bluuebunny/crossref_metadata_2025 huggingface dataset:
Quail as the chimeric counterpart of the chicken: Morphology and ontogeny of the bursa of Fabricius
Authors: Nagy Nándor , Magyar Attila , Tóth Miklós , Oláh Imre
Published: February 2004

The quail is the chimeric and parabiotic counterpart of the chicken, thus increasing the value of quail in the field of developmental biology. Quail bursa of Fabricius was studied by light microscopy, electron microscopy, and immunocytochemical methods. The basic cellular composition and structural framework are comparable with those of the chicken bursa. One of the major structural differences is the absence of the continuous cortico‐medullary arch. In addition to the epithelial reticular cell the bursal secretory dendritic cell is the other medullary‐specific bursal cell. The bursal secretory dendritic cell is a highly elongated cell which expresses vimentin intermediate filaments and produces secretory granules. The substance of the granules can be visualized by NIC2 monoclonal antibody, which was produced against guinea fowl bursal secretory dendritic cell. The released granular content appears on the lateral surface of the bursal secretory dendritic cell and is gradually solubilized. Thus, the NIC2‐positive substance may occur in membrane‐bound and solubilized forms in the isolated environment of the medulla. The bursal secretory dendritic cell establishes membrane contact areas with the B cells; therefore, they may influence B‐cell maturation by cell contact and chemical (humoral) product. During embryogenesis bursal secretory dendritic cell precursors enter the epithelium and 1) induce epithelial bud formation, and 2) produce an NIC2‐positive substance. Senescent bursal secretory dendritic cells can be phagocytic and migrate into the follicle‐associated epithelium. This physiological turnover of the bursal secretory dendritic cell represents a novel pathway of macrophage formation from dendritic cells. J. Morphol. 259:328–339, 2004. © 2004 Wiley‐Liss, Inc.
Larval development of Phoronis pallida (Phoronida): Implications for morphological convergence and divergence among larval body plans
Authors: Santagata Scott
Published: February 2004

Morphological variation among larval body plans must be placed into a phylogenetic and ecological context to assess whether similar morphologies are the result of phylogenetic constraints or convergent selective pressures. Investigations are needed of the diverse larval forms within the Lophotrochozoa, especially the larvae of phoronids and brachiopods. The actinotroch larva of Phoronis pallida (Phoronida) was reared in the laboratory to metamorphic competence. Larval development and growth were followed with video microscopy, SEM, and confocal microscopy. Early developmental features were similar to other phoronid species. Gastrulation was accomplished by embolic invagination of the vegetal hemisphere. Mesenchymal cells were found in the remaining blastocoelic space after invagination began. Mesenchymal cells formed the body wall musculature during the differentiation of larval features. Body wall musculature served as the framework from which all other larval muscles proliferated. Larval growth correlated best with developmental stage rather than age. Consistent with other phoronid species, differentiation of juvenile tissues occurred most rapidly at the latest stages of larval development. The minimum precompetency period of P. pallida was estimated to be ∼4–6 weeks. Previously published studies have documented that the planktonic embryos of P. pallida develop faster than the brooded embryos of P. vancouverensis. However, these data showed that the difference in developmental rate between the two species decreased in succeeding larval stages. There may be convergent selective pressures that result in similar timing to metamorphic competence among phoronid and brachiopod planktotrophic larval types. Morphological differences between these larval types result from heterochronic developmental shifts in the differentiation of juvenile tissue. Similarities in the larval morphology of phoronids and basal deuterostomes are likely the result of functional and developmental constraints rather than a shared (recent) evolutionary origin. These constraints are imposed by the functional design of embryological stages, feeding structures, and swimming structures. J. Morphol. 259:347–358, 2004. © 2004 Wiley‐Liss, Inc.
Comparative electron microscopic study of the stomach of Orchestia cavimana and Arcitalitrus sylvaticus (crustacea: Amphipoda)
Authors: Štrus Jasna , Storch Volker
Published: February 2004

The functional morphology of stomachs of the European semiterrestrial amphipod Orchestia cavimana and of the Australian terrestrial species Arcitalitrus sylvaticus was studied by electron microscopy. The stomach of the two amphipod species is divided longitudinally into a spacious dorsal food channel and two ventral filtration channels. Additionally, a prominent helically oriented circulation channel is situated on each lateral side of the stomach, forming a semicircular channel separated from the food channel by spines. The food channel conveys coarse food particles directly into the midgut through a funnel. The filtration channels receive fine material filtered through primary and secondary filters. Material forced through the secondary filters by the pressure of the laterally located inferolateralia eventually reaches the openings of the midgut glands. Washing of filters and soaking of ingested food items with enzymes probably is achieved by a forward stream of digestive juice from the midgut glands and conveyed through the circulatory channels. The specializations of the stomach of the two species of Amphipoda investigated are described and compared to the pertinent structures of Mysidacea and Isopoda. J. Morphol. 259:340–346, 2004. © 2004 Wiley‐Liss, Inc.
A comparison between the accuracy of voxel‐based morphometry and hippocampal volumetry in Alzheimer's disease
Authors: Testa Cristina , Laakso Mikko P. , Sabattoli Francesca , Rossi Roberta , Beltramello Alberto , Soininen Hilkka , Frisoni Giovanni B.
Published: February 2004

Purpose

To compare the accuracy of voxel‐based morphometry (VBM) and region of interest (ROI)‐based hippocampal volumetry to detect medial temporal lobe atrophy in Alzheimer's disease (AD).

Materials and Methods

A total of 27 AD patients (age 74 ± 9 years; 22 women; Mini‐Mental State Exam [MMSE] 21 ± 4) and 25 controls (age 70 ± 8; 16 women; MMSE 29 ± 1) were studied. Accuracy of VBM to detect gray matter loss in those seven AD patients and 11 controls with similar ROI‐based hippocampal measures and of ROI‐based volumetry to detect gray matter loss in those four AD patients and five controls with similar VBM‐based hippocampal measures was assessed. VBM was performed with statistical parametric mapping (SPM99).

