bluuebunny/crossref_metadata_2025 huggingface dataset:
Differential regulation of imprinting in the murine embryo and placenta by theDlk1-Dio3imprinting control region
Authors: Lin Shau-Ping , Coan Phil , da Rocha Simao Teixeira , Seitz Herve , Cavaille Jerome , Teng Pi-Wen , Takada Shuji , Ferguson-Smith Anne C.
Published: December 2006

Genomic imprinting is an epigenetic mechanism controlling parental-origin-specific gene expression. Perturbing the parental origin of the distal portion of mouse chromosome 12 causes alterations in the dosage of imprinted genes resulting in embryonic lethality and developmental abnormalities of both embryo and placenta. A 1 Mb imprinted domain identified on distal chromosome 12 contains three paternally expressed protein-coding genes and multiple non-coding RNA genes, including snoRNAs and microRNAs,expressed from the maternally inherited chromosome. An intergenic,parental-origin-specific differentially methylated region, the IG-DMR, which is unmethylated on the maternally inherited chromosome, is necessary for the repression of the paternally expressed protein-coding genes and for activation of the maternally expressed non-coding RNAs: its absence causes the maternal chromosome to behave like the paternally inherited one. Here, we characterise the developmental consequences of this epigenotype switch and compare these with phenotypes associated with paternal uniparental disomy of mouse chromosome 12. The results show that the embryonic defects described for uniparental disomy embryos can be attributed to this one cluster of imprinted genes on distal chromosome 12 and that these defects alone, and not the mutant placenta, can cause prenatal lethality. In the placenta, the absence of the IG-DMR has no phenotypic consequence. Loss of repression of the protein-coding genes occurs but the non-coding RNAs are not repressed on the maternally inherited chromosome. This indicates that the mechanism of action of the IG-DMR is different in the embryo and the placenta and suggests that the epigenetic control of imprinting differs in these two lineages.
Ascl1 defines sequentially generated lineage-restricted neuronal and oligodendrocyte precursor cells in the spinal cord
Authors: Battiste James , Helms Amy W. , Kim Euiseok J. , Savage Trisha K. , Lagace Diane C. , Mandyam Chitra D. , Eisch Amelia J. , Miyoshi Goichi , Johnson Jane E.
Published: December 2006

The neural basic helix-loop-helix transcription factor Ascl1 (previously Mash1) is present in ventricular zone cells in restricted domains throughout the developing nervous system. This study uses genetic fate mapping to define the stage and neural lineages in the developing spinal cord that are derived from Ascl1-expressing cells. We find that Ascl1 is present in progenitors to both neurons and oligodendrocytes, but not astrocytes. Temporal control of the fate-mapping paradigm reveals rapid cell-cycle exit and differentiation of Ascl1-expressing cells. At embryonic day 11, Ascl1 identifies neuronal-restricted precursor cells that become dorsal horn neurons in the superficial laminae. By contrast, at embryonic day 16, Ascl1 identifies oligodendrocyte-restricted precursor cells that distribute throughout the spinal cord. These data demonstrate that sequentially generated Ascl1-expressing progenitors give rise first to dorsal horn interneurons and subsequently to late-born oligodendrocytes. Furthermore, Ascl1-null cells in the spinal cord have a diminished capacity to undergo neuronal differentiation, with a subset of these cells retaining characteristics of immature glial cells.
Extrinsic and intrinsic mechanisms directing epithelial cell sheet replacement duringDrosophilametamorphosis
Authors: Ninov Nikolay , Chiarelli Dominic A. , Martín-Blanco Enrique
Published: December 2006

The fusion of epithelial sheets is an essential morphogenetic event. Here,we study the development of the abdomen of Drosophila as a model of bounded epithelia expansion and uncover a complex multistep process for the generation of the adult epidermis from histoblasts, founder cells that replace the larval cells during metamorphosis. We find that histoblasts experience a biphasic cell cycle and emit apical projections that direct their invasive planar intercalation in between larval cells. Coordinately, the larval cells extrude from the epithelia by apical constriction of an actomyosin ring and as a consequence die by apoptosis and are removed by circulating haemocytes. We demonstrate that the proliferation of histoblasts and the death of larval cells are triggered by two independent extrinsic Ecdysone hormonal pulses. Finally, we show that histoblast spreading and the death of larval cells depend on a mutual exchange of signals and are non-autonomous processes.
Intraflagellar transport is essential for endochondral bone formation
Authors: Haycraft Courtney J. , Zhang Qihong , Song Buer , Jackson Walker S. , Detloff Peter J. , Serra Rosa , Yoder Bradley K.
Published: December 2006

While cilia are present on most cells in the mammalian body, their functional importance has only recently been discovered. Cilia formation requires intraflagellar transport (IFT), and mutations disrupting the IFT process result in loss of cilia and mid-gestation lethality with developmental defects that include polydactyly and abnormal neural tube patterning. The early lethality in IFT mutants has hindered research efforts to study the role of this organelle at later developmental stages. Thus, to investigate the role of cilia during limb development, we generated a conditional allele of the IFT protein Ift88 (polaris). Using the Cre-lox system, we disrupted cilia on different cell populations within the developing limb. While deleting cilia in regions of the limb ectoderm had no overt effect on patterning, disruption in the mesenchyme resulted in extensive polydactyly with loss of anteroposterior digit patterning and shortening of the proximodistal axis. The digit patterning abnormalities were associated with aberrant Shh pathway activity, whereas defects in limb outgrowth were due in part to disruption of Ihh signaling during endochondral bone formation. In addition, the limbs of mesenchymal cilia mutants have ectopic domains of cells that resemble chondrocytes derived from the perichondrium, which is not typical of Indian hedgehog mutants. Overall these data provide evidence that IFT is essential for normal formation of the appendicular skeleton through disruption of multiple signaling pathways.
Comparative analysis of Hox downstream genes inDrosophila
Authors: Hueber Stefanie D. , Bezdan Daniela , Henz Stefan R. , Blank Martina , Wu Haijia , Lohmann Ingrid
Published: December 2006

