bluuebunny/crossref_metadata_2025 huggingface dataset:
The hydrolytic degradation of polydioxanone (PDSII) sutures. Part II: Micromechanisms of deformation
Authors: Ping Ooi Chui , Cameron Ruth E.
Published: August 2002

An investigation of morphological changes in hydrolytically degraded polydioxanone sutures (PDSII) during in situ tensile deformation is reported. These changes were followed with the use of real‐time small‐angle X‐ray scattering with simultaneous tensile deformation experiments. The results are correlated with separately performed wide‐angle X‐ray scattering, and optical microscopy experiments on the effects of tensile strain. Interlamellar and interfibrillar amorphous regions appear to control the microdeformation and the shape of the stress‐strain curves of PDSII sutures. The fibers deform initially by interlamellar separation and at larger strains by fibrillar slip. Fractures appear to be caused by the catastrophic growth of cracks. The degradation of interlamellar tie chains in PDSII sutures does not appear to significantly affect the progress of deformation, because deformation through interlamellar separation is largely unchanged by degradation. Conversely, hydration and scission of interfibrillar tie chains lowers the load required to propagate interfibrillar shear. The fibers fail at progressively lower strains as degradation proceeds. © 2002 Wiley Periodicals, Inc. J Biomed Mater Res (Appl Biomater) 63: 291–298, 2002
Porous structure of Purevision™ versus Focus® Night&Day™ and conventional hydrogel contact lenses
Authors: López‐Alemany Antonio , Compañ Vicente , Refojo Miguel F.
Published: August 2002

The surface and bulk structures of hydrogel contact lenses that contain siloxane moieties, Purevision™ (balafilcon A) and Focus®Night&Day™ (lotrafilcon A), were investigated. Standard hydrogel lenses of low (Seequence®), medium (Acuvue®), and high water content (Precision UV®) were used as controls. All the lenses were dehydrated in a series of ethanol solutions of increased concentration, critical‐point dried in CO2, and sputter coated with gold/palladium before they were examined by scanning electron microscopy. Of all lenses examined, only the balafilcon lenses presented, in addition to the polymer network porosity characteristic of all hydrogels, a macroporous structure that was observed on the front and back surfaces, and in their bulk. The average diameter of the macropores appears to be much larger, from one to several orders of magnitude, than the network porosity of standard hydrogel lenses. The macropores might contribute to the gas and water permeability of these lenses, as well as to their mobility on the cornea. © 2002 Wiley Periodicals, Inc. J Biomed Mater Res (Appl Biomater) 63: 319–325, 2002
In vitro dissolution of melt‐derived 45S5 and sol‐gel derived 58S bioactive glasses
Authors: Sepulveda P. , Jones J. R. , Hench L. L.
Published: August 2002

Effects of powder type, particle size (5–20 μm; 90–300 μm; 90–710 μm), and type of dissolution medium on the dissolution behavior of bioactive glasses were investigated in vitro using melt‐derived 45S5 and sol‐gel derived 58S bioactive glass powders. Dissolution studies were performed in simulated body fluid and in α‐MEM based cell culture medium at 37°C under dynamic conditions (1 Hz) for periods of 30 min, 1, 2, 4, 8, 17, and 22 h. The concentrations of elements dissolved from the glasses were evaluated using inductively coupled plasma analysis. The reacted powders were analyzed for bioactivity using Fourier transform infrared spectrometry to observe the formation of a calcium phosphate layer on the surface. The non‐porous surfaces of melt‐derived 45S5 glass powders exhibited lower dissolution rates and rate of surface layer formation than 58S gel‐glass powders. The rates of dissolution for both types of powders were lower in culture medium, compared to simulated body fluid , and increased as the particle size decreased. Thus, particle size range, glass type, and powder volume fraction can be used as a means to control the release rate of active ions that stimulate the gene expression and cellular response for tissue proliferation and repair. © 2002 Wiley Periodicals, Inc. J Biomed Mater Res 61: 301–311, 2002
Regeneration of canine peroneal nerve with the use of a polyglycolic acid–collagen tube filled with laminin‐soaked collagen sponge: a comparative study of collagen sponge and collagen fibers as filling materials for nerve conduits
Authors: Toba Toshinari , Nakamura Tatsuo , Shimizu Yasuhiko , Matsumoto Kazuya , Ohnishi Katsunori , Fukuda Seijun , Yoshitani Makoto , Ueda Hiroki , Hori Yoshio , Endo Katsuaki
Published: August 2002

A novel artificial nerve conduit was developed and its efficiency was evaluated on the basis of promotion of peripheral nerve regeneration across an 80‐mm gap in dogs. The nerve conduit was made of a polyglycolic acid–collagen tube filled with laminin‐soaked collagen sponge. Conduits filled with either sponge‐ or fiber‐form collagen were implanted into an 80‐mm gap of the peroneal nerve (five dogs for each form). Twelve months postoperatively nerve regeneration was superior in the sponge group both morphometrically (percentage of neural tissue: fiber: 39.7 ± 5.2, sponge: 43.0 ± 4.5, n=3) and electrophysiologically (fiber: CMAP 1.06 ± 0.077, SEP 1.32 ± 0.127 sponge: CMAP 1.04 ± 0.106, SEP 1.24 ± 0.197, n=5), although these differences were not statistically significant. The observed regeneration was complementary to successful results reported previously in the same model, in which collagen fibers exclusively were used. The results indicate a possible superiority of collagen sponge over collagen fibers as filling materials. In addition, the mass‐producibility, superior scaffolding potential, and capacity for gradual release of soluble factors of the sponge provide make it an attractive alternative to fine fibers, which are both technologically difficult and costly to produce. This newly developed nerve conduit has the potential to enhance peripheral nerve regeneration across longer gaps commonly encountered in clinical settings. © 2001 John Wiley & Sons, Inc. J Biomed Mater Res (Appl Biomater) 58: 622–630, 2001
Tobramycin‐containing bone cement and systemic cefazolin in a one‐stage revision. Treatment of infection in a rabbit model
Authors: Nijhof Marc W. , Fleer Andre , Hardus Karin , Vogely H. Charles , Schouls Leo M. , Verbout Abraham J. , Dhert Wouter J. A.
Published: August 2002

