bluuebunny/crossref_metadata_2025_split huggingface dataset:
Surface engineering of living myoblasts via selective periodate oxidation
Authors: De Bank P. A. , Kellam B. , Kendall D. A. , Shakesheff K. M. 
Published: January 2003

AbstractCell surface molecules are vital for normal cell activity. To study the functions of these molecules or manipulate cell behavior, the ability to decorate cell surfaces with bioactive molecules of our choosing is a potentially powerful technique. Here, we describe the molecular engineering of living L6 myoblast monolayers via selective periodate oxidation of sialic acid residues and the application of this surface modification in the artificial aggregation of cells. The aldehyde groups generated by this reaction were used to selectively ligate a model molecule, biotin hydrazide, to the cell surfaces. Flow cytometry analysis after staining with fluorescently conjugated avidin revealed a concentration‐dependent increase in fluorescence compared to untreated cells with a maximal shift of 345.1 ± 27.4‐fold and an EC50 of 17.4 ± 1.1 μM. This mild oxidation reaction did not affect cell number, viability, or morphology. We then compared this chemical technique with the metabolic incorporation of reactive cell surface ketone groups using N‐levulinoylmannosamine (ManLev). In this cell line, only a 22.3‐fold fluorescence shift was observed compared to untreated cells when myoblasts were incubated with a high concentration of ManLev for 48 hours. Periodate oxidation was then used to modify myoblast surfaces to induce cell aggregation. Crosslinking biotinylated myoblasts, which do not spontaneously aggregate in culture, with avidin resulted in the rapid formation of millimeter‐sized, multicellular structures. These data indicate that sodium periodate treatment is an effective, noncytotoxic method for the in vitro molecular engineering of living cell surfaces with the potential for cell biology and tissue engineering applications. © 2003 Wiley Periodicals, Inc. Biotechnol Bioeng 81: 800–808, 2003.
An EPR study of the secretion of G‐CSF heterologous protein from Pichia pastoris
Authors: Anžur Lasnik M. , Nemec M. , Romih R. , Schara M. 
Published: January 2003

AbstractThe biologically active protein known as human granulocyte colony stimulating factor (G‐CSF) can be efficiently secreted from the transformed GS115 Pichia pastoris (GS115/pPIC9/G‐CSF), which contains an α‐mating‐factor prepro signal sequence and an alcohol oxidase I promoter, both introduced using the pPIC9 plasmid. To study the P. pastoris G‐CSF protein‐secretion process, changes to the plasma membrane's lateral domain structure were monitored using electron paramagnetic resonance (EPR). The GS115 and its transformed analogue show that the plasma membrane can be described by fluid‐disordered and fluid‐ordered lateral domains. The relative proportion of these domains is a sensitive parameter that is able to describe the membrane's involvement in the protein‐excretion process. Here, P. pastoris GS115 served as a control for us to compare with the GS115/pPIC9/G‐CSF heterologous protein‐secreting cells. Electron paramagnetic resonance studies using the spin‐probe 5‐doxyl methyl ester of palmitic acid [MeFASL (10,3)] showed that during cultivation in a glycerol medium all types of cells had a relatively high proportion of cell‐membrane fluid‐disordered domains, while during the production phase the G‐CSF protein‐secreting cells showed a decrease in the proportion of fluid‐disordered domains. We ascribe this effect to the vesicular lipid exchange caused by the fusion of secretary vesicles with the plasma membrane during exocytosis. Using electron microscopy and immunocytochemistry intracellular vesicles containing the G‐CSF protein were detected. Our studies support the exocytotic mechanism of the heterologous protein secretion. © 2003 Wiley Periodicals, Inc. Biotechnol Bioeng 81: 768–774, 2003.
Quantitative comparison of transient growth of Saccharomyces cerevisiae, Saccharomyces kluyveri, and Kluyveromyces lactis
Authors: Herwig Christoph , von Stockar Urs 
Published: January 2003

