An Investigation of Freezing of Supercooled Water on Anti-Freeze Protein Modified Surfaces

This work investigates how functionalization of aluminium surfaces with natural type III Anti-Freeze Protein (AFP) affects the mechanism of heterogeneous ice nucleation. First the bulk ice nucleation properties of distilled water and aqueous solution of AFP were evaluated by differential scanning calorimetry. Then the modified surface was characterized by Secondary Ions Mass Spectroscopy (SIMS), Fourier Transform InfraRed (FTIR) spectroscopy and contact angle measurement. Freezing experiments were then conducted in which water droplets underwent a slow controlled cooling. This study shows that compared to uncoated aluminium, the anti-freeze proteins functionalized surfaces exhibit a higher and narrower range of freezing temperature. It was found that these proteins that keep living organisms from freezing in cold environment act in the opposite way once immobilized on surfaces by promoting ice nucleation. Some suggestions regarding the mechanism of action of the observed phenomena were proposed based on the Classical Nucleation Theory (CNT).     

On the influence of prenatal hypoxia on formation of the orexinergic system and sleep–wake cycle in early ontogenesis of rats

The role of orexin in the organization of the sleep–wake cycle (SWC) is well known. The aim of this study was to examine the timing of the orexinergic system formation in rat postnatal ontogenesis and to assess the role of orexin A in the SWC organization under normal conditions and after prenatal hypoxia undergone on days 14 and 19 of embryogenesis. The SWC was investigated in 30-day-old rats with electrodes implanted into the somatosensory and occipital cortex. Immunoreactivity within the orexigenic structures of the lateral hypothalamus was analyzed. It was shown that in control 14-day-old animals the orexinergic structures were in their formative stage, whereas in 30-day-old rats they were already as formed as in adults. In 14-day-old rats, prenatal hypoxia evoked retarded formation of the orexinergic system. In 30-day-old animals, hypoxia undergone in the prenatal period increased the activity of the orexinergic system, which was higher in animals exposed to hypoxia on day 19 than on day 14 of gestation. In 30-day-old rats, these changes were reflected in the SWC formation in the form of shorter slow-wave sleep, more fitful sleep and increased number of transitions from slow- to fast-wave sleep. The results obtained are discussed in the light of the adaptive-compensatory role of the orexigenic system in postnatal ontogenesis after prenatal damage to the central nervous system.     

Characterisation of antioxidants in photosynthetic and non‐photosynthetic leaf tissues of variegated Pelargonium zonale plants

Hydrogen peroxide is an important signalling molecule, involved in regulation of numerous metabolic processes in plants. The most important sources of H₂O₂ in photosynthetically active cells are chloroplasts and peroxisomes. Here we employed variegated Pelargonium zonale to characterise and compare enzymatic and non‐enzymatic components of the antioxidative system in autotrophic and heterotrophic leaf tissues at (sub)cellular level under optimal growth conditions. The results revealed that both leaf tissues had specific strategies to regulate H₂O₂ levels. In photosynthetic cells, the redox regulatory system was based on ascorbate, and on the activities of thylakoid‐bound ascorbate peroxidase (tAPX) and catalase. In this leaf tissue, ascorbate was predominantly localised in the nucleus, peroxisomes, plastids and mitochondria. On the other hand, non‐photosynthetic cells contained higher glutathione content, mostly located in mitochondria. The enzymatic antioxidative system in non‐photosynthetic cells relied on the ascorbate–glutathione cycle and both Mn and Cu/Zn superoxide dismutase. Interestingly, higher content of ascorbate and glutathione, and higher activities of APX in the cytosol of non‐photosynthetic leaf cells compared to the photosynthetic ones, suggest the importance of this compartment in H₂O₂ regulation. Together, these results imply different regulation of processes linked with H₂O₂ signalling at subcellular level. Thus, we propose green‐white variegated leaves as an excellent system for examination of redox signal transduction and redox communication between two cell types, autotrophic and heterotrophic, within the same organ.     

Diencephalo-telencephalic changes of tyrosine hydroxylase in rats and grass frogs after sleep deprivation

Based on sleep deprivation-produced changes of electrographic parameters of the wakefulness-sleep cycle (WSC) in rats and frogs (Rana temporaria), dynamics of activity of tyrosine hydroxylase, the key enzyme of dopamine synthesis, was studied immunohistochemically in substantia nigra and nigrostriatal pathway in rats and in striatum, paraventricular organ, and extrahypothalamic pathways in frogs. Changes in dynamics of tyrosine hydroxylase in rats and in frogs are revealed after the 6-h sleep deprivation and after 2 h of postdeprivation sleep. This allows determining the degree of participation of corticostriatal neuroregulatory and hypothalamo-pituitary neurosecretory systems and their role in regulation of WSC. Possible evolutionary peculiarities of morphofunctional differences in homoiothermal and poikilothermal animals are discussed.     

Long-term impact of war on healthcare costs: an eight-country study

Objective

Exposure to war can negatively affect health and may impact on healthcare costs. Estimating these costs and identifying their predictors is important for appropriate service planning. We aimed to measure use of health services in an adult population who had experienced war in the former-Yugoslavia on average 8 years previously, and to identify characteristics associated with the use and costs of healthcare.

