The presence of the WGD motif in CC8 heterodimeric disintegrin increases its inhibitory effect on alphaII(b)beta3, alpha(v)beta3, and alpha5beta1 integrins

Two highly homologous dimeric disintegrins, CC5 and CC8, have been isolated from the venom of the North African sand viper Cerastes cerastes. CC5 is a homodimer containing an RGD motif in its subunits. CC8 is a heterodimer. The CC8A and CC8B subunits contain RGD and WGD tripeptide sequence in their respective integrin-binding loops. Both CC5 and CC8 inhibited platelet aggregation and the adhesion of cells expressing integrins alphaII(b)beta3, alpha(v)beta3, and alpha5beta1 to appropriate ligands. However, the inhibitory activity of CC8 was at least 1 order of magnitude higher than that of CC5. Enhanced activity of CC8 over CC5 was also observed in the induction of LIBS epitopes on beta1 and beta3 integrins. Synthetic peptides in which the arginyl residue of the RGD motif had been replaced with tryptophans exhibited increased inhibitory activity toward integrins alpha5beta1, alphaII(b)beta3, and alpha(v)beta3. Moreover, alanine substitution of the aspartic acid of the WGD motif of these peptides decreased their inhibitory ability, whereas the same substitution in the RGD sequence almost completely abolished the activity of the peptides. We conclude that the WGD motif enhances the inhibitory activity of disintegrins toward alphaII(b)beta3, alpha(v)beta3, and alpha5beta1 integrins.     

Survival and Competitiveness of Bradyrhizobium japonicum Strains 20 Years after Introduction into Field Locations in Poland

It was previously demonstrated that there are no indigenous strains of Bradyrhizobium japonicum forming nitrogen-fixing root nodule symbioses with soybean plants in arable field soils in Poland. However, bacteria currently classified within this species are present (together with Bradyrhizobium canariense) as indigenous populations of strains specific for nodulation of legumes in the Genisteae tribe. These rhizobia, infecting legumes such as lupins, are well established in Polish soils. The studies described here were based on soybean nodulation field experiments, established at the Poznań University of Life Sciences Experiment Station in Gorzyń, Poland, and initiated in the spring of 1994. Long-term research was then conducted in order to study the relation between B. japonicum USDA 110 and USDA 123, introduced together into the same location, where no soybean rhizobia were earlier detected, and nodulation and competitive success were followed over time. Here we report the extra-long-term saprophytic survival of B. japonicum strains nodulating soybeans that were introduced as inoculants 20 years earlier and where soybeans were not grown for the next 17 years. The strains remained viable and symbiotically competent, and molecular and immunochemical methods showed that the strains were undistinguishable from the original inoculum strains USDA 110 and USDA 123. We also show that the strains had balanced numbers and their mobility in soil was low. To our knowledge, this is the first report showing the extra-long-term persistence of soybean-nodulating strains introduced into Polish soils and the first analyzing the long-term competitive relations of USDA 110 and USDA 123 after the two strains, neither of which was native, were introduced into the environment almost 2 decades ago.     

Somatostatin-like immunoreactivity in the amygdala of the pig

The distribution and morphology of neurons containing somatostatin (SOM) was investigated in the amygdala (CA) of the pig. The SOM-immunoreactive (SOM-IR) cell bodies and fibres were present in all subdivisions of the porcine CA, however, their number and density varied depending on the nucleus studied. The highest density of SOM-positive somata was observed in the layer III of the cortical nuclei, in the anterior (magnocellular) part of the basomedial nucleus and in the caudal (large-celled) part of the lateral nucleus. Moderate to high numbers of SOM-IR cells were also observed in the medial and basolateral nuclei. Many labeled neurons were also consistently observed in the lateral part of the central nucleus. In the remaining CA regions, the density of SOM-positive cell bodies varied from moderate to low. In any CA region studied SOM-IR neurons formed heterogeneous population consisting of small, rounded or slightly elongated cell bodies, with a few poorly branched smooth dendrites. In general, morphological features of these cells clearly resembled the non-pyramidal Golgi type II interneurons. The routine double-labeling studies with antisera directed against SOM and neuropeptide Y (NPY) demonstrated that a large number of SOM-IR cell bodies and fibers in all studied CA areas contained simultaneously NPY. In contrast, co-localization of SOM and cholecystokinin (CCK) or SOM and vasoactive intestinal polypeptide (VIP) was never seen in cell bodies and fibres in any of nuclei studied. In conclusion, SOM-IR neurons of the porcine amygdala form large and heterogeneous subpopulation of, most probably, interneurons that often contain additionally NPY. On the other hand, CCK- and/or VIP-IR neurons belonged to another, discrete subpopulations of porcine CA neurons.     

