bluuebunny/crossref_metadata_2025 huggingface dataset:
Noise suppression of spectrum‐sliced light using semiconductor optical amplifiers
Authors: Yamatoya Takeshi , Koyama Fumio
Published: January 2003

The spectrum slicing method for slicing incoherent light by using an optical filter can easily generate large‐scale multiwavelength light in bundles, but has the problem of beat noise in the spontaneously emitted light. We proposed noise suppression of spectrum‐sliced light by using the characteristic of the gain saturation of the semiconductor optical amplifier (SOA). In this paper, we quantitatively measure noise suppression by gain saturation of this SOA and improve the noise suppression in the high‐frequency domain by lengthening the SOA. We also measure the bit error rate (BER) of the spectrum‐sliced light that was noise suppressed by a gain‐saturated SOA and demonstrate its possible application to light sources for high‐speed optical transmission. Finally, we propose and demonstrate a gain‐saturated SOA modulator that simultaneously suppresses noise and modulates. © 2003 Wiley Periodicals, Inc. Electron Comm Jpn Pt 2, 86(2): 28–35, 2003; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/ecjb.10077
Audio data hiding using band elimination
Authors: Ikeda Mikio , Toyoshima Ryouzoh , Takeda Kazuya , Itakura Fumitada
Published: January 2003

In this paper, we will propose a data hiding technique for audio signals using the combinations of center frequencies and bandwidths of band elimination. For that reason, we have investigated, by subjective evaluation, the detection limits of the center frequencies and bandwidths of band elimination. From the results of the subjective evaluation, the center frequencies and bandwidths of band elimination for data hiding are determined. As a result of actually performing data hiding, it was learned that the degradation due to process distortion was scarcely perceived. When data of about 10.8 bits/s per channel are actually embedded in an audio signal of CD quality, the extraction error of the data can be reduced below 1%. However, when MPEG coding of low bit rate is performed on a TSP signal, it was learned that the extraction error becomes larger. © 2003 Wiley Periodicals, Inc. Electron Comm Jpn Pt 2, 86(2): 57–67, 2003; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/ecjb.10120
The secret‐key block cipher MKC1
Authors: Matsukawa Koichi , Kobayashi Kunikatsu
Published: January 2003

This paper proposes a new secret‐key block cipher algorithm called MKC1. MKC1 is a Feistel‐type block cipher with variable block length, key length, and number of rounds. The merits of this algorithm are that numerous structures can be formed by selecting the encryption functions and that cellular automata, which can ensure an immense number of structures, and power functions over a finite field, which have strong resistance to differential and linear attacks, are used for the encryption functions. These features increase MKC1's resistance to exhaustive key searches and analytical cryptanalysis. In addition, rules that are relatively powerful as encryption functions among the total of 256 rules for one‐dimensional, two‐state, neighborhood‐three cellular automata were found to practically correspond to Class 3 of Wolfram's classification scheme. The results of computer experiments indicate that MKC1 is several times faster than DES. The industrial value of this algorithm was confirmed by implementing it in an automatic electronic ticket system using a contactless IC card. © 2003 Wiley Periodicals, Inc. Electron Comm Jpn Pt 2, 86(2): 68–83, 2003; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/ecjb.10123
A study on performance of pitch extraction by BPFP method under pitch variation, wave disturbance, and noise
Authors: Miyabayashi Hideo , Funada Tetsuo
Published: January 2003

Detection of the pitch frequency as the basic characteristic of the voice is one of the most important research subjects in voice analysis and synthesis. In this paper we study, in comparison with the cepstrum method and the modified correlation method, how the BPFP method using bandpass filter pairs and the BPFP‐NN method combining the BPFP banks and the neural network (NN) proposed by the authors perform in the presence of pitch disturbance, waveform variation, and added noise when applied to voiced/unvoiced (U/V) detection and pitch extraction. According to the experimental results, the BPFP method and the BPFP‐NN method exhibit more stable performance and more effective pitch extraction with U/V detection than the typical cepstrum method and modified correlation method, and the effect is particularly evident at low frequencies due to the logarithmic expression of the center frequency spacing of the BPFP banks. © 2003 Wiley Periodicals, Inc. Electron Comm Jpn Pt 3, 86(5): 24–35, 2003; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/ecjc.10045
Identification of PINCH in Schwann cells and DRG neurons: Shuttling and signaling after nerve injury
Authors: Campana W. Marie , Myers Robert R. , Rearden Ann
Published: January 2003

Particularly interesting new cysteine‐histidine rich protein (PINCH) is a double zinc finger domain (LIM)‐only adapter protein that functions to recruit the integrin‐linked kinase (ILK) to sites of integrin activation. Genetic studies have shown that PINCH and ILK are required for integrin signaling. Since integrin activation is associated with Schwann cell migration, neurite outgrowth and regeneration, this study examined PINCH in the normal peripheral nervous system and after chronic constriction injury (CCI) in adult Sprague‐Dawley rats. Immunohistochemistry identified PINCH immunoreactivity in cell bodies of dorsal root ganglia (DRG) neurons, axons, satellite cells, and Schwann cells. PINCH immunostaining was localized to the membrane of uninjured DRG cell bodies consistent with its localization at a site of integrin activation. In contrast, 5 days following CCI, PINCH immunostaining was diffuse throughout the DRG cell cytoplasm. Confocal microscopy of primary and transformed Schwann cells localized PINCH in cytoplasmic, perinuclear and nuclear areas. Examination of the PINCH sequence revealed a putative leucine‐rich nuclear export signal (NES) and an overlapping basic nuclear localization signal (NLS). To demonstrate nuclear export of PINCH, rabbit anti‐PINCH IgG was microinjected into Schwann cell nuclei and allowed to combine with PINCH contained within the nucleus. Immunofluorescence showed that the PINCH and anti‐PINCH IgG complex rapidly translocated to the cytoplasm. Treatment with leptomycin B caused nuclear accumulation of PINCH, indicating that the CRM1 pathway mediates nuclear export of PINCH. ILK activity in Schwann cells was enhanced by platelet‐derived growth factor (PDGF) and tumor necrosis factor α. PINCH immunoprecipitates from PDGF‐ and TNFα‐stimulated Schwann cells contained several high‐molecular‐weight threonine‐phosphorylated proteins. Taken together, these results indicate that PINCH is an abundant shuttling/signaling protein in Schwann cells and DRG neurons. GLIA 41:213–223, 2003. © 2003 Wiley‐Liss, Inc.
Olfactory ensheathing cells of the lamina propria in vivo and in vitro
Authors: Au Edmund , Roskams A. Jane
Published: January 2003

