A finite element based method to determine the properties of planar soft tissue.

A finite element based method to determine the incremental elastic material properties of planar membranes was developed and evaluated. The method is applicable to tissues that exhibit inhomogeneity, geometric and material nonlinearity, and anisotropy. Markers are placed on the tissue to form a four-node quadrilateral element. The specimen is loaded to an initial reference state, then three incremental loading sets are applied and the nodal displacements recorded. One of these loadings must include shear. These data are used to solve an over-determined system of equations for the tangent stiffness matrix. The method was first verified using analytical data. Next, data obtained from a latex rubber sheet were used to evaluate experimental procedures. Finally, experiments conducted on preconditioned rat skin revealed nonlinear orthotropic behavior. The vector norm comparing the applied and calculated nodal force vectors was used to evaluate the accuracy of the solutions.     

Bacterial and phytoplankton responses to nutrient amendments in a boreal lake differ according to season and to taxonomic resolution

Nutrient limitation and resource competition in bacterial and phytoplankton communities may appear different when considering different levels of taxonomic resolution. Nutrient amendment experiments conducted in a boreal lake on three occasions during one open water season revealed complex responses in overall bacterioplankton and phytoplankton abundance and biovolume. In general, bacteria were dominant in spring, while phytoplankton was clearly the predominant group in autumn. Seasonal differences in the community composition of bacteria and phytoplankton were mainly related to changes in observed taxa, while the differences across nutrient treatments within an experiment were due to changes in relative contributions of certain higher- and lower-level phylogenetic groups. Of the main bacterioplankton phyla, only Actinobacteria had a treatment response that was visible even at the phylum level throughout the season. With increasing resolution (from 75 to 99% sequence similarity) major responses to nutrient amendments appeared using 454 pyrosequencing data of 16S rRNA amplicons. This further revealed that OTUs (defined by 97% sequence similarity) annotated to the same highly resolved freshwater groups appeared to occur during different seasons and were showing treatment-dependent differentiation, indicating that OTUs within these groups were not ecologically coherent. Similarly, phytoplankton species from the same genera responded differently to nutrient amendments even though biovolumes of the majority of taxa increased when both nitrogen and phosphorus were added simultaneously. The bacterioplankton and phytoplankton community compositions showed concurrent trajectories that could be seen in synchronous succession patterns over the season. Overall, our data revealed that the response of both communities to nutrient changes was highly dependent on season and that contradictory results may be obtained when using different taxonomic resolutions.     

Improved in vitro development rates of sheep somatic nuclear transfer embryos by using a reverse-order zona-free cloning method

This paper describes a modified zona-free cloning protocol for examining the effects of short-term exposure of donor nuclei to maternal chromosomal components and associated factors. In vitro matured zona-free sheep oocytes were enucleated by micromanipulator-assisted aspiration either before or after fusion with adult fibroblast or granulosa donor cells. Subsequent kinetics of donor nuclei and maternal chromatin as well as in vitro embryo development rates were recorded. The effect of an additional activation stimulus in connection with the reverse-order cloning (fusion before enucleation) and the feasibility of manual enucleation by metal blade were also studied. As a result of the simultaneous fusion and activation, most donor nuclei remained in interphase but swelled in size. Maternal chromosomes reached anaphase II-telophase II stages within 1-2 h of activation, effectively facilitating telophase enucleation with both the micromanipulator-assisted aspiration and manual bisection. A significantly higher development rate to the blastocyst stage was achieved with the reverse-order protocol, suggesting further investigation into the possible role of oocyte nucleus-associated factors in reprogramming is warranted. Overall, the reverse-order zona-free cloning method was efficient in the production of transferable cloned sheep blastocysts and may offer yet another choice of methodology in the practical application of nuclear transfer technology.     

Alpha-methylated derivatives of 2-arachidonoyl glycerol: synthesis, CB1 receptor activity, and enzymatic stability

Alpha-methylated analogues of the endogenous cannabinoid, 2-arachidonoyl glycerol (2-AG), were synthesized aiming to the improved enzymatic stability of 2-AG. In addition, the CB1 activity properties of fluoro derivatives of 2-AG were studied. The CB1 receptor activity was determined by the [35S]GTPgammaS binding assay, and the enzymatic stability of alpha-methylated analogues was determined in rat cerebellar membranes. The results indicate that even if the alpha-methylated 2-AG derivatives are slightly weaker CB1 receptor agonists than 2-AG, they are clearly more stable than 2-AG. In addition, the results showed that the replacement of the hydroxyl group(s) of 2-AG by fluorine does not improve the CB1 activity of 2-AG.     

