Circadian organization of a subarctic rodent, the northern red-backed vole (Clethrionomys rutilus)

Arctic and subarctic environments are exposed to extreme light: dark (LD) regimes, including periods of constant light (LL) and constant dark (DD) and large daily changes in day length, but very little is known about circadian rhythms of mammals at high latitudes. The authors investigated the circadian rhythms of a subarctic population of northern red-backed voles (Clethrionomys rutilus). Both wild-caught and third-generation laboratory-bred animals showed predominantly nocturnal patterns of wheel running when exposed to a 16:8 LD cycle. In LL and DD conditions, animals displayed large phenotypic variation in circadian rhythms. Compared to wheel-running rhythms under a 16:8 LD cycle, the robustness of circadian activity rhythms decreased among all animals tested in LL and DD (i.e., decreased chi-squared periodogram waveform amplitude). A large segment of the population became noncircadian (60% in DD, 72% in LL) within 8 weeks of exposure to constant lighting conditions, of which the majority became ultradian, with a few individuals becoming arrhythmic, indicating highly labile circadian organization. Wild-caught and laboratory-bred animals that remained circadian in wheel running displayed free-running periods between 23.3 and 24.8 h. A phase-response curve to light pulses in DD showed significant phase delays at circadian times 12 and 15, indicating the capacity to entrain to rapidly changing day lengths at high latitudes. Whether this phenotypic variation in circadian organization, with circadian, ultradian, and arrhythmic wheel-running activity patterns in constant lighting conditions, is a novel adaptation to life in the arctic remains to be elucidated.     

A prevalent POLG CAG microsatellite length allele in humans and African great apes

The human nuclear gene for the catalytic subunit of mitochondrial DNA polymerase (POLG) contains within its coding region a CAG microsatellite encoding a polyglutamine repeat. Previous studies demonstrated an association between length variation at this repeat and male infertility, suggesting a mechanism whereby the prevalent (CAG)₁₀ allele, which occurs at a frequency of >80% in different populations, could be maintained by selection. Sequence analysis of the POLG CAG microsatellite region of more than 1000 human chromosomes reveals that virtually all allelic variation at the locus is accounted for by length variation of the CAG repeat. Analysis of POLG from African great apes shows that a prevalent length allele is present in each species, although its exact length is species-specific. In common chimpanzee (Pan troglodytes) a number of different sequence variants contribute to the prevalent length allele, strongly supporting the idea that the length of the POLG microsatellite region, rather than its exact nucleotide or amino acid sequence, is what is maintained. Analysis of POLG in other primates indicates that the repeat has expanded from a shorter, glutamine-rich sequence, present in the common ancestor of Old and New World monkeys.     

Effects of Acute Lorazepam Administration on Aminergic Activity in Normal Elderly Subjects: Relationship to Performance Effects and Apolipoprotein Genotype

The effects of acute lorazepam challenges on plasma (p) HVA, MHPG, and 5-HIAA, and their relationship to drug-induced cognitive and motor deficits and the apolipoprotein (APOE)-epsilon4 allele were examined. Eighteen healthy elderly (8 epsilon4 carriers) received placebo or acute oral lorazepam doses (0.5 mg or 1 mg) in random sequence, 1-week apart. Cognitive assessment and plasma levels of pHVA, pMHPG, and p5-HIAA were determined at baseline and at 1, 2.5, and 5 h postchallenge. There was no drug-to-placebo difference in monoamine levels and no consistent relationship between changes in monoamine levels and cognitive performance, regardless of epsilon4 status. However, the 1.0 mg dose increased p5-HIAA in epsilon4 carriers, whereas it caused a reduction in noncarriers. Higher baseline pMHPG and p5-HIAA levels were associated with better baseline memory. The epsilon4 allele may modulate the effect of lorazepam on p5-HIAA, but further studies are needed to confirm this finding and elucidate its possible significance.     

Hyperhydricity in micropropagated carnation shoots: the role of oxidative stress

The physiology of hyperhydricity in relation to oxidative stress, mineral nutrients, antioxidant enzymes and ethylene has been studied in three micropropagated carnation cultivars under experimentally induced hyperhydricity. A marked increase in Fe content in comparison with normal tissues was observed in the hyperhydric tissues from the three cultivars. The levels of ethylene, solute leakage and malondialdehyde content were also significantly higher in the hyperhydric tissues. In relation to the time course of H₂O₂ production measured by fluorescence quenching, a similar trend could be observed for the three cultivars, with a clear increase in the generation of hydrogen peroxide in hyperhydric tissues. The activities of all the antioxidative enzymes studied, except lipoxygenase, were higher in the hyperhydric shoots. Phenylalanine ammonia-lyase (PAL) showed a significant decrease in activity in the hyperhydric tissues in comparison with the controls for the three cultivars. Soluble guaiacol peroxidase had a strong increase in activity in hyperhydric shoots of the three cultivars. These results provide, for the first time, direct evidence of H₂O₂ generation in hyperhydric tissues, characterize the response of the antioxidant system to an oxidative stress during hyperhydricity in carnation leaves and point to the accumulation of toxic forms of oxygen as the inducer of some of the abnormalities observed.     