Olfactory ensheathing cells (OECs) continuously support the regeneration of olfactory receptor neurons (ORNs). In addition, OECs promote regeneration of neurons within the CNS in a number of transplantation paradigms, but details of exactly how they support regeneration remain elusive. The majority of studies using OECs to promote regeneration have thus far focused on understanding the cell biology of OECs purified from the olfactory bulb (OB). Here we show that a population of OECs similar to those obtained from the OB is present in the lamina propria (LP) beneath the olfactory epithelium (OE). These OECs are the first glial cells encountered by the axons of developing ORNs as they exit the OE and display distinct and variable expression of p75, S100β, GFAP, and O4, characteristic markers of bulb OECs. Once purified in vitro, they display Schwann cell‐like and astrocyte‐like properties and expand rapidly. In addition to resembling OB‐OECs, LP‐OECs also express a unique combination of developmentally important proteins—CD 44, β1 integrin, P200, Notch 3, NG2, VEGF, and PACAP and CREB binding protein (CBP/p300)—not previously reported in OB‐OECs. These data suggest that LP‐OECs, like OB‐OECs, are a developmentally distinct class of glia that are capable of both immature and mature function, depending on environmental stimuli, within the adult nervous system. GLIA 41:224–236, 2003. © 2003 Wiley‐Liss, Inc.
Nodal sodium channel domain integrity depends on the conformation of the paranodal junction, not on the presence of transverse bands
Authors: Rosenbluth Jack , Dupree Jeffrey L. , Popko Brian
Published: January 2003

Our understanding of the role that axoglial interactions play in node of Ranvier formation and maintenance remains incomplete. Previous studies of CNS myelinated fibers of CGT‐null mice showed abnormalities in the arrangement of paranodal myelin loops and absence of a conspicuous component of the paranodal junction, the ridge‐like intercellular transverse bands. Axolemmal sodium channel domains were largely preserved at nodes of Ranvier but displayed some abnormalities in form. Using a combination of freeze‐fracture and immunocytochemical methods, we have found additional evidence documenting abnormalities in the size, shape, and location of axolemmal sodium channel clusters in CGT‐null mice as well as evidence that these nodal abnormalities are complementary to the organization of paranodal myelin loops, despite the absence of transverse bands. We conclude that the differentiated form of the nodal axolemma and the distribution of axolemmal sodium channels depend on the conformation of paranodal axoglial contacts but not on the presence of transverse bands at the sites of contact. GLIA 41:318–325, 2003. © 2003 Wiley‐Liss, Inc.
Upregulation of gp130 and STAT3 activation in the rat hippocampus following transient forebrain ischemia
Authors: Choi Jeong‐Sun , Kim Seong Yun , Cha Jung‐Ho , Choi Yun‐Sik , Sung Ki‐Wug , Oh Seong Taek , Kim Ok Nyu , Chung Jin‐Woong , Chun Myung‐Hoon , Lee Sang Bok , Lee Mun‐Yong
Published: January 2003

To determine whether the pathophysiological processes after transient forebrain ischemia are mediated via a signal pathway involving gp130 (a signal transducer for the interleukin‐6 family), we analyzed changes in the expression of gp130 and its downstream transcription factor, signal transducer and activator of transcription factor 3 (STAT3), in the rat hippocampus of a four‐vessel occlusive ischemia model. Expression of gp130 mRNA was restricted to neurons of the pyramidal cell and granule cell layers in control animals. Four hours after ischemic injury, astrocytes expressed gp130 mRNA. Expression of gp130 increased preferentially in the CA1 and dentate hilar regions, and was maintained for at least 2 weeks. Increase in gp130 expression was accompanied by the activation of STAT3 following ischemic injury. Four hours after injury, STAT3 and phosphorylated STAT3 (pSTAT3) were observed in the nuclei of the dentate hilar region, and sequentially in the CA1 region at day 1. By day 3, STAT3 immunoreactivity markedly increased in these areas, where small cells with the morphology of astrocytes showed nuclear and cytoplasmic STAT3 and nuclear pSTAT3 immunoreactivities. These patterns were especially maintained in the CA1 area until 14 days of reperfusion. Double‐labeling experiments revealed that the cells expressing STAT3 and pSTAT3 were glial fibrillary acidic protein‐expressing reactive astrocytes. These results show a coordinated and long‐lasting upregulation of gp130 mRNA and STAT3 activation in reactive astrocytes of the postischemic hippocampus, indicating that they may be involved in the astrocytic response to an ischemic insult. GLIA 41:237–246, 2003. © 2003 Wiley‐Liss, Inc.
Highly differential expression of SN1, a bidirectional glutamine transporter, in astroglia and endothelium in the developing rat brain
Authors: Boulland Jean‐Luc , Rafiki Amina , Levy Line M. , Storm‐Mathisen Jon , Chaudhry Farrukh A.
Published: January 2003

