Ce1-xRuxO2-delta (x=0.05, 0.10): A New High Oxygen Storage Material and Pt, Pd-Free Three-Way Catalyst

Nanocrystalline Ce1-xRuxO2-delta (x = 0.05 and 0.10) of 8-10 nm sizes have been synthesized by hydrothermal method using melamine as complexing agent. Compounds have been characterized by powder X-ray diffraction (XRD), transmission electron microscopy (TEM), X-ray photoelectron spectroscopy (XPS), and energy-dispersive X-ray analysis (EDX) and their structures have been refined by the Rietveld method.The compounds crystallize in fluorite structure and the composition is Ce1-xRuxO2-x/2 where Ru is in +4 state and Ce is in mixed-valence (+3, +4) state. Substitution of Ru4+ ion in CeO2 activated the lattice oxygen. Ce1-xRuxO2-x/2 reversibly releases 0.22[O] and 0.42[O] for x = 0.05 and 0.10, respectively, which is higher than the maximumpossible OSC of 0.22 [O] observed for Ce0.50Zr0.50O2. Utilization of Higher OSC of Ce1-xRuxO2-delta (x = 0.05 and 0.10) is also reflected in terms of low-temperature CO oxidation with these catalysts, both in the presence and absence of feed oxygen. The Ru4+ ion acts as an active center for reducing molecules (CO, hydrocarbon ``HC'') and oxide ion vacancy acts as an active center for O-2 and NO, leading to low-temperature NO conversion to N-2. Thus due to Ru4+ ion, Ce1-xRuxO2-delta is not just a high oxygen storage material but also shows high activity toward CO, hydrocarbon ``HC'' oxidation, and NO reduction by CO at low temperature with high N-2 selectivity for three-way catalysis.

Isolation and Estimation of Cytokinins and Cytokinin-Containing Transfer RNAs from Cucumis

N6-({Delta}2-Isopentenyl) adenosine antibodies were used for the isolation of free cytokinins and cytokinin-containing tRNAs from parts of Cucumis sativus L. var. Guntur seedlings and for the estimation of cytokinins in them. Immobilized N6-({Delta}2-isopentenyl) adenosine antibodies retained tRNAs containing N6-({Delta}2-isopentenyl) adenosine and N6-(4-hydroxy-3-methylbut-2-enyl) adenosine with equal efficiencies. There were at least five cytokinins in the free form in cucumber seedlings. N6-(4-Hydroxy-3-methylbut-2-enyl) adenosine, N6-({Delta}2-isopentenyl) adenosine, and N6-({Delta}2-isopentenyl) adenine were present at least to the extent of 80, 23, and 9 nanograms, respectively, in the cotyledons and 40, 6, and 3 nanograms, respectively, in the decotyledonated seedlings per gram of tissue. Only two cytokinins were found in the tRNAs of cucumber cotyledons, namely N6-({Delta}2-isopentenyl) adenosine and N6-(4-hydroxy-3-methylbut-2-enyl) adenosine in amounts of 12 and 318 nanograms, respectively, per gram of tissue. Immunoaffinity chromatographic analysis of radiolabeled aminoacyl tRNAs from cucumber cotyledons showed that tRNAPhe and tRNATyr contained cytokinins whereas tRNAAla did not.

Living on the edge : Population genetics of Finno-Ugric-speaking humans in North Eurasia

The major aim of this thesis was to examine the origins and distribution of uniparental and autosomal genetic variation among the Finno-Ugric-speaking human populations living in Boreal and Arctic regions of North Eurasia. In more detail, I aimed to disentangle the underlying molecular and population genetic factors which have produced the patterns of uniparental and autosomal genetic diversity in these populations. Among Finno-Ugrics the genetic amalgamation and clinal distribution of West and East Eurasian gene pools were observed within uniparental markers. This admixture indicates that North Eurasia was colonized through Central Asia/ South Siberia by human groups already carrying both West and East Eurasian lineages. The complex combination of founder effects, gene flow and genetic drift underlying the genetic diversity of the Finno-Ugric- speaking populations were emphasized by low haplotype diversity within and among uniparental and biparental markers. A high prevalence of lactase persistence allele among the North Eurasian Finno- Ugric agriculturalist populations was also shown indicating a local adaptation to subsistence change with lactose rich diet. Moreover, the haplotype background of lactase persistence allele among the Finno- Ugric-speakers strongly suggested that the lactase persistence T-13910 mutation was introduced independently more than once to the North Eurasian gene pool. A significant difference in genetic diversity, haplotype structure and LD distribution within the cytochrome P450 CYP2C and CYP2D regions revealed the unique gene pool of the Finno-Ugric Saami created mainly by population genetic processes compared to other Europeans and sub-Saharan Mandenka population. From all studied populations the Saami showed also significantly the highest allele frequency of a CYP2C19 gene mutation causing variable drug reactions. The diversity patterns observed within CYP2C and CYP2D regions emphasize the strong effect of demographic history shaping genetic diversity and LD especially among such small and constant size populations as the Finno-Ugric-speaking Saami. Moreover, the increased LD in Saami due to genetic drift and/or admixture was shown to offer an advantage for further attempts to identify alleles associated to common complex pharmacogenetic traits.

