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  • Springer  (25)
  • American Institute of Physics  (6)
  • American Physical Society  (1)
  • 1995-1999  (32)
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Year
  • 1
    ISSN: 1432-0827
    Keywords: Key words: Transfection — Gene transfer — Adenovirus — Polylysine-conjugate — Skeletal cells.
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Physics
    Notes: Abstract. In order to assess efficient DNA gene transfer into human primary cell cultures derived from the skeleton we tested two viral-based procedures. First, replication-deficient recombinant adenoviruses (ADV) were used to infect post-confluent human marrow stromal fibroblasts (HMSF) and human trabecular bone (HTB) cells. Both cell types were readily infected by modified adenoviral vectors carrying a reporter gene making this virus an attractive candidate to facilitate DNA gene transfer. In a second approach we coincubated DNA with ADV that had polylysine (PLL) covalently attached. With this ADV/PLL/DNA complex, very efficient gene transfer into multilayered HMSF and HTB cell cultures was observed, and DNA coincubated with unmodified ADV failed to be effectively transferred. These data imply that the covalently bound PLL more effectively binds exogenous DNA, resulting in a highly efficient internalization event in both cell types. Thus, this latter method has many advantages over conventional ADV gene transfer procedures. It is simple, rapid, and it does not require engineering of DNA into the viral genome, thereby allowing transfer of large fragments of DNA.
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  • 2
    Electronic Resource
    Electronic Resource
    Springer
    Journal of molecular evolution 49 (1999), S. 769-779 
    ISSN: 1432-1432
    Keywords: Key Words: Branchiopods — Hemoglobin — Concerted evolution — Crustaceans — Introns
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract. The extracellular hemoglobins of cladocerans derive from the aggregation of 12 two-domain globin subunits that are apparently encoded by four genes. This study establishes that at least some of these genes occur as a tandem array in both Daphnia magna and Daphnia exilis. The genes share a uniform structure; a bridge intron separates two globin domains which each include three exons and two introns. Introns are small, averaging just 77 bp, but a longer sequence (2.2–3.2 kb) separates adjacent globin genes. A survey of structural diversity in globin genes from other daphniids revealed three independent cases of intron loss, but exon lengths were identical, excepting a 3-bp insertion in exon 5 of Simocephalus. Heterogeneity in the extent of nucleotide divergence was marked among exons, largely as a result of the pronounced diversification of the terminal exon. This variation reflected, in part, varying exposure to concerted evolution. Conversion events were frequent in exons 1–4 but were absent from exons 5 and 6. Because of this difference, the results of phylogenetic analyses were strongly affected by the sequences employed in this construction. Phylogenies based on total nucleotide divergence in exons 1–4 revealed affinities among all genes isolated from a single species, reflecting the impact of gene conversion events. In contrast, phylogenies based on total nucleotide divergence in exons 5 and 6 revealed affinities among orthologous genes from different taxa.
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  • 3
    ISSN: 1432-0827
    Keywords: Key words: Bone sialoprotein — UMR106-01 BSP — YY1 motif — CCAAT — TATA motif.
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Physics
    Notes: Abstract. Bone sialoprotein (BSP) is a noncollagenous matrix glycoprotein localized predominantly in mineralized tissues but also detected in extraskeletal sites undergoing focal mineralization. We have previously characterized the human BSP gene and have shown that the upstream sequence contains inverted TATA and CCAAT motifs at the expected locations from the transcriptional start site (J. M. Kerr et al. [13]) and a potential YY1 binding motif located within the first 30 bp of intron 1 of the human gene. Deletion analyses of the human BSP promoter/exon 1 sequence fused to a CAT reporter gene indicate that CCAAT enhances basal transcription of BSP in transiently transfected rat UMR106-01 BSP osteosarcoma and rat skin fibroblasts. Though this enhancing activity was lost with inclusion of 68 bp of intron containing a YY1 motif in these constructs, reporter activity in the UMR106-01-BSP cells was elevated four- to seven-fold relative to that of rat fibroblasts. Gel electrophoretic mobility shift, UV-crosslinking, and southwestern experiments indicate that YY1 is present only in the extracts of nuclei isolated from the UMR cells and may contribute to the elevated transcriptional activity of the human BSP promoter construct in UMR106-01-BSP.
