Growth and branching morphogenesis of rat collecting duct anlagen in the absence of metanephrogenic mesenchyme

The growth and differentiation of the epithelium in many tissues is mediated by interactions with the adjacent mesenchyme, but the mechanisms responsible remain undefined. To identify the factors involved in the growth and branching morphogenesis of ureteric bud, which is the collecting duct anlagen, buds from 13-gestation-day rat embryos were separated from the metanephrogenic mesenchyme and explanted to culture dishes coated with gelled type I collagen in a defined medium. Under these conditions buds attached to the substrate and grew out without indication of cell senescence. When buds were instead suspended in gelled type I collagen, branching morphogenesis was observed despite the absence of mesenchyme although it was not as extensive as in vivo. Since growth occurred much more slowly in culture than expected, culture conditions were varied in attempts to accelerate the process. Despite extensive screening of matrices and growth factors, only epidermal and endothelial cell growth factors stimulated growth to a significant extent. Transforming growth factor-beta, on the other hand, was a potent inhibitor of growth. Homogenates from tumors that caricature metanephrogenic mesenchyme were highly mitogenic for bud cells and, thus, will be a source of material for characterizing regulatory factors involved in renal growth. These studies show that growth and branching morphogenesis of the ureteric bud can occur without direct cell-cell interactions with the metanephrogenic mesenchyme and that matrices and factors secreted by the mesenchyme may mediated these activities in vivo.     

Analysis of intercellular washing fluids of potato tubers and detection of increased proteolytic activity upon fungal infection

We present the first procedure for extracting intercellular fluids of potato ( Solanum tuberosum L. cv. Spunta) tubers. Intercellular washing fluids were isolated from healthy and Fusarium ‐infected potato tissue. The electrophoretic pattern using SDS‐PAGE indicated differences between the fluids from the two tissues. A significant extracellular proteolytic activity was accumulated during the infection with Fusarium solani f. sp. eumartii . A major proteolytic band with an apparent molecular mass of 70 kDa and another of approximately 30 kDa were detected after separation of intercellular fluids by casein gel electrophoresis. Proteolytic activity was principally inhibited by diisopropylfluorophosphate, which is indicative of the involvement of serine protease(s). In vitro degradation assay indicated that specific potato proteins from healthy tubers were hydrolyzed by fluid proteases from infected tubers. The biological role of such activity in potato‐ Fusarium interaction is still unknown. Our results suggest that the intercellular serine protease has a fungal origin.     

Flower morphology and biology of Myrsine laetevirens, structural and evolutionary implications of anemophily in Myrsinaceae

This study deals with the phenology, pollination biology and floral morphology of Myrsine laetevirens , a neotropical dioecious tree. In Punta Lara (Argentina), its flowering period occurs during January-February. Both pistillate and staminate flowers are small, with a yellowish-green perianth and produce neither nectar nor odour. Staminate flowers have five stamens and a vestigial gynoecium while pistillate flowers, have non-functional anthers and a conspicuous stigma. The floral characteristics of staminate and pistillate plants are related to the syndrome of anemophily. Fruit set in inflorescences covered with mesh bags and observations prove that animals are not involved in the pollination process. A provisional cladistic analysis of Myrsinaceae shows that dioecy evolved as one of the most recent apomorphies of Myrsine and is part of the anemophilous syndrome.     