The efficacy of tobramycin‐containing bone cement with that of systemic cefazolin for treatment of infection in a one‐stage revision model is compared. In addition, the value of detecting bacterial DNA after antibiotic treatment was investigated. An implant was inserted into the right tibia of rabbits after inoculation with Staphylococcus aureus. At 28 days, the implant was removed. Subsequently, either plain bone cement with or without systemic administration of cefazolin, or tobramycin‐containing bone cement was injected into the medullary canal. The tibiae were cultured 14 days after revision (Day 42), and showed a significant decrease in bacterial counts for both antibiotic groups compared with the control group (p≤0.05). The rate of infection in the tobramycin‐cement group was slightly higher (2/9) than in the cefazolin group (0/8), although the difference was not significant. Persistence of bacterial DNA after antibiotic treatment may be the result of delayed clearance of DNA and not a sign of active infection. This animal model shows that in a one‐stage revision tobramycin‐containing bone cement can reduce size and rate of infection, although systemic cefazolin may be more efficacious. Therefore, the use of antibiotic‐containing bone cement combined with systemic antibiotic might provide optimal treatment. © 2001 John Wiley & Sons, Inc. J Biomed Mater Res (Appl Biomater) 58: 747–753, 2001
Evaluation of the biocompatibility of a chitosan scaffold in mice
Authors: VandeVord Pamela J. , Matthew Howard W. T. , DeSilva Stephen P. , Mayton Lois , Wu Bin , Wooley Paul H.
Published: August 2002

Chitosan scaffolds appear to be suitable for a variety of tissue engineering applications. This study addressed the biocompatibility of chitosan in a mouse implantation model. Porous chitosan scaffolds were implanted in mice, and animals were sacrificed after 1, 2, 4, 8, or 12 weeks. Macroscopic inspection of the implantation site revealed no pathological inflammatory responses. Histological assessment indicated marked neutrophil accumulation within the implant, which resolved with increasing implantation time. Gram staining and limulus assays revealed no evidence of infection or endotoxin. Collagen was observed within the chitosan pore spaces, indicating that connective tissue matrix was deposited within the implant. Angiogenic activity associated with the external implant surface was also observed. Cellular immune responses were determined by lymphocyte proliferation assays, and antibody responses were measured using ELISA techniques. These assays indicated a very low incidence of chitosan‐specific reactions. Although there was a large migration of neutrophils into the implantation area, there were minimal signs of any inflammatory reaction to the material itself. This preliminary study demonstrates that chitosan has a high degree of biocompatibility in this animal model. Overall, the findings suggest that chitosan may be suitable for the development of implantable materials. © 2001 Wiley Periodicals, Inc. J Biomed Mater Res 59: 585–590, 2002
Electrically communicating three‐dimensional cardiac tissue mimic fabricated by layered cultured cardiomyocyte sheets
Authors: Shimizu Tatsuya , Yamato Masayuki , Akutsu Takumitsu , Shibata Takahiko , Isoi Yuki , Kikuchi Akihiko , Umezu Mitsuo , Okano Teruo
Published: August 2002

Recent progress in stem cell biology is likely to provide implantable sources of human cardiomyocytes in the near future. This possibility has encouraged cardiac tissue engineering. To construct heart‐like tissue, we have exploited the capabilities of novel cell culture surfaces grafted with a temperature‐responsive polymer, poly(N‐isopropylacrylamide) (PIPAAm), to produce intact viable monolayer cell sheets simply by reducing culture temperature. Cultured chick embryonic cardiomyocyte sheets detached from PIPAAm‐grafted surfaces were layered into tissue‐like laminate stacks using hydrophilic support and transfer membranes. The layered cell sheets rapidly adhered to each other, establishing cell‐to‐cell connections characteristic of heart tissue, including desmosomes and intercalated disks. Bilayer cell sheets pulsed spontaneously and synchronously, altering their characteristic pulsing frequency with applied electric stimulation transmitted across the sheets. These results demonstrate that electrically communicative three‐dimensional cardiac constructs can be achieved by stacking monolayer cardiomyocyte sheets. Cardiac tissue engineering based on this technology will facilitate new in vitro heart models and may prove useful for in vivo cardiovascular tissue repair. © 2002 Wiley Periodicals, Inc. J Biomed Mater Res 60: 110–117, 2002
Chromium(VI) interaction with plant and animal mitochondrial bioenergetics: A comparative study
Authors: Fernandes M. A. S. , Santos M. S. , Alpoim M. C. , Madeira V. M. C. , Vicente J. A. F.
Published: August 2002

