Seedling responses of three Australian tree species to toxic concentrations of zinc in solution culture

A frequently desired outcome when rehabilitating Zn toxic sites in Australia is to establish a self-sustaining native ecosystem. Hence, it is important to understand the tolerance of Australian native plants to high concentrations of Zn. Very little is known about the responses of Australian native plants, and trees in particular, to toxic concentrations of Zn. Acacia holosericea, Eucalyptus camaldulensis and Melaleuca leucadendra plants were grown in dilute solution culture for 10 weeks. The seedlings (42 days old) were exposed to six Zn treatments viz., 0.5, 5, 10, 25, 50 and 100 muM. The order of tolerance to toxic concentrations of Zn was E. camaldulensis > A. holosericea > M. leucadendra, the critical external concentrations being approximately 20, 12 and 1.5 muM, respectively. Tissue Zn concentrations increased as solution Zn increased for all species. Root tissue concentrations were higher than shoot tissue concentrations at all solution Zn concentrations. The critical tissue Zn concentrations were approximately 85 and 110 mug g(-1) DM for M. leucadendra, 115 and 155 mug g(-1) DM for A. holosericea and 415 and 370 mug g(-1) DM for E. camaldulensis for the youngest fully expanded leaf and total shoots, respectively. The results from this paper provide the first comprehensive combination of growth responses, critical external concentrations, critical tissue concentrations and plant toxicity symptoms for three important Australian genera, viz., Eucalyptus, Acacia and Melaleuca, for use in the rehabilitation of potentially Zn toxic sites.

Microtubule integrity is essential for apical polarization and epithelial morphogenesis in the thyroid

In this study, we examined the contribution of microtubules to epithelial morphogenesis in primary thyroid cell cultures. Thyroid follicles consist of a single layer of polarized epithelial cells surrounding a closed compartment, the follicular lumen. Freshly isolated porcine thyroid cells aggregate and reorganize to form follicles when grown in primary cultures. Follicular reorganization is principally a morphogenetic process that entails the assembly of biochemically distinct apical and basolateral membrane domains, delimited by tight junctions. The establishment of cell surface polarity during folliculogenesis coincided with the polarized redistribution of microtubules, predominantly in the developing apical poles of cells. Disruption of microtubule integrity using either colchicine or nocodazole caused loss of defined apical membrane domains, tight junctions and follicular lumina. Apical membrane and tight junction markers became randomly distributed at the outer surfaces of aggregates. In contrast, the basolateral surface markers, E-cadherin and Na+,K+-ATPase, remained correctly localized at sites of cell-cell contact and at the free surfaces of cell aggregates. These findings demonstrate that microtubules play a necessary role in thyroid epithelial morphogenesis. Specifically, microtubules are essential to preserve the correct localization of apical membrane components within enclosed cellular aggregates, a situation that is also likely to pertain where lumina must be formed from solid aggregates of epithelial precursors. (C) 2001 Wiley-Liss, Inc.

On the importance function in splitting simulation

The splitting method is a simulation technique for the estimation of very small probabilities. In this technique, the sample paths are split into multiple copies, at various stages in the simulation. Of vital importance to the efficiency of the method is the Importance Function (IF). This function governs the placement of the thresholds or surfaces at which the paths are split. We derive a characterisation of the optimal IF and show that for multi-dimensional models the natural choice for the IF is usually not optimal. We also show how nearly optimal splitting surfaces can be derived or simulated using reverse time analysis. Our numerical experiments illustrate that by using the optimal IF, one can obtain a significant improvement in simulation efficiency.