AbstractA multitude of metabolic regulations occur in yeast, particularly under dynamic process conditions, such as under sudden glucose excess. However, quantification of regulations and classification of yeast strains under these conditions have yet to be elucidated, which requires high‐frequency and consistent quantification of the metabolic response. The present study aimed at quantifying the dynamic regulation of the central metabolism of strains Saccharomyces cerevisiae, S. kluyveri, and Kluyveromyces lactis upon sudden glucose excess, accomplished by a shift‐up in dilution rate inside of the oxidative region using a small metabolic flux model. It was found that, under transient growth conditions, S. kluyveri behaved like K. lactis, while classification using steady‐state conditions would position S. kluyveri close to S. cerevisiae. For transient conditions and based on the observation whether excess glucose is initially used for catabolism (energy) or anabolism (carbon), we propose to classify strains into energy‐driven, such as S. cerevisiae, and carbon‐driven, such as S. kluyveri and K. lactis, strains. Furthermore, it was found that the delayed onset of fermentative catabolism in carbon‐driven strains is a consequence of low catabolic flux and the initial shunt of glucose in non‐nitrogen‐containing biomass constituents. The MFA model suggests that energy limitation forced the cell to ultimately increase catabolic flux, while the capacity of oxidative catabolism is not sufficient to process this flux oxidatively. The combination of transient experiments and its exploitation with reconciled intrinsic rates using a small metabolic model could corroborate earlier findings of metabolic regulations, such as tight glucose control in carbon‐driven strains and transient changes in biomass composition, as well as explore new regulations, such as assimilation of ethanol before glucose. The benefit from using small metabolic flux models is the richness of information and the enhanced insight into intrinsic metabolic pathways without a priori knowledge of adaptation kinetics. Used in an online context, this approach serves as an efficient tool for strain characterization and physiological studies. © 2003 Wiley Periodicals. Biotechnol Bioeng 81: 837–847, 2003.
Solid‐phase peptide synthesis by ion‐paired α‐chymotrypsin in nonaqueous media
Authors: Altreuter David H. , Dordick Jonathan S. , Clark Douglas S. 
Published: January 2003

AbstractSolid‐phase synthesis of dipeptides in low‐water media was achieved using AOT ion‐paired α‐chymotrypsin solubilized in organic solvents. Multiple solvents and systematic variation of water activity, aw, were used to examine the rate of coupling between N‐α‐benzyloxycarbonyl‐L‐phenylalanine methyl ester (Z‐Phe‐OMe) and leucine as a function of the reaction medium for both solid‐phase and solution‐phase reactions. In solution, the observed maximum reaction rate in a given solvent generally correlated with measures of hydrophobicity such as the log of the 1‐octanol/water partitioning coefficient (log P) and the Hildebrand solubility parameter. The maximum rate for solution‐phase synthesis (13 mmol/h g‐enzyme) was obtained in a 90/10 (v/v) isooctane/tetrahydrofuran solvent mixture at an aw of 0.30. For the synthesis of dipeptides from solid‐phase leucine residues, the highest synthetic rates (0.14–1.3 mmol/h g‐enzyme) were confined to solvent environments that fell inside abruptly defined regions of solvent parameter space (e.g., log P > 2.3 and normalized electron acceptance index <0.13). The maximum rate for solid‐phase synthesis was obtained in a 90/10 (v/v) isooctane/tetrahydrofuran solvent mixture at an aw of 0.14. In 90/10 and 70/30 (v/v) isooctane/tetrahydrofuran environments with aw set to 0.14, seven different N‐protected dipeptides were synthesized on commercially available Tentagel support with yields of 74–98% in 24 h. © 2003 Wiley Periodicals, Inc. Biotechnol Bioeng 81: 809–817, 2003.
Self‐reflection and self‐practice in training of Cognitive Behaviour Therapy: an overview
Authors: Laireiter Anton‐Rupert , Willutzki Ulrike 
Published: January 2003