The mechanism of Cr(VI)‐induced toxicity in plants and animals has been assessed for mitochondrial bioenergetics and membrane damage in turnip root and rat liver mitochondria. By using succinate as the respiratory substrate, ADP/O and respiratory control ratio (RCR) were depressed as a function of Cr(VI) concentration. State 3 and uncoupled respiration were also depressed by Cr(VI). Rat mitochondria revealed a higher sensitivity to Cr(VI), as compared to turnip mitochondria. Rat mitochondrial state 4 respiration rate triplicated in contrast to negligible stimulation of turnip state 4 respiration. Chromium(VI) inhibited the activity of the NADH‐ubiquinone oxidoreductase (complex I) from rat liver mitochondria and succinate‐dehydrogenases (complex II) from plant and animal mitochondria. In rat liver mitochondria, complex I was more sensitive to Cr(VI) than complex II. The activity of cytochrome c oxidase (complex IV) was not sensitive to Cr(VI). Unique for plant mitochondria, exogenous NADH uncoupled respiration was unaffected by Cr(VI), indicating that the NADH dehydrogenase of the outer leaflet of the plant inner membrane, in addition to complexes III and IV, were insensitive to Cr(VI). The ATPase activity (complex V) was stimulated in rat liver mitochondria, but inhibited in turnip root mitochondria. In both, turnip and rat mitochondria, Cr(VI) depressed mitochondrial succinate‐dependent transmembrane potential (Δψ) and phosphorylation efficiency, but it neither affected mitochondrial membrane permeabilization to protons (H+) nor induced membrane lipid peroxidation. However, Cr(VI) induced mitochondrial membrane permeabilization to K+, an effect that was more pronounced in turnip root than in rat liver mitochondria. In conclusion, Cr(VI)‐induced perturbations of mitochondrial bioenergetics compromises energy‐dependent biochemical processes and, therefore, may contribute to the basal mechanism underlying its toxic effects in plant and animal cells. © 2002 Wiley Periodicals, Inc. J Biochem Mol Toxicol 16:53–63, 2002; Published online in Wiley Interscience (www.interscience.wiley.com). DOI 10.1002/jbt.10025
Four patients with both thrombotic thrombocytopenic purpura and autoimmune thrombocytopenic purpura: The concept of a mixed immune thrombocytopenia syndrome and indications for plasma exchange
Authors: Baron Beverly W. , Martin Marisa Saint , Sucharetza Brian S. , Jeon Hye‐Ran , Baron Joseph M.
Published: August 2002

Autoimmune thrombocytopenic purpura (ATP) and thrombotic thrombocytopenic purpura (TTP) are each well recognized clinical syndromes which may appear as single episodes or may have chronic relapsing courses. We present four patients negative for human immunodeficiency virus (HIV) infection who appear to have both diagnoses with either concomitant or intermingled episodes, and we review seven additional patients reported in the literature with similar features. All four of our patients are female, two have underlying connective tissue disorders, and their ATP processes came to our attention because of incomplete response of the platelet count to plasma exchange therapy (PEX) during a TTP phase (Cases 1 and 2) or development of thrombocytopenia in the absence of microangiopathy on the background of prior typical TTP episodes (Cases 3 and 4). Recognition of the ATP diagnosis in each case resulted in discontinuation of PEX (Cases 1 and 2) or not instituting PEX (Cases 3 and 4). In each instance, a satisfactory rise in platelet count followed treatment for ATP. Based upon this experience, we conclude that some individuals may have a mixed immune thrombocytopenia syndrome; careful analysis of the mechanism of thrombocytopenia, especially in recurrent episodes and in patients who respond incompletely to PEX for TTP, is important when deciding whether to initiate or continue PEX, or to consider therapies appropriate for other mechanisms of thrombocytopenia. J. Clin. Apheresis 16:179–185, 2001. © 2001 Wiley‐Liss, Inc.
Ethanolamine is a co‐mitogenic factor for proliferation of primary hepatocytes
Authors: Ajioka Itsuki , Akaike Toshihiro , Watanabe Yoshifumi
Published: August 2002

Mature adult parenchymal hepatocytes can enter the S phase in the presence of growth factors such as HGF and EGF, but rarely proliferate in culture. We hypothesized that the cell cycle of hepatocytes in culture is restricted before G2/M phase and we attempted to identify the factor that induces cell cycle progression. We found that the conditioned medium from long‐term cultured hepatocytes contained co‐mitogenic activity with other growth factors, which was attributed to ethanolamine (Etn). Etn induced not only DNA synthesis but also cell replication of cultured hepatocytes with various other growth factors. Etn and HGF synergistically induced cyclin D1, A and B expression, however, only cyclin B but not cyclin A formed a complex with Cdc2. In addition, Etn combined with HGF enhanced PKCβII expression and translocated PKCβII to the plasma membrane, and induced filopodia formation, which was inhibited by an antisense oligonucleotide against PKCβII. In addition, blocking the cytoskeleton rearrangement with inhibitors (colchicine, cytochalasin D, or chlerythrine (a specific PKC inhibitor)) inhibited cyclin expression and cell proliferation. Although Etn enhanced the downstream product, cellular phosphatidylethanolamine (PE), PE itself did not show any Etn‐like activities on hepatocytes. Taken together, our results indicate that Etn functions as a co‐replication factor to promote the cell cycle of mature hepatocytes to G2/M phase in the presence of growth factors. The activity is thought to be mediated by PKCβII‐dependent cyclin B expression. J. Cell. Biochem. 84: 249–263, 2002. © 2001 Wiley‐Liss, Inc.
Differential maintenance and de novo methylating activity by three DNA methyltransferases in aging and immortalized fibroblasts
Authors: Lopatina Nadejda , Haskell Joyce F. , Andrews Lucy G. , Poole Joseph C. , Saldanha Sabita , Tollefsbol Trygve
Published: August 2002

