Acoustic Features Distinguishing Emotions in Swedish Speech.

Few studies have examined which acoustic features of speech can be used to distinguish between different emotions, and how combinations of acoustic parameters contribute to identification of emotions. The aim of the present study was to investigate which acoustic parameters in Swedish speech are most important for differentiation between, and identification of, the emotions anger, fear, happiness, sadness, and surprise in Swedish sentences. One-way ANOVAs were used to compare acoustic parameters between the emotions and both simple and multiple logistic regression models were used to examine the contribution of different acoustic parameters to differentiation between emotions. Results showed differences between emotions for several acoustic parameters in Swedish speech: surprise was the most distinct emotion, with significant differences compared to the other emotions across a range of acoustic parameters, while anger and happiness did not differ from each other on any parameter. The logistic regression models showed that fear was the best-predicted emotion while happiness was most difficult to predict. Frequency- and spectral-balance-related parameters were best at predicting fear. Amplitude- and temporal-related parameters were most important for surprise, while a combination of frequency-, amplitude- and spectral balance-related parameters are important for sadness. Assuming that there are similarities between acoustic models and how listeners infer emotions in speech, results suggest that individuals with hearing loss, who lack abilities of frequency detection, may compared to normal hearing individuals have difficulties in identifying fear in Swedish speech. Since happiness and fear relied primarily on amplitude- and spectral-balance-related parameters, detection of them are probably facilitated more by hearing aid use.

A nationwide cohort study of the risk of chronic obstructive pulmonary disease in coeliac disease.

OBJECTIVE: Chronic obstructive pulmonary disease (COPD) continues to be an important cause of morbidity, mortality and healthcare costs in the western world. Although smoking is an important trigger of COPD, other factors such as chronic inflammation and malnutrition are known to influence its development. Because coeliac disease (CD) is characterized both by dysregulated inflammation and malnutrition, the possibility of an association between CD and COPD was investigated. METHODS: Through biopsy data from all Swedish pathology departments, we identified 10 990 individuals with CD who were biopsied between 1987 and 2008 (Marsh 3: villous atrophy). As controls, 54 129 reference individuals matched for age, sex, county and calendar year of first biopsy were selected. Cox regression analysis was then performed to estimate hazard ratios (HRs) for having a diagnosis of COPD according to the Swedish Patient Register. RESULTS: During follow-up, 380 individuals with CD (3.5%) and 1391 (2.6%) controls had an incident diagnosis of COPD, which corresponds to an HR of 1.24 (95% CI: 1.10-1.38) and an excess risk of COPD of 79/100 000 person-years in CD. The risk increase remained 5 years after biopsy (HR = 1.17; 95% CI: 1.00-1.37). Risk estimates did not change with adjustment for type 1 diabetes, thyroid disease, rheumatoid arthritis, country of birth or level of education. Men with CD were at a higher risk of COPD (HR = 1.39; 95% CI: 1.18-1.62) than women with CD (HR = 1.11; 95% CI: 0.94-1.30). Of note, CD was also associated with COPD before CD diagnosis (odds ratio = 1.22; 95% CI: 1.02-1.46). Conclusion. Patients with CD seem to be at a moderately increased risk of COPD both before and after CD diagnosis.

Recombinant protein expression and solubility screening in Escherichia coli: a comparative study.

Producing soluble proteins in Escherichia coli is still a major bottleneck for structural proteomics. Therefore, screening for soluble expression on a small scale is an attractive way of identifying constructs that are likely to be amenable to structural analysis. A variety of expression-screening methods have been developed within the Structural Proteomics In Europe (SPINE) consortium and to assist the further refinement of such approaches, eight laboratories participating in the network have benchmarked their protocols. For this study, the solubility profiles of a common set of 96 His(6)-tagged proteins were assessed by expression screening in E. coli. The level of soluble expression for each target was scored according to estimated protein yield. By reference to a subset of the proteins, it is demonstrated that the small-scale result can provide a useful indicator of the amount of soluble protein likely to be produced on a large scale (i.e. sufficient for structural studies). In general, there was agreement between the different groups as to which targets were not soluble and which were the most soluble. However, for a large number of the targets there were wide discrepancies in the results reported from the different screening methods, which is correlated with variations in the procedures and the range of parameters explored. Given finite resources, it appears that the question of how to most effectively explore ;expression space' is similar to several other multi-parameter problems faced by crystallographers, such as crystallization.

