bluuebunny/crossref_metadata_2025 huggingface dataset:
Confirmation of Mycobacterium tuberculosis infection by flow cytometry after ex vivo incubation of peripheral blood T cells with an ESAT‐6‐derived peptide pool
Authors: Tesfa Lydia , Koch Franz‐Wilhelm , Pankow Wulf , Volk Hans‐Dieter , Kern Florian
Published: June 2004

Background

The presence of a T‐cell response to the early secretory antigenic target‐6 (ESAT‐6) indicates previous infection with or exposure to Mycobacterium tuberculosis. Measuring this response is useful for identifying individuals infected with M. tuberculosis. It was also reported that the frequencies of ESAT‐6‐specific T cells correlate with disease state. Established procedures measure secreted T‐cell cytokines following whole blood stimulation with recombinant ESAT‐6 protein or use Elispot as a read‐out.

Methods

A single ESAT‐6‐ spanning pool of overlapping peptides (15 amino acids length with 11 overlaps) was used for overnight stimulation of peripheral blood mononuclear cells (PBMCs) from 15 patients infected with Mycobacterium tuberculosis and 11 healthy controls. T‐cell responses were rated positive if interferon‐γ (IFN‐γ)‐producing T cells were identified above background level, using 4‐color cytokine flow cytometry.

Results

Thirteen of 15 (87%) patients, but none of the healthy controls, had a positive CD4 T‐cell response to the ESAT‐6 spanning peptide pool. The frequencies of IFN‐γ‐producing cells varied between 1 and 167 per 10,000 CD4 T cells. The test performed as well as the tests described in the literature.

Conclusions

Cytokine flow cytometry following PBMC stimulation with an ESAT‐6 spanning peptide pool is a useful laboratory test for ESAT‐6‐specific T cells combining precise counting and multi‐parameter phenotyping. © 2004 Wiley‐Liss, Inc.
Tetraspan and beta‐1 integrins expression pattern of the epithelial lung adenocarcinoma cell line A549 and its sensitivity to divalent cations
Authors: Massin Frédéric , Rubinstein Eric , Faure Gilbert C. , Martinet Yves , Boucheix Claude , Béné Marie C.
Published: June 2004

Background

Tetraspans are ubiquitous integral transmembrane molecules associated on the cell surface with such adhesion molecules as integrins. Their expression has been shown to vary in tumors, but has seldom been described on lung tumoral epithelial cells, and the conditions required for a proper association of tetraspans and integrins have not yet been fully explored.

Methods

We investigated the expression of 10 tetraspans and six β1 integrins on the tumoral lung epithelial cell line A549. Cells were examined both in quantitative flow cytometry and as monolayers, under normal or chelated conditions, in order to determine the cation dependency of their expression.

Results

Five tetraspans and four β1 integrins are expressed on the membrane of A549 cells. Both quantitative and qualitative surface and cytoplasmic modifications of this pattern were induced in chelating conditions, suggesting the importance of divalent cations for the expression of these molecules.

Conclusions

These data indicate that a specific pattern of tetraspans and integrins, relying strongly on the availability of divalent cations in the microenvironment, is expressed by tumoral epithelial cells. © 2004 Wiley‐Liss, Inc.
MARCO expression on pediatric alveolar macrophages
Authors: Bunn Hazel J. , Kobzik Lester , Grigg Jonathan
Published: June 2004

Background

Phagocytosis of unopsonized pollutant particles by alveolar macrophages (AM) occurs via the macrophage receptor with collagenous structure (MARCO). In a previous study, we demonstrated that the laser scanning cytometer (LSC) can be used to characterize receptor expression in pediatric AM. Since we have demonstrated the presence of ultrafine carbon particles within AM obtained from young infants, we sought to determine MARCO expression by pediatric AM using the LSC, hypothesizing that MARCO expression, and hence the ability to phagocytize pollutant particles, is mature in early infancy.

Methods

AM from 17 healthy children undergoing elective surgery, between 0.1 and 14.1 years of age, were obtained by bronchoalveolar lavage. Cytocentrifuged AM were dual immunostained with monoclonal antibodies to the panmacrophage marker CD68, and either MARCO or an appropriate isotypic control. Expression of MARCO was quantified using LSC and adjusted for cell area. Intensity of immunostaining was compared to each child's own isotypic control.

Results

MARCO was present on 98% of pediatric AM. There was no association between age and either intensity of MARCO expression or the proportion of cells expressing MARCO.

Conclusions

We conclude that there is no developmental immaturity in the major receptor responsible for unopsonized particle uptake in healthy young children. © 2004 Wiley‐Liss, Inc.
Cytometric bead assay of cytokines in sepsis: A clinical evaluation
Authors: Vedrine Christophe , Caraion Cristina , Lambert Claude , Genin Christian
Published: June 2004

Background

The clinical relevancy of an attractive new multiparametric method, cytometric beads assay (CBA), was evaluated for the monitoring of cytokines in sepsis.

Methods

A total of 52 samples (26 patients) were simultaneously tested by CBA and chemiluminescence (IL‐6, IL‐8, IL‐1β, and TNFα) or ELISA (IL‐10, IL‐12 p40, IL‐12 p70, and soluble TNFα‐RI).

Results

CBA standard curves were linear from 20–5,000 ng/L except for IL‐1β (40–5,000 ng/L). IL‐6 and IL‐8 were detected in 41 and 48 samples (44–5,000 ng/L and 22–5,000 ng/L), respectively, and out of range in six samples. IL‐10 and IL‐1β were detected in 14 and 15 samples (21–1,548 ng/L and 29–582 ng/L), respectively. TNFα was rarely detected, and IL‐12 p70 was never detected. Accuracy and repeatability were good (CV for IL‐6 was 2.1–8.7%; for IL‐8 3.7–5.3%; for IL‐10 2.5–9.1%; for IL‐12 5.2–6.5%; for IL‐1 β 4–12%; and for TNFα 3–19.3%). Reproducibility of four standard curves and 20 samples (−1.1 to +1.03%) was excellent between tests done at two‐ to six‐week intervals. CBA values were correlated (r2 > 0.89; P < 0.001) with our reference methods, but were lower on CBA (TNFα 45% ± 8; IL‐6 49% ± 7), probably due to interactions with patients' serum, as confirmed by spiking diluted standards in one patient's serum with end stage renal failure and high levels of TNF soluble receptor (i.e., TNFα levels 34 and 21%).

