Brain Dopamine and Serotonin Receptors in the Perception of Pain. Positron Emission Tomography Studies in Healthy Subjects

The role of dopamine and serotonin in spinal pain regulation is well established. However, little is known concerning the role of brain dopamine and serotonin in the perception of pain in humans. The aim of this study was to assess the potential role of brain dopamine and serotonin in determining experimental pain sensitivity in humans using positron emission tomography (PET) and psychophysical methods. A total of 39 healthy subjects participated in the study, and PET imaging was performed to assess brain dopamine D2/D3 and serotonin 5-HT_1A receptor availability. In a separate session, sensitivity to pain and touch was assessed with traditional psychophysical methods, allowing the evaluation of potential associations between D2/D3 and 5-HT_1A binding and psychophysical responses. The subjects’ responses were also analyzed according to Signal Detection Theory, which enables separate assessment of the subject’s discriminative capacity (sensory factor) and response criterion (non-sensory factor). The study found that the D2/D3 receptor binding in the right putamen was inversely correlated with pain threshold and response criterion. 5-HT_1A binding in cingulate cortex, inferior temporal gyrus and medial prefrontal cortex was inversely correlated with discriminative capacity for touch. Additionally, the response criterion for pain and intensity rating of suprathreshold pain were inversely correlated with 5-HT_1A binding in multiple brain areas. The results suggest that brain D2/D3 receptors and 5-HT_1A receptors modulate sensitivity to pain and that the pain modulatory effects may, at least partly, be attributed to influences on the response criterion. 5-HT_1A receptors are also involved in the regulation of touch by having an effect on discriminative capacity.

CARBOXYLATION OF SILVER NANOPARTICLES FOR THE IMMOBILIZATION OF β-GALACTOSIDASE AND ITS EFFICACY IN GALACTO-OLIGOSACCHARIDES PRODUCTION

The present study investigated the carboxylation of silver nanoparticles (AgNPs) by 1:3 nitric acid-sulfuric acid mixtures for immobilizing Aspergillus oryzae β-galactosidase. Carboxylated AgNPs retained 93% enzyme upon immobilization and the enzyme did not leach out appreciably from the modified nanosupport in the presence of 100 mmol L-1 NaCl. Atomic force micrograph revealed the binding of β-galactosidase on the modified AgNPs. The optimal pH for soluble and carboxylated AgNPs adsorbed β-galactosidase (IβG) was observed at pH 4.5 while the optimal operating temperature was broadened from 50 ºC to 60 ºC for IβG. Michaelis constant, Km was increased two and a half fold for IβG while Vmax decreases slightly as compared to soluble enzyme. β-galactosidase immobilized on surface functionalized AgNPs retained 70% biocatalytic activity even at 4% galactose concentration as compared to enzyme in solution. Our study showed that IβG produces greater amount of galacto-oligosaccharides at higher temperatures (50 ºC and 60 ºC) from 0.1 mol L-1 lactose solution at pH 4.5 as compared to previous reports.

Sensitive Troponin Assays: Diagnostic and Prognostic Use in Cardiology

Cardiac troponins (cTns) are the recommended biochemical markers in the diagnosis of myocardial infarction (MI). They are very sensitive and tissue-specific but are limited by their delayed appearance in the circulation. Biochemical markers with more rapid release kinetics, e.g. myoglobin and especially heart-type fatty acid-binding protein (H-FABP), have been used to enhance the early identification of MI. The implementation of cTns into clinical practice has shown that cardiomyocyte injury occurs in many other clinical conditions than MI. The aim of this study was to evaluate the impact of modern and highly sensitive cTnI assays on the early diagnosis of MI. In a patient cohort with suspected MI, such a sensitive cTnI assay enhanced the early diagnostic accuracy when compared to a less sensitive cTnI assay and to myoglobin. When compared to H-FABP during the early hours after symptom onset, the sensitive cTnI assay showed at least similar and, after 6 hours, superior diagnostic accuracy. A positive cTnI test result had superior prognostic value when compared to H-FABP, even among early presenters. The prognostic value of cTn in acute heart failure (AHF) was evaluated in 364 patients who participated in the FINN-AKVA study. The patients presented with AHF but no acute coronary syndrome (ACS). Up to half of the patients had elevated cTn levels which were associated with higher 6-month mortality. The magnitude of cTn elevation was directly proportional to mortality. Finally, the clinical spectrum of cTnI elevations was evaluated in 991 cTnI positive emergency department (ED) patients. 83% of the patients had MI and 17% had cTnI elevation due to other clinical conditions. The latter patient group was characterized by lower absolute cTnI levels and – importantly – higher in-hospital mortality when compared to the MI patients. In conclusion, the use of a highly sensitive cTnI assay enhances the early diagnostic accuracy and risk stratification in suspected MI patients. Cardiac troponin elevations are highly prevalent also in other acute clinical conditions and indicate an adverse outcome of these patients.

Equivalence and translation strategies in multilingual thesaurus construction

In the present dissertation, multilingual thesauri were approached as cultural products and the focus was twofold: On the empirical level the focus was placed on the translatability of certain British-English social science indexing terms into the Finnish language and culture at a concept, a term and an indexing term level. On the theoretical level the focus was placed on the aim of translation and on the concept of equivalence. In accordance with modern communicative and dynamic translation theories the interest was on the human dimension. The study is qualitative. In this study, equivalence was understood in a similar way to how dynamic, functional equivalence is commonly understood in translation studies. Translating was seen as a decision-making process, where a translator often has different kinds of possibilities to choose in order to fulfil the function of the translation. Accordingly, and as a starting point for the construction of the empirical part, the function of the source text was considered to be the same or similar to the function of the target text, that is, a functional thesaurus both in source and target context. Further, the study approached the challenges of multilingual thesaurus construction from the perspectives of semantics and pragmatics. In semantic analysis the focus was on what the words conventionally mean and in pragmatics on the ‘invisible’ meaning - or how we recognise what is meant even when it is not actually said (or written). Languages and ideas expressed by languages are created mainly in accordance with expressional needs of the surrounding culture and thesauri were considered to reflect several subcultures and consequently the discourses which represent them. The research material consisted of different kinds of potential discourses: dictionaries, database records, and thesauri, Finnish versus British social science researches, Finnish versus British indexers, simulated indexing tasks with five articles and Finnish versus British thesaurus constructors. In practice, the professional background of the two last mentioned groups was rather similar. It became even more clear that all the material types had their own characteristics, although naturally not entirely separate from each other. It is further noteworthy that the different types and origins of research material were not used to represent true comparison pairs, and that the aim of triangulation of methods and material was to gain a holistic view. The general research questions were: 1. Can differences be found between Finnish and British discourses regarding family roles as thesaurus terms, and if so, what kinds of differences and which are the implications for multilingual thesaurus construction? 2. What is the pragmatic indexing term equivalence? The first question studied how the same topic (family roles) was represented in different contexts and by different users, and further focused on how the possible differences were handled in multilingual thesaurus construction. The second question was based on findings of the previous one, and answered to the final question as to what kinds of factors should be considered when defining translation equivalence in multilingual thesaurus construction. The study used multiple cases and several data collection and analysis methods aiming at theoretical replication and complementarity. The empirical material and analysis consisted of focused interviews (with Finnish and British social scientists, thesaurus constructors and indexers), simulated indexing tasks with Finnish and British indexers, semantic component analysis of dictionary definitions and translations, coword analysis and datasets retrieved in databases, and discourse analysis of thesauri. As a terminological starting point a topic and case family roles was selected. The results were clear: 1) It was possible to identify different discourses. There also existed subdiscourses. For example within the group of social scientists the orientation to qualitative versus quantitative research had an impact on the way they reacted to the studied words and discourses, and indexers placed more emphasis on the information seekers whereas thesaurus constructors approached the construction problems from a more material based solution. The differences between the different specialist groups i.e. the social scientists, the indexers and the thesaurus constructors were often greater than between the different geo-cultural groups i.e. Finnish versus British. The differences occurred as a result of different translation aims, diverging expectations for multilingual thesauri and variety of practices. For multilingual thesaurus construction this means severe challenges. The clearly ambiguous concept of multilingual thesaurus as well as different construction and translation strategies should be considered more precisely in order to shed light on focus and equivalence types, which are clearly not self-evident. The research also revealed the close connection between the aims of multilingual thesauri and the pragmatic indexing term equivalence. 2) The pragmatic indexing term equivalence is very much context-depended. Although thesaurus term equivalence is defined and standardised in the field of library and information science (LIS), it is not understood in one established way and the current LIS tools are inadequate to provide enough analytical tools for both constructing and studying different kinds of multilingual thesauri as well as their indexing term equivalence. The tools provided in translation science were more practical and theoretical, and especially the division of different meanings of a word provided a useful tool in analysing the pragmatic equivalence, which often differs from the ideal model represented in thesaurus construction literature. The study thus showed that the variety of different discourses should be acknowledged, there is a need for operationalisation of new types of multilingual thesauri, and the factors influencing pragmatic indexing term equivalence should be discussed more precisely than is traditionally done.

Fibroblast Growth Factor 8 and its Receptors in The Growth and Angiogenesis of Experimental Breast Cancer . With Special Reference to Thrombospondin-1 as a Novel Target Gene for FGF-8

The growth of breast cancer is regulated by hormones and growth factors. Recently, aberrant fibroblast growth factor (FGF) signalling has been strongly implicated in promoting the progression of breast cancer and is thought to have a role in the development of endocrine resistant disease. FGFs mediate their auto- and paracrine signals through binding to FGF receptors 1-4 (FGFR1-4) and their isoforms. Specific targets of FGFs in breast cancer cells and the differential role of FGFRs, however, are poorly described. FGF-8 is expressed at elevated levels in breast cancer, and it has been shown to act as an angiogenic, growth promoting factor in experimental models of breast cancer. Furthermore, it plays an important role in mediating androgen effects in prostate cancer and in some breast cancer cell lines. We aimed to study testosterone (Te) and FGF-8 regulated genes in Shionogi 115 (S115) breast cancer cells, characterise FGF-8 activated intracellular signalling pathways and clarify the role of FGFR1, -2 and -3 in these cells. Thrombospondin-1 (TSP-1), an endogenous inhibitor of angiogenesis, was recognised as a Te and FGF-8 regulated gene. Te repression of TSP-1 was androgen receptor (AR)-dependent. It required de novo protein synthesis, but it was independent of FGF-8 expression. FGF-8, in turn, downregulated TSP-1 transcription by activating the ERK and PI3K pathways, and the effect could be reversed by specific kinase inhibitors. Differential FGFR1-3 action was studied by silencing each receptor by shRNA expression in S115 cells. FGFR1 expression was a prerequisite for the growth of S115 tumours, whereas FGFR2 expression alone was not able to promote tumour growth. High FGFR1 expression led to a growth advantage that was associated with strong ERK activation, increased angiogenesis and reduced apoptosis, and all of these effects could be reversed by an FGFR inhibitor. Taken together, the results of this thesis show that FGF-8 and FGFRs contribute strongly to the regulation of the growth and angiogenesis of experimental breast cancer and support the evidence for FGF-FGFR signalling as one of the major players in breast cancers.

TEMPERATURE AND SUBSTRATE INFLUENCES ON SEED GERMINATION AND SEEDLING FORMATION IN Callisthene fasciculata Mart. (VOCHYSIACEAE) IN THE LABORATORY1

ABSTRACTCallisthene fasciculata Mart. is a tree belonging to the Vochysiaceae family. Its wood is moderately heavy and resistant and used to make poles, beams, and other structures. The aim of this work was to evaluate seed germination and the initial growth of seedlings of C. fasciculata at different temperatures and in different substrates. Seeds were collected from fruits in the Pantanal de Miranda, Mato Grosso do Sul state, Brazil. In one experiment, the seeds were subjected to constant temperatures of 20, 25, 30 and 35 °C and to alternating temperatures of 20-30 and 25-35 °C (on paper substrate). In another experiment, the seeds were subjected to temperatures of 20 and 25 °C on three substrates (sand, vermiculite and between paper) in a germinator. The experiment had a randomized design, with four replicates of 25 seeds per treatment. The F-values obtained for germination indicated no significant effect of substrate or temperature on the final germination percentage. The analyses revealed no effect of a treatment interaction (temperature x substrate) on either germination or average germination time; however, a treatment interaction effect was observed on the germination speed index. The treatment combinations yielding the best performance were between paper substrate at 20 °C and sand substrate at 25 °C. There was a significant effect of the interaction between temperature and substrate on seedling growth, with increased root growth observed in the between paper substrate at 25 °C and increased aerial component growth in both sand at 20 °C and vermiculite at 25 °C. The between paper treatment at 25 °C yielded the greatest final seedling size. Between paper is the most recommended substrate for the production of seedlings due to its ease of handling and lower probability of contamination.