Microsatellite diversity associated with ecological factors in Hordeum spontaneum populations in Israel

Microsatellite diversity at 18 loci was analysed in 94 individual plants of 10 wild barley, Hordeum spontaneum (C. Koch) Thell., populations sampled from Israel across a southward transect of increasing aridity. Allelic distribution in populations was not distributed randomly. Estimates of mean gene diversity were highest in stressful arid-hot environments. Sixty-four per cent of the genetic variation was partitioned within populations and 36% between populations. Associations between ecogeographical variables and gene diversity, H(e), were established in nine microsatellite loci. By employing principle component analysis we reduced the number of ecogeographical variables to three principal components including water factors, temperature and geography. At three loci, stepwise multiple regression analysis explained significantly the gene diversity by a single principal component (water factors). Based on these observations it is suggested that simple sequence repeats are not necessarily biologically neutral.     

Genome Sequence of Lactobacillus crispatus ST1

Lactobacillus crispatus is a common member of the beneficial microbiota present in the vertebrate gastrointestinal and human genitourinary tracts. Here, we report the genome sequence of L. crispatus ST1, a chicken isolate displaying strong adherence to vaginal epithelial cells.Lactobacillus crispatus can persist in the vertebrate gastrointestinal tract and is among the most prevalent species of the Lactobacillus-dominated human vaginal microbiota (2, 9, 13, 14). It belongs to the so-called acidophilus group (3), which has attracted interest because some of its species are important factors in the production of fermented foods (12) and some can, at least transiently, colonize the human host (2, 9, 13, 14). Moreover, some specific strains, mainly L. acidophilus NCFM and L. johnsonii NCC 533, have received prominence as intestinal-health-promoting microbes (4). Although the genomes of seven members of the acidophilus complex have been sequenced to date (12), the genome sequences of L. crispatus and other predominant lactobacillar species in the urogenital flora have mostly remained obscure. Vaginal lactobacilli can have an important role in controlling the health of the host (2, 14). They can, for example, positively influence and stabilize the host''s vaginal microbiota via the production of compounds that are acidic or exert a direct inhibiting action toward pathogenic bacteria (2, 14). In addition to the antimicrobial compounds, the competitive exclusion of pathogens is another mechanism by which the host''s microbiota can be balanced (2). L. crispatus ST1 was originally isolated from the crop of a chicken, and PCR profiling of L. crispatus isolates has verified it to be an abundant colonizer of the chicken crop (6, 8). It also displays a strong protein-dependent adhesion to the epithelial cells of the human vagina and has been shown to inhibit the adhesion of avian pathogenic Escherichia coli (6, 7).The genome was sequenced (18× coverage) using a 454 pyrosequencer with GS FLX chemistry (Roche). The contig order was confirmed and gaps were filled by sequencing PCR fragments from the genomic DNA template using ABI 3730 and Big Dye chemistry (Applied Biosystems). Genomic data were processed using the Staden Package (11) and gsAssembler (Roche). Coding sequences (CDSs) were predicted using Glimmer3 (5) followed by manual curation of the start sites. The remaining intergenic regions were reanalyzed for missed CDSs by using BlastX (1). Annotation transfer was performed based on a BlastP search, followed by Blannotator analysis using default settings (http://ekhidna.biocenter.helsinki.fi/poxo/blannotator) and manual verification. Orthologous groups between the different lactobacillar proteomes were identified using OrthoMCL (10).The genome of L. crispatus ST1 consists of a single circular chromosome 2.04 Mbp in size, with an overall G+C content of 37%, without any plasmids. There are 64 tRNA genes, 4 rRNA operons, and 2 CRISPR loci. Out of the 2,024 predicted CDSs, a putative function was assigned to 77%, whereas 10% of the CDSs were annotated as conserved and 13% as novel. Based on the orthologous grouping, 302 (15%) of the CDSs encoded by ST1 have no detectable homologs in any of the Lactobacillus proteomes published to date.     