Functional diversification of body parts is dependent on the formation of specialized structures along the various body axes. In animals,region-specific morphogenesis along the anteroposterior axis is controlled by a group of conserved transcription factors encoded by the Hox genes. Although it has long been assumed that Hox proteins carry out their function by regulating distinct sets of downstream genes, only a small number of such genes have been found, with very few having direct roles in controlling cellular behavior. We have quantitatively identified hundreds of Hox downstream genes in Drosophila by microarray analysis, and validated many of them by in situ hybridizations on loss- and gain-of-function mutants. One important finding is that Hox proteins, despite their similar DNA-binding properties in vitro, have highly specific effects on the transcriptome in vivo, because expression of many downstream genes respond primarily to a single Hox protein. In addition, a large fraction of downstream genes encodes realizator functions, which directly affect morphogenetic processes, such as orientation and rate of cell divisions, cell-cell adhesion and communication,cell shape and migration, or cell death. Focusing on these realizators, we provide a framework for the morphogenesis of the maxillary segment. As the genomic organization of Hox genes and the interaction of Hox proteins with specific co-factors are conserved in vertebrates and invertebrates, and similar classes of downstream genes are regulated by Hox proteins across the metazoan phylogeny, our findings represent a first step toward a mechanistic understanding of morphological diversification within a species as well as between species.
Temperature-dependent effects of cadmium and purine nucleotides on mitochondrial aconitase from a marine ectotherm,Crassostrea virginica: a role of temperature in oxidative stress and allosteric enzyme regulation
Authors: Cherkasov Anton A. , Overton Robert A. , Sokolov Eugene P. , Sokolova Inna M.
Published: December 2006

Temperature and heavy metals such as cadmium (Cd) are important environmental stressors that can strongly affect mitochondrial function of marine poikilotherms. In this study, we investigated the combined effects of temperature (20°C and 30°C) and Cd stress on production of reactive oxygen species (ROS) and oxidative stress in a marine poikilotherm Crassostrea virginica (the eastern oyster) using mitochondrial aconitase as a sensitive biomarker of oxidative damage. We also assessed potential involvement of mitochondrial uncoupling proteins (UCPs) in antioxidant protection in oyster mitochondria using purine nucleotides (GDP,ATP and ADP) as specific inhibitors, and free fatty acids as stimulators, of UCPs. Our results show that exposure to Cd results in elevated ROS production and oxidative damage as indicated by aconitase inactivation which is particularly pronounced at elevated temperature. Unexpectedly, oyster mitochondrial aconitase was inhibited by physiologically relevant levels of ATP (IC50=1.93 and 3.04 mmol l-1 at 20°C and 30°C, respectively), suggesting that allosteric regulation of aconitase by this nucleotide may be involved in regulation of the tricarboxylic acid flux in oysters. Aconitase was less sensitive to ATP inhibition at 30°C than at 20°C, consistent with the elevated metabolic flux at higher temperatures. ADP and GDP also inhibited mitochondrial aconitase but at the levels well above the physiological concentrations of these nucleotides (6-11 mmol l-1). Our study shows expression of at least three UCP isoforms in C. virginica gill tissues but provides no indication that UCPs protect mitochondrial aconitase from oxidative inactivation in oysters. Overall, the results of this study indicate that temperature stress exaggerates toxicity of Cd leading to elevated oxidative stress in mitochondria, which may have important implications for survival of poikilotherms in polluted environments during seasonal warming and/or global climate change, and suggest a novel temperature-dependent mechanism of allosteric regulation of TCA flux in oyster mitochondria.
Electromyography of the buccal musculature of octopus (Octopus bimaculoides): a test of the function of the muscle articulation in support and movement
Authors: Uyeno Theodore A. , Kier William M.
Published: December 2006

The buccal mass musculature of the octopus (Octopus bimaculoides)was studied with electromyography to test the predictions of a previous morphological study in which we suggested that the muscles of the buccal mass serve as both the effectors of movement and as the joint itself, forming a new category of flexible joint termed a `muscle articulation'. The predictions of muscle function were tested by correlating muscle electrical activity in isolated buccal masses with spontaneous beak movements. Bipolar electromyography electrodes were implanted in the various beak muscles and beak position was recorded simultaneously with an electronic movement monitor(N=14). The results are consistent with the hypothesis that the lateral mandibular muscles produce opening movements of the beaks and provide the first definitive explanation of the opening mechanism. The results are also consistent with the hypothesis that the superior mandibular muscle functions primarily in closing. Co-contraction of the lateral mandibular muscles and the superior mandibular muscles was also observed, suggesting that these muscles may also stabilize the beaks during movement or provide a means of controlling the location of the pivot between the beaks. This study provides an important first test of the predictions of the role of the complex musculature found in muscle articulations such as the cephalopod buccal mass.
Flower tracking in hawkmoths: behavior and energetics
Authors: Sprayberry Jordanna D. H. , Daniel Thomas L.
Published: December 2006