Structural requirements for the glycolipid receptor of human uropathogenic Escherichia coli

The binding of uropathogenic Escherichia coli to the globo series of glycolipids via P pili is a critical step in the infectious process that is mediated by a human-specific PapG adhesin. Three classes of PapG adhesins exist with different binding specificities to Galα4Gal-containing glycolipids. The structural basis for PapG recognition of the human glycolipid receptor globoside was investigated by using soluble saccharide analogues as inhibitors of bacterial haemagglutination. The minimum binding epitope was confirmed as the Galα4Gal moiety, but parts of the GalNAcβ and glucose residues, which flank the Galα4Gal in globoside (GbO₄), were also shown to be important for strong binding. Furthermore, the same five hydroxyl groups of Galα4Gal in globotriasyl ceramide that were recognized by a previously characterized PapG variant were also recognized by the human-specific PapG in binding the GbO₄ that dominates In the human kidney. Saccharide analogues that blocked haemagglutination also blocked the adherence of human uropathogenic E. coli to human kidney sections. Knowledge of the molecular details of the PapG-GbO₄ interaction will make it possible to design antiadherence therapeutics.     

Regulation of glutathione synthesis in leaves of transgenic poplar (Populus tremula X P. alba) overexpressing glutathione synthetase

The poplar hybrid Populus tremula X P. alba was transformed with the Escherichia coli gene for glutathione synthetase ( gsh II ) targetted to the cytosol. Leaves of five lines of transgenic plants exhibited glutathione synthetase activities 15- to 60-fold higher than those of wild-type plants. Total glutathione levels and GSH/GSSG ratios were similar in transgenic and wild-type plants. Precursor feeding experiments with cysteine and γ-glutamylcysteine suggest that glutathione synthesis in the cytoplasm is controlled by a multistep procedure that includes (i) the availability of cysteine, (ii) the availability of γ-glutamylcysteine, and (iii) regulation of the activities of both γ-glutamylcysteine synthetase and glutathione synthetase. However step (ii) may set an upper limit for the cellular glutathione content.     

Interaction of bracken-fern extract with vitamin C in human submandibular gland and oral epithelium cell lines

The consumption of bracken-fern (Pteridium aquilinum) as food is associated with a high incidence of cancer in humans and animals. Thus far, the carcinogenic effects of bracken-fern consumption could be related to chromosome aberrations verified in animal and in human peripheral lymphocytes. We tested the in vitro effects of vitamin C (10 and 100 microg/ml) on the reversibility of DNA damage caused by bracken-fern on human submandibular gland (HSG) cells and on oral epithelium cells (OSCC-3) previously exposed to bracken-fern extract. DNA damage (i.e. nuclei with increased levels of DNA migration) was determined by comet assay, cell morphology was evaluated by light microscopy and cellular degeneration was assessed by the acridine orange/ethidium bromide fluorescent-dyeing test. Results showed that vitamin C alone did not reduce DNA damage caused by bracken-fern in HSG and OSSC-3 cells. However, at a higher concentration (100 microg/ml), vitamin C induced DNA damage in both cell lines. Moreover, vitamin C (10 and 100 microg/ml) together with bracken-fern extract showed synergistic effects on the frequency of DNA damage in HSG cells. In addition, cells treated with bracken-fern extract or vitamin C alone, or with their association, showed apoptosis morphological features, such as chromatin condensation, cytoplasmic volume loss, changes in membrane symmetry and the appearance of vacuoles; these alterations were observed in both cell lines. These results demonstrate that bracken-fern extract was cytotoxic to HSG and OSCC-3 cells, causing cell death by apoptosis, and that vitamin was not able to revert these effects.     

Regulation of the water status in three co-occurring phreatophytes at the southern fringe of the Taklamakan Desert

Aims We investigated the regulation of the water status in three predominant perennial C₃ phreatophytes (Alhagi sparsifolia, Populus euphratica, Tamarix ramosissima) at typical sites of their occurrence at the southern fringe of the hyperarid Taklamakan Desert (north-west China).Methods In the foreland of the river oasis of Qira (Cele), we determined meteorological variables, plant biomass production, plant water potentials (Ψ L) and the water flux through the plants. We calculated the hydraulic conductance on the flow path from the soil to the leaves (k SL) and tested the effects of k SL, Ψ L and the leaf-to-air difference in the partial pressure of water vapour (Δ w) on stomatal regulation using regression analyses.Important findings Despite high values of plant water potential at the point of turgor loss, all plants sustained Ψ L at levels that were high enough to maintain transpiration throughout the growing season. In A. sparsifolia, stomatal resistance (r s; related to leaf area or leaf mass) was most closely correlated with k SL; whereas in P. euphratica, ~70% of the variation in r s was explained by Δ w. In T. ramosissima, leaf area-related r s was significantly correlated with Ψ L and k SL. The regulation mechanisms are in accordance with the growth patterns and the occurrence of the species in relation to their distance to the ground water.     

