Cleavage in<Emphasis Type="Italic"> Nemertoderma westbladi</Emphasis> (Nemertodermatida) and its phylogenetic significance

Recent phylogenetic analyses of ribosomal and protein coding nuclear genes place the marine worms within the Nemertodermatida as one of the oldest lineages among the bilaterian animals. We studied the early embryonic cleavage in Nemertoderma westbladi to provide the first account of nemertodermatid early development. Live embryos were studied with interference microscopy and fixed embryos were either sectioned or studied with confocal laser scanning microscopy. Initially the divisions in the embryo are radial, but then micromeres are shifted clockwise generating a spiral pattern. The four-cell stage is characterized by duets of macromeres and micromeres and thus resembles the duet cleavage reported from members of the Acoela. However, subsequent stages differ from the acoel duet pattern and also from quartet spiral cleavage. The optimization of the cleavage pattern on current phylogenetic hypotheses with Nemertodermatida and Acoela as early bilaterian branches is discussed.     

The mechanism of EDHF-mediated responses in subcutaneous small arteries from healthy pregnant women

We studied the importance of endothelium-derived hyperpolarizing factor (EDHF) vs. nitric oxide (NO) and prostacyclin (PGI(2)) in bradykinin (BK)-induced relaxation in isolated small subcutaneous arteries from normal pregnant women. We also explored the contribution of cytochrome P-450 (CYP450) product of arachidonic acid (AA) metabolism, hydrogen peroxide (H(2)O(2)), and gap junctions that have been suggested to be involved in EDHF-mediated responses. Isolated arteries obtained from subcutaneous fat biopsies of normal pregnant women (n = 30) undergoing planned cesarean section were mounted in a wire-myography system. In norepinephrine-constricted vessels, incubation with N(G)-nitro-L-arginine methyl ester (L-NAME) resulted in a significant reduction in relaxation to BK. Simultaneous incubation with L-NAME and indomethacin failed to modify this response further. BK-mediated dilatation in the presence of K(+)-modified solution was decreased to similar level as obtained after incubation with L-NAME. Incubation with L-NAME abolished BK-induced responses in K(+)-modified solution. Sulfaphenazole, a specific inhibitor of CYP450 epoxygenase, and catalase (an enzyme that decomposes H(2)O(2)) did not affect the EDHF-mediated relaxation because concentration-response curves to BK were similar in arteries after incubation with L-NAME vs. L-NAME + sulfaphenazole and L-NAME + catalase. The inhibitor of gap junctions, 18 alpha-glycyrrhetinic acid, significantly reduced BK-mediated relaxation both without and with incubation with L-NAME. We found that both NO and EDHF, but not PGI(2), are involved in the endothelium-dependent dilatation to BK. BK-induced relaxation is almost equally mediated by NO and EDHF. CYP450 epoxygenase metabolites of AA or H(2)O(2) do not account for EDHF-mediated response; however, gap junctions are involved in the EDHF-mediated responses to BK in subcutaneous small arteries in normal pregnancy.     

Detection of intracellular bacteria in the buds of Scotch pine (Pinus sylvestris L.) by in situ hybridization

Bacterial isolates were obtained from pine (Pinus sylvestris L.) tissue cultures and identified as Methylobacterium extorquens and Pseudomonas synxantha. The existence of bacteria in pine buds was investigated by 16S rRNA in situ hybridization. Bacteria inhabited the buds of every tree examined, primarily colonizing the cells of scale primordia and resin ducts.     