As hovering feeders, hawkmoths cope with flower motions by tracking those motions to maintain contact with the nectary. This study examined the tracking, feeding and energetic performance of Manduca sexta feeding from flowers moving at varied frequencies and in different directions. In general we found that tracking performance decreased as frequency increased; M. sexta tracked flowers moving at 1 Hz best. While feeding rates were highest for stationary flowers, they remained relatively constant for all tested frequencies of flower motion. Calculations of net energy gain showed that energy expenditure to track flowers is minimal compared to energy intake;therefore, patterns of net energy gain mimicked patterns of feeding rate. The direction effects of flower motion were greater than the frequency effects. While M. sexta appeared equally capable of tracking flowers moving in the horizontal and vertical motion axes, they demonstrated poor ability to track flowers moving in the looming axis. Additionally, both feeding rates and net energy gain were lower for looming axis flower motions.
Dipole hearing measurements in elasmobranch fishes
Authors: Casper Brandon M. , Mann David A.
Published: December 2006

The hearing thresholds of the horn shark Heterodontus francisciand the white-spotted bamboo shark Chiloscyllium plagiosum were measured using auditory evoked potentials (AEP) in response to a dipole sound stimulus. The audiograms were similar between the two species with lower frequencies yielding lower particle acceleration thresholds. The particle acceleration audiograms showed more sensitive hearing at low frequencies than previous elasmobranch audiograms, except for the lemon shark Negaprion brevirsotris. Auditory evoked potential signals were also recorded while the dipole stimulus was moved to different locations above the head and body. The strongest AEP signals were recorded from the area around the parietal fossa, supporting previous experiments that suggested this region is important for elasmobranch hearing. This is the first time that hearing experiments have been conducted using a dipole stimulus with elasmobranchs, which more closely mimics the natural sounds of swimming prey.
Echolocation signals of wild harbour porpoises,Phocoena phocoena
Authors: Villadsgaard Anne , Wahlberg Magnus , Tougaard Jakob
Published: December 2006

Field recordings of harbour porpoises (Phocoena phocoena) were made in the inner Danish waters with a vertical array of three or four hydrophones. The back-calculated source level ranged from 178 to 205 dB re 1μPa pp @ 1 m with a mean source level of 191 dB re 1 μPa pp @ 1 m. The maximum source level was more than 30 dB above what has been measured from captive animals, while the spectral and temporal properties were comparable. Calculations based on the sonar equation indicate that harbour porpoises,using these high click intensities, should be capable of detecting fish and nets and should be detectable by porpoise detectors over significantly larger distances than had previously been assumed. Harbour porpoises in this study preferred a relatively constant inter-click interval of about 60 ms, but intervals up to 200 ms and down to 30 ms were also recorded.
No trade-off between biting and suction feeding performance in clariid catfishes
Authors: Van Wassenbergh Sam , Herrel Anthony , Adriaens Dominique , Aerts Peter
Published: December 2006

It is generally assumed that biting performance trades off with suction performance in fish because both feeding types may place conflicting demands on the cranial musculo-skeletal system. However, the functional consequences of morphological adaptations enhancing biting on the mechanics and performance of suction feeding in fish remain obscure. In this study, suction feeding performance was compared between three clariid catfish species differing considerably in their biting capacity, by measuring the velocity of a standardized prey being sucked into the buccal cavity using high-speed cineradiography. In addition, buccal volume changes during prey capture were quantified by ellipse modelling. As all species were able to accelerate the prey to similar peak velocities, our results demonstrate the possibility for catfishes to increase bite performance considerably without compromising suction performance. The amount of buccal expansion in the ventral direction is approximately equal for all species. Consequently, the system generating expansion through ventral rotation of the lower jaw, hyoid and pectoral girdle is apparently not constrained (mechanically or architectonically) by the hypertrophy of the jaw adductors. As the effect of a reduced magnitude of lateral expansion (suspensorium abduction) on suction performance in Clariidae appears to be negligible (for example in Gymnallabes typus), these data demonstrate the dominant role of ventral expansion for producing suction in these fish.
Dual mechanisms for nitric oxide control of large arteries in the estuarine crocodile Crocodylus porosus
Authors: Broughton Brad R. S. , Donald John A.
Published: December 2006

In reptiles, accumulating evidence suggests that nitric oxide (NO) induces a potent relaxation in the systemic vasculature. However, very few studies have examined the source from which NO is derived. Therefore, the present study used both anatomical and physiological approaches to establish whether NO-mediated vasodilation is via an endothelial or neural NO pathway in the large arteries of the estuarine crocodile Crocodylus porosus. Specific endothelial nitric oxide synthase (NOS) staining was observed in aortic endothelial cells following nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) histochemistry and endothelial NOS immunohistochemistry (IHC), suggesting that an endothelial NO pathway is involved in vascular control. This finding was supported by in vitroorgan bath physiology, which demonstrated that the relaxation induced by acetylcholine (10-5 mol l-1) was abolished in the presence of the NOS inhibitor, N-omega-nitro-l-arginine(l-NNA; 10-4 mol l-1), the soluble guanylyl cyclase inhibitor, 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ;10-5 mol l-1), or when the endothelium was removed. Interestingly, evidence for a neural NO pathway was also identified in large arteries of the crocodile. Neural NOS was located in perivascular nerves of the major blood vessels following NADPH-d histochemistry and neural NOS IHC and in isolated aortic rings, l-NNA and ODQ, but not the removal of the endothelium, abolished the relaxation effect of the neural NOS agonist,nicotine (3×10-4 mol l-1). Thus, we conclude that the large arteries of C. porosus are potentially regulated by NO-derived from both endothelial and neural NOS.
Associations between basal metabolic rate and reproductive performance in C57BL/6J mice
Authors: Johnston Sarah L. , Souter Donna M. , Erwin Susan S. , Tolkamp Bert J. , Yearsley Jonathan M. , Gordon Iain J. , Illius Andrew W. , Kyriazakis Ilias , Speakman John R.
Published: December 2006