Results

The area under the curve was 0.96 (95% C.I., 0.92–1.00) for VBM, 0.89 (95% C.I., 0.80–0.98) for ROI‐based hippocampal measures, and 0.99 (95% C.I., 0.96–1.00) for both. In subjects with similar ROI‐based hippocampal measures, VBM detected atrophy in AD patients at P < 0.0001, while in subjects with similar VBM‐based hippocampal measure, volumetry was not significant (P = 0.11). Both measures independently contributed to discrimination (P = 0.004 and P = 0.032) in a logistic regression model.

Conclusion

These results indicate that VBM is more accurate, but the combination of both methods provides the highest accuracy for detection of hippocampal atrophy in AD. J. Magn. Reson. Imaging 2004;19:274–282. © 2004 Wiley‐Liss, Inc.
Gadopentetate dimeglumine‐enhanced three‐dimensional MR‐angiography: Dosing, safety, and efficacy
Authors: Goyen Mathias , Debatin Jörg F.
Published: February 2004

Noninvasiveness, inherent three‐dimensionality allowing reformations in any desired plane, and safe contrast agents, coupled with high diagnostic accuracy have driven the rise in popularity of contrast‐enhanced MR angiography (CE‐MRA) within the medical community. Reflecting its dominant market share as a paramagnetic contrast agent, gadopentetate dimeglumine (Gd‐DTPA) has been used for the majority of clinically‐performed MRA exams. Over the period January 1994 to February 2002, a total of 172 original studies describing the use of gadolinium‐enhanced MRA in more than three human subjects were identified. Of these, 117 described the use of Gd‐DTPA as the contrast agent for MRA. A total of 4046 subjects who received Gd‐DTPA for MRA are described in these studies. Analysis of these data demonstrate Gd‐DTPA to be a safe contrast agent for MRA when applied in a dose ranging from 0.1 to 0.3 mmol/kg of bodyweight. The documented clinical results show Gd‐DTPA to be efficacious in the assessment of the arterial system. The effectiveness of Gd‐DTPA‐enhanced MRA extends beyond the detection, localization, and characterization of arterial disease, and encompasses choice and planning of appropriate therapy, as well as evaluation of therapeutic effectiveness. J. Magn. Reson. Imaging 2004;19:261–273. © 2004 Wiley‐Liss, Inc.
Limits of 8‐Tesla magnetic resonance imaging spatial resolution of the deoxygenated cerebral microvasculature
Authors: Dashner Roger A. , Kangarlu Allahyar , Clark David L. , RayChaudhury Abhik , Chakeres Donald W.
Published: February 2004

Purpose

To quantify the minimum magnetic resonance imaging (MRI) spatial resolution of the visible deoxygenated microscopic vessels of the human brain at 8 T.

Materials and Methods

This study compared 8‐T gradient echo (GE) images of a human cadaver brain having an in‐plane resolution of 195 × 195 μm to corresponding digital photographs of 205 cryomicrotome sections of the same cadaver brain, along with summed images of 25 contiguous cryomicrotome sections. One‐millimeter‐thick GE images of a 1‐cm‐thick unfixed whole coronal brain section were acquired using techniques similar to those commonly utilized for 8‐T human imaging in vivo.

Results

There was excellent MR visualization of the deoxygenated microscopic vessels within the brain down to a resolution of approximately 100 μm.

Conclusion

By taking advantage of magnetic susceptibility‐based blood oxygenation level‐dependent (BOLD) contrast, deoxygenated microscopic blood vessels smaller than the pixel dimensions used for imaging can be visualized using a whole‐body 8‐T MRI system. J. Magn. Reson. Imaging 2004;19:303–307. © 2004 Wiley‐Liss, Inc.
Adrenal lesions with heterogeneous suppression on chemical shift imaging: Clinical implications
Authors: Gabriel Helena , Pizzitola Victor , McComb Erin N. , Wiley Elizabeth , Miller Frank H.
Published: February 2004

Purpose

To investigate the frequency and significance of adrenal lesions that demonstrate heterogeneous suppression on chemical shift magnetic resonance imaging (MRI).

Materials and Methods

A retrospective search of adrenal lesions identified on MR from November 1997–July 2001 was performed. The adrenal lesions were classified as having suppression typical for an adenoma, nonsuppression, or atypical heterogeneous suppression. Lesions with heterogeneous suppression were further reviewed. Follow‐up for the heterogeneous lesions consisted of imaging studies or pathology from surgical excision.

Results

Adrenal lesions were identified in 242 patients. Heterogeneously suppressing lesions were seen in 34 (14%) patients. Several different patterns of heterogeneous suppression were identified. Imaging or pathologic follow‐up was available for 18 of the heterogeneously suppressing lesions (one patient had both). Fifteen patients with follow‐up imaging showed stability of the lesion over a significant interval, suggesting a benign lesion. Pathology was available for four patients revealing two patients with adenomas and two patients with nodular hyperplasia. Thus, all 18 patients with heterogeneous suppression had a benign lesion.

Conclusion

Adrenal lesions that have a heterogeneous pattern of suppression on out‐of‐phase chemical shift MR images are a common finding. Our small series suggest that these lesions are probably benign. J. Magn. Reson. Imaging 2004;19:308–316. © 2004 Wiley‐Liss, Inc.
Comparison of three methods for generating group statistical inferences from independent component analysis of functional magnetic resonance imaging data
Authors: Schmithorst Vincent J. , Holland Scott K.
Published: February 2004

Purpose

To evaluate the relative effectiveness of three previously proposed methods of performing group independent component analysis (ICA) of functional magnetic resonance imaging (fMRI) data.

