Novel conditional mouse models for targeted kidney research : Emphasis on the analysis of the biological function of nephrin

Nephrin is a transmembrane protein belonging to the immunoglobulin superfamily and is expressed primarily in the podocytes, which are highly differentiated epithelial cells needed for primary urine formation in the kidney. Mutations leading to nephrin loss abrogate podocyte morphology, and result in massive protein loss into urine and consequent early death in humans carrying specific mutations in this gene. The disease phenotype is closely replicated in respective mouse models. The purpose of this thesis was to generate novel inducible mouse-lines, which allow targeted gene deletion in a time and tissue-specific manner. A proof of principle model for succesful gene therapy for this disease was generated, which allowed podocyte specific transgene replacement to rescue gene deficient mice from perinatal lethality. Furthermore, the phenotypic consequences of nephrin restoration in the kidney and nephrin deficiency in the testis, brain and pancreas in rescued mice were investigated. A novel podocyte-specific construct was achieved by using standard cloning techniques to provide an inducible tool for in vitro and in vivo gene targeting. Using modified constructs and microinjection procedures two novel transgenic mouse-lines were generated. First, a mouse-line with doxycycline inducible expression of Cre recombinase that allows podocyte-specific gene deletion was generated. Second, a mouse-line with doxycycline inducible expression of rat nephrin, which allows podocyte-specific nephrin over-expression was made. Furthermore, it was possible to rescue nephrin deficient mice from perinatal lethality by cross-breeding them with a mouse-line with inducible rat nephrin expression that restored the missing endogenous nephrin only in the kidney after doxycycline treatment. The rescued mice were smaller, infertile, showed genital malformations and developed distinct histological abnormalities in the kidney with an altered molecular composition of the podocytes. Histological changes were also found in the testis, cerebellum and pancreas. The expression of another molecule with limited tissue expression, densin, was localized to the plasma membranes of Sertoli cells in the testis by immunofluorescence staining. Densin may be an essential adherens junction protein between Sertoli cells and developing germ cells and these junctions share similar protein assembly with kidney podocytes. This single, binary conditional construct serves as a cost- and time-efficient tool to increase the understanding of podocyte-specific key proteins in health and disease. The results verified a tightly controlled inducible podocyte-specific transgene expression in vitro and in vivo as expected. These novel mouse-lines with doxycycline inducible Cre recombinase and with rat nephrin expression will be useful for conditional gene targeting of essential podocyte proteins and to study in detail their functions in the adult mice. This is important for future diagnostic and pharmacologic development platforms.

A cross-linguistic study of lexical iconicity and its manifestation in bird names

Abstract This dissertation is a cross-linguistic study of lexical iconicity. The study is based on a genealogically stratified sample of 237 languages. The aim is to contribute with an empirical study to the growing dialogue focusing on different forms of lexical iconicity. The conceptual framework of the present study is based on an analysis of types and means of lexical iconicity in the sample languages. Archaeological and cultural evidence are used to tie lexical iconicity to its context. Phenomena related to lexical iconicity are studied both cross-linguistically and language-specifically. The cognitive difference between imitation and symbolism is essential. Lexical iconicity is not only about the iconic relationship between form and referents, but also about how certain iconic properties may become conventional, means used to create sound symbolism. All the sample languages show some evidence of lexical iconicity, demonstrating that it is a universal feature. Nine comparisons of onomatopoeic verbs and nouns, with samples varying between six and 141 languages, show that typologically highly different languages use similar means for creating words based on sound imitation. Two cross-linguistic comparisons of bird names demonstrate that a vast majority of the Eurasian names of the common cuckoo and the world-wide names of crow and raven of the 141 genera are onomatopoeic.

