Showing papers by "Ludwig Maximilian University of Munich published in 1995"

Toward a better definition of the restless legs syndrome

Abstract: A large International Restless Legs Syndrome (RLS) Study Group has been formed. As its first task, the group has taken upon itself the role of definig the clinical features of the RLS. As minimal criteria for diagnosis, the group proposes the following four features: (a) desire to move the extremities, often associated with paresthesias/dysesthesias; (b) motor restlessness; (c) worsening of symptoms at rest with at least temporary relief by activity, and (d) worsening of symptoms in the evening or night. Other features commonly seen in RLS include sleep disturbance, periodic limb movements in sleep and similar involuntary movements while awake, a normal neurological examination in the idiopathic from, a tendency for the symptoms to be worse in middle to older age, and, in some cases, a family history suggestive of an autosomal dominant mode of inheritance.

A gene (PEX) with homologies to endopeptidases is mutated in patients with X–linked hypophosphatemic rickets

Abstract: X–linked hypophosphatemic rickets (HYP) is a dominant disorder characterised by impaired phosphate uptake in the kidney, which is likely to be caused by abnormal regulation of sodium phosphate cotransport in the proximal tubules. By positional cloning, we have isolated a candidate gene from the HYP region in Xp22.1. This gene exhibits homology to a family of endopeptidase genes, members of which are involved in the degradation or activation of a variety of peptide hormones. This gene (which we have called PEX) is composed of multiple exons which span at least five cosmids. Intragenic non–overlapping deletions from four different families and three mutations (two splice sites and one frameshift) have been detected in HYP patients, which suggest that the PEX gene is involved in the HYP disorder.

Time structure of the activity in neural network models

Abstract: Several neural network models in continuous time are reconsidered in the framework of a general mean-field theory which is exact in the limit of a large and fully connected network. The theory assumes pointlike spikes which are generated by a renewal process. The effect of spikes on a receiving neuron is described by a linear response kernel which is the dominant term in a weak-coupling expansion. It is shown that the resulting ``spike response model'' is the most general renewal model with linear inputs. The standard integrate-and-fire model forms a special case. In a network structure with several pools of identical spiking neurons, the global states and the dynamic evolution are determined by a nonlinear integral equation which describes the effective interaction within and between different pools. We derive explicit stability criteria for stationary (incoherent) and oscillatory (coherent) solutions. It is shown that the stationary state of noiseless systems is ``almost always'' unstable. Noise suppresses fast oscillations and stabilizes the system. Furthermore, collective oscillations are stable only if the firing occurs while the synaptic potential is increasing. In particular, collective oscillations in a network with delayless excitatory interaction are at most semistable. Inhibitory interactions with short delays or excitatory interactions with long delays lead to stable oscillations. Our general results allow a straightforward application to different network models with spiking neurons. Furthermore, the theory allows an estimation of the errors introduced in firing rate or ``graded-response'' models.

Mirror Symmetry, Mirror Map and Applications to Calabi-Yau Hypersurfaces

Abstract: Mirror Symmetry, Picard-Fuchs equations and instanton corrected Yukawa couplings are discussed within the framework of toric geometry. It allows to establish mirror symmetry of Calabi-Yau spaces for which the mirror manifold had been unavailable in previous constructions. Mirror maps and Yukawa couplings are explicitly given for several examples with two and three moduli.

Cardiac Release of Cytokines and Inflammatory Responses in Acute Myocardial Infarction

Abstract: Background In animal models of myocardial infarction (MI), inflammatory responses compromise microcirculation during reperfusion and restrict functional recovery. To investigate cardiac inflammator...

A stress-inducible 72-kDa heat-shock protein (HSP72) is expressed on the surface of human tumor cells, but not on normal cells.

Abstract: It is suggested that members of the heat-shock protein (HSP) 70 and 90 families are involved in intracellular antigen processing and the presentation of cell-membrane-anchored antigens. We show that non-lethal heat shock (41.8 degrees C) causes comparable rates of HSP72 (about 20x) and HSP73 (about 3x) synthesis in both tumor (including human Ewing's sarcoma, ES and osteosarcoma cells, HOS58) and normal cells (including EBV-transformed B-LCL, PBL and fibroblasts derived from healthy human volunteers). However, following non-lethal heat stress and a recovery period at 37 degrees C, flow cytometric analysis with a specific MAb showed HSP72 to be expressed only on the cell surface of tumor cells. The cell-surface localization of HSP72 was confirmed by Western-blot analysis of separated membranes and by immunoprecipitation with the HSP72-specific MAb. In addition, co-incubation of untreated tumor cells with supernatants from lethally heat-shocked cells, which contain HSP72, did not lead to HSP72 cell-surface expression. Thus, non-specific association of HSP72 molecules with the outer plasma membrane is unlikely. In conclusion, despite comparable cytoplasmic HSP72 induction, human tumor cells differ from normal cells in their capacity to express HSP72 on their surface. This might imply clinical application as a means to target a stress-inducible, tumor-specific immune response.