Conclusions

Finally, these results suggest that IL‐6, IL‐8, IL‐1β, and Il‐10 are clinically promising for sepsis evaluation. However, sensitivity of TNFα has to be improved and IL‐12 p70 should be replaced with more relevant parameters such as TNF‐R, procalcitonin, or neopterin. © 2004 Wiley‐Liss, Inc.
Flow cytometric test for beryllium sensitivity
Authors: Milovanova Tatyana N. , Popma Sicco H. , Cherian Sindhu , Moore Jonni S. , Rossman Milton D.
Published: June 2004

Background

Chronic beryllium disease (CBD) is an occupational granulomatous disorder characterized by hypersensitivity to beryllium, mediated by CD4+ T lymphocytes, and predominantly affects the lungs. In this disorder, lymphocyte proliferative responses to beryllium, measured by 3H thymidine incorporation, are used for diagnosis of CBD, for screening asymptomatic workers or former workers to detect unrecognized disease, and for surveillance as a bioassay to detect abnormal exposures. Problems with test variability and the use of radioactivity have recently led to the search for alternative methods.

Methods

We applied a 5,6‐carboxyfluorescein diacetate succinimidyl ester flow cytometric technique for measurement of mitogen‐ and antigen‐induced T‐lymphocyte proliferation to a group of beryllium‐exposed sensitized individuals and beryllium‐unexposed controls.

Results

We detected mitogen and antigen proliferative responses in CD3+, CD4+, and CD8+ subpopulations. Phytohemagglutinin and Candida stimulated CD4+ and CD8+ T‐cell responses, but beryllium appeared to stimulate only CD3+/CD4+ responses.

Conclusions

This technique may provide a sensitive, nonradioactive alternative to the traditional proliferation tests that measure beryllium sensitivity. It offers the added specificity of enabling phenotypic description of the responding cell type and may prove to be easier to standardize for clinical use. © 2004 Wiley‐Liss, Inc.
Extra copies of c‐myc are more pronounced in nodular melanomas than in superficial spreading melanomas as revealed by fluorescence in situ hybridisation
Authors: Treszl Andrea , Ádány Róza , Rákosy Zsuzsa , Kardos László , Bégány Ágnes , Gilde Katalin , Balázs Margit
Published: June 2004

Background

Amplification of c‐myc is a common genetic alteration and associated with a poor prognosis in a variety of cancers. Extra copies of the gene have been found in large numbers of melanoma metastases, but only few primary tumours have been studied. We investigated the c‐myc copy number alterations in two different subtypes of primary melanomas with different biological behaviours.

Methods

Fluorescence in situ hybridisation was performed using c‐myc and centromeric 8 (C8) probes on 68 lesions (28 nodular melanomas [NMs], 26 superficial spreading melanomas [SSMs], and 14 metastases). To assess the ploidy pattern, copy number distribution of seven different chromosomes was also investigated.

Results

All tumours showed aneuploid populations for at least three chromosomes. Whereas 61% of the NMs exhibited extra c‐myc copies, only 27% of SSMs showed increased gene dosage. The c‐myc/C8 ratio exceeding 1.5 was significantly higher in NMs (P = 0.01). High level amplification was seen only in NMs. An elevated c‐myc/C8 ratio was higher than 1.5 in only four metastases.

Conclusion

Our data show that c‐myc copy number alterations differ in the two melanoma subtypes and are associated with the advanced stage of the disease. The less frequent amplification of the c‐myc gene in metastatic lesions indicates that it may play an important role in the development of an invasive potential rather than in the metastatic process. © 2004 Wiley‐Liss, Inc.
CD4–c‐kit–CD3ϵ–IL‐2Rα+ Peyer's patch cells are a novel cell subset which secrete IL‐5 in response to IL‐2: implications for their role in IgA production
Authors: Kuraoka Masayuki , Hashiguchi Masaaki , Hachimura Satoshi , Kaminogawa Shuichi
Published: June 2004

In this study, we examined which cell population contributes to IL‐5 production by Peyer's patch (PP) cells. Thy1.2– fraction of PP cells, but not those of splenocytes, secreted IL‐5 in response to IL‐2. We found that CD3ϵ–IL‐2Rα+ cells purified from the Thy1.2–B220– fraction of PP cells secreted IL‐5 when stimulated with IL‐2. CD3ϵ–IL‐2Rα+ cells were subdivided into CD4+ and CD4– populations or c‐kit+ and c‐kit– populations, and only the CD4– and c‐kit– CD3ϵ–IL‐2Rα+ cells secreted IL‐5 in response to IL‐2. CD3ϵ–IL‐2Rα+ cells did not express NK cell‐markers and exhibited a lymphoid morphology. We have therefore identified CD3ϵ–IL‐2Rα+ cells as a unique lymphoid population that are not classified into conventional IL‐5‐producing cell populations, such as T cells, mast cells and NK cells. Depletion of CD3ϵ–IL‐2Rα+ cells from PP resulted in reduced IL‐5 production. Furthermore, IgA secretion by B cells was increased when PP B cells were cocultured with CD3ϵ–IL‐2Rα+ cells. Taken together, these results suggest that the novel subset of CD4–c‐kit–CD3ϵ–IL‐2Rα+ PP cells are capable of secreting a high level of IL‐5 in response to IL‐2, contribute markedly to IL‐5 production and help IgA secretion by B cells.
Apoptosis via the B cell antigen receptor requires Bax translocation and involves mitochondrial depolarization, cytochrome C release, and caspase‐9 activation
Authors: Eldering Eric , Mackus Wendelina J. M. , Derks Ingrid A. M. , Evers Ludo M. , Beuling Esther , Teeling Peter , Lens Susanne M. A. , van Oers Marinus H. J. , van Lier René A. W.
Published: June 2004

