Post-hatching dynamics of plasma biochemistry in free-living European starlings (Sturnus vulgaris)

This is the first study of plasma biochemical parameters in free-living altricial birds during an entire developmental period in a nest, represented by European starling (Sturnus vulgaris). Dynamics of postnatal changes from hatching until close to fledging (days 1 to 15) were registered. Parameters of protein metabolism represented by total proteins, albumin and globulin concentrations increased continuously during the observed developmental period. There were two peaks in uric acid concentration on days 5 and 11. To the contrary, the creatinine content did not change throughout the observed period and increased only on day 15. Creatine kinase activity gradually increased until day 11 and then fell before fledging. Parameters of lipid metabolism (concentration of total lipids, triacylglycerols and nonesterified fatty acids) in plasma increased gradually reaching a plateau between days 8 and 11 and then declined on day 15. The cholesterol concentration pattern was similar to maximum value on day 11, then consecutively decreased. Concentration of glucose increased until day 8 and remained unchanged until fledging. Whereas calcium reached the highest concentration during days 8 and 11, phosphorus peaked earlier on day 5. The activity of alkaline phosphatase was similar to the pattern found in calcium concentration. Presented data showed an increase in both protein and lipid metabolism during the phase of rapid growth. A remarkable decrease in parameters of lipid metabolism before fledging may reflect increased physical activity and changes in nutrition.     

The structure of DNA-DOPC aggregates formed in presence of calcium and magnesium ions: A small-angle synchrotron X-ray diffraction study

The structure of aggregates formed due to DNA interaction with dioleoylphosphatidylcholine (DOPC) vesicles in presence of Ca²⁺ and Mg²⁺ cations was investigated using synchrotron small-angle X-ray diffraction. For DOPC/DNA = 1:1 mol/base and in the range of concentration of the cation²⁺ 0-76.5 mM, the diffractograms show the coexistence of two lamellar phases: L^x phase with repeat distance d_Lx ∼ 8.26-7.39 nm identified as a phase where the DNA strands are intercalated in water layers between adjacent lipid bilayers, and L_DOPC phase with repeat distance d_DOPC ∼ 6.45-5.65 nm identified as a phase of partially dehydrated DOPC bilayers without any divalent cations and DNA strands. The coexistence of these phases was investigated as a function of DOPC/DNA molar ratio, length of DNA fragments and temperature. If the amount of lipid increases, the fraction of partially dehydrated L_DOPC phase is limited, depends on the portion of DNA in the sample and also on the length of DNA fragments. Thermal behaviour of DOPC + DNA + Ca²⁺ aggregates was investigated in the range 20-80 °C. The transversal thermal expansivities of both phases were evaluated.     

Synthesis of the tetrasaccharide side chain of the major glycoprotein of the Bacillus anthracis exosporium

An alpha-glycoside of the tetrasaccharide sequence beta-Ant-(1-->3)-alpha-l-Rhap-(1-->3)-alpha-l-Rhap-(1-->2)-alpha-l-Rhap whose aglycon allows conjugation to suitable carriers was synthesized. The NMR characteristics of the compound are virtually identical with those of the alpha-anomer of the tetrasaccharide isolated from the major glycoprotein of the Bacillus anthracis exosporium. Thus, the correct structure of the natural product has been proven by chemical synthesis.     

Molecular recognition in the P2Y14 receptor: Probing the structurally permissive terminal sugar moiety of uridine-5′-diphosphoglucose

The P2Y₁₄ receptor, a nucleotide signaling protein, is activated by uridine-5′-diphosphoglucose 1 and other uracil nucleotides. We have determined that the glucose moiety of 1 is the most structurally permissive region for designing analogues of this P2Y₁₄ agonist. For example, the carboxylate group of uridine-5′-diphosphoglucuronic acid proved to be suitable for flexible substitution by chain extension through an amide linkage. Functionalized congeners containing terminal 2-acylaminoethylamides prepared by this strategy retained P2Y₁₄ activity, and molecular modeling predicted close proximity of this chain to the second extracellular loop of the receptor. In addition, replacement of glucose with other sugars did not diminish P2Y₁₄ potency. For example, the [5′′]ribose derivative had an EC₅₀ of 0.24 μM. Selective monofluorination of the glucose moiety indicated a role for the 2′′- and 6′′-hydroxyl groups of 1 in receptor recognition. The β-glucoside was twofold less potent than the native α-isomer, but methylene replacement of the 1′′-oxygen abolished activity. Replacement of the ribose ring system with cyclopentyl or rigid bicyclo[3.1.0]hexane groups abolished activity. Uridine-5′-diphosphoglucose also activates the P2Y₂ receptor, but the 2-thio analogue and several of the potent modified-glucose analogues were P2Y₁₄-selective.     