The transmitters glutamate and GABA also subserve trophic action and are required for normal development of the brain. They are formed from glutamine, which may be synthesized in glia or extracted from the blood. In the adult, the glutamine transporter SN1 is expressed in the astroglia. SN1 works in both directions, depending on the concentration gradients of its substrates and cotransported ions, and is thought to regulate extracellular glutamine and to supply the neurons with the transmitter precursor. In this article, we have quantified the expression and studied the localization of SN1 at different developmental stages. SN1 is expressed in astroglia throughout the CNS from embryonic stages through adulthood. No indication of SN1 staining in neuronal elements has been obtained at any stage. Quantitative immunoblotting of whole brain extracts demonstrates increasing expression of SN1 from P0, reaching a peak at P14, twice the adult level. A moderate and slower rise and fall of the expression levels of SN1 occurs in the cerebellum. Strong transient SN1‐like staining is also found in Bergmann glia and vascular endothelium in the first postnatal weeks. Strong intracellular staining in the same time period suggests a high rate of SN1 synthesis in the early postnatal period. This coincides with the increasing levels of glutamate and GABA in the CNS and with the time course of synaptogenesis. This study suggests that the expression of SN1 is highly regulated, correlating with the demand for glutamine during the critical period of development. GLIA 41:260–275, 2003. © 2003 Wiley‐Liss, Inc.
Recovery from lactacidosis‐induced glial cell swelling with the aid of exogenous anion channels
Authors: Nabekura Takashi , Morishima Shigeru , Cover Timothy L. , Mori Shin‐Ichiro , Kannan Hiroshi , Komune Shizuo , Okada Yasunobu
Published: January 2003

Hypotonic challenge induces transient swelling in glial cells, which is typically followed by a regulatory volume decrease (RVD). In contrast, lactic acidosis (lactacidosis) induces persistent cell swelling in astrocytes without an accompanying RVD. In the present study, we studied the mechanisms by which lactacidosis interferes with normal volume regulation in rat astrocytic glioma C6 cells. Following exposure of C6 cells to a hypotonic challenge, a current was detected that exhibited properties consistent with those of volume‐sensitive outwardly rectifying (VSOR) anion channels. When exposed to in vitro conditions designed to simulate lactacidosis, C6 cells failed to respond to hypotonic stress with an RVD, and VSOR anion currents were not activated. When added to C6 cells, an anion channel‐forming protein purified from Helicobacter pylori, VacA, was found to form anion‐selective channels in the plasma membrane, and the activity of the VacA channel was not affected by lactacidosis (pH 6.2). Cells preincubated with VacA and then exposed to lactacidotic conditions underwent transient swelling followed by RVD. In contrast, application of a cation ionophore, gramicidin, failed to inhibit lactacidosis‐induced persistent cell swelling. From these results, we conclude that inhibition of a volume‐sensitive anion channel contributes to persistent swelling induced by lactacidosis in glial cells. Introduction of anion channel activity into glial cells might provide a novel approach for treating cerebral edema, which is associated with lactacidosis in cerebral ischemia or head injury. GLIA 41:247–259, 2003. © 2003 Wiley‐Liss, Inc.
Oligodendrocyte precursor hypercellularity and abnormal retina development in mice overexpressing PDGF‐B in myelinating tracts
Authors: Forsberg‐Nilsson Karin , Erlandsson Anna , Zhang Xiao‐Qun , Ueda Hideho , Svensson Kristian , Nistér Monica , Trapp Bruce D. , Peterson Alan C. , Westermark Bengt
Published: January 2003

Platelet‐derived growth factor (PDGF) influences the generation of neurons and glia during embryogenesis and in early postnatal life. In an attempt to determine the consequences of an overexpression of PDGF‐B during the first weeks of life, we targeted transgenic expression of a human PDGF‐B cDNA to myelinating tracts using the promoter region of the myelin basic protein (MBP) gene. Transgenic mRNA and protein were expressed in the brain and the expression profile of the human PDGF‐B during early postnatal development closely paralleled that of the endogenous mouse MBP gene. The gross morphological appearance of transgenic brains was normal but at the cellular level several phenotypic alterations could be identified. In white matter tracts such as the corpus callosum and cerebellar medulla, there was a marked hypercellularity. The number of oligodendrocyte precursors was increased and astrocytes were more abundant. In adult mice carrying the MBP–PDGF‐B transgene, however, myelination appeared normal and the amount of oligodendrocytes was similar to that of control littermates. In addition to the phenotypic alterations in the brain, investigation of eye structure revealed a striking disorganization of retinal architecture. The retina was folded with cells collected in papillar or follicular‐like structures. Retinal whole mount preparations after India ink perfusion revealed capillary disorganization with large‐caliber vessels supporting only a few fine branches. Our observations strengthen the notion that PDGF is an important effector molecule in postnatal CNS development. GLIA 41:276–289, 2003. © 2003 Wiley‐Liss, Inc.
Expression of protein zero is increased in lesioned axon pathways in the central nervous system of adult zebrafish
Authors: Schweitzer Jörn , Becker Thomas , Becker Catherina G. , Schachner Melitta
Published: January 2003

The immunoglobulin superfamily molecule protein zero (P0) is important for myelin formation and may also play a role in adult axon regeneration, since it promotes neurite outgrowth in vitro. Moreover, it is expressed in the regenerating central nervous system (CNS) of fish, but not in the nonregenerating CNS of mammals. We identified a P0 homolog in zebrafish. Cell type‐specific expression of P0 begins in the ventromedial hindbrain and the optic chiasm at 3–5 days of development. Later (at 4 weeks) expression has spread throughout the optic system and spinal cord. This is consistent with a role for P0 in CNS myelination during development. In the adult CNS, glial cells constitutively express P0 mRNA. After an optic nerve crush, expression is increased within 2 days in the entire optic pathway. Expression peaks at 1 to 2 months and remains elevated for at least 6 months postlesion. After enucleation, P0 mRNA expression is also upregulated but fails to reach the high levels observed in crush‐lesioned animals at 4 weeks postlesion. Spinal cord transection leads to increased expression of P0 mRNA in the spinal cord caudal to the lesion site. The glial upregulation of P0 mRNA expression after a lesion of the adult zebrafish CNS suggests roles for P0 in promoting axon regeneration and remyelination after injury. GLIA 41:301–317, 2003. © 2003 Wiley‐Liss, Inc.
Astrocytes are the major intracerebral source of macrophage inflammatory protein‐3α/CCL20 in relapsing experimental autoimmune encephalomyelitis and in vitro
Authors: Ambrosini Elena , Columba‐Cabezas Sandra , Serafini Barbara , Muscella Antonella , Aloisi Francesca
Published: January 2003

