Immunochemical studies of Salmonella Dakar and Salmonella Telaviv O-antigens (serogroup O:28)

Salmonella Dakar and Salmonella Telaviv bacteria belong to serogroup O:28, which represents 107 serovars and possesses only the epitope O28. Salmonella Telaviv has the subfactors O28(1) and O28(2) , whereas S. Dakar has O28(1) and O28(3) . So far, only limited serological and immunological information for this serogroup is available in the literature. Knowledge of the structures of their O-polysaccharides and the immunochemical investigations performed in this work allowed to reveal the nature of subfactor O28(1) as attributed to the presence of 3-linked (or 3,4-disubstituted) α-d-GalpNAc in the main chains of S. Dakar and S. Telaviv O-polysaccharides. An explanation for the cross-reactions between Salmonella enterica O28 O-antigens and other Salmonella O-polysaccharides and their structural similarity to Escherichia coli O-serogroups is also given.     

Endothelin induces rapid, dynamin-mediated budding of endothelial caveolae rich in ET-B

Clathrin-independent trafficking pathways for internalizing G protein-coupled receptors (GPCRs) remain undefined. Clathrin-mediated endocytosis of receptors including ligand-engaged GPCRs can be very rapid and comprehensive (<10 min). Caveolae-mediated endocytosis of ligands and antibodies has been reported to be much slower in cell culture (≫10 min). Little is known about the role of physiological ligands and specific GPCRs in regulating caveolae trafficking. Here, we find that one receptor for endothelin, ET-B but not ET-A, resides on endothelial cell surfaces in both tissue and cell culture primarily concentrated within caveolae. Reconstituted cell-free budding assays show that endothelins (ETs) induce the fission of caveolae from endothelial plasma membranes purified from rat lungs. Electron microcopy of lung tissue sections and tissue subcellular fractionation both show that endothelin administered intravascularly in rats also induces a significant loss of caveolae at the luminal surface of lung vascular endothelium. Endothelial cells in culture show that ET stimulates very rapid internalization of caveolae and cargo including caveolin, caveolae-targeting antibody, and itself. The ET-B inhibitor BQ788, but not the ET-A inhibitor BQ123, blocks the ET-induced budding of caveolae. Both the pharmacological inhibitor Dynasore and the genetic dominant negative K44A mutant of dynamin prevent this induced budding and internalization of caveolae. Also shRNA lentivirus knockdown of caveolin-1 expression prevents rapid internalization of ET and ET-B. It appears that endothelin can engage ET-B already highly concentrated in caveolae of endothelial cells to induce very rapid caveolae fission and endocytosis. This transport requires active dynamin function. Caveolae trafficking may occur more rapidly than previously documented when it is stimulated by a specific ligand to signaling receptors already located in caveolae before ligand engagement.     

Analysis of mercury levels in historical bone material from syphilitic subjects--pilot studies (short report)

The aim of the present work is to study the remains of seven individuals with typical symptoms of tertiary syphilis in terms of mercury content in bones, thereby verifying whether they were subjected to treatment and, if they were, how long their organisms were exposed to toxic mercury fumes. Mercury was used, mainly in the Middle Ages and in the early modern period, as a preventive measure in case of individuals suffering from syphilis, a venereal disease, and also leprosy. Syphilitic patients treated this way should demonstrate increased concentration of mercury in their bones. The skeletons studied in the present work originate from various archaeological sites in southern and north-central Poland. The analyses concerned individuals with diagnosed syphilis as well as healthy individuals who constituted the control group. The analyses were performed by the LA-ICP-MS technique, with the use of laser Nd: YAG, Macro, 266 nm, New Wave, USA, coupled with Spectrometer Elan DRC-e Perkin Elmer, USA. The content analysis of the studied bone material revealed with high probability that the contact method of mercurial treatment was used only in the case of two women from north-central Poland, deceased at the turn of the 15th century at the earliest.     

A Study on Selected Physiological Parameters of Plants Grown Under Lithium Supplementation