Pain among employees : risk factors and consequences

The overall objective of this study was to gain epidemiological knowledge about pain among employee populations. More specifically, the aims were to assess the prevalence of pain, to identify socio-economic risk groups and work-related psychosocial risk factors, and to assess the consequences in terms of health-related functioning and sickness absence. The study was carried out among the municipal employees of the City of Helsinki. Data comprised questionnaire survey conducted in years 2000-2002 and register data on sickness absence. Altogether 8960 40-60 year old employees participated to the survey (response rate 67%). Pain is common among ageing employees. Approximately 29 per cent of employees reported chronic pain and 15 per cent acute pain, and about seven per cent reported moderately or severely limiting disabling chronic pain. Pain was more common among those with lower level of education or in a low occupational class. -- Psychosocial work environment was associated with pain reports. Job strain, bullying at workplace, and problems in combining work and home duties were associated with pain among women. Among men combining work and home duties was not associated with pain, whereas organizational injustice showed associations. Pain affects functional capacity and predicts sickness absence. Those with pain reported lower level of both mental and physical functioning than those with no pain, physical functioning being more strongly affected than mental. Bodily location of pain or whether pain was acute or chronic had only minor impact on the variation in functioning, whereas the simple count of painful locations was associated with widest variation. Pain accounted for eight per cent of short term (1-3 day) sickness absence spells among men and 13 per cent among women. Of absence spells lasting between four and 14 days pain accounted for 23 per cent among women and 25 per cent among men, corresponding figures for over 14 day absence spells being 37 and 30 per cent. The association between pain and sickness absence was relatively independent of physical and psychosocial work factors, especially among women. The results of this study provide a picture of the epidemiology of pain among employees. Pain is a significant problem that seriously affects work ability. Information on risk groups can be utilized to make prevention measures more effective among those at high risk, and to decrease pain rates and thereby narrow the differences between socio-economic groups. Furthermore, the work-related psychosocial risk factors identified in this study are potentially modifiable, and it should be possible to target interventions on decreasing pain rates among employees.

