Thiol-inducing and immunoregulatory effects of flavonoids in peripheral blood mononuclear cells from patients with end-stage diabetic nephropathy

The cellular thiol status and its relationship to T-cell activation and cytokine synthesis of mononuclear cells was investigated in patients with end-stage diabetic nephropathy (ESDN) undergoing dialysis treatment. The functional effects of thiol repair by in vitro and in vivo treatment with flavonoids were elucidated. The thiol status of peripheral blood lymphocytes from 30 ESDN patients on hemodialysis and healthy controls was determined by flow cytometry. T-cell activation in response to pokeweed mitogen was analyzed by CD69 expression; cytokines were determined in cell culture supernatants. In result, compared to age-matched healthy subjects, a significant thiol deficiency in ESDN patients was obvious. The lowered total intracellular thiol levels correlated directly to a significant diminished T-cell activation and an elevated synthesis of TNF-alpha in the patient group. The treatment with flavonoids led to a restoration of the thiol status within 72 h in vitro and in vivo. This effect showed a biphasic kinetic that first utilized cell surface thiols and secondly intracellular thiols. In parallel, the T-cell activation was improved substantially along with a significant decrease in TNF-alpha release. These data provide the rational for clinical trials using flavonoids in ESDN to normalize immunoregulatory defects via restoration of the cellular thiol status.     

Fast large scale oligonucleotide selection using the longest common factor approach

We present a fast method that selects oligonucleotide probes (such as DNA 25-mers) for microarray experiments on a truly large scale. For example, reliable oligos for human genes can be found within four days, a speedup of one to two orders of magnitude compared to previous approaches. This speed is attained by using the longest common substring as a specificity measure for candidate oligos. We present a space- and time-efficient algorithm, based on a suffix array with additional information, to compute matching statistics (lengths of longest matches) between all candidate oligos and all remaining sequences. With the matching statistics available, we show how to incorporate constraints such as oligo length, melting temperature, and self-complementarity into the selection process at a postprocessing stage. As a result, we can now design custom oligos for any sequenced genome, just as the technology for on-site chip synthesis is becoming increasingly mature.     

Effective proliferation of human regulatory T cells requires a strong costimulatory CD28 signal that cannot be substituted by IL-2

The strength of immune repression by regulatory T (Treg) cells is thought to depend on the efficiency of Treg cell activation. The stimuli and their individual strength required to activate resting human Treg cells, however, have so far not been elucidated in detail. We reveal here that induction of proliferation of human CD4(+)C25(+) Treg cells requires an extraordinary strong CD28 costimulatory signal in addition to TCR/CD3 engagement. CD28 costimulation, noteworthy, cannot be substituted by IL-2 to induce proliferation of Treg cells, which is in contrast to CD4(+)CD25(-) T cells. IL-2, in contrast, prevents spontaneous apoptosis of Treg cells, but does not initiate their amplification. IL-2 and CD28 costimulation clearly exhibit disparate effects on Treg cells which are in contrast to those on CD4(+)CD25(-) T cells. Moreover, the prerequisites for Treg cell proliferation differ strikingly from those for effector T cells, implying a balanced orchestration in initiating and limiting a T cell immune response. In addition, data are of relevance for the design of therapeutic strategies involving IL-2 administration and CD28 costimulation.     

Caterpillar regurgitant induces pore formation in plant membranes

Formation of channel-like pores in a plant membrane was induced within seconds after application of an aqueous solution containing regurgitant of the insect larvae Spodoptera littoralis. Gated pore currents recorded on the tonoplast of the Charophyte Chara corallina displayed conductances up to several hundred pS. A voltage-dependent gating reaction supports the assumption that pore-forming molecules have amphipathic properties. Regurgitant samples separated into masses smaller or larger than 3kDa were evaluated by patch-clamp and mass spectroscopy. Fractions containing peptides larger than 3kDa constituted pores of large conductances, peptides smaller than 3kDa constituted pores of small conductances. Peptide-free eluates did not constitute conducting pores, indicating that pore-forming components in regurgitant are membrane-spanning oligopeptides.     

Synergism between mutational meltdown and Red Queen in parthenogenetic biotypes of the freshwater planarian Schmidtea polychroa

Do parasites and accumulation of deleterious mutations act synergistically in balancing the costs of sex? We addressed this possibility in the freshwater planarian flatworm Schmidtea polychroa. Sexual and parthenogenetic forms of this species sometimes coexist but show no ecological separation. Previous studies indicate that in a mixed sexual/ parthenogenetic population in Lago di Caldonazzo (N. Italy) parthenogens get more frequently infected with parasites. At the same time, they suffer from higher embryo mortality, which has been interpreted as a sign of accumulation of deleterious mutations. In the present study, we test whether these two factors are correlated, by focusing on the differences among the clonal lineages of a predominantly parthenogenetic subpopulation. Our results suggest that, for two out of three parasite types found, the infections are positively associated with the indirect measure of host mutation load.     

Spatial behaviour in the Aquatic Warbler (Acrocephalus paludicola) during mating and breeding

Summary The range use of both sexes of the Aquatic Warbler was studied during two breeding seasons by radio-tracking. Males used home ranges of up to 8 ha. These home ranges over-lapped by up to 74 %, (mean 51 %) between individuals. During the mating period and prior to nesting, females used isolated sites of 2.8 to 6.4 ha (mean: 4.2 ha) within activity ranges of 100 to 160 ha (mean: 120 ha). Home ranges of different females overlapped by up to 20 %. However, during nest-building and incubation, the average size of home ranges in females was only 1.6 ha. Since no constant ranges were observed, we conclude that Aquatic Warblers are not territorial in the breeding season.