AbstractThe present paper deals with a developing training element in cognitive behaviour therapy (CBT), the self‐practice of therapeutic techniques and the self‐exploration of the person of the therapist. Initially, the current status of this training element in CBT is discussed, and a short note on its terminology is presented. Then an overview of the most important objectives of self‐practice and self‐reflection in CBT, and the concepts and methods of practice of this element is given. The concepts focus on two major aims, the self‐exploration of the person of the therapist within and beyond his or her therapeutic practice, and the self‐application of therapeutic techniques (= self‐practice). In the second part of the paper empirical studies related to the outcome of self‐reflection and self‐practice on the development of the therapist and her or his therapeutic practice are reviewed. The few empirical studies show that trainees experience in their own view substantial professional and personal gains from this training tool, with the professional impact being more important than the personal one. The most important outcomes evaluated by subjective data from the trainees are improvements in self‐insight and self‐awareness and a better understanding of the therapist's role and the therapeutic change process. Additionally, a better understanding of CBT methods and of general therapeutic skills, such as empathy and role‐taking‐competencies, are reported by the trainees. The paper comes to the conclusion that self‐exploration and self‐practice are important components of CBT training. Consequences and recommendations for its integration into training courses for CBT are discussed as well as the necessity for more research in this area. Copyright © 2002 John Wiley & Sons, Ltd.
Trainee clinical psychologists' adaptation and professional functioning: a longitudinal study
Authors: Kuyken Willem , Peters Emmanuelle , Power Michael J. , Lavender Tony 
Published: January 2003

AbstractClinical psychology training is a professionally formative period, which provides an opportunity to enable trainees to learn good self‐care skills and mature approaches to learning. If realized, this can support lifelong learning and high levels of professional functioning. This study sought to use transactional coping theory and experiential learning theory to improve our understanding of what factors predict changes in psychological adaptation and professional functioning over the course of clinical psychology training. A mixed prospective within‐persons and cross‐sectional between‐persons design was used. A sample of 183 trainee clinical psychologists (60.2% response rate) from 15 British clinical psychology training programmes participated at time one, 167 of whom participated at time 2 a year later (91.3% of the time 1 sample). They completed measures of appraisal, coping, social support and professional functioning at times 1 and 2. Path analyses suggested that trainees who appraised the demands of training as manageable, and reported greater access to appropriate support, engaged in less avoidance coping, reported fewer problems of psychological adaptation and were more likely to approach the task of learning and working appropriately and resiliently. Implications for clinical psychology training are suggested. Copyright © 2003 John Wiley & Sons, Ltd.
Identification of genes induced in regenerating Xenopus tadpole tails by using the differential display method
Authors: Ishino Tomoko , Shirai Mie , Kunieda Takekazu , Sekimizu Kazuhisa , Natori Shunji , Kubo Takeo 
Published: January 2003

AbstractTo identify candidate gene(s) involved in the tail regeneration of Xenopus laevis tadpoles, we used the differential display method to isolate four genes (clones 1, 2, 13a, and 13b) whose expression is induced in regenerating tadpole tails. Among them, clones 13a and 13b were found to encode the Xenopus homologues of the α1 chain of type XVIII collagen and neuronal pentraxin I, respectively. Expression of clone 2 and neuronal pentraxin I genes increased dramatically in the blastema 3 days after amputation, whereas that for the clone 1 and type XVIII collagen genes was induced gradually after amputation. In situ hybridization revealed that the neuronal pentraxin I gene is expressed specifically in the regenerating tail epidermis but not in the normal tail epidermis or the most distal margin of the tail blastema, suggesting that it has a tissue‐inductive role in tail regeneration. Expression of the four genes was induced in the limb and in the tail blastema, suggesting that they are involved in the regeneration of both organs. Finally, expression of clone 2 and neuronal pentraxin I genes was scarce during embryonic stages in comparison to the tail blastema, suggesting that their main functions are in organ regeneration. Our results demonstrate unique features of spatial and temporal gene expression patterns during Xenopus tadpole tail regeneration. Developmental Dynamics 226:317–325, 2003. © 2003 Wiley‐Liss, Inc.
Recruitment of postmitotic neurons into the regenerating spinal cord of urodeles
Authors: Zhang Fang , Ferretti Patrizia , Clarke Jonathan D.W. 
Published: January 2003

AbstractBy using fluorescent tracers, we have investigated the origin of the cells that form the regenerating spinal cord after tail amputation in urodele amphibians. We show that spinal cord cells immediately adjacent to the amputation plane die and are removed by phagocytic cells. Spinal cells just anterior to these dying cells are destined to make the majority of the regenerating cord. The largest contribution is likely to come from the radial ependymal cells, but we also demonstrate that postmitotic neurons in this location can translocate into the regenerating cord. These neurons integrate into the regenerate structure and survive for at least 4 weeks. We find no evidence that these translocated neurons dedifferentiate and divide during this regeneration process. We discuss the possibility that these neurons survive long term in the regenerate cord and become part of the functional neuronal circuitry. Developmental Dynamics 226:341–348, 2003. © 2003 Wiley‐Liss, Inc.
Bone formation during forelimb regeneration: A microtomography (microCT) analysis
Authors: Stock Stuart R. , Blackburn David , Gradassi Michael , Simon Hans‐Georg 
Published: January 2003