Macrophage inflammatory protein‐3α/CCL20 is a recently identified chemokine that binds to CCR6 and acts as a chemoattractant for memory/differentiated T‐cells, B‐cells, and immature dendritic cells. We have previously reported that CCL20 and CCR6 mRNAs are expressed in the CNS of SJL mice with experimental autoimmune encephalomyelitis (EAE) and that CCL20 is produced by CNS‐infiltrating leukocytes at disease onset and, additionally, by intraparenchymal astrocyte‐like cells during disease relapses. In this study, we provide further immunohistochemical evidence that astrocytes represent the main CNS source of CCL20 during EAE. Moreover, we show that the proinflammatory cytokines interleukin‐1β and tumor necrosis factor‐α, but not interferon‐γ, induce expression of CCL20 mRNA and secretion of CCL20 protein in cultures of mouse brain‐derived astrocytes. We also show that supernatants from cytokine‐activated astrocytes stimulate the migration of polarized T helper cells and that this effect is partially inhibited by anti‐CCL20 antibody. These findings suggest that, through secretion of CCL20, astrocytes could play an important role in orchestrating the recruitment of specific leukocyte subsets to the inflamed CNS and in regulating CNS‐targeted immune responses. GLIA 41:290–300, 2003. © 2003 Wiley‐Liss, Inc.
Synthesis of high specific activity 35S‐labelled N‐methanesulfonyl farnesylcysteine and a photoactive analog
Authors: Kale Tamara A. , Raab Conrad , Yu Nathan , Aquino Evelyn , Dean Dennis C. , Distefano Mark D.
Published: January 2003

Prenylated cysteine analogs, which mimic the prenylated cysteine residue of prenylated GTP‐binding proteins (G‐proteins), have been used in a variety of contexts for the study of prenylated G‐protein behavior. In earlier work in this area, we prepared the photoactive analog [35S]4 and showed that it labelled RhoGDI upon photolysis; those results were consistent with the idea that GDI contains an isoprenoid binding site. Here, we describe the preparation of [35S]N‐methanesulfonyl labelled analogs (1a and 2a) of N‐acetyl farnesylcysteine and its methyl ester together with an improved synthetic procedure for photoactive analogs 3 and 4; specific activities of ∼1100 Ci/mmol were achieved. Compounds 1a and 2a in unlabelled form were used as competitors in photolysis reactions to show that the methanesulfonamido group is a reasonable acetamide substitution. Additional experiments show that the photoactive ester [35S]3 can cross‐link GDI in both purified form and crude bacterial extract. However, the extent of cross‐linking obtained with the ester ([35S]3) is significantly less than that observed with the free acid ([35S]4) despite the fact that the esterified form probably more closely reflects the structure of the C‐terminus of a prenylated protein; using the GDI·Cdc42 co‐crystal structure, the structural basis for these results is discussed. Copyright © 2002 John Wiley & Sons, Ltd.
Preparation of 4‐[11C]methylmetaraminol, a potential PET tracer for assessment of myocardial sympathetic innervation
Authors: Langer Oliver , Forngren Tobias , Sandell Johan , Dollé Frédéric , Långström Bengt , Någren Kjell , Halldin Christer
Published: January 2003

The false adrenergic neurotransmitter [11C]meta‐hydroxyephedrine ([11C]HED) is currently the PET tracer of choice for assessment of myocardial sympathetic innervation. The molecule is metabolised in the 4‐position of the aromatic ring. The resulting radiolabelled metabolites need to be measured in order to obtain an arterial input function. Our aim was the development of a PET tracer with an increased metabolic stability relative to [11C]HED. We selected 4‐methylmetaraminol as a candidate molecule for radiolabelling with 11C (t1/2 20.4 min). Our radiosynthetic approach towards 4‐[11C]methylmetaraminol involved a palladium‐catalyzed cross‐coupling reaction of a protected 4‐trimethylstannyl derivative of metaraminol with [11C]methyl iodide followed by removal of the protective groups. 4‐[11C]methylmetaraminol was obtained in a final decay‐corrected radiochemical yield of 20–25% within a synthesis time of 60–80 min. The specific radioactivity at the end of the synthesis ranged from 18–37 to GBq/μmol. The unlabelled reference molecule, 4‐methylmetaraminol, was prepared in a 5‐step synthesis starting from metaraminol. A biological evaluation of 4‐[11C]methylmetaraminol is in progress and the results will be reported elsewhere. Copyright © 2002 John Wiley & Sons, Ltd.
Synthesis and characterization of the potent cannabinoid agonist [naphthyl‐3H] WIN 55212‐2 at high specific activity
Authors: Seguin Richard J. , Filer Crist N.
Published: January 2003

[Naphthyl‐3H] WIN 55212‐2 was prepared at high specific activity by the catalytic tritiation of dibromo precursor 3. Product 4 was characterized by chromatography as well as tritium NMR and proven to be a useful radioligand. Copyright © 2002 John Wiley & Sons, Ltd.
Synthesis and biological characteristics of the Technetium‐99m triamide derivatives of mercaptoacetyltriglycine (MAG3)
Authors: Okarvi S. M. , Adriaens P. , Verbruggen A. M.
Published: January 2003