Various routes to apoptosis can be active during B cell development. In a model system of mature B cells, differences in caspase‐3 processing have suggested that antigen receptor (BCR)‐mediated apoptosis may involve a zVAD‐insensitive initiator protease(s). In search of the events leading to caspase‐3 activation, we now establish that both CD95‐ and BCR‐mediated apoptosis depend on Bax activation and cytochrome C (cytC) release. Nevertheless, the timing and caspase‐dependence of mitochondrial membrane depolarization differed considerably after CD95‐ or BCR‐triggering. To delineate events subsequent to cytC release, we compared apoptosis induced via BCR triggering and via direct mitochondrial depolarization by CCCP. In both cases, partial processing of caspase‐3 was observed in the presence of zVAD. By expression in 293 cells we addressed the potential of candidate initiator caspases to function in the presence of zVAD, and found that caspase‐9 efficiently processed caspase‐3, while caspase‐2 or –8 were inactive. Finally, retroviral expression of dominant‐negative caspase‐9 inhibited both CD95‐ and BCR‐mediated apoptosis. In conclusion, we obtained no evidence for involvement of a BCR‐specific protease. Instead, our data show for the first time that the BCR‐signal causes Bax translocation, followed by mitochondrial depolarization, and cytC release. Subsequent caspase‐9 activation can solely account for events further downstream.
The effects of endothelin‐1 on degranulation, cytokine, and growth factor production by skin‐derived mast cells
Authors: Matsushima Hironori , Yamada Nobuo , Matsue Hiroyuki , Shimada Shinji
Published: June 2004

Endothelin‐1 (ET‐1), originally described as a vasoconstrictor, is now known to be involved in pathogenesis of various disorders including vascular, inflammatory, and fibrotic diseases. Recentstudies suggest that mast cells are also involved in the same pathological conditions. In this study, we tested a hypothesis that ET‐1 would affect mast cell functions and contribute to such disease conditions, using fetal skin‐derived cultured mast cells (FSMC) and bone marrow‐derived cultured mast cells (BMMC). FSMC expressed ET receptors (ETA and ETB) at mRNA and protein levels, whereas BMMC expressed lower levels of ETA, and little, if any, ETB. ET‐1 induced degranulation by FSMC, but not by BMMC through ETA‐mediated pathways. ET‐1 at different concentrations exerted the reciprocal effects on degranulation by IgE‐bound FSMC. Furthermore, ET‐1 induced TNF‐α and IL‐6 production by FSMC, but not by BMMC, and significantly enhanced VEGF production and TGF‐β1 mRNA expression by FSMC. Finally, ET‐1 was produced by FSMC, but not by BMMC in response to Toll‐like receptor ligands. These results indicate contrasting impacts of ET‐1 on distinct mast cell populations. We propose that ET‐1 may participate in pathological conditions of various disorders via its multi‐functional effects on mast cells under certain conditions.
HSP60 induces self‐tolerance to repeated HSP60 stimulation and cross‐tolerance to other pro‐inflammatory stimuli
Authors: Kilmartin Barry , Reen Denis J.
Published: June 2004

In addition to their primary function as intracellular chaperone proteins, the immunomodulatory properties of heat shock proteins (HSP), including their role as adjuvants for vaccines, have become a focus of intense research interest. Interestingly, the effect of chronic exposure to an endogenous immunomodulator and initiator of inflammation such as autologous HSP60 has as yet remained uncharacterized. In this study, we demonstrate that pretreatment of monocytes with human HSP60 results in a suppression of TNF‐α production on restimulation with HSP60. Furthermore, desensitization with HSP60 inhibits TNF‐α expression in these cells in response to LPS stimulation, thereby inducing "cross‐tolerance". In contrast to TNF‐α suppression, IL‐1β expression was augmented in HSP60‐pretreated monocytes on restimulation, while being suppressed in THP‐1 cells. Addition of an anti‐IL‐10 neutralizing antibody had no significant effect on HSP60‐ or LPS‐induced tolerance.HSP60 priming of monocytes also results in significant down‐regulation of HLA‐DR, CD86 and Toll‐like receptor 4 expression, but minimally up‐regulates CD80 expression, similar to that previously reported with LPS. By identifying a previously unrecognized "tolerizing" effect of extended exposure to autologous HSP60 on the innate immune system, as opposed to its recently identified pro‐inflammatory stimulatory capacity, this study highlights a further level of complexity of our understanding of the biological activities of HSP.
Effect of Substitution of Methyl Groups on the Luminescence Performance of IrIII Complexes: Preparation, Structures, Electrochemistry, Photophysical Properties and Their Applications in Organic Light‐Emitting Diodes (OLEDs)
Authors: Jung Sungouk , Kang Youngjin , Kim Hyung‐Sun , Kim Yun‐Hi , Lee Chang‐Lyoul , Kim Jang‐Joo , Lee Sung‐Koo , Kwon Soon‐Ki
Published: June 2004

