Investigation of unmatured hematopoietic cell migration in a mouse model

The objective of this work was to examine the migration of transplanted bone marrow hematopoietic lin⁻ cell population using the BALB/c mouse contact hypersensitivity model in vivo and to determine the time point at which they reach the site of injury (paw edema) as well as other undamaged organs, such as liver and spleen. Female BALB/c mice with induced contact hypersensitivity reaction were intravenously injected with 1×10⁶ cells/mouse lin⁻ cells, labeled with PKH67. The presence of lin⁻ stained cells in mouse tissue sections was evaluated by fluorescent microscopy. After one hour, the labeled cells were found in mice paw edema and liver, and after 4 hours in spleen tissue. Migrated hematopoietic lin⁻ cells remained in liver tissue for 48 h, and in spleen and paw edema at least for 72 h. Migrated stained cells in untreated paw were not found. The results prove that bone marrow unmatured hematopoietic cells are first found in paw edema, where they participate in the inhibition of tissue inflammation; these cells subsequently migrate to the liver and are found in the spleen shortly afterwards.     

Ecological impact of Eurasian beaver (<Emphasis Type="Italic">Castor fiber</Emphasis>) activity on macroinvertebrate communities in Lithuanian trout streams

Our study found that beaver activity affects macroinvertebrate assemblages of both beaver ponds and downstream sites. The percentage composition of the invertebrate faunae of beaver ponds was strikingly different from the invertebrate faunae of upstream forested and downstream sites. The number of EPT (ephemeropteran, plecopteran, trichopteran) taxa in the upstream forested sites in all streams was higher than in beaver pond and downstream sites. Statistically significant differences were found in absolute and relative abundances of EPT and Chironomidae between different streams sites. The absolute and relative abundance of pollution-sensitive EPT was significantly higher in forested sites than in beaver pond and downstream sites in all measured streams. Beaver ponds had a significantly higher absolute and relative abundance of Chironomidae compared with upstream forested and downstream sites. We found that Plecoptera and Coleoptera were absent from beaver pond sites. The absolute abundance of Plecoptera was significantly higher in upstream forested sites than in downstream sites in all three streams. Gatherers were the dominant functional feeding group in relative abundance in all three habitat types. The percentage of gatherers was higher in beaver ponds than in forested and downstream sites.     

Mediated amperometry reveals two distinct modes of yeast responses to glucose

Mediated amperometry was exploited to monitor intracellular redox activity without cell disruption. Continuous measurements of menadione-mediated glucose currents at carbon paste electrodes with various immobilized intact wild type yeasts (Saccharomyces cerevisiae, Candida pulcherrima, Clavispora lusitaniae, Wickerhamomyces anomalus, Pichia guilliermondii, Kluyveromyces lactis var. lactis, Debaryomyces hansenii, Candida zeymolaydes and Candida tropicalis) revealed two distinct and previously unreported modes of development of the currents during the first 2 to 3 min. after subjection to glucose. A correlation among the values of the currents and the capacities of wild type yeasts to secrete various substances was observed.     

Litter‐dwelling beetles (Insecta: Coleoptera) can survive in clear‐cutting during subsequent soil ploughing

1. A 2‐year study of litter dwelling beetles was conducted in different mature pine stands and clear‐cuts in Lithuania using the litter sifting method. We hypothesized that clear‐cutting and subsequent ploughing would increase species diversity and the abundance of beetles, and also would encourage the immediate appearance of early‐successional beetle species replacing late successional species in the clear‐cuts.
2. We did not confirm a hypothesis regarding increase in the number of species and abundance of beetles in clear‐cuts.
3. Our hypothesis regarding the immediate appearance of early successional species and disappearance of late‐successional species in clear‐cuts was confirmed.
4. We also revealed that subsequent soil ploughing in clear‐cuts did not accelerate this process, which was linked to the possibility of late successional species surviving in the undisturbed spaces between the strips of ploughed soil.
5. The present study shows that late‐successional forest litter‐dwelling coleopteran species of old pine forests have a better chance of survival in a ploughed clear‐cut, at least in northern Europe, than has been reported in other studies.

