Hyperglycemia increases endothelial superoxide that impairs smooth muscle cell Na+-K+-ATPase activity

Nitric oxide (NO) plays an important role in the control of numerous vascular functions including basal Na+-K+-ATPase activity in arterial tissue. Hyperglycemia inhibits Na+-K+-ATPase activity in rabbit aorta, in part, through diminished bioactivity of NO. The precise mechanism(s) for such observations, however, are not yet clear. The purpose of this study was to examine the role of superoxide in modulating NO-mediated control of Na+-K+-ATPase in response to hyperglycemia. Rabbit aorta incubated with hyperglycemic glucose concentrations (44 mM) demonstrated a 50% reduction in Na+-K+-ATPase activity that was abrogated by superoxide dismutase. Hyperglycemia also produced a 50% increase in steady-state vascular superoxide measured by lucigenin-enhanced chemiluminescence that was closely associated with reduced Na+-K+-ATPase activity. Specifically, the hyperglycemia-induced increase in vascular superoxide was endothelium dependent, inhibited by L-arginine, and stimulated by N(omega)-nitro-L-arginine. Aldose reductase inhibition with zopolrestat also inhibited the hyperglycemia-induced increase in vascular superoxide. In each manipulation of vascular superoxide, a reciprocal change in Na+-K+-ATPase activity was observed. Finally, a commercially available preparation of Na+-K+-ATPase was inhibited by pyrogallol, a superoxide generator. These data suggest that hyperglycemia induces an increase in endothelial superoxide that inhibits the stimulatory effect of NO on vascular Na+-K+-ATPase activity.     

Efficiency of embryoid body formation and hematopoietic development from embryonic stem cells in different culture systems

Embryonic stem (ES) cells have tremendous potential as a cell source for cell-based therapies. Realization of that potential will depend on our ability to understand and manipulate the factors that influence cell fate decisions and to develop scalable methods of cell production. We compared four standard ES cell differentiation culture systems by measuring aspects of embryoid body (EB) formation efficiency and cell proliferation, and by tracking development of a specific differentiated tissue type-blood-using functional (colony-forming cell) and phenotypic (Flk-1 and CD34 expression) assays. We report that individual murine ES cells form EBs with an efficiency of 42 +/- 9%, but this value is rarely obtained because of EB aggregation-a process whereby two or more individual ES cells or EBs fuse to form a single, larger cell aggregate. Regardless of whether EBs were generated from a single ES cell in methylcellulose or liquid suspension culture, or aggregates of ES cells in hanging drop culture, they grew to a similar maximum cell number of 28,000 +/- 9,000 cells per EB. Among the three methods for EB generation in suspension culture there were no differences in the kinetics or frequency of hematopoietic development. Thus, initiating EBs with a single ES cell and preventing EB aggregation should allow for maximum yield of differentiated cells in the EB system. EB differentiation cultures were also compared to attached differentiation culture using the same outputs. Attached colonies were not similarly limited in cell number; however, hematopoietic development in attached culture was impaired. The percentage of early Flk-1 and CD34 expressing cells was dramatically lower than in EBs cultured in suspension, whereas hematopoietic colony formation was almost completely inhibited. These results provide a foundation for development of efficient, scalable bioprocesses for ES cell differentiation, and inform novel methods for the production of hematopoietic tissues.     

Inter-hemispheric comparison of bottom-up effects on community structure: Insights revealed using the comparative-experimental approach

The comparative-experimental approach uses identically designed, replicated experiments at different sites along environmental gradients in order to gain insight into the changing dynamics of communities with changing environmental conditions. Such studies reveal how ecological processes vary in intensity and interact to produce community structure. Early emphases were on the community consequences of shifting top-down impacts, competition and disturbance with environmental stress. Recent advances include the more precise quantification of gradients and thus a better understanding of species responses to the environment, and the revelation that bottom-up forces can vary significantly on within-region scales, with major consequences for the impact of top-down forces and thus community dynamics. Here the use of the method to examine the role of geographic location (coastal ecosystems in different hemispheres) and oceanographic conditions (upwelling vs downwelling) on these bottom-up/top-down linkages is advanced. We show that a bottom-up factor (prey recruitment) and a top-down effect (predation rate) vary consistently with oceanographic conditions within each coastal ecosystem, and also between geographic locations (New Zealand, Oregon). In general, both recruitment and predation rates are higher in Oregon. It is postulated that these differences are common responses to oceanographic variation, and that between-hemisphere differences result from the stronger and more persistent upwelling in the California Current ecosystem.     

"I speak a different dialect": teen explanatory models of difference and disability

What do teens with disabilities believe about their conditions, and what do they understand to be the causes, correlates, and consequences of disability? We elicited a cultural explanatory model (EM) of disability from a longitudinal sample of 23 European American adolescents with varied cognitive disabilities and delay. We asked teens how they were similar to or different from others; the name of this difference; its causes, severity, course, effects, associated problems and benefits; and need for treatment. IQ and type of disability strongly affected quality of responses only from the lowest functioning teens. A majority of teens had a reasonably rich and coherent EM, blending typical and disability themes of cultural knowledge and identity. The EM is a window into social context (schools, services, parents, and peers) as well as personal experience. Eliciting explanatory models from teens with disabilities is not only possible but also can enhance understanding of identity, family influence, and appropriate services.     