A series of dimethyl‐substituted tris(pyridylphenyl)iridium(III) derivatives [(n‐MePy−n′‐MePh)3Ir] [n = 3, n′ = 4 (1); n = 4, n′ = 4 (2); n = 4, n′ = 5 (3); n = 5, n′ = 4 (4); n = 5, n′ = 5 (5)] have been synthesized and characterized to investigate the effect of the substitution of methyl groups on the solid‐state structure and photo‐ and electroluminescence. The absorption, emission, cyclic voltammetry and electroluminescent performance of 1−5 have also been systematically evaluated. The structures of 2 and 4 have been determined by a single‐crystal X‐ray diffraction analysis. Under reflux (> 200 °C) in glycerol solution, fac‐type complexes with a distorted octahedral geometry are predominantly formed as the major components in all cases. Electrochemical studies showed much smaller oxidation potentials relative to Ir(ppy)3 (Hppy = 2‐phenylpyridine). All complexes exhibit intense green photoluminescence (PL), which has been attributed to metal‐to‐ligand charge transfer (MLCT) triplet emission. The maximum emission wavelengths of thin films of 1, 3, 4 and 5 at room temperature are in the range 529−536 nm, while 2 displays a blue‐shifted emission band (λmax = 512 nm) with a higher PL quantum efficiency (ΦPL = 0.52) than those of complexes 1 and 3−5; this is attributed to a decrease of the intermolecular interactions. Multilayered organic light‐emitting diodes (OLEDs) were fabricated by using three (2, 3 and 4) of these IrIII derivatives as dopant materials. The electroluminescence (EL) spectra of the devices, which have the maximum peaks at 509−522 nm, with shoulder peaks near 552 nm, are consistent with the PL spectra in solution at 298 K. The devices show operating voltages at 1 mA/cm2 of 4.9, 5.6, 5.1, and 4.6 V for Ir(ppy)3, 2, 3, and 4, respectively. In particular, the device with 2 shows a higher external quantum efficiency (ηext = 11% at 1 mA/cm2) and brightness (4543 cd/m2 at 20 mA/cm2) than Ir(ppy)3 (ηext = 6.0% at 1 mA/cm2; 3156 cd/m2 at 20 mA/cm2) and other Ir(dmppy)3 derivatives, (dmppy = dimethyl‐substituted ppy), under the same conditions. The methyl groups at the meta (Ph) and para (Py) positions to the Ir metal atom have a great influence on absorption, emission, redox potentials and electroluminescence. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2004)
Solution Synthesis of Two Orthogonally Protected Lactosides as Tetravalent Disaccharide‐Based Scaffolds
Authors: Castoldi Sergio , Cravini Massimiliano , Micheli Fabrizio , Piga Elisabetta , Russo Giovanni , Seneci Pierfausto , Lay Luigi
Published: June 2004

Two tetravalent lactosidic scaffolds have been synthesised in solution from commercial lactose. Careful manipulation of the protecting groups allowed us to orthogonally protect four OH groups for their use as diversity sites for the development of broad screening libraries of sugar mimics. The selective access to each of these hydroxy groups has been demonstrated on scaffold 2 by deprotection and functionalisation with p‐fluorophenyl isocyanate. Finally, the 6‐OH derivative of compound 2 was covalently attached to a polymeric support. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2004)
Reduction of Carbonyl Compounds by Lanthanide Metal/2‐Propanol: In‐situ Generation of Samarium Isopropyloxide for Stereoselective Meerwein−Ponndorf−Verley Reduction
Authors: Fukuzawa Shin‐ichi , Nakano Narihito , Saitoh Takahide
Published: June 2004

The reduction of ketones and aldehydes with lanthanide metals (La, Ce, Sm, Yb) and a catalytic amount of iodine (5 mol %) in iPrOH proceeded smoothly to produce the corresponding alcohols as the major products in good yield, while in THF, methanol, and ethanol the pinacols were mainly produced. The yields of alcohols were improved most effectively by the use of Sm metal, the amount of pinacol produced thus being minimized. The actual reducing agent may be samarium isopropyloxide, which mediates the Meerwein−Ponndorf−Verley‐type hydride‐transfer reaction, since the reaction can be carried out catalytically with respect to samarium. The stereoselectivities of the reductions of the 2‐ and 4‐substituted cyclohexanones with Sm/iPrOH were higher than those achieved with SmI2/iPrOH or Sm/H2O. The asymmetric reduction of acetophenone could be achieved to give the 1‐phenylethanol in up to 95% ee in the presence of the chiral ligand 7. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2004)
Synthesis of Free and Nα‐Fmoc‐/Nγ‐Boc‐Protected (2S,4S)‐ and (2S,4R)‐4‐Aminopipecolic Acids
Authors: Machetti Fabrizio , Cordero Franca M. , De Sarlo Francesco , Papini Anna M. , Alcaro Maria C. , Brandi Alberto
Published: June 2004

The preparation of (2S,4S)‐ and (2S,4R)‐4‐aminopipecolic acid, a conformationally constrained basic amino acid bearing orthogonal Nα/Nγ‐protection suitable for solid‐phase peptide synthesis, is reported. These amino acids were synthesised from enantiopure ethyl (2S)‐4‐oxo‐1‐(1‐phenylethyl)piperidine‐2‐carboxylate (1) with introduction of the side‐chain amino group by reductive amination, followed by protection/deprotection of the functional groups. A mixture of Fmoc‐Sly(Boc)‐OH 6 and 7 was used to establish their compatibility in solid‐phase peptide synthesis. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2004)
Non‐Dipolar Behavior of Mesoionic Heterocycles: Synthesis and Tautomerism of 2‐Alkylthioisomünchnones
Authors: Ávalos Martín , Babiano Reyes , Cintas Pedro , Díaz Jesús , Jiménez José L. , López Ignacio , Palacios Juan C.
Published: June 2004

This paper describes a general preparation of a series of 1,3‐thiazolium‐4‐olates, each bearing an alkyl group at C‐2, through reactions between N‐arylthiocarboxamides and α‐haloacyl halides. Unlike the 2‐aryl‐substituted derivatives, such alkylated mesoionic compounds exist in equilibria with their non‐dipolar tautomers, the corresponding 2‐alkylidene‐1,3‐thiazolidin‐4‐ones. The unambiguous characterization of such tautomers and their relative stabilities have now been assessed by spectroscopic and computational studies. The presence of o,o′‐disubstituted aryl groups at N‐3 of the heterocyclic ring slows down free rotation around the N−Ar bond, thus opening access to a promising class of non‐biaryl atropisomers. Finally, treatment of N‐arylthioformamides with α‐haloacyl halides gives rise to N‐acylthioformamides instead of the corresponding mesoionic species. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2004)
1,8‐Diazabicyclo[6.6.6]eicosane, 1,8‐Diazabicyclo[6.6.5]nonadecane and 1,8‐Diazabicyclo[6.6.4]octadecane and Their Diprotonated Forms
Authors: Pool Brett , Balalaie Saeed , Kunze Andreas , Schilling Gerhard , Bischof Peter , Gleiter Rolf
Published: June 2004

