A quantum chemical investigation of the metal centres in cytochrome c oxidase

Quantum effects are often of key importance for the function of biological systems at molecular level. Cellular respiration, where energy is extracted from the reduction of molecular oxygen to water, is no exception. In this work, the end station of the electron transport chain in mitochondria, cytochrome c oxidase, is investigated using quantum chemical methodology. Cytochrome c oxidase contains two haems, haem a and haem a3. Haem a3, with its copper companion, CuB, is involved in the final reduction of oxygen into water. This binuclear centre receives the necessary electrons from haem a. Haem a, in turn, receives its electrons from a copper ion pair in the vicinity, called CuA. Density functional theory (DFT) has been used to clarify the charge and spin distributions of haem a, as well as changes in these during redox activity. Upon reduction, the added electron is shown to be evenly distributed over the entire haem structure, important for the accommodation of the prosthetic group within the protein. At the same time, the spin distribution of the open-shell oxidised state is more localised to the central iron. The exact spin density distribution has been disputed in the literature, however, different experiments indicating different distributions of the unpaired electron. The apparent contradiction is shown to be due to the false assumption of a unit amount of unpaired electron density; in fact, the oxidised state has about 1.3 unpaired electrons. The validity of the DFT results have been corroborated by wave function based coupled cluster calculations. Point charges, for use in classical force field based simulations, have been parameterised for the four metal centres, using a newly developed methodology. In the procedure, the subsystem for which point charges are to be obtained, is surrounded by an outer region, with the purpose of stabilising the inner region, both electronically and structurally. Finally, the possibility of vibrational promotion of the electron transfer step between haem a and a3 has been investigated. Calculating the full vibrational spectra, at DFT level, of a combined model of the two haems, revealed several normal modes that do shift electron density between the haems. The magnitude of the shift was found to be moderate, at most. The proposed mechanism could have an assisting role in the electron transfer, which still seems to be dominated by electron tunnelling.

Aza-Wittigringslutningar vid syntes av sammansmälta kväveheterocykler

Denna pro gradu avhandling är en litteraturstudie av intramolekylära aza-Wittigringslutningar vid syntes av sammansmälta kväveheterocykler. I arbetet behandlas material huvudsakligen från 1980 och framöver. Aza-Wittigreaktionen påminner om den analoga Wittigreaktionen. Aza-Wittigreaktioner har nästan uteslutande gjorts mellan karbonylgrupper och iminofosforaner. Reaktionsmekanismen sker enligt en tvåstegsaddition, som inleds av iminokvävets nukleofila attack till karbonylkolet och avslutas av att en zwitterjonisk betain bildar en azoxafosfetanintermediär. Intermediären sönderfaller spontant till en iminoprodukt och fosfinoxid. Reaktionen görs ofta under milda betingelser d.v.s. vid rumstemperatur och dessutom utan dyr arbetsutrustning Ett vanligt sätt att framställa iminofosforanen är från azid genom Staudingerreaktion och ofta kombineras Staudinger- och aza-Wittigreaktionerna så att iminofosforanen inte isoleras före ringslutningen. Alla andra ringslutningsreaktioner än aza-Wittig, såsom elektrocykliska ringslutningar, har uteslutits ur detta arbete. Materialet i litteraturstudien har indelats enligt vilken typ av karbonylgrupp iminofosforanen reagerar med. På så vis åskådliggörs hurudana typiska produktmolekyler som erhållits med ringslutning till en viss typ av karbonylgrupp. Det har visat sig att det är förmånligt om karbonylkolet har elektronunderskott och iminofosforanens kväve har elektronöverskott. Den ringslutande molekylens entropi och närbelägna substituenters elektroniska och steriska natur samt produktmolekylens termodynamiska fördelaktighet inverkar tillsammans på reaktionens gång. Ifall reaktionen förväntas ske långsamt är det bättre att använda sig av alkyliminofosforaner än aryliminofosforaner. Valet av lösningsmedel har nästan uteslutande lämnats oförklarat i de behandlade publikationerna men i de flesta fall har ortoxylen eller toluen varit goda lösningsmedel. Oönskad tetrazolbildning av aziden kan minimeras genom användning av opolärt lösningsmedel. Likaså kan förmånligt placerade kväveskyddsgrupper hindra intramolekylära vätebindningar. På senare tid har aza-Wittigreaktionen allt mera tillämpats vid framställning av farmakologiska produkter, vilket ökat intresset för att framställa stora mängder närbesläktade produktmolekyler. Dylika molekylbibliotek har med fördel framställts i fastfas varvid reningen av produkten underlättats märkbart. Ett nytt område inom aza-Wittigsyntetiken är asymmetriska reaktioner, vilka säkert kommer att få mera uppmärksamhet i framtiden. I denna litteraturstudie framkom det att många av de utförda synteserna på området kunde upprepas med större variation och systematik gällande reagens och reaktionsbetingelser.