Genomic methylation, which influences many cellular processes such as gene expression and chromatin organization, generally declines with cellular senescence although some genes undergo paradoxical hypermethylation during cellular aging and immortalization. To explore potential mechanisms for this process, we analyzed the methylating activity of three DNA methyltransferases (Dnmts) in aging and immortalized WI‐38 fibroblasts. Overall maintenance methylating activity by the Dnmts greatly decreased during cellular senescence. In immortalized WI‐38 cells, maintenance methylating activity was similar to that of normal young cells. Combined de novo methylation activity of the Dnmts initially decreased but later increased as WI‐38 cells aged and was strikingly elevated in immortalized cells. To further elucidate the mechanisms for changes in DNA methylation in aging and immortalized cells, the individual Dnmts were separated and individually assessed for maintenance and de novo methylating activity. We resolved three Dnmt fractions, one of which was the major maintenance methyltransferase, Dnmt1, which declined steadily in activity with cellular senescence and immortalization. However, a more basic Dnmt, which has significant de novo methylating activity, increased markedly in activity in aging and immortalized cells. We have identified this methyltransferase as Dnmt3b which has an important role in neoplastic transformation but its role in cellular senescence and immortalization has not previously been reported. An acidic Dnmt we isolated also had increased de novo methylating activity in senescent and immortalized WI‐38 cells. These studies indicate that reduced genome‐wide methylation in aging cells may be attributed to attenuated Dnmt1 activity but that regional or gene‐localized hypermethylation in aging and immortalized cells may be linked to increased de novo methylation by Dnmts other than the maintenance methyltransferase. J. Cell. Biochem. 84: 324–334, 2002. © 2001 Wiley‐Liss, Inc.
Functional receptor for platelet‐derived growth factor in rat embryonic heart‐derived myocytes: Role of sequestered Ca2+ stores in receptor signaling and antagonism by arginine vasopressin
Authors: Brostrom Margaret A. , Meiners Sally , Brostrom Charles O.
Published: August 2002

Platelet‐derived growth factor (PDGF) is established to function importantly in the growth, development, and function of most cardiovascular tissues. However, evidence that the factor participates directly in the growth and development of the mammalian myocardium is lacking. H9c2 rat embryonic ventricular myocytes were found to respond to PDGF‐BB with a rapid mobilization of cell‐associated Ca2+ and increased rates of protein synthesis, followed by markedly increased rates of DNA synthesis. PDGF acted as a full mitogen for these myocytes. Evidence is provided that documents the expression of classical PDGF‐β, but not PDGF‐α, receptors in H9c2 cells. Scatchard analysis revealed the presence of 44,000 β‐receptors per myocyte. Cell shortening and clustering of plasmalemmal β‐receptors occurred within 30 min of exposure to PDGF‐BB. Treatment was also associated with a transient increase in the rate of synthesis of GRP78/BiP, consistent with a transitory release of Ca2+ from the sarcoplasmic/endoplasmic reticulum [S(E)R]. Increased rates of protein synthesis at early times of PDGF treatment were additive with those occurring in response to arginine vasopressin, indicating different mechanisms of translational upregulation by these agents. The mitogenic effects of PDGF were delayed by vasopressin, which causes H9c2 myocytes to undergo hypertrophy while promoting the persistent depletion of S(E)R Ca2+ stores. In the presence of PDGF, vasopressin did not induce hypertrophy. As compared to untreated myocytes, DNA synthesis in PDGF‐treated myocytes was optimized at lower extracellular Ca2+ concentrations and was significantly less sensitive to inhibition by ionomycin. H9c2 cells appear to provide a useful embryonic cardiomyocyte model in which to examine both PDGF‐activated proliferative and vasopressin‐activated hypertrophic events and the importance of transient vs. sustained Ca2+ release in these events. J. Cell. Biochem. 84: 736–749, 2002. © 2002 Wiley‐Liss, Inc.
Nuclear localization and DNA interaction of protein disulfide isomerase ERp57 in mammalian cells
Authors: Coppari Sabina , Altieri Fabio , Ferraro Anna , Chichiarelli Silvia , Eufemi Margherita , Turano Carlo
Published: August 2002

Protein disulfide isomerase ERp57 is localized predominantly in the endoplasmic reticulum, but is also present in the cytosol and, according to preliminary evidence, in the nucleus of avian cells. Conclusive evidence of its nuclear localization and of its interaction with DNA in vivo in mammalian cells is provided here on the basis of DNA–protein cross‐linking experiments performed with two different cross‐linking agents on viable HeLa and 3T3 cells. Nuclear ERp57 could also be detected by immunofluorescence in HeLa cells, where it showed an intracellular distribution clearly different from that of an homologous protein, located exclusively in the endoplasmic reticulum. Mammalian ERp57 resembles the avian protein in its recognition of S/MAR‐like DNA sequences and in its association with the nuclear matrix. It can be hypothesized that ERp57, which is known to associate with other proteins, in particular STAT3 and calreticulin, may contribute to their nuclear import, DNA binding, or other functions that they fulfil inside the nucleus. J. Cell. Biochem. 85: 325–333, 2002. © 2002 Wiley‐Liss, Inc.
Mapping of intrinsic bent DNA sites in the upstream region of DNA puff BhC4‐1 amplified gene
Authors: Fiorini Adriana , Basso Luiz Roberto , Paçó‐Larson Maria Luisa , Fernandez Maria Aparecida
Published: August 2002