Preattentive auditory information processing under exposure to the 902 MHz GSM mobile phone electromagnetic field: A mismatch negativity (MMN) study

Previous studies on the effects of the mobile phone electromagnetic field (EMF) on various event‐related potential (ERP) components have yielded inconsistent and even contradictory results, and often failed in replication. The mismatch negativity (MMN) is an auditory ERP component elicited by infrequent (deviant) stimuli differing in some physical features from the repetitive frequent (standard) stimuli in a sound sequence. The MMN provides a sensitive measure for cortical auditory stimulus feature discrimination, regardless of attention and other contaminating factors. In this study, MMN responses to duration, intensity, frequency, and gap changes were recorded in healthy young adults (n = 17), using a multifeature paradigm including several types of auditory change in the same stimulus sequence, while a GSM mobile phone was placed on either ear with the EMF (902 MHz pulsed at 217 Hz; SAR_1g = 1.14 W/kg, SAR_10g = 0.82 W/kg, peak value = 1.21 W/kg, measured with an SAM phantom) on or off. An MMN was elicited by all deviant types, while its amplitude and latency showed no significant differences due to EMF exposure for any deviant types. In the present study, we found no conclusive evidence that acute exposure to GSM mobile phone EMF affects cortical auditory change detection processing reflected by the MMN. Bioelectromagnetics 30:241–248, 2009. © 2009 Wiley‐Liss, Inc.     

Neolithic land use in the northern Boreal zone: high-resolution multiproxy analyses from Lake Huhdasjärvi,south-eastern Finland

Two high-resolution pollen and charcoal analyses were constructed from sediments obtained from a small bay in eastern Finland in order to gain information on human activity during the Neolithic Stone Age, 5200–1800 bc. We used measurements of loss on ignition (LOI), magnetic susceptibility and geochemical analyses to describe the sedimentological characteristics. Palaeomagnetic dating and measurements of ¹³⁷Cs-activity were supported by ¹⁴C-datings. The analyses revealed human activity between 4400 and 3200 bc, which is synchronous with archaeological cultures defined through different stages of Comb Ware pottery types and Middle Neolithic pottery types with asbestos as a primary temper. Direct evidence of Hordeum cultivation was dated to 4040–3930 cal bc. According to the pollen data, more significant effort was put into the production of fibres from hemp and lime than the actual cultivation of food.     

A comparison of established and new approaches in ovine and bovine nuclear transfer

Several breakthroughs in nuclear transfer research were first achieved in sheep, although cattle soon became the main livestock species of interest. However, sheep still offer significant advantages both in basic and applied research. With increased interest in cloning of livestock, new approaches have been developed for both sheep and cattle nuclear transfer technology. These include methods for zona-free nuclear transfer that can be performed with or without the use of micromanipulator. Here we describe four different nuclear transfer methods including the traditional micromanipulation-assisted method in sheep, zona-free method in sheep in which the order of enucleation and nucleus delivery have been reversed ("reverse-order" cloning) and zona free manual cloning methods ("hand-made cloning") for embryonic and somatic cloning in cattle. The purpose of this paper is to encourage people to familiarize themselves with these different methods available and to help them choose and test the method most suitable for their particular circumstances.     

Discovery of Triterpenoids as Reversible Inhibitors of α/β-hydrolase Domain Containing 12 (ABHD12)

Background

α/β-hydrolase domain containing (ABHD)12 is a recently discovered serine hydrolase that acts in vivo as a lysophospholipase for lysophosphatidylserine. Dysfunctional ABHD12 has been linked to the rare neurodegenerative disorder called PHARC (polyneuropathy, hearing loss, ataxia, retinosis pigmentosa, cataract). In vitro, ABHD12 has been implicated in the metabolism of the endocannabinoid 2-arachidonoylglycerol (2-AG). Further studies on ABHD12 function are hampered as no selective inhibitor have been identified to date. In contrast to the situation with the other endocannabinoid hydrolases, ABHD12 has remained a challenging target for inhibitor development as no crystal structures are available to facilitate drug design.

Methodology/Principal Findings

Here we report the unexpected discovery that certain triterpene-based structures inhibit human ABHD12 hydrolase activity in a reversible manner, the best compounds showing submicromolar potency. Based on structure activity relationship (SAR) data collected for 68 natural and synthetic triterpenoid structures, a pharmacophore model has been constructed. A pentacyclic triterpene backbone with carboxyl group at position 17, small hydrophobic substituent at the position 4, hydrogen bond donor or acceptor at position 3 accompanied with four axial methyl substituents was found crucial for ABHD12 inhibitor activity. Although the triterpenoids typically may have multiple protein targets, we witnessed unprecedented selectivity for ABHD12 among the metabolic serine hydrolases, as activity-based protein profiling of mouse brain membrane proteome indicated that the representative ABHD12 inhibitors did not inhibit other serine hydrolases, nor did they target cannabinoid receptors.

Conclusions/Significance

We have identified reversibly-acting triterpene-based inhibitors that show remarkable selectivity for ABHD12 over other metabolic serine hydrolases. Based on SAR data, we have constructed the first pharmacophore model of ABHD12 inhibitors. This model should pave the way for further discovery of novel lead structures for ABHD12 selective inhibitors.