A number of MAG3‐derivatives, containing the three‐amide functions modified by inverting the sequence –CO–NH– to –NH–CO– in the first‐ and/or second‐amide functions, have been labelled with 99mTc in order to study the renal characteristics of the resulting MAG3‐derivatives versus the reference 99mTc‐MAG3. The 99mTc‐MAG3‐derivatives displayed HPLC‐profiles similar to that of 99mTc‐MAG3. Furthermore, in mice they exhibited biological behaviour comparable or even superior to 99mTc‐MAG3, which indicates that the sequence of the first‐ and/or second‐amide bond in 99mTc‐MAG3 affects the biological behaviour only to a limited degree. However, in baboon the plasma clearance of the 99mTc‐MAG3‐derivatives was relatively slower than 99mTc‐MAG3, which underlines the inter‐species variability. Copyright © 2002 John Wiley & Sons, Ltd.
Synthesis of PNU‐243922 labelled with 14C
Authors: Felicini C. , Fontana E. , Giribone D. , M. Nilsson B. , Österlund K. , Paptchikhine A.
Published: January 2003

The arylpiperazine PNU‐243922 (1) has been labelled with 14C. A two‐step sequence starting from the 14C labelled alcohol 6a led to radiochemically pure (>98%) [14C]PNU‐243922 with a specific activity of 546 MBq/mmol Copyright © 2002 John Wiley & Sons, Ltd.
Synthesis of N‐(piperidin‐1‐yl)‐5‐(4‐methoxyphenyl)‐1‐(2‐chlorophenyl)‐4‐[18F]fluoro‐1H‐pyrazole‐3‐carboxamide by nucleophilic [18F] fluorination: a PET radiotracer for studying CB1 cannabinoid receptors
Authors: Katoch‐Rouse Reeti , Horti Andrew G.
Published: January 2003

The feasibility of nucleophilic displacement of bromide in the 4‐bromopyrazole ring with [18F]fluoride has been demonstrated by the synthesis of two radiolabeled compounds: N‐(piperidin‐1‐yl)‐5‐(4‐methoxyphenyl)‐1‐(2‐chlorophenyl)‐4‐[18F]fluoro‐1H‐pyrazole‐3‐carboxamide, ([18F] NIDA‐42033) 1b and 1‐(2‐chlorophenyl)‐4‐[18F]fluoro‐5‐(4‐methoxyphenyl)‐1H‐pyrazole‐3‐carboxylic acid, ethyl ester 4. The radiochemical yields were in the range of 1–6%. [18F]NIDA‐42033, a potential radiotracer for the study of CB1 cannabinoid receptors in the animal brain by positron emission tomography, has been synthesized in sufficient quantities with specific radioactivity greater than 2500 mCi/μmol and radiochemical purity >95%. Copyright © 2002 John Wiley & Sons, Ltd.
Preparation of 1,2,3,4,6‐penta‐O‐galloyl‐[U‐14C]‐D‐glucopyranose
Authors: Chen Yumin , E. Hagerman Ann , E. Minto Robert
Published: January 2003

The synthesis of 1,2,3,4,6‐penta‐O‐galloyl‐[U‐14C]‐D‐glucopyranose is described. [U‐14C] glucopyranose was reacted with tri‐O‐benzylgallic acid forming 1,2,3,4,6‐penta(tri‐O‐benzylgalloyl)‐[U‐14C]‐D‐glucopyranose as chromatograpically separable anomers. Removal of the benzyl group by catalytic hydrogenation afforded 1,2,3,4,6‐penta‐O‐galloyl‐[U‐14C]‐D‐glucopyranose in 54% overall yield. Copyright © 2002 John Wiley & Sons, Ltd.
Origin of the conformation dependence of protein charge‐state distributions in electrospray ionization mass spectrometry
Authors: Grandori Rita
Published: January 2003

The influence of tertiary structure on the electrospray ionization mass spectra of proteins is a well known and broadly exploited phenomenon. However, the underlying mechanism is not well understood. This paper discusses the bases and the implications of the two current hypotheses (solvent accessibility and Coulombic repulsions), pointing out the remaining open questions. Evidence reported here supports a third hypothesis, i.e. that intramolecular interactions in folded proteins play a key role in determining the observed charge‐state distributions. It is proposed that native protein structures stabilize to a large extent pre‐existing charges of the opposite polarity to the net charge of the ion, preventing their neutralization during the electrospray process. Thus, the higher charge states of unfolded conformations, relative to the folded structure, would not derive from a more extensive ionization of the former, but rather from a higher content of neutralizing charges in the latter. This interpretation allows several other problematic observations to be explained, including the different shapes of the spectra of folded and unfolded proteins, the discrepancies between observed and predicted gas‐phase reactivity of protein ions and the apparent inconsistency of positive‐ and negative‐ion mode results. Copyright © 2003 John Wiley & Sons, Ltd.
Feasibility of a liquid‐phase microextraction sample clean‐up and liquid chromatographic/mass spectrometric screening method for selected anabolic steroid glucuronides in biological samples
Authors: Kuuranne Tiia , Kotiaho Tapio , Pedersen‐Bjergaard Stig , Einar Rasmussen Knut , Leinonen Antti , Westwood Steven , Kostiainen Risto
Published: January 2003

