Class III beta-tubulin in human development and cancer

The differential cellular expression of class III beta-tubulin isotype (betaIII) is reviewed in the context of human embryological development and neoplasia. As compared to somatic organs and tissues, betaIII is abundant in the central and peripheral nervous systems (CNS and PNS) where it is prominently expressed during fetal and postnatal development. As exemplified in cerebellar and sympathoadrenal neurogenesis, the distribution of betaIII is neuron-associated, exhibiting distinct temporospatial gradients according to the regional neuroepithelia of origin. However, transient expression of this protein is also present in the subventricular zones of the CNS comprising putative neuronal- and/or glial precursor cells, as well as in Kulchitsky neuroendocrine cells of the fetal respiratory epithelium. This temporally restricted, potentially non-neuronal expression may have implications in the identification of presumptive neurons derived from embryonic stem cells. In adult tissues, the distribution of betaIII is almost exclusively neuron-specific. Altered patterns of expression are noted in cancer. In "embryonal"- and "adult-type" neuronal tumors of the CNS and PNS, betaIII is associated with neuronal differentiation and decreased cell proliferation. In contrast, the presence of betaIII in gliomas and lung cancer is associated with an ascending histological grade of malignancy. Thus, betaIII expression in neuronal tumors is differentiation-dependent, while in non-neuronal tumors it is aberrant and/or represents "dedifferentiation" associated with the acquisition of progenitor-like phenotypic properties. Increased expression in various epithelial cancer cell lines is associated with chemoresistance to taxanes. Because betaIII is present in subpopulations of neoplastic, but not in normal differentiated glial or somatic epithelial cells, the elucidation of mechanisms responsible for the altered expression of this isotype may provide insights into the role of the microtubule cytoskeleton in tumorigenesis and tumor progression.     

Extrusion/spheronization of pectin-based formulations. I. Screening of important factors

This study investigated the possibility of producing pectin-based pellets by extrusion/spheronization. The study also identified factors influencing the process and the characteristics of the resulting product. Three types of pectin with different degrees of amid and methoxyl substitution were studied in combination with different granulation liquids (water, calcium chloride, citric acid, and ethanol) and/or microcrystalline cellulose. Pellets were prepared in a power-consumption-controlled, twinscrew extruder; then they were spheronized and dried. The products were characterized by image analysis, sieving analysis, and disintegration and dissolution tests. The results were evaluated by multivariate analysis. Different additives, either in the granulation liquid or in the powder mixture, influenced the ability of the extruded mass to form pellets (the processability) with this technique. However, the various pectin types responded to modifications to a different extent. Short, nearly spherical pellets are obtained with granulation liquids, such as ethanol, that reduce the swelling ability of pectin. Pellets produced with ethanol are, however, mechanically weak and tend to ditintegrate. Pectin molecules with a high degree of free carboxylic acid groups seem to be more sensitive to changes in the granulation liquid. Addition of microcrystalline cellulose as an extrusion aid generally resulted in improvements in shape and size. It was demonstrated that the processability of pectin as well as the characteristics of the products can be influenced in different ways during the process (eg, adding substances to the granulation liquid or to the powder mixture).     

Extrusion/spheronization of pectin-based formulations. II. Effect of additive concentration in the granulation liquid

Purpose. The aim of this study was to improve the formation of spherical pectin pellets by investigating the effect of additive concentration in the granulation liquid on the shape and size of the products as well as by identifying an optimal additive concentration.Methods. High-methoxylated, low-methoxylated, and amidated low-methoxylated pectin types were evaluated in combination with different concentrations of methanol, ethanol, citric acid, lactic acid, and calcium chloride. Pellets were prepared in a power-consumption-controlled twin-screw extruder, then spheronized and dried. The moisture content of the extrudate was determined, and the final products were characterized by image analysis and sieving analysis. A cloud point test was employed for the identification of an optimal additive concentration.Results. The concentration of additive in the granulation liquid affected the moisture content of the extrudate and the shape, size, and mechanical stability of the pectin pellets. Improvements in the pellet characteristics are dependent on the pectin type employed. The 2 low-methoxylated pectins were more sensitive to concentration changes than was the high-methoxylated type. Above a certain threshold concentration, the quality of the pellets are improved. This additive concentration differs according to type of pectin and type of additive.Conclusion. It was demonstrated that there is a concentration-dependent interaction between pectin and substances added to the granulation liquid that can be utilized to improve the formation of spherical pectin pellets.     