Redox-linked proton transfer by cytochrome c oxidase

The respiratory chain is found in the inner mitochondrial membrane of higher organisms and in the plasma membrane of many bacteria. It consists of several membrane-spanning enzymes, which conserve the energy that is liberated from the degradation of food molecules as an electrochemical proton gradient across the membrane. The proton gradient can later be utilized by the cell for different energy requiring processes, e.g. ATP production, cellular motion or active transport of ions. The difference in proton concentration between the two sides of the membrane is a result of the translocation of protons by the enzymes of the respiratory chain, from the negatively charged (N-side) to the positively charged side (P-side) of the lipid bilayer, against the proton concentration gradient. The endergonic proton transfer is driven by the flow of electrons through the enzymes of the respiratory chain, from low redox-potential electron donors to acceptors of higher potential, and ultimately to oxygen. Cytochrome c oxidase is the last enzyme in the respiratory chain and catalyzes the reduction of dioxygen to water. The redox reaction is coupled to proton transport across the membrane by a yet unresolved mechanism. Cytochrome c oxidase has two proton-conducting pathways through which protons are taken up to the interior part of the enzyme from the N-side of the membrane. The K-pathway transfers merely substrate protons, which are consumed in the process of water formation at the catalytic site. The D-pathway transfers both substrate protons and protons that are pumped to the P-side of the membrane. This thesis focuses on the role of two conserved amino acids in proton translocation by cytochrome c oxidase, glutamate 278 and tryptophan 164. Glu278 is located at the end of the D-pathway and is thought to constitute the branching point for substrate and pumped protons. In this work, it was shown that although Glu278 has an important role in the proton transfer mechanism, its presence is not an obligatory requirement. Alternative structural solutions in the area around Glu278, much like the ones present in some distantly related heme-copper oxidases, could in the absence of Glu278 support the formation of a long hydrogen-bonded water chain through which proton transfer from the D-pathway to the catalytic site is possible. The other studied amino acid, Trp164, is hydrogen bonded to the ∆-propionate of heme a3 of the catalytic site. Mutation of this amino acid showed that it may be involved in regulation of proton access to a proton acceptor, a pump site, from which the proton later is expelled to the P-side of the membrane. The ion pair that is formed by the ∆-propionate of heme a3 and arginine 473 is likely to form a gate-like structure, which regulates proton mobility to the P-side of the membrane. The same gate may also be part of an exit path through which water molecules produced at the catalytically active site are removed towards the external side of the membrane. Time-resolved optical and electrometrical experiments with the Trp164 to phenylalanine mutant revealed a so far undetected step in the proton pumping mechanism. During the A to PR transition of the catalytic cycle, a proton is transferred from Glu278 to the pump site, located somewhere in the vicinity of the ∆-propionate of heme a3. A mechanism for proton pumping by cytochrome c oxidase is proposed on the basis of the presented results and the mechanism is discussed in relation to some relevant experimental data. A common proton pumping mechanism for all members of the heme-copper oxidase family is moreover considered.

Traditional medicinal uses and biological activities of some plant extracts of African Combretum Loefl., Terminalia L. and Pteleopsis Engl. species

In Africa various species of Combretum, Terminalia and Pteleopsis are used in traditional medicine. Despite of this, some species of these genera have still not been studied for their biological effects to validate their traditional uses. The aim of this work has been to document the ethnomedicinal uses of several species of Combretum and Terminalia in Mbeya region, south-western Tanzania, and to use this information for finding species with good antimicrobial and cytotoxic potential. During a five weeks expedition to Tanzania in spring 1999 sixteen different species of Combretum and Terminalia, as well as Pteleopsis myrtifolia were collected from various locations in the districts of Mbeya, Iringa and Dar-es-Salaam. Traditional healers in seven different villages in the Mbeya region were interviewed in Swahili and Nyakyusa on the medicinal uses of Combretum and Terminalia species shown to them. A questionnaire was used during the interviews. The results of the interviews correlated well between different villages, the same species being used in similar ways in different villages. Of the ten species shown to the healers six were frequently used for treatment of skin diseases, bacterial infections, diarrhea, oedema and wounds. The dried plants were most commonly prepared into hot water decoctions or mixed into maize porridge, Ugali. Infusions made from dried or fresh plant material were also common. Wounds and topical infections were treated with ointments made from the dried plant material mixed with sheep fat. Twenty-one extracts of six species of Combretum and four of Terminalia, collected from Tanzania, were screened for their antibacterial effects against two gram-negative and five gram-positive bacteria, as well as the yeast, Candida albicans, using an agar diffusion method. Most of the screened plants showed substantial antimicrobial activity. A methanolic root extract of T. sambesiaca showed the most potent antibacterial effects of all the plant species screened, and gave a MIC value of 0.9 mg/ml against Enterobacter aerogenes. Also root extracts of T. sericea and T. kaiserana gave excellent antimicrobial effects, and notably a hot water extract of T. sericea was as potent as extracts of this species made from EtOH and MeOH. Thus, the traditional way of preparing T. sericea into hot water decoctions seems to extract antimicrobial compounds. Thirty-five extracts of five species of Terminalia, ten of Combretum and Pteleopsis myrtifolia were screened for their antifungal effects against five species of yeast (Candida spp.) and Cryptococcus neoformans. The species differed from each other to their antifungal effects, some being very effective whereas others showed no antifungal effects. The most effective extracts showed antifungal effects comparable to the standard antibiotics itraconazol and amphotericin B. Species of Terminalia gave in general stronger antifungal effects than those of Combretum. The best effects were obtained with methanolic root extracts of T. sambesiaca, T. sericea and T. kaiserana, and this investigation indicates that decoctions of these species might be used for treatment of HIV-related fungal infections. Twenty-seven crude extracts of eight species of Combretum, five of Terminalia and Pteleopsis myrtifolia were evaluated for their cytotoxic effects against human cancer cell lines (HeLa, cervical carcinoma; MCF 7, breast carcinoma, T 24 bladder carcinoma) and one endothelial cell line (BBCE, bovine brain capillary endothelial cells). The most outstanding effects were obtained with a leaf extract of Combretum fragrans, which nearly totally inhibited the proliferation of T 24 and HeLa cells at a concentration of 25 ug/ml and inhibited 60 % of the growth of the HeLa cells at a concentration of 4.3 ug/ml. The species of Terminalia were less cytotoxically potent than the Combretum species, although T. sericea and T. sambesiaca gave good cytotoxic effects (< 30 % proliferation). In summary this study indicates that some of the species of Terminalia, Combretum and Pteleopsis, used in Tanzanian traditional medicine, are powerful inhibitors of both microbial and cancer cell growth. In depth studies would be needed to find the active compounds behind these biological activities.