Exposure of sunflower and maize plants to increasing concentrations of lithium (0?C50?mg Li dm^?3) in a nutrient solution induced changes in biomass, leaf area and photosynthetic pigment accumulation, as well as levels of lipid peroxidation. The highest applied lithium dose (50?mg Li dm^?3) evoked a significant reduction in the shoot biomass for both examined species, as well as necrotic spots and a reduction of the leaf area in sunflower plants. An enrichment of a nutrient solution with 5?C50?mg Li dm^?3 did not significantly affect chlorophylls a and b and the carotenoid content in sunflower plants. However, in maize, a significant decrease in all pigment content under highest used lithium concentration was noted. The levels of lipid peroxidation of the cell membranes in leaves of sunflower plants and the roots of maize increased significantly in the presence of 50?mg Li dm^?3, which suggests disturbances of the membrane integrity and pro-oxidant properties of the excess lithium ions. Nonetheless, in maize, an increase of shoot biomass and leaf area in the presence of 5?mg Li dm^?3 was found. An analysis of the metal content indicated that lithium accumulated significantly in sunflower and maize shoots in a dose-dependent manner, but differences occurred between species. The sunflower plants accumulated considerably greater amounts of this metal than maize. The potassium content in shoots remained unchanged under lithium treatments, except for a significant increase in the potassium levels for sunflower plants grown in the presence of 50?mg Li dm^?3. These results suggest that lithium at 50?mg Li dm^?3 is toxic to both plant species, but the symptoms of toxicity are species-specific. Moreover, the lithium influence on plants is dose-dependent and its ions can exert toxicity at high concentrations (50?mg Li dm^?3) or stimulate growth at low concentrations (5?mg Li dm^?3).     

Production of bioactive phenolic acids and furanocoumarins in in vitro cultures of Ruta graveolens L. and Ruta graveolens ssp. divaricata (Tenore) Gams. under different light conditions

Extracts from the biomass of Ruta graveolens and Ruta graveolens ssp. divaricata cultured in vitro under different light conditions (far-red, red and blue light, UV-A irradiation, in darkness and white light) were tested for the amounts of free phenolic acids and cinnamic acid (twelve compounds) as well as furanocoumarins and umbelliferone (seven compounds) using HPLC methods. Total amounts of the investigated groups of compounds in the cultures of both plants increased from 2.6 to 6.7 times, depending on light quality, and the maximum values reached were 106.50 and 1,276.74?mg?100?g^?1 DW (in R. graveolens), and 106.97 and 262.54?mg?100?g^?1 DW (in the subspecies), respectively. Both white light and blue light were equally beneficial for the total production of phenolic acids in cultures of both plants, whereas the total production of furanocoumarins was clearly better stimulated by blue light in R. graveolens and by darkness in the subspecies (i.e. the amounts were respectively 1.44 and 1.7 times higher than in the biomass cultivated under white light). The amounts of individual compounds in both plant cultures increased from about 2.2 to 26.3 times depending on light quality. The following bioactive compounds were obtained in quantities which are of interest from a practical perspective: in R. graveolens culture??protocatechuic acid (45?mg?100?g^?1 DW), isopimpinellin (about 500?mg?100?g^?1 DW) and bergapten (about 270?mg?100?g^?1 DW), and in the subspecies culture: p-coumaric acid (70?mg?100?g^?1 DW) and isopimpinellin (about 210?mg?100?g^?1 DW).     

Insights into the regulation of eukaryotic elongation factor 2 kinase and the interplay between its domains

eEF2K (eukaryotic elongation factor 2 kinase) is a Ca2+/CaM (calmodulin)-dependent protein kinase which regulates the translation elongation machinery. eEF2K belongs to the small group of so-called 'α-kinases' which are distinct from the main eukaryotic protein kinase superfamily. In addition to the α-kinase catalytic domain, other domains have been identified in eEF2K: a CaM-binding region, N-terminal to the kinase domain; a C-terminal region containing several predicted α-helices (resembling SEL1 domains); and a probably rather unstructured 'linker' region connecting them. In the present paper, we demonstrate: (i) that several highly conserved residues, implicated in binding ATP or metal ions, are critical for eEF2K activity; (ii) that Ca2+/CaM enhance the ability of eEF2K to bind to ATP, providing the first insight into the allosteric control of eEF2K; (iii) that the CaM-binding/α-kinase domain of eEF2K itself possesses autokinase activity, but is unable to phosphorylate substrates in trans; (iv) that phosphorylation of these substrates requires the SEL1-like domains of eEF2K; and (v) that highly conserved residues in the C-terminal tip of eEF2K are essential for the phosphorylation of eEF2, but not a peptide substrate. On the basis of these findings, we propose a model for the functional organization and control of eEF2K.     

