannich type reaction as a key step. Second, an array of phosphonodipeptides and their homologs was synthesized through multistep reactions including condensation of phthalic anhydride with glycine or β‐alanine, chlorination of N‐blocked amino acids, coupling of acid chloride with α‐aminophosphonates and sequential hydrazinolysis. Finally, new classes of phosphonodipeptide conjugates of ursolic acid and their homologs were obtained by condensation of 3β‐acetoxy‐urs‐12‐en‐28‐oyl chloride with phosphonodipeptides and their homologs. © 2008 Wiley Periodicals, Inc. Heteroatom Chem 19:55–65, 2008; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/hc.20396
Bismuth triflate catalyzed prins‐type cyclization in ionic liquid: Synthesis of 4‐tetrahydropyranol derivatives
Authors: Murty M. S. R. , Rajasekhar K. , Harikrishna V. , Yadav J. S.
Published: January 2008

The reaction of aldehydes with homoallylic alcohols in the presence of catalytic amount of bismuth triflate in [bmim]PF6generates 4‐tetrahydropyranol derivatives in excellent yield and with high diastereoselectivity. © 2008 Wiley Periodicals, Inc. Heteroatom Chem 19:104–106, 2008; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/hc.20403
Diastereoselective addition of hypophosphorous acid to N‐(R)‐α‐methylbenzyl‐substituted schiff bases
Authors: Lewkowski Jarosław , Karpowicz Rafał , Rybarczyk Małgorzata
Published: January 2008

The addition of hypophosphorous acid to an azomethine bond of N‐(R)‐α‐methylbenzyl Schiff bases of a variety of aldehydes led to the formation of N‐(R)‐α‐methylbenzylamino‐(S)‐methanephosphonous acids in 100% diastereoselectivity. This fact allows us to suggest the probable mechanism of the Strecker‐like reaction between hypophosphorous acid, an aldehyde, and (R)‐α‐methylbenzylamine. © 2008 Wiley Periodicals, Inc. Heteroatom Chem 19:35–37, 2008; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/hc.20406
Synthesis of peptidyl phosphonates containing 5‐(4′‐carboxyphenyl)‐10,15,20‐tritolylporphyrin
Authors: Habdas Jan , Boduszek Bogdan
Published: January 2008

The synthesis and characterization of three new porphyrin‐peptidyl‐phosphonate derivatives, containing a diphenyl 3‐pyridylmethyl‐phosphonate moiety, is described. These compounds could serve as potential photosensitizers for the PDT method of tumor therapy and displayed activity as inhibitors of aminopeptidase N. © 2008 Wiley Periodicals, Inc. Heteroatom Chem 19:107–111, 2008; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/hc.20407
Comparison of the reactivity of N‐(p‐toluenesulfonyl)‐sulfinimidoyl fluorides and chlorides toward triphenylphosphine
Authors: Pashinnik Valeriy E. , Borovikov Alexei V. , Shermolovich Yuriy G.
Published: January 2008

The path of the reaction of aryl‐N‐(p‐toluenesulfonyl)‐sulfinimidoyl fluorides and triphenylphosphine is highly dependent on the order of the reactants addition. Addition of triphenylphos‐phine to aryl‐N‐(p‐toluenesulfonyl)‐sulfinimidoyl fluorides results in the formation of triphenyl(aryl‐thio)phosphonium salts of N,N′‐bis(p‐toluenesul‐fonyl)aryl‐sulfinamidines and triphenyldifluorophosphorane. By changing the reagent addition order, we obtained triphenyldifluorophosphorane, P,P,P‐triphenyl‐N‐(p‐toluenesulfonyl)‐phosphine imide, and diaryl disulfides. The outcome of the reaction aryl‐N‐(arenesulfonyl)‐sulfinimidoyl chlorides and triphenylphosphine does not depend on the order of addition of the reactants. P,P,P‐Triphenyl‐N‐(arenesulfonyl)‐phosphine imides, triphenyldichloro‐phosphorane, and diaryl disulfides were formed as a result. © 2008 Wiley Periodicals, Inc. Heteroatom Chem 19:66–71, 2008; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/hc.20408
Photochemical reaction of silyl‐substituted 1,4‐disila(dewar‐benzene) with isocyanide and phenylacetylene
Authors: Oikawa Toru , Nakata Norio , Matsumoto Takeshi , Kabe Yoshio , Sekiguchi Akira
Published: January 2008

The irradiation of silyl‐substituted 1,4‐disila(Dewar‐benzene) 1 with light of wavelength λ > 320 nm in the presence of 2,6‐dimethylphenyl‐isocyanide or phenylacetylene produced 1,4‐bis(di‐tert‐butylmethylsilyl)‐2,3,5,6‐tetraethyl‐7‐(2,6‐dime‐thylphenylimino)‐1,4‐disilabicyclo[2.2.1]hepta‐2,5‐diene 5 or 1,4‐bis(di‐tert‐butylmethylsilyl)‐2,3,5,6‐tetraethyl‐1‐(2‐phenylethynyl)‐1,4‐disilacyclohexa‐2,5‐diene 6, respectively. The molecular structures of 5 and 6 were determined by spectroscopic data and X‐ray crystallography. © 2008 Wiley Periodicals, Inc. Heteroatom Chem 19:87–92, 2008; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/hc.20411