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  • 4
    Electronic Resource
    Electronic Resource
    Springer
    European journal of clinical pharmacology 54 (1998), S. 567-571 
    ISSN: 1432-1041
    Keywords: Key words Troglitazone ; Hepatic insufficiency
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology , Medicine
    Notes: Abstract Objective: Troglitazone is an agonist of the peroxisome proliferator-activated receptor-γ (PPAR-γ), which has been shown to improve the metabolic control of type 2 diabetes. Troglitazone undergoes hepatic metabolism to an inactive sulphate conjugate and an oxidative quinone metabolite with minor activity. The objective of this study was to compare the pharmacokinetics of troglitazone in patients with hepatic insufficiency and normal subjects. Methods: Three groups of eight subjects with normal liver function and moderate or severe hepatic impairment (Pugh-Child classification) completed this open study. Subjects received a single 400-mg dose of troglitazone 30 min after breakfast. Plasma concentrations of troglitazone and its metabolites were measured and standard pharmacokinetic parameters derived. Results: A 46% increase in area under the plasma concentration-time curve (AUClast) was observed for troglitazone, together with a 154% increase for the quinone metabolite in the patients with moderate hepatic impairment compared with normal subjects, but these did not reach statistical significance. Corresponding increases of 18% and 53% in the severe group also failed to reach statistical significance. For the sulphate conjugate, the AUClast values for both moderate and severe hepatic impairment were in the order of fourfold higher than those in the normal group. There were reductions in the maximum observed plasma concentration (Cmax) of troglitazone to 61% of the normal group in the severe group for troglitazone, and twofold increases in sulphate metabolite Cmax in the moderate and severe groups. There was an approximately threefold increase in the half-life of the sulphate conjugate in subjects with both moderate and severe impairment of liver function compared with normal individuals. First times to maximum concentrations of troglitazone, its sulphate conjugate and the quinone metabolite were significantly longer in all severely impaired subjects compared with those with normal hepatic function, although the range was wide in all cases. Plasma protein binding was high in all subjects measured (mean unbound fraction range 0.7–5.1%), but there were insufficient samples to compare across groups. Conclusion: The formation of metabolites of troglitazone following a single dose is not impaired in the presence of reduced liver function although the capacity to eliminate the metabolites is altered. The clinical significance of the effect of liver disease on the conjugates is not clear.
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  • 5
    Electronic Resource
    Electronic Resource
    Springer
    European journal of clinical pharmacology 55 (1999), S. 101-104 
    ISSN: 1432-1041
    Keywords: Key words Troglitazone ; Erythropoietic ; parameters ; Plasma volume
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology , Medicine
    Notes: Abstract Objective: Troglitazone is a new anti-diabetic agent for the treatment of type 2 diabetes. In placebo-controlled trials troglitazone improves glycaemic control, reduces hyperinsulinaemia and has beneficial effects on blood lipids. However, minor, reversible reductions in erythrocyte count, haemoglobin and haematocrit with no associated clinical symptoms have been observed in some troglitazone-treated patients. The primary objective of the present study was to determine if these changes could be explained by a decrease in red cell mass or change in plasma volume. Methods: Twenty-four healthy males were randomized in a double-blind manner to troglitazone (200 or 600 mg per day) or placebo for 6 weeks. Blood samples for the measurement of red cell mass and plasma volume were obtained in the 2 weeks prior to treatment and after 6 weeks of treatment. Reticulocyte and erythrocyte counts, haemoglobin and haematocrit were also measured. Results: At the end of the treatment period there were no statistically significant changes in red cell mass. Similarly there were no changes in reticulocyte count, erythropoietin or soluble transferrin receptors. These data indicate that troglitazone does not affect erythropoiesis. In addition, troglitazone was not associated with increased red blood cell destruction or haemolysis. There was a trend towards increased plasma volume in the troglitazone groups: increases of 2.5 ml · kg−1 (5.7% increase) in the troglitazone 200 mg group and 3.4 ml · kg−1 (7.8% increase) in the troglitazone 600 mg group were observed compared with placebo. Conclusion: These data suggest that dilutional effects related to a modest increase in plasma volume may explain the haematological changes seen in other clinical trials with high doses of troglitazone, although this study has shown that the changes in plasma volume are not statistically significant.