Method

War-affected community samples in Bosnia-Herzegovina, Croatia, Kosovo, FYR Macedonia, and Serbia were recruited through a random walk technique. Refugees in Germany, Italy and the UK were contacted through registers, organisations and networking. Current service use was measured for the previous three months and combined with unit costs for each country for the year 2006/7. A two-part approach was used, to identify predictors of service use with a multiple logistic regression model and predictors of cost with a generalised linear regression model.

Results

3,313 participants were interviewed in Balkan countries and 854 refugees in Western European countries. In the Balkan countries, traumatic events and mental health status were related to greater service use while in Western countries these associations were not found. Participants in Balkan countries with post traumatic stress disorder (PTSD) had costs that were 63% higher (p = 0.005) than those without PTSD. Distress experienced during the most traumatic war event was associated with higher costs (p = 0.013). In Western European countries costs were 76% higher if non-PTSD anxiety disorders were present (0.027) and 63% higher for mood disorders (p = 0.006).

Conclusions

War experiences and their effects on mental health are associated with increased health care costs even many years later, especially for those who stayed in the area of conflict. Focussing on the mental health impact of war is important for many reasons including those of an economic nature.     

Molecular analysis of carbohydrate N-acetylgalactosamine 4-O sulfotransferase 8 (CHST8) as a candidate gene for bovine spongiform encephalopathy susceptibility

Endogenous prion proteins (PrP) play the central role in the pathogenesis of transmissible spongiform encephalopathies. The carbohydrate N -acetylgalactosamine 4-O sulfotransferase 8 (CHST8) promotes the conversion of the cellular PrP^C into the pathogenic PrP^d. Six sequence variants within the CHST8 gene were identified by comparative sequencing and genotyped for a sample of 623 animals comprising bovine spongiform encephalopathy (BSE)-affected and healthy control cows representing German Fleckvieh (German Simmental), German Holstein (Holstein-Friesian) and Brown Swiss. Significant differences in the allele, genotype and haplotype frequencies between BSE-affected and healthy cows indicate an association of sequence variant g.37254017G>T with the development of the disease in Brown Swiss cattle.     

Zinc‐induced oxidative stress in Verbascum thapsus is caused by an accumulation of reactive oxygen species and quinhydrone in the cell wall

Oxidative stress is one aspect of metal toxicity. Zinc, although unable to perform univalent oxido‐reduction reactions, can induce the oxidative damage of cellular components and alter antioxidative systems. Verbascum thapsus L. plants that were grown hydroponically were exposed to 1 and 5 mM Zn²⁺. Reactive oxygen species (ROS) accumulation was demonstrated by the fluorescent probe H₂DCFDA and EPR measurements. The extent of zinc‐induced oxidative damage was assessed by measuring the level of protein carbonylation. Activities and isoform profiles of some antioxidant enzymes and the changes in ascorbate and total phenolic contents of leaves and roots were determined. Stunted growth because of zinc accumulation, preferentially in the roots, was accompanied by H₂O₂ production in the leaf and root apoplasts. Increased EPR signals of the endogenous oxidant quinhydrone, ^?CH₃ and ^?OH, were found in the cell walls of zinc‐treated plants. The activities of the antioxidative enzymes ascorbate peroxidase (APX) (EC 1.11.1.11), soluble superoxide dismutase (SOD) (EC 1.15.1.1), peroxidase (POD), (EC 1.11.1.7) and monodehydroascorbate reductase (EC 1.6.5.4) were increased; those of glutathione reductase (EC 1.6.4.2), dehydroascorbate reductase (EC 1.8.5.1) and ascorbate oxidase (AAO) (EC 1.10.3.3) were decreased with zinc treatment. Zinc induced a cell‐wall‐bound SOD isoform in both organs. Leaves accumulated more ascorbate and phenolics in comparison to roots. We propose a mechanism for zinc‐promoted oxidative stress in V. thapsus L. through the generation of charge transfer complexes and quinhydrone because of phenoxyl radical stabilisation by Zn²⁺ in the cell wall. Our results suggest that the SOD and APX responses are mediated by ROS accumulation in the apoplast. The importance of the POD/Phe/AA (ascorbic acid) scavenging system in the apoplast is also discussed.     

Orexin-Immunopositive Structures in the Thalamic Reticular Nucleus

Journal of Evolutionary Biochemistry and Physiology - Orexin-A and orexin-B were immunolocalized in laboratory rodents (adult Wistar and Sprague–Dawley rats, C57Bl/6J mice) in neurons of...     

Engineering the respiratory complex I to energy-converting NADPH:ubiquinone oxidoreductase

The respiratory complex I couples the electron transfer from NADH to ubiquinone with a translocation of protons across the membrane. Its nucleotide-binding site is made up of a unique Rossmann fold to accommodate the binding of the substrate NADH and of the primary electron acceptor flavin mononucleotide. Binding of NADH includes interactions of the hydroxyl groups of the adenosine ribose with a conserved glutamic acid residue. Structural analysis revealed that due to steric hindrance and electrostatic repulsion, this residue most likely prevents the binding of NADPH, which is a poor substrate of the complex. We produced several variants with mutations at this position exhibiting up to 200-fold enhanced catalytic efficiency with NADPH. The reaction of the variants with NAD(P)H is coupled with proton translocation in an inhibitor-sensitive manner. Thus, we have created an energy-converting NADPH:ubiquinone oxidoreductase, an activity so far not found in nature. Remarkably, the oxidation of NAD(P)H by the variants leads to an enhanced production of reactive oxygen species.