Spatial isolation and genetic differentiation in naturally fragmented plant populations of the Swiss Alps

Aims The effect of anthropogenic landscape fragmentation on the genetic diversity and adaptive potential of plant populations is a major issue in conservation biology. However, little is known about the partitioning of genetic diversity in alpine species, which occur in naturally fragmented habitats. Here, we investigate molecular patterns of three alpine plants (Epilobium fleischeri, Geum reptans and Campanula thyrsoides) across Switzerland and ask whether spatial isolation has led to high levels of population differentiation, increasing over distance, and a decrease of within-population variability. We further hypothesize that the contrasting potential for long-distance dispersal (LDD) of seed in these species will considerably influence and explain diversity partitioning.Methods For each study species, we sampled 20–23 individuals from each of 20–32 populations across entire Switzerland. We applied Random Amplified Polymorphic Dimorphism markers to assess genetic diversity within (Nei's expected heterozygosity, H e; percentage of polymorphic bands, P p) and among (analysis of molecular variance, Φ st) populations and correlated population size and altitude with within-population diversity. Spatial patterns of genetic relatedness were investigated using Mantel tests and standardized major axis regression as well as unweighted pair group method with arithmetic mean cluster analyses and Monmonier's algorithm. To avoid known biases, we standardized the numbers of populations, individuals and markers using multiple random reductions. We modelled LDD with a high alpine wind data set using the terminal velocity and height of seed release as key parameters. Additionally, we assessed a number of important life-history traits and factors that potentially influence genetic diversity partitioning (e.g. breeding system, longevity and population size).Important findings For all three species, we found a significant isolation-by-distance relationship but only a moderately high differentiation among populations (Φ st : 22.7, 14.8 and 16.8%, for E. fleischeri, G. reptans and C. thyrsoides, respectively). Within-population diversity (H e : 0.19–0.21, P p : 62–75%) was not reduced in comparison to known results from lowland species and even small populations with <50 reproductive individuals contained high levels of genetic diversity. We further found no indication that a high long-distance seed dispersal potential enhances genetic connectivity among populations. Gene flow seems to have a strong stochastic component causing large dissimilarity between population pairs irrespective of the spatial distance. Our results suggest that other life-history traits, especially the breeding system, may play an important role in genetic diversity partitioning. We conclude that spatial isolation in the alpine environment has a strong influence on population relatedness but that a number of factors can considerably influence the strength of this relationship.     

Gypsy moth mating disruption in open landscapes

1 Aerial applications of Disrupt II, a plastic laminated flake formulation containing a racemic form of the gypsy moth sex pheromone, disparlure, achieved > 99% reduction of mating among females on individual, isolated trees surrounded by an area cleared of trees.
2 These results support the use of mating disruption to eradicate isolated gypsy moth populations in open landscapes, such as parks, residential areas and commercial settings.
3 Mating success in both treated and untreated areas varied with the initial distance between males and females. When the initial distance between males and females was < 5 cm in an area receiving a dosage of 37.5 g of racemic disparlure per ha, mating success was reduced by 27% compared with a similar deployment in an untreated area. Mating was eliminated in areas treated at the same dosage when males and females were initially deployed 1 m apart but on separate trees.
4 This suggests that mating disruption may not be an effective tactic for gypsy moth eradication in cases where the infestation is concentrated on a small number of trees and males and females are in close proximity in space and time.