Copper,zinc, and selenium in human blood and urine after injection of sodium 2,3-dimercaptopropane-1-sulfonate: a study on subjects with dental amalgam

This study investigated the effects of a single dose of intravenously administered sodium 2,3-dimercaptopropane-1-sulfonate (DMPS) on the essential elements copper, zinc, and selenium in human blood and urine. The possible role of dental amalgam was also addressed. Eighty individuals, divided in four groups according to the presence or absence of dental amalgam fillings and symptoms self-related to such fillings, were given DMPS (2 mg/kg body wt) and 500 mL Ringer’s acetate intravenously. Urine and blood were collected prior to the injection, and thereafter at intervals over a 24-h period. Cu, Zn, and Se concentrations were determined by atomic absorption spectrometry methods. A statistically significant increase in the concentrations of Cu and Zn in urine was observed 30 and 120 min after the DMPS injection compared to the preinjection concentrations. The concentrations of Se were not affected. The cumulated excretion over 24 h after DMPS injection constitutes only from 0.1% to 0.7% of the body content of these elements. There was no effect of different amalgam statuses on Cu and Zn excretion. We found a temporary decrease (4–7%) in the concentrations of Cu, Zn, and Se in blood 15 and 30 min after DMPS, but this seems to be the result of dilution factors. Administration of a single dose of DMPS does not affect the body stores of the essential elements Cu, Zn, and Se.     

Induction and Assessment of Ischemia-reperfusion Injury in Langendorff-perfused Rat Hearts

The biochemical events surrounding ischemia reperfusion injury in the acute setting are of great importance to furthering novel treatment options for myocardial infarction and cardiac complications of thoracic surgery. The ability of certain drugs to precondition the myocardium against ischemia reperfusion injury has led to multiple clinical trials, with little success. The isolated heart model allows acute observation of the functional effects of ischemia reperfusion injury in real time, including the effects of various pharmacological interventions administered at any time-point before or within the ischemia-reperfusion injury window. Since brief periods of ischemia can precondition the heart against ischemic injury, in situ aortic cannulation is performed to allow for functional assessment of non-preconditioned myocardium. A saline filled balloon is placed into the left ventricle to allow for real-time measurement of pressure generation. Ischemic injury is simulated by the cessation of perfusion buffer flow, followed by reperfusion. The duration of both ischemia and reperfusion can be modulated to examine biochemical events at any given time-point. Although the Langendorff isolated heart model does not allow for the consideration of systemic events affecting ischemia and reperfusion, it is an excellent model for the examination of acute functional and biochemical events within the window of ischemia reperfusion injury as well as the effect of pharmacological intervention on cardiac pre- and postconditioning. The goal of this protocol is to demonstrate how to perform in situ aortic cannulation and heart excision followed by ischemia/reperfusion injury in the Langendorff model.     

Investigating the effects of topography and clonality on genetic structuring within a large Norwegian population of Arabidopsis lyrata

Background and Aims

The gene flow through pollen or seeds governs the extent of spatial genetic structure in plant populations. Another factor that can contribute to this pattern is clonal growth. The perennial species Arabidopsis lyrata ssp. petraea (Brassicaceae) is a self-incompatible, clonal species found in disjunctive populations in central and northern Europe.

Methods

Fourteen microsatellite markers were employed to study the level of kinship and clonality in a high-altitude mountain valley at Spiterstulen, Norway. The population has a continuous distribution along the banks of the River Visa for about 1·5 km. A total of 17 (10 m × 10 m) squares were laid out in a north–south transect following the river on both sides.

Key Results

It is shown that clonal growth is far more common than previously shown in this species, although the overall size of the genets is small (mean diameter = 6·4 cm). Across the whole population there is no indication of isolation by distance, and spatial genetic structure is only visible on fine spatial scales. In addition, no effect of the river on the spatial distribution of genotypes was found.

Conclusions

Unexpectedly, the data show that populations of small perennials like A. lyrata can behave like panmictic units across relatively large areas at local sites, as opposed to earlier findings in central Europe.     

The Bacterial Peptide Pheromone Plantaricin A Permeabilizes Cancerous, but not Normal, Rat Pituitary Cells and Differentiates between the Outer and Inner Membrane Leaflet

Plantaricin A (PlnA) is a 26-mer peptide pheromone with membrane-permeabilizing, strain-specific antibacterial activity, produced by Lactobacillus plantarum C11. We investigated the membrane-permeabilizing effects of PlnA on cultured cancerous and normal rat anterior pituitary cells using patch-clamp techniques and microfluorometry (fura-2). Cancerous cells displayed massive permeabilization within 5 s after exposure to 10–100 μm PlnA. The membrane depolarized to nearly 0 mV, and the membrane resistance decreased to a mere fraction of the initial value after less than 1 min. In outside-out membrane patches, 10 μm PlnA induced membrane currents reversing at 0 mV, which is compatible with an unspecific conductance increase. The d and l forms of the peptide had similar potency, indicating a nonchiral mechanism for the membrane-permeabilizing effect. Surprisingly, inside-out patches were insensitive to 1 mm PlnA. Primary cultures of normal rat anterior pituitary cells were also insensitive to the peptide. Thus, PlnA differentiates between plasma membranes and membrane leaflets. Microfluorometric recordings of [Ca²⁺] _i and cytosolic concentration of fluorochrome verified the rapid permeabilizing effect of PlnA on cancerous cells and the insensitivity of normal pituitary cells.     