We have identified bent DNA sites in the distal and proximal DNA puff BhC4‐1 amplified gene promoter region of Bradysia hygida. The 2D modeling of the 3D DNA path and the ENDS ratio values calculated in this promoter region resulted in the identification of ten pronounced bent sites named BhC4B − 9 to + 1. The 1847 bp fragment (− 3697 to − 1850) in relation to the transcription start site shows multiple bending sites, BhC4B − 9 to BhC4B − 4, with periodicity ∼300 bp. The analysis of the other identified bent region, starting at position − 957, reveals that the BhC4B + 1 bent site colocalizes with the putative BhC4‐1 minimal promoter. The sequence analysis of bent site BhC4B − 4 shows a distribution of dA•dT at ∼10 bp intervals between the middle of each tract, but intervals with more than one turn, ∼20 bp, two helix turns, were detected in the other bent sites described here. The bent sites BhC4B − 6 and BhC4B − 4, contain two consensus sequences, with 60 bp each. The apparent molecular weight of fragments in the BhC4‐1 promoter region were estimated in agarose gels and compared with the data obtained in polyacrylamide gels without and with ethidium bromide. The mobility reduction ratios (R‐values) were determined, and a high R‐value, 1.80, for a 1215 bp fragment in the distal promoter region and a 1.23 significant R‐value for a 662 bp fragment in the proximal segment were found. To further analyze the predicted bent DNA sites in these fragments, the 2D trajectories of the 3D DNA path and other parameters, AT percentage, roll angle, ENDS ratio and ΔG, were determined. The role of these bent sites in the BhC4‐1 transcription regulation is discussed. © 2001 Wiley‐Liss, Inc.
Protein tyrosine phosphatase SHP‐1 specifically recognizes C‐terminal residues of its substrates via helix α0
Authors: Yang Jian , Cheng Zhiliang , Niu Tianqi , Liang Xiaoshan , Zhao Zhizhuang Joe , Zhou G. Wayne
Published: August 2002

The catalytic domain of protein tyrosine phosphatase SHP‐1 possesses distinct substrate specificity. It recognizes the P‐3 to P‐5 residues of its substrates via the β5‐loop‐β6 region. To study the substrate specificity further, we determined the structure of the catalytic domain of SHP‐1 (C455S) complexed with a less‐favorable‐substrate peptide originated from SIRPα. The complex has disordered N‐terminal peptide structure and reduced interactions between the N‐terminal peptide and the β5‐loop‐β6 region. This could be the basis for the lower affinity of peptide pY427 for the catalytic domain of SHP‐1. In addition, by comparing the SHP‐1/less‐favorable peptide complex structure with the SHP‐1/substrate complex structures, we identified a novel substrate‐recognition site in the catalytic domain of SHP‐1. This site was formed by helix α0 and the α5‐loop‐α6 motif of SHP‐1, and specifically bound residues at the P + 4 and further C‐terminal positions of peptide substrates. © 2001 Wiley‐Liss, Inc.
Assessment of promoter elements of the germ cell‐specific proacrosin gene
Authors: Schulten Hans‐Jürgen , Nayernia Karim , Reim Kerstin , Engel Wolfgang , Burfeind Peter
Published: August 2002

The testis‐specific proacrosin gene encodes for a fertilization‐promoting protein. In mouse and rat it is first transcribed in late pachytene spermatocytes and revealed to be translationally regulated. Former proacrosin promoter studies demonstrated that elements necessary for conducting a stage and temporal‐specific expression of the gene are located within 0.9 kb upstream of the translational start codon. In the present study we analyzed putative cis‐acting elements located in this promoter region for their specific binding properties to nuclear factors assumed to be involved in proacrosin gene regulation. Supplement of specific antibodies in electrophoretic mobility shift assays (EMSA) revealed that two Y‐box proteins and the transcription factors CREM and YY1 interact with proacrosin promoter elements. The Y‐box proteins, antigenically related to the frog Y‐box proteins FRGY1 and FRGY2, bound to the Y‐box (55–66 bp upstream of the ATG initiation codon) in brain and testis nuclear extracts, respectively. CREM bound to three elements (30–37, 252–259, and 717–724 bp upstream of ATG). The ubiquitous transcription factor YY1 bound to a conserved element in the central proacrosin promoter (457–473 bp upstream of ATG) and showed almost germ cell‐specific truncates in EMSA. These results suggest that the identified factors are involved in proacrosin gene regulation. J. Cell. Biochem. 83: 155–162, 2001. © 2001 Wiley‐Liss, Inc.
Phosphoinositide‐specific phospholipase Cβ1 expression is not linked to nerve growth factor‐induced differentiation, cell survival or cell cycle control in PC12 rat pheocromocytoma cells
Authors: Bortul Roberta , Aluigi Michela , Tazzari Pier Luigi , Tabellini Giovanna , Baldini Giovanna , Bareggi Renato , Narducci Paola , Martelli Alberto M.
Published: August 2002