Preliminary characterisation and extraction of anterior adhesive secretion in monogenean (platyhelminth) parasites

Secreted anterior adhesives, used for temporary attachment to epithelial surfaces of fishes (skin and gills) by some monogenean (platyhelminth) parasites have been partially characterised. Adhesive is composed of protein. Amino acid composition has been determined for seven monopisthocotylean monogeneans. Six of these belong to the Monocotylidae and one species, Entobdella soleae (van Beneden et Hesse, 1864) Johnston, 1929, is a member of the Capsalidae. Histochemistry shows that the adhesive does not contain polysaccharides, including acid mucins, or lipids. The adhesive before secretion and in its secreted form contains no dihydroxyphenylalanine (dopa). Secreted adhesive is highly insoluble, but has a soft consistency and is mechanically removable from glass surfaces. Generally there are high levels of glycine and alanine, low levels of tyrosine and methionine, and histidine is often absent. However, amino acid content varies between species, the biggest differences evident when the monocotylid monogeneans were compared with E. soleae. Monogenean adhesive shows similarity in amino acid profile with adhesives from starfish, limpets and barnacles. However, there are some differences in individual amino acids in the temporary adhesive secretions of, on the one hand, the monogeneans and, on the other hand, the starfish and limpets. These differences may reflect the fact that monogeneans, unlike starfish and barnacles, attach to living tissue (tissue adhesion). A method of extracting unsecreted adhesive was investigated for use in further characterisation studies on monogenean glues.

Empirical evidence for ultrametric structure in multi-layer perceptron error surfaces

Combinatorial optimization problems share an interesting property with spin glass systems in that their state spaces can exhibit ultrametric structure. We use sampling methods to analyse the error surfaces of feedforward multi-layer perceptron neural networks learning encoder problems. The third order statistics of these points of attraction are examined and found to be arranged in a highly ultrametric way. This is a unique result for a finite, continuous parameter space. The implications of this result are discussed.

Avian paramyxoviruses and influenza viruses isolated from mallard ducks (Anas platyrhynchos) in New Zealand

A comprehensive study using virological and serological approaches was carried out to determine the status of live healthy mallard ducks (Anas platyrhynchos) in New Zealand for infections with avian paramyxoviruses (APMV) and influenza viruses (AIV). Thirty-three viruses isolated from 321 tracheal and cloacal swabs were characterized as: 6 AIV (two H5N2 and four H4N6), 10 APMV-1 and 17 APMV-4. Of 335 sera samples tested for AIV antibodies, 109 (32.5%) sera were positive by nucleoprotein-blocking ELISA (NP-B-ELISA). Serum samples (315) were examined for antibody to APMV-1, -2, -3, -4, -6, -7, -8, -9 by the haemagglutination inhibition test. The largest number of reactions, with titres up to greater than or equal to 1/64, was to APMV-1 (93.1%), followed by APMV-6 (85.1%), APMV-8 (56%), APMV-4 (51.7%), APMV-7 (47%), APMV-9 (15.9%), APMV-2 (13.3%) and APMV-3 (6.0%). All of the H5N2 isolates of AIV and the APMV-1 isolates from this and earlier New Zealand studies had low pathogenicity indices assessed by the Intravenous Pathogenicity Index (IVPI) with the result 0.00 and Intracerebral Pathogenicity Index (ICPI) with results 0.00-0.16. Partial genomic and antigenic analyses were also consistent with the isolates being non-pathogenic. Phylogenetic analysis of the 10 APMV-1 isolates showed 9 to be most similar to the reference APMV-1 strain D26/76 originally isolated in Japan and also to the Que/66 strain, which was isolated in Australia. The other isolate was very similar to a virus (MC 110/77) obtained from a shelduck in France.

New W-isotope evidence for rapid terrestrial accretion and very early core formation

The short-lived Hf-182-W-182-isotope system is an ideal clock to trace core formation and accretion processes of planets. Planetary accretion and metal/silicate fractionation chronologies are calculated relative to the chondritic Hf-182-W-182-isotope evolution. Here, we report new high-precision W-isotope data for the carbonaceous chondrite Allende that are much less radiogenic than previously reported and are in good agreement with published internal Hf-W chronometry of enstatite chondrites. If the W-isotope composition of terrestrial rocks, representing the bulk silicate Earth, is homogeneous and 2.24 epsilon(182W) units more radiogenic than that of the bulk Earth, metal/silicate differentiation of the Earth occurred very early. The new W-isotope data constrain the mean time of terrestrial core formation to 34 million years after the start of solar system accretion. Early terrestrial core formation implies rapid terrestrial accretion, thus permitting formation of the Moon by giant impact while Hf-182 was still alive. This could explain why lunar W-isotopes are more radiogenic than the terrestrial value. Copyright (C) 2002 Elsevier Science Ltd.