The octadecanoic pathway: signal molecules for the regulation of secondary pathways.

Abstract: Plant defense against microbial pathogens and herbivores relies heavily on the induction of defense proteins and low molecular weight antibiotics. The signals between perception of the aggression, gene activation, and the subsequent biosynthesis of secondary compounds are assumed to be pentacylic oxylipin derivatives. The rapid, but transient, synthesis of cis-jasmonic acid was demonstrated after insect attack on a food plant and by microbial elicitor addition to plant suspension cultures. This effect is highly specific and not caused by a number of environmental stresses such as light, heavy metals, or cold or heat shock. Elicitation of Eschscholtzia cell cultures also led to a rapid alkalinization of the growth medium prior to jasmonate formation. Inhibition of this alkalinization process by the protein kinase inhibitor staurosporine also inhibited jasmonate formation. The induction of specific enzymes in the benzo[c]phenanthridine alkaloid pathway leading to the antimicrobial sanguinarine was induced to a qualitatively and quantitatively similar extent by fungal elicitor, methyl jasmonate, and its linolenic acid-derived precursor 12-oxophytodienoic acid. It is herein proposed that a second oxylipid cascade may exist in plants starting from linoleic acid via 15,16-dihydro-12-oxophytodienoic acid to 9,10-dihydrojasmonate. Experiments with synthetic trihomojasmonate demonstrated that beta-oxidation is not a prerequisite for biological activity and that 12-oxophytodienoic acid and derivatives are most likely fully active as signal transducers. Octadecanoic acid-derived compounds are essential elements in modulating the synthesis of antibiotic compounds and are thus integral to plant defense.

The evolution of vortices in vertical shear. I: Initially barotropic vortices

Abstract: The behaviour of initially-barotropic vortices in vertically-sheared environmental flows is investigated. the strength and structure of the vortices used are representative of tropical cyclones. the calculations are performed using a primitive-equation numerical model on an f-plane. It is found that the initial response of the vortex to the vertical shear is to tilt in the plane of the shear. As soon as a tilt is established, the upper- and lower-level centres begin to rotate cyclonically about the mid-level centre. This rotation can be understood in terms of upper- and lower-level potential-vorticity anomalies which are displaced in the horizontal relative to one another. the flow associated with the vertical projection of each anomaly advects the other anomaly, leading to the observed cyclonic rotation. the rotation rate decreases with time, so that the direction of tilt becomes constant, but the magnitude of the tilt continues to increase. We argue that the observed rotation acts to oppose the destructive action of the vertical shear on the vortex, even in the absence of diabatic processes. The role of the vertical circulation is considered in detail. It is shown that the vertical circulation develops in a manner which is consistent with the model flow remaining balanced. It is found that the mesoscale nature of the vertical circulation leads to a distortion of the axisymmetric vortex. This results in the inner core having a smaller vertical tilt than the outer region. the vertical circulation does not act on a large enough scale to explain why the vortex is not destroyed by the vertical shear. The behaviour of the vortex is found to depend on various parameters. Results are presented where the vertical shear, the strength and size of the vortex, the Coriolis parameter, and the static stability are varied. With the exception of the vertical shear, altering any of these parameters alters the vertical penetration of a potential-vorticity anomaly. the results show that increasing the penetration depth leads to an increase in the rotation rate of the upper- and lower-level vortex centres about the mid-level centre, and to a reduction in the magnitude of the vertical tilt.

High‐level Biosynthetic Substitution of Methionine in Proteins by its Analogs 2‐Aminohexanoic Acid, Selenomethionine, Telluromethionine and Ethionine in Escherichia coli