Protein-RNA interactions: a structural analysis

A detailed computational analysis of 32 protein–RNA complexes is presented. A number of physical and chemical properties of the intermolecular interfaces are calculated and compared with those observed in protein–double-stranded DNA and protein–single-stranded DNA complexes. The interface properties of the protein–RNA complexes reveal the diverse nature of the binding sites. van der Waals contacts played a more prevalent role than hydrogen bond contacts, and preferential binding to guanine and uracil was observed. The positively charged residue, arginine, and the single aromatic residues, phenylalanine and tyrosine, all played key roles in the RNA binding sites. A comparison between protein–RNA and protein–DNA complexes showed that whilst base and backbone contacts (both hydrogen bonding and van der Waals) were observed with equal frequency in the protein–RNA complexes, backbone contacts were more dominant in the protein–DNA complexes. Although similar modes of secondary structure interactions have been observed in RNA and DNA binding proteins, the current analysis emphasises the differences that exist between the two types of nucleic acid binding protein at the atomic contact level.     

A small outbreak of listeriosis potentially linked to the consumption of imitation crab meat

A small outbreak of listeriosis involving two previously healthy adults occurred in Ontario. Food samples obtained from the refrigerator of the patients included imitation crab meat, canned black olives, macaroni and vegetable salad, spaghetti sauce with meatballs, mayonnaise and water. All of the samples except the water contained Listeria monocytogenes. The three most heavily contaminated samples were the imitation crab meat, the olives and the salad which contained 2.1 x 109, 1.1 x 107 and 1.3 x 106 cfu g-1, respectively. L. monocytogenes serotype 1/2b was isolated from the patients, as well as from the opened and unopened imitation crab meat. Molecular typing of the isolates by both randomly amplified polymorphic DNA (RAPD) and pulsed-field gel electrophoresis (PFGE) typing demonstrated the imitation crab meat and clinical strains to be indistinguishable. Challenge studies performed with a pool of L. monocytogenes strains showed that imitation crab meat, but not olives, supported growth of the organism. In this study we have shown for the first time the potential involvement of imitation crab meat in a small outbreak of listeriosis. In terms of disease prevention, temperature control is critical to prevent or reduce the growth of this foodborne pathogen. In addition, with refrigerated products having a long (> 30 d) shelf life, additional safety factors must be used to prevent the growth of foodborne pathogens such as L. monocytogenes.     

Shallow genetic and morphological divergence among seaperches in the South Pacific (family Scorpaenidae; genus Helicolenus)

The phylogenetic relationships among populations of seaperch, Helicolenus spp., in the south-west Pacific were examined with mtDNA markers. Parts of the cytochrome b gene [459 base pair (bp)] and the control region (448 bp) were sequenced in 58 specimens from the south-west Pacific and four specimens of Helicolenus lengerichi from Chile. Only one clade was recognized in New Zealand coastal waters, despite a wide range of colour morphs. This clade also occurred in the mid Tasman Sea on the Norfolk Ridge and around Tasmania and Victoria. A second sympatric clade was identified around Tasmania and Victoria and to the west of New Zealand. A third allopatric clade was identified to the north of New Zealand and in deep water on the Chatham Rise and a fourth clade on the Foundation Seamounts and the Louisville Ridge. Helicolenus lengerichi from Chile formed a fifth clade. Assuming a molecular clock, the clades were estimated to have diverged c. 0·7–2·6 million years ago. Only two clades, around Tasmania and Victoria, were separated using morphology, colour (in live) and dorsal-fin soft ray counts and were confirmed as Helicolenus percoides and Helicolenus barathri . Two characters, orbit diameter and colour variation, previously used to identify two species in New Zealand waters were unreliable characters for species discrimination. Principle component analyses of 11 morphological measures from 67 individuals did not delineate the clades. A canonical discriminant analysis was able to separate four of the five clades, but mean discriminate probabilities were low (77·6%), except for the five Chilean specimens of H. lengerichi (100%).     

Sonic Hedgehog Controls the Phenotypic Fate and Therapeutic Efficacy of Grafted Neural Precursor Cells in a Model of Nigrostriatal Neurodegeneration

The expression of soluble growth and survival promoting factors by neural precursor cells (NPCs) is suggested to be a prominent mechanism underlying the protective and regenerative effects of these cells after transplantation. Nevertheless, how and to what extent specific NPC-expressed factors contribute to therapeutic effects is not well understood. Using RNA silencing, the current study investigated the roles of two donor NPC molecules, namely glial cell-line derived neurotrophic factor (GDNF) and sonic hedgehog (SHH), in the protection of substantia nigra dopamine neurons in rats treated with 6-hydroxydopamine (6-OHDA). Analyses indicate that as opposed to the knock-down of GDNF, SHH inhibition caused a profound decline in nigrostriatal neuroprotection. Further, SHH silencing also curbed endogenous neurogenesis and the migration of host brdU⁺/dcx⁺ neural precursors into the striatum, which was present in the animals receiving control or GDNF silenced NPCs. A change in graft phenotype, mainly reflected by a reduced proportion of undifferentiated nestin⁺ cells, as well as a significantly greater host microglial activity, suggested an important role for these processes in the attenuation of neuroprotection and neurogenesis upon SHH silencing. Overall these studies reveal core mechanisms fundamental to grafted NPC-based therapeutic effects, and delineate the particular contributions of two graft-expressed molecules, SHH and GDNF, in mediating midbrain dopamine neuron protection, and host plasticity after NPC transplantation.