Lipase-catalyzed regioselective synthesis of steryl (6′-<Emphasis Type="Italic">O</Emphasis>-acyl)glucosides

The regioselective acylation of cholesteryl β-d-glucoside, at the C-6 of the glucose moiety, was achieved using microbial lipases in organic solvents. With palmitic acid as an acyl donor 81 or 63% conversions of cholesteryl glucoside to its 6′-O-palmitoyl derivative were obtained using Candida antarctica or Rhizomucor miehei enzymes, respectively. High yields (64–92%) were also obtained with fatty acids 6:0–22:0 and 16:1 (n-7). The synthesis of cholesteryl (6′-O-palmitoyl)glucoside was also achieved via transesterification, using mono-, di- and tri-palmitoylglycerols or methyl and ethyl palmitate as acyl sources. With R. miehei lipase transesterification between methyl palmitate (80 mM) and cholesteryl glucoside (1 mM) proceeded after 24 h with a nearly quantitative yield (97%).     

Antigens HLA-G, sHLA- G and sHLA- class I in reproductive failure

It can be supposed that relation between HLA-G polymorphism and sHLA-G protein expression are associated with successful embryo implantation and pregnancy maintenance. The aim of the study was the estimation specific differences in expression of sHLA-G and sHLA- class I antigens in women with reproductive failure in comparison with fertile women. The study sample enrolled 80 women, divided into 2 groups. The study group (B) enrolled 60 women with reproductive failure including 20 women with 3 recurrent spontaneous abortions in the first trimester of pregnancy (RSA), 20 women with empty sac (ES) and 20 women with 3 consecutive in-vitro fertilization failures (IVFf). The control group (C) enrolled 20 fertile women with at least 2 children. Soluble HLA- class I antigens (sHLA-I) and soluble HLA-G (sHLA-G) were determined using ELISA test kits from IBio Vendor Labolatory Medicine, Inc. HLA-G allele found in individuals in our study were identified by comparing the obtained bp sequences of exon 2., 3. and 4. with bp sequences of HLA-G antigen published at the Nolan Research Institute website. The highest concentration of sHLA-I is noted among women with HLA-G 10401 allele which differed significantly for the mean sHLA-I concentration calculated for all the remaining alleles (p<0.0001). The most prevalent alleles were: HLA-G 10101, 10102 and 10108 with sHLA-I concentrations among women bearing those alleles significantly lower in comparison to the HLA-G 10401 carriers (p<0.001). Allele 10101 and 10102 was related to the lower significantly plasma sHLA-I concentrations than 10108 allele (p<0.02). Lowest mean sHLA-G values were observed in the IVFf group with significant difference from the remaining groups (p<0.05). To conclude, sHLA-G molecules is associated to certain HLA-G alleles and imply that sHLA-G levels are under genetic control. Low concentration sHLA-G seems to be prognostically important in IVF failure.     

Is the later arrival of young male red-breasted flycatchers (Ficedula parva) related to their physical condition?

Intraspecific variation in the arrival time of migratory birds to breeding grounds is common. Although this phenomenon has been explained in various ways, the condition-dependency of arrival is often evoked. I analyzed the arrival time of male red-breasted flycatchers Ficedula parva, a long-distance migratory passerine, in relation to age, body size and body condition. Data were collected over the course of 5 years in the primeval Białowieża Forest of northeast Poland. Each of these years, young males arrived at the breeding grounds significantly later in the year than did older males. Young males also exhibited significantly shorter wings and lower body condition than older males. Settlement speed was significantly related to age, but not to wing length or body condition. Only the arrival time of old males was related to the body mass and condition. Later arrival of young males could be explained by the lack of experience or by avoidance of both aggression and competition from older males, and both explanations are thought to conserve energy for breeding. Young male red-breasted flycatchers exhibit delayed plumage maturation, and this duller plumage supports the strategy of restraint in the arrival time of young males in this species.     