The mechanism of contribution of integrin linked kinase (ILK) to epithelial-mesenchymal transition (EMT)

Integrin linked kinase (ILK) is ubiquitously expressed serine/threonine protein kinase, a binding partner of β1 and β3 integrin subunit as a cytoplasmic effector of integrin receptors that functionally links them to the actin cytoskeleton.We postulate that ILK is important enzyme involved in epithelial-mesenchymal transition (EMT) a critical event in the process of cancer progression. Commonly used EMT molecular markers include among others increased expression of N-cadherin and vimentin, nuclear localization of β-catenin, and the decrease of E-cadherin synthesis. In this study we were able to show that N-cadherin expression in melanoma cells is dependent on ILK signaling and the translocation of β-catenin to the nucleus. Silencing of ILK expression by siRNA significantly inhibited the stabilization and subsequent nuclear translocation of β-catenin and the expression of N-cadherin, a crucial molecule in the EMT, which facilitates association with fibroblast and endothelial cells during invasion of various cancers. The results allow to cautiously speculate on the important role of ILK in the cross-talk between integrins and cadherins accompanying EMT in melanoma.     

Synthesis of a 3'-naphthamido-LacNAc fluorescein conjugate with high selectivity and affinity for galectin-3

Described is the synthesis of a fluorescent LacNAc derivative appended with a 3'-deoxy-3'-naphthamido functionality, 2-(fluorescein-5/6-amido)ethyl 3-deoxy-3-(2-naphthamido)-beta-D-galactopyranosyl-(1-->4)-2-acetamido-2-deoxy-beta-D-glucopyranoside, which confers high affinity (Kd 170 nM) and selectivity for galectin-3 via a stacking interaction with Arg144. Its use as a selective and sensitive galectin-3 probe is demonstrated with fluorescence polarization measurements.     

Inhibitor binding to the plasmepsin IV aspartic protease from Plasmodium falciparum

Plasmepsin IV (Plm IV) is one of the aspartic proteases present in the food vacuole of the malaria parasite Plasmodium falciparum involved in host hemoglobin degradation by the parasite. Using a series of previously synthesized plasmepsin inhibitors [Ersmark, K., et al. (2005) J. Med. Chem. 48, 6090-106], we report here experimental data and theoretical analysis of their inhibitory activity toward Plm IV. All compounds share a 1,2-dihydroxyethylene unit as the transition state mimic. They possess symmetric P1 and P1' side chains and either a diacylhydrazine, a five-membered oxadiazole ring, or a retroamide at the P2 and P2' positions. Experimental binding affinities are compared to those predicted by the linear interaction energy (LIE) method and an empirical scoring function, using both a crystal structure and a homology model for the enzyme. Molecular dynamics (MD) simulations of the modeled complexes allow a rational interpretation of the structural determinants for inhibitor binding. A ligand bearing a P2 and P2' symmetric oxadiazole which is devoid of amide bonds is identified both experimentally and theoretically as the most potent inhibitor of Plm IV. For the P2 and P2' asymmetric compounds, the results are consistent with earlier predictions regarding the mode of binding of this class of inhibitors to Plm II. Theoretical estimation of selectivity for some compounds is also reported. Significant features of the Plm IV binding pocket are discussed in comparison to related enzymes, and the results obtained here should be helpful for further optimization of inhibitors.     

Chemical Composition of the Essential Oils from Different Morphological Parts of Pinus cembra L.

The essential oils from needles, twigs, bark, wood, and cones of Pinus cembra were analyzed by GC‐FID, GC/MS, and ¹H‐NMR spectroscopy. More than 130 compounds were identified. The oils differed in the quantitative composition. The principal components of the oil from twigs with needles were α‐pinene (36.3%), limonene (22.7%) and β‐phellandrene (12.0%). The needle oil was dominated by α‐pinene (48.4%), whereas in the oil from bark and in the oil from twigs without needles there were limonene (36.2% and 33.6%, resp.) and β‐phellandrene (18.8% and 17.1%, resp.). The main constituents of the wood oil as well as cone oil were α‐pinene (35.2% and 39.0%, resp.) and β‐pinene (10.4% and 18.9%, resp.). The wood oil and the cone oil contained large amounts of oxygenated diterpenes in comparison with needle, twig, and bark oils.     