Recent reports have highlighted that phosphoinositide‐specific phospholipase Cβ1 expression is linked to neuronal differentiation in different experimental models. We sought to determine whether or not this is also true for nerve growth factor (NGF)‐induced neuronal differentiation of rat PC12 cells. However, we did not find differences in the expression of both the forms of phosphoinositide‐specific phospholipase Cβ1 (a and b) during sympathetic differentiation of these cells. Also, PC12 cell clones stably overexpressing phosphoinositide‐specific phospholipase Cβ1 were not more susceptible to the differentiating effect of NGF. Furthermore, since it is well established that phosphoinositide‐specific phospholipase Cβ1 affects cell proliferation, we investigated whether or not PC12 cell clones stably overexpressing phosphoinositide‐specific phospholipase Cβ1 showed differences in survival to serum deprivation and cell cycle, when compared to wild type cells. Nevertheless, we did not find any differences in these parameters between wild type cells and the overexpressing clones. Interestingly, in PC12 cells the overexpressed phosphoinositide‐specific phospholipase Cβ1 did not localize to the nucleus, but by immunofluorescence analysis, was detected in the cytoplasm. Therefore, our findings may represent another important clue to the fact that only when it is located within the nucleus phosphoinositide‐specific phospholipase Cβ1 is able to influence cell proliferation. J. Cell. Biochem. 84: 56–67, 2002. © 2001 Wiley‐Liss, Inc.
Role of bifurcation in the bond shifting of cyclooctatetraene
Authors: Castaño Obis , Palmeiro Raúl , Frutos Luis Manuel , Luisandrés José
Published: August 2002

The present study of the cyclooctatetraene potential energy surface shows the presence of a bifurcation (valley ridge inflection point) in the intrinsic reaction coordinate path between the two transition states of D8h and D4h symmetries. This result is of capital importance for the correct understanding of the bond shifting and ring inversion processes in this compound. © 2002 Wiley Periodicals, Inc. J Comput Chem 23: 732–736, 2002
Relativistic energy‐consistent pseudopotentials—Recent developments
Authors: Stoll Hermann , Metz Bernhard , Dolg Michael
Published: August 2002

The direct adjustment of two‐component pseudopotentials (scalar‐relativistic + spin‐orbit potentials), to atomic total energy valence spectra derived from four‐component multiconfiguration Dirac–Hartree–Fock all‐electron calculations based on the Dirac–Coulomb–Breit Hamiltonian, has been made a routine tool for an efficient treatment of heavy main‐group elements. Both large‐core (nsp valence shell) and small‐core ((n − 1)spd nsp valence shell) potentials have been generated for all the post‐d elements of groups 13–17. At the example of lead and bismuth compounds (PbHal, BiH, BiO, BiHal (Hal = F, Cl, Br, I)), we show how small‐core and large‐core potentials can be combined in accurate, yet computationally economic, spin‐free‐state‐shifted relativistic electronic structure calculations of molecular ground and excited states. © 2002 Wiley Periodicals, Inc. J Comput Chem 23: 767–778, 2002
Spin‐orbit coupling of DFT/MRCI wavefunctions: Method, test calculations, and application to thiophene
Authors: Kleinschmidt Martin , Tatchen Jörg , Marian Christel M.
Published: August 2002

During the past decade the one‐center mean‐field approximation has proven to be a very appropriate framework for the accurate description of spin‐orbit effects at the correlated all‐electron level. Here, a new efficient code, SPOCK, is introduced that calculates spin‐orbit matrix elements in the one‐center mean‐field approximation for multireference CI wave functions. For the first time, the computation of spin‐dependent interactions within a Kohn‐Sham orbital based CI (DFT/MRCI) scheme1 is made possible. The latter approach is suitable for large scale systems with up to 100–200 valence electrons. Test calculations are performed on well‐known diatomic molecules and the thiocarbonyl pyranthione. Spin‐orbit matrix elements show good agreement with their Hartree‐Fock orbital based counterparts but are obtained at considerably lower expense, thus demonstrating the power of the method. As an application singlet‐triplet couplings in thiophene are investigated that are important for the photophysics and photochemistry. Spin‐orbit matrix elements between all π → π* excited states are found to be small. Considerably larger spin‐orbit matrix elements are observed only for cases in which π → σ* excited configurations are involved. © 2002 Wiley Periodicals, Inc. J Comput Chem 23: 824–833, 2002
Calculation of stacking geometries by a modified MM2 method
Authors: Kamishima Mayumi , Kojima Masaaki , Yoshikawa Yuzo
Published: August 2002

The MM2 method has been modified using a three‐center charge model aimed at determining stacking geometries. The stacking interactions for the benzene dimer, porphine dimer, and bis(4,5‐diazafluoren‐9‐one thiosemicarbazonato)palladium(II) have been assessed with both the three‐center charge model and the conventional single‐center charge model using the MM2 method. The modified MM2 method always gave better results than the conventional method, and the obtained structures by the former method are in good agreement with the proposed structures from those theoretical and experimental results. The possible advantages of using the three‐center charge model for complex systems are also discussed. © 2001 John Wiley & Sons, Inc. J Comput Chem 22: 835–845, 2001
Real‐time identification of Pseudomonas aeruginosa direct from clinical samples using a rapid extraction method and polymerase chain reaction (PCR)
Authors: Jaffe Richard I. , Lane Janae D. , Bates Christopher W.
Published: August 2002