The regional differences between countries in traffic safety: A cross-cultural study and Turkish Case

Road traffic accidents are a large problem everywhere in the world. However, regional differences in traffic safety between countries are considerable. For example, traffic safety records are much worse in Southern Europe and the Middle East than in Northern and Western Europe. Despite the large regional differences in traffic safety, factors contributing to different accident risk figures in different countries and regions have remained largely unstudied. The general aim of this study was to investigate regional differences in traffic safety between Southern European/Middle Eastern (i.e., Greece, Iran, Turkey) and Northern/Western European (i.e., Finland, Great Britain, The Netherlands) countries and to identify factors related to these differences. We conducted seven sub-studies in which I applied a traffic culture framework, including a multi-level approach, to traffic safety. We used aggregated level data (national statistics), surveys among drivers, and data on traffic accidents and fatalities in the analyses. In the first study, we investigated the influence of macro level factors (i.e., economic, societal, and cultural) on traffic safety across countries. The results showed that a high GNP per capita and conservatism correlated with a low number of traffic fatalities, whereas a high degree of uncertainty avoidance, neuroticism, and egalitarianism correlated with a high number of traffic fatalities. In the second, third, and fourth studies, we examined whether the conceptualisation of road user characteristics (i.e., driver behaviour and performance) varied across traffic cultures and how these factors determined overall safety, and the differences between countries in traffic safety. The results showed that the factorial agreement for driver behaviour (i.e., aggressive driving) and performance (i.e., safety skills) was unsatisfactory in Greece, Iran, and Turkey, where the lack of social tolerance and interpersonal aggressive violations seem to be important characteristics of driving. In addition, we found that driver behaviour (i.e., aggressive violations and errors) mediated the relationship between culture/country and accidents. Besides, drivers from "dangerous" Southern European countries and Iran scored higher on aggressive violations and errors than did drivers from "safe" Northern European countries. However, "speeding" appeared to be a "pan-cultural" problem in traffic. Similarly, aggressive driving seems largely depend on road users' interactions and drivers' interpretation (i.e., cognitive biases) of the behaviour of others in every country involved in the study. Moreover, in all countries, a risky general driving style was mostly related to being young and male. The results of the fifth and sixth studies showed that among young Turkish drivers, gender stereotypes (i.e., masculinity and femininity) greatly influence driver behaviour and performance. Feminine drivers were safety-oriented whereas masculine drivers were skill-oriented and risky drivers. Since everyday driving tasks involve not only erroneous (i.e., risky or dangerous driving) or correct performance (i.e., normal habitual driving), but also "positive" driver behaviours, we developed a reliable scale for measuring "positive" driver behaviours among Turkish drivers in the seventh study. Consequently, I revised Reason's model [Reason, J. T., 1990. Human error. Cambridge University Press: New York] of aberrant driver behaviour to represent a general driving style, including all possible intentional behaviours in traffic while evaluating the differences between countries in traffic safety. The results emphasise the importance of economic, societal and cultural factors, general driving style and skills, which are related to exposure, cognitive biases as well as age, sex, and gender, in differences between countries in traffic safety.

Trust, participation and health among ageing people : A cross-sectional cohort study of men and women born in 1926-30, 1936-40, and 1946-50 in the Lahti region, southern Finland

The purpose of the present study was to explore the associations between good self-rated health and economic and social factors in different regions among ageing people in the Päijät-Häme region in southern Finland. The data of this study were collected in 2002 as part of the research and development project Ikihyvä 2002 2012 (Good Ageing in Lahti region GOAL project). The baseline data set consisted of 2,815 participants born in 1926 30, 1936 40, and 1946 50. The response rate was 66 %. According to the previous studies, trust in other people and social participation as the main aspects of social capital are associated with self-rated health. In addition, socioeconomic position (SEP) and self-rated health are associated, but all SEP indicators do not have identical associations with health. However, there is a lack of knowledge of the health associations and regional differences with these factors, especially among ageing people. Regarding these questions, the present study gives new information. According to the results of this study, self-perceived adequacy of income was significantly associated with good self-rated health, especially in the urban areas. Similar associations were found in the rural areas, though education was also considered an important factor. Adequacy of income was an even stronger predictor of good health than the actual income. Women had better self-rated health than men only in the urban areas. The youngest respondents had quite equally better self-rated health than the others. Social participation and access to help when needed were associated with good self-rated health, especially in the urban area and the sparsely populated rural areas. The result was comparable in the rural population centres. The correlation of trust with self-rated health was significant in the urban area. High social capital was associated with good self-rated health in the urban area. The association was quite similar in the other areas, though it was statistically insignificant. High social capital consisted of co-existent high social participation and high trust. The association of traditionalism (low participation and high trust) with self-rated health was also substantial in the urban area. The associations of self-rated health with low social capital (low participation and low trust) and the miniaturisation of community (high participation and low trust) were less significant. From the forms of single participation, going to art exhibitions, theatre, movies, and concerts among women, and studying and self-development among men were positively related to self-rated health. Unexpectedly, among women, active participation in religious events and voluntary work was negatively associated with self-rated health. This may indicate a coping method with ill-health. As a whole, only minor variations in self-rated health were found between the areas. However, the significance of the factors associated with self-rated health varied according to the areas. Economic factors, especially self-perceived adequacy of income was strongly associated with good self-rated health. Also when adjusting for economic and several other background factors social factors (particularly high social capital, social participation, and access to help when needed) were associated with self-rated health. Thus, economic and social factors have a significant relation with the health of the ageing, and improving these factors may have favourable effects on health among ageing people.