Materials and Methods

Data were generated via computer simulation. Components were added to a varying number of subjects between 1 and 20, and intersubject variability was simulated for both the added sources and their associated time courses. Three methods of group ICA analyses were performed: across‐subject averaging, subject‐wise concatenation, and row‐wise concatenation (e.g., across time courses).

Results

Concatenating across subjects provided the best overall performance in terms of accurate estimation of the sources and associated time courses. Averaging across subjects provided accurate estimation (R > 0.9) of the time courses when the sources were present in a sufficient fraction (about 15%) of 100 subjects. Concatenating across time courses was shown not to be a feasible method when unique sources were added to the data from each subject, simulating the effects of motion and susceptibility artifacts.

Conclusion

Subject‐wise concatenation should be used when computationally feasible. For studies involving a large number of subjects, across‐subject averaging provides an acceptable alternative and reduces the computational load. J. Magn. Reson. Imaging 2004;19:365–368. © 2004 Wiley‐Liss, Inc.
Comparison of conventional fast spin echo, single‐shot two‐dimensional and three‐dimensional half‐fourier RARE for T2‐weighted female pelvic imaging
Authors: Sugimura Hiroshi , Yamaguchi Kenichiro , Furukoji Eiji , Tamura Shozo , Sakae Tatefumi , Koga Haruyuki , Machida Yoshio , Kitane Shinichi
Published: February 2004

Purpose

To evaluate the usefulness of the three‐dimensional half‐Fourier RARE sequence in comparison with single‐shot two‐dimensional half‐Fourier RARE and conventional fast spin echo (FSE) for female pelvic imaging.

Materials and Methods

Imaging with all sequences was performed in 146 patients with 166 focal lesions on a 1.5‐T system. The images were compared on the basis of quality, lesion conspicuity, and lesion to the uterus contrast‐to‐noise ratio (CNR).

Results

The sharpness of intrapelvic organs on the three‐dimensional half‐Fourier RARE sequence was better than that on two‐dimensional half‐Fourier RARE and worse than that on FSE. Motion‐related artifacts for three‐dimensional half‐Fourier RARE were more frequent than those for two‐dimensional half‐Fourier RARE. There was no statistical difference between the three‐dimensional half‐Fourier RARE sequence and FSE in regard to lesion conspicuity and overall image quality. The CNR of leiomyoma to myometrium and cervical cancer to cervical stroma was the highest with three‐dimensional half‐Fourier RARE (P< 0.05).

Conclusion

The three‐dimensional half‐Fourier RARE sequence generates images with higher contrast and better image resolution than two‐dimensional‐RARE. The three‐dimensional data set provided images that can be observed in any orientation without acquiring an additional scan by using the multiplanar reconstruction (MPR) method. J. Magn. Reson. Imaging 2004;19:349–355. © 2004 Wiley‐Liss, Inc.
1H magnetic resonance spectroscopy of preinvasive and invasive cervical cancer: In vivo–ex vivo profiles and effect of tumor load
Authors: Mahon Marrita M. , Cox I. Jane , Dina Roberto , Soutter FRCOG W. Patrick , McIndoe MRCOG G. Angus , Williams Andreanna D. , deSouza Nandita M.
Published: February 2004

Purpose

To compare in vivo 1H magnetic resonance (MR) spectra of preinvasive and invasive cervical lesions with ex vivo magic angle spinning (MAS) spectra of intact biopsies from the same subjects and to establish the effects of tumor load in the tissue sampled on the findings.

Materials and Methods

A total of 51 subjects (nine with normal cervix, 10 with cervical intraepithelial neoplasia [CIN], and 32 with cervical cancer) underwent endovaginal MR at 1.5 T. Single‐voxel (3.4 cm3) 1H MR spectra were acquired and voxel tumor load was calculated (tumor volume within voxel as a percentage of voxel volume). Resonances from triglycerides –CH2 and –CH3 and choline‐containing compounds (Cho) were correlated with voxel tumor load. Biopsies analyzed by 1H MAS‐MR spectroscopy (MRS) had metabolite levels correlated with tumor load in the sample at histology.

Results

In vivo studies detected Cho in normal, CIN, and cancer patients with no significant differences in levels (P = 0.93); levels were independent of voxel tumor load. Triglyceride –CH2 and –CH3 signals in‐phase with Cho were present in 77% and 29%, respectively, of cancer subjects (but not in normal women or those with CIN), but did not correlate with voxel tumor load. Ex vivo cancer biopsies showed levels of triglycerides –CH2 and –CH3 and of Cho that were significantly greater than in normal or CIN biopsies (P < 0.05); levels were independent of the tumor load in the sample. The presence of –CH2 in vivo predicted the presence of cancer with a sensitivity and specificity of 77.4% and 93.8% respectively, positive (PPV) and negative (NPV) predictive values were 96% and 68.2%; for –CH2 ex vivo, sensitivity was 100%; specificity, 69%; PPV, 82%; and NPV, 100%.

Conclusion

Elevated lipid levels are detected by MRS in vivo and ex vivo in cervical cancer and are independent of tumor load in the volume of tissue sampled. J. Magn. Reson. Imaging 2004;19:356–364. © 2004 Wiley‐Liss, Inc.
Usefulness of contrast kinetics for predicting and monitoring tissue changes in muscle following thermal therapy in long survival studies
Authors: Cheng Hai‐Ling Margaret , Purcell Carrie M. , Bilbao Juan M. , Plewes Donald B.
Published: February 2004

Purpose

To investigate Gd‐DTPA kinetics as indicators of subacute and subchronic histopathological changes following focused ultrasound (FUS) thermal therapy for improved evaluation.