Anabolic androgenic steroids (AAS) are metabolized extensively in the human body, resulting mainly in the formation of glucuronide conjugates. Current detection methods for AAS are based on gas chromatographic/mass spectrometric (GC/MS) analysis of the hydrolyzed steroid aglycones. These analyses require laborious sample preparation steps and are therefore time consuming. Our interest was to develop a rapid and straightforward method for intact steroid glucuronides in biological samples, using liquid‐phase microextraction (LPME) sample clean‐up and concentration method combined with liquid chromatographic/tandem mass spectrometric (LC/MS/MS) analysis. The applicability of LPME was optimized for 13 steroid glucuronides, and compared with conventional liquid–liquid extraction (LLE) and solid‐phase extraction (SPE) procedures. An LC/MS/MS method was developed for the quantitative detection of AAS glucuronides, using a deuterium‐labeled steroid glucuronide as the internal standard. LPME, owing to its high specificity, was shown to be better suited than conventional LLE and SPE for the clean‐up of urinary AAS glucuronides. The LPME/LC/MS/MS method was fast and reliable, offering acceptable reproducibility and linearity with detection limits in the range 2–20 ng ml−1 for most of the selected AAS glucuronides. The method was successfully applied to in vitro metabolic studies, and also tested with an authentic forensic urine sample. For a urine matrix the method still has some unsolved problems with specificity, which should be overcome before the method can be reliably used for doping analysis, but still offering additional and complementary data for current GC/MS analyses. Copyright © 2003 John Wiley & Sons, Ltd.
Liquid chromatographic/electrospray ionization tandem mass spectrometric study of the phenolic composition of cocoa (Theobroma cacao)
Authors: Sánchez‐Rabaneda Ferran , Jáuregui Olga , Casals Isidre , Andrés‐Lacueva Cristina , Izquierdo‐Pulido Maria , Lamuela‐Raventós Rosa M.
Published: January 2003

Liquid chromatography coupled with ionspray mass spectrometry in the tandem mode (LC/MS/MS) with negative ion detection was used for the identification of a variety of phenolic compounds in a cocoa sample. Gradient elution with water and acetonitrile, both containing 0.1% HCOOH, was used. Standard solutions of 31 phenolic compounds, including benzoic and cinnamic acids and flavonoid compounds, were studied in the negative ion mode using MS/MS product ion scans. At low collisional activation, the deprotonated molecule [M − H]− was observed for all the compounds studied. For cinnamic and benzoic acids, losses of CO2 or formation of [M − CH3]−· in the case of methoxylated compounds were observed. However, for flavonol and flavone glycosides, the spectra present both the deprotonated molecule [M − H]− of the glycoside and the ion corresponding to the deprotonated aglycone [A − H]−. The latter ion is formed by loss of the rhamnose, glucose, galactose or arabinose residue from the glycosides. Different fragmentation patterns were observed in MS/MS experiments for flavone‐C‐glycosides which showed fragmentation in the sugar part. Fragmentation of aglycones provided characteristic ions for each family of flavonoids. The optimum LC/MS/MS conditions were applied to the characterization of a cocoa sample that had been subjected to an extraction/clean‐up procedure which involved chromatography on Sephadex LH20 and thin‐layer chromatographic monitoring. In addition to compounds described in the literature, such as epicatechin and catechin, quercetin, isoquercitrin (quercetin‐3‐O‐glucoside) and quercetin‐3‐O‐arabinose, other compounds were identified for the first time in cocoa samples, such as hyperoside (quercetin‐3‐O‐galactoside), naringenin, luteolin, apigenin and some O‐glucosides and C‐glucosides of these compounds. Copyright © 2003 John Wiley & Sons, Ltd.
Clustering of nucleosides in the presence of alkali metals: Biologically relevant quartets of guanosine, deoxyguanosine and uridine observed by ESI‐MS/MS
Authors: Aggerholm Tenna , Nanita Sergio C. , Koch Kim J. , Cooks R. Graham
Published: January 2003

Electrospray ionization (ESI) mass spectra of nucleosides, recorded in the presence of alkali metals, display alkali metal ion‐bound quartets and other clusters that may have implications for understanding non‐covalent interactions in DNA and RNA. The tetramers of guanosine and deoxyguanosine and also their metaclusters (clusters of clusters), cationized by alkali metals, were observed as unusually abundant magic number clusters. The observation of these species in the gas phase parallels previous condensed‐phase studies, which show that guanine derivatives can form quartets and metaclusters of quartets in solution in the presence of metal cations. This parallel behavior and also internal evidence suggest that bonding in the guanosine tetramers involves the bases rather than the sugar units. The nucleobases thymine and uracil are known to form magic number pentameric adducts with K+, Cs+ and NH4+ in the gas phase. In sharp contrast, we now show that the nucleosides uridine and deoxythymidine do not form the pentameric clusters characteristic of the corresponding bases. More subtle effects of the sugars are evident in the fact that adenosine and cytidine form numerous higher order clusters with alkali metals, whereas deoxyadenosine and deoxycytidine show no clustering. It is suggested that hydrogen bonding between the bases in the tetramers of dG and rG are the dominant interactions in the clusters, hence changing the ribose group to deoxyribose (and vice versa) generally has little effect. However, the additional hydroxyl group of RNA nucleosides enhances the non‐selective formation of higher‐order aggregates for adenosine and cytidine and results in the lack of highly stable magic number clusters. Some clusters are the result of aggregation in the course of ionization (ESI) whereas others appear to be intrinsic to the solution being examined. Copyright © 2003 John Wiley & Sons, Ltd.
Liquid chromatographic/tandem mass spectrometric method for the determination of the tobacco‐specific nitrosamine metabolite NNAL in smokers' urine
Authors: Byrd Gary D. , Ogden Michael W.
Published: January 2003