Pseudomonas aeruginosa has emerged as one of the most problematic Gram‐negative nosocomial pathogens. Bacteremia caused by P. aeruginosa is clinically indistinguishable from other Gram‐negative infections although the mortality rate is higher. This microorganism is also inherently resistant to common antibiotics. Standard bacterial identification and susceptibility testing is normally a 48‐hour process and difficulty sometimes exists in rapidly and accurately identifying antimicrobial resistance. The Polymerase Chain Reaction (PCR) is a rapid and simple process for the amplification of target DNA sequences. However, many sample preparation methods are unsuitable for the clinical laboratory because they are not cost effective, take too long to perform, or do not provide a good template for PCR. Our goal was to provide same‐day results to facilitate rapid diagnosis. In this report, we have utilized our rapid DNA extraction method to generate bacterial DNA direct from clinical samples for PCR. The lower detection level for P. aeruginosa was estimated to be 10 CFU/ml. In addition, we wanted to compare the results of a new rapid‐cycle DNA thermocycler that uses continuous fluorescence monitoring with the results of standard thermocycling. We tested 40 clinical isolates of P. aeruginosa and 18 non‐P. aeruginosa isolates received in a blinded fashion. Coded data revealed that there was 100% correlation in both the rapid‐cycle DNA thermocycling and standard thermocycling when compared to standard clinical laboratory results. In addition, total results turn‐around time was less than 1 hour. Specific identification of P. aeruginosa was determined using intragenic primer sets for bacterial 16S rRNA and Pseudomonas outer‐membrane lipoprotein gene sequences. The total cost of our extraction method and PCR was $2.22 per sample. The accuracy and rapidness of this DNA‐extraction method, with its PCR‐based identification system, make it an ideal candidate for use in the clinical laboratory. J. Clin. Lab. Anal. 15:131–137, 2001. © 2001 Wiley‐Liss, Inc.
Multiplex polymerase chain reaction: A practical approach
Authors: Markoulatos P. , Siafakas N. , Moncany M.
Published: August 2002

Considerable time and effort can be saved by simultaneously amplifying multiple sequences in a single reaction, a process referred to as multiplex polymerase chain reaction (PCR). Multiplex PCR requires that primers lead to amplification of unique regions of DNA, both in individual pairs and in combinations of many primers, under a single set of reaction conditions. In addition, methods must be available for the analysis of each individual amplification product from the mixture of all the products. Multiplex PCR is becoming a rapid and convenient screening assay in both the clinical and the research laboratory. The development of an efficient multiplex PCR usually requires strategic planning and multiple attempts to optimize reaction conditions. For a successful multiplex PCR assay, the relative concentration of the primers, concentration of the PCR buffer, balance between the magnesium chloride and deoxynucleotide concentrations, cycling temperatures, and amount of template DNA and Taq DNA polymerase are important. An optimal combination of annealing temperature and buffer concentration is essential in multiplex PCR to obtain highly specific amplification products. Magnesium chloride concentration needs only to be proportional to the amount of dNTP, while adjusting primer concentration for each target sequence is also essential. The list of various factors that can influence the reaction is by no means complete. Optimization of the parameters discussed in the present review should provide a practical approach toward resolving the common problems encountered in multiplex PCR (such as spurious amplification products, uneven or no amplification of some target sequences, and difficulties in reproducing some results). Thorough evaluation and validation of new multiplex PCR procedures is essential. The sensitivity and specificity must be thoroughly evaluated using standardized purified nucleic acids. Where available, full use should be made of external and internal quality controls, which must be rigorously applied. As the number of microbial agents detectable by PCR increases, it will become highly desirable for practical purposes to achieve simultaneous detection of multiple agents that cause similar or identical clinical syndromes and/or share similar epidemiological features. J. Clin. Lab. Anal. 16:47–51, 2002. © 2002 Wiley‐Liss, Inc.
Ganglioside agglutination immunoassay for rapid detection of autoantibodies in immune‐mediated neuropathy
Authors: Alaedini Armin , Wirguin Itzhak , Latov Norman
Published: August 2002

Elevated levels of serum autoantibodies directed against gangliosides are closely associated with acute and chronic autoimmune neuropathies. An agglutination immunoassay using polystyrene microparticles coated with a total extract of brain gangliosides was used to test patient sera for the presence of anti‐ganglioside antibodies. Results were compared with those obtained by ELISA for anti‐GM1 and anti‐GQ1b ganglioside antibodies. Eight of the twelve sera from patients with multifocal motor neuropathy and seven of the thirteen sera from patients with Guillain‐Barré syndrome were positive for the presence of anti‐ganglioside antibodies by the ganglioside agglutination immunoassay. The assay compared favorably with the ELISA system in sensitivity and specificity, while requiring a fraction of the time and cost to perform. The new assay can serve as a rapid and effective method for detecting or screening for anti‐ganglioside antibodies in patients with acute or chronic immune‐mediated neuropathies. It would be particularly useful for detecting antibodies that react with multiple gangliosides, or with minor or as yet uncharacterized gangliosides. J. Clin. Lab. Anal. 15:96–99, 2001. © 2001 Wiley‐Liss, Inc.
Guidelines for enhancing homework compliance
Authors: Tompkins Michael A.
Published: August 2002

The ability to enhance compliance with therapeutic tasks such as homework may be a crucial therapeutic skill that determines the success of psychotherapy in real‐world clinical settings. This article presents guidelines for enhancing homework compliance through careful attention to: features of homework assignments, such as whether they are meaningful and doable and include a clear rationale and backup plan; and the therapist's manner when developing and reviewing homework assignments, such as whether that therapist is curious, collaborative, and reinforces all behavior that supports the homework's purposes. Clinical examples are used to illustrate the application of the guidelines. © 2002 Wiley Periodicals, Inc. J Clin Psychol/In Session 58: 565–576, 2002
Pretreatment correlates of the therapeutic bond
Authors: Saunders Stephen M.
Published: August 2002