Materials and Methods

A total of 18 FUS lesions were created in the thigh muscle of five rabbits under magnetic resonance (MR) guidance at 1.5 Tesla. The rabbits were killed at different times: 40 hours, three days, and seven days. All lesions were analyzed histologically. An analysis of the uptake kinetics of Gd‐DTPA, injected within two hours postheating and before sacrifice, was performed. The resulting kinetic maps, permeability (Ktrans) and leakage space (ve), were correlated to T2‐weighted MR and histology.

Results

Images of Ktrans and ve better differentiate subacute and subchronic changes not visible on conventional MR in the days following therapy and are consistent with the histopathology observed. In particular, the border between nonviable and viable tissue is well demarcated. The extent of damage is best indicated on ve, whereas the borders of inflammation are shown on Ktrans. The total lesion extent is relatively stable over the 7 days posttherapy and can be predicted by ve or T2‐weighted MR at early times after heating.

Conclusion

Our results suggest that Gd‐DTPA kinetics can complement conventional MR for improved evaluation of FUS thermal therapy by providing finer differentiation of necrotic states, inflammation, and repair processes. J. Magn. Reson. Imaging 2004;19:329–341. © 2004 Wiley‐Liss, Inc.
Cardiac magnetic resonance parallel imaging at 3.0 Tesla: Technical feasibility and advantages
Authors: McGee Kiaran P. , Debbins Josef P. , Boskamp Ed B. , Blawat LeRoy , Angelos Lisa , King Kevin F.
Published: February 2004

Purpose

To quantify changes in signal‐to‐noise ratio (SNR), contrast‐to‐noise ratio (CNR), specific absorption rate (SAR), RF power deposition, and imaging time in cardiac magnetic resonance imaging with and without the application of parallel imaging at 1.5 T and 3.0 T.

Materials and Methods

Phantom and volunteer data were acquired at 1.5 T and 3.0 T with and without parallel imaging.

Results

Doubling field strength increased phantom SNR by a factor of 1.83. In volunteer data, SNR and CNR values increased by factors of 1.86 and 1.35, respectively. Parallel imaging (reduction factor = 2) decreased phantom SNR by a factor of 1.84 and 2.07 when compared to the full acquisition at 1.5 T and 3.0 T, respectively. In volunteers, SNR and CNR decreased by factors of 2.65 and 2.05 at 1.5 T and 1.99 and 1.75 at 3.0 T, respectively. Doubling the field strength produces a nine‐fold increase in SAR (0.0751 to 0.674 W/kg). Parallel imaging reduced the total RF power deposition by a factor of two at both field strengths.

Conclusions

Parallel imaging decreases total scan time at the expense of SNR and CNR. These losses are compensated at higher field strengths. Parallel imaging is effective at reducing total power deposition by reducing total scan time. J. Magn. Reson. Imaging 2004;19:291–297. © 2004 Wiley‐Liss, Inc.
Characterization of α‐ and γ‐glutamyl dipeptides by negative ion collision‐induced dissociation
Authors: Harrison Alex G.
Published: February 2004

The low‐energy CID mass spectra of the [M − H]− ions of a variety of dipeptides containing glutamic acid have been obtained using cone‐voltage collisional activation. Dipeptides with the γ‐linkage, H‐Glu(Xxx‐OH)‐OH, are readily distinguished from those with the α‐linkage, H‐Glu‐Xxx‐OH, by the much more prominent elimination of H‐Xxx‐OH from the [M − H]− ions of the former isomers, resulting in formation of m/z 128, presumably deprotonated pyroglutamic acid. Dipeptides with the reverse linkage, H‐Xxx‐Glu‐OH, show distinctive fragmentation reactions of the [M − H]− ions including enhanced elimination of CO2 and formation of deprotonated glutamic acid. Exchange of the labile hydrogens for deuterium has shown that there is considerable interchange of C‐bonded hydrogens with labile (N‐ and O‐bonded) hydrogens prior to most fragmentation reactions. All dipeptides show loss of H2O from [M − H]−. MS3 studies show that the [M − H − H2O]− ion derived from H‐Glu‐Gly‐OH has the structure of deprotonated pyroglutamylglycine while the [M − H − H2O]− ions derived from H‐Glu(Gly‐OH)‐OH and H‐Gly‐Glu‐OH show a different fragmentation behaviour indicating distinct structures for the fragment ions. Copyright © 2004 John Wiley & Sons, Ltd.
Rearrangement process occurring in the fragmentation of adefovir derivatives
Authors: Chen Xiaoyan , Xing Jie , Zhong Dafang
Published: February 2004

The fragmentation of the antiviral drug adefovir dipivoxil and its two active metabolites, adefovir and monopivoxil adefovir, was investigated using both ion trap and triple‐quadrupole mass spectrometers. Fragment ions due to loss of 30 Da were observed and attributed to an unanticipated rearrangement process by loss of formaldehyde. The proposed mechanism is supported with the aid of three newly synthesized adefovir derivatives and with accurate mass measurement. Other fragmentations by loss of a pivaloyl group, loss of water, C—P bond cleavage and C—O bond cleavage were also observed for adefovir derivatives. It was concluded that the compounds containing a >POO—CHR—OCO— group generally displayed a rearrangement reaction by loss of RCHO in collision‐induced dissociation, and the process generally required an activation energy lower than for a direct bond cleavage. Copyright © 2004 John Wiley & Sons, Ltd.
Mass spectra of N‐substituted cantharidinimides
Authors: Lin Pen‐Y. , Lin Shaw‐T. , Liang David W.‐M.
Published: February 2004