A specific and rapid liquid chromatographic/tandem mass spectrometric (LC/MS/MS) method was developed and validated for NNAL, a metabolite of the tobacco‐specific nitrosamine metabolite NNK. The metabolite was detected in smokers' urine with a limit of quantitation (LOQ) of 20 pg ml−1 and a linear range up to 1000 pg ml−1. The method features a single solid‐phase extraction step and MS/MS monitoring following electrospray ionization. Fragmentation pathways for the protonated molecular ion are proposed. The sample preparation is simpler than that for gas chromatographic methods reported in the literature and maintains sensitivity adequate for determining NNAL in smokers' urine. By using enzyme hydrolysis to determine total NNAL in urine, the amount of NNAL‐glucuronide was calculated. A standard pooled smokers' urine sample used for development gave values of 176 ± 8 pg ml−1 free NNAL and 675 ± 26 pg ml−1 total NNAL following enzyme hydrolysis. The method was applied to a group of seven smokers; the free NNAL level for the group was 101–256 pg ml−1 with NNAL‐glucuronides at 247–566 pg ml−1. The ratio of conjugated to free NNAL was in the range 0.98–2.95. The variability in total daily amount of NNAL excreted (ng per 24 h) had RSDs of 6–21% for free NNAL, 7–22% for conjugated NNAL and 6–20% for total NNAL excreted. When normalized to the number of cigarettes smoked, the amounts of NNAL excreted per cigarette smoked were in the range of amounts of NNK yields reported for cigarettes in the literature. Copyright © 2003 John Wiley & Sons, Ltd.
Human and simian immunodeficiency virus‐infected chimpanzees do not have increased intracellular levels of β‐chemokines in contrast to infected humans
Authors: Ondoa Pascale , Vereecken Chris , Fransen Katrien , Colebunders Robert , van der Groen Guido , Heeney Jonathan L. , Kestens Luc
Published: January 2003

This study was undertaken to explain why chimpanzees infected with HIV‐1 (human immunodeficiency virus type 1) or SIVcpz (simian immunodeficiency virus of chimpanzee) are relatively resistant to AIDS (acquired immunodeficiency syndrome). The numbers of β‐chemokine‐positive cells were compared between uninfected and infected humans and chimpanzees using three‐color cytofluorometry. In humans, the percentage of β‐chemokine‐positive cells was significantly higher in CD8+ T and natural killer (NK) cells than in CD4+ T cells in both uninfected and HIV‐1‐infected individuals. In the presence of HIV‐1 infection, however, both CD8+ and CD4+ T cell subsets contained significantly more β‐chemokine‐positive cells than in the absence of infection. Interestingly, in chimpanzees two important differences were noted. First, their percentage of β‐chemokine‐positive CD8+ T and NK cells was significantly higher than in uninfected humans. Second, in contrast to humans, infection with either HIV‐1 or with SIVcpz was not associated with increased numbers of β‐chemokine‐positive cells. These results indicate that: constitutive high levels of intracellular β‐chemokines in chimpanzees' CD8 lymphocytes and NK cells do not necessarily correspond to lower levels of virus replication during the chronic phase of infection; and increased percentages of β‐chemokine‐positive cells in HIV‐infection are not a correlate of disease resistance. The data suggest that neither pre‐nor post‐exposure levels of intracellular β‐chemokines are correlated with the subsequent control of disease progression. J. Med. Virol. 69:297–305, 2003. © 2003 Wiley‐Liss, Inc.
Hepatitis B virus genotypes and serotypes in western India: Lack of clinical significance
Authors: Gandhe Swati S. , Chadha Mandeep S. , Arankalle Vidya A.
Published: January 2003

To determine hepatitis B virus genotype and subtype distribution among HBV infected individuals with different clinical manifestations in western India, serum samples from 19 asymptomatic hepatitis B surface antigen carriers, 30 chronic hepatitis B patients, 8 acute hepatitis B patients, 5 fulminant hepatitis B patients, and with circulating HBV DNA were genotyped and subtyped on the basis of the nucleotide sequence analysis of S region of the HBV genome. Genotype D was the predominant genotype circulating in western India (57/62; 91.93%). All 19 asymptomatic hepatitis B surface antigen carriers, 8 acute hepatitis B patients, 5 fulminant hepatic failure patients and 25/30 chronic hepatitis B patients were circulating genotype D and ayw3/ayw2 subtypes. HBV genotype A was prevalent in 8% (5/62) of the total number of patients and all belonged to chronic hepatitis B category. Subtyping analysis showed that all genotype A isolates were of subtype adw2. As most of the patients from different clinical categories were infected with HBV genotype D, it is concluded that this genotype did not influence the outcome of HBV infection. J. Med. Virol. 69:324–330, 2003. © 2003 Wiley‐Liss, Inc.
Changes in hypervariable region 1 of the envelope 2 glycoprotein of hepatitis C virus in children and adults with humoral immune defects
Authors: Gaud Umesh , Langer Barbara , Petropoulou Theoni , Thomas Howard C. , Karayiannis Peter
Published: January 2003

The N‐terminal end of the hepatitis C virus (HCV) envelope glycoprotein E2 contains a stretch of 27 amino acids that exhibit increased variability. This hypervariable region 1 (HVR‐1), as it is normally referred to, is thought to contain epitopes that come under humoral immune attack. In the present study, 10 patients (5 children and 5 adults) with humoral immune defects and chronic HCV infection were investigated, to see how HVR‐1 sequences behave over time in these patients who are unable to produce antibodies. Amplicons of this region showed little or no variation at all over time, indicating that quasispecies variation in this region is driven by the host's humoral immune response. J. Med. Virol. 69:350–356, 2003. © 2003 Wiley‐Liss, Inc.
Processing of hepatitis C virus core protein is regulated by its C‐terminal sequence
Authors: Kato Takanobu , Miyamoto Michiko , Furusaka Akihiro , Date Tomoko , Yasui Kotaro , Kato Junko , Matsushima Shozo , Komatsu Tatsuji , Wakita Takaji
Published: January 2003