Mobile learning and video experience in delivering digital content

In recent years, the worldwide distribution of smartphone devices has been growing rapidly. Mobile technologies are evolving fast, a situation which provides new possibilities for mobile learning applications. Along with new delivery methods, this development enables new concepts for learning. This study focuses on the effectiveness and experience of a mobile learning video promoting the key features of a specific device. Through relevant learning theories, mobile technologies and empirical findings, the thesis presents the key elements for a mobile learning video that are essential for effective learning. This study also explores how previous experience with mobile services and knowledge of a mobile handset relate to final learning results. Moreover, this study discusses the optimal delivery mechanisms for a mobile video. The target group for the study consists of twenty employees of a Sanoma Company. The main findings show that the individual experience of learning and the actual learning results may differ and that the design for certain video elements, such as sound and the presentation of technical features, can have an impact on the experience and effectiveness of a mobile learning video. Moreover, a video delivery method based on cloud technologies and HTML5 is suggested to be used in parallel with standalone applications.

Immediate Decision-Making and Information Needs in Intensive Care Coordination

The aim of this study was to develop a theoretical model for information integration to support the deci¬sion making of intensive care charge nurses, and physicians in charge – that is, ICU shift leaders. The study focused on the ad hoc decision-making and immediate information needs of shift leaders during the management of an intensive care unit’s (ICU) daily activities. The term ‘ad hoc decision-making’ was defined as critical judgements that are needed for a specific purpose at a precise moment with the goal of ensuring instant and adequate patient care and a fluent flow of ICU activities. Data collection and research analysis methods were tested in the identification of ICU shift leaders’ ad hoc decision-making. Decision-making of ICU charge nurses (n = 12) and physicians in charge (n = 8) was observed using a think-aloud technique in two university-affiliated Finnish ICUs for adults. The ad hoc decisions of ICU shift leaders were identified using an application of protocol analysis. In the next phase, a structured online question¬naire was developed to evaluate the immediate information needs of ICU shift leaders. A national survey was conducted in all Finnish, university-affiliated hospital ICUs for adults (n = 17). The questionnaire was sent to all charge nurses (n = 515) and physicians in charge (n = 223). Altogether, 257 charge nurses (50%) and 96 physicians in charge (43%) responded to the survey. The survey was also tested internationally in 16 Greek ICUs. From Greece, 50 charge nurses out of 240 (21%) responded to the survey. A think-aloud technique and protocol analysis were found to be applicable for the identification of the ad hoc decision-making of ICU shift leaders. During one day shift leaders made over 200 ad hoc decisions. Ad hoc decisions were made horizontally, related to the whole intensive care process, and vertically, concerning single intensive care incidents. Most of the ICU shift leaders’ ad hoc decisions were related to human resources and know-how, patient information and vital signs, and special treatments. Commonly, this ad hoc decision-making involved several multiprofessional decisions that constituted a bundle of immediate decisions and various information needs. Some of these immediate information needs were shared between the charge nurses and the physicians in charge. The majority of which concerned patient admission, the organisation and management of work, and staff allocation. In general, the information needs of charge nurses were more varied than those of physicians. It was found that many ad hoc deci-sions made by the physicians in charge produced several information needs for ICU charge nurses. This meant that before the task at hand was completed, various kinds of information was sought by the charge nurses to support the decision-making process. Most of the immediate information needs of charge nurses were related to the organisation and management of work and human resources, whereas the information needs of the physicians in charge mainly concerned direct patient care. Thus, information needs differ between professionals even if the goal of decision-making is the same. The results of the international survey confirmed these study results for charge nurses. Both in Finland and in Greece the information needs of charge nurses focused on the organisation and management of work and human resources. Many of the most crucial information needs of Finnish and Greek ICU charge nurses were common. In conclusion, it was found that ICU shift leaders make hundreds of ad hoc decisions during the course of a day related to the allocation of resources and organisation of patient care. The ad hoc decision-making of ICU shift leaders is a complex multi-professional process, which requires a lot of immediate information. Real-time support for information related to patient admission, the organisation and man¬agement of work, and allocation of staff resources is especially needed. The preliminary information integration model can be applied when real-time enterprise resource planning systems are developed for intensive care daily management

Antimicrobial resistance and detection of mecA and blaZ genes in coagulase-negative Staphylococcus isolated from bovine mastitis