The mass spectra of a series of N‐substituted cantharidinimides were examined. The feature of this series compounds is a sequential double hydrogen transfer from the oxabicycloheptane unit to either the carbonyl group of the succinimide unit or the nitrogen atom of the pyridyl or thiazolyl substituent through space. The ability of the N‐substituent to accept a hydrogen atom possibly leads to the different fragmentation pathway. Copyright © 2004 John Wiley & Sons, Ltd.
Analysis of native amino acid and peptide enantiomers by high‐performance liquid chromatography/atmospheric pressure chemical ionization mass spectrometry
Authors: Desai Meera J. , Armstrong Daniel W.
Published: February 2004

High‐performance liquid chromatography (HPLC) coupled to atmospheric pressure chemical ionization (APCI) mass spectrometry was used for the separation and detection of amino acid and peptide enantiomers. With detection limits as low as 250 pg, 25 amino acids enantiomers were baseline resolved on a Chirobiotic T chiral stationary phase. APCI demonstrated an order of magnitude better sensitivity over electrospray ionization (ESI) for free amino acids and low molecular mass peptides at the high LC flow‐rates necessary for rapid analysis. As the peptide chain length increased (peptides with Mr ≥ 300 Da), however, ESI proved to be the more ideal atmospheric pressure ionization source. A mobile phase consisting of 1% (w/w) ammonium trifluoroacetate in methanol and 0.1% (w/w) formic acid in water increased the sensitivity of the APCI method significantly. A step gradient was then used to separate simultaneously all 19 native protein amino acid enantiomers in less than 20 min using extracted ion chromatograms. Copyright © 2004 John Wiley & Sons, Ltd.
Sequential exoproteolysis as a structural probe: a cautionary note
Authors: Beynon Robert J.
Published: February 2004

In a recent paper, Villaneuvaet al.(J. Mass Spectrom.2002;37: 974) described the use of exoproteases as probes of higher order structure in proteins. Their model assumes that the proteins are attacked sequentially from either the N‐terminus or the C‐terminus, depending on the type of exoprotease (aminopeptidase or carboxypeptidase) used. The products of this presumed exoproteolysis were then analysed by matrix‐assisted laser desorption/ionization time‐of‐flight mass spectrometry. The pattern of fragments obtained was mapped on to the primary sequence of the protein, and the exoproteolysis was interpreted as comprising a series of fast and slow phases, the rates of the different phases being directly related to the higher order structure of particular segments of the protein. Here, it is shown that this explanation is unlikely, that both kinetic and practical considerations suggest that alternative explanations for the data should be sought, and that exoproteolysis will perhaps not be as valuable as a conformational probe as the authors suggest. Copyright © 2004 John Wiley & Sons, Ltd.
Bromazepam determination in human plasma by high‐performance liquid chromatography coupled to tandem mass spectrometry: a highly sensitive and specific tool for bioequivalence studies
Authors: Laurito Tiago L. , Mendes Gustavo D. , Santagada Vincenzo , Caliendo Giuseppe , de Moraes Maria Elisabete A. , De Nucci Gilberto
Published: February 2004

A rapid, sensitive and specific method to quantify bromazepam in human plasma using diazepam as the internal standard (IS) is described. The analyte and the IS were extracted from plasma by liquid–liquid extraction using diethyl ether–hexane (80 : 20, v/v). The extracts were analyzed by high‐performance liquid chromatography (HPLC) coupled to electrospray tandem mass spectrometry (MS/MS). Chromatography was performed isocratically on a Genesis C18 analytical column (100 × 2.1 mm i.d., film thickness 4 µ m). The method had a chromatographic run time of 5.0 min and a linear calibration curve over the range 5.0–150 ng ml−1 (r2 > 0.9952). The limit of quantification was 5 ng ml−1. This HPLC/MS/MS procedure was used to assess the bioequivalence of two bromazepam 6 mg tablet formulations (bromazepam from Medley SA Indústria Farmacêutica as the test formulation and Lexotan from Produtos Roche Químico e Farmacêutico SA as the reference formulation). A single 6 mg dose of each formulation was administered to 24 healthy volunteers (12 males and 12 females). The study was conducted using an open, randomized, two‐period crossover design with a 3 week washout interval. Since the 90% CI for Cmax, AUClast, AUC0–240 h (linear) and AUC(0–∞) ratios were all inside the 80–125% interval proposed by the US Food and Drug Administration, it was concluded that the bromazepam formulation from Medley is bioequivalent to the Lexotan formulation for both the rate and the extent of absorption. Copyright © 2004 John Wiley & Sons, Ltd.
Improved detection of multi‐phosphorylated peptides in the presence of phosphoric acid in liquid chromatography/mass spectrometry
Authors: Kim Jeongkwon , Camp David G. , Smith Richard D.
Published: February 2004

In contrast to lower phosphorylation states (e.g. the tryptic monophosphopeptide FQpSEEQQQTEDELQDK from bovine β‐casein), the specific detection of multi‐phosphorylated peptides (e.g. the tetraphosphopeptide RELEELNVPGEIVEpSLpSpSpSEESITR from tryptic digestion of bovine β‐casein) has often been problematic for liquid chromatographic mass spectrometric (LC/MS) analysis owing to their high affinity for adsorption to exposed surfaces. We observed an enhancement in the overall detection of phosphopeptides on addition of phosphoric acid (0.1–1.0%) to the sample solution; a 10‐fold increase in sensitivity was determined for the detection of two tryptic phosphopeptides and also a significant improvement in the detection of the tetraphosphopeptide. Using capillary LC with ion trap tandem MS for detection and identification, the achievable detection limits were 50 fmol and 50 pmol for the monophosphopeptide and the tetraphosphopeptide, respectively. Phosphoric acid is believed to act as a blocking agent to available silanol groups on both the silica capillary surface and the C18‐bonded stationary phase silica surface. Copyright © 2004 John Wiley & Sons, Ltd.
Full hydrodynamic simulation of GaAs MESFETs
Authors: Aste Andreas , Vahldieck Rüdiger , Rohner Marcel
Published: February 2004