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  • 6
    Electronic Resource
    Electronic Resource
    Springer
    Journal of industrial microbiology and biotechnology 16 (1996), S. 102-109 
    ISSN: 1476-5535
    Keywords: Millet ; Pennisetum typhoides ; liquefaction ; saccharification ; baker's yeast ; Saccharomyces cerevisiae ; fermentation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract A fermentation medium based on millet (Pennisetum typhoides) flour hydrolysate and a four-phase feeding strategy for fed-batch production of baker's yeast,Saccharomyces cerevisiae, are presented. Millet flour was prepared by dry-milling and sieving of whole grain. A 25% (w/v) flour mash was liquefied with a thermostable 1,4-α-d-glucanohydrolase (EC 3.2.1.1) in the presence of 100 ppm Ca2+, at 80°C, pH 6.1–6.3, for 1 h. The liquefied mash was saccharified with 1,4-α-d-glucan glucohydrolase (EC 3.2.1.3) at 55°C, pH 5.5, for 2 h. An average of 75% of the flour was hydrolysed and about 82% of the hydrolysate was glucose. The feeding profile, which was based on a model with desired specific growth rate range of 0.18–0.23 h−1, biomass yield coefficient of 0.5 g g−1 and feed substrate concentration of 200 g L−1, was implemented manually using the millet flour hydrolysate in test experiments and glucose feed in control experiments. The fermentation off-gas was analyzed on-line by mass spectrometry for the calculation of carbon dioxide production rate, oxygen up-take rate and the respiratory quotient. Off-line determination of biomass, ethanol and glucose were done, respectively, by dry weight, gas chromatography and spectrophotometry. Cell mass concentrations of 49.9–51.9 g L−1 were achieved in all experiments within 27 h of which the last 15 h were in the fedbatch mode. The average biomass yields for the millet flour and glucose media were 0.48 and 0.49 g g−1, respectively. No significant differences were observed between the dough-leavening activities of the products of the test and the control media and a commercial preparation of instant active dry yeast. Millet flour hydrolysate was established to be a satisfactory low cost replacement for glucose in the production of baking quality yeast.
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  • 7
    ISSN: 1572-8951
    Keywords: MD ; DNA ; protein-DNA ; simulation ; solvent ; lambda repressor ; operator ; GROMOS ; reduced charge model
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Abstract The results of a recent nanosecond (ns) molecular dynamics (MD) simulation of the d(CGCGAATTCGCG) double helix in water and a 100 ps MD study of the λ repressor-operator complex are described. The DNA simulations are analyzed in terms of the structural dynamics, fluctuations in the groove width and bending of the helical axis. The results indicate that the ns dynamical trajectory progresses through a series of three substates of B form DNA, with lifetimes of the order of hundreds of picoseconds (ps). An incipient dynamical equilibrium is evident. A comparison of the calculated axis bending with that observed in corresponding crystal structure data is presented. Simulation of the DNA in complex with the protein and that of the free DNA in solution, starting from the crystal conformation, reveal the dynamical changes that occur on complex formation.
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  • 8
    ISSN: 1573-5001
    Keywords: Selective labeling ; Glutamine ; 15N ; Superoxide dismutase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract The glutamine in position 69 is one of only three conserved active-site amino acid differencesbetween Fe- and Mn-containing superoxide dismutases (SODs). We have refined theconditions for extremely selective labeling of the side chains of glutamine with 15N, and thusobtained dramatically simplified spectra, despite the large size of Fe-SOD. The improvedresolution afforded by such highly specific labeling permits the use of direct 15N detectionto observe and assign Gln 69, even though its distance to the paramagnetic Fe2+ is only 5Å. Selective glutamine side-chain labeling is inexpensive and has general utility forlarge (and paramagnet-containing) proteins.
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  • 9
    ISSN: 1871-4528
    Keywords: Dumas combustion ; Solanum tuberosum L
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary Calibrations are provided to relate near infrared reflectance (NIR) data to total N concentration in potato leaves, stems, and tubers, estimated by Dumas combustion. Separate calibrations are necessary for each tissue-type. the NIR models simulate Dumas [N] estimates accurately, quickly and cheaply and have been shown to be stable. The methods of deriving the calibrations, correcting them and then providing independent tests are described.
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  • 10
    ISSN: 1617-4623
    Keywords: Key words RNA cleavage ; Hammerhead ribozymes ; Antisense RNA ; Antiviral agents ; In vivo expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  Ribozymes of the hammerhead class can be designed to cleave a target RNA in a sequence-specific manner and can potentially be used to specifically modulate gene activity. We have targeted the tobacco mosaic virus (TMV) genome with a ribozyme containing three catalytic hammerhead domains embedded within a 1 kb antisense RNA. The ribozyme was able to cleave TMV RNA at all three target sites in vitro at 25° C. Transgenic tobacco plants were generated which expressed the ribozyme or the corresponding antisense constructs directed at the TMV genome. Six of 38 independent transgenic plant lines expressing the ribozyme and 6 of 39 plant lines expressing the antisense gene showed some level of protection against TMV infection. Homozygous progeny of some lines were highly resistant to TMV; at least 50% of the plants remained asymptomatic even when challenged with high levels of TMV. These plants also displayed resistance to infection with TMV RNA or the related tomato mosaic virus (ToMV). In contrast, hemizygous plants of the same lines displayed only very weak resistance when inoculated with low amounts of TMV and no resistance against high inoculation levels. Resistance in homozygous plants was not overcome by a TMV strain which was altered at the three target sites to abolish ribozyme-mediated cleavage, suggesting that the ribozyme conferred resistance primarily by an antisense mechanism.
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