Within-population spatial variation in pollinator visitation rates, pollen limitation on seed set, and flower longevity in an alpine species

Pollen limitation through insufficient pollen deposition on stigmas caused by too infrequent pollinator visitation may influence the reproductive outcome of plants. In this study we investigated how pollinator visitation rate, the degree of pollen limitation, and flower longevity varied spatially among three sites at different altitudes within a population of the dwarf shrub Dryas octopetala L. in alpine southern Norway. Significant pollen limitation on seed set only occurred at the mid-elevation site, while seed set at the other sites appeared to be mainly resource limited, thus indicating a spatial variation in pollen limitation. There was no association between the spatial variation in the extent of pollen limitation and pollinator visitation rate to flowers. However, pollinator visitation rates were related to flower longevity of Dryas; sites with low visitation rates had long-lived flowers and vice versa. Thus, our results suggest within-population spatial co-variation between pollinator visitation rates, pollen limitation, and a developmental response to these factors, flower longevity.     

Nuclear Receptor Liver X Receptor Is O-GlcNAc-modified in Response to Glucose

Post-translational modification of nucleocytoplasmic proteins by O-linked β-N-acetylglucosamine (O-GlcNAc) has for the last 25 years emerged as an essential glucose-sensing mechanism. The liver X receptors (LXRs) function as nutritional sensors for cholesterol-regulating lipid metabolism, glucose homeostasis, and inflammation. LXRs are shown to be post-translationally modified by phosphorylation, acetylation, and sumoylation, affecting their target gene specificity, stability, and transactivating and transrepressional activity, respectively. In the present study, we show for the first time that LXRα and LXRβ are targets for glucose-hexosamine-derived O-GlcNAc modification in human Huh7 cells. Furthermore, we observed increased hepatic LXRα O-GlcNAcylation in vivo in refed mice and in streptozotocin-induced refed diabetic mice. Importantly, induction of LXRα O-GlcNAcylation in both mouse models was concomitant with increased expression of the lipogenic gene SREBP-1c (sterol regulatory element-binding protein 1c). Furthermore, glucose increased LXR/retinoic acid receptor-dependent activation of luciferase reporter activity driven by the mouse SREBP-1c promoter via the hexosamine biosynthetic pathway in Huh7 cells. Altogether, our results suggest that O-GlcNAcylation of LXR is a novel mechanism by which LXR acts as a glucose sensor affecting LXR-dependent gene expression, substantiating the crucial role of LXR as a nutritional sensor in lipid and glucose metabolism.     

Plant volatiles mediate attraction to host and non-host plant in apple fruit moth, Argyresthia conjugella

Plant volatiles mediate host finding in insect herbivores and lead to host fidelity and habitat‐specific mating, generating premating reproductive isolation and facilitating sympatric divergence. The apple fruit moth, Argyresthia conjugella Zeller (Lepidoptera: Argyresthiidae), is a particularly suitable species to study the cues and behavioural mechanisms leading to colonization of a new host: it recurrently oviposits on the non‐host plant, apple Malus domestica Borkh. (Rosaceae), where the larvae cannot complete their development. The larval host of the apple fruit moth (Lepidoptera, Argyresthiidae), is rowan Sorbus aucuparia L. (Rosaceae). Fruit setting in rowan, however, fluctuates strongly over large areas in Scandinavia. Every 2–4 years, when too few rowanberries are available for egg laying in forests, apple fruit moth females oviposit instead on apple in nearby orchards, but not on other fruits, such as pear or plum. This poses the question of which cues mediate attraction to rowan and apple, and how apple fruit moth discriminates rowan from apple. Chemical analysis and antennal recordings showed that 11 out of 15 rowan volatiles eliciting an antennal response in A. conjugella females co‐occur in rowan and apple headspace, in a different proportion. In the field, A. conjugella was attracted to several of these plant volatiles, especially to 2‐phenyl ethanol, methyl salicylate, and decanal. Addition of anethole to 2‐phenyl ethanol had a strong synergistic effect, the 1 : 1 blend is a powerful attractant for A. conjugella males and females. These results confirm that volatiles common to both plants may account for a host switch in A. conjugella from rowan to apple. Some of the most attractive compounds, including 2‐phenyl ethanol, anethole, and decanal, which have been found in several apple cultivars, were not present in the headspace of the apple cultivar, Aroma, which is also susceptible to attack by A. conjugella. This supports the idea that the odour signal from apple is suboptimal for attraction of A. conjugella, but is nonetheless sufficient for attraction, during times when rowan is not available for egg laying.