A finite difference upwind discretization scheme in two dimensions is presented in detail for the transient simulation of the highly coupled non‐linear partial differential equations of the full hydrodynamic model, providing thereby a practical engineering tool for improved charge carrier transport simulations at high electric fields and frequencies. The discretization scheme preserves the conservation and transportive properties of the equations. The hydrodynamic model is able to describe inertia effects which play an increasing role in different fields of micro‐ and optoelectronics, where simplified charge transport models like the drift–diffusion model and the energy balance model are no longer applicable. Results of extensive numerical simulations are shown for a two‐dimensional MESFET device. A comparison of the hydrodynamic model to the commonly used energy balance model is given and the accuracy of the results is discussed. Copyright © 2004 John Wiley & Sons, Ltd.
Partial element equivalent circuit (PEEC) models for on‐chip passives and interconnects
Authors: Verbeek Menno E.
Published: February 2004

The accurate simulation of the electromagnetic behaviour of interconnects and on‐chip inductors is becoming more and more important for chip designs. For this purpose, we investigate the use of the electric field integral equation, which can be used to construct a full‐wave electromagnetic model. To facilitate the combined analysis of the chip's circuit and the electromagnetic behaviour, the electromagnetic model can be formulated as an equivalent circuit. We have conducted some simple experiments to see the method at work and to obtain an idea of the frequencies for which retardation effects are important. Copyright © 2004 John Wiley & Sons, Ltd.
Calcium channel blockers in the treatment of disease
Authors: Elmslie Keith S.
Published: February 2004

Over the last 20 years the combination of patch clamp and molecular biology techniques have resulted in an explosion in our knowledge of the different calcium channel types and their roles in physiology. A crucial component to this advance has been the discovery of specific blockers for different calcium channel types. These blockers have not only permitted researchers to assign specific functions to different channel types, but their specificity allowed them to be used to treat diseases that resulted from enhanced calcium channel function. In some cases, the enhanced calcium channel function is secondary to other problems leading to increased cellular excitability. The specificity of calcium channel blockers, however, has provided a means to treat symptoms of the pathophysiology until more effective treatments become available to address the underlying problems. The goal of this review is to introduce the various types of calcium channels, their primary physiologic roles, drugs that block these channels, and diseases that are currently being treated with these drugs. © 2004 Wiley‐Liss, Inc.
Asymmetric localization of LGN but not AGS3, two homologs of Drosophila pins, in dividing human neural progenitor cells
Authors: Fuja Tannin J. , Schwartz Philip H. , Darcy Dan , Bryant Peter J.
Published: February 2004

Human neural progenitor cells (hNPCs) can be recovered from postmortem human brains and used to study the molecular basis of neurogenesis. Human NPCs are being used to investigate the molecular basis of cell fate determination during stem cell divisions, based on comparison with the Drosophila model system. Drosophila neuroblasts and sensory organ precursors undergo well‐defined asymmetric cell divisions (ACD), under the control of a genetically defined set of apical and basal determinants that are localized tightly and dynamically during division. We show by indirect immunofluorescence, confocal microscopy, and time‐lapse video‐microscopy that LGN and AGS3, two human homologs of the Drosophila ACD determinant Pins, have distinct patterns of localization in hNPCs. When cells are grown under conditions favoring proliferation, LGN is distributed asymmetrically in a cell cycle‐dependent manner; it localizes to one side of the dividing cell and segregates into one of the daughter cells. When the cells are grown under conditions favoring differentiation, LGN accumulates in double foci similar to those containing the mitotic apparatus protein NuMA, and in a pattern shown previously for LGN and NuMA in differentiated cells. AGS3, a slightly more distant Pins homolog than LGN, does not show asymmetric localization in these cells. The progenitor cell marker nestin also localizes asymmetrically in colcemid‐treated hNPCs and colocalizes with LGN. The results suggest that hNPCs undergo ACD and that similar molecular pathways may underlie these divisions in Drosophila and human cells. © 2004 Wiley‐Liss, Inc.
Somatodendritic localization and mRNA association of the splicing regulatory protein Sam68 in the hippocampus and cortex
Authors: Grange Julien , Boyer Véronique , Fabian‐Fine Ruth , Fredj Naïla Ben , Sadoul Rémy , Goldberg Yves
Published: February 2004

The RNA‐binding protein Sam68 has been implicated in the signal‐dependent processing of pre‐mRNA and in the utilization of intron‐containing retroviral mRNAs. Sam68 is predominantly nuclear but exhibits remarkable binding affinity for signalling proteins located at the membrane. We have investigated the subcellular distribution of Sam68 in adult rat cortex and hippocampus. Subcellular fractionation showed that the protein was most abundant in nuclei but also was present at a significant level in the cytosol and membrane fractions, including light and synaptic membranes derived from crude synaptosomes. Sam68 extracted from the synaptosomal fraction cosedimented with polysomes on sucrose gradients. In agreement with these findings, immunohistochemical staining indicated that Sam68 was concentrated in neuronal nuclei but was also detectable in the soma and dendrites. Sam68 immunoreactivity examined at the ultrastructural level was found to associate with dendritic microtubules, endoplasmic reticulum, and free polyribosomes, sometimes close to synapses. A combination of immunoprecipitation and RT‐PCR directly confirmed that Sam68 was bound to polyadenylated mRNA in cortical lysates. The αCaMKII mRNA was identified as one of the coprecipitated transcripts; in contrast, the gephyrin and NR1‐1 mRNAs were not coprecipitated, indicating a certain degree of sequence specificity in the association. In electrophoretic mobility shift assays, recombinant GST‐Sam68 as well as brain‐derived Sam68 bound with high affinity to the αCaMKII 3′ untranslated region. These results suggest that Sam68 may accompany and, conceivably, regulate mature mRNAs during nuclear export, somatodendritic transport, and translation. © 2004 Wiley‐Liss, Inc.
Structure and function of long‐lived olfactory organotypic cultures from postnatal mice
Authors: Josephson E.M. , Yilma S. , Vodyanoy V. , Morrison E.E.
Published: February 2004