Ephrin-A5 induces rounding, blebbing and deadhesion of EphA3-expressing 293T and melanoma cells by CrkII and Rho-mediated signalling

Eph receptor tyrosine kinases and ephrins regulate morphogenesis in the developing embryo where they effect adhesion and motility of interacting cells. Although scarcely expressed in adult tissues, Eph receptors and ephrins are overexpressed in a range of tumours. In malignant melanoma, increased Eph and ephrin expression levels correlate with metastatic progression. We have examined cellular and biochemical responses of EphA3-expressing melanoma cell lines and human epithelial kidney 293T cells to stimulation with polymeric ephrin-A5 in solution and with surfaces of defined ephrin-A5 densities. Within minutes, rapid reorganisation of the actin and myosin cytoskeleton occurs through activation of RhoA, leading to the retraction of cellular protrusions, membrane blebbing and detachment, but not apoptosis. These responses are inhibited by monomeric ephrin-A5, showing that receptor clustering is required for this EphA3 response. Furthermore, the adapter CrkII, which associates with tyrosine-phosphorylated EphA3 in vitro, is recruited in vivo to ephrin-A5-stimulated EphA3. Expression of an SH3-domain mutated CrkII ablates cell rounding, blebbing and detachment. Our results suggest that recruitment of CrkII and activation of Rho signalling are responsible for EphA3-mediated cell rounding, blebbing and de-adhesion, and that ephrin-A5-mediated receptor clustering and EphA3 tyrosine kinase activity are essential for this response.

Endodontic sequelae of dental erosion

Background. The incidence of, pulp involvement in patients with excessive wear has not been extensively documented. Methods: Clinical, records of 448 patients with excessive tooth wear were reviewed and 52 cases (11.6 per cent) with near or frank pulp exposures or root canal treatments were found and their numbers and sites were tabulated. Light microscopy of study models was used to determine aetiology at each site of exposure as. attrition, erosion or abrasion, scanning electron microscopy (SEM) was performed on some individual teeth. Results: Forty sites of near exposure and 57 sites of frank exposures or root canal treatments were found, some cases had both types of exposure. The commonest sites exposed by erosion were the palatal surfaces of maxillary, and the incisal surfaces of mandibular anterior teeth. Posterior teeth were not commonly affected. Toothbrush abrasion had exacerbated softie lesions as shown by SEM. Conclusions: Endodontic sequelae were found in 11 per cent of tooth wear patients as late stages of dental erosion. Near and frank exposures of the pulp thus constitute a small but significant, problem for,the Australian dental profession's concern in the of the tooth wear cases.

In vivo and in vitro models demonstrate a role for caveolin-1 in the pathogenesis of ischaemic acute renal failure

Caveolae and their proteins, the caveolins, transport macromolecules; compartmentalize signalling molecules; and are involved in various repair processes. There is little information regarding their role in the pathogenesis of significant renal syndromes such as acute renal failure (ARF). In this study, an in vivo rat model of 30 min bilateral renal ischaemia followed by reperfusion times from 4 h to 1 week was used to map the temporal and spatial association between caveolin-1 and tubular epithelial damage (desquamation, apoptosis, necrosis). An in vitro model of ischaemic ARF was also studied, where cultured renal tubular epithelial cells or arterial endothelial cells were subjected to injury initiators modelled on ischaemia-reperfusion (hypoxia, serum deprivation, free radical damage or hypoxia-hyperoxia). Expression of caveolin proteins was investigated using immunohistochemistry, immunoelectron microscopy, and immunoblots of whole cell, membrane or cytosol protein extracts. In vivo, healthy kidney had abundant caveolin-1 in vascular endothelial cells and also some expression in membrane surfaces of distal tubular epithelium. In the kidneys of ARF animals, punctate cytoplasmic localization of caveolin-1 was identified, with high intensity expression in injured proximal tubules that were losing basement membrane adhesion or were apoptotic, 24 h to 4 days after ischaemia-reperfusion. Western immunoblots indicated a marked increase in caveolin-1 expression in the cortex where some proximal tubular injury was located. In vitro, the main treatment-induced change in both cell types was translocation of caveolin-1 from the original plasma membrane site into membrane-associated sites in the cytoplasm. Overall, expression levels did not alter for whole cell extracts and the protein remained membrane-bound, as indicated by cell fractionation analyses. Caveolin-1 was also found to localize intensely within apoptotic cells. The results are indicative of a role for caveolin-1 in ARF-induced renal injury. Whether it functions for cell repair or death remains to be elucidated. Copyright (C) 2003 John Wiley Sons, Ltd.