Basal metabolic rate (BMR) is highly variable, both between and within species. One hypothesis is that this variation may be linked to the capacity for sustained rate of energy expenditure, leading to associations between high BMR and performance during energy-demanding periods of life history, such as reproduction. However, despite the attractive nature of this hypothesis,previous studies have failed to show an association between BMR and fecundity. Our approach was to mate 304 C57BL/6J mice and allow them to wean pups before measuring BMR by indirect calorimetry. We did not find an association between BMR and litter mass, size or pup mass at birth or weaning that could not be accounted for by the body mass of the dam. There was also no relationship between BMR (or BMR corrected for body mass) and birth or weaning success,losses during weaning, or sex ratio. However, a significant relationship was found between BMR and gestational weight loss indicative of foetal resorption. This suggests that during pregnancy the available energy may be limited and partitioned away from the growing foetus and towards maintenance of the mother. In this context, a high BMR may actually be disadvantageous,conflicting with the idea that high BMR may bring reproductive benefits.
Echolocation and passive listening by foraging mouse-eared batsMyotis myotisandM. blythii
Authors: Russo Danilo , Jones Gareth , Arlettaz Raphaël
Published: December 2006

The two sibling mouse-eared bats, Myotis myotis and M. blythii, cope with similar orientation tasks, but separate their trophic niche by hunting in species-specific foraging microhabitats. Previous work has shown that both species rely largely on passive listening to detect and glean prey from substrates, and studies on other bat species have suggested that echolocation is `switched off' during passive listening. We tested the hypothesis that mouse-eared bats continuously emit echolocation calls while approaching prey. Echolocation may be needed for orientation while simultaneously listening for prey. Because these sibling species forage in different microhabitats and eat different prey, we also compared their echolocation behaviour and related it to their ecology. Both species used echolocation throughout prey approach, corroborating a functional role for echolocation during gleaning. Captive bats of both species emitted similar orientation calls, and pulse rate increased during prey approach. Between the search to approach phases, call amplitude showed a sudden, dramatic drop and bats adopted `whispering echolocation' by emitting weak calls. Whispering echolocation may reduce the risks of masking prey-generated sounds during passive listening, the mouse-eared bats' main detection tactic; it may also avoid alerting ultrasound-sensitive prey. In several cases M. myotisemitted a loud buzz made of 2-18 components when landing. We hypothesise that the buzz, absent in M. blythii at least when gleaning from the same substrate, is used to assess the distance from ground and refine the landing manoeuvre. Our findings have implications for niche separation between sibling species of echolocating bats, support a role for echolocation during passive listening and suggest a functional role for buzzes in landing control.
Calcium-regulated fusion of yolk granules is important for yolk degradation during early embryogenesis of Rhodnius prolixusStahl
Authors: Ramos I. B. , Miranda K. , de Souza W. , Oliveira D. M. P. , Lima A. P. C. A. , Sorgine M. H. F. , Machado E. A.
Published: December 2006

This study examined the process of membrane fusion of yolk granules (YGs)during early embryogenesis of Rhodnius prolixus. We show that eggs collected at days 0 and 3 after oviposition contain different populations of YGs, for example day-3 eggs are enriched in large YGs (LYGs). Day-3 eggs also contain the highest free [Ca2+] during early embryogenesis of this insect. In vitro incubations of day-0 YGs with [Ca2+]similar to those found in day-3 eggs resulted in the formation of LYGs, as observed in vivo. Fractionation of LYGs and small YGs (SYGs) and their subsequent incubation with the fluorescent membrane marker PKH67 showed a calcium-dependent transference of fluorescence from SYGs to LYGs, possibly as the result of membrane fusion. Acid phosphatase and H+-PPase activities were remarkably increased in day-3 LYGs and in calcium-treated day-0 LYGs. Both fractions were found to contain vitellins as major components, and incubation of YGs with calcium induced yolk proteolysis in vitro. Altogether, our results suggest that calcium-induced membrane fusion events take part in yolk degradation, leading to the assembly of the yolk mobilization machinery.
Emancipation Discourse in the Late 18th Century: Christian Wilhelm von Dohm on the Jews (1781)
Authors: Crouter Richard
Published: December 2006