Abstract: We have utilized a T7 polymerase/promoter system for the high-level incorporation of methionine analogs with suitable labels for structural research (X-ray and NMR studies) on recombinant annexin V produced in Escherichia coli. Here, we describe, to our knowledge, the first biosynthetic high-level substitution of methionine by 2-aminohexanoic acid (norleucine), ethionine and telluromethionine in a protein. The replacement has been confirmed by electrospray mass spectroscopy, amino acid analysis and X-ray structural analysis. Conditions for expression were optimized concerning the frequency of appearance of revertants, high-level replacement and maximal protein yield. For the incorporation of norleucine and ethionine, E. coli B834 (DE3)(hsd metB), which is auxotrophic for methionine, was grown under methionine-limited conditions with an excess of the analog in the culture medium, and the expression of protein under the control of the T7 promoter was induced after the methionine supply had been exhausted. The factor limiting the high-level incorporation of telluromethionine into protein is its sensitivity towards oxidation. To overcome this problem, bacteria were grown with a limited amount of methionine, harvested after its exhaustion and resuspended in fresh media without methionine; telluromethionine was added and protein synthesis induced. Under these conditions, significant amounts of protein can be expressed before telluromethionine has been completely degraded (within hours). Biosynthetic incorporation of heavy atoms such as tellurium into recombinant proteins can accelerate the process of obtaining heavy-atom derivatives suitable for X-ray structural analysis, supplementing the traditional trial-and-error preparation of heavy-atom derivatives for the method of multiple isomorphous replacement. Furthermore, the successful high-level incorporation of amino acid analogs can provide single-atom mutations for the detailed study of the structure and function of proteins.

Structure and functional expression of a new member of the tetrodotoxin-sensitive voltage-activated sodium channel family from human neuroendocrine cells.

Abstract: A member of a new subclass of the voltage-activated sodium channel genes has been cloned from the human medullary thyroid carcinoma (hMTC) cell line. The cDNA of hNE-Na (human neuroendocrine sodium channel) encodes a 1977 amino acid protein which phylogenetically represents a link between sodium channels isolated from skeletal muscle and brain. The hNE-Na alpha subunit was transiently expressed in human embryonic kidney cells either alone or in combination with the human sodium channel beta 1 subunit. The channel exhibited rapid activation and inactivation kinetics, and was blocked by tetrodotoxin and cadmium with IC50 values of 24.5 nM and 1.1 mM, respectively. Action potentials were generated in cells expressing high levels of hNE-Na. Northern blot and reverse transcription-polymerase chain reaction (RT-PCR) analyses demonstrated its expression in hMTC cells, in a C-cell carcinoma, and in thyroid and adrenal gland. Transcripts were not identified in pituitary gland, brain, heart, liver or kidney, indicating that the hNE-Na is a sodium channel solely expressed in neuroendocrine cells.

Sequestration and recycling of beta 2-adrenergic receptors permit receptor resensitization.

Abstract: Stimulation of beta 2-adrenergic receptors in intact cells causes, first, rapid functional uncoupling from Gs, which is triggered by receptor phosphorylation, and, second, somewhat slower sequestration of the receptors to an internal compartment. The present study addresses a possible role of sequestration in the resensitization of desensitized beta 2-adrenergic receptors in human A431 cells. Exposure of these cells to isoproterenol caused rapid phosphorylation, desensitization (as assessed in adenylyl cyclase assays), and sequestration of the receptors. Subsequent removal of the agonist led to recycling of the receptors to the cell surface, dephosphorylation, and restoration of receptor function. These effects occurred without any change in the total receptor number. The rate constant of agonist-induced sequestration was 0.03/min; the rate constant of receptor recycling was 0.06/min and was not markedly altered by the presence of agonist. Blockade of sequestration with concanavalin A or 0.6 M sucrose prevented receptor dephosphorylation as well as receptor resensitization. Inhibition of protein phosphatases with calyculin A caused a similar blockade of beta 2-adrenergic receptor resensitization; the effects of maximally effective concentrations of concanavalin A and calyculin A were not additive. Monensin impaired recycling of desensitized beta 2-adrenergic receptors to the cell surface and also prevented receptor resensitization. We conclude that sequestration of beta 2-adrenergic receptors, followed by dephosphorylation and recycling to the cell surface, may serve to restore the function of desensitized receptors.