Variation in tracheid cross-sectional dimensions and wood viscoelasticity extent and control methods

Printing papers have been the main product of the Finnish paper industry. To improve properties and economy of printing papers, controlling of tracheid cross-sectional dimensions and wood viscoelasticity are examined in this study. Controlling is understood as any procedure which yields raw material classes with distinct properties and small internal variation. Tracheid cross-sectional dimensions, i.e., cell wall thickness and radial and tangential diameters can be controlled with methods such as sorting wood into pulpwood and sawmill chips, sorting of logs according to tree social status and fractionation of fibres. These control methods were analysed in this study with simulations, which were based on measured tracheid cross-sectional dimensions. A SilviScan device was used to measure the data set from five Norway spruce (Picea abies) and five Scots pine (Pinus sylvestris) trunks. The simulation results indicate that the sawmill chips and top pulpwood assortments have quite similar cross-sectional dimensions. Norway spruce and Scots pine are on average also relatively similar in their cross-sectional dimensions. The distributions of these species are somewhat different, but from a practical point of view, the differences are probably of minor importance. The controlling of tracheid cross-sectional dimensions can be done most efficiently with methods that can separate fibres into earlywood and latewood. Sorting of logs or partitioning of logs into juvenile and mature wood were markedly less efficient control methods than fractionation of fibres. Wood viscoelasticity affects energy consumption in mechanical pulping, and is thus an interesting control target when improving energy efficiency of the process. A literature study was made to evaluate the possibility of using viscoelasticity in controlling. The study indicates that there is considerable variation in viscoelastic properties within tree species, but unfortunately, the viscoelastic properties of important raw material lots such as top pulpwood or sawmill chips are not known. Viscoelastic properties of wood depend mainly on lignin, but also on microfibrillar angle, width of cellulose crystals and tracheid cross-sectional dimensions.

The virulence of Finnish Pyrenophora teres f. teres isolates and its implications for resistance breeding

In Finland, barley, Hordeum vulgare L., covers 50 % of the total acreage devoted to cereal cultivation. The most common disease of barley in Finland is net blotch, a foliar disease caused by the ascomycete Pyrenophora teres Drechsler. Disease resistance based on plant genes is an environmentally friendly and economical way to manage plant diseases caused by biotic stresses. Development of a disease resistance breeding programme is dependent on knowledge of the pathogen. In addition to information on the epidemiology and virulence of a pathogen, knowledge on how the pathogen evolves and the nature of the risks that might arise in the future are essential issues that need to be taken into account to achieve the final breeding aims. The main objectives of this study were to establish reliable and efficient testing methods for Pyrenophora teres f. teres virulence screening, and to understand the role of virulence of P. teres f. teres in Finland from a disease resistance breeding point of view. The virulence of P. teres was studied by testing 239 Finnish P. teres f. teres isolates collected between 1994 2007 originating from 19 locations, and 200 P. teres progeny isolates originating from artificially produced P. teres matings. According to the results of this study, screening for P. teres f. teres isolates on barley seedlings under greenhouse conditions is a feasible and cost efficient method to describe the virulence spectrum of the pathogen. Inoculum concentration and the seedling leaf used to gauge virulence had significant effects. Barley grain size, morphological traits of P. teres isolates, spore production and growth rate on agar did not affect the expression of virulence. A common barley differential set to characterize the P. teres virulence was developed and is recommended to be used globally. The virulence spectrum of Finnish P. teres f. teres isolates collected in 1994-2007 was constant both within and between the years. The results indicated differences in the pathogen s aggressiveness and in barley genotypes resistance. However, differences in virulence were rarely significant. Unlike in laboratory conditions, no indications of changes in virulence caused by the sexual reproduction have been observed in Finnish barley fields. In Finland, durable net blotch resistance has been achieved by introducing resistance from other barley varieties using traditional crossing methods, including wide crossing, and testing the breeding material at early generations at several sites under natural infection pressure. Novel resistance is available, which is recommended to minimize the risk of selection of virulent isolates and breakdown of currently deployed resistance.