Weit häufiger zitiert als tatsächlich besprochen und analysiert muß Christian Wilhelm von Dohms Abhandlung aus dem Jahr 1781 – ein politisches Gegenstück zu Lessings Nathan der Weise – als Meilenstein auf dem Weg der jüdischen Emanzipation angesehen werden. Vor dem Hintergrund seiner Studien der Rechte, der Ökonomie und der Statistik in Göttingen trugen Dohms Argumente einer vergleichenden Geschichtsbetrachtung, seine Anlehnung an Naturrechtsentwürfe und seine realistische Einschätzung menschlichen Charakters nachweislich mehr zur Ausformung eines modernen Pluralismus, in dem Bürgerrechte für Juden ihren Platz finden, bei, als es zeitgenössische Diskurse christlicher Theologen vermochten – inklusive der Position, die Schleiermacher 1799 einnahm, als er gegen eine Überprüfung religiöser Identität als Basis für die Anerkennung jüdischer Bürgerrechte eintrat.
„Das Geschichtliche auszuschließen sei doch gegen das christl. Gefühl“ – Karl von Hases Jenaer Seminar 1850–1883 als Tradierungsort liberaler Bürgertheologie (Erster Teil: SS 1850)
Authors: Wischmeyer Johannes
Published: December 2006

Hohe Überlieferungsdisziplin einer akademischen Institution und ein wenig Glück auf seiten des Historikers sind die Voraussetzung dafür, daß sich im Aktenmaterial eines Lehrstuhls bedeutende Quellen aus dem 19. Jahrhundert finden bzw. identifizieren lassen, die generationenlang den Weg ins reguläre Universitätsarchiv gescheut haben. Im Besitz des Jenaer Lehrstuhls für Kirchengeschichte haben zwei handschriftliche Protokollbücher überdauert, die einen lückenlosen Überblick über dreiunddreißig Jahre Arbeit der dogmatisch-historischen Klasse des Jenaer theologischen Seminars unter der Leitung des Kirchenhistorikers Karl von Hase geben. Sie werden hier erstmals ediert.
Interlaboratory Comparison of the Thickness of the Destroyed Surface Layer of Closed-Cell Foam Insulation Specimens
Authors: Stovall TK
Published: December 2006

The preparation of closed-cell foam insulation test specimens can affect the results of a number of hygrothermal property measurements. In particular, the thickness of the destroyed surface layer affects the measurement of gas diffusion coefficients and thermal conductivity. Multiple specimen preparation methods are in use, as well as multiple methods to measure the thickness of the destroyed surface layer. A ruggedness test, including an interlaboratory comparison, was conducted by an ASTM technical committee to examine the variations due to both of these factors. The results are important in understanding the likely range in values for each preparation technique. The results also demonstrate a sensitivity to the measurement technique.
The Line Method for Petrographic Determination of the Quantity of Fly Ash and Ground-granulated Blast Furnace Slag in Hardened Concrete and Blended Cement
Authors: Fox JM , Miller PT
Published: December 2006

Research was undertaken using thin sections and immersion mounts with the objective of using a modified line count procedure for the quantitative petrographic determination of fly ash and ground-granulated blast furnace slag (GGBFS) in hardened concrete and blended cements. The line method used in this study is a variation of standard microscopic point count methods and yields an average number frequency that is a linear function of the weight percent of fly ash or GGBFS of total cementitious materials. Multiple traverses were counted on thin sections of concrete over areas of paste only and in immersion slides of blended cements containing known amounts of fly ash and GGBFS. The average number of particles in a traverse was computed, the traverse length being equal to the width of the field of view of the petrographic microscope. For hardened concrete, only pozzolan particles were counted. For blended cements, all particles were counted and the ratio of pozzolan particles to total particles was determined. A linear relationship between the average number of pozzolan particles for a traverse line and the weight percent of pozzolan makes it possible to estimate the fly ash and GGBFS content of unknowns. Results of the research indicate that the line method is a rapid and accurate method for petrographic determination of fly ash and GGBFS in hardened concrete and blended cements. This technique and variations of it have been used by other researchers and it has the potential to be a standard test procedure.
Modellierung von Kraft- und Beschleunigungsaufnehmern für die Stoßkalibrierung (Modelling Force and Acceleration Transducers for Shock Calibrations)
Authors: Link Alfred , Kobusch Michael , Bruns Thomas , Elster Clemens
Published: December 2006

Neuere Auswerteverfahren für die Stoßkalibrierung von Beschleunigungsaufnehmern werden vorgestellt, die auf der Modellierung des Aufnehmerverhaltens basieren. Die zur Identifikation dieser Modelle entwickelten Techniken einschließlich der Bestimmung resultierender Messunsicherheiten werden erläutert. Die Übertragung auf den Fall der Stoßkalibrierung von Kraftaufnehmern wird diskutiert und anhand der Auswertung von Messungen an der Kraftstoßkalibriereinrichtung der Physikalisch-Technischen Bundesanstalt illustriert.
Experimentelle Bestimmung von Massenträgheitsmomenten als Rückführungsgröße für die dynamische Drehmomentkalibrierung (Experimental Determination of Moments of Inertia for Traceability in Dynamic Torque Calibration)
Authors: Bruns Thomas , Wedmann Achim
Published: December 2006

Die Physikalisch-Technische Bundesanstalt (PTB) entwickelt neue Verfahren für die Kalibrierung von Drehmomentaufnehmern mit zeitlich variierendem Drehmoment. Die Rückführung erfolgt dabei auf die Größen Massenträgheitsmoment und Winkelbeschleunigung. Die vorliegende Arbeit beschreibt die experimentelle Bestimmung des beteiligten Massenträgheitsmomentes der Normal-Messeinrichtung (NME). Zur Messung wurde ein Torsionspendel aufgebaut und dessen Schwingung mit dem interferometrischen Messsystem der NME erfasst.
Cloning and expression of chicken 20-hydroxysteroid dehydrogenase
Authors: Bryndová J , Klusoňová P , Kučka M , Mazancová-Vagnerová K , Mikšík I , Pácha J
Published: December 2006