D* D pi and B* B pi couplings in QCD

Abstract: We calculate the ${\mathit{D}}^{\mathrm{*}}$D\ensuremath{\pi} and ${\mathit{B}}^{\mathrm{*}}$B\ensuremath{\pi} couplings using QCD sum rules on the light cone. In this approach, the large-distance dynamics is incorporated in a set of pion wave functions. We take into account two-particle and three-particle wave functions of twist 2, 3, and 4. The resulting values of the coupling constants are ${\mathit{g}}_{\mathit{D}}^{\mathrm{*}}$D\ensuremath{\pi}=12.5\ifmmode\pm\else\textpm\fi{}1 and ${\mathit{g}}_{\mathit{B}}^{\mathrm{*}}$B\ensuremath{\pi}=29\ifmmode\pm\else\textpm\fi{}3. From this we predict the partial width \ensuremath{\Gamma}(${\mathit{D}}^{\mathrm{*}+}$\ensuremath{\rightarrow}${\mathit{D}}^{0}$${\mathrm{\ensuremath{\pi}}}^{+}$)=32\ifmmode\pm\else\textpm\fi{}5 keV. We also discuss the soft-pion limit of the sum rules which is equivalent to the external axial field approach employed in earlier calculations. Furthermore, using ${\mathit{g}}_{\mathit{B}}^{\mathrm{*}}$B\ensuremath{\pi} and ${\mathit{g}}_{\mathit{D}}^{\mathrm{*}}$D\ensuremath{\pi} the pole dominance model for the B\ensuremath{\rightarrow}\ensuremath{\pi} and D\ensuremath{\rightarrow}\ensuremath{\pi} semileptonic form factors is compared with the direct calculation of these form factors in the same framework of ligh-cone sum rules.

Effects of forest liming on soil processes

Abstract: On the basis of a field experiment in Norway spruce with acid irrigation and compensatory liming of the soil surface (Hoglwald, S-Bavaria), liming effects are described as lime dissolution rate, transformation of carbonate buffer to exchange buffer, time required for deacidification of soil and drainage water, mobilization of Cu and Pb, changes in soil organisms, humus decomposition, and nitrogen turnover. It was shown that lime dissolution followed an exponentially decreasing curve. 4 t ha-1 dolomitic lime were dissolved within 6 years. Additional acid irrigation of 4 kmol H+ ha-1 yr-1 as sulphuric acid speeded up the lime dissolution to about 4 years. After dissolution of lime about 70% of Ca and about 30% of Mg, both originating from lime dissolution, are retained in the surface humus layer, loading the exchange buffer capacity there. Liming acted as a protection against acid irrigation but the extension of soil deacidification downwards proceeded slowly due to the high base neutralizing capacity of protonated functional groups of the organic matter. The main depth effect is caused by Mg translocation. A significant increase of organic Cu complexes occurred due to mobilization of water soluble humus decomposition products. The effect of liming on litter decomposing organisms is demonstrated with microorganisms, collembolae and earthworms regarding the abundance and the structure of dominance. It was shown that liming may induce unusually large changes in biocenoses of forest soils. The decay of surface humus accounted for 7.2 t ha-1 or 23% of the store within 7 years. Within the same time span, liming caused a loss of about 170 kg N ha-1 or 14% of the store of the surface humus layer. The nitrate concentration in the drainage water thus increased by about 50 to 60 mg NO3 - L-1. Site-specific conditions are discussed, which produce such negative liming effects as increased nitrate concentration of seepage, humus decay and heavy metal mobilization. Redistribution of tree roots, induction of boron deficiency and root rot are also considered. It is indicated that liming may aggravate the increasing problem of nitrate contamination of forest ground water resources which is associated with deposition of atmogenous nitrogen compounds. Some recommendations are given regarding forest practice.

Use of polymerase chain reaction epitope tagging for protein tagging in Saccharomyces cerevisiae

Abstract: Epitope tagging is the insertion of a short stretch of amino acids constituting an epitope into another protein. Tagged proteins can be identified by Western, immunoprecipitation and immunofluorescence assays using pre-existing antibodies. We have designed vectors containing the URA3 gene flanked by direct repeats of epitope tags. We use the polymerase chain reaction (PCR) to amplify the tag-URA3-tag cassette such that the ends of the PCR fragments possess homology to the gene of interest. In vivo recombination is then used to direct integration of the fragment to the location of interest, and transformants are selected by their Ura+ phenotype. Finally, selection for Ura- cells on 5-fluoro-orotic acid plates yields cells where recombination between the repeated epitopes has 'popped out' the URA3 gene, leaving a single copy of the epitope at the desired location. PCR epitope tagging (PET) provides a rapid and direct technique for tagging that does not require any cloning steps. We have used PET to tag three Saccharomyces cerevisiae proteins, Cln1, Sic1 and Est1.

Ribosomal DNA phylogeny of the major extant arthropod classes and the evolution of myriapods

Abstract: THE evolutionary relationships among arthropods are of particular interest because the best-studied model system for ontogenetic pattern formation, the insect Drosophila, is a member of this phylum. Evolutionary inferences about the developmental mechanisms that have led to the various designs of the arthropod body plan depend on a knowledge of the phylogenetic framework of arthropod evolution. Based on morphological evidence1–3, but also on palaeonto-logical considerations4, the sister group of the insects is believed to be found among the myriapods. Using nuclear ribosomal gene sequences for constructing a molecular phylogeny, we provide strong evidence that the crustaceans and not the myriapods should be considered to be the sister group of the insects. Moreover, the degree of sequence divergence suggests that the diversification of the myriapods occurred during the Cambrian, Our findings have general implications for the course of land colonization by the different arthropod groups, as well as for the interpretation of primitive and derived features of arthropod morphology.