The preparation of the title compounds 9−11 was achieved by catalytic hydrogenation of 1,8‐diazabicyclo[6.6.6]eicosa‐4,11‐diyne (13), 1,8‐diazabicyclo[6.6.5]nonadeca‐4,11‐diyne (14), and 1,8‐diazabicyclo[6.6.4]octadeca‐4,11‐diyne (15), respectively. As catalyst we used Pd(OH)2 on carbon. NMR studies on 9−11 revealed an in/in conformation at the bridgehead positions. Treatment of 9−11 with an excess of trifluoroacetic acid yielded, in a kinetically controlled reaction, mixtures of the out/out and in/out diprotonated species [9·2H]2+, [10·2H]2+ and [11·2H]2+. The thermodynamically controlled species, the corresponding in/in conformers, are formed at higher temperatures and longer reaction times. The experimental observations are supported by AM1 calculations. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2004)
Highly Enantioselective Synthesis of No‐Carrier‐Added 6‐[18F]Fluoro‐L‐dopa by Chiral Phase‐Transfer Alkylation
Authors: Lemaire Christian , Gillet Steve , Guillouet Stéphane , Plenevaux Alain , Aerts Joël , Luxen André
Published: June 2004

[18F]Fluoro‐L‐dopa, an important radiopharmaceutical for positron emission tomography (PET), has been synthesized using a phase‐transfer alkylation reaction. A chiral quaternary ammonium salt derived from a Cinchona alkaloid served as phase‐transfer catalyst for the enantioselective alkylation of a glycine derivative. The active methylene group of this Schiff‐base substrate was deprotonated with cesium hydroxide and rapidly alkylated by the 2‐[18F]fluoro‐4,5‐dimethoxybenzyl halide (X = Br, I). The reaction proceeded with high yield (> 90%) at 0 °C or room temperature in various solvents such as toluene or dichloromethane. Preparation of the [18F]alkylating agent on a solid support was developed. After labelling, the labeled [18F]fluoroveratraldehyde was trapped on a tC18 cartridge and then converted on the cartridge into the corresponding benzyl halide derivatives by addition of aqueous sodium borohydride and gaseous hydrobromic or ‐iodic acid. Hydrolysis and purification by preparative HPLC made 6‐[18F]fluoro‐L‐dopa ready for human injection in a 25−30% decay‐corrected radiochemical yield in a synthesis time of 100 min. The product was found to be chemically, radiochemically and enantiomerically pure (ee > 95%). (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2004)
Telomerization of Butadiene withL‐Arabinose andD‐Xylose in DMF: Selective Formation of their Monooctadienyl Glycosides
Authors: Estrine Boris , Bouquillon Sandrine , Hénin Françoise , Muzart Jacques
Published: June 2004

Conditions to achieve the palladium‐catalysed telomerization of butadiene withL‐arabinose andD‐xylose as telogens have been identified. With DMF as solvent, optimised ratios of substrate, reactants and catalytic system allowed the selective grafting of one octadienyl chain onto the anomeric hydroxy group. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2004)
Transannular Reactions of Two Non‐Parallel 1,3‐Butadiyne Units − Syntheses, Structures and Protonation Reactions of 1‐Isopropyl‐1‐azacyclopentadeca‐3,5,11,13‐tetrayne and 1‐Isopropyl‐1‐azacyclohexadeca‐3,5,12,14‐tetrayne
Authors: Schmidt Erik M. , Gleiter Rolf , Rominger Frank
Published: June 2004

The syntheses of 1‐isopropyl‐1‐azacyclopentadeca‐3,5,11,13‐tetrayne (15a) and 1‐isopropyl‐1‐azacyclohexadeca‐3,5,12,14‐tetrayne (15b) were accomplished in a stepwise approach. The key intermediates were 1,14‐dibromotetradeca‐2,4,10,12‐tetrayne (14a) and 1,15‐dibromopentadeca‐2,4,11,13‐tetrayne (14b). The ring closure to 15a and 15b was achieved by reaction with isopropylamine. X‐ray investigations on single crystals of 15a and 15b revealed a non‐parallel orientation of the 1,3‐butadiyne units. The reaction of 15b with concd. HCl in ethanol yielded 5,12‐dichloro‐2‐isopropyl‐1,2,3,6,7,8,9,10‐octahydrocyclonona[e]isoindole (16c) and 5‐chloro‐2‐isopropyl‐2,3,6,7,8,9,10,11‐octahydrocyclonona[e]isoindol‐12(1H)‐one (17c). A mechanism for the reaction of 15b with HCl is proposed. The reaction of 15a with concd. HCl in ethanol gives 5‐chloro‐2‐isopropyl‐2,3,7,8,9,10‐hexahydrocycloocta[e]isoindol‐11(6H)‐one (24c). (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2004)
Synthesis of 3‐Substituted Dihydro‐1‐phenylamino‐1H‐pyrrolo[1,2‐a]imidazole‐2,5(3H,6H)‐diones from α‐Amino Acid Phenylhydrazides and Levulinic Acid
Authors: Verardo Giancarlo , Geatti Paola , Merli Marcello , Castellarin Elena E.
Published: June 2004