The ligand specificity and activation of steroid receptors depend considerably on the enzymatic activities involved in local pre-receptor synthesis and the metabolism of the steroids. Several enzymes in particular, steroid dehydrogenases have been shown to participate in this process. Here we report the isolation of 20-hydroxysteroid dehydrogenase (ch20HSD) cDNA from chicken intestine and the distribution of ch20HSD mRNA and 20-reductase activity in various avian tissues. Using a reverse transcription PCR and comparison with the known sequences of mammalian 20βHSDs, we have isolated a new ch20HSD cDNA. This cDNA predicted 276 amino acid residues that shared about 75% homology with mammalian 20βHSD. Sequences specific to the short-chain dehydrogenase/reductase superfamily (SDR) were found, the Gly-X-X-X-Gly-X-Gly cofactor-binding motif (residues 11–17) and the catalytic activity motif Tyr-X-X-X-Lys (residues 193–197). The cDNA coding for ch20HSD was expressed in Escherichia coli by placing it under isopropylthiogalactoside (IPTG) inducible control. Both the IPTG cells of E. coli and the isolated recombinant protein reduced progesterone to 20-dihydroprogesterone, corticosterone to 20-dihydrocorticosterone and 5α-dihydrotestosterone to its 3-ol derivative. The 20-reductase and 3-reductase activities of ch20HSD catalyzed both 3α/β- and 20α/20β-epimers. The mRNA transcripts of ch20HSD were found in the kidney, colon, and testes; weaker expression was also found in the heart, ovaries, oviduct, brain, liver, and ileum. 20-Reductase activity has been proven in tissue slices of kidney, colon, ileum, liver, oviduct, testis, and ovary; whereas the activity was nearly absent in the heart and brain. A similar distribution of 20-reductase activity was found in tissue homogenates measured under Vmax conditions. These results suggest that chicken 20HSD is the latest member of the SDR superfamily to be found, is expressed in many avian tissues and whose precise role remains to be determined.
Modulation of human estrogen receptor α F promoter by a protein kinase C/c-Src-dependent mechanism in osteoblast-like cells
Authors: Longo Maurizio , Peruzzi Barbara , Fortunati Dario , De Luca Veronica , Denger Stefanie , Caselli Gianfranco , Migliaccio Silvia , Teti Anna
Published: December 2006

The human estrogen receptor α (ERα ) gene is driven by multiple promoters, of which the F promoter alone is found to be active in primary osteoblasts. The study was aimed at identifying new regulatory pathways affecting transcription of the receptor in this cell lineage. We generated human osteoblast-like cells, Saos-2, stably transfected with a luciferase-reporter gene downstream of the human ERα F promoter (Saos F-Luc), and assayed the reporter response to differentiation-related signals. Over-confluence, shown to stimulate osteoblast differentiation, caused a time-dependent increase of F-promoter activity and correlated with an inactivation of protein kinase C α (PKCα ). PKC downregulation, obtained by long-term treatment with phorbol 12-myristate 13-acetate (PMA), resulted in promoter stimulation at similar levels in sub-confluent cells. The F promoter contains a putative PMA-responsive AP-1 site, but AP-1 activation was unremarkable in over-confluent cells. Treatment with PP1, a specific inhibitor of the non-receptor tyrosine-kinase c-Src, which is a negative regulator of osteoblast differentiation, showed that the activity of this kinase inhibits the F promoter. In PP1-treated cells, F-promoter activity was not further increased by PMA. Treatment with the generic kinase inhibitor 4-dimethylaminopyridine (DMAP) resulted in a dose-dependent induction of the promoter, which matched a parallel decrease of active c-Src. The effect was c-Src dependent, as DMAP caused no further promoter induction in PP1-treated cells. Overexpression of exogenous human ERα resulted in modest promoter stimulation, which required the ligand-independent activator function 1 of the receptor. In murine primary osteoblasts, additional ERα signal was observed upon induction of F promoter. In conclusion, we demonstrated a robust PKC/c-Src-dependent and estrogen-independent mechanism modulating transcription of ERα in osteoblasts, probably affecting estrogen responsiveness during cell differentiation.
The hormone replacement therapy drug tibolone acts very similar to medroxyprogesterone acetate in an estrogen-and progesterone-responsive endometrial cancer cell line
Authors: Hanifi-Moghaddam P , Sijmons B , Ott M C , van IJcken W F J , Nowzari D , Kuhne E C M , van der Spek P , Kloosterboer H J , Burger C W , Blok L J
Published: December 2006

Tibolone, a steroidogenic compound with both estrogenic and progestagenic properties, is used as an alternative for estrogen or estrogen plus progesterone hormone therapy for the treatment of symptoms associated with menopause and osteoporosis. We have evaluated whether the effect of tibolone on a human endometrial cell line is similar to, or comparable with, the effect of estradiol (E2), medroxyprogesterone acetate (MPA) or E2 + MPA treatment. Using stable transfection techniques, the estrogen receptor (ER) expressing human endometrial cancer cell line, ECC1, was altered to also express both progesterone receptors (PRs). These cells were then used to assess growth regulation and expression profiling (Affymetrix U133plus2) under the influence of E2 (1 nM), MPA (1 nM), E2 + MPA or tibolone (100 nM). Growth assessment and comparison of profiles indicate that tibolone behaves predominantly like MPA. Furthermore, regulation of prereplication complex genes, such as the minichromosome maintenance genes, could be involved in the observed strong inhibition of growth by tibolone as well as MPA. In addition, in total, 15 genes were found to be specific for tibolone treatment. These genes were predominantly involved in regulation of the cell cycle and differentiation.
Impact of uncoupling protein-2 overexpression on proinsulin processing
Authors: Kashemsant Narudee , Chan Catherine B
Published: December 2006