James W. Gerard: His image of imperial Germany, 1913-1918

The subject of this study was a typical, if in some respects well qualified, U.S. ambassadorial appointee for his time, the early twentieth century: an attorney, judge, and politician who served competently in his one diplomatic assignment, in Berlin, before returning to private life.—Ed.

Structure of the HERG K+ channel S5P extracellular linker - Role of an amphipathic alpha-helix in C-type inactivation

The HERG K+ channel has very unusual kinetic behavior that includes slow activation but rapid inactivation. These features are critical for normal cardiac repolarization as well as in preventing lethal ventricular arrhythmias. Mutagenesis studies have shown that the extracellular peptide linker joining the fifth transmembrane domain to the pore helix is critical for rapid inactivation of the HERG K+ channel. This peptide linker is also considerably longer in HERG K+ channels, 40 amino acids, than in most other voltage-gated K+ channels. In this study we show that a synthetic 42-residue peptide corresponding to this linker region of the HERG K+ channel does not have defined structural elements in aqueous solution; however, it displays two well defined helical regions when in the presence of SDS micelles. The helices correspond to Trp(585)-Ile(593) and Gly(604)-Tyr(611) of the channel. The Trp(585)-Ile(593) helix has distinct hydrophilic and hydrophobic surfaces. The Gly(604)-Tyr(611) helix corresponds to an N-terminal extension of the pore helix. Electrophysiological studies of HERG currents following application of exogenous S5P peptides show that the amphipathic helix in the S5P linker interacts with the pore region of the channel in a voltage-dependent manner.

Enhanced effector and memory CTL responses generated by incorporation of receptor activator of NF-kappa B(RANK)/RANK ligand costimulatory molecules into dendritic cell immunogens expressing a human tumor-specific antigen

The outcome of dendritic cell (DC) presentation of Ag to T cells via the TCR/MHC synapse is determined by second signaling through CD80/86 and, importantly, by ligation of costimulatory ligands and receptors located at the DC and T cell surfaces. Downstream signaling triggered by costimulatory molecule ligation results in reciprocal DC and T cell activation and survival, which predisposes to enhanced T cell-mediated immune responses. In this study, we used adenoviral vectors to express a model tumor Ag (the E7 oncoprotein of human papillomavirus 16) with or without coexpression of receptor activator of NF-kappaB (RANK)/RANK ligand (RANKL) or CD40/CD40L costimulatory molecules, and used these transgenic DCs to immunize mice for the generation of E7-directed CD8(+) T cell responses. We show that coexpression of RANK/RANKL, but not CD40/CD40L, in E7-expressing DCs augmented E7-specific IFN-gamma-secreting effector and memory T cells and E7-specific CTLs. These responses were also augmented by coexpression of T cell costimulatory molecules (RANKL and CD40L) or DC costimulatory molecules (RANK and CD40) in the E7-expressing DC immunogens. Augmentation of CTL responses correlated with up-regulation of CD80 and CD86 expression in DCs transduced with costimulatory molecules, suggesting a mechanism for enhanced T cell activation/survival. These results have generic implications for improved tumor Ag-expressing DC vaccines, and specific implications for a DC-based vaccine approach for human papillomavirus 16-associated cervical carcinoma.