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Räumliches Verhalten des Seggenrohrsängers (Acrocephalus paludicola) während Partnerfindung und Bebrütung

Zusammenfassung In zwei aufeinanderfolgenden Brutzeiten wurde die Raumnutzung von Seggenrohrsängern mittels Telemetrie untersucht. Die ermittelte Größe der Streifgebiete von Männchen betrug in der gesamten Brutzeit bis zu 8 ha. In dem Streifgebiet eines einzelnen Männchens wurden bis zu elf telemetrierte Männchen festgestellt. Die Überlappungen der Streifgebiete telemetrierter Männchen betrugen durchschnittlich 51 %, maximal 74 %. Weibchen nutzten während der Paarungszeit nicht zusammenhängende inselartige Areale von insgesamt 2,8 bis 6,4 ha (Mittelwert 4,2 ha) innerhalb von Aktionsräumen von 110 bis 160 ha (Mittelwert 120 ha). Diese Areale überlappten zu 20 % mit den Aufenthaltsgebieten anderer Weibchen. Während des Brütens sank die Größe der Streifgebiete der Weibchen auf durchschnittlich 1,6 ha. Da im Beobachtungszeitraum keine konstant genutzten Aufenthaltsgebiete vorlagen, schließen wir, daß sich Seggenrohrsänger in der Brutzeit nicht territorial verhalten.

     

Evolution of ITS1 rDNA in the Digenea (Platyhelminthes: Trematoda): 3′ End Sequence Conservation and Its Phylogenetic Utility

A comparison of ribosomal internal transcribed spacer 1 (ITS1) elements of digenetic trematodes (Platyhelminthes) including unidentified digeneans isolated from Cyathura carinata (Crustacea: Isopoda) revealed DNA sequence similarities at more than half of the spacer at its 3′ end. Primary sequence similarity was shown to be associated with secondary structure conservation, which suggested that similarity is due to identity by descent and not chance. Using an analysis of apomorphies, the sequence data were shown to produce a distinct phylogenetic signal. This was confirmed by the consistency of results of different tree reconstruction methods such as distance approaches, maximum parsimony, and maximum likelihood. Morphological evidence additionally supported the phylogenetic tree based on ITS1 data and the inferred phylogenetic position of the unidentified digeneans of C. carinata met the expectations from known trematode life-cycle patterns. Although ribosomal ITS1 elements are generally believed to be too variable for phylogenetic analysis above the species or genus level, the overall consistency of the results of this study strongly suggests that this is not the case in digenetic trematodes. Here, 3′ end ITS1 sequence data seem to provide a valuable tool for elucidating phylogenetic relationships of a broad range of phylogenetically distinct taxa. Received: 20 October 1997 / Accepted: 24 March 1998     

Calcium-ganglioside interactions and synaptic plasticity: effect of calcium on specific ganglioside/peptide (valinomycin, gramicidin A)-complexes in mixed mono- and bilayers.

A controlled exchange of calcium between the extracellular space (mM Ca2+) and the neuroplasm (microM Ca2+) is considered to be an essential prerequisite for almost every stage of neuronal activity. Our research interest is focused on those compounds, which due to their physico-chemical properties and localization within the synaptic membrane might fulfill the task as neuromodulators for functional synaptic proteins. Because of this specific binding properties towards calcium and their peculiar interactions with calcium in model systems gangliosides (amphiphilic sialic acid containing glycosphingolipids) are favorite candidates for a functional involvement in synaptic transmission of information. In this study we used monolayers to investigate the molecular packing and surface potential at the air/water interface, the interaction of gangliosides with the depsipeptide valinomycin (= monovalent ion carrier), and its influenceability by calcium. Furthermore we looked at calcium effects on the single channel conductance and mean channel life-time of the monovalent ion channel gramicidin A in mixed PC/ganglioside bilayers. In pure ganglioside monolayers the addition of 0.01 mM Ca2+ induces monolayer condensation, a rise in collapse pressure (= higher film stability), a shift of phase transition (= change of conformation), and a more negative head group potential (change of electric properties). In mixed ganglioside-valinomycin monolayers the addition of Ca2+ causes phase separation and/or aggregate formation between the ganglioside and the peptide. Single channel conductance fluctuations as well as mean channel life-time were analyzed for gramicidin A incorporated into binary mixed black lipid membranes of negatively charged gangliosides (GM1, GD1a, GT1b, GMix) and neutral lecithin (DOPC) in different molar ratios. At monovalent electrolyte concentrations up to < 250 mM CsCl the single channel conductance was significantly larger in the negatively charged mixed DOPC/ganglioside membranes than in the neutral DOPC membrane. Additionally, in the presence of gangliosides the mean channel life-time is increased. The addition of calcium (0.05 mM) induced a reduction of single channel conductance of gramicidin A in DOPC- and mixed DOPC/ganglioside membranes. These physico-chemical data in connection with new electromicroscopical evidences for a precise localization of calcium, a calcium pump (Ca(2+)-ATPase), a clustered arrangement of gangliosides in synaptic terminals, and biochemical results with regard to activatory nature of exogenous gangliosides for neuronal protein phosphorylation and ATPases, support the hypothesis of a modulatory function of gangliosides in synaptic transmission.