This study examined the association between pretreatment client characteristics and the therapeutic bond, which is defined as the emotional aspect of the therapeutic alliance. Three types of interpersonal problems and four indicators of emotional distress were measured and compared to the therapeutic bond and its subscales (role investment, empathic resonance, and mutual affirmation) measured early in treatment. Interpersonal problems related to being overly detached were associated with a poorer quality bond and, in particular, with a tendency to feel as if the therapist does not understand what the client is thinking and feeling (lower empathic resonance). Higher levels of distress were correlated negatively with the therapeutic bond. Analyses indicated that interpersonal problems were better predictors of the bond than was distress. The relevance of these findings for practitioners is discussed. © 2001 John Wiley & Sons, Inc. J Clin Psychol 57: 1339–1352, 2001.
Who benefits most from a broadly targeted prevention program? Differential efficacy across populations in the Teen Outreach program
Authors: Allen Joseph P. , Philliber Susan
Published: August 2002

This study examined a highly successful, well‐documented, national program to prevent teenage pregnancy and school failure—the Teen Outreach program—to address a fundamental question: How well can a developmentally focused, broadly targeted prevention program address the needs of those students within the program who are at the highest risk of problematic behavior. The hypothesis that the developmental focus of a broadly targeted intervention would lead it to have greater program efficacy among those young people who began the program at greatest risk was examined with multisite data collected on more than 3,300 Teen Outreach and comparison group students. Results confirmed prior findings regarding the overall efficacy of the Teen Outreach program, and indicated that the program appeared most effective for those students at greatest initial risk of the problem behaviors being targeted. Implications for the targeting of the Teen Outreach program specifically and of similar primary prevention programs more generally are discussed. © 2001 John Wiley & Sons, Inc.
Extracellular matrix regulates induction of alkaline phosphatase expression by ascorbic acid in human fibroblasts
Authors: Abe Tatsuya , Abe Yukiko , Aida Yoshitomi , Hara Yoshitaka , Maeda Katsumasa
Published: August 2002

During wound healing and inflammation, fibroblasts express elevated alkaline phosphatase (ALP), but are not in contact with collagen fibrils in the fibronectin (FN)‐rich granulation tissue. We hypothesized that the extracellular matrix (ECM) environment might influence the induction of ALP in fibroblasts. Here we tested this hypothesis by studying the ALP‐inductive response of normal human gingival fibroblasts to ascorbic acid (AsA). AsA induced ALP activity and protein in cells in conventional monolayer culture. This induction was inhibited by blocking‐antibodies to the FN receptor α5β1 integrin and by the proline analog 3,4‐dehydroproline (DHP). DHP prevented cells from arranging FN fibrils into a pericellular network and reduced the activity of cell spreading on FN. Plating of cells on FN facilitated the up‐regulation by AsA of ALP expression, but did not substitute for AsA. In contrast, AsA did not cause ALP induction in cells cultured on and in polymerized type I collagen gels. Collagen fibrils inhibited the up‐regulation by AsA of ALP expression in cells plated on FN. These results indicate that the ECM regulates the induction of ALP expression by AsA in fibroblasts: FN enables them to express ALP in response to AsA through interaction with integrin α5β1, whereas type I collagen fibrils cause the suppression of ALP expression and overcome FN. © 2001 Wiley‐Liss, Inc.
Evidence for a physiological role for membrane rafts in human platelets
Authors: Gousset Karine , Wolkers Willem F. , Tsvetkova Nelly M. , Oliver Ann E. , Field Cara L. , Walker Naomi J. , Crowe John H. , Tablin Fern
Published: August 2002

We have investigated raft formation in human platelets in response to cell activation. Lipid phase separation and domain formation were detected using the fluorescent dye 1,1′‐dioctadecyl‐3,3,3′,3′‐tetramethyl‐indocarbocyanine perchlorate (diI‐C18) that preferentially partitions into gel‐like lipid domains. We showed that when human platelets are activated by cold and physiological agonists, rafts coalesce into visible aggregates. These events were disrupted by depletion of membrane cholesterol. Using Fourier transform infrared spectroscopy (FTIR), we measured a thermal phase transition at around 30°C in intact platelets, which we have assigned as the liquid‐ordered to the liquid‐disordered phase transition of rafts. Phase separation of the phospholipid and the sphingomyelin‐enriched rafts could be observed as two phase transitions at around 15 and 30°C, respectively. The higher transition, assigned to the rafts, was greatly enhanced with removal of membrane cholesterol. Detergent‐resistant membranes (DRMs) were enriched in cholesterol (50%) and sphingomyelin (20%). The multi‐functional platelet receptor CD36 selectively partitioned into DRMs, whereas the GPI‐linked protein CD55 and the major platelet integrin αIIbβ3a did not, which suggests that the clustering of proteins within rafts is a regulated process dependent on specific lipid protein interactions. We suggest that raft aggregation is a dynamic, reversible physiological event triggered by cell activation. J. Cell. Physiol. 190: 117–128, 2002. © 2002 Wiley‐Liss, Inc.
Role of Id family proteins in growth control
Authors: Yokota Yoshifumi , Mori Seiichi
Published: August 2002

Id proteins (inhibitors of DNA binding/differentiation) are negative regulators of basic helix‐loop‐helix (bHLH) type transcription factors, which promote the differentiation of various cell types. In addition to their “classical” ability to inhibit cell differentiation, they are able to stimulate cell cycle progression. These facts suggest that Id proteins play a role in keeping precursor cells immature and in expanding the cell population size during development. In vitro as well as in vivo analyses in the last several years have shown that Id proteins have more complex activities; they induce apoptosis or function as survival factors, depending on the cell context. Furthermore, dysregulated expression of Id proteins has been reported in several human tumors and seems to be related to the malignant character of tumors. Here, we summarize and discuss the biological activities of Id proteins from the standpoint of cell growth control. J. Cell. Physiol. 190: 21–28, 2002. © 2002 Wiley‐Liss, Inc.