The present study evaluated the pheno- and genotypical antimicrobial resistance profile of coagulase-negative Staphylococcus (CNS) species isolated from dairy cows milk, specially concerning to oxacillin. Of 100 CNS isolates, the S. xylosus was the prevalent species, followed by S. cohnii, S. hominis, S. capitis and S. haemolyticus. Only 6% were phenotypically susceptible to the antimicrobial agents tested in disk diffusion assay. Penicillin and ampicillin resistance rates were significantly higher than others antimicrobials. Four isolates were positive to mecA gene (4%), all represented by the S. xylosus species. The blaZ gene was detected in 16% of the isolates (16/100). It was noticed that all mecA + were also positive to this gene and the presence of both genes was correlated to phenotypic beta-lactamic resistance. We conclude that CNS species from bovine milk presented significantly distinct antimicrobial resistance profiles, evaluated by phenotypic and genotypic tests, which has implications for treatment and management decisions.

Professional sports and gambling regulation in Finland - Examining the institutional factors of gambling affecting professional sport clubs in Finland

This study examines how to institutional environment of gambling is currently in motion both in Europe and Finland. Furthermore, it examines the criticism by Finnish professional sport clubs directed towards the national gambling monopolies, especially Veikkaus Oy. This criticism addresses the acclaimed issue of low or non-existing sponsorship funds coming to the clubs despite the clubs’ duties to promote Veikkaus Oy in their stadiums etc. In essence the main research objective was to examine the interaction and institutional environments of both Finnish professional sport clubs and gambling regulation. This was done through three sub-objectives: 1) to analyze professional sport as business and its institutional environment 2) to analyze the institutions of gambling in their current state and their potential future 3) to evaluate the potential impact of an institutional change in gambling legislation to the professional sport clubs The findings from Finland were then compared to those of Denmark where an institutional change had occurred in gambling regulation. Empirical data was collected through multiple interviews. Interviewees represented sport clubs (7), sport association (1), sport league (1), Finnish monopoly representatives (2), commercial gambling providers (1), Danish monopoly system representatives (1), Danish sport club (1). In addition a vast amount of secondary data (e.g. Green and white books by EU, court decisions, a variety of studies etc.). Theoretically this study combines the aspects of institutional theory with the theory of professional sports as business. This proved to be a rather new approach and no published literature was found to have done specifically this. The findings of this study are twofold, on the European level it is clear that the momentum if towards a more liberated gambling market while Finland is at the moment trying to go the opposite direction and uphold its monopoly. From the sport club’s level the findings suggest that currently sport clubs do not directly benefit from the funds originated from Veikkaus Oy as these funds are more or less used on the association/league levels. However, the clubs themselves are also lacking in self-criticism as they are lacking in clear sponsorship packages/programs which Veikkaus Oy might be interested in participating. If liberation of the gambling market would occur it is highly possible that that the largest clubs in football and ice-hockey would be the main beneficiaries while smaller clubs and sports could possibly be worse off than currently. These interpretations were well supported by the findings from Denmark.

Biomolecular screening for inhibitors of butyrylcholinesterase : identification and characterization using in vitro and in silico tools

Drug discovery is a continuous process where researchers are constantly trying to find new and better drugs for the treatment of various conditions. Alzheimer’s disease, a neurodegenerative disease mostly affecting the elderly, has a complex etiology with several possible drug targets. Some of these targets have been known for years while other new targets and theories have emerged more recently. Cholinesterase inhibitors are the major class of drugs currently used for the symptomatic treatment of Alzheimer’s disease. In the Alzheimer’s disease brain there is a deficit of acetylcholine and an impairment in signal transmission. Acetylcholinesterase has therefore been the main target as this is the main enzyme hydrolysing acetylcholine and ending neurotransmission. It is believed that by inhibiting acetylcholinesterase the cholinergic signalling can be enhanced and the cognitive symptoms that arise in Alzheimer’s disease can be improved. Butyrylcholinesterase, the second enzyme of the cholinesterase family, has more recently attracted interest among researchers. Its function is still not fully known, but it is believed to play a role in several diseases, one of them being Alzheimer’s disease. In this contribution the aim has primarily been to identify butyrylcholinesterase inhibitors to be used as drug molecules or molecular probes in the future. Both synthetic and natural compounds in diverse and targeted screening libraries have been used for this purpose. The active compounds have been further characterized regarding their potencies, cytotoxicity, and furthermore, in two of the publications, the inhibitors ability to also inhibit Aβ aggregation in an attempt to discover bifunctional compounds. Further, in silico methods were used to evaluate the binding position of the active compounds with the enzyme targets. Mostly to differentiate between the selectivity towards acetylcholinesterase and butyrylcholinesterase, but also to assess the structural features required for enzyme inhibition. We also evaluated the compounds, active and non-active, in chemical space using the web-based tool ChemGPS-NP to try and determine the relevant chemical space occupied by cholinesterase inhibitors. In this study, we have succeeded in finding potent butyrylcholinesterase inhibitors with a diverse set of structures, nine chemical classes in total. In addition, some of the compounds are bifunctional as they also inhibit Aβ aggregation. The data gathered from all publications regarding the chemical space occupied by butyrylcholinesterase inhibitors we believe will give an insight into the chemically active space occupied by this type of inhibitors and will hopefully facilitate future screening and result in an even deeper knowledge of butyrylcholinesterase inhibitors.

