The thermal properties of some nests of the Eurasian Tree Sparrow Passer montanus

Using an electronic apparatus simulating a bird roosting in a nest at night, we examined the insulating qualities of Eurasian Tree Sparrow (Passer montanus) nests built in nest boxes under winter conditions. Nests of different construction were compared with an empty box, and with roosting in open air. Energy savings in an empty box accounted for 18%, in boxes with incomplete nests for 23% and in boxes with complete nests up to 36%. The insulating value of nests mostly depended on their completeness and the proportion of feathers in the lining.     

Arg-Ile-Tyr (RIY) derived from rapeseed protein decreases food intake and gastric emptying after oral administration in mice

We previously reported that a bioactive tripeptide Arg-Ile-Tyr (RIY), which has been isolated as an inhibitor for angiotensin I-converting enzyme from the subtilisin digest of rapeseed protein, decreased blood pressure. In this study, we also found that RIY dose-dependently decreased food intake at a dose of 150 mg/kg after oral administration in fasted ddY male mice. The anorexigenic action of RIY was blocked by a cholecystokinin-1 CCK1 receptor antagonist, lorglumide. RIY also decreased the gastric emptying rate at a dose of 150 mg/kg and the RIY-induced delay of gastric emptying was blocked by lorglumide. However, RIY had no affinity for CCK1 receptor. Taken together, RIY decreased food intake and gastric emptying by stimulating CCK release.     

Density of Sgt1-immunopositive neurons is decreased in the cerebral cortex of Alzheimer's disease brain

Sgt1 was discovered as a protein required for the mitotic activity of kinetochore and for the activity of ubiquitin ligase in yeast [Kitagawa, K., Skowyra, D., Elledge, S.J., Harper, J.W., Hieter, P., 1999. SGT1 encodes an essential component of the yeast kinetochore assembly pathway and a novel subunit of the SCF ubiquitin ligase complex. Mol. Cell 4, 21-33.]. Later, Sgt1 was identified in different organisms including mammals where it was found at high level in the brain. To understand Sgt1 function in this tissue we analyzed its localization in human brain by immunohistochemistry. In normal brain we observed Sgt1-immunostaining in Purkinje cells of the cerebellum, in granule cells of the dentate gyrus of the hippocampus and in multiple neurons of the cortex. By Western blotting we found a higher level of this protein in the cortex than in the cerebellum. Subsequent morphometric analyses showed that the density of Sgt1-immunopositive neurons varied in different cortical regions. The highest density of Sgt1-immunopositive cells was seen in the temporal cortex (from 1.2% to 5.7%), and the lowest - in the entorhinal cortex (from 0 to 1.1% of all neurons). We next compared the density of Sgt1-immunopositive neurons in cortical layers of healthy aged and Alzheimer's disease (AD) brain sections. A significant decrease in Sgt1-immunopositive neurons was found in the temporal (up to 25-fold), angular (up to 11-fold) and posterior cingulate cortex (up to five-fold). In the entorhinal and precentral cortex the reduction of Sgt1-immunopositive neurons was only about two-fold in AD brains as compared to healthy aged ones. The presence of Sgt1 in post-mitotic neurons indicates the involvement of this protein in a process different from that required for activity of the kinetochore. Decreased immunostaining in AD cortex point to Sgt1 as a possible marker of neurons degenerating in AD.     

Prognostic significance of Ki67-negative blast cell clone in the high risk group of children treated for acute myeloid leukaemia

The aim of this study was to demonstrate the value of immunocytochemical staining of Ki67 antigen expression in blast cells of children with acute myeloid leukemia (AML) and to evaluate its correlation with treatment failure. The material included bone marrow specimens obtained during induction treatment from 46 children treated for AML between 1998-2003. Immunocytochemical staining for Ki67 was based on the ABC technique. Expression of Ki67 antigen on day 0 of induction treatment was confirmed in all patients. The percentage of immunopositive blasts ranged from 88.4% to 99.8% (mean 91.8%). On day 15, according to chemotherapy response, patients were divided into two groups: G1-36 children who responded to induction treatment and reached remission (blast level 5%, low risk group) and G2-10 patients who did not meet remission criterion (blast level > 5%) and were assigned to the high risk (HR) group. Out of 10 children assigned to this group, Ki67 expression in blast cells was confirmed in 4 cases. The fraction of immunopositive blasts ranged from 78.4% to 88.6%. In the other 6 cases, blasts were Ki67-negative. In 12-month period after beginning the treatment, 18 cases of treatment failure (including 7 deceases) were observed in both groups. Five deaths, observed in the HR group, concerned the patients characterized by Ki67-negative blasts. The results indicate a possible correlation between the Ki67-immunonegative blast pattern on day 15 of treatment induction and early decease of AML children assigned to HR group.     

Expression of adenosine receptors in cardiac fibroblasts as a function of insulin and glucose level

Adenosine among other factors is known to regulate the growth and function of cardiac fibroblasts (CFs). Its action is mediated by cell-surface receptors linked to a variety of signaling systems. The goal of present work was to examine the effects of glucose and insulin on adenosine receptors (ARs) mRNA and protein level in primary culture of rat CFs by means of real-time PCR and Western blot. Elevated glucose level increased the expression of A(1)-AR, A(2A)-AR, decreased the expression of A(3)-AR, and had no effect on A(2B)-AR expression. On the other hand insulin suppressed the expression of A(1)-AR, and A(2B)-AR, and had no effect on A(2A)-AR and A(3)-AR expression. Our measurements showed that accumulation of cAMP in response to ARs agonists correlated well with the changes in receptors expression level. These results indicate that changes in glucose and insulin level independently and differentially regulate the ARs expression and functional state in CFs.     