Development of an optical immunosensor based on the fluorescence of Cyanine-5 for veterinarian diagnostics

A model optical immunosensor was developed to quantify an antibody present in a sample by measuring the fluorescence of Cyanine-5 conjugated with the antibody, using a competitive and a sandwich immunoreaction configuration, with the antigen immobilised in controlled pore glass beads. At pH 2, 94% of the antigen-antibody complex was dissociated, allowing reutilisation. Photobleaching had no effect on the fluorescence. This model system was used to detect Brucella sp. infection and could quantify anti-Brucella sp. antibodies in ovine serum samples in the range from 0.005 to 0.11 mg ml(-1).     

TFF1 is membrane-associated in breast carcinoma cell line MCF-7

Trefoil factor family (TFF) domain peptides, products of mucin-secreting epithelial cells, are thought to influence mucosal integrity. Molecular studies revealed that mammalian TFFs lack transmembrane domains. Using immunocytochemistry and FACS analysis we demonstrated the association of TFF1 with the cell membrane in MCF-7 (a breast adenocarcinoma cell line), and tested the hypothesis that glycosylphosphatidylinositol (GPI) linkage is the mechanism for this association. Cleavage of GPI anchorage using phospholipase C did not affect TFF1 binding to the cell membrane. Our results demonstrate for the first time that TFF1 is associated with the cell membrane of MCF-7 cells and is not linked via a GPI anchor.     

Non-Invasive monitoring of diaphragmatic timing by means of surface contact sensors: An experimental study in dogs

Background

The predictive role of many cytokines has not been well defined in Acute Respiratory Distress Syndrome (ARDS).

Methods

We measured prospectively IL-4, IL-6, IL-6 receptor, IL-8, and IL-10, in the serum and bronchoalveolar lavage fluid (BALF) in 59 patients who were admitted to ICU in order to identify predictive factors for the course and outcome of ARDS. The patients were divided into three groups: those fulfilling the criteria for ARDS (n = 20, group A), those at risk for ARDS and developed ARDS within 48 hours (n = 12, group B), and those at risk for ARDS but never developed ARDS (n = 27, group C).

Results

An excellent negative predictive value for ARDS development was found for IL-6 in BALF and serum (100% and 95%, respectively). IL-8 in BALF and IL-8 and IL-10 serum levels were higher in non-survivors in all studied groups, and were associated with a high negative predictive value. A significant correlation was found between IL-8 and APACHE score (r = 0.60, p < 0.0001). Similarly, IL-6 and IL-6r were highly correlated with PaO2/FiO2 (r = -0.27, p < 0.05 and r = -0.55, p < 0.0001, respectively).

Conclusions

BALF and serum levels of the studied cytokines on admission may provide valuable information for ARDS development in patients at risk, and outcome in patients either in ARDS or in at risk for ARDS.     

The human model: a genetic dissection of immunity to infection in natural conditions

Tremendous progress has been achieved in developmental, cellular and molecular immunology in the past 20 years, largely due to studies using the mouse as a model system and the arrival of molecular genetics. Immunology is now faced with a difficult challenge. What are the functions of the individual cells and molecules in achieving immunity to infection? Renewed interest in animal models of disease has provided considerable insight in this area, but such models of infection suffer from the inherent limitation of being experimental. In humans, the complex host-environment interaction occurs in natural, as opposed to experimental, conditions. The human model is therefore an indispensable complement to animal models, as it allows an observational genetic dissection of immunity to infection.     

Two different proteins that compete for binding to thrombin have opposite kinetic and thermodynamic profiles

Thrombin binds thrombomodulin (TM) at anion binding exosite 1, an allosteric site far from the thrombin active site. A monoclonal antibody (mAb) has been isolated that competes with TM for binding to thrombin. Complete binding kinetic and thermodynamic profiles for these two protein-protein interactions have been generated. Binding kinetics were measured by Biacore. Although both interactions have similar K(D)s, TM binding is rapid and reversible while binding of the mAb is slow and nearly irreversible. The enthalpic contribution to the DeltaG(bind) was measured by isothermal titration calorimetry and van't Hoff analysis. The contribution to the DeltaG(bind) from electrostatic steering was assessed from the dependence of the k(a) on ionic strength. Release of solvent H(2)O molecules from the interface was assessed by monitoring the decrease in amide solvent accessibility at the interface upon protein-protein binding. The mAb binding is enthalpy driven and has a slow k(d). TM binding appears to be entropy driven and has a fast k(a). The favorable entropy of the thrombin-TM interaction seems to be derived from electrostatic steering and a contribution from solvent release. The two interactions have remarkably different thermodynamic driving forces for competing reactions. The possibility that optimization of binding kinetics for a particular function may be reflected in different thermodynamic driving forces is discussed.