Lectin chromatography/mass spectrometry discovery workflow identifies putative biomarkers of aggressive breast cancers

We used a lectin chromatography/MS-based approach to screen conditioned medium from a panel of luminal (less aggressive) and triple negative (more aggressive) breast cancer cell lines (n=5/subtype). The samples were fractionated using the lectins Aleuria aurantia (AAL) and Sambucus nigra agglutinin (SNA), which recognize fucose and sialic acid, respectively. The bound fractions were enzymatically N-deglycosylated and analyzed by LC-MS/MS. In total, we identified 533 glycoproteins, ～90% of which were components of the cell surface or extracellular matrix. We observed 1011 glycosites, 100 of which were solely detected in ≥3 triple negative lines. Statistical analyses suggested that a number of these glycosites were triple negative-specific and thus potential biomarkers for this tumor subtype. An analysis of RNaseq data revealed that approximately half of the mRNAs encoding the protein scaffolds that carried potential biomarker glycosites were up-regulated in triple negative vs luminal cell lines, and that a number of genes encoding fucosyl- or sialyltransferases were differentially expressed between the two subtypes, suggesting that alterations in glycosylation may also drive candidate identification. Notably, the glycoproteins from which these putative biomarker candidates were derived are involved in cancer-related processes. Thus, they may represent novel therapeutic targets for this aggressive tumor subtype.     

Ceramide profiles in the brain of rats with diabetes induced by streptozotocin

Diabetes is associated with disturbances of brain activity and cognitive impairment. We hypothesize that ceramides may constitute an important contribution to diabetes-linked neuro-dysfunction. In our study we used rats injected with streptozotocin (STZ) as a model of severe hyperglycemia. Using the gas-liquid chromatography technique we found a significant increase of ceramide content in brains and a decrease in plasma of diabetic rats. The inhibitor of serine palmitoyltransferase, myriocin, reduced ceramide generation in hyperglycemic brains, although injected alone it exerted a paradoxical effect of ceramide upregulation. Myriocin had no impact on ceramide concentration in the plasma of either control or diabetic rats. The level of ceramide saturated fatty acids was elevated whereas the level of ceramide poly-unsaturated fatty acids was downregulated in brains of all experimental groups. The concentration of ceramide mono-unsaturated fatty acids remained unchanged. The pattern of individual ceramide species was altered depending on treatment. We noted an STZ-evoked increase of brain ceramide C16:0, C18:0 and C20:0 and a strong decline in ceramide C18:2 fatty acid levels. Some changes of brain ceramide pattern were modified by myriocin. We found a decreased amount of total ceramide-ω-6 fatty acids in STZ-treated rat brains and no changes in ceramide-ω-3 concentration. We conclude that ceramides may be important mediators of diabetes-accompanied brain dysfunction.     

MHC class II derived recombinant T cell receptor ligands protect DBA/1LacJ mice from collagen-induced arthritis

We previously demonstrated the therapeutic effects of MHC class II derived recombinant T cell receptor ligands (RTL), single-chain two domain complexes of the alpha1 and beta1 domains of MHC class II molecules genetically linked with an immunodominant peptide, in experimental autoimmune encephalomyelitis. In the current study, we produced a monomeric murine I-Aq-derived RTL construct covalently linked with bovine collagen type II peptide (bCII257-270) suitable for use in DBA/1LacJ mice that develop collagen-induced arthritis (CIA), an animal model of human rheumatoid arthritis, after immunization with bCII protein in CFA. In this study, we demonstrate that the I-Aq-derived RTLs reduced the incidence of the disease, suppressed the clinical and histological signs of CIA and induced long-term modulation of T cells specific for arthritogenic Ags. Our results showed that the I-Aq/bCII257-270 molecule could systemically reduce proinflammatory IL-17 and IFN-gamma production and significantly increase anti-inflammatory IL-10, IL-13, and FoxP3 gene expression in splenocytes. Moreover, I-Aq/bCII257-270 molecule could also selectively inhibit IL-1beta, IL-6, and IL-23 expression in local joint tissue. This is the first report demonstrating effective prevention of joint inflammation and clinical signs of CIA with an I-Aq-derived RTL, thus supporting the possible clinical use of this approach for treating rheumatoid arthritis in humans.     

Decrease in sex difference in premature mortality during system transformation in Poland

In recent years, sex differences in mortality have followed diverse patterns of change in developed countries. As there is no analogous evidence from Poland, the aim of this study was to describe the pattern of change in excess male mortality among Polish inhabitants aged 35-64 during 1995-2002, when the major socioeconomic transformation occurred, and compare it with sex differences in mortality observed in the late 1980s. During the study period, excess male mortality decreased significantly, independently of age and educational level. The reduction in mortality was observed in both sexes, but its magnitude was greater in men. These changes resulted mainly from a reduction in mortality rates due to cardiovascular disease and lung cancer in males and a concomitant increase in mortality rates due to lung cancer and suicides in females. Although, in general, excess male mortality decreased, social gradients related to this phenomenon increased. Subjects (in particularly men) who had graduated from university benefited the most, their magnitude of reduction in mortality rates being the greatest. Changes in social environment during the transformation period in Poland are suggested as major determinants of these changes, but further studies are needed.