Polyprotein processing of plus‐strand RNA viruses is important in the regulation of gene production and replication. The core protein of hepatitis C virus (HCV), constructing the viral particle, is processed from its precursor polyprotein and observed as two forms, p23 and p21. Production of p21 by cleavage at the C‐terminus of p23 is considered crucial to viral assembly and replication. In this study, this processing step was compared between clones isolated from two patients with fulminant hepatitis and from five patients with chronic hepatitis by an in vitro translation assay and cell transfection assay. The p21 core protein was predominant from the clone isolated from one of the fulminant hepatitis patient (p21 core protein production was 65.98%), while p23 was abundant with clones from five chronic hepatitis patients (p21 core protein production was 7.11±1.62%) and clone from another fulminant hepatitis patient (p21 core protein production was 13.36%). Investigations with chimeric and mutation‐introduced constructs revealed that four amino acid residues in the C‐terminus of the core region are responsible for this difference. The data suggest that core protein processing is regulated by C‐terminus mutations. J. Med. Virol. 69:357–366, 2003. © 2003 Wiley‐Liss, Inc.
Quantitation of cytomegalovirus (CMV) DNA by real‐time PCR for occurrence of CMV disease in HIV‐infected patients receiving highly active antiretroviral therapy
Authors: Gourlain Karine , Salmon Dominique , Gault Elyanne , Leport Catherine , Katlama Christine , Matheron Sophie , Costagliola Dominique , Mazeron Marie‐Christine , Fillet Anne‐Marie
Published: January 2003

In HIV‐infected patients treated with highly active antiretroviral therapy (HAART) included in the Predivir cohort, we have evaluated the usefulness of CMV DNA quantitation by a TaqMan® PCR assay from peripheral blood leukocytes (PBLs) to predict CMV disease occurrence. In parallel with the immune restoration after treatment by HAART, the percentage of positive samples decreased progressively from 7.3% at Day 0 to 3.5% at Month 12. Among the CMV markers, the smallest concordance with PBL CMV TaqMan® PCR, as evaluated by kappa, was observed with pp65 antigenemia, whereas concordance with all other CMV markers was high. Among the 16 patients with CMV DNA copies at least once >100/150,000 cells, CMV disease occurred in six during follow‐up, whereas among the 159 patients with CMV DNA copies always <10/150,000 cells, CMV disease occurred in three and among the seven patients with CMV DNA copies >10 and <100 occurred in only one. In univariate Cox models, all the CMV markers including PBL CMV TaqMan® PCR >10/150,000 cells (RR: 27.6, IC95: 7.1–107.2), the CD4 cell count <75 cells/mm3 and the HIV viral load >100,000 copies/ml were predictive for CMV disease. In a stepwise multivariate analysis, which should be interpreted with caution due to the small number of events (n = 10), three covariates were associated independently with CMV disease: pp65 antigenemia >100 nuclei/200,000, PBL CMV TaqMan® PCR >10 copies/150,000 cells and HIV viral load remaining or increasing >100,000 copies/ml. J. Med. Virol. 69:401–407, 2003. © 2003 Wiley‐Liss, Inc.
Induction of cellular immunity to varicella‐zoster virus glycoproteins tested with pernasal coadministration of escherichia coli enterotoxin in mice
Authors: Tsuji Takao , Shiraki Kimiyasu , Sato Hitoshi , Sasaki Keiko , Arita Michiko , Kato Michio , Takahashi Tsuyoshi , Ochi Sadayuki , Ichinose Yoshio , Yokochi Takashi , Asano Yoshizo
Published: January 2003

A mutant of Escherichia coli enterotoxin promotes the induction of cellular immunity to a live varicella vaccine (the Oka strain) as a mucosal adjuvant in mice. An investigation was carried out to determine which of the purified glycoproteins of the virus among three induced cellular immunity with a single nasal administration. Spleen cells from mice immunized nasally with the vaccine and toxin produced interleukin‐2 (IL‐2) at the same level on restimulation in vitro with glycoprotein H: glycoprotein L (gH:gL), gB, and gE:gI, but not IL‐4. The spleen cells from mice immunized with gH:gL, gB, or gE:gI and toxin produced IL‐2 on restimulation with gH:gL, gB, or gE:gI, respectively, and the vaccine, but not IL‐4. Immunization with gH:gL and the toxin showed increased thymidine uptake and production of IL‐2 and interferon‐γ (IFN‐γ) of the spleen cells, but not IL‐4, depending on the dose of gH:gL used for immunization and restimulation in vitro. Purified gE:gI and gB have been reported to be the strongest stimulators of cellular immunity to varicella upon subcutaneous injection and are useful as a subunit vaccine. All the glycoproteins tested are excellent stimulators of cellular immunity to the virus and itself on nasal co‐immunization with the toxin. J. Med. Virol. 69:451–458, 2003. © 2003 Wiley‐Liss, Inc.
Apolipoprotein E expression after spinal cord injury in the mouse
Authors: Seitz Alexander , Kragol Maja , Aglow Elsa , Showe Louise , Heber‐Katz Ellen
Published: January 2003

Apolipoprotein E (apo‐E), a protein involved in lipid metabolism and cholesterol transport, has been found to be up‐regulated in CNS injury and is associated with Alzheimer's disease in humans. In this study, we show that apo‐E is also up‐regulated after complete spinal cord transection in the C57BL/6 mouse. In the uninjured cord, the cellular localization of apo‐E protein is in astrocytes, in individual neurons throughout the laminae except for the dorsal horn, and in endothelial cells of capillaries in the immediate vicinity of those neurons. After injury, RNA levels are elevated as early as 4 days and reach a maximal level between 1 and 2 weeks. Protein levels follow closely but remain up‐regulated beyond 3 weeks. Early on, the protein can be found in neutrophils and macrophages at the injury site and only at later times in astrocytes during the remodeling of white matter tracts, most prominently in degenerating parts of the fasciculus gracilis. © 2002 Wiley‐Liss, Inc.