Population dynamics of blue mussels in a variable environment at the edge of their range

Spatial and temporal variation in the abundance of species can often be ascribed to spatial and temporal variation in the surrounding environment. Knowledge of how biotic and abiotic factors operate over different spatial and temporal scales in determining distribution, abundance, and structure of populations lies at the heart of ecology. The major part of the current ecological theory stems from studies carried out in central parts of the distributional range of species, whereas knowledge of how marginal populations function is inadequate. Understanding how marginal populations, living at the edge of their range, function is however in a key position to advance ecology and evolutionary biology as scientific disciplines. My thesis focuses on the factors affecting dynamics of marginal populations of blue mussels (Mytilus edulis) living close to their tolerance limits with regard to salinity. The thesis aims to highlight the dynamics at the edge of the range and contrast these with dynamics in more central parts of the range in order to understand the potential interplay between the central and the marginal part in the focal system. The objectives of the thesis are approached by studies on: (1) factors affecting regional patterns of the species, (2) long-term temporal dynamics of the focal species spaced along a regional salinity gradient, (3) selective predation by increasing populations of roach (Rutilus rutilus) when feeding on their main food item, the blue mussel, (4) the primary and secondary effects of local wave exposure gradients and (5) the role of small-scale habitat heterogeneity as determinants of large-scale pattern. The thesis shows that populations of blue mussels are largely determined by large scale changes in sea water salinity, affecting mainly recruitment success and longevity of local populations. In opposite to the traditional view, the thesis strongly indicate that vertebrate predators strongly affect abundance and size structure of blue mussel populations, and that the role of these predators increases towards the margin where populations are increasingly top-down controlled. The thesis also indicates that the positive role of biogenic habitat modifiers increases towards the marginal areas, where populations of blue mussels are largely recruitment limited. Finally, the thesis shows that local blue mussel populations are strongly dependent on high water turbulence, and therefore, dense populations are constrained to offshore habitats. Finally, the thesis suggests that ongoing sedimentation of rocky shores is detrimental for the species, affecting recruitment success and post-recruit survival, pushing stable mussel beds towards offshore areas. Ongoing large scale changes in the Baltic Sea, especially dilution processes with attendant effects, are predicted to substantially contract the distributional range of the mussel, but also affect more central populations. The thesis shows that in order to understand the functioning of marginal populations, research should (1) strive for multi-scale approaches in order to link ecosystem patterns with ecosystem processes, and (2) challenge the prevailing tenets that origin from research carried out in central areas that may not be valid at the edge.

From actin monomers to bundles: The roles of twinfilin and α-actinin in Drosophila melanogaster development