The first synapse in the olfactory pathway mediates a significant transfer of information given the restricted association of specific olfactory receptor neurons with specific glomeruli in the olfactory bulb. To understand better how this connection is made and what the functional capacities of the participating cells are, we created a long‐lived culture system composed of olfactory epithelium and olfactory bulb tissues. Using the roller tube method of culturing, we grew epithelium‐bulb cocultures, explanted from 1–4‐day‐old Swiss Webster mice, on Aclar for periods ranging from 18 hr to 68 days. The explants flattened so that in some areas the culture was only a few cells thick, making individual cells distinguishable. From 107 cultures studied, we identified the following cell types by expression of specific markers (oldest culture expressing marker, days in vitro, DIV): olfactory receptor neurons (neural cell adhesion molecule, 42 DIV); mature receptor neurons (olfactory marker protein, 28 DIV); postmitotic olfactory receptor neurons and olfactory bulb neurons (β‐tubulin, 68 DIV); astrocytes (glial fibrillary acidic protein, glutamate/aspartate transporter, 68 DIV); olfactory horizontal basal cells (cytokeratin, 22 DIV). Neuronal processes formed glomeruli in 2–4‐week‐old cultures. We also recorded electro‐olfactography responses to puffs of vapor collected over an odorant mixture containing ethyl butyrate, eugenol, (+) carvone, and (−) carvone from cultures as old as 21 DIV. These features of our olfactory culture system make this model useful for studying properties of immature and mature olfactory receptor neurons, pathfinding strategies of receptor axons, and mechanisms of information transfer in the olfactory glomerulus. © 2004 Wiley‐Liss, Inc.
Glucose deprivation increases hydrogen peroxide level in immunostimulated rat primary astrocytes
Authors: Choi Ji Woong , Shin Chan Young , Yoo Byoung Kwon , Choi Min Sik , Lee Woo Jong , Han Byoung Hee , Kim Won‐Ki , Kim Hyoung‐Chun , Ko Kwang Ho
Published: February 2004

Activated astrocytes produce a large amount of bioactive molecules, including reactive oxygen and nitrogen species. Astrocytes are in general resistant to those reactive species. However, we previously reported that immunostimulated astrocytes became highly vulnerable to metabolic insults, such as glucose deprivation. In this study, we investigated whether H2O2 production was associated with the increased vulnerability. Glucose deprivation for up to 8 hr did not change the intracellular level of H2O2 in astrocytes. Treatment with lipopolysaccharide plus interferon‐γ for 48 hr evoked astroglial H2O2 production; however, no apparent death or injury was observed in immunostimulated astrocytes. Glucose deprivation after 48 hr of immunostimulation markedly increased H2O2 level, depleted adenosine triphosphate (ATP), and enhanced lactate dehydrogenase (LDH) release. The ATP depletion and LDH release were in part prevented by catalase, mannitol, and N‐acetyl‐L‐cysteine. The enhanced level of H2O2 in glucose‐deprived immunostimulated astrocytes appeared to be secondary to the depletion of reduced glutathione. 4‐(2‐Aminoethyl)bebzenesulfonyl fluoride (AEBSF), an inhibitor of NADPH oxidase, reduced H2O2 level and LDH release in glucose‐deprived immunostimulated astrocytes. H2O2, either endogenously produced or exogenously added, depolarized mitochondrial transmembrane potential in glucose‐deprived astrocytes, leading to their ATP depletion and death. The present results strongly indicate that glucose deprivation causes deterioration of immunostimulated astrocytes by increasing the intracellular concentration of H2O2. © 2004 Wiley‐Liss, Inc.
JNK activation by tetrahydrobiopterin: Implication for Parkinson's disease
Authors: Choi Hyun Jin , Lee So Yeon , Cho Yuri , Hwang Onyou
Published: February 2004

Parkinson's disease (PD) is a progressive neurologic disease associated with selective degeneration of dopaminergic neurons in the substantia nigra. Despite extensive studies to understand the underlying cause of dopaminergic degeneration, the pathologic factors leading to this neuronal loss in PD remain obscure. We have observed previously that tetrahydrobiopterin (BH4) exerts selective toxicity and oxidative stress on dopaminergic cells, suggesting that BH4 might participate endogenously in dopaminergic neurodegeneration in PD. We investigated signaling events leading to BH4 toxicity in dopaminergic CATH.a cells. We show that c‐Jun N‐terminal kinase (JNK), but not extracellular signal‐regulated kinase (ERK) or p38 mitogen‐activated protein kinase (MAPK), is phosphorylated significantly by BH4 exposure. BH4 also leads to c‐Jun phosphorylation and an increase in c‐Jun protein level. The JNK inhibitor SP600125 protects cells against BH4 toxicity and inhibits cytochrome c release and apoptotic nuclear condensation induced by BH4. These data indicate that activation of the JNK pathway is important in mediating BH4‐induced dopaminergic cell death. © 2004 Wiley‐Liss, Inc.
Binding and transport of [3H](2S,4R)‐ 4‐methylglutamate, a new ligand for glutamate transporters, demonstrate labeling of EAAT1 in cultured murine astrocytes
Authors: Apricò Karina , Beart Philip M. , Crawford Duncan , O'Shea Ross D.
Published: February 2004