α‐Amino acid phenylhydrazides 1 readily react with levulinic acid to produce the imidazolidin‐4‐one intermediates 4, which undergo a second ring closure to afford the dihydro‐1H‐pyrrolo[1,2‐a]imidazole‐2,5‐dione derivatives 5. It has been established that the solvent polarity has a great influence on the rate of the second condensation reaction, but not on the first. A mechanism, supported by experimental evidence, has been proposed to explain how the imidazolidin‐4‐one intermediates 4, obtained as expected as a mixture of two diastereoisomers, give a single isomer for the bicyclic derivatives 5; the absolute stereochemistry of these compounds has been determined by X‐ray crystallographic analysis. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2004)
Synthesis, Self‐Assembled Monolayers and Alkaline Earth Metal Ion Recognition of p‐tert‐Butylcalix[4]arene Derivatives
Authors: Zhang Sheng , Song Fayi , Echegoyen Luis
Published: June 2004

The synthesis and self‐assembled monolayer (SAM) formation of three conformational isomers of p‐tert‐butylcalix[4]crown‐6 derivatives 3a−3c and one p‐tert‐butylcalix[4]arene derivative (3d) are reported. The SAMs of these compounds were prepared and characterized by electrochemistry, reflection‐absorption infrared spectroscopy (RAIRS) and contact‐angle measurements. The recognition properties of these SAMs with metal cations were systematically investigated using electrochemical techniques and impedance spectroscopy. The monolayers exhibit selective binding and sensing abilities with alkaline earth metal cations such as Ca2+ and Ba2+ over alkali metal ions. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2004)
Nucleophilic Reactivities of Ketene Acetals
Authors: Tokuyasu Takahiro , Mayr Herbert
Published: June 2004

The kinetics of the reactions of ketene acetals with benzhydrylium ions have been followed photometrically. The reactions proceed with rate‐determining C−C bond formation that is considerably faster than expected for outer‐sphere electron transfer processes. Structure−reactivity relationships are discussed. The excellent correlations between the observed second‐order rate constants (log k2) and the previously reported electrophilicity parameters (E) of benzhydrylium ions suggest that several of the compounds investigated in this work can be used as reference nucleophiles for the development of electrophilicity scales. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2004)
Pyrrolizidine Alkaloids by Intramolecular Palladium‐Catalysed Allylic Alkylation: Synthesis of (±)‐Isoretronecanol
Authors: Lemaire Sébastien , Giambastiani Giuliano , Prestat Guillaume , Poli Giovanni
Published: June 2004

An efficient and stereoconvergent approach to 3‐substituted hexahydroindol‐2‐one derivatives by palladium‐catalysed intramolecular allylic alkylation has been developed. Subsequently, the straightforward conversion of the hexahydroindol‐2‐one 7d into the alkaloid (±)‐isoretronecanol has been performed. The synthesis entails 11 steps starting from 1,3‐cyclohexadiene, affording the final target in a 29% overall yield. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2004)
Carbon Dioxide Fixation by Lithium Amides: DFT Studies on the Reaction Mechanism of the Formation of Lithium Carbamates
Authors: Lill Sten O. Nilsson , Köhn Uwe , Anders Ernst
Published: June 2004

A realistic picture of the complete insertion reaction for a representative example of the formation of carbamates from lithium amides is given by combining experimental information with detailed DFT investigations. The chiral lithium (S)‐2‐(pyrrolidin‐1‐ylmethyl)pyrrolidide (2) reacts with CO2 to form its corresponding carbamate. In this reaction the second pyrrolidine ring functions as an intramolecular ligand, and therefore, “symmetrical” as well as “unsymmetrical” dimers of 2 have to be taken into account. On the energetically preferred reaction pathway this amide exists predominantly as a (solvated) dimer 4. Attack by CO2 occurs in an “end‐on” manner to give several heterodimers 5 which are composed of a lithium amide and a lithium carbamate. This exergonic step is rate determining, and the activation barrier was estimated to be about 10 kcal mol−1. A second lithium‐mediated CO2 insertion into a nitrogen−lithium bond yields the encounter complex 5d−(CO2), which then goes on to form several almost isoenergetic homodimeric carbamates 6. This strongly exergonic insertion reaction has a barrier of 9.7 kcal mol−1. We also report on novel lithium‐mediated CO2 insertion reactions into oxygen−lithium bonds in lithium carbamates. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2004)
Preparation of α,β‐Unsaturated γ‐Keto Aldehydes and New Tetronic Acid and Pyridazine Derivatives by Oxidative Transformations of Alkoxyallene‐Based Dihydrofurans
Authors: Flögel Oliver , Reißig Hans‐Ulrich
Published: June 2004

Oxidation of 3‐alkoxy‐substituted dihydrofuran derivatives 6 and 11 with DDQ unexpectedly provided α,β‐unsaturated γ‐keto aldehydes 10 and 12. A mechanism for this new oxidative ring‐cleavage is presented. Since α,β‐unsaturated γ‐keto aldehydes are versatile intermediates, other 3‐methoxy‐substituted dihydrofuran derivatives 24, 26, and 28 were prepared from lithiated methoxyallene and the corresponding aldehydes. Oxidation of dihydrofuran derivatives with DDQ and subsequent treatment with sodium chlorite furnished hydroxy‐substituted tetronic acid derivatives, such as 30 and 31. Condensation of 30 with hydrazine provided the unsaturated pyridazinone derivative 32. A second route to pyridazine derivatives involves DDQ‐mediated oxidation of dihydrofurans and reaction of the products with hydrazine hydrate. This leads to 4‐methoxypyridazines 33, 34, 35, and 36 in good overall yields. The oxidative transformations of dihydrofuran derivatives reported here demonstrate new examples of reactivity umpolung; the lithiated alkoxyallenes are equivalents of the unusual synthons B and C, which represent anions of malondialdehyde or malonaldehydic acid. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2004)
Cover Picture: The High‐Intrinsic Diels−Alder Reactivity of (−)‐Galiellalactone; Generating Four Quaternary Carbon Centers under Mild Conditions (Eur. J. Org. Chem. 13/2004)
Authors: von Nussbaum Franz , Hanke Roman , Fahrig Thomas , Benet‐Buchholz Jordi
Published: June 2004