The actin cytoskeleton is essential for a large variety of cell biological processes. Actin exists in either a monomeric or a filamentous form, and it is very important for many cellular functions that the local balance between these two actin populations is properly regulated. A large number of proteins participate in the regulation of actin dynamics in the cell, and twinfilin, one of the proteins examined in this thesis, belongs to this category. The second level of regulation involves proteins that crosslink or bundle actin filaments, thereby providing the cell with a certain shape. α-Actinin, the second protein studied, mainly acts as an actin crosslinking protein. Both proteins are conserved in organisms ranging from yeast to mammals. In this thesis, the roles of twinfilin and α-actinin in development were examined using Drosophila melanogaster as a model organism. Twinfilin is an actin monomer binding protein that is structurally related to cofilin. In vitro, twinfilin reduces actin polymerisation by sequestering actin monomers. The Drosophila twinfilin (twf) gene was identified and found to encode a protein functionally similar to yeast and mammalian twinfilins. A strong hypomorphic twf mutation was identified, and flies homozygous for this allele were viable and fertile. The adult twf mutant flies displayed reduced viability, a rough eye phenotype and severely malformed bristles. The shape of the adult bristle is determined by the actin bundles that are regularly spaced around the perimeter of the developing pupal bristles. Examination of the twf pupal bristles revealed an increased level of filamentous actin, which in turn resulted in splitting and displacement of the actin bundles. The bristle defect was rescued by twf overexpression in developing bristles. The Twinfilin protein was localised at sites of actin filament assembly, where it was required to limit actin polymerisation. A genetic interaction between twinfilin and twinstar (the gene encoding Cofilin) was detected, consistent with the model predicting that both proteins act to limit the amount of filamentous actin. α-Actinin has been implicated in several diverse cell biological processes. In Drosophila, the only function for α-actinin yet known is in the organisation of the muscle sarcomere. Muscle and non-muscle cells utilise different α-actinin isoforms, which in Drosophila are produced by alternative splicing of a single gene. In this work, novel α-actinin deletion alleles, including ActnΔ233, were generated, which specifically disrupted the transcript encoding the non-muscle α-actinin isoform. Nevertheless, ActnΔ233 homozygous mutant flies were viable and fertile with no obvious defects. By comparing α-actinin protein distribution in wild type and ActnΔ233 mutant animals, it could be concluded that non-muscle α-actinin is the only isoform expressed in young embryos, in the embryonic central nervous system and in various actin-rich structures of the ovarian germline cells. In the ActnΔ233 mutant, α-actinin was detected not only in muscle tissue, but also in embryonic epidermal cells and in certain follicle cell populations in the ovaries. The population of α-actinin protein present in non-muscle cells of the ActnΔ233 mutant is referred to as FC-α-actinin (Follicle Cell). The follicular epithelium in the Drosophila ovary is a well characterised model system for studies on patterning and morphogenesis. Therefore, α-actinin expression, regulation and function in this tissue were further analysed. Examination of the α-actinin localisation pattern revealed that the basal actin fibres of the main body follicle cells underwent an organised remodelling during the final stages of oogenesis. This involved the assembly of a transient adhesion site in the posterior of the cell, in which α-actinin and Enabled (Ena) accumulated. Follicle cells genetically manipulated to lack all α-actinin isoforms failed to remodel their cytoskeleton and translocate Ena to the posterior of the cell, while the actin fibres as such were not affected. Neither was epithelial morphogenesis disrupted. The reorganisation of the basal actin cytoskeleton was also disturbed following ectopic expression of Decapentaplegic (Dpp) or as a result of a heat shock. At late oogenesis, the main body follicle cells express both non-muscle α-actinin and FC-α-actinin, while the dorsal anterior follicle cells express only non-muscle α-actinin. The dorsal anterior cells are patterned by the Dpp and Epidermal growth factor receptor (EGFR) signalling pathways, and they will ultimately secrete the dorsal appendages of the egg. Experiments involving ectopic activation of EGFR and Dpp signalling showed that FC-α-actinin is negatively regulated by combined EGFR and Dpp signalling. Ubiquitous overexpression of the adult muscle-specific α-actinin isoform induced the formation of aberrant actin bundles in migrating follicle cells that did not normally express FC-α-actinin, provided that the EGFR signalling pathway was activated in the cells. Taken together, this work contributes new data to our knowledge of α-actinin function and regulation in Drosophila. The cytoskeletal remodelling shown to depend on α-actinin function provides the first evidence that α-actinin has a role in the organisation of the cytoskeleton in a non-muscle tissue. Furthermore, the cytoskeletal remodelling constitutes a previously undescribed morphogenetic event, which may provide us with a model system for in vivo studies on adhesion dynamics in Drosophila.

Spatial ecology of a specialist insect herbivore : the leaf-mining moth Tischeria ekebladella on the pedunculate oak Quercus robur

Herbivorous insects comprise a major part of terrestrial biodiversity, and their interactions with their host plants and natural enemies are of vast ecological importance. A large body of research demonstrates that the ecology and evolution of these insects may be affected by trophic interactions, by abiotic influences, and by intraspecific processes, but so far research on these individual aspects has rarely been combined. This thesis uses the leaf-mining moth Tischeria ekebladella and the pedunculate oak (Quercus robur) as a case study to assess how spatial variation in trophic interactions and the physical distribution of host trees jointly affect the distribution, dynamics and evolution of a host-specific herbivore. With respect to habitat quality, Tischeria ekebladella experiences abundant variation at several spatial scales. Most of this variation occurs at small scales notably among leaves and shoots within individual trees. While hypothetically this could cause moths to evolve an ability to select leaves and shoots of high quality, I did not find any coupling between female preference and offspring performance. Based on my studies on temporal variation in resource quality I therefore propose that unpredictable temporal changes in the relative rankings of individual resource units may render it difficult for females to predict the fate of their developing offspring. With respect to intraspecific processes, my results suggest that limited moth dispersal in relation to the spatial distribution of oak trees plays a key role in determining the regional distribution of Tischeria ekebladella. The distribution of the moth is aggregated at the landscape level, where local leaf miner populations are less likely to be present where oaks are scarce. A modelling exercise based on empirical dispersal estimates revealed that the moth population on Wattkast an island in south-western Finland is spatially structured overall, but that the relative importance of local and regional processes on tree-specific moth dynamics varies drastically across the landscape. To conclude, my work in the oak-Tischeria ekebladella system demonstrates that the local abundance and regional distribution of a herbivore may be more strongly influenced by the spatial location of host trees than by their relative quality. Hence, it reveals the importance of considering spatial context in the study of herbivorous insects, and forms a bridge between the classical fields of plant-insect interactions and spatial ecology.