Protein cross-linking with oxidative enzymes and transglutaminase : Effects in meat protein systems

The effects of tyrosinase, laccase and transglutaminase (TG) were studied in different meat protein systems. The study was focused on the effects of the enzymes on the gel formation properties of myofibrils, and on the textural and water-holding properties of the heated meat systems. The cross-linking efficiency of a novel Trichoderma reesei tyrosinase was compared to that of the commercial Agaricus bisporus tyrosinase. Trichoderma tyrosinase was found to be superior compared to the Agaricus enzyme in its protein cross-linking efficiency and in the incorporation of a small molecule into a complex proteinaceous substrate. Tyrosinase, laccase and TG all polymerised myofibrillar proteins, but laccase was also found to cause protein fragmentation. A positive connection between covalent cross-link and gel formation was observed with tyrosinase and TG. Laccase was able to increase the gel formation only slightly. With an excessive laccase dosage the gel formation declined due to protein fragmentation. Tyrosinase, laccase and TG had different effects on the texture and water-holding of the heated chicken breast meat homogenates. Tyrosinase improved the firmness of the homogenate gels free of phosphate and with a low amount of meat. TG improved the firmness of all studied homogenates. Laccase weakened the gel firmness of the low-meat, low-salt and low-salt/phosphate homogenates and maintained the firmness on the control level in the homogenate free of phosphate. Tyrosinase was the only enzyme capable of reducing the weight loss in the homogenates containing a low amount of meat and a low amount of NaCl. TG was the only enzyme that could positively affect the firmness of the homogenate gel containing both low NaCl and phosphate amounts. In pilot scale the test products were made of coarsely ground chicken breast fillet with a moderate amount of salt. Increasining the amount of meat, salt and TG contents favoured the development of firmness of the test products. The evaporation loss decreased slightly along with increasing TG and NaCl amounts in the experimental conditions used, indicating a positive interaction between these two factors. In this work it was shown that tyrosinase, laccase and TG affected the same myofibrillar proteins, i.e. myosin and troponin T. However, these enzymes had distinguishable effects on the gel formation of a myofibril system as well as on the textural and water-holding properties of the finely ground meat homogenates, reflecting distinctions at least in the reaction mechanisms and target amino acid availability in the protein substrates for these enzymes.

Fatty acids, fibre, carotenoids and tocopherols in relation to glucose metabolism in subjects at high risk for type 2 diabetes : a cross-sectional analysis

Fatty acids, fibre, carotenoids and tocopherols in relation to glucose metabolism in subjects at high risk for type 2 diabetes a cross-sectional analysis Type 2 diabetes (T2D) is a heterogeneous disorder of carbohydrate, lipid and protein metabolism, resulting from genetics, environmental influences and interactions between these. The disease is characterized by insulin resistance, β-cell dysfunction, hepatic glucose overproduction and disordered fat mobilization and storage. The literature on associations between dietary factors and glucose metabolism is inconsistent. One factor behind the discrepant results may be genetic heterogeneity of study populations. Data on nutrient-gene interactions in relation to glucose metabolism are scarce. Thus, investigating high-risk populations and exploring nutrient-gene interactions are essential for improving the understanding of T2D aetiology. Ideally, this information could help to develop prevention programmes that take into account the genetic predisposition to the disease. In this study, associations between measures of glucose metabolism predicting T2D and fatty acids, antioxidative nutrients and fibre were examined in a high-risk population, i.e., in non-diabetic relatives of affected patients. Interactions between the PPARG Pro12Ala polymorphism and fatty acids on glucose metabolism were taken into consideration. This common polymorphism plays an important role in the regulation of glucose metabolism. The inverse associations observed between dietary fibre and insulin resistance are consistent with the prevailing recommendations urging increased intake of fibre to prevent T2D. Beneficial associations observed between the intake of carotenoids and glucose levels stress that a high consumption of vegetables, fruits and berries rich in carotenoids might also play a role in the prevention of T2D. Whether tocopherols have an independent association with glucose metabolism remains questionable. Observed interactions between fatty acids and glucose metabolism suggest that a high intake of palmitic acid is associated with high fasting glucose levels mainly in female Ala allele carriers. Furthermore, the PPARG Pro12Ala polymorphism may modify the metabolic response to dietary marine fat. The beneficial associations of high intake of marine n 3 fatty acids with insulin resistance and glucose levels may be restricted to carriers of the Ala allele. The findings pertain to subjects with a family history of T2D, and the cross-sectional nature of the study precludes inferences about causality. Results nevertheless show that associations of dietary factors with glucose metabolism may be modulated by the genetic makeup of an individual. Additional research is warranted to elucidate the role of probably numerous nutrient-gene interactions, some of which may be sex-specific, in the aetiology of T2D.