Common functionally important motions of the nucleotide‐binding domain of Hsp70

The 70 kDa heat shock proteins (Hsp70) are a family of molecular chaperones involved in protein folding, aggregate prevention, and protein disaggregation. They consist of the substrate‐binding domain (SBD) that binds client substrates, and the nucleotide‐binding domain (NBD), whose cycles of nucleotide hydrolysis and exchange underpin the activity of the chaperone. To characterize the structure–function relationships that link the binding state of the NBD to its conformational behavior, we analyzed the dynamics of the NBD of the Hsp70 chaperone from Bos taurus (PDB 3C7N:B) by all‐atom canonical molecular dynamics simulations. It was found that essential motions within the NBD fall into three major classes: the mutual class, reflecting tendencies common to all binding states, and the ADP‐ and ATP‐unique classes, which reflect conformational trends that are unique to either the ADP‐ or ATP‐bound states, respectively. “Mutual” class motions generally describe “in‐plane” and/or “out‐of‐plane” (scissor‐like) rotation of the subdomains within the NBD. This result is consistent with experimental nuclear magnetic resonance data on the NBD. The “unique” class motions target specific regions on the NBD, usually surface loops or sites involved in nucleotide binding and are, therefore, expected to be involved in allostery and signal transmission. For all classes, and especially for those of the “unique” type, regions of enhanced mobility can be identified; these are termed “hot spots,” and their locations generally parallel those found by NMR spectroscopy. The presence of magnesium and potassium cations in the nucleotide‐binding pocket was also found to influence the dynamics of the NBD significantly. Proteins 2015; 83:282–299. © 2014 Wiley Periodicals, Inc.     

Influence of environmental abiotic factors on the content of saponins in plants

Saponins are a large group of secondary metabolites occurring in significant amounts in many plant species. However, the saponin content of plants is variable and it can be influenced by the surrounding environment. The local geoclimate, seasonal changes, external conditions such as light, temperature, humidity and soil fertility, as well as cultivation techniques, affect both the quantitative amount and qualitative composition of saponins. Such variation substantially impacts on the quality and properties of wild and cultivated plants exploited for pharmaceutical, nutritional and industrial applications. This review summarizes the available data on the effects of abiotic environmental factors on saponin level in plants, especially those of considerable economic importance, highlighting current problems such as the reduction in natural plant resources, over-exploitation and destruction of wild habitats, climate shifts as well as the consequences of the growing demand for plant-derived medicinal and industrial products. The need for a theoretical basis for a reasonable harvest, attempts at the domestication of wild plant species and the development of new agricultural technologies allowing high production under optimized conditions are also discussed.     

Identification of sites in the cysteine-rich domain of the class P-III snake venom metalloproteinases responsible for inhibition of platelet function

Atrolysin A and jararhagin are class P-III snake venom metalloproteinases (SVMPs) with three distinct domains: a metalloproteinase, a disintegrin-like and a cysteine-rich. The metalloproteinase and the disintegrin-like domains of atrolysin A and jararhagin contain peptide sequences that interact with alpha2beta1 integrin and inhibit the platelet responses to collagen. Recently, the recombinant cysteine-rich domain of atrolysin A was shown to have similar effects, but the sequence(s) responsible for this is unknown. In this report, we demonstrate two complete peptide sequences from the homologous cysteine-rich domains of atrolysin A and jararhagin that inhibit both platelet aggregation by collagen and adhesion of alpha2-expressing K562 cells to this protein. In addition, the peptide effects on platelets do not seem to involve an inhibition of GPVI. These results identify, for the first time, sites in the cysteine-rich domain of SVMPs that inhibit cell responses to collagen and reveal the complexity of the potential biological effects of these enzymes with multifunctional domains.     

Estimation of Interaction Between Human Keratinocytes and Xenogenic Collagen in vitro

This paper describes the interaction observed between human keratinocytes and xenogenic collagen in vitro modified by HCl. Human keratinocytes were cultivated for 3–10 days, on modified and control support. Their growth, morphology and interaction with support were analyzed. It was found that on both control and experimental (modified) collagen cells proliferated in a similar way. Within 3–10 days, the culture became multilayered and mature and differentiation of cells was visible. Using electron microscope elements of basal membrane interacting with support were seen. On modified support processes of cells penetrating the support are occasionally seen. By use of the immunofluorescent, cytochemical techniques was found the presence of: BP-180 (antigen), β₄ integrin, laminin 5 and collagen IV, VII, VIIc. On the modified support the above listed elements appeared between 3 and 7 days of culture, whereas on the control between 7th and 10th days. On 10th day of culture, the presence of elements of basal membranes became less evident. Results give some hope for using xenogenic, modified collagen as support of keratinocytes culture in process of human skin engineering.