Balancing creativity and risk intelligence in professional knowledge-intensive business services

The main strengths of professional knowledge-intensive business services (P-KIBS) are knowledge and creativity which needs to be fostered, maintained and supported. The process of managing P-KIBS companies deals with financial, operational and strategic risks. That is why it is reasonable to apply risk management techniques and frameworks in this context. A significant challenge hides in choosing reasonable ways of implementing risk management, which will not limit creative ability in organization, and furthermore will contribute to the process. This choice is related to a risk intelligent approach which becomes a justified way of finding the required balance. On a theoretical level the field of managing both creativity and risk intelligence as a balanced process remains understudied in particular within KIBS industry. For instance, there appears to be a wide range of separate models for innovation and risk management, but very little discussion in terms of trying to find the right balance between them. This study aims to shed light on the importance of well-managed combination of these concepts. The research purpose of the present study is to find out how the balance between creativity and risk intelligence can be managed in P-KIBS. The methodological approach utilized in the study is strictly conceptual without empirical aspects. The research purpose can be achieved through answering the following research supporting questions: 1. What are the characteristics and role of creativity as a component of innovation process in a P-KIBS company? 2. What are the characteristics and role of risk intelligence as an approach towards risk management process implementation in a P-KIBS company? 3. How can risk intelligence and creativity be balanced in P-KIBS? The main theoretical contribution of the study conceals in a proposed creativity and risk intelligence stage process framework. It is designed as an algorithm that can be applied on organizational canvas. It consists of several distinct stages specified by actors involved, their roles and implications. Additional stage-wise description provides detailed tasks for each of the enterprise levels, while combining strategies into one. The insights driven from the framework can be utilized by a vast range of specialists from strategists to risk managers, and from innovation managers to entrepreneurs. Any business that is designing and delivering knowledge service can potentially gain valuable thoughts and expand conceptual understanding from the present report. Risk intelligence in the current study is a unique way of emphasizing the role of creativity in professional knowledge-intensive industry and a worthy technique for making profound decisions towards risks.

Fructan degradation and hydrolytic activity in tuberous roots of Viguiera discolor Baker (Asteraceae), a herbaceous species from the cerrado

Fructans of the inulin type are the major reserve carbohydrates in tuberous roots of Viguiera discolor, a perennial herb native to the cerrado. Changes in molecular mass of the polymer, followed by releasing free fructose suggested that hydrolysis could be related to the sprouting of the buds after the dormant period, when aerial parts of the plant are naturally absent. Excision of aerial parts resulted in the increase of fructan 1-exohydrolase (1-FEH) activity in tuberous roots after sprouting. 1-FEH was partially purified from this material by binding to ConA-Sepharose and the highest activity was detected at pH 5.4 and between 20 and 40 °C. Values of Km for V. discolor inulin could not be determined since no saturation was observed up to 10%. The study of the kinetics of the 1-FEH activity showed that it does not follow Michaelis-Menten and apparently presents allosteric behaviour, as data fits in the Hill equation. The 1-FEH from V. discolor is a glycoprotein, more active on low molecular mass fructans than on high molecular mass inulin from the same species.