Defining Adequate Vitamin D Intake : Cross-sectional and Intervention Studies

D-vitamiini ylläpitää normaalia luun kasvua ja uudistumista koko elämän ajan. Suomessa, kuten monissa muissakin länsimaissa, väestön D-vitamiinitilanne on riittämätön – talvisin osalla jopa puutteellinen. Tässä väitöskirjassa on tutkittu, lisääkö D-vitamiini luumassan kertymistä kasvuiässä, ja ylläpitäkö D-vitamiini luuston tasapainoista aineenvaihduntaa aikuisiällä. Nämä vaikutukset saattavat ehkäisi osteoporoosin kehittymistä eri ikäkausina. Väitöskirjatyössä tutkittiin erisuuruisten D-vitamiinilisäysten vaikutuksia kolmessa eri ikäryhmässä, jotka olivat 11-12 -vuotiaat tytöt (N=228), 21-49 -vuotiaat miehet (N=54) ja 65-85 -vuotiaat naiset (N=52). Tutkittavat satunnaistettiin ryhmiin, jotka nauttivat joko lumevalmistetta tai 5-20 µg D3-vitamiinia vitamiinilisänä. Tutkimukset olivat kaksoissokkoutettuja. Tutkimuksen aikana tutkittavilta otettiin paastoveri- ja virtsanäytteitä. Lisäksi he täyttivät tutkimuslomakkeen taustatietojen kartoittamiseksi sekä frekvenssikyselylomakkeen kalsiumin ja D-vitamiinin saannin selvittämiseksi. Tyttöjen luunmineraalitiheys (BMD) mitattiin DXA–laitteella ja miesten volumetrinen luuntiheys pQCT-menetelmällä. Näytteistä määritettiin mm. seerumin 25-hydroksi-D-vitamiinin (=S-25-OHD), lisäkilpirauhashormonin (=S-PTH) ja luun aineenvaihduntaa kuvaavien merkkiaineiden pitoisuuksia. Murrosikäisten tyttöjen poikkileikkaustutkimuksessa S-25-OHD- ja luun muodostusmerkkiaineen pitoisuudet vaihtelivat kuukausien välillä; suurimmat pitoisuudet mitattiin syyskuussa ja pienimmät maaliskuussa, mikä kuvastaa vuodenaikaisvaihtelua. Vastaava vaihtelu havaittiin lannerangan ja reisiluun BMD:ssä. D-vitamiinilisäyksellä oli myönteinen vaikutus tyttöjen luumassan lisääntymiseen. Suurin D-vitamiinilisä (10 µg/vrk) lisäsi luumassaa 17.2% enemmän reisiluussa ja 12.5% enemmän lannerangassa verrattuna lumevalmistetta nauttivien tyttöjen vastaaviin tuloksiin, mutta tulos riippui hoitomyöntyvyydestä. D-vitamiinin vaikutus luustoon välittyi vähentyneen luun hajotuksen kautta. Tutkimustuloksiin perustuen riittävä D-vitamiinin saanti murrosikäisille tytöille on 15 µg/vrk. D-vitamiinilisän vaikutus 65-85 -vuotiaiden naisten S-25-OHD-pitoisuuteen vakioitui kuudessa viikossa annoksen ollessa 5-20 µg/vrk. Näillä D-vitamiiniannoksilla ei saavutettu tavoiteltavaa S-25-OHD-pitoisuutta, joka on 80 nmol/l. Arvioimme, että 60 nmol/l -pitoisuuden, jota esiintyy kesäisin tämän ikäryhmän suomalaisilla, tämän ikäryhmän naiset saavuttaisivat 24 µg:n päivittäisellä D-vitamiinin saannilla. Terveillä miehillä havaittiin vuodenaikaisvaihtelu S-25-OHD- ja S-PTH-pitoisuudessa sekä luun hajotusta kuvaavassa merkkiainepitoisuudessa. Toisaalta vaihtelua ei havaittu radiuksen volumetrisessä luuntiheydessä eikä luun muodostusmerkkiaineen pitoisuudessa. Vuodenaikaisvaihtelu estettiin 17 µg:n päivittäisellä D-vitamiinin saannilla, mutta tämän ei havaittu vaikuttavan radiuksen luuntiheyteen kuusi kuukautta kestävän tutkimuksen aikana. Yhteenvetona todetaan, että D-vitamiinin saanti on edelleenkin riittämätöntä tutkimusten kohderyhmillä. Tämä näkyy S-25-OHD- ja PTH-pitoisuuden sekä luunaineenvaihduntaa kuvaavien merkkiaineiden vuodenaikaisvaihteluna, mikä on haitallista luuston hyvinvoinnille. D-vitamiinin saantia tulisi lisätä, jotta vähintäänkin riittävä D-vitamiinitilanne (S-25-OHD>50 nmol/l) tai mahdollisesti jopa tavoiteltava D-vitaminitilanne (S-25-OHD≥80 nmol/l) saavutettaisiin. Jotta D-vitamiinin saannin lisääminen olisi kaikissa ikäryhmissä mahdollista, on suunniteltava nykyistä enemmän D-vitamiinilla täydennettyjä elintarvikkeita.