Expression of soluble recombinant TGF-beta type II receptor fused with the Fc portion of human IgG1 (sTbetaRII-Fc) in NS0 cells

We have constructed and expressed recombinant chimeric soluble TGF-beta type II receptor fused with the Fc portion of human IgG1 (sTbetaRII-Fc) in NS0 mouse myeloma cells and isolated cell lines constitutively secreting very high levels of biologically active protein. The GS-NS0 expression system takes advantage of the strong human cytomegalovirus immediate early promoter expression vector and glutamine synthetase as a selectable marker. The recombinant chimeric receptor could be produced in high amounts and efficiently purified by one step chromatography on a protein A column. Biochemical studies revealed that recombinant sTbetaRII-Fc binds native TGF-beta1 and TGF-beta3 isoforms and neutralizes their activity in vitro.     

Pyridine-2,6-Bis(Thiocarboxylic Acid) Produced by Pseudomonas stutzeri KC Reduces and Precipitates Selenium and Tellurium Oxyanions

The siderophore of Pseudomonas stutzeri KC, pyridine-2,6-bis(thiocarboxylic acid) (pdtc), is shown to detoxify selenium and tellurium oxyanions in bacterial cultures. A mechanism for pdtc's detoxification of tellurite and selenite is proposed. The mechanism is based upon determination using mass spectrometry and energy-dispersive X-ray spectrometry of the chemical structures of compounds formed during initial reactions of tellurite and selenite with pdtc. Selenite and tellurite are reduced by pdtc or its hydrolysis product H₂S, forming zero-valent pdtc selenides and pdtc tellurides that precipitate from solution. These insoluble compounds then hydrolyze, releasing nanometer-sized particles of elemental selenium or tellurium. Electron microscopy studies showed both extracellular precipitation and internal deposition of these metalloids by bacterial cells. The precipitates formed with synthetic pdtc were similar to those formed in pdtc-producing cultures of P. stutzeri KC. Culture filtrates of P. stutzeri KC containing pdtc were also active in removing selenite and precipitating elemental selenium and tellurium. The pdtc-producing wild-type strain KC conferred higher tolerance against selenite and tellurite toxicity than a pdtc-negative mutant strain, CTN1. These observations support the hypothesis that pdtc not only functions as a siderophore but also is involved in an initial line of defense against toxicity from various metals and metalloids.     

High-affinity binding of the NC1 domain of collagen VII to laminin 5 and collagen IV

Anchoring functions of collagen VII depend on its ability to form homotypic fibrils and to bind to other macromolecules to form heterotypic complexes. Biosensor-based binding assays were employed to analyze the kinetics of the NC1 domain-mediated binding of collagen VII to laminin 5, collagen IV, and collagen I. We showed that collagen VII interacts with laminin 5 and collagen IV with a Kd value of 10(-9) M. In contrast, the NC1-mediated binding to collagen I was weak with a Kd value of 10(-6) M. Binding assays also showed that the NC1 domain utilizes the same region to bind to both laminin 5 and collagen IV. We postulate that the ability of the NC1 domains to bind with high affinities to laminin 5 and collagen IV facilitates stabilization of the structure of the basement membrane itself and that the NC1-collagen I interaction may be less important for stabilization of the dermal-epidermal junction.     

Verteporfin, photofrin II, and merocyanine 540 as PDT photosensitizers against melanoma cells

The efficiency of photodynamic effect (PDE) for Photofrin II (PfII), Verteporfin, and Merocyanine 540 (MC540) was compared against neoplastic cells. Triplet state lifetimes and singlet molecular oxygen quantum yields were correlated with biological effect. PfII triplet lifetime was two times longer than that of Verteporfin, however, its singlet molecular oxygen quantum yield was two times lower in comparison with Verteporfin. High singlet molecular oxygen quantum yield of Verteporfin resulted in high biological efficacy. To achieve 50% mortality of cells four times lower light dose and five times lower concentration of Verteporfin were applied in comparison with PfII. The same level of cell damage was reached using 10 times higher light dose and two times higher concentration of MC540 in comparison with PfII. Our results confirm that singlet molecular oxygen based mechanism, prevalent for Verteporfin and PfII, was highly effective against melanoma cells. Verteporfin can be used at small doses with high cellular damage efficiency.     

Nucleic acid-binding properties of the RRM-containing protein RDM1

RDM1 (RAD52 Motif 1) is a vertebrate protein involved in the cellular response to the anti-cancer drug cisplatin. In addition to an RNA recognition motif, RDM1 contains a small amino acid motif, named RD motif, which it shares with the recombination and repair protein, RAD52. RDM1 binds to single- and double-stranded DNA, and recognizes DNA distortions induced by cisplatin adducts in vitro. Here, we have performed an in-depth analysis of the nucleic acid-binding properties of RDM1 using gel-shift assays and electron microscopy. We show that RDM1 possesses acidic pH-dependent DNA-binding activity and that it binds RNA as well as DNA, and we present evidence from competition gel-shift experiments that RDM1 may be capable of discrimination between the two nucleic acids. Based on reported studies of RAD52, we have generated an RDM1 variant mutated in its RD motif. We find that the L119GF --> AAA mutation affects the mode of RDM1 binding to single-stranded DNA.