The influence of eutrophication on sexual selection in sticklebacks

Jordens ekologiska system undergår för tillfället stora förändringar pga. människans aktiviteter. Ett växande antal studier visar att dessa förändringar påverkar naturliga och sexuella urvalet och därmed evolutiva processer. Målet med detta arbete var att undersöka effekterna av omgivningsförändringar på sexuella urvalet genom att använda den ökade övergödningen inom storpiggen Gasterosteus aculeatus lekområden som modell system. Sexuella urvalet är en viktig evolutiv kraft med följder på populations- och artnivå (Kapitel 1). Avhandlingens olika delar fokuserar på övergödningens effekter på upptäckandet av partners, användningen av visuella- och doftsignaler i partnersval, och fördelningen av parningsframgången mellan bobyggande hanar. I Kapitel II och III simuleras hur grumlighet orsakad av fytoplankton påverkar hastigheten med vilken potentiella partners påträffas, genom effekter på synligheten. Resultaten visar att normala algblomningar i Östersjön har en måttlig effekt på finnandet av potentiella partners. Detta tyder på att algblomningarna troligen inte kommer att minska på selektiva parningen pga. ökade sökkostnader. I Kapitel IV visas att storspiggen ändrar relativa användningen av olika signaler när vattnets grumlighet ökar; visuella signaler minskar i betydelse medan doftsignaler ökar i betydelse. Samtidigt underlättas användandet av doftsignaler av ändringar i vattnets kemiska sammansättning då fotosyntesen intensifieras (Kapitel V). Lek i övergödda vatten kan ändå vara kostsamt både på individ- och populationsnivån, då parasiterade hanar, som troligen är dåligt genetiskt anpassade till sin miljö, lyckas få mer ägg i sina bon än friskare hanar som troligen är av högre genetisk kvalitet (Kapitel VI). Övergödningen påverkar således partnersval och konkurrensen om partners genom att påverka upptäckandet av potentiella partners, evalueringen av partners och fördelningen av partners inom lekområdena. De följder detta kan ha för evolutionen av sexuellt selekterad egenskaper och för populationers dynamik och livskraft är dock oklara. Avhandlingen visar på svårigheten att förutse följderna av omgivningsförändringar för sexuella urvalet och effekterna på individ och populationsnivå.

Molecular mechanisms of bacteriophage phi6 RNA-dependent RNA polymerase and its utilization in biotechnology

Double-stranded RNA (dsRNA) viruses encode only a single protein species that contains RNA-dependent RNA polymerase (RdRP) motifs. This protein is a central component in the life cycle of a dsRNA virus, carrying out both RNA transcription and replication. The architecture of viral RdRPs resembles that of a 'cupped right hand' with fingers, palm and thumb domains. Those applying de novo initiation have additional structural features, including a flexible C-terminal domain that constitutes the priming platform. Moreover, viral RdRPs must be able to interact with the incoming 3'-terminus of the template and position it so that a productive binary complex is formed. Bacteriophage phi6 of the Cystoviridae family is to date one of the best studied dsRNA viruses. The purified recombinant phi6 RdRP is highly active in vitro and possesses both RNA replication and transcription activities. The extensive biochemical observations and the atomic level crystal structure of the phi6 RdRP provides an excellent platform for in-depth studies of RNA replication in vitro. In this thesis, targeted structure-based mutagenesis, enzymatic assays and molecular mapping of phi6 RdRP and its RNA were used to elucidate the formation of productive RNA-polymerase binary complexes. The positively charged rim of the template tunnel was shown to have a significant role in the engagement of highly structured ssRNA molecules, whereas specific interactions further down in the template tunnel promote ssRNA entry to the catalytic site. This work demonstrated that by aiding the formation of a stable binary complex with optimized RNA templates, the overall polymerization activity of the phi6 RdRP can be greatly enhanced. Furthermore, proteolyzed phi6 RdRPs that possess a nick in the polypeptide chain at the hinge region, which is part of the extended loop, were better suited for catalysis at higher temperatures whilst favouring back-primed initiation. The clipped C-terminus remains associated with the main body of the polymerase and the hinge region, although structurally disordered, is involved in the control of C-terminal domain displacement. The accumulated knowhow on bacteriophage phi6 was utilized in the development of two technologies for the production of dsRNA: (i) an in vitro system that combines the T7 RNA polymerase and the phi6 RdRP to generate dsRNA molecules of practically unlimited length, and (ii) an in vivo RNA replication system based on restricted infection with phi6 polymerase complexes in bacterial cells to produce virtually unlimited amounts of dsRNA. The pools of small interfering RNAs derived from dsRNA produced by these systems were validated and shown to efficiently decrease the expression of both exogenous and endogenous targets.

Long-term clinical outcome after liver transplantation

With transplant rejection rendered a minor concern and survival rates after liver transplantation (LT) steadily improving, long-term complications are attracting more attention. Current immunosuppressive therapies, together with other factors, are accompanied by considerable long-term toxicity, which clinically manifests as renal dysfunction, high risk for cardiovascular disease, and cancer. This thesis investigates the incidence, causes, and risk factors for such renal dysfunction, cardiovascular risk, and cancer after LT. Long-term effects of LT are further addressed by surveying the quality of life and employment status of LT recipients. The consecutive patients included had undergone LT at Helsinki University Hospital from 1982 onwards. Data regarding renal function – creatinine and estimated glomerular filtration rate (GFR) – were recorded before and repeatedly after LT in 396 patients. The presence of hypertension, dyslipidemia, diabetes, impaired fasting glucose, and overweight/obesity before and 5 years after LT was determined among 77 patients transplanted for acute liver failure. The entire cohort of LT patients (540 patients), including both children and adults, was linked with the Finnish Cancer Registry, and numbers of cancers observed were compared to site-specific expected numbers based on national cancer incidence rates stratified by age, gender, and calendar time. Health-related quality of life (HRQoL), measured by the 15D instrument, and employment status were surveyed among all adult patients alive in 2007 (401 patients). The response rate was 89%. Posttransplant cardiovascular risk factor prevalence and HRQoL were compared with that in the age- and gender-matched Finnish general population. The cumulative risk for chronic kidney disease increased from 10% at 5 years to 16% at 10 years following LT. GFR up to 10 years after LT could be predicted by the GFR at 1 year. In patients transplanted for chronic liver disease, a moderate correlation of pretransplant GFR with later GFR was also evident, whereas in acute liver failure patients after LT, even severe pretransplant renal dysfunction often recovered. By 5 years after LT, 71% of acute liver failure patients were receiving antihypertensive medications, 61% were exhibiting dyslipidemia, 10% were diabetic, 32% were overweight, and 13% obese. Compared with the general population, only hypertension displayed a significantly elevated prevalence among patients – 2.7-fold – whereas patients exhibited 30% less dyslipidemia and 71% less impaired fasting glucose. The cumulative incidence of cancer was 5% at 5 years and 13% at 10. Compared with the general population, patients were subject to a 2.6-fold cancer risk, with non-melanoma skin cancer (standardized incidence ratio, SIR, 38.5) and non-Hodgkin lymphoma (SIR 13.9) being the predominant malignancies. Non-Hodgkin lymphoma was associated with male gender, young age, and the immediate posttransplant period, whereas old age and antibody induction therapy raised skin-cancer risk. HRQoL deviated clinically unimportantly from the values in the general population, but significant deficits among patients were evident in some physical domains. HRQoL did not seem to decrease with longer follow-up. Although 87% of patients reported improved working capacity, data on return to working life showed marked age-dependency: Among patients aged less than 40 at LT, 70 to 80% returned to work, among those aged 40 to 50, 55%, and among those above 50, 15% to 28%. The most common cause for unemployment was early retirement before LT. Those patients employed exhibited better HRQoL than those unemployed. In conclusion, although renal impairment, hypertension, and cancer are evidently common after LT and increase with time, patients’ quality of life remains comparable with that of the general population.

Understanding the chemistry of atmospheric aerosol particle formation via observations of physical properties

It is widely accepted that the global climate is heating up due to human activities, such as burning of fossil fuels. Therefore we find ourselves forced to make decisions on what measures, if any, need to be taken to decrease our warming effect on the planet before any irrevocable damage occurs. Research is being conducted in a variety of fields to better understand all relevant processes governing Earth s climate, and to assess the relative roles of anthropogenic and biogenic emissions into the atmosphere. One of the least well quantified problems is the impact of small aerosol particles (both of anthropogenic and biogenic origin) on climate, through reflecting solar radiation and their ability to act as condensation nuclei for cloud droplets. In this thesis, the compounds driving the biogenic formation of new particles in the atmosphere have been examined through detailed measurements. As directly measuring the composition of these newly formed particles is extremely difficult, the approach was to indirectly study their different characteristics by measuring the hygroscopicity (water uptake) and volatility (evaporation) of particles between 10 and 50 nm. To study the first steps of the formation process in the sub-3 nm range, the nucleation of gaseous precursors to small clusters, the chemical composition of ambient naturally charged ions were measured. The ion measurements were performed with a newly developed mass spectrometer, which was first characterized in the laboratory before being deployed at a boreal forest measurement site. It was also successfully compared to similar, low-resolution instruments. The ambient measurements showed that sulfuric acid clusters dominate the negative ion spectrum during new particle formation events. Sulfuric acid/ammonia clusters were detected in ambient air for the first time in this work. Even though sulfuric acid is believed to be the most important gas phase precursor driving the initial cluster formation, measurements of the hygroscopicity and volatility of growing 10-50 nm particles in Hyytiälä showed an increasing role of organic vapors of a variety of oxidation levels. This work has provided additional insights into the compounds participating both in the initial formation and subsequent growth of atmospheric new aerosol particles. It will hopefully prove an important step in understanding atmospheric gas-to-particle conversion, which, by influencing cloud properties, can have important climate impacts. All available knowledge needs to be constantly updated, summarized, and brought to the attention of our decision-makers. Only by increasing our understanding of all the relevant processes can we build reliable models to predict the long-term effects of decisions made today.

