P-Glycoprotein is localized in intermediate-density membrane microdomains distinct from classical lipid rafts and caveolar domains

P-glycoprotein (Pgp), a member of the ATP-binding cassette (ABC) superfamily responsible for the ATP-driven extrusion of diverse hydrophobic molecules from cells, is a cause of multidrug resistance in human tumours. Pgp can also operate as a phospholipid and glycosphingolipid flippase, and has been functionally linked to cholesterol, suggesting that it might be associated with sphingolipid-cholesterol microdomains in cell membranes. We have used nonionic detergent extraction and density gradient centrifugation of extracts from the multidrug-resistant Chinese hamster ovary cell line, CH(R)B30, to address this question. Our data indicate that Pgp is localized in intermediate-density membrane microdomains different from classical lipid rafts enriched in Src-family kinases. We demonstrate that Brij-96 can selectively isolate the Pgp domains, separating them from the caveolar and classical lipid rafts. Pgp was found entirely in the Brij-96-insoluble domains, and only partially in the Triton X-100-insoluble membrane microdomains. We studied the sensitivity of these domains to cholesterol removal, as well as their relationship to GM(1) ganglioside- and caveolin-1-enriched caveolar domains. We found that the buoyant density of the Brij-96-based Pgp-containing microdomains was sensitive to cholesterol removal by methyl-beta-cyclodextrin. The Brij-96 domains retained their structural integrity after cholesterol depletion while, in contrast, the Triton X-100-based caveolin-1/GM(1) microdomains did not. Using confocal fluorescence microscopy, we determined that caveolin-1 and GM(1) colocalized, while Pgp and caveolin-1, or Pgp and GM(1), did not. Our results suggest that Pgp does not interact directly with caveolin-1, and is localized in intermediate-density domains, distinct from classical lipid rafts and caveolae, which can be isolated using Brij-96.     

Simulation of hurricane-like disturbances on a Caribbean seagrass bed

We tested the hypothesis that hurricanes cause changes in the plant community structure of Caribbean seagrass beds by acting selectively on populations of rooted macrophytes, which include seagrasses and rhizophytic algae. We also tested the hypothesis that susceptibility to elimination of the rooted macrophytes by the disturbance depends on differences in their growth forms. Two commonly registered disturbances by hurricanes in shallow seagrass beds are burial and sediment removal, which were simulated in marked plots of 1.1 × 1.1 m, at two randomly selected stations in Puerto Morelos tropical reef lagoon. The treatments consisted of control (no disturbance), two levels of burial, and two levels of sediment removal, with four replicates per treatment per station. The experiment was initiated in July 2002 and, 2 months afterwards, the densities of the populations of macrophytes were measured in experimental units of 0.8 × 0.8 m within the plots. MDS analysis showed that both sediment removal and burial caused changes in the species composition of the seagrass community. At one station, burial had a greater impact than sediment removal, whereas at the other station, the degree of impact of both types of disturbance was similar. Some macrophytes were consistently removed more than others, supporting the selective elimination hypothesis. Populations of Thalassia testudinum Banks ex König, Halimeda spp., and spongy algae (Avrainvillea spp. and Cladocephalus spp.) were, in almost all cases, undamaged by experimental manipulations. The populations of Syringodium filiforme Kütz., brush-like algae (Penicillus spp. and Rhipocephalus spp.), and Udotea spp. were reduced by more than 70%, when averaged across all manipulations and stations. A comparative analysis of growth forms of the above-mentioned macrophytes suggested that a solid, deeply anchored root-rhizome or rhizoid system, combined with a flexible or modular above-ground structure, is an advantageous characteristic to resist perturbation by hurricanes or storms.     

Characterisation of community structure of bacteria in parallel mesophilic and thermophilic pilot scale anaerobe sludge digesters

The aim of the present work was to compare the microbial communities of a mesophilic and a thermophilic pilot scale anaerobe sludge digester. For studying the communities cultivation independent chemotaxonomical methods (RQ and PLFA analyses) and T-RFLP were applied. Microbial communities of the mesophilic and thermophilic pilot digesters showed considerable differences, both concerning the species present, and their abundance. A Methanosarcina sp. dominated the thermophilic, while a Methanosaeta sp. the mesophilic digester among Archaea. Species diversity of Bacteria was reduced in the thermophilic digester. Based on the quinone patterns in both digesters the dominance of sulphate reducing respiratory bacteria could be detected. The PLFA profiles of the digester communities were similar though in minor components characteristic differences were shown. Level of branched chain fatty acids is slightly lower in the thermophilic digester that reports less Gram positive bacteria. The relative ratio of fatty acids characteristic to Enterobacteriaceae, Bacteroidetes and Clostridia shows differences between the two digesters: their importance generally decreased under thermophilic conditions. The sulphate reducer marker (15:1 and 17:1) fatty acids are present in low quantity in both digesters.     

Bioelectromagnetic field effects on cancer cells and mice tumors

We present possibilities and trends of ELF bioelectromagnetic effects in the mT amplitude range on cancer cells and on mice bearing tumors. In contrast to invasive electrochemotherapy and electrogenetherapy, using mostly needle electrodes and single high-amplitude electropulses for treatment, extremely low-frequency (ELF) pulsating electromagnetic fields (PEMF) and sinusoidal electromagnetic fields (SEMF) induce tumor cell apoptosis, inhibit angiogenesis, impede proliferation of neoplastic cells, and cause necrosis non invasively, whereas human lymphocytes are negligibly affected. Our successful results in killing cancer cells-analyzed by trypan blue staining or by flow cytometry-and of the inhibition of MX-1 tumors in mice by 15-20?mT, 50?Hz treatment in a solenoid coil also in the presence of bleomycin are presented in comparison to similar experimental results from the literature. In conclusion, the synergistic combinations of PEMF or SEMF with hyperthermia (41.5°C) and/or cancerostatic agents presented in the tables for cells and mice offer a basis for further development of an adjuvant treatment for patients suffering from malignant tumors and metastases pending the near-term development of suitable solenoids of 45-60?cm in diameter, producing >20?mT in their cores.     