Laser-triggered high-voltage plasma switching with diffractive optics.

High-power lasers can be used to induce ionization of gases and thereby enable rapid triggering of electrical discharge devices, potentially faster than any devices based on mechanical or solid-state switching. With diffractive optical elements (DOEs) the laser light can conveniently be directed to positions within the gas so that an electrical discharge between two high-voltage electrodes is triggered reliably and rapidly. Here we report on two different types of DOE used for creating an electrical discharge in pure argon for potential high-voltage applications. One is the diffractive equivalent of a conventional axicon that yields an extended, and continuous, high-intensity focal region between the electrodes. The other is a multiple-focal-distance kinoform--a DOE that is designed to produce a linear array of 20 discrete foci, with high peak intensities, between the electrodes. We show that DOEs enable efficient, rapid switching and may provide increased flexibility in the design of novel electrode configurations.

Beta-cell apoptosis in an accelerated model of autoimmune diabetes.

BACKGROUND: The non-obese diabetic (NOD) mouse is a model of human type 1 diabetes in which autoreactive T cells mediate destruction of pancreatic islet beta cells. Although known to be triggered by cytotoxic T cells, apoptosis has not been unequivocally localized to beta cells in spontaneously diabetic NOD mice. We created a model of accelerated beta-cell destruction mediated by T cells from spontaneously diabetic NOD mice to facilitate the direct detection of apoptosis in beta cells. MATERIALS AND METHODS: NOD.scid (severe combined immunodeficiency) mice were crossed with bm1 mice transgenically expressing the costimulatory molecule B7-1 (CD80) in their beta cells, to generate B7-1 NOD.scid mice. Apoptosis in islet cells was measured as DNA strand breakage by the TdT-mediated-dUTP-nick end labeling (TUNEL) technique. RESULTS: Adoptive transfer of splenocytes from spontaneously diabetic NOD mice into B7-1 NOD.scid mice caused diabetes in recipients within 12-16 days. Mononuclear cell infiltration and apoptosis were significantly greater in the islets of B7-1 NOD.scid mice than in nontransgenic NOD.scid mice. Dual immunolabeling for TUNEL and either B-7 or insulin, or the T cell markers CD4 and CD8, and colocalization by confocal microscopy clearly demonstrated apoptosis in beta cells as well in a relatively larger number of infiltrating T cells. The clearance time of apoptotic beta cells was estimated to be less than 6 min. CONCLUSIONS: B7-1 transgenic beta cells undergo apoptosis during their accelerated destruction in response to NOD mouse effector T cells. Rapid clearance implies that beta cells undergoing apoptosis would be detected only rarely during more protracted disease in spontaneously diabetic NOD mice.

Preserved catalytic activity in an engineered ribonucleotide reductase R2 protein with a nonphysiological radical transfer pathway. The importance of hydrogen bond connections between the participating residues.

A hydrogen-bonded catalytic radical transfer pathway in Escherichia coli ribonucleotide reductase (RNR) is evident from the three-dimensional structures of the R1 and R2 proteins, phylogenetic studies, and site-directed mutagenesis experiments. Current knowledge of electron transfer processes is difficult to apply to the very long radical transfer pathway in RNR. To explore the importance of the hydrogen bonds between the participating residues, we converted the protein R2 residue Asp237, one of the conserved residues along the radical transfer route, to an asparagine and a glutamate residue in two separate mutant proteins. In this study, we show that the D237E mutant is catalytically active and has hydrogen bond connections similar to that of the wild type protein. This is the first reported mutant protein that affects the radical transfer pathway while catalytic activity is preserved. The D237N mutant is catalytically inactive, and its tyrosyl radical is unstable, although the mutant can form a diferric-oxo iron center and a R1-R2 complex. The data strongly support our hypothesis that an absolute requirement for radical transfer during catalysis in ribonucleotide reductase is an intact hydrogen-bonded pathway between the radical site in protein R2 and the substrate binding site in R1. Our data thus strongly favor the idea that the electron transfer mechanism in RNR is coupled with proton transfer, i.e. a radical transfer mechanism.