Transporters for L‐glutamate (excitatory amino acid transporters; EAATs), localized to astrocytes, are involved intimately in intermediary metabolism within the brain. Because (2S,4R)‐4‐methylglutamate (4MG) has affinity for glial EAATs, we employed [3H]4MG to define the characteristics of EAATs in cultured murine astrocytes and describe new approaches to analyze EAAT function. Specific binding of [3H]4MG in astrocytic membranes at 4°C represented 90% of total binding. Binding was rapid (apparent t1/2 ∼7 min) and saturable. Saturation and Scatchard analyses indicated a single binding site (nH = 0.8) with a Kd of 6.0 ± 1.5 μM and Bmax = 9.7 ± 2.9 pmol/mg protein. Binding of [3H]4MG to astrocytic homogenates was Na+‐dependent and inhibited by K+. Compounds acting at EAATs, such as L‐glutamate (Glu), D‐aspartate (D‐Asp), L‐(2S,3S,4R)‐2‐(carboxycyclopropyl)glycine and L‐trans‐pyrrolidine‐2,4‐dicarboxylate displaced binding to nonspecific levels. L‐Serine‐O‐sulphate, an EAAT1‐preferring ligand, fully displaced binding of [3H]4MG. In contrast, inhibitors having preferential affinity for EAAT2, L‐threo‐3‐methylglutamate, dihydrokainate, and kainate, were relatively ineffective binding displacers. Agonists and antagonists for Glu receptors failed to significantly inhibit [3H]4MG binding. Studies with [3H]D‐Asp reinforced evidence that [3H]4MG was binding to EAATs. These data were consistent with Western blot analyses, which indicated abundant expression of EAAT1 but not EAAT2. [3H]4MG was also accumulated rapidly (apparent t1/2 ∼4 min) into whole astrocytes by a sodium‐ and temperature‐sensitive process (Km of 146 ± 24 μM, Vmax = 336 ± 27 nmol/mg protein/min), which possessed an EAAT1‐like pharmacologic profile. These findings confirm that 4MG is a substrate for EAAT1 and that the binding assay developed using [3H]4MG can be utilized in various preparations including cultured astrocytes. © 2004 Wiley‐Liss, Inc.
Increased expression of the MBP mRNA binding protein HnRNP A2 during oligodendrocyte differentiation
Authors: Maggipinto M. , Rabiner C. , Kidd G.J. , Hawkins A.J. , Smith R. , Barbarese E.
Published: February 2004

Heterogeneous nuclear ribonucleoprotein (hnRNP) A2, a trans‐acting factor that mediates intracellular trafficking of myelin basic protein (MBP) mRNA to the myelin compartment in oligodendrocytes, is most abundant in the nucleus, but shuttles between the nucleus and cytoplasm. In the cytoplasm, it is associated with granules that transport mRNA from the cell body to the processes of oligodendrocytes. We found that the overall level of hnRNP A2 increased in oligodendrocytes as they differentiated into MBP‐positive cells, and that this augmentation was reflected primarily in the cytoplasmic pool of hnRNP A2 present in the form of granules. The extranuclear distribution of hnRNP A2 was also observed in brain during the period of myelination in vivo. Methylation and phosphorylation have been implicated previously in the nuclear to cytoplasmic distribution of hnRNPs, so we used drugs that block methylation and phosphorylation of hnRNPs to assess their effect on hnRNP A2 distribution and mRNA trafficking. Cultures treated with adenosine dialdehyde (AdOx), an inhibitor of S‐adenosyl‐L‐homocysteine hydrolase, or with 5,6‐dichloro‐1‐β‐D‐ribofuranosylbenzimidazole (DRB), a drug that inhibits casein kinase 2 (CK2), maintained the preferential nuclear distribution of hnRNP A2. Treatment with either drug affected the transport of RNA trafficking granules that remained confined to the cell body. © 2004 Wiley‐Liss, Inc.
Distinctive expression of midkine in the repair period of rat brain during neurogenesis: Immunohistochemical and immunoelectron microscopic observations
Authors: Kikuchi‐Horie Kae , Kawakami Emiko , Kamata Makiko , Wada Manabu , Hu Jian‐Guo , Nakagawa Hachiro , Ohara Kazuhiko , Watabe Kazuhiko , Oyanagi Kiyomitsu
Published: February 2004

Distinctive expression of midkine (MK) was observed during the repair period of fetal brain neuroepithelium. MK is a heparin‐binding growth factor that occurs as a product of a retinoic acid‐inducible gene, and has a molecular mass of 13 kDa. MK expression was examined immunohistochemically and by immunoelectron microscopy during a period of repair in developing rat brain at the neurogenesis stage. Injury was induced in rat fetuses by transplacental administration of ethylnitrosourea (ENU) on embryonic Day (E) 16, and histological changes were examined up to 48 hr thereafter (i.e., up to E 18). In normal rat fetuses, MK immunostaining was observed in the cytoplasm and radial and horizontal processes of all cells in the neuroepithelium (NE), subventricular zone (SV), and intermediate zone (IMZ). In ENU‐administered brains, cells in the NE, SV, and IMZ were damaged severely, especially 16–24 hr after ENU administration. The remaining neuroepithelial cells, with the exception of those in M‐phase and the tips of processes at the ventricular surface, were negative for MK immunohistochemistry 16–24 hr after the administration of ENU. Forty‐eight hours after the administration, the cytoplasm and processes of cells in the NE, SV, and IMZ were MK immunopositive. Our previous data reported that the cell cycle of most NE cells is synchronized to the S‐phase 16 hr after ENU administration and to the M‐phase at 24 hr, and many NE cells were recovered 48 hr after ENU administration. The previous results taken together with the present results indicate that: (1) MK expression does not increase during the repair period of the NE, being different from adults; (2) MK expression is likely to be suppressed at S‐phase according to the condition of the NE; and (3) MK expression is not essential for every cell cycle phase of NE cells; but (4) is necessary to maintain the M‐phase of NE cells. © 2004 Wiley‐Liss, Inc.