     

Volume of Hsp90 ligand binding and the unfolding phase diagram as a function of pressure and temperature

Volume changes that accompany protein unfolding and ligand binding are important but largely neglected thermodynamic parameters that may facilitate rational drug design. Here, we determined the volume of lead compound ICPD47 binding to an anticancer target, heat shock protein 90 N-terminal domain, using a pressure shift assay (PressureFluor). The ligand exhibited a stabilizing effect on the protein by increasing its melting pressure and temperature. The Gibbs free energy of unfolding depends on the absence or presence of ligand and has an elliptical shape. Ellipse size increases upon addition of the strongly binding ligand, which stabilizes the protein. The three-dimensional (3D) ellipsoidal surface of the Gibbs free energy of unfolding was calculated with increasing ligand concentrations. The negative volume of ligand binding was relatively large and significantly exceeded the volume of protein unfolding. The pressure shift assay technique could be used to determine the volume changes associated with both protein unfolding as well as ligand binding to protein.     

High-Yield Expression in Escherichia coli,Purification and Application of Budding Yeast K2 Killer Protein

Saccharomyces cerevisiae K2 toxin is a highly active extracellular protein, important as a biocontrol agent for biotechnological applications in the wine industry. This protein is produced at negligible levels in yeast, making difficult to isolate it in amounts sufficient for investigation and generation of analysis tools. In this work, we demonstrate the use of a bacterial system for expression of the recombinant K2 protein, suitable for generation of antibodies specific for toxin of the yeast origin. Synthesis of the full-length S. cerevisiae K2 preprotoxin in Escherichia coli was found to be toxic to the host cell, resulting in diminished growth. Such effect was abolished by the introduction of the C-terminal truncation into K2 protein, directing it into non-toxic inclusion body fraction. The obtained protein is of limited solubility thus, facilitating the purification by simple and efficient chromatography-free procedure. The protein aggregates were successfully refolded into a soluble form yielding sufficient amounts of a tag-less truncated K2 protein suitable for polyclonal antibody production. Antibodies were raised in rabbit and found to be specific for detection of both antigen and native S. cerevisiae K2 toxin.     

An efficient and regioselective biocatalytic synthesis of aromatic N-oxides by using a soluble di-iron monooxygenase PmlABCDEF produced in the Pseudomonas species

Here, we present an improved whole-cell biocatalysis system for the synthesis of heteroaromatic N-oxides based on the production of a soluble di-iron monooxygenase PmlABCDEF in Pseudomonas sp. MIL9 and Pseudomonas putida KT2440. The presented biocatalysis system performs under environmentally benign conditions, features a straightforward and inexpensive procedure and possesses a high substrate conversion and product yield. The capacity of gram-scale production was reached in the simple shake-flask cultivation. The template substrates (pyridine, pyrazine, 2-aminopyrimidine) have been converted into pyridine-1-oxide, pyrazine-1-oxide and 2-aminopyrimidine-1-oxide in product titres of 18.0, 19.1 and 18.3 g l^-1, respectively. To our knowledge, this is the highest reported productivity of aromatic N-oxides using biocatalysis methods. Moreover, comparing to the chemical method of aromatic N-oxides synthesis based on meta-chloroperoxybenzoic acid, the developed approach is applicable for a regioselective oxidation that is an additional advantageous option in the preparation of the anticipated N-oxides.     

Response of juveniles of seven forest tree species and their populations to different combinations of simulated climate change-related stressors: spring-frost,heat, drought,increased UV radiation and ozone concentration under elevated CO2 level

Journal of Plant Research - The study aimed to assess response of juvenile progeny of seven forest tree species, Pinus sylvestris, Picea abies, Betula pendula, Alnus glutinosa, Populus tremula,...