Binding of allosteric effectors to ribonucleotide reductase protein R1: reduction of active-site cysteines promotes substrate binding.

BACKGROUND: Ribonucleotide reductase (RNR) is an essential enzyme in DNA synthesis, catalyzing all de novo synthesis of deoxyribonucleotides. The enzyme comprises two dimers, termed R1 and R2, and contains the redox active cysteine residues, Cys462 and Cys225. The reduction of ribonucleotides to deoxyribonucleotides involves the transfer of free radicals. The pathway for the radical has previously been suggested from crystallographic results, and is supported by site-directed mutagenesis studies. Most RNRs are allosterically regulated through two different nucleotide-binding sites: one site controls general activity and the other controls substrate specificity. Our aim has been to crystallographically demonstrate substrate binding and to locate the two effector-binding sites. RESULTS: We report here the first crystal structure of RNR R1 in a reduced form. The structure shows that upon reduction of the redox active cysteines, the sulfur atom of Cys462 becomes deeply buried. The more accessible Cys225 moves to the former position of Cys462 making room for the substrate. In addition, the structures of R1 in complexes with effector, effector analog and effector plus substrate provide information about these binding sites. The substrate GDP binds in a cleft between two domains with its beta-phosphate bound to the N termini of two helices; the ribose forms hydrogen bonds to conserved residues. Binding of dTTP at the allosteric substrate specificity site stabilizes three loops close to the dimer interface and the active site, whereas the general allosteric binding site is positioned far from the active site. CONCLUSIONS: Binding of substrate at the active site of the enzyme is structurally regulated in two ways: binding of the correct substrate is regulated by the binding of allosteric effectors and binding of the actual substrate occurs primarily when the active-site cysteines are reduced. One of the loops stabilized upon binding of dTTP participates in the formation of the substrate-binding site through direct interaction with the nucleotide base. The general allosteric effector site, located far from the active site, appears to regulate subunit interactions within the holoenzyme.

Two conserved tyrosine residues in protein R1 participate in an intermolecular electron transfer in ribonucleotide reductase.

The enzyme ribonucleotide reductase consists of two nonidentical proteins, R1 and R2, which are each inactive alone. R1 contains the active site and R2 contains a stable tyrosyl radical essential for catalysis. The reduction of ribonucleotides is radical-based, and a long range electron transfer chain between the active site in R1 and the radical in R2 has been suggested. To find evidence for such an electron transfer chain in Escherichia coli ribonucleotide reductase, we converted two conserved tyrosines in R1 into phenylalanines by site-directed mutagenesis. The mutant proteins were shown to be enzymatically inactive. In addition, the mechanism-based inhibitor 2'-azido-2'-deoxy-CDP was incapable of scavenging the R2 radical, and no azido-CDP-derived radical intermediate was formed. We also show that the loss of enzymatic activity was not due to impaired R1-R2 complex formation or substrate binding. Based on these results, we predict that the two tyrosines, Tyr-730 and Tyr-731, are part of a hydrogen-bonded network that constitutes an electron transfer pathway in ribonucleotide reductase. It is demonstrated that there is no electron delocalization over these tyrosines in the resting wild-type complex.

Measuring and modeling the proximity effect in direct-write electron-beam lithography kinoforms.

The proximity effect in successively developed direct-write electron-beam lithography gratings is measured. The grating relief shapes are obtained from the measured power in several of the gratings' diffraction orders. Describing the proximity effect by a convolution with a double Gaussian point-spread function, we determine the parameters of the point-spread function. The writing part of the point-spread function is found to increase significantly with increasing development time, the background part much less.

Proximity-compensated blazed transmission grating manufacture with direct-writing, electron-beam lithography.

Proximity-compensated, as well as uncompensated, blazed transmission gratings with periods of 4, 8, and 16 µm were manufactured with direct-writing, electron-beam lithography in positive resist. The compensated gratings performed better than the uncompensated ones. For the 4-µm compensated grating the measured diffraction efficiency was 67%. It was 35% for the uncompensated grating. The compensation was made by repeated convolutions in the spatial domain with the electron-beam point spread function. We determined this function by retrieving the phase from the measured diffraction pattern of the uncompensated gratings.