Influence of Environmental Factors on Essential Oil Variability in Origanum compactum Benth. Growing Wild in Morocco

The present study aimed to evaluate the influence of environmental factors on essential oils (EOs) composition of Origanum compactum populations sampled all over the distribution area of the species in Morocco, and to determine the extent of the chemical profiles throughout the geographical distribution of the species. The chemical compositions were submitted to canonical correlation analysis and canonical discriminant analysis that indicated a significant relationship between oil components and some environmental factors. According to their chemical composition and edapho‐climatic characteristics, two major groups of populations were differentiated. The first group was composed of samples growing in regions with humid climate, clayey, sandy, and alkaline soils. These samples showed high thymol, α‐terpineol, linalool, and carvacryl methyl oxide content. The second group consisted of plants belonging to semi‐arid climate, and growing at high altitudes and silty soils. These samples were characterized by high carvacrol, α‐thujene, α‐terpinene, and myrcene content. However, populations exposed to sub‐humid climate, appeared less homogeneous and belong mainly either to the first or second group. A significant correlation between some edaphic factors (pH, K₂O content, soil texture) and the EOs yield of O. compactum plants was evidenced. In spite of the correlation obtained for the oil composition with edapho‐climatic factors and the variance explained by the environmental data set, the observed EO diversity might be also genetically determined.     

Characterization and cytotoxic effect of aqua-(2,2′,2′′-nitrilotriacetato)-oxo-vanadium salts on human osteosarcoma cells

The use of protonated N-heterocyclic compound, i.e. 2,2′-bipyridinium cation, [bpyH⁺], enabled to obtain the new nitrilotriacetate oxidovanadium(IV) salt of the stoichiometry [bpyH][VO(nta)(H₂O)]H₂O. The X-ray measurements have revealed that the compound comprises the discrete mononuclear [VO(nta)(H₂O)]^? coordination ion that can be rarely found among other known compounds containing nitrilotriacetate oxidovanadium(IV) moieties. The antitumor activity of [bpyH][VO(nta)(H₂O)]H₂O and its phenanthroline analogue, [phenH][VO(nta)(H₂O)](H₂O)_0.5, towards human osteosarcoma cell lines (MG-63 and HOS) has been assessed (the LDH and BrdU tests) and referred to cis-Pt(NH₃)₂Cl₂ (used as a positive control). The compounds exert a stronger cytotoxic effect on MG-63 and HOS cells than in untransformed human osteoblast cell line. Thus, the [VO(nta)(H₂O)]^? containing coordination compounds can be considered as possible antitumor agents in the osteosarcoma model of bone-related cells in culture.     

Microbial biodiversity of meadows under different modes of land use: catabolic and genetic fingerprinting

The main goal of the study was to find differences in the bacterial community structure resulting from different ways of meadow management in order to get the first insight into microbial biodiversity in meadow samples. The next generation sequencing technique (454-pyrosequencing) was accompanied with the community level physiological profiling (CLPP) method in order to acquire combined knowledge of both genetic and catabolic bacterial fingerprinting of two studied meadows (hayland and pasture). Soil samples (FAO: Mollic Gleysol) were taken in April 2015 from the surface layer (0–20 cm). Significant differences of the bacterial community structure between the two analyzed meadows resulted from different land mode were evidenced by pyrosequencing and CLPP techniques. It was found that Alpha- and Gammaproteobacteria dominated in the hayland, whereas Delta- and Betaproteobacteria prevailed in the pasture. Additionally, the hayland displayed lower Firmicutes diversity than the pasture. Predominant bacterial taxa: Acidobacteria, together with Chloroflexi and Bacteroidetes seemed to be insensitive to the mode of land use, because their abundance remained at a similar level in the both studied meadows. The CLPP analysis confirmed much faster degradation of the carbon sources by microorganisms from the hayland rather than from the pasture. Amino acids were the most favoured carbon source groups utilized by microorganisms in contrast to carbohydrates, which were utilized to the lowest extent. The study clearly proved that the consequences of even moderate anthropogenic management are always changes in bacterial community structure and their metabolic activity. Bacterial taxa that are sensitive and resistant on modes of land use were determined.