AbstractIn our study of bone regeneration in the forelimbs of mature newts (Notophthalmus viridescens), we used noninvasive X‐ray microtomography (microCT) to image regenerating limbs from 37 to 85 days and matching (contralateral) controls. We compared the patterns of regenerated and existing (nonregenerated) bone, investigating in particular the onset of mineralization of specific bones, the level of mineral present, and the lengths of the different bones. Overall, we find that the missing limb skeletal elements are restored in a proximal‐to‐distal direction, which reiterates the developmental patterning program. However, in contrast to this proximal–distal sequence, the portion of the humerus distal to the amputation site fails to ossify in synchrony with the regenerating radius and ulna. This finding suggests that the replacement of cartilage with mineralized bone close to the amputation site is delayed with respect to other regenerating skeletal elements. Developmental Dynamics 226:410–417, 2003. © 2003 Wiley‐Liss, Inc.
Mediolateral intercalation in planarians revealed by grafting experiments
Authors: Saito Yumi , Koinuma Satoshi , Watanabe Kenji , Agata Kiyokazu 
Published: January 2003

AbstractWe investigated how planarians organize their left–right axis by using ectopic grafting. Planarians have three body axes: anteroposterior (A‐P), dorsoventral (D‐V), and left–right (L‐R). When a small piece is implanted into an ectopic region along the A‐P and D‐V axes, intercalary structures are always formed to compensate for positional gaps. There are two hypotheses regarding L‐R axis formation in this organism: first, that the left and right sides of the animal may be recognized as different parts, and L‐R intercalation can induce midline structures (asymmetry hypothesis); second, that both sides may have symmetrical positional values, and mediolateral (M‐L) intercalation creates positional values along the L‐R axis (symmetry hypothesis). We performed ectopic grafting experiments in the head region of the planarian, Dugesia japonica, to examine these hypotheses. A left lateral fragment containing a left auricle was implanted into the medial region of the host. Ectopic structures were always formed only on the left side of the graft, where lateral tissues abutted onto the medial tissues. However, no morphologic change was induced on the right side of the graft, where left‐sided tissues faced onto right‐sided tissues. Molecular marker analyses indicated that ectopic structures formed on the left side of the graft were induced by M‐L intercalation, supporting the “symmetry hypothesis.” When the midline tissues were implanted into a lateral region, they induced a complete ectopic head, demonstrating that M‐L intercalation may be sufficient to establish the L‐R axis in planarians. Developmental Dynamics 226:334–340, 2003.© 2003 Wiley‐Liss, Inc.
Old questions, new tools, and some answers to the mystery of fin regeneration
Authors: Akimenko Marie‐Andrée , Marí‐Beffa Manuel , Becerra José , Géraudie Jacqueline 
Published: January 2003

AbstractPluridisciplinary approaches led to the notion that fin regeneration is an intricate phenomenon involving epithelial–mesenchymal and reciprocal exchanges throughout the process as well as interactions between ray and interray tissue. The establishment of a blastema after fin amputation is the first event leading to the reconstruction of the missing part of the fin. Here, we review our knowledge on the origin of the blastema, its formation and growth, and of the mechanisms that control differentiation and patterning of the regenerate. Our current understanding results from studies of fin regeneration performed in various teleost fish over the past century. We also report the recent breakthroughs that have been made in the past decade with the arrival of a new model, the zebrafish, Danio rerio, which now offers the possibility to combine cytologic, molecular, and genetic analyses and open new perspectives in this field. Developmental Dynamics 226:190–201, 2003. © 2003 Wiley‐Liss, Inc.
Intercalary regeneration in planarians
Authors: Agata K. , Tanaka T. , Kobayashi C. , Kato K. , Saitoh Y. 
Published: January 2003