Regulation of HSF2 and its function in mitosis

The cell is continuously subjected to various forms of external and intrinsic proteindamaging stresses, including hyperthermia, pathophysiological states, as well as cell differentiation and proliferation. Proteindamaging stresses result in denaturation and improper folding of proteins, leading to the formation of toxic aggregates that are detrimental for various pathological conditions, including Alzheimer’s and Huntington’s diseases. In order to maintain protein homeostasis, cells have developed different cytoprotective mechanisms, one of which is the evolutionary well-conserved heat shock response. The heat shock response results in the expression of heat shock proteins (Hsps), which act as molecular chaperones that bind to misfolded proteins, facilitate their refolding and prevent the formation of protein aggregates. Stress-induced expression of Hsps is mediated by a family of transcription factors, the heat shock factors, HSFs. Of the four HSFs found in vertebrates, HSF1-4, HSF1 is the major stress-responsive factor that is required for the induction of the heat shock response. HSF2 cannot alone induce Hsps, but modulates the heat shock response by forming heterotrimers with HSF1. HSFs are not only involved in the heat shock response, but they have also been found to have a function in development, neurodegenerative disorders, cancer, and longevity. Therefore, insight into how HSFs are regulated is important for the understanding of both normal physiological and disease processes. The activity of HSF1 is mainly regulated by intricate post-translational modifications, whereas the activity of HSF2 is concentrationdependent. However, there is only limited understanding of how the abundance of HSF2 is regulated. This study describes two different means of how HSF2 levels are regulated. In the first study it was shown that microRNA miR-18, a member of the miR-17~92 cluster, directly regulates Hsf2 mRNA stability and thus protein levels. HSF2 has earlier been shown to play a profound role in the regulation of male germ cell maturation during the spermatogenesis. The effect on miR-18 on HSF2 was examined in vivo by transfecting intact seminiferous tubules, and it was found that inhibition of miR-18 resulted in increased HSF2 levels and modified expression of the HSF2 targets Ssty2 and Speer4a. HSF2 has earlier been reported to modulate the heat shock response by forming heterotrimers with HSF1. In the second study, it was shown that HSF2 is cleared off the Hsp70 promoter and degraded by the ubiquitinproteasome pathway upon acute stress. By silencing components of the anaphase promoting complex/cyclosome (APC/C), including the co-activators Cdc20 and Cdh1, it was shown that APC/C mediates the heatinduced ubiquitylation of HSF2. Furthermore, down-regulation of Cdc20 was shown to alter the expression of heat shock-responsive genes. Next, we studied if APC/C-Cdc20, which controls cell cycle progression, also regulates HSF2 during the cell cycle. We found that both HSF2 mRNA and protein levels decreased during mitosis in several but not all human cell lines, indicating that HSF2 has a function in mitotic cells. Interestingly, although transcription is globally repressed during mitosis, mainly due to the displacement of RNA polymerase II and transcription factors, including HSF1, from the mitotic chromatin, HSF2 is capable of binding DNA during mitosis. Thus, during mitosis the heat shock response is impaired, leaving mitotic cells vulnerable to proteotoxic stress. However, in HSF2-deficient mitotic cells the Hsp70 promoter is accessible to both HSF1 and RNA polymerase II, allowing for stress-inducible Hsp expression to occur. As a consequence HSF2-deficient mitotic cells have a survival advantage upon acute heat stress. The results, presented in this thesis contribute to the understanding of the regulatory mechanisms of HSF2 and its function in the heat shock response in both interphase and mitotic cells.

Correlation of discocyte frequency and ATP concentration in preserved blood: A morphological indicator of red blood cell viability

Red blood cells (RBC) are viable if kept in an adequate preservative solution, although gradual changes in morphology and metabolism may occur. There is a gradual decrease in adenosine-5'-triphosphate (ATP) concentration, pH, glucose consumption, and enzyme activity during preservation. The normal discocyte shapes are initially replaced by echinocytes and stomatocytes and, at final stages, by spherocytes, the last step before splenic sequestration. Post-transfusional survival has been correlated with the ATP concentration. RBC preserved in ADSOL, a solution containing adenine, dextrose, sodium chloride, and mannitol, are viable for transfusion for up to 6 weeks. Erythrocytes from 10 blood units taken from healthy adult donors were preserved for 12 weeks in ADSOL at 4oC. We now report a significant correlation (r2 = 0.98) between the percentage of discocytes (89 to 7%) and ATP (100 to 10%) concentration in ADSOL-preserved RBC. The results suggest that the percent of discocyte shapes used as an indicator of ATP concentration may be a useful indicator for quality control of RBC viability in centers which have limited assay facilities.