Annual pollen traps reveal the complexity of climatic control on pollen productivity in Europe and the Caucasus

Annual PAR (pollen accumulation rates; grains cm^?2 year^?1) were studied with modified Tauber traps situated in ten regions, in Poland (Roztocze), the Czech Republic (two regions in Krkono?e, two in ?umava), Switzerland (4 regions in the Alps), and Georgia (Lagodekhi). The time-series are 10–16 years long, all ending in 2007. We calculated correlations between pollen data and climate. Pollen data are PAR summarized per region (4–7 traps selected per region) for each pollen type (9–14 per region) using log-transformed, detrended medians. Climate data are monthly temperature and precipitation measured at nearby stations, and their averages over all possible 2- to 6-month windows falling within the 20-month window ending with August, just prior to the yearly pollen-trap collection. Most PAR/climate relationships were found to differ both among pollen types and among regions, the latter probably due to differences among the study regions in the habitats of plant populations. Results shared by a number of regions can be summarized as follows. Summer warmth was found to enhance the following year’s PAR of Picea, Pinus non-cembra, Larix and Fagus. Cool summers, in contrast, increase the PAR of Abies, Alnus viridis and Gramineae in the following year, while wet summers promote PAR of Quercus and Gramineae. Wetness and warmth in general were found to enhance PAR of Salix. Precipitation was found to be more important for PAR of Alnus glutinosa-type than temperature. Weather did not have an impact on the PAR of Gramineae, and possibly of Cyperaceae in the same year. Care is advised when extrapolating our results to PAR in pollen sequences, because there are large errors associated with PAR from sediments, due to the effects of taphonomy and sedimentation and high uncertainty in dating. In addition, in pollen sequences that have decadal to centennial rather than near-annual resolution, plant-interaction effects may easily out-weigh the weather signal.     

Proteomic Analysis of the Maternal Preoptic Area in Rats

The behavior of female rats changes profoundly as they become mothers. The brain region that plays a central role in this regulation is the preoptic area, and lesions in this area eliminates maternal behaviors in rodents. The molecular background of the behavioral changes has not been established yet; therefore, in the present study, we applied proteomics to compare protein level changes associated with maternal care in the rat preoptic area. Using 2-dimensional fluorescence gel electrophoresis followed by identification of altered spots with mass spectrometry, 12 proteins were found to be significantly increased, and 6 proteins showed a significantly reduced level in mothers. These results show some similarities with a previous proteomics study of the maternal medial prefrontal cortex and genomics approaches applied to the preoptic area. Gene ontological analysis suggested that most altered proteins are involved in glucose metabolism and neuroplasticity. These proteins may support the maintenance of increased neuronal activity in the preoptic area, and morphological changes in preoptic neuronal circuits are known to take place in mothers. An increase in the level of alpha-crystallin B chain (Cryab) was confirmed by Western blotting. This small heat shock protein may also contribute to maintaining the increased activity of preoptic neurons by stabilizing protein structures. Common regulator and target analysis of the altered proteins suggested a role of prolactin in the molecular changes in the preoptic area. These results first identified the protein level changes in the maternal preoptic area. The altered proteins contribute to the maintenance of maternal behaviors and may also be relevant to postpartum depression, which can occur as a molecular level maladaptation to motherhood.

     

Patient-specific in silico endovascular repair of abdominal aortic aneurysms: application and validation

Biomechanics and Modeling in Mechanobiology - Non-negligible postinterventional complication rates after endovascular aneurysm repair (EVAR) leave room for further improvements. Since the potential...     

SPATIAL AND TEMPORAL VARIABILITY IN BIOMASS AND PRODUCTION OF PSAMMOPHYTIC HALIMEDA INCRASSATA (BRYOPSIDALES,CHLOROPHYTA) IN A CARIBBEAN REEF LAGOON1

Estimates for the production of calcium carbonate by Halimeda spp. have been based on limited measures in small areas or over short periods, subsequently extrapolated over larger temporal and spatial scales. The accuracy of these extrapolations depends on the variations in time and space of the parameters used for the derivations of the production, which were evaluated in the present study for Halimeda incrassata (J. Ellis) J. V. Lamour. in the Puerto Morelos reef lagoon, Mexican Caribbean. Growth, biomass, and CaCO₃ content of the branches of the thalli were determined at 1–3 monthly intervals from November 1997 until June 1999, using the marking technique with the coloring agent Alizarin Red‐S. Biomass exhibited great variability (V=25.4%), and fluctuations were closely related to changes in thallus density. Growth and CaCO₃‐content changes were related to the solar cycle, having coefficients of 15.4% and 2.5%, respectively. Additionally, calcified biomasses of the thalli were determined over a large spatial (31 stations in ～7 km² area) and time (6 y) scale. Spatial variability in calcified biomass was 59%, and the coefficient of variation attained its highest value (69%) for the samples collected over a 6 y period, from 1990 to 1996 (except 1995). Based on overall average values, branches of H. incrassata in the study area had a mean turnover of 30 d, with an annual production of 815 g CaCO₃ per square meter.