Similarities and distinctions in Y chromosome gene pool of Western Slavs

Analysis of Y chromosome Y‐STRs has proven to be a useful tool in the field of population genetics, especially in the case of closely related populations. We collected DNA samples from 169 males of Czech origin, 80 males of Slovakian origin, and 142 males dwelling Northern Poland. We performed Y‐STR analysis of 12 loci in the samples collected (PowerPlex Y system from Promega) and compared the Y chromosome haplotype frequencies between the populations investigated. Also, we used Y‐STR data available from the literature for comparison purposes. We observed significant differences between Y chromosome pools of Czechs and Slovaks compared to other Slavic and European populations. At the same time we were able to point to a specific group of Y‐STR haplotypes belonging to an R1a haplogroup that seems to be shared by Slavic populations dwelling in Central Europe. The observed Y chromosome diversity may be explained by taking into consideration archeological and historical data regarding early Slav migrations. Am J Phys Anthropol 142:540–548, 2010. © 2010 Wiley‐Liss, Inc.     

Knowledge and Perceptions of,and Attitudes to,Bats by People Living around Arabuko-Sokoke Forest,Malindi-Kenya

Bat populations continue to decline worldwide because of myriad human activities. To enhance bat conservation, human behavior needs to change. Such change can occur, in part, through an understanding of what motivates human actions toward bats. We used a Bat Attitude Questionnaire (BAQ) to investigate attitudes toward bats in people (n = 394) living around Arabuko-Sokoke Forest (ASF), Kenya. Belief in myths seemed to prevail among those surveyed and that these myths were significantly associated with low tolerance of bats (ordinal regression, p < 0.05). Older and more educated people reported more positive attitudes toward bats than others. Multiple linear regression revealed that females showed more negative attitudes toward, and more belief in, myths about bats than males (both p < 0.01). Ordinal regression showed that hostile behavior toward bats was more common among males (p < 0.001). Nearly one-third of the respondents reported actively killing bats or destroying bat roosts. A similar proportion did not see any benefits of bats to humans, while the majority of respondents associated bats with the destruction of farmers’ fruits. To address prevailing negative attitudes about bats, we recommend conducting evening school-based bat study tours, specifically targeting youths in schools around the forest, to examine whether physical contacts with bats would help positively influence human attitudes toward bats. Additionally, since mango (Mangifera indica) farming is an important source of income to people around ASF, an assessment of the quantity of mangoes destroyed by bats would be important to determine whether there is a need for controlling bat access to crops through appropriate schemes.     

Impact of mitochondrial beta-oxidation in fatty acid-mediated inhibition of glioma cell proliferation

Tetradecylthioacetic acid (TTA), which cannot be beta-oxidized, exerts growth-limiting properties in glioma cells. In order to investigate the importance of modulated lipid metabolism and alterations in mitochondrial properties in this cell death process, we incubated glioma cells both with TTA and the oxidizable fatty acid palmitic acid (PA), in the presence of L-carnitine and the carnitine palmitoyltransferase inhibitors etomoxir and aminocarnitine. L-carnitine partly abolished the PA-mediated growth reduction of glioma cells, whereas etomoxir and aminocarnitine enhanced the antiproliferative effect of PA. The production of acid-soluble products increased and the incorporation of PA into glycerolipids decreased after L-carnitine supplementation. L-carnitine was found to enhance the antiproliferative effect of TTA, but did not affect the incorporation of TTA into glycerolipids, or ceramide. PDMP, sphingosine 1-phosphate, desipramine, fumonisin B(1), and L-cycloserine were able not to rescue the glioma cells from PA and TTA-induced growth inhibition, suggesting that increased ceramide production is not important in the growth reduction. TTA-mediated growth inhibition was accompanied with an increased uptake of PA and increased incorporation of PA into triacylglycerol (TG). Our data suggest that mitochondrial functions are involved in fatty acid-mediated growth inhibition. Whether there is a causal relationship between TG accumulation and the apoptotic process remains to be determined.     

