Beta-catenin defines head versus tail identity during planarian regeneration and homeostasis

After amputation, freshwater planarians properly regenerate a head or tail from the resulting anterior or posterior wound. The mechanisms that differentiate anterior from posterior and direct the replacement of the appropriate missing body parts are unknown. We found that in the planarian Schmidtea mediterranea, RNA interference (RNAi) of beta-catenin or dishevelled causes the inappropriate regeneration of a head instead of a tail at posterior amputations. Conversely, RNAi of the beta-catenin antagonist adenomatous polyposis coli results in the regeneration of a tail at anterior wounds. In addition, the silencing of beta-catenin is sufficient to transform the tail of uncut adult animals into a head. We suggest that beta-catenin functions as a molecular switch to specify and maintain anteroposterior identity during regeneration and homeostasis in planarians.     

SMEDWI-2 is a PIWI-like protein that regulates planarian stem cells

We have identified two genes, smedwi-1 and smedwi-2, expressed in the dividing adult stem cells (neoblasts) of the planarian Schmidtea mediterranea. Both genes encode proteins that belong to the Argonaute/PIWI protein family and that share highest homology with those proteins defined by Drosophila PIWI. RNA interference (RNAi) of smedwi-2 blocks regeneration, even though neoblasts are present, irradiation-sensitive, and capable of proliferating in response to wounding; smedwi-2(RNAi) neoblast progeny migrate to sites of cell turnover but, unlike normal cells, fail at replacing aged tissue. We suggest that SMEDWI-2 functions within dividing neoblasts to support the generation of cells that promote regeneration and homeostasis.     

Clonal production and organization of inhibitory interneurons in the neocortex

The neocortex contains excitatory neurons and inhibitory interneurons. Clones of neocortical excitatory neurons originating from the same progenitor cell are spatially organized and contribute to the formation of functional microcircuits. In contrast, relatively little is known about the production and organization of neocortical inhibitory interneurons. We found that neocortical inhibitory interneurons were produced as spatially organized clonal units in the developing ventral telencephalon. Furthermore, clonally related interneurons did not randomly disperse but formed spatially isolated clusters in the neocortex. Individual clonal clusters consisting of interneurons expressing the same or distinct neurochemical markers exhibited clear vertical or horizontal organization. These results suggest that the lineage relationship plays a pivotal role in the organization of inhibitory interneurons in the neocortex.     

Expression of 14-3-3 σ protein in normal and neoplastic canine mammary gland

14-3-3 σ protein is a negative cell cycle regulator, with both reduced and elevated levels associated with cancer in humans. This study assessed the expression of this protein in canine mammary tissues using immunohistochemistry and Western blotting. 14-3-3 σ was detected in 97% of the mammary tissue samples examined and was found in both myoepithelial (MECs) and epithelial (ECs) cells. Expression levels were elevated and reduced in neoplastic ECs and MECs, respectively (P < 0.001). Intense expression of 14-3-3 σ was detected in neoplastic ECs infiltrating blood vessels and lymph nodes and suggests a possible role for this protein in the malignant transformation of mammary neoplasms. Moreover, double immunostaining for 14-3-3 σ and the MEC – specific marker p63, confirmed that 14-3-3 σ is a highly sensitive marker of MECs since all p63 – positive cells were also positive for 14-3-3 σ. However, this protein is not exclusive to MECs as ECs also labelled positively.     

Predation of artificial nests in a fragmented landscape in the tropical region of Los Tuxtlas, Mexico

Predation rates of artificial nests were investigated in a fragmented landscape in the lowlands of Los Tuxtlas in southern Mexico. Hen and plasticine eggs were used to assess predation pressure in four habitats: the interior of forest fragments, the forest-pasture edge, corridors of residual forest vegetation and linear strips of live fences across pastures. Three sites per habitat were used in three experimental trials. Hen and plasticine ground nests with three eggs each were alternated every 50 m along transects at each site. Predation rates on each type of nest were monitored for 9 days. Survey of potential avian and mammalian potential nest predators were conducted at each site prior to the experimental trails. Readings of amount of light illuminating the ground were taken by each nest at each site to assess exposure of nests. In general, average predation rates were significantly higher for both hen and plasticine nests in the forest-pasture edge and in the corridors than in the interior of the forest fragments. While birds and mammals were the principal predators on hen eggs in the forests, mammals were responsible for the majority (?70%) of eggs damaged at the other habitats. Surveys of potential nest predators showed that avian and mammalian potential nest predators were significantly more common at the forest-pasture edges and at the other habitats than in the forests. Readings of light reaching the ground suggest that concealment of nests by the vegetation may play an important role in predation risk. Our results are consistent with reports from other Neotropical rainforests indicating an increase of artificial nest predation pressures from forest interior to open habitats. Restoration of forest fragments, allowing the vegetation to grow along the forest-pasture edge and the planting of arboreal crops at the forest-pasture edges may be measures that could increase cover and nest protection.     

Dall''s sheep responses to overflights by helicopter and fixed-wing aircraft

High rates of behavioural disruption caused by human activities could jeopardize the body condition and reproductive success of wildlife. I exposed Dall's sheep (Ovis dalli dalli) of the Yukon Territory to experimental overflights by a fixed-wing aircraft and a helicopter. Aircraft approaches that were more direct (as determined by the aircraft's elevation and horizontal distance from sheep) were more likely to elicit fleeing or to disrupt resting. Latency to resume feeding or resting after fixed-wing overflights was longer during more direct approaches. During indirect approaches by helicopters, sheep far from rocky slopes were much more likely to flee than sheep on rocky slopes. Sheep did not flee while nearby helicopters flew along the opposite side of a ridge, presumably because the obstructive cover buffered disturbing stimuli. Results provide preliminary parameters for predicting energetic and fitness costs incurred as a function of overflight rates, and can help mitigate disturbance by guiding temporal and spatial restrictions to aircraft.     

