The effect of interrogative function on intonation in spontaneous and read Finnish

Goals This study aims to map the effect of interrogative function on the intonation of spontaneous and read Finnish. Earlier research shows that the most prominent feature in Finnish question intonation is an appeal to the listener. Question word questions typically start with a high peak which is followed by falling intonation. In yes/no questions, F0 remains on a high level until the word carrying sentence stress and then falls. Final rises are mainly found in intonation clichés such as "Ai mitä?" ("What?") These earlier results are based on read speech and enacted dialogues. In this study, questions and statements found in spontaneous dialogues were compared. These utterances were also compared with read versions of the same utterances. Fundamental frequency values were compared using a mixed model. Contours were also grouped using auditory and visual inspection. Thus it was possible to compare frequencies of contour types according to utterance type and speech style. The position of questions in the F0 distribution of the whole material was also investigated in this study. Method The material consisted of four spontaneous dialogues and their read versions. The speakers were young adults from the Helsinki metropolitan area, four females and four males. The whole material was first divided into broad dialogue function categories arising from the material and F0 curves were calculated for each category. After this, 277 questions and 244 statements were selected for closer inspection. Values reflecting F0 distribution and contour shape were measured from the F0 contours of these utterances. A mixed model was used to analyse the differences. Utterance type, question type, speech style and speaker gender were used as fixed effects. The frequencies of F0 contour types were compared using a Chi square test. Additional material in this study came from eight young female speakers in central Finland. Results and conclusions In the mixed model analysis, significant differences were found both between questions and statements and between spontaneous and read speech. Generally, utterance type affected the variables reflecting contour type while speech style affected the variables reflecting F0 distribution. The effect of question type was not clearly visible. In read speech the contours resembled earlier results more closely. Speakers had different strategies in differentiating between questions and statements. In the whole material, F0 was slightly higher in questions than in statements. The effect of dialectal background could be seen in the contour types. The results show that interrogative function affects intonation in both spontaneous and read Finnish.

Behaviour of methylmethacrylate-acrylonitrile random copolymers in dilute solution

Studies on the dilute solution properties of methylmethacrylate-acrylonitrile random copolymers of three different compositions, 0.236, 0.5 and 0.74 mole fraction (m.f.) of acrylonitrile (AN) designated as MAa, MAb and MAc, respectively, have been made in good solvents and theta solvents. MAa has been studied in benzene (Bz) and ethylacetate (EAc). MAb in acetonitrile (MeCN), dimethyl sulphoxide (DMSO) and a binary solvent mixture of Bz and dimentyl formamide (DMF) in the volume ratio 6.5:1 designated as BM1 and MAc in MeCN, DMSO and Bz + DMF in the volume ratio 1.667:1 designated as BM2. The Mark-Houwink exponent ‘a’ reveals that Bz is a theta solvent for MAa at 20°C. For MAb and MAc, BM1 and BM2, respectively have ‘a’ values of 0.5 at all three temperatures studied (30°, 40° and 50°C). It is not clear whether they represent theta states or preferential adsorption plays a role complicating the behaviour in solution. The values of A2 are very low in MeCN considering that it is a very good solvent for the copolymer, ‘a’ values for MAb and MAc being 0.75 and 0.7, respectively.

Calculating the Fickian diffusivity for a lattice-based random walk with agents and obstacles of different shapes and sizes

Random walk models are often used to interpret experimental observations of the motion of biological cells and molecules. A key aim in applying a random walk model to mimic an in vitro experiment is to estimate the Fickian diffusivity (or Fickian diffusion coefficient),D. However, many in vivo experiments are complicated by the fact that the motion of cells and molecules is hindered by the presence of obstacles. Crowded transport processes have been modeled using repeated stochastic simulations in which a motile agent undergoes a random walk on a lattice that is populated by immobile obstacles. Early studies considered the most straightforward case in which the motile agent and the obstacles are the same size. More recent studies considered stochastic random walk simulations describing the motion of an agent through an environment populated by obstacles of different shapes and sizes. Here, we build on previous simulation studies by analyzing a general class of lattice-based random walk models with agents and obstacles of various shapes and sizes. Our analysis provides exact calculations of the Fickian diffusivity, allowing us to draw conclusions about the role of the size, shape and density of the obstacles, as well as examining the role of the size and shape of the motile agent. Since our analysis is exact, we calculateDdirectly without the need for random walk simulations. In summary, we find that the shape, size and density of obstacles has a major influence on the exact Fickian diffusivity. Furthermore, our results indicate that the difference in diffusivity for symmetric and asymmetric obstacles is significant.