Induced cell death of preimplantation mouse embryos cultured in vitro evaluated by comet assay

The occurrence of apoptosis in mouse preimplantation embryos was analyzed using DNA staining (Hoechst 33342, PI) for the visualization of nuclear changes and by the comet assay, a single-cell gel electrophoresis assay, modified for the analysis of blastocysts. Mouse preimplantation embryos isolated 56 h after superovulation were cultured in vitro for 64 h. Apoptosis was induced by treatment with camptothecin and actinomycin D during the first 15 h of culture. After culture in vitro, a number of embryos were stained and analyzed using morphological criteria. The remaining embryos were examined using the comet assay for the detection of DNA fragmentation. The proportion of damaged embryos in experimental groups, in comparison to controls, was dependent on the dose of apoptosis inductor. At high doses (camptothecin, microg/ml and actinomycin D, 0.05 microg/ml) over 90% (chi-square test, P<0.001) of embryos had apoptotic comets, at medium doses (camptothecin, 0.01 microg/ml and actinomycin D, 0.005 microg/ml) comets appeared only in 30-70% of embryos (camptothecin, P<0.01 and actinomycin D, P<0.001). At low doses (camptothecin, 0.001 microg/ml and actinomycin D, 0.0005 microg/ml) the increase in damaged embryos was not statistically significant. Hoechst/PI staining showed a higher percentage of damaged blastomeres at high doses. Morphological changes correlated with the outcome of the comet assay. Our results show that comet assay is an appropriate method for studying apoptosis in preimplantation embryos, and it appears to be more sensitive than the classically used morphological analyses.     

Effects of non-ionic surfactants N-alkyl-N,N-dimethylamine-N-oxides on the structure of a phospholipid bilayer: small-angle X-ray diffraction study

Effects of non-ionic surfactants N-alkyl-N,N-dimethylamine-N-oxides (C(n)NO, n is the number of alkyl carbons) on the structure of egg yolk phosphatidylcholine (EYPC) bilayers in the lamellar fluid phase was studied by small-angle X-ray diffraction as a function of H(2)O:EYPC and C(n)NO:EYPC molar ratios. The bilayer thickness d(L) and the lipid surface area at the bilayer-aqueous interface S(L) were calculated from the repeat period, d of the lamellar phase, based on the model that water and EYPC + CnNO molecules form separated layers and that their molecular volumes are additive. In the studied range of m=CnNO:EYPC molar ratios up to 1:1, d(L) and S(L) change linearly. The slopes Delta L = delta dL/ delta m and Delta S= delta S L / delta m are equal to -0.876 +/- 0.027 nm and 0.347 +/- 0.006 nm2 for C(6)NO, -1.025+/-0.060 nm and 0.433+/-0.025 nm(2) for C(8)NO, -0.836+/-0.046 nm and 0.405+/-0.018 nm(2) for C(10)NO, -0.604+/-0.015 nm and 0.375+/-0.007 nm(2) for C(12)NO, -0.279+/-0.031 nm and 0.318+/-0.005 nm(2) for C(14)NO, -0.0865+/-0.070 nm and 0.2963 +/-0.014 nm(2) for C(16)NO, and -0.040+/-0.022 nm and 0.297+/- 0.002 nm(2) for C(18)NO, respectively, at full bilayer hydration. The peak-peak distance in the bilayer electron density profile, which relates to the P-P distance d(PP), obtained from the first four diffraction peaks by the Fourier transform also depends linearly on m, and the slope Delta PP = delta dPP/delta m is -0.528+/-0.065 nm for C(6)NO, -0.680+/-0.018 nm for C(8)NO, -0.573+/-0.021 nm for C(10)NO, -0.369+/-0.075 nm for C(12)NO, -0.190+/-0.015 for C(14)NO, -0.088+/-0.016 nm for C(16)NO and -0.094+/-0.016 nm for C(18)NO. The effects of C(n)NO on Delta(L), Delta(S) and Delta(PP) are the results of C(n)NO insertion into EYPC bilayers and depend on the hydrophobic mismatch between C(n)NO and EYPC hydrocarbon chains and on the lateral interactions of C(n)NO and EYPC in the bilayer.     

In vitro identification of mitochondrial oxidative stress production by time-resolved fluorescence imaging of glioma cells

Oxidative phosphorylation and glycolysis are important features, by which cells could bypass oxidative stress. The level of oxidative stress, and the ability of cells to promote oxidative phosphorylation or glycolysis, significantly determined proliferation or cell demise. In the present work, we have employed selective mitochondrial probe MitoTracker? Orange CMTM/Ros (MTO) to estimate the level of oxidative stress in cancer cells at different stressed conditions. MTO is partially sensitive to decrease of mitochondrial membrane potential and to reactive oxygen species (ROS) generated in mitochondria. We have demonstrated, that fluorescence lifetime of MTO is much more sensitive to oxidative stress than intensity-based approaches. This method was validated in different cancer cell lines. Our approach revealed, at relatively low ROS levels, that Gö 6976, a protein kinase C (PKC) α inhibitor, and rottlerin, an indirect PKCδ inhibitor, increased mitochondrial ROS level in glioma cell. Their involvement in oxidative phosphorylation and apoptosis was investigated with oxygen consumption rate estimation, western blot and flow-cytometric analysis. Our study brings new insight to identify feeble differences in ROS production in living cells.