Countercurrent supercritical fluid extraction and fractionation of high-added-value compounds from a hexane extract of olive leaves

Countercurrent supercritical fluid extraction (CC-SFE) at a pilot scale plant was used for fractionation of high-added-value products from a raw extract of olive leaves in hexane. Compounds found in the raw extract were waxes, hydrocarbons, squalene, beta-carotene, triglycerides, alpha-tocopherol, beta-sitosterol, and alcohols. The CC-SFE extraction process was investigated according to a 2(3) full factorial experimental design using the following variables and ranges: extraction pressure, 75-200 bar; extraction temperature, 35-50 degrees C; and ethanol as modifier, 0-10%. Data were analyzed in terms of extraction yield, enrichment, recovery, and selectivity. Higher extraction yields were attained at 200 bar. For most of the compounds analyzed enrichment was attained at the same conditions, that is, 75 bar, 35 degrees C, and 10% ethanol. Hydrocarbons were usually recovered in the separators, whereas waxes and alpha-tocopherol remain in the raffinate. Selectivity data reveal that alpha-tocopherol is the most easily separable compound. The influence of the experimental factors on the recovery of all the compounds was studied by means of regression models. The best fitted model was attained for beta-sitosterol, with R2 = 99.25%.     

Ultrasensitive detection of genetically modified maize DNA by capillary gel electrophoresis with laser-induced fluorescence using different fluorescent intercalating dyes

In this work, four different fluorescent intercalating dyes are compared for the ultrasensitive CGE-LIF detection of DNA from transgenic maize in flours. The fluorescent intercalating dyes compared are YOPRO-1, SYBR-Green-I, Ethidium bromide (EthBr), and EnhanCE. For all the four dyes optimum concentrations are established, and efficient separations of DNA fragments ranging in size from 80 to 1000 bp are obtained. The comparative study demonstrates that SYBR-Green-I and YOPRO-1 provide better limits of detection (LODs) than EnhanCE or EthBr (i.e., LODs are, respectively, 700, 1000, 11300, and 97400 zmol, calculated for a 200-bp DNA fragment). Separations using YOPRO-1 are faster than those using SYBR-Green-I (30 min vs 47 min for the analysis of the 80-1000 bp DNA fragments). Also, separations using YOPRO-1 are more efficient than those using SYBR-Green-I (e.g., 2.4 x 10(6) plates/m vs 1.6 x 10(6) plates/m, respectively, calculated for the 200-bp fragment). Also, buffer depletion and cost per analysis are worse with SYBR-Green-I than with YOPRO-1. Therefore, YOPRO-1 was selected as the preferred intercalating dye. Using this fluorescent compound, analysis time reproducibility for the CGE-LIF separation of the DNA fragments is determined to be better than 1.7% (% RSD, n = 10) within the same day, and better than 1.9% (% RSD, n = 30) for three different days. Moreover, the fluorescence signal obtained using this dye is shown to vary linearly with the DNA concentration in the range studied, i.e., 1-500 ng/microL. It is demonstrated that by using this method 0.01% of transgenic maize can be detected in flour by direct injection of the PCR-amplified sample.     

Microscopic postmortem changes in adrenal glands of the domestic fowl

Eighty-four male white leghorn chickens were killed by CO2 gas to determine the type, rate, and sequence of microscopic postmortem changes in the adrenal glands of dry and wet intact carcasses. They were held at 29 or 18 C with 50% relative humidity for different times postmortem. The sequence of microscopic postmortem changes was similar in all chickens except at 18 C, when karyorrhexis of cortical and medullary cells was observed. Cellular changes occurred earlier at 29 C than at 18 C and in dry chickens but not in chickens wet with detergent solution before storage, although slight quantitative and qualitative differences between wet and dry chickens were noted. Medullary cells underwent postmortem changes earlier than cortical cells. Nuclei of medullary cells decreased in size, with chromatin clumping leading to pyknosis, followed by cytoplasmic vacuolation, cellular shrinkage, and finally karyolysis and cell dissociation. Cortical cells had nuclear chromatin marginated, nuclei reduced in size initially, and some nuclear fading, followed by pyknosis and karyolysis. Karyorrhexis was not a prominent feature of cortical and medullary cells, although it occasionally occurred before pyknosis. Cytoplasm of cortical cells remained eosinophilic, granular, and vacuolated, but vacuoles became finer later. Pyknotic medullary cells with vacuolated cytoplasm were observed as early as 3 hr postmortem, regardless of the temperature. Diffuse pyknosis of medullary cells was noted at 18 hr in chickens held at 29 C and at 48 hr in chickens held at 18 C. Marked cortical pyknosis was noted only at 36 hr in wet chickens held at 29 C, when bacterial invasion started. Dry chickens held 36 hr at 29 C had diffuse cellular dissociation, karyolysis and cytoplasmic acidophilia, and marked bacterial invasion. Erythrocytes were pyknotic and had cytoplasmolysis. It was concluded that adrenal glands may still be useful for histopathological examination before 18 hr at 29 C and before 48 hr at 18 C.