Pengarna eller livet? : En kvalitativ och longitudinell studie om långtidsarbetslösa unga i ett aktörsperspektiv

Your money or your life? A qualitative follow-up study of the young unemployed from an actor perspective is a qualitative and longitudinal study following 36 unemployed young people in Helsinki over a span of ten years. The purpose of the study is to shed light on how a few young people view employment/unemployment and their lives and future, how they as unemployed perceive their encounters with society, and how society supports them. Four so-called key informants were followed at a finer level of empirical detail. They were chosen for the thematic interviews because of their different personalities, starting points and preferences. Although some differences were expected, what the results show is quite striking. The individual stories raise a number of questions about differences between young people, about society s view of the young unemployed, and about the principles behind the so-called activation policy and how society s support is distributed. The key informants descriptions underline that the group young unemployed does not consist of individuals who are alike but that life is complex, that paid work and unemployment can be perceived very differently, and that background and unofficial support can have consequences for self-perception and for ways of looking at the future, vocational choices, paid work and activation policy. Margaret S. Archer s theory of Morphogenesis and Barbara Cruikshank s theory of constructing democracies compose the study s theoretical framework. The key informants stories give a picture of a formal support system that, even though it puts part of the responsibility for unemployment on the individuals themselves, in the name of fairness and equality, treats them in an impersonal way, not giving their personal situation and wishes much weight. As a consequence, those who share the dominant values of society do well, while others who do not are faced with difficulties. The bigger the gap between society s and the individual s values, the bigger the risk to be met by little understanding and by penalties. And vice versa: Those who initially have the right values and know how to deal with authorities get heard and their opinions get accepted. The informants ask for a more personal encounter, which could improve both the atmosphere and the clients experiences of being heard. Still the risk of having a more individualistic system should be addressed, as a new system might generate new winners, but just as well give new losers. Finally, we have to ask if the so-called activation policy is looking for answers primarily to a macro-level problem on the micro-level. If it does not produce more jobs, its support for the unemployed will be insignificant. It is not enough to think about what to do at the grassroots level to make the system more functional and support job-seeking. If the current rate of unemployment endures, the quality of life of the unemployed should be addressed. A first step could be taken by placing less guilt on the unemployed. Instead of talking about activating the unemployed, discussion should be targeted at removing structural impediments to employment. If we want to have less polarisation between the those with paid work and those without, who often struggle with low incomes, we need to include the macro-level in the discussion. What does high unemployment mean in a work-based society, where the individual s self-perception and important social forms of support are linked to labour income? And what can be done at the macro-level to change this undesirable condition at the micro-level? Keywords: Unemployment, Youth, Public interventions, Activation policy, Individual actors, Qualitative, Longitudinal, Holistic, Helsinki, Finland

Good Lives, Hidden Miseries : An Ethnography of Uncertainty in a Finnish Village

This is an ethnographic study, in the field of medical anthropology, of village life among farmers in southwest Finland. It is based on 12 months of field work conducted 2002-2003 in a coastal village. The study discusses how social and cultural change affects the life of farmers, how they experience it and how they act in order to deal with the it. Using social suffering as a methodological approach the study seeks to investigate how change is related to lived experiences, idioms of distress, and narratives. Its aim has been to draw a locally specific picture of what matters are at stake in the local moral world that these farmers inhabit, and how they emerge as creative actors within it. A central assumption made about change is that it is two-fold; both a constructive force which gives birth to something new, and also a process that brings about uncertainty regarding the future. Uncertainty is understood as an existential condition of human life that demands a response, both causing suffering and transforming it. The possibility for positive outcomes in the future enables one to understand this small suffering of everyday life both as a consequence of social change, which fragments and destroys, and as an answer to it - as something that is positively meaningful. Suffering is seen to engage individuals to ensure continuity, in spite of the odds, and to sustain hope regarding the future. When the fieldwork was initiated Finland had been a member of the European Union for seven years and farmers felt it had substantially impacted on their working conditions. They complained about the restrictions placed on their autonomy and that their knowledge was neither recognised, nor respected by the bureaucrats of the EU system. New regulations require them to work in a manner that is morally unacceptable to them and financial insecurity has become more prominent. All these changes indicate the potential loss of the home and of the ability to ensure continuity of the family farm. Although the study initially focused on getting a general picture of working conditions and the nature of farming life, during the course of the fieldwork there was repeated mention of a perceived high prevalence of cancer in the area. This cancer talk is replete with metaphors that reveal cultural meanings tied to the farming life and the core values of autonomy, endurance and permanence. It also forms the basis of a shared identity and a means of delivering a moral message about the fragmentation of the good life; the loss of control; and the invasion of the foreign. This thesis formed part of the research project Expressions of Suffering. Ethnographies of Illness Experiences in Contemporary Finnish Contexts funded by the Academy of Finland. It opens up a vital perspective on the multiplicity and variety of the experience of suffering and that it is particularly through the use of the ethnographic method that these experiences can be brought to light. Keywords: suffering, uncertainty, phenomenology, habitus, agency, cancer, farming

Stor Anna, Liten Anna och tio andra personporträtt : Om innehållsliga och språkliga mönster i en mediegenres kvinnobeskrivningar

The aim of this licentiate thesis is to analyse how femininity is constructed in twelve portrait interviews of women in the dailies Dagens Nyheter (Stockholm) and Hufvudstadsbladet (Helsinki) in September 1996, and to explore the portrait interview as a media genre. The qualitative analysis has a feminist and constructionist perspective and is connected to critical text analysis. It was carried out on two levels: first, femininity is identified on the linguistic level by choice of words, and second on the level of content (topical motifs/themes). The portrait interview as a genre constitutes a third dimension in the analysis: The aim is not towards the identification of femininity, but rather towards the identification of the portrait interview a relatively unexplored media genre. References (Swedish: omtal) to the principal character (or protagonist) are traced mainly through reference chains which consist of names, pronouns and substantive phrases. The interviewees were referred to by their full names in Dagens Nyheter (with the exception of the oldest and youngest interviewees, both of whom were mainly referred to by their first names), while the style of reference varied more in Hufvudstadsbladet. The position of the principal character was also analysed through her relation in the text to minor characters from her working life and from her private life. These minor characters maintained their subordinate positions in all of the portraits except that of the youngest principal character, in which the subsidiary voices became at least as strong as the voice of the principal character. Three frequently-recurring topical motifs occurred in the portraits: The first involved explanations for the principal character s success divided into three categories, agent, affect and ambition, the second concerned using journeys or trips as symbols for turning points in life, and the third referred to the ambiguity in the contradiction between private (family/other private life) and public (work) life. This ambiguity is connected to the portrait interview as a text type (genre) which features conclusions at the end of portraits, which in turn is characteristic of reportage. However, the analysis showed that the conclusions of the portrait interviews often also included elements of ambiguity. This was evident in the contradictions be14 tween private and public life that arose in the portrait interviews that focused on these two spheres. The portraits that focused on the principal character s public life showed ambiguity on a more general level concerning questions about being a woman and having a profession, and they often ended with a description of some details of her private life. The women in the portraits were all constructed as being successful, in terms of having achieved direct success, reflective success or success in the form of life wisdom. The women of direct success were described as ambitious individuals with no sidetracks on their life paths, while those of reflective success were described as active heroines who had received help from different agents, who could use their affects as enriching ingredients in life, but who in the end had control over their own lives (life stories). The elderly women were constructed as having achieved life wisdom and their portraits were focused upon the past. The portrait interview as a genre is characterised by journalistic freedom (in relation to the more strict news genre), by a now room (Swedish nurum ) where the journalist meets the principal character (usually via spoken dialogue that she or he transforms into written text to be read by a mass-media audience) and by the relatively closed structure of the portrait. The portrait is relatively independent in relation to the news genre and in relation to the context of what has previously been written, what is being written at the time and what will be written in the future the principal character does not need to belong to the newspaper s usual gallery of actors. Furthermore, the principal character is constructed as being independent in relation to the subsidiary characters and other media actors. The conflict is within the principal character herself and within her life story, unlike the news genre in which equal actors are in conflict with each other. The portrait is also independent in relation to the news lifespan; the publishing timetable is not as strict as in the news genre, but is still dependent on the factors initiating the portrait. The enclosures consist of a raw analysis of two of twelve portrait interviews and of copies of all portraits.

Samarbete genom lagstiftning. Samarbetsförfarandets organisation, procedur och innehåll

Samarbetslagen är en omstridd lag. Innan lagen trädde i kraft i början av år 1979 fanns på arbetstagarsidan förhoppningar om att skapa en lag om reell företagsdemokrati medan man från arbetsgivarhåll var rädd att personalen skulle få ett alltför stort inflytande på beslutsfattandet. En oro fanns också att medbestämmandet skulle byråkratisera beslutsfattandet inom företagen. I dag anser många arbetstagare att samarbetsförfarandet endast används vid massuppsägningar, nedskärningar och rationaliseringar men i övrigt glömts bort. Samarbetsförfarandet och regleringen av det är en viktig och aktuell fråga, eftersom samarbetsförfarandet utgör det forum i samarbetet mellan arbetsgivare och personal där nedskärningar och massuppsägningar diskuteras. Avhandlingens syfte är att analysera hur samarbetsförfarandet fungerar utifrån samarbetslagen. Målet är att granska samarbetsförfarandet ur ett perspektiv där samarbetsförfarandet ses som en potentiell möjlighet och ett verktyg till framgångsrik företagsverksamhet och personalledarskap. I analysen utgår författaren från samarbetslagens 1 §. Syftet med samarbetsförfarandet är att utveckla företagets verksamhet och arbetsförhållanden, att effektivera samarbetet mellan arbetsgivare och personal samt personalens interna samverkan. Samarbetsförfarandet skall utöka personalens möjligheter att påverka behandlingen av ärenden som gäller deras arbete och arbetsplats i enlighet med lagen. Enligt samarbetsförfarandet skall arbetsgivaren rådgöra med arbetstagarna och deras företrädare om de viktigaste ärendena som påverkar personalens ställning och arbetsförhållande innan besluten fattas. Avhandlingen granskar hur lagen tillämpas i dag och hur lagstiftningen kan utvecklas för att effektivera samarbetet i framtiden.