Successive development optimization of resist kinoforms manufactured with direct-writing, electron-beam lithography.

It is shown that multilevel SAL 110 resist kinoforms can be developed stepwise. Measurements of the kinoform diffraction pattern, performed between the development steps, permitted correct final developments to be made. No significant relief shape degradation was observed for development times as high as 25 min. The results imply that the electron-beam exposure doses, and hence the exposure time, can be reduced by a factor of 3 compared with doses used currently.

Nd:YAG laser machining with multilevel resist kinoforms.

We demonstrate that resist kinoforms can be used for laser micromachining. A 10-level resist kinoform, manufactured by electron-beam lithography, was shown to have a diffraction efficiency of 68%. Nine diffraction-limited holes were simultaneously drilled in 0.10-mm-thick stainless steel. Marking in a silicon wafer is also demonstrated.

Chemotherapeutic drug sensitivity testing of human leukemia cells in vitro using a semiautomated fluorometric assay.

A simple and reproducible semiautomated fluorometric method for drug sensitivity testing of leukemic cells in microculture is described. The assay is based on hydrolysis of nonfluorescent fluorescein diacetate (FDA) to a strongly fluorescent product (fluorescein) by cells with intact plasma membranes after 72 hr of culture and was in the present study applied to acute lymphocytic leukemia (ALL) cell lines and specimens from patients with lymphocytic and myelocytic leukemia. FDA fluorescence was linearly related to viable cell number within a wide range of cell densities (3-4 logs) as well as in the presence of different added proportions of dead cells. The assay reliably detects high and low grade resistance to vincristine (vcr) and daunorubicin, respectively, as well as the subsequent reversal of vcr resistance by cyclosporin A and the calcium channel blocker verapamil. Using ALL cell lines, drug sensitivity was in good correspondence with data obtained by the microculture tetrazolium assay. Furthermore, drug sensitivity data of fresh leukemia cells from patients with leukemia were readily obtained. The results indicate that the presently described method is applicable for simple and reliable chemosensitivity testing of leukemia cell lines as well as tumor specimens from patients with leukemia.

Multilevel phase holograms manufactured by electron-beam lithography.

Ten-level transmission phase holograms (kinoforms) manufactured in one resist layer by electron-beam lithography are reported for the first time to our knowledge. The measured hologram diffraction efficiencies were 70% for the two resist materials used. This corresponds to 82% of the maximum theoretical value for these holograms and is, to our knowledge, the highest reported to date.

Single photomask, multilevel kinoforms in quartz and photoresist: manufacture and evaluation.

Kinoforms manufactured in photoresist by photolithographic techniques using a single, ten-level, grey scale photomask, exposed in a specially designed laser exposure system, are described. Kinoforms designed for uniform as well as for partial Gaussian beam illumination are discussed. The highest measured diffraction efficiency was 55%. Photoresist kinoforms were transferred into quartz substrates by reactive ion etching. The highest measured diffraction efficiency for the resulting all-quartz kinoforms was 53%.

Lipoproteins and metabolic control in hypertensive type II diabetics treated with clonidine.

Twenty patients with type II diabetes mellitus and hypertension (WHO stages I and II) participated in a 3-month double-blind cross-over study to evaluate the effects of clonidine (75-300 micrograms daily) on blood pressure, glycemic control and plasma lipoproteins. Already after 1 month's treatment with clonidine the systolic and diastolic blood pressures had decreased, from 168/103 to 161/98 mmHg (p less than 0.01). Fasting blood glucose and HbA1c concentrations were unaffected by 3 months' treatment. Similarly, plasma lipid and lipoprotein concentrations remained unchanged throughout the study (i.e. mean high and low density lipoprotein cholesterol concentrations were 0.89 and 3.87 mmol/l on placebo vs. 0.90 and 3.98 mmol/l on clonidine). Adverse effects were mild and tolerable, and consisted mainly of dryness of the mouth. We conclude that clonidine lowers the blood pressure in patients with type II diabetes without any adverse effects on glycemic control or plasma lipoproteins.