High-resolution melt-curve analysis of random-amplified-polymorphic-DNA markers, for the characterisation of pathogenic Leptospira.

A new test for pathogenic Leptospira isolates, based on RAPD-PCR and high-resolution melt (HRM) analysis (which measures the melting temperature of amplicons in real time, using a fluorescent DNA-binding dye), has recently been developed. A characteristic profile of the amplicons can be used to define serovars or detect genotypes. Ten serovars, of leptospires from the species Leptospira interrogans (serovars Australis, Robinsoni, Hardjo, Pomona, Zanoni, Copenhageni and Szwajizak), L. borgpetersenii (serovar Arborea), L. kirschneri (serovar Cynopteri) and L. weilii (serovar Celledoni), were typed against 13 previously published RAPD primers, using a real-time cycler (the Corbett Life Science RotorGene 6000) and the optimised reagents from a commercial kit (Quantace SensiMix). RAPD-HRM at specific temperatures generated defining amplicon melt profiles for each of the tested serovars. These profiles were evaluated as difference-curve graphs generated using the RotorGene software package, with a cut-off of at least 8 'U' (plus or minus). The results demonstrated that RAPD-HRM can be used to measure serovar diversity and establish identity, with a high degree of stability. The characterisation of Leptospira serotypes using a DNA-based methodology is now possible. As an objective and relatively inexpensive and rapid method of serovar identification, at least for cultured isolates, RAPD-HRM assays show convincing potentia.

Random Breath Testing Operation and Effectiveness in 1987

• In December 1986 funds were approved to double the intensity of random breath testing (RBT) and provide publicity support for police efforts. These changes were considered necessary to make RBT effective. • RBT methods were changed in the metropolitan area to enable block testing (pulling over a block of traffic rather than one or two cars), deployment of police to cut off escape routes, and testing by traffic patrols in all police subdivisions. Additional operators were trained for country RBT. • A publicity campaign was developed, aimed mainly at male drivers aged 18-50. The campaign consisted of the “cardsharp” television commercials, radio commercials, newspaper articles, posters and pamphlets. • Increased testing and the publicity campaigns were launched on 10 April 1987. • Police tests increased by 92.5% in May – December 1987, compared with the same period in the previous four years. • The detection rate for drinking drivers picked up by police who were cutting off escape routes was comparatively high, indicating that drivers were attempting to avoid RBT, and that this police method was effective at detecting these drivers. • A telephone survey indicated that drivers were aware of the messages of the publicity campaign. • The telephone survey also indicated that the target group had been exposed to high levels of RBT, as planned, and that fear of apprehension was the major factor deterring them from drink driving. • A roadside survey of driver blood alcohol concentrations (BACs) by the University of Adelaide’s Road Accident Research Unit (RARU) showed that, between 10p.m. and 3a.m., the proportion of drivers in Adelaide with a BAC greater than or equal to 0/08 decreased by 42%. • Drivers under 21 were identified as a possible problem area. • Fatalities in the twelve month period commencing May 1987 decreased by 18% in comparison with the previous twelve month period, and by 13% in comparison with the average of the previous two twelve month periods (commencing May 1985 and May 1986). There are indications that this trend is continuing. • It is concluded that the increase in RBT, plus publicity, was successful in achieving its aims of reductions in drink driving and accidents.

Random Breath Testing Operation and Effectiveness in 1985

Random breath testing (RBT) was introduced in South Australia in 1981 with the intention of reducing the incidence of accidents involving alcohol. In April 1985, a Select Committee of the Upper House which had been established to “review the operation of random breath testing in this State and any other associated matters and report accordingly” presented its report. After consideration of this report, the Government introduced extensive amendments to those sections of the Motor Vehicles Act (MVA) and Road Traffic Act (RTA) which deal with RBT and drink driving penalties. The amended section 47da of the RTA requires that: “(5) The Minister shall cause a report to be prepared within three months after the end of each calendar year on the operation and effectiveness of this section and related sections during that calendar year. (6) The Minister shall, within 12 sitting days after receipt of a report under subsection (5), cause copies of the report to be laid before each House of Parliament.” This is the first such report. Whilst it deals with RBT over a full year, the changed procedures and improved flexibility allowed by the revision to the RTA were only introduced late in 1985 and then only to the extent that the existing resources would allow.

Learning with symmetric label noise: The importance of being unhinged

Convex potential minimisation is the de facto approach to binary classification. However, Long and Servedio [2008] proved that under symmetric label noise (SLN), minimisation of any convex potential over a linear function class can result in classification performance equivalent to random guessing. This ostensibly shows that convex losses are not SLN-robust. In this paper, we propose a convex, classification-calibrated loss and prove that it is SLN-robust. The loss avoids the Long and Servedio [2008] result by virtue of being negatively unbounded. The loss is a modification of the hinge loss, where one does not clamp at zero; hence, we call it the unhinged loss. We show that the optimal unhinged solution is equivalent to that of a strongly regularised SVM, and is the limiting solution for any convex potential; this implies that strong l2 regularisation makes most standard learners SLN-robust. Experiments confirm the unhinged loss’ SLN-robustness.

Frequency and temperature-dependent mobility of a charged carrier on a randomly interrupted strand

A randomly interrupted strand model of a one-dimensional conductor is considered. An exact analytical expression is obtained for the temperature-dependent ac mobility for a finite segment drawn at random, taking into account the reflecting barriers at the two open ends. The real part of mobility shows a broad resonance as a function of both frequency and temperature, and vanishes quadratically in the dc limit. The frequency (temperature) maximum shifts to higher values for higher temperatures (frequencies).

High-resolution melt-curve analysis of random amplified polymorphic DNA (RAPD-HRM) for the characterisation of pathogenic leptospires: intra-serovar divergence, interserovar convergence, and evidence of attenuation in Leptospira reference collections.

High-resolution melt-curve analysis of random amplified polymorphic DNA (RAPD-HRM) is a novel technology that has emerged as a possible method to characterise leptospires to serovar level. RAPD-HRM has recently been used to measure intra-serovar convergence between strains of the same serovar as well as inter-serovar divergence between strains of different serovars. The results indicate that intra-serovar heterogeneity and inter-serovar homogeneity may limit the application of RAPD-HRM in routine diagnostics. They also indicate that genetic attenuation of aged, high-passage-number isolates could undermine the use of RAPD-HRM or any other molecular technology. Such genetic attenuation may account for a general decrease seen in titres of rabbit hyperimmune antibodies over time. Before RAPD-HRM can be further advanced as a routine diagnostic tool, strains more representative of the wild-type serovars of a given region need to be identified. Further, RAPD-HRM analysis of reference strains indicates that the routine renewal of reference collections, with new isolates, may be needed to maintain the genetic integrity of the collections.

Biophysical Characterization of Oxidized Phospholipids : Implications for Altered Membrane Structure and Function

The present study aims to elucidate the modifications in the structure and functionality of the phospholipid matrix of biological membranes brought about by free radical-mediated oxidative damage of its molecular constituents. To this end, the surface properties of two oxidatively modified phospholipids bearing an aldehyde or carboxyl function at the end of truncated sn-2 acyl chain were studied using a Langmuir balance. The results obtained reveal both oxidized species to have a significant impact on the structural dynamics of phospholipid monolayers, as illustrated by the progressive changes in force-area isotherms with increasing mole fraction of the oxidized lipid component. Moreover, surface potential measurements revealed considerable modifications in the electric properties of oxidized phospholipid containing monolayers during film compression, suggesting a packing state-controlled reorientation of the intramolecular electric dipoles of the lipid headgroups and acyl chains. Based on the above findings, a model describing the conformational state of oxidized phospholipid molecules in biological membranes is proposed, involving the protrusion of the acyl chains bearing the polar functional groups out from the hydrocarbon phase to the surrounding aqueous medium. Oxidative modifications alter profoundly the physicochemical properties of unsaturated phospholipids and are therefore readily anticipated to have important implications for their interactions with membrane-associating molecules. Along these lines, the carboxyl group bearing lipid was observed to bind avidly the peripheral membrane protein cytochrome c. The binding was reversed following increase in ionic strength or addition of polyanionic ATP, thus suggesting it to be driven by electrostatic interactions between cationic residues of the protein and the deprotonated lipid carboxyl exposed to the aqueous phase. The presence of aldehyde function bearing oxidized phospholipid was observed to enhance the intercalation of four antimicrobial peptides into phospholipid monolayers and liposomal bilayers. Partitioning of the peptides to monolayers was markedly attenuated by the aldehyde scavenger methoxyamine, revealing it to be mediated by the carbonyl moiety possibly through efficient hydrogen bonding or, alternatively, formation of covalent adduct in form of a Schiff base between the lipid aldehydes and primary amine groups of the peptide molecules. Lastly, both oxidized phospholipid species were observed to bind with high affinity three small membrane-partitioning therapeutic agents, viz. chlorpromazine, haloperidol, and doxorubicin. In conclusion, the results of studies conducted using biomimetic model systems support the notion that oxidative damage influences the molecular architecture as well as the bulk physicochemical properties of phospholipid membranes. Further, common polar functional groups carried by phospholipids subjected to oxidation were observed to act as molecular binding sites at the lipid-water interface. It is thus plausible that oxidized phospholipid species may elicit cellular level effects by modulating integration of various membrane-embedded and surface-associated proteins and peptides, whose conformational state, oligomerization, and functionality is known to be controlled by highly specific lipid-protein interactions and proper physical state of the membrane environment.

Expression and function of angiotensins in the regulation of intraocular pressure - an experimental study

Glaucoma is a multifactorial long-term ocular neuropathy associated with progressive loss of the visual field, retinal nerve fiber structural abnormalities and optic disc changes. Like arterial hypertension it is usually a symptomless disease, but if left untreated leads to visual disability and eventual blindness. All therapies currently used aim to lower intraocular pressure (IOP) in order to minimize cell death. Drugs with new mechanisms of action could protect glaucomatous eyes against blindness. Renin-angiotensin system (RAS) is known to regulate systemic blood pressure and compounds acting on it are in wide clinical use in the treatment of hypertension and heart failure but not yet in ophthalmological use. There are only few previous studies concerning intraocular RAS, though evidence is accumulating that drugs antagonizing RAS can also lower IOP, the only treatable risk factor in glaucoma. The main aim of this experimental study was to clarify the expression of the renin-angiotensin system in the eye tissues and to test its potential oculohypotensive effects and mechanisms. In addition, the possible relationship between the development of hypertension and IOP was evaluated in animal models. In conclusion, a novel angiotensin receptor type (Mas), as well as ACE2 enzyme- producing agonists for Mas, were described for the first time in the eye structures participating in the regulation of IOP. In addition, a Mas receptor agonist significantly reduced even normal IOP. The effect was abolished by a specific receptor antagonist. Intraocular, local RAS would thus to be involved in the regulation of IOP, probably even more in pathological conditions such as glaucoma though there was no unambiguous relationship between arterial and ocular hypertension. The findings suggest the potential as antiglaucomatous drugs of agents which increase ACE2 activity and the formation of angiotensin (1-7), or activate Mas receptors.

The mechanisms of human renal epithelial cell modulation of autologous dendritic cell phenotype and function

Proximal tubule epithelial cells (PTEC) of the kidney line the proximal tubule downstream of the glomerulus and play a major role in the re-absorption of small molecular weight proteins that may pass through the glomerular filtration process. In the perturbed disease state PTEC also contribute to the inflammatory disease process via both positive and negative mechanisms via the production of inflammatory cytokines which chemo-attract leukocytes and the subsequent down-modulation of these cells to prevent uncontrolled inflammatory responses. It is well established that dendritic cells are responsible for the initiation and direction of adaptive immune responses. Both resident and infiltrating dendritic cells are localised within the tubulointerstitium of the renal cortex, in close apposition to PTEC, in inflammatory disease states. We previously demonstrated that inflammatory PTEC are able to modulate autologous human dendritic cell phenotype and functional responses. Here we extend these findings to characterise the mechanisms of this PTEC immune-modulation using primary human PTEC and autologous monocyte-derived dendritic cells (MoDC) as the model system. We demonstrate that PTEC express three inhibitory molecules: (i) cell surface PD-L1 that induces MoDC expression of PD-L1; (ii) intracellular IDO that maintains the expression of MoDC CD14, drives the expression of CD80, PD-L1 and IL-10 by MoDC and inhibits T cell stimulatory capacity; and (iii) soluble HLA-G (sHLA-G) that inhibits HLA-DR and induces IL-10 expression by MoDC. Collectively the results demonstrate that primary human PTEC are able to modulate autologous DC phenotype and function via multiple complex pathways. Further dissection of these pathways is essential to target therapeutic strategies in the treatment of inflammatory kidney disorders.

Milk casein-derived tripeptides : A special focus on blood pressure and vascular function

Increased consumption of low-fat milk products is inversely associated with the risk of hypertension. The beneficial effect of milk on blood pressure is attributed to high calcium and potassium content but also to specific peptide sequences, which are cleaved from milk protein during gastrointestinal digestion, fermentation of milk with proteolytic starter cultures or enzymatic hydrolysis. Milk products fermented with Lactobacillus helveticus contain casein-derived tripeptides isoleucine-proline-proline (Ile-Pro-Pro) and valine-proline-proline (Val-Pro-Pro), which have been shown to possess antihypertensive effects in humans and in experimental animals. The aim of the present series of studies was to investigate the effects of tripeptides Ile-Pro-Pro and Val-Pro-Pro or fermented milk products containing them on vascular function and blood pressure and to elucidate the mechanisms behind them by using different experimental models of hypertension. Another aim was to characterize the acute effects of tripeptides on blood pressure and arterial stiffness in mildly hypertensive humans. Ile-Pro-Pro and Val-Pro-Pro or fermented milk products containing them attenuated the development of hypertension in two experimental models of hypertension, spontaneously hypertensive rat (SHR) and type 2 diabetic Goto-Kakizaki (GK) rat fed with high-salt diet. Significant differences in systolic blood pressure (SBP) were seen after 8 weeks treatment with tripeptide-containing products compared to control product. Plant sterols did not enhance this effect. Two differently produced tripeptide powders produced a similar attenuating effect on SBP in SHR. In mildly hypertensive subjects, a single administration of tripeptide- and plant sterol-containing fermented milk product decreased both SBP and diastolic blood pressure (DBP) over a period of 8 hours. Protective effect of tripeptides Ile-Pro-Pro and Val-Pro-Pro and fermented milk products containing them on vascular function was demonstrated in in vitro studies and long-term experimental studies. The effect was shown to be endothelium-dependent and possibly involving endothelium-derived hyperpolarizing factor (EDHF). In the clinical study, single administration of tripeptide-containing fermented milk product did not affect measures of arterial stiffness. Long-term treatment with fermented milk product containing Ile-Pro-Pro and Val-Pro-Pro inhibited angiotensin-converting enzyme (ACE) and decreased aldosterone levels thus showing beneficial effects on the renin-angiotensin system (RAS) in SHR and GK. No changes in the components of RAS were observed by the single administration of the same product in mildly hypertensive subjects. Increased levels of cGMP, NOx and citrulline suggest increased nitric oxide (NO) production by the tripeptides. Taken together, Ile-Pro-Pro and Val-Pro-Pro -containing products attenuate the development of hypertension after long-term treatment in experimental models of hypertension and possess an acute antihypertensive effect in mildly hypertensive subjects. In addition, these tripeptides show endothelium-mediated beneficial effects on vascular function. Attenuation of blood pressure increase by the tripeptides in experimental animals involves RAS, but its role in the antihypertensive effect in humans remains to be elucidated.