Cross-layer Assisted TCP Algorithms for Vertical Handoff

The ever expanding growth of the wireless access to the Internet in recent years has led to the proliferation of wireless and mobile devices to connect to the Internet. This has created the possibility of mobile devices equipped with multiple radio interfaces to connect to the Internet using any of several wireless access network technologies such as GPRS, WLAN and WiMAX in order to get the connectivity best suited for the application. These access networks are highly heterogeneous and they vary widely in their characteristics such as bandwidth, propagation delay and geographical coverage. The mechanism by which a mobile device switches between these access networks during an ongoing connection is referred to as vertical handoff and it often results in an abrupt and significant change in the access link characteristics. The most common Internet applications such as Web browsing and e-mail make use of the Transmission Control Protocol (TCP) as their transport protocol and the behaviour of TCP depends on the end-to-end path characteristics such as bandwidth and round-trip time (RTT). As the wireless access link is most likely the bottleneck of a TCP end-to-end path, the abrupt changes in the link characteristics due to a vertical handoff may affect TCP behaviour adversely degrading the performance of the application. The focus of this thesis is to study the effect of a vertical handoff on TCP behaviour and to propose algorithms that improve the handoff behaviour of TCP using cross-layer information about the changes in the access link characteristics. We begin this study by identifying the various problems of TCP due to a vertical handoff based on extensive simulation experiments. We use this study as a basis to develop cross-layer assisted TCP algorithms in handoff scenarios involving GPRS and WLAN access networks. We then extend the scope of the study by developing cross-layer assisted TCP algorithms in a broader context applicable to a wide range of bandwidth and delay changes during a handoff. And finally, the algorithms developed here are shown to be easily extendable to the multiple-TCP flow scenario. We evaluate the proposed algorithms by comparison with standard TCP (TCP SACK) and show that the proposed algorithms are effective in improving TCP behavior in vertical handoff involving a wide range of bandwidth and delay of the access networks. Our algorithms are easy to implement in real systems and they involve modifications to the TCP sender algorithm only. The proposed algorithms are conservative in nature and they do not adversely affect the performance of TCP in the absence of cross-layer information.