Familial aggregation of type 1 diabetes and diabetic nephropathy in Finland

Type 1 diabetes (T1D) is a common, multifactorial disease with strong familial clustering. In Finland, the incidence of T1D among children aged 14 years or under is the highest in the world. The increase in incidence has been approximately 2.4% per year. Although most new T1D cases are sporadic the first-degree relatives are at an increased risk of developing the same disease. This study was designed to examine the familial aggregation of T1D and one of its serious complications, diabetic nephropathy (DN). More specifically the study aimed (1) to determine the concordance rates of T1D in monozygotic (MZ) and dizygotic (DZ) twins and to estimate the relative contributions of genetic and environmental factors to the variability in liability to T1D as well as to study the age at onset of diabetes in twins; (2) to obtain long-term empirical estimates of the risk of T1D among siblings of T1D patients and the factors related to this risk, especially the effect of age at onset of diabetes in the proband and the birth cohort effect; (3) to establish if DN is aggregating in a Finnish population-based cohort of families with multiple cases of T1D, and to assess its magnitude and particularly to find out whether the risk of DN in siblings is varying according to the severity of DN in the proband and/or the age at onset of T1D: (4) to assess the recurrence risk of T1D in the offspring of a Finnish population-based cohort of patients with childhood onset T1D, and to investigate potential sex-related effects in the transmission of T1D from the diabetic parents to their offspring as well as to study whether there is a temporal trend in the incidence. The study population comprised of the Finnish Young Twin Cohort (22,650 twin pairs), a population-based cohort of patients with T1D diagnosed at the age of 17 years or earlier between 1965 and 1979 (n=5,144) and all their siblings (n=10,168) and offspring (n=5,291). A polygenic, multifactorial liability model was fitted to the twin data. Kaplan-Meier analyses were used to provide the cumulative incidence for the development of T1D and DN. Cox s proportional hazards models were fitted to the data. Poisson regression analysis was used to evaluate temporal trends in incidence. Standardized incidence ratios (SIRs) between the first-degree relatives of T1D patients and background population were determined. The twin study showed that the vast majority of affected MZ twin pairs remained discordant. Pairwise concordance for T1D was 27.3% in MZ and 3.8% in DZ twins. The probandwise concordance estimates were 42.9% and 7.4%, respectively. The model with additive genetic and individual environmental effects was the best-fitting liability model to T1D, with 88% of the phenotypic variance due to genetic factors. The second paper showed that the 50-year cumulative incidence of T1D in the siblings of diabetic probands was 6.9%. A young age at diagnosis in the probands considerably increased the risk. If the proband was diagnosed at the age of 0-4, 5-9, 10-14, 15 or more, the corresponding 40-year cumulative risks were 13.2%, 7.8%, 4.7% and 3.4%. The cumulative incidence increased with increasing birth year. However, SIR among children aged 14 years or under was approximately 12 throughout the follow-up. The third paper showed that diabetic siblings of the probands with nephropathy had a 2.3 times higher risk of DN compared with siblings of probands free of nephropathy. The presence of end stage renal disease (ESRD) in the proband increases the risk three-fold for diabetic siblings. Being diagnosed with diabetes during puberty (10-14) or a few years before (5-9) increased the susceptibility for DN in the siblings. The fourth paper revealed that of the offspring of male probands, 7.8% were affected by the age of 20 compared with 5.3% of the offspring of female probands. Offspring of fathers with T1D have 1.7 times greater risk to be affected with T1D than the offspring of mothers with T1D. The excess risk in the offspring of male fathers manifested itself through the higher risk the younger the father was when diagnosed with T1D. Young age at onset of diabetes in fathers increased the risk of T1D greatly in the offspring, but no such pattern was seen in the offspring of diabetic mothers. The SIR among offspring aged 14 years or under remained fairly constant throughout the follow-up, approximately 10. The present study has provided new knowledge on T1D recurrence risk in the first-degree relatives and the risk factors modifying the risk. Twin data demonstrated high genetic liability for T1D and increased heritability. The vast majority of affected MZ twin pairs, however, remain discordant for T1D. This study confirmed the drastic impact of the young age at onset of diabetes in the probands on the increased risk of T1D in the first-degree relatives. The only exception was the absence of this pattern in the offspring of T1D mothers. Both the sibling and the offspring recurrence risk studies revealed dynamic changes in the cumulative incidence of T1D in the first-degree relatives. SIRs among the first-degree relatives of T1D patients seems to remain fairly constant. The study demonstrates that the penetrance of the susceptibility genes for T1D may be low, although strongly influenced by the environmental factors. Presence of familial aggregation of DN was confirmed for the first time in a population-based study. Although the majority of the sibling pairs with T1D were discordant for DN, its presence in one sibling doubles and presence of ESRD triples the risk of DN in the other diabetic sibling. An encouraging observation was that although the proportion of children to be diagnosed with T1D at the age of 4 or under is increasing, they seem to have a decreased risk of DN or at least delayed onset.

Obesity, Smoking and Dieting

Lihavuus ja ylipaino ovat viime vuosikymmeninä yleistyneet; jo yli puolet länsimaiden väestöstä on ylipainoisia ja viidennes lihavia. Varsinkin nuorilla ylipainon lisääntyminen on ollut nopeaa. Ylipaino, erityisesti yhdistettynä vyötärölihavuuteen, sekä tupakointi lisäävät sairastavuutta sydän- ja verisuonisairauksiin, metabolisiin sairauksiin, kuten diabetekseen, sekä moniin syöpiin. Lihavuus ja tupakointi ovatkin kehittyneiden maiden tärkeimpiä ehkäistävissä olevia kuolinsyitä. Samanaikaisesti ylipainon kanssa laihduttaminen ja jopa terveydelle haitalliset laihdutusmenetelmät, kuten tupakointi painonhallintakeinona on tullut yhä yleisemmäksi. Nopeaan painonpudotukseen tähtäävällä laihduttamisella on usein terveydelle haitallisia seurauksia kuten painon nousu yli alkuperäisen painon ja kehon rasvajakauman muuttuminen epäterveellisemmäksi. Kolme neljännestä merkittävästi laihduttaneista kertoo painon nousseen takaisin. Tupakoinnin ja toistuvan laihduttamisen vaikutukset ylipainon ja lihavuuden kehittymiselle kytkeytyvät toisiinsa. Tässä väitöskirjatyössä tutkittiin toistuvan laihduttamisen ja tupakoinnin vaikutusta kehon painoon ja lisäksi tupakoinnin vaikutusta vyötärölihavuuden kehittymiseen. Työn toisena tavoitteena oli tutkia, kuinka voimakkