bluuebunny/crossref_metadata_2025_split huggingface dataset:
Cross‐linking density alters early metabolic activities in chondrocytes encapsulated in poly(ethylene glycol) hydrogels and cultured in the rotating wall vessel
Authors: Villanueva Idalis , Klement Brenda J. , von Deutsch Daniel , Bryant Stephanie J. 
Published: September 2008

AbstractIn designing a tissue engineering strategy for cartilage repair, selection of both the bioreactor, and scaffold is important to the development of a mechanically functional tissue. The hydrodynamic environment associated with many bioreactors enhances nutrient transport, but also introduces fluid shear stress, which may influence cellular response. This study examined the combined effects of hydrogel cross‐linking and the hydrodynamic environment on early chondrocyte response. Specifically, chondrocytes were encapsulated in poly(ethylene glycol) (PEG) hydrogels having two different cross‐linked structures, corresponding to a low and high cross‐linking density. Both cross‐linked gels yielded high water contents (92% and 79%, respectively) and mesh sizes of 150 and 60 Å respectively. Cell‐laden PEG hydrogels were cultured in rotating wall vessels (RWV) or under static cultures for up to 5 days. Rotating cultures yielded low fluid shear stresses (≤0.11 Pa) at the hydrogel periphery indicating a laminar hydrodynamic environment. Chondrocyte response was measured through total DNA content, total nitric oxide (NO) production, and matrix deposition for glycosaminoglycans (GAG). In static cultures, gel cross‐linking had no effect on DNA content, NO production, or GAG production; although GAG production increased with culture time for both cross‐linked gels. In rotating cultures, DNA content increased, NO production decreased, and overall GAG production decreased when compared to static controls for the low cross‐linked gels. For the high cross‐linked gels, the hydrodynamic environment had no effect on DNA content, but exhibited similar results to the low cross‐linked gel for NO production, and matrix production. Our findings demonstrated that at early culture times, when there is limited matrix production, the hydrodynamic environment dramatically influences cell response in a manner dependent on the gel cross‐linking, which may impact long‐term tissue development. Biotechnol. Bioeng. 2009;102: 1242–1250. © 2008 Wiley Periodicals, Inc.
Engineering and expression of a chimeric transferrin receptor monoclonal antibody for blood–brain barrier delivery in the mouse
Authors: Boado Ruben J. , Zhang Yun , Wang Yuntao , Pardridge William M. 
Published: September 2008

AbstractProtein therapeutics may be delivered across the blood–brain barrier (BBB) by genetic fusion to a BBB molecular Trojan horse. The latter is an endogenous peptide or a peptidomimetic monoclonal antibody (MAb) against a BBB receptor, such as the insulin receptor or the transferrin receptor (TfR). Fusion proteins have been engineered with the MAb against the human insulin receptor (HIR). However, the HIRMAb is not active against the rodent insulin receptor, and cannot be used for drug delivery across the mouse BBB. The rat 8D3 MAb against the mouse TfR is active as a drug delivery system in the mouse, and the present studies describe the cloning and sequencing of the variable region of the heavy chain (VH) and light chain (VL) of the rat 8D3 TfRMAb. The VH and VL were fused to the constant region of mouse IgG1 heavy chain and mouse kappa light chain, respectively, to produce a new chimeric TfRMAb. The chimeric TfRMAb was expressed in COS cells following dual transfection with the heavy and light chain expression plasmids, and was purified by protein G affinity chromatography. The affinity of the chimeric TfRMAb for the murine TfR was equal to the 8D3 MAb using a radio‐receptor assay and mouse fibroblasts. The chimeric TfRMAb was radio‐labeled and injected into mice for a pharmacokinetics study of the clearance of the chimeric TfRMAb. The chimeric TfRMAb was rapidly taken up by mouse brain in vivo at a level comparable to the rat 8D3 MAb. In summary, these studies describe the genetic engineering, expression, and validation of a chimeric TfRMAb with high activity for the mouse TfR, which can be used in future engineering of therapeutic fusion proteins for BBB drug delivery in the mouse. Biotechnol. Bioeng. 2009;102: 1251–1258. © 2008 Wiley Periodicals, Inc.
Molecular engineering of exocytic vesicle traffic enhances the productivity of Chinese hamster ovary cells
Authors: Peng Ren‐Wang , Fussenegger Martin 
Published: September 2008

AbstractA complex vesicle trafficking system manages the precise and regulated distribution of proteins, membranes and other molecular cargo between cellular compartments as well as the secretion of (heterologous) proteins in mammalian cells. Sec1/Munc18 (SM) proteins are key components of the system by regulating membrane fusion. However, it is not clear how SM proteins contribute to the overall exocytosis. Here, functional analysis of the SM protein Sly1 and Munc18c suggested a united, positive impact upon SNARE‐based fusion of ER‐to‐Golgi‐ and Golgi‐to‐plasma membrane‐addressed exocytic vesicles and increased the secretory capacity of different therapeutic proteins in Chinese hamster ovary cells up to 40 pg/cell/day. Sly1‐ and Munc18c‐based vesicle traffic engineering cooperated with Xbp‐1‐mediated ER/Golgi organelle engineering. Our study supports a model for united function of SM proteins in stimulating vesicle trafficking machinery and provides a generic secretion engineering strategy to improve biopharmaceutical manufacturing of important protein therapeutics. Biotechnol. Bioeng. 2009;102: 1170‐1181. © 2008 Wiley Periodicals, Inc.
Redirecting the inactivation pathway of penicillin amidase and increasing amoxicillin production via a thermophilic molecular chaperone
Authors: Bergeron Lisa M. , Tokatlian Talar , Gomez Lizabeth , Clark Douglas S. 
Published: September 2008

AbstractWe have previously shown that a single‐subunit thermosome from Methanocaldococcus jannaschii (rTHS) can stabilize enzymes in semi‐aqueous media (Bergeron et al., 2008b). In the present study, rTHS was used to stabilize penicillin amidase (PGA) in methanol–water mixtures. Including methanol in the reaction medium for amoxicillin synthesis can suppress unwanted hydrolysis reactions but inactivate PGA. Inactivation and reactivation pathways proposed for PGA illustrate the predictability of enzyme stabilization by rTHS in co‐solvents. Calcium was necessary for reversible dissociation of the two PGA subunits in methanol–water and the presence of calcium resulted in an enhancement of chaperone‐assisted stabilization. rTHS also acted as a stabilizer in the enzymatic synthesis of the β‐lactam antibiotic amoxicillin. rTHS stabilized PGA, increasing its half‐life in 35% methanol by fivefold at 37°C. Stabilization by rTHS was enhanced but did not require the presence of ATP. Including rTHS in fed‐batch reactions performed in methanol–water resulted in nearly 4 times more amoxicillin than when the reaction was run without rTHS, and over threefold higher selectivity towards amoxicillin synthesis compared to aqueous conditions without rTHS. The thermosome and other thermophilic chaperones may thus be generally useful for stabilizing enzymes in their soluble form and expanding the range of conditions suitable for biocatalysis. Biotechnol. Bioeng. 2009;102: 417–424. © 2008 Wiley Periodicals, Inc.
Effect of sodium dodecylbenzene sulfonate on subtilisin Carlsberg proteolysis of an immobilized ovalbumin film
Authors: Foose Ladan L. , Blanch Harvey W. , Radke C.J. 
Published: September 2008

AbstractEnzymatic degradation of immobilized ovalbumin multilayer films by subtilisin Carlsberg was investigated using in situ ellipsometry. Changes in the substrate cleavage rate in the presence of an anionic surfactant, sodium dodecylbenzene sulfonate (SDBS), were assessed. Exposure of the protein film to SDBS prior to introduction of the enzyme increased the measured proteolysis rate threefold. Surfactant increased the measured film thickness, absorbing into the protein film and causing swelling. Surfactant‐induced film swelling was reversible upon aqueous rinsing. Nevertheless, exposure of enzyme to the surfactant‐rinsed film increased the proteolysis rate, most likely due to irreversible conformational changes induced in the substrate film by the surfactant. Simultaneous addition of SDBS with enzyme after the initial surfactant exposure did not produce additional protein‐removal benefit. Biotechnol. Bioeng. 2009;102: 1273–1277. © 2008 Wiley Periodicals, Inc.
Modeling electrostatic exclusion effects during ion exchange chromatography of monoclonal antibodies
Authors: Zydney Andrew L. , Harinarayan Chithkala , van Reis Robert 
Published: September 2008

AbstractRecent experimental studies have shown a reduction in dynamic‐ binding capacity for both monoclonal antibodies and antigen‐binding fragments at very low conductivity, conditions that should generate the greatest electrostatic attraction. This behavior has been attributed to the steric and electrostatic exclusion of the charged protein from the entrance of the resin pores. This manuscript presents a quantitative mathematical description of this phenomenon. The protein partition coefficient was evaluated using models for the partitioning of a charged sphere into a charged cylindrical pore, with the pore size distribution evaluated by inverse size exclusion chromatography. The results were in very good agreement with experimental data for batch protein uptake and dynamic‐binding capacity over a range of pH and conductivity. This theoretical framework provides important insights into the behavior of ion exchange chromatography for protein purification. Biotechnol. Bioeng. 2009;102: 1131–1140. © 2008 Wiley Periodicals, Inc.
Attachment anxiety, intra‐group (dis)respect, actual efforts, and group donation
Authors: Erez Ayelet , Sleebos Ed , Mikulincer Mario , Van Ijzendoorn Marinus H. , Ellemers Naomi , Kroonenberg Pieter M. 
Published: September 2008

AbstractThe current study examines attachment‐style differences in responses to inductions of group respect and disrespect. Participants completed a scale assessing attachment anxiety and avoidance, performed group tasks, and received high, average, or low respect feedback from group members. Then we assessed commitment to this group, actual effort expenditure on behalf of the group, and money donation to the group. For participants scoring relatively high on attachment anxiety, high group respect heightened group commitment and effort expenditure on behalf of the group, whereas group disrespect led to lower group commitment but to more money donation to the group and higher effort expenditure. Participants who scored relatively low on attachment anxiety were not significantly affected by group respect or disrespect. The implications of attachment theory for group dynamics were discussed. Copyright © 2008 John Wiley & Sons, Ltd.
Experimental and numerical characterization of the flowfield in the large‐scale UW live fire research facility
Authors: Devaud C. B. , Weisinger J. , Johnson D. A. , Weckman E. J. 
Published: September 2008

AbstractRecently, a large‐scale (19.5m × 15.4m × 12.8m) live fire research facility (LFRF) has been built at the University of Waterloo with the capabilities of conducting controlled and systematic experimental studies to investigate the behaviour of large fuel spill fires in crosswinds. The present work is focused on characterizing the wind generated by banked fans and the flowfield in the LFRF using experimental techniques and large eddy simulation (LES). Two configurations are examined: the empty enclosure and the enclosure with a large blocking object (cylinder) aligned perpendicular to the flow direction.Detailed velocity measurements are provided throughout the facility. The experimental results show that the flowfield inside the empty enclosure is consistent with the behaviour of a bluff wall jet.Three‐dimensional LES are performed for the same two configurations. The experimental velocity profiles are used to set the inflow conditions in the calculations. The numerical predictions are all within the experimental uncertainty in the core region for the empty enclosure. For the case with the cylinder, the size of the wake is very well reproduced by the simulations and the shedding frequency corresponds to the value given in published experimental studies of a flow over free standing cylinders. The present LES model is also capable of predicting more detailed flow characteristics such as the stagnation and recirculation region, compared with what was achievable in the large‐scale experiments. In the wake the time‐averaged velocities from the LES results are overpredicted. Possible sources of discrepancy are discussed. Copyright © 2008 John Wiley & Sons, Ltd.
Identification of candidate tumor suppressor genes inactivated by promoter methylation in melanoma
Authors: Bonazzi Vanessa F. , Irwin Darryl , Hayward Nicholas K. 
Published: September 2008

AbstractTumor suppressor genes (TSGs) are sometimes inactivated by transcriptional silencing through promoter hypermethylation. To identify novel methylated TSGs in melanoma, we carried out global mRNA expression profiling on a panel of 12 melanoma cell lines treated with a combination of 5‐Aza‐2‐deoxycytidine (5AzadC) and an inhibitor of histone deacetylase, Trichostatin A. Reactivation of gene expression after drug treatment was assessed using Illumina whole‐genome microarrays. After qRT‐PCR confirmation, we followed up 8 genes (AKAP12, ARHGEF16, ARHGAP27, ENC1, PPP1R3C, PPP1R14C, RARRES1, and TP53INP1) by quantitative DNA methylation analysis using mass spectrometry of base‐specific cleaved amplification products in panels of melanoma cell lines and fresh tumors. PPP1R3C, ENC1, RARRES1, and TP53INP1, showed reduced mRNA expression in 35–59% of the melanoma cell lines compared to melanocytes and which was correlated with a high proportion of promoter methylation (>40–60%). The same genes also showed extensive promoter methylation in 6–25% of the tumor samples, thus confirming them as novel candidate TSGs in melanoma. © 2008 Wiley‐Liss, Inc.
Integration of genomic and gene expression data of childhood ALL without known aberrations identifies subgroups with specific genetic hallmarks
Authors: Bungaro Silvia , Dell'Orto Marta Campo , Zangrando Andrea , Basso Dario , Gorletta Tatiana , Lo Nigro Luca , Leszl Anna , Young Bryan D. , Basso Giuseppe , Bicciato Silvio , Biondi Andrea , te Kronnie Gertruy , Cazzaniga Giovanni 
Published: September 2008

AbstractPediatric acute lymphoblastic leukemia (ALL) comprises genetically distinct subtypes. However, 25% of cases still lack defined genetic hallmarks. To identify genomic aberrancies in childhood ALL patients nonclassifiable by conventional methods, we performed a single nucleotide polymorphisms (SNP) array‐based genomic analysis of leukemic cells from 29 cases. The vast majority of cases analyzed (19/24, 79%) showed genomic abnormalities; at least one of them affected either genes involved in cell cycle regulation or in B‐cell development. The most relevant abnormalities were CDKN2A/9p21 deletions (7/24, 29%), ETV6 (TEL)/12p13 deletions (3/24, 12%), and intrachromosomal amplifications of chromosome 21 (iAMP21) (3/24, 12%). To identify variation in expression of genes directly or indirectly affected by recurrent genomic alterations, we integrated genomic and gene expression data generated by microarray analyses of the same samples. SMAD1 emerged as a down‐regulated gene in CDKN2A homozygous deleted cases compared with nondeleted. The JAG1 gene, encoding the Jagged 1 ligand of the Notch receptor, was among a list of differentially expressed (up‐regulated) genes in ETV6‐deleted cases. Our findings demonstrate that integration of genomic analysis and gene expression profiling can identify genetic lesions undetected by routine methods and potential novel pathways involved in B‐progenitor ALL pathogenesis. © 2008 Wiley‐Liss, Inc.
Regulation of Schwann cell function by the extracellular matrix
Authors: Chernousov Michael A. , Yu Wei‐Ming , Chen Zu‐Lin , Carey David J. , Strickland Sidney 
Published: September 2008

AbstractLaminins and collagens are extracellular matrix proteins that play essential roles in peripheral nervous system development. Laminin signals regulate Schwann cell proliferation and survival as well as actin cytoskeleton dynamics, which are essential steps for radial sorting and myelination of peripheral axons by Schwann cells. Collagen and their receptors promote Schwann cell adhesion, spreading, and myelination as well as neurite outgrowth. In this article, we will review the recent advances in the studies of laminin and collagen function in Schwann cell development. © 2008 Wiley‐Liss, Inc.
Molecular domains of myelinated axons in the peripheral nervous system
Authors: Salzer James L. , Brophy Peter J. , Peles Elior 
Published: September 2008

AbstractMyelinated axons are organized into a series of specialized domains with distinct molecular compositions and functions. These domains, which include the node of Ranvier, the flanking paranodal junctions, the juxtaparanodes, and the internode, form as the result of interactions with myelinating Schwann cells. This domain organization is essential for action potential propagation by saltatory conduction and for the overall function and integrity of the axon. © 2008 Wiley‐Liss, Inc.
Molecular mechanisms of inherited demyelinating neuropathies
Authors: Scherer Steven S. , Wrabetz Lawrence 
Published: September 2008

AbstractThe past 15 years have witnessed the identification of more than 25 genes responsible for inherited neuropathies in humans, many associated with primary alterations of the myelin sheath. A remarkable body of work in patients, as well as animal and cellular models, has defined the clinical and molecular genetics of these illnesses and shed light on how mutations in associated genes produce the heterogeneity of dysmyelinating and demyelinating phenotypes. Here, we review selected recent developments from work on the molecular mechanisms of these disorders and their implications for treatment strategies. © 2008 Wiley‐Liss, Inc.
The function of RhoGTPases in axon ensheathment and myelination
Authors: Feltri M. Laura , Suter Ueli , Relvas João B. 
Published: September 2008