Hyperproinsulinemia is observed in type 2 diabetic patients. We hypothesized that the induction of uncoupling protein-2 (UCP2) would impair processing of proinsulin to mature insulin and potentially contribute to hyperproinsulinemia, based on the evidence that hormone processing is an ATP-dependent process and UCP2 up-regulation can suppress cellular ATP production. UCP2 was overexpressed (UCP2-OE) by twofold in INS-1 cells by means of plasmid transfection. Although UCP2-OE reduced glucose-stimulated insulin secretion and cellular ATP content, no effects on proinsulin processing, as measured by western blotting, were observed. To increase the demand for insulin, we then cultured UCP2-OE and control INS-1 cells in medium containing 20 mM KCl for 24 h. High K+ markedly reduced glucose-stimulated insulin secretion from control cells, indicating inability of cells to meet secretory demand. Independent of UCP2 expression, high K+ reduced preproinsulin mRNA expression but had no effect on ATP content despite increasing ATP synthase expression. In UCP2-OE cells, high K+decreased total cellular insulin species content and increased the ratio of proinsulin to insulin, indicating an impairment of processing. We conclude that UCP2-OE can negatively impact proinsulin processing, possibly by ATP-dependent alteration of the granule environment or reduction of Ca2+availability, particularly when cells are chronically stimulated to secrete insulin.
In vivo transfection of rat liver discloses binding sites conveying GH-dependent and female-specific gene expression
Authors: Gardmo Cissi , Mode Agneta
Published: December 2006

The sexually dimorphic mode of GH secretion leads to a sex-differentiated expression of many hepatic target genes. Expression of the a1bg gene in rat liver is specifically induced by the female pattern of GH secretion. In this study, we have used the a1bg promoter in in vivo transfection experiments to investigate molecular mechanisms of GH-mediated female-specific hepatic gene regulation. Rat liver transfection was achieved by rapid tail vein injection of large volumes of plasmid solution. Expression of reporter constructs showed that the 160 bp proximal part of the a1bg promoter contained elements directing sex-specific expression. In vitro footprinting analysis and electromobility shift assays identified binding of hepatic nuclear factor 6 (HNF6), signal transducer and activator of transcriptions (Stat5) and nuclear factor 1 (NF1) in liver nuclear extracts to the 160 bp proximal promoter. Transfection of mutated and/or deletion constructs showed that HNF6 and NF1 binding markedly enhanced expression in female livers, whereas Stat5 reduces the sex difference by enhancing expression more strongly in male than in female rat liver. Based on our present results, we propose that adjacent binding sites for NF1 and HNF6 constitute a gene regulatory unit of importance for transducing the female-specific effect of GH in rat liver.
Experimental hyperthyroidism and central mediators of stress axis and thyroid axis activity in common carp (Cyprinus carpio L.)
Authors: Geven Edwin J W , Verkaar Folkert , Flik Gert , Klaren Peter H M
Published: December 2006

The effect of experimental hyperthyroidism, realized by T4 injection, on central mediators of the hypothalamo–pituitary–interrenal axis (HPI-axis) in common carp (Cyprinus carpio L.) was studied. Our results show that hyperthyroidism evokes a marked 3.2-fold reduction in basal plasma cortisol levels. Corticotropin-releasing hormone-binding protein (CRH-BP) mRNA levels in the hypothalamus, measured by real-time quantitative PCR, were significantly elevated by 40%, but CRH, urotensin-I, prepro-TRH, prohormone convertase-1 (PC1), and POMC mRNA levels were unchanged. In the pituitary pars distalis, PC1, CRH receptor-1, and POMC mRNA levels were unaffected, as was ACTH content. Plasma α-MSH concentrations were significantly elevated by 30% in hyperthyroid fish, and this was reflected in PC1 and POMC mRNA levels in pituitary pars intermedia that were increased 1.5- and 2.4-fold respectively. The α-MSH content of the pars intermedia was unchanged. Hyperthyroidism has profound effects on the basal levels of a central mediator, i.e., CRH-BP, of HPI-axis function in unstressed carp in vivo, and we conclude that HPI- and hypothalamo–pituitary–thyroid-axis functions are strongly interrelated. We suggest that the changes in plasma cortisol, thyroid hormone, and α-MSH levels reflect their concerted actions on energy metabolism.
DNA damage and its repair in lymphocytes and thyroid nodule cells during radioiodine therapy in patients with hyperthyroidism
Authors: Grzesiuk W , Nieminuszczy J , Kruszewski M , Iwanienko T , Plazińska M , Bogdańska M , Bar-Andziak E , Królicki L , Grzesiuk E
Published: December 2006