AbstractHow can a planarian regenerate its entire body from a small portion of its body? Neoblasts, the totipotent stem cells of planarian, are assumed to be able to produce all missing cell types. However, we do not know how the cell fate of these cells is controlled during regeneration. Our recent studies with molecular markers suggest that intercalary regeneration is the fundamental principle in planarian regeneration. Here, we introduce the intercalation induced by ectopic grafting along the anteroposterior (A‐P), dorsoventral (D‐V), and left–right (L‐R) axes. Blastema formation is evoked by ectopic D‐V interactions after wound closure. Intercalation between the blastema and stump induces rearrangement of the positional identities along the A‐P axis. Consequently, totipotent stem cells change their differentiation patterns according to the newly rearranged positional identities along the A‐P, D‐V, and L‐R axes. According to the classic view, the blastema is regarded as the place where undifferentiated cells accumulate and regenerative events occur. Here, we propose a new interpretation, i.e., that the blastema may work as a signaling center inducing intercalary regeneration. Also, the roles of molecules and genes involved in intercalary regeneration are discussed. Developmental Dynamics 226:308–316, 2003. © 2003 Wiley‐Liss, Inc.
Rearrangements and increased expression of cyclin D1 (CCND1) in neuroblastoma
Authors: Molenaar Jan J. , van Sluis Peter , Boon Kathy , Versteeg Rogier , Caron Huib N. 
Published: January 2003

AbstractCyclin D1 regulates G1 cell cycle progression by controlling the phosphorylation of the retinoblastoma protein. This pathway is frequently deregulated in many malignancies. In neuroblastoma, however, no consistent G1 cell cycle checkpoint aberrations have been found. We examined the possible deregulation of cyclin D1 (CCND1) in this tumor. mRNA expression profiles of neuroblastoma generated by SAGE (Serial Analysis of Gene Expression) revealed a high expression of CCND1 in a subset of neuroblastoma cell lines and tumors. The CCND1 expression level can be 0.3% of the total cellular mRNA. Northern blot analysis of CCND1 expression showed a relative overexpression in 16 of 23 neuroblastoma cell lines and 10 of 15 tumor samples. In the majority of cases, the high CCND1 mRNA levels also led to high CCND1 protein levels. In the search for mechanisms causing this relative overexpression, we screened for amplifications and rearrangements of CCND1. Five amplifications were found in 202 neuroblastoma tumors and cell lines. Analysis of the 3′‐UTR of CCND1 showed a rearrangement in 1 of 96 tumors. These clonal aberrations of CCND1 together with the high expression suggest a role for deregulated CCND1 activity in neuroblastoma tumorigenesis. © 2003 Wiley‐Liss, Inc.
Cryptic chromosomal aberrations leading to an AML1/ETO rearrangement are frequently caused by small insertions
Authors: Gamerdinger Ulrike , Teigler‐Schlegel Andrea , Pils Sabine , Bruch Jochen , Viehmann Susanne , Keller Monika , Jauch Anna , Harbott Jochen 
Published: January 2003

AbstractThe translocation t(8;21)(q22;q22), which results in the fusion of the AML1 (RUNX1) and ETO (CBFA2T1) genes, is a recurrent aberration in acute myeloid leukemia (AML), preferentially correlated with FAB M2, and has the highest incidence in childhood AML. Because of the favorable prognosis, the evidence of the t(8;21) or the AML1/ETO fusion gene is mandatory in most of the therapy trials, allowing the stratification of the patients to the correct risk group in terms of treatment. Here we present six out of 59 children with AML who were positive for AML1/ETO by RT‐PCR, but showed no evidence of the classical t(8;21)(q22;q22) by conventional cytogenetics. Because of the discrepancies between molecular and cytogenetic analyses, these six patients were further investigated by fluorescence in situ hybridization analysis. Small hidden interstitial insertions resulting in an AML1/ETO rearrangement were detected in five (8.5%) of the 59 patients, whereas the sixth patient showed a cryptic three‐way translocation. The insertions could be characterized as ins(21;8) in three patients and ins(8;21) in the remaining two. Additionally, three of the patients showed secondary chromosome aberrations leading to a higher complexity of the karyotype. In conclusion, the combination of more than one standard technique in the analysis of AML1/ETO is useful to reveal the overall frequency of cryptic chromosome rearrangements and permits a better understanding of the mechanisms involved in the generation of this fusion gene. © 2003 Wiley‐Liss, Inc.