The cover picture shows a further chapter in the excitingDiels−Alder story around the natural product (−)‐galiellalactone. This story started with Steglich′s proposal on the Diels−Alder biosynthesis of galiellalactone, that was then experimentally confirmed by T. Anke, H. Anke, and Sterner. The producer of galiellalactone, the ascomycete Galiella rufa, is shown in the background (photo by Pamela Kaminski). Now a surprising intrinsic Diels−Alder reactivity of galiellalactone has been found. Remarkable Diels−Alder transformations are triggered by acylation of the natural product. In the presented case four quaternary carbon centers are generated under mild conditions, yielding a dimer of unprecedented architecture. Details are discussed in the article by F. von Nussbaum et al. on page 2783 ff.
Numerical methods for large‐eddy simulation in general co‐ordinates
Authors: Tang Gefeng , Yang Zhiyin , McGuirk James J.
Published: June 2004

Large scale unsteady motions in many practical engineering flows play a very important role and it is very unlikely that these unsteady flow features can be captured within the framework of Reynolds averaged Navier–Stokes approach. Large‐eddy simulation (LES) has become, arguably, the only practical numerical tool for predicting those flows more accurately since it is still not realistic to apply DNS to practical engineering flows with the current and near future available computing power.

Numerical methods for the LES of turbulent flows in complex geometry have been developed and applied to predict practical engineering flows successfully. The method is based on body‐fitted curvilinear co‐ordinates with the contravariant velocity components of the general Navier–Stokes equations discretized on a staggered orthogonal mesh. For incompressible flow simulations the main source of computational expense is due to the solution of a Poisson equation for pressure. This is especially true for flows in complex geometry. A multigrid 3D pressure solver is developed to speed up the solution. In addition, the Poisson equation for pressure takes a simpler form with no cross‐derivatives when orthogonal mesh is used and hence resulting in increased convergence rate and producing more accurate solutions. Copyright © 2004 John Wiley & Sons, Ltd.
Robust handling of non‐linear constraints for GA optimization of aerodynamic shapes
Authors: Peigin S. , Epstein B.
Published: June 2004

A new approach to the robust handling of non‐linear constraints for GAs (genetic algorithms) optimization is proposed. A specific feature of the approach consists of the change in the conventional search strategy by employing search paths which pass through both feasible and infeasible points (contrary to the traditional approach where only feasible points may be included in a path). The method (driven by full Navier–Stokes computations) was applied to the problem of multiobjective optimization of aerodynamic shapes subject to various geometrical and aerodynamic constraints. The results demonstrated that the method retains high robustness of conventional GAs while keeping CFD computational volume to an acceptable level, which allowed the algorithm to be used in a demanding engineering environment. Copyright © 2004 John Wiley & Sons, Ltd.
Novel method for studying myelination in vivo reveals that EDTA is a potent inhibitor of myelin protein and mRNA expression during development of the rat sciatic nerve
Authors: Meintanis Stathis , Thomaidou Dimitra , Jessen Kristjan R. , Mirsky Rhona , Matsas Rebecca
Published: June 2004

To probe the effects of possible inhibitors or enhancers of in vivo myelination, we have modified a technique widely used in studies of the developing neuromuscular system that involves incorporation of test compounds into a silicon rubber solution, which solidifies on contact with air. U‐shaped rubber implants are inserted around the sciatic nerve of 1‐day‐old rats and left in place for 24–48 h. Sections from the region of the nerve lying within the implant, with or without the test compound, are then immunolabeled, examined with in situ hybridization or electron microscopy. Application of EDTA (440 μg/implant) in this way strongly suppressed the levels of the myelin‐associated molecules protein P0, myelin basic protein (MBP), and galactocerebroside (Galc). mRNA levels for P0 and the myelin‐related transcription factor Krox‐20 were also reduced, further supporting association of the EDTA‐induced effect with the myelinating Schwann cells. In contrast, no obvious differences were observed in either neurofilament (NF) protein or glial fibrillary acidic protein (GFAP) expression, suggesting absence of influence on axons or nonmyelinating Schwann cells. Despite the severely altered molecular composition of myelin in the presence of EDTA, examination in the electron microscope did not reveal any apparent ultrastructural changes in the myelin sheaths or nerve development. This work introduces a novel method for studying nerve development and shows that EDTA, which chelates divalent cations such as Ca2+ and Mg2+, strongly and selectively reduces levels of molecules, which, on postnatal days 1–4, are expressed in myelinating cells at much higher levels than in cells not engaged in myelination. © 2004 Wiley‐Liss, Inc.
K+channel blockade impairs remyelination in the cuprizone model
Authors: Bacia Andrzej , Wollmann Robert , Soliven Betty
Published: June 2004

The adult CNS has the capacity to remyelinate following metabolic, toxic and autoimmune demyelinating insults. In cuprizone‐induced demyelination, spontaneous remyelination occurs after the cessation of cuprizone diet. We used the cuprizone model to investigate the role of glial K+channels in oligodendroglial (OLG) regeneration and remyelination in vivo. We found that treatment with 4‐aminopyridine (4‐AP), a broad‐spectrum K+channel antagonist, results in: (1) decreased number of oligodendroglial progenitors (OP) and OLGs; (2) diminished astrogliosis; and (3) decreased remyelination in the corpus callosum based on the immunoreactivity to myelin basic protein (MBP), Rip monoclonal antibody, and by electron microscopy. Our findings support the concept that glial K+channels play an important role during OLG regeneration and remyelination, a crucial factor to be considered during the development of therapeutic strategies to facilitate recovery in demyelinating diseases and spinal cord injury. © 2004 Wiley‐Liss, Inc.
Effects of diabetes mellitus on astrocyte GFAP and glutamate transporters in the CNS
Authors: Coleman Elaine , Judd Robert , Hoe Lori , Dennis John , Posner Philip
Published: June 2004