Salivary cortisol levels throughout childhood and adolescence: relation with age, pubertal stage, and weight.

Abstract: The measurement of cortisol in saliva has become a reliable tool for both the scientist and the clinician for studying adrenal cortical function in the adult. We have measured salivary cortisol in samples from 138 healthy infants, children, and adolescents, and from 14 adults. Saliva samples were obtained at home using a cotton swab and a saliva-collecting tube at 800, 1300, and 1800 h before meals. Cortisol was measured using a time-resolved fluorescent immunoassay. Cortisol levels in saliva ranged from less than 2 nmol/L up to more than 100 nmol/L. Cortisol levels were age-dependent. Interestingly, after the age of 6 y, cortisol levels correlated significantly with pubertal stages (analysis of variance). No sex difference was found. In addition, cortisol morning levels and daily cortisol levels (area under the curve from three measurements) increased with body weight and body mass index. The highest cortisol levels were measured in saliva of children younger than 1 y. No circadian variation was evident before the age of 9 mo. After 1 y of age, salivary cortisol levels varied in a circadian fashion. The measurement of salivary cortisol levels is an attractive way of testing adrenal function in infants and children. It provides a reliable tool for the determination of the physiology and developmental characteristics of cortisol metabolism.

Flexible bat echolocation: the influence of individual, habitat and conspecifics on sonar signal design

Abstract: Acoustic signals which are used in animal communication must carry a variety of information and are therefore highly flexible. Echolocation has probably such functions and could prove as flexible. Measurable variabitlity can indicate flexibility in a behaviour. To quantify variability in bat sonar and relate to behavioural and environmental factors, I recorded echolocation calls of Euderma maculatum, Eptesicus fuscus, Lasiurus borealis and L. cinereus while the bats hunted in their natural habitat. I analysed 3390 search phase calls emitted by 16 known and 16 unknown individuals foraging in different environmental and behvioural situations. All four species used mainly multiharmonic signals that showed considerable intra- and inter-individual variability in the five signal variables I analysed (call duration, call interval, highest and lowest frequency and frequency with maximum energy) and also in the shape of the sonagram. A nested multivariate analysis of variance identified the influences of individual, hunting site, close conspecifics and of each observation on the frequency with maximum energy in the calls, and on other variables measured. Individual bats differed in multiple comparisons, most often in the main call frequency and least often in call interval. In a discriminant function analysis with resubstitution, 56–76% of a species' calls were assigned to the correct individual. Distinct individual call patterns were recorded in special situations in all species and the size of foraging areas in forested areas influenced temporal and spectral call structure. Echolocation behaviour was influenced by the presence of conspecifics. When bats were hunting together, call duration decreased and call interval increased in all species, but spectral effects were less pronounced. The role of morphometric differences as the source of individually distinct vocalizations is discussed. I also examined signal adaptations to long range echolocation and the influence of obstacle distance on echolocation call design. My results allow to discuss the problems of echo recognition and jamming avoidance in vespertilionid bats.

High prevalence of RET rearrangement in thyroid tumors of children from Belarus after the Chernobyl reactor accident

Abstract: RET rearrangement was studied in papillary thyroid carcinomas (PTC) of children exposed to radioactive fallout in Belarus after the Chernobyl accident. To detect RET rearrangement in small tissue samples from thyroidectomy specimen (12 PTC of children; 2 PTC and 1 follicular carcinoma of adults; non-tumorous thyroid tissue of 4 children and 4 adults as controls), a RT-multiplex PCR was developed using primers suited to amplify fragments in different quantities depending on the presence or absence of RET rearrangements in the tissues. The type of rearrangement was determined by RT-PCR and direct sequencing using primers for ret/PTC1, 2 and 3. Two-thirds of the papillary thyroid carcinomas of the children revealed a RET rearrangement, with ret/PTC3 being more frequent by a factor of 3 than ret/PTC1. ret/PTC2 was not detected. All RET rearrangement-positive tumors had lymph node metastasis while half of the tumors with wild-type cRET had not. More than half of the cases with ret/PTC3 expressed not only the ELE/RET transcript as expected, but also the RET/ELE transcript. Intrachromosomal rearrangement involving RET and the adjacent H4 or ELE gene on chromosome no. 10 is a very frequent event in thyroid cancer of children of the Chernobyl-contaminated zone of Belarus.