Function and regulation of Cdk7

Understanding the process of cell division is crucial for modern cancer medicine due to the central role of uncontrolled cell division in this disease. Cancer involves unrestrained proliferation as a result of cells loosing normal control and being driven through the cell cycle, where they normally would be non-dividing or quiescent. Progression through the cell cycle is thought to be dependent on the sequential activation of cyclin-dependent kinases (Cdks). The full activation of Cdks requires the phosphorylation of a conserved residue (threonine-160 on human Cdk2) on the T-loop of the kinase domain. In metazoan species, a trimeric complex consisting of Cdk7, cyclin H and Mat1 has been suggested to be the T-loop kinase of several Cdks. In addition, Cdk7 have also been implicated in the regulation of transcription. Cdk7, cyclin H, and Mat1 can be found as subunits of general transcription factor TFIIH. Cdk7, in this context, phosphorylates the Carboxy-terminal domain (CTD) of the large subunit of RNA polymerase II (RNA pol II), specifically on serine-5 residues of the CTD repeat. The regulation of Cdk7 in these and other functions is not well known and the unambiguous characterization of the in vivo role of Cdk7 in both T-loop activation and CTD serine-5 phosphorylation has proved challenging. In this study, the fission yeast Cdk7-cyclin H homologous complex, Mcs6-Mcs2, is identified as the in vivo T-loop kinase of Cdk1(Cdc2). It also identifies multiple levels of regulation of Mcs6 kinase activity, i.e. association with Pmh1, a novel fission yeast protein that is the apparent homolog of metazoan Mat1, and T-loop phosphorylation of Mcs6, mediated by Csk1, a monomeric T-loop kinase with similarity to Cak1 of budding yeast. In addition, Skp1, a component of the SCF (Skp1-Cullin-F box protein) ubiquitin ligase is identified by its interactions with Mcs2 and Pmh1. The Skp1 association with Mcs2 and Pmh1 is however SCF independent and does not involve proteolytic degradation but may reflect a novel mechanism to modulate the activity or complex assembly of Mcs6. In addition to Cdk7, also Cdk8 has been shown to have CTD serine-5 kinase activity in vitro. Cdk8 is not essential in yeast but has been shown to function as a transcriptional regulator. The function of Cdk8 is unknown in flies and mammals. This prompted the investigation of murine Cdk8 and its potential role as a redundant CTD serine-5 kinase. We find that Cdk8 is required for development prior to implantation, at a time that is co-incident with a burst of Cdk8 expression during normal development. The results does not support a role of Cdk8 as a serine-5 CTD kinase in vivo but rather shows an unexpected requirement for Cdk8, early in mammalian development. The results presented in this thesis extends our current knowledge of the regulation of the cell cycle by characterizing the function of two distinct cell cycle regulating T-loop kinases, including the unambiguous identification of Mcs6, the fission yeast Cdk7 homolog, as the T-loop kinase of Cdk1. The results also indicate that the function of Mcs6 is conserved from fission yeast to human Cdk7 and suggests novel mechanisms by which the distinct functions of Cdk7 and Mcs6 could be regulated. These findings are important for our understanding of how progression of the cell cycle and proper transcription is controlled, during normal development and tissue homeostasis but also under condition where cells have escaped these control mechanisms e.g. cancer.