Radioiodine treatment of hyperthyroid patients with autonomous thyroid nodule leads to cellular DNA damage not only in thyrocytes but also in peripheral blood lymphocytes. The purpose of this study was to evaluate DNA breakage and base damage in thyrocytes and lymphocytes in patients treated with 131-I. In all the patients thyroid scintiscan was performed using 131-I. Damage to DNA was estimated by comet assay. Samples were taken before radioiodine treatment, and 12 and 54 days afterwards. Our results indicate high diversity in the level of DNA damage among the individual patients. However, in all cases, after 54 days the level of DNA damage in lymphocytes was similar or even lower than that in the controls. In contrast, in hot nodule the DNA damage persisted until the 54th day after 131-I application. Differences in the type of DNA damage between thyrocytes and lymphocytes were also observed. In lymphocytes there was more base damage, whereas in thyrocytes single strand breaks prevailed. This may indicate different mechanisms of DNA damage induction and/or DNA repair.
Molecular cloning and characterization of α1-soluble guanylyl cyclase gene promoter in rat pituitary cells
Authors: Jiang Yonghua , Stojilkovic Stanko S
Published: December 2006

Soluble guanylyl cyclase is a cytosolic enzyme which catalyzes conversion of GTP to the second messenger cyclic GMP. The transcriptional regulation at the promoter levels of four soluble guanylyl cyclase subunits, termed α1, α2, β1, and β2, is largely unknown. In this study, we identified the transcription start site of α1-soluble guanylyl cyclase gene in rat pituitary cells and cloned the 3.5 kb 5′-promoter. Sequence analysis of this TATA-less promoter revealed the presence of several putative-binding sites for transcriptional factors, including CCAAT site at −41 to −32 and Sp1 site at −34 to −24. Transfection of pituitary cells with constructs of variable lengths confirmed the relevance of different promoter regions in the control of transcriptional activity. Among them, the −49 to + 156 region was critical for basal transcriptional activity. Electrophoretic mobility shift assay using nuclear proteins extracted from normal and immortalized pituitary cells indicated that the CCAAT/Sp1 site within the −49 to + 156 region was able to specifically interact with CCAAT-binding factor and Sp1. These two sites were partly overlapped and both of them conferred stimulatory effects. The in vivo recruitment of CCAAT-binding factor and Sp1 was confirmed by chromatin immunoprecipitation. These results indicate that the composite CCAAT/Sp1 cis-element contributes to the expression of α1-sGC subunit in resting pituitary cells.
Angiotensin II does not stimulate proliferation of rat thyroid PC Cl3 cell line
Authors: Romano Simona , Muscella Antonella , Storelli Carlo , Marsigliante Santo
Published: December 2006

In PC Cl3 cells, a rat thyroid cell line, angiotensin (Ang II) activates the atypical protein kinase C-ζ (PKC-ζ) and the extracellular signal-regulated kinase (ERK) pathways. We here studied the Ang II effects on PC Cl3 cell proliferation. It was found that Ang II: (1) induced the phosphorylation of protein kinase B (PKB), (2) induced the growth-related early gene c-fos expression, (3) enhanced the cyclin E and p27kip expression, (4) had no effects on Cdk2, and (5) did not affect the transition from G0/G1 to S phase. Inhibition of phosphoinositide-3kinase by LY294002 further increased the effect of Ang II on p27kip induction, whilst PKCs inhibition by GF109203X decreased such effect. The role of PKC-ζ was recognized by the use of a synthetic myristoylated peptide with sequences based on the endogenous PKC-ζ pseudosubstrate and by PKC-ζ downregulation using the small interfering RNA (siRNA). Insulin had a replicating effect on PC Cl3 cells, induced the phosphorylation of PKB, decreased p27kip expression and had no effect on the PKC-ζ cytosol-to-membrane translocation. PC Cl3 cell proliferation was induced more potently by simultaneous stimulation with insulin and Ang II than by stimulation with insulin alone, and the effect on p27kip expression was similar to that obtained with insulin only. These observations demonstrate that in PC Cl3 cells Ang II causes a block in G1 phase, although both ERK and PKB pathways are activated, and this effect may be due to the upregulation of p27kip and PKC-ζ operativity.
A possible role for melanocortin peptides in longitudinal growth
Authors: Yeh James K , Evans Jodi F , Niu Qing-Tian , Aloia John F
Published: December 2006

Clinical and in vitro data suggest a link between the elevation of the melanocortin peptide, ACTH, and longitudinal growth. Overproduction of ACTH in familial glucocorticoid deficiency (FGD) is associated with increased growth and ACTH increases the differentiation of chondrocytes along the endochondral pathway in vitro. Using the leptin-deficient obese (ob/ob) mouse along with lean control littermates (n = 9–10), we investigated the effects of adrenalectomy (ADX)-induced elevated ACTH with and without peripheral administration of the MC3-R-specific agonist, γ2-melanocyte stimulating hormone (γ2-MSH), on longitudinal growth. Naso-anal and tibial growth were measured together with growth plate parameters; both total and zonal heights together with the proliferative index. Data were analyzed using two-way ANOVA with post hoc comparisons made using the Bonferroni correction. ADX significantly increased naso-anal length in lean mice and ADX plus γ2-MSH administration significantly increased naso-anal length above ADX alone in ob/ob mice. γ2-MSH administration to ADX lean and ob/ob mice significantly increased tibial length. In ob/ob mice, these changes occurred in the context of reduced food intake. Analysis of total and zonal growth plate heights suggest an increase in hypertrophic differentiation and an overall increase in growth plate turnover in ADX lean and ob/ob mice. These in vivo data show that ADX enhances linear growth and the results of γ2-MSH treatment suggest that the melanocortin system plays a role in linear growth.