AbstractRhoGTPases are molecular switches that integrate extracellular signals to perform diverse cellular responses. This ability relies on the network of proteins regulating RhoGTPases activity and localization, and on the interaction of RhoGTPases with many different cellular effectors. Myelination is an ideal place for RhoGTPases regulation, as it is the result of fine orchestration of many stimuli from at least two cell types. Recent work has revealed that RhoGTPases are required for Schwann cells to sort, ensheath, and myelinate axons. Here, we will review these recent advances showing the critical roles for RhoGTPases in various aspects of Schwann development and myelination, including the recent discovery of their involvement in Charcot‐Marie‐Tooth disease. Comparison with potential roles of RhoGTPases in central nervous system myelination will be drawn. © 2008 Wiley‐Liss, Inc.
Neuregulin‐1, a key axonal signal that drives Schwann cell growth and differentiation
Authors: Birchmeier Carmen , Nave Klaus‐Armin 
Published: September 2008

AbstractInteractions between neuronal and glial cells are crucial for establishing a functional nervous system. Many aspects of Schwann cell development and physiology are regulated by neuronal signals; possibly the most spectacular is the elaboration of the myelin sheath. An extensive line of research has revealed that one neuronal factor, termed “neuregulin”, promotes Schwann cell growth and survival, migration along the extending axon, and myelination. The versatility of glial responses elicited by this factor is thus clearly astounding. © 2008 Wiley‐Liss, Inc.
Pituicyte modulation of neurohormone output
Authors: Rosso Lia , Mienville Jean‐Marc 
Published: September 2008

AbstractPituicytes have long been suspected to play a role in the regulation of neurohypophysial hormone output. This role has been mainly ascribed to morphological changes in these cells and subsequent modifications of their tight structural relationships with surrounding nerve terminals and capillaries. These entirely reversible changes are brought about by physiological states such as parturition, lactation, or dehydration, and it was inferred that they should facilitate neurohormone output, based on concerted analyses ofin vitro,in situ, andex vivoexperiments. Pituicyte stellation, thein vitrocounterpart of these morphological changes, can be induced by β‐adrenergic or A1‐adenosine receptor activation, and appears to result from inhibition of the small GTPase RhoA. Actin depolymerization is the key event allowing stellation. Vasopressin and oxytocin reverse stellation and return pituicytes to their basal shape by activating Cdc42, another small GTPase that reorganizes the actin cytoskeleton in a cortical position. Adenosine and neurohormones also have opposite actions on the efflux of taurine, a local messenger that is released by pituicytes in hypotonic conditions and accordingly inhibits vasopressin output from axon terminals. As adenosine is likely generated from endogenous ATP co‐released with neurohormones and broken down by local ectoATPases, these data suggest a subtle balance between a positive and a negative feedback on vasopressin output operated, respectively, by adenosine and vasopressin to maintain hydromineral homeostasis. A theoretical scenario is presented to account for the putative sequence of pituicyte‐related events following disturbance of the hydromineral system. © 2008 Wiley‐Liss, Inc.
Negative regulation of myelination: Relevance for development, injury, and demyelinating disease
Authors: Jessen Kristján R. , Mirsky Rhona 
Published: September 2008

AbstractDedifferentiation of myelinating Schwann cells is a key feature of nerve injury and demyelinating neuropathies. We review recent evidence that this dedifferentiation depends on activation of specific intracellular signaling molecules that drive the dedifferentiation program. In particular, we discuss the idea that Schwann cells contain negative transcriptional regulators of myelination that functionally complement positive regulators such as Krox‐20, and that myelination is therefore determined by a balance between two opposing transcriptional programs. Negative transcriptional regulators should be expressed prior to myelination, downregulated as myelination starts but reactivated as Schwann cells dedifferentiate following injury. The clearest evidence for a factor that works in this way relates to c‐Jun, while other factors may include Notch, Sox‐2, Pax‐3, Id2, Krox‐24, and Egr‐3. The role of cell–cell signals such as neuregulin‐1 and cytoplasmic signaling pathways such as the extracellular‐related kinase (ERK)1/2 pathway in promoting dedifferentiation of myelinating cells is also discussed. We also review evidence that neurotrophin 3 (NT3), purinergic signaling, and nitric oxide synthase are involved in suppressing myelination. The realization that myelination is subject to negative as well as positive controls contributes significantly to the understanding of Schwann cell plasticity. Negative regulators are likely to have a major role during injury, because they promote the transformation of damaged nerves to an environment that fosters neuronal survival and axonal regrowth. In neuropathies, however, activation of these pathways is likely to be harmful because they may be key contributors to demyelination, a situation which would open new routes for clinical intervention. © 2008 Wiley‐Liss, Inc.