Diabetes mellitus increases the risk of central nervous system (CNS) disorders such as stroke, seizures, dementia, and cognitive impairment. The cellular mechanisms responsible for the increased risk of these disorders are incompletely understood. Astrocytes are proving critical for normal CNS function, and alterations in their activity could contribute to diabetes‐related disturbances in the brain. We examined the effects of streptozotocin (STZ)‐induced diabetes in rats on the level of the astrocyte intermediate filament protein, glial fibrillary acidic protein (GFAP), number of astrocytes, and levels of the astrocyte glutamate transporters, glutamate transporter‐1 (GLT‐1) and glutamate/aspartate transporter (GLAST), in the cerebral cortex, hippocampus, and cerebellum by Western blotting (WB) and immunohistochemistry (IH). Studies were carried out at 4 and 8 weeks of diabetes duration. Diabetes resulted in a significant decrease in GFAP protein levels (WB) in the hippocampus and cerebellum at 4 weeks and in the cerebral cortex, hippocampus and cerebellum by 8 weeks. Attenuated GFAP immunoreactivity (IH) was evident in the hippocampus, cerebellum and white matter regions such as the corpus callosum and external capsule at both 4 and 8 weeks of diabetes. Astrocyte cell counts of adjacent sections immunoreactive for S‐100B were not different between control and diabetic animals. No significant differences were noted in astrocyte glutamate transporter levels in the cerebral cortex, hippocampus, or cerebellum at either time period (WB, IH). With the expanding list of astrocyte functions in the CNS, the role of astrocytes in diabetes‐induced CNS disorders clearly warrants further investigation. © 2004 Wiley‐Liss, Inc.
Gap junction channels coordinate the propagation of intercellular Ca2+ signals generated by P2Y receptor activation
Authors: Suadicani S.O. , Flores C.E. , Urban‐Maldonado M. , Beelitz M. , Scemes E.
Published: June 2004

Astrocytes express gap junction proteins and multiple types of P2Y receptors (P2YRs) that contribute to the propagation of intercellular Ca2+ waves (ICW). To gain access to the role played by gap junctional communication in ICW propagation generated by P2YR activation, we selectively expressed P2Y1,2,4R subtypes and Cx43 in the human 1321N1 astrocytoma cell line, which lacks endogenous P2 receptors. Fluorescence recovery after photobleaching revealed that 1321N1 cells are poorly dye‐coupled and do not propagate ICW. Forced expression of Cx43 in 1321N1 cells (which did not show functional hemichannels) increased dye coupling and allowed short‐range ICW transmission that was mainly mediated by intercellular diffusion of Ca2+ generated in the stimulated cells. Astrocytoma clones expressing each of the P2YR subtypes were also able to propagate ICWs that were likely dependent on IP3 generation. These waves exhibited properties particular to each P2YR subtype. Co‐expression of eGFP‐hCx43 and P2Y1R modified the properties of P2Y1R‐generated ICW to those characteristics of P2Y2R. Increased coupling in P2Y4R clones induced by expression of eGFP‐hCx43 abolished the ICWs observed in uncoupled P2Y4R clones. No changes in the behavior of ICWs generated in P2Y2R clones were observed after forced expression of Cx43. These data indicate that in 1321N1 cells gap junctional communication provides intercellular integration of Ca2+ signals generated by P2YR activation, thus coordinating the propagation of intercellular calcium waves. © 2004 Wiley‐Liss, Inc.
Estimation of multiscale neurophysiologic parameters by electroencephalographic means
Authors: Robinson P.A. , Rennie C.J. , Rowe D.L. , O'Connor S.C.
Published: June 2004

It is shown that new model‐based electroencephalographic (EEG) methods can quantify neurophysiologic parameters that underlie EEG generation in ways that are complementary to and consistent with standard physiologic techniques. This is done by isolating parameter ranges that give good matches between model predictions and a variety of experimental EEG‐related phenomena simultaneously. Resulting constraints range from the submicrometer synaptic level to length scales of tens of centimeters, and from timescales of around 1 ms to 1 s or more, and are found to be consistent with independent physiologic and anatomic measures. In the process, a new method of obtaining model parameters from the data is developed, including a Monte Carlo implementation for use when not all input data are available. Overall, the approaches used are complementary to other methods, constraining allowable parameter ranges in different ways and leading to much tighter constraints overall. EEG methods often provide the most restrictive individual constraints. This approach opens a new, noninvasive window on quantitative brain analysis, with the ability to monitor temporal changes, and the potential to map spatial variations. Unlike traditional phenomenologic quantitative EEG measures, the methods proposed here are based explicitly on physiology and anatomy. Hum. Brain Mapping 23:53–72, 2004. © 2004 Wiley‐Liss, Inc.
Monocular visual activation patterns in albinism as revealed by functional magnetic resonance imaging
Authors: Schmitz Bernd , Käsmann‐Kellner Barbara , Schäfer Torsten , Krick Christoph M. , Grön Georg , Backens Martin , Reith Wolfgang
Published: June 2004

Human albinism is characterized by a disturbance of the chiasmatic projection system leading to predominant representation of just one eye in the contralateral hemisphere. Patients show congenital nystagmus without perceiving oscillopsia. The purpose of the present study was to demonstrate the consequences of atypical chiasmatic crossing with monocular visual stimulation using functional magnetic resonance imaging (fMRI). Sixteen patients with albinism and fifteen normally pigmented controls were stimulated with a monocular visual activation paradigm using flickering checkerboards. In patients, we observed contralaterally dominated activation of visual cortices correlating to clinical albinism parameters. This confirms albinism as a continuous range of hypopigmentation disorders. Additionally, albinos showed activation of the superior colliculus and of visual motion areas although the stimulus was stationary. Activation of visual motion areas is due probably to congenital nystagmus without a conscious correlate like oscillopsia. Hum. Brain Mapping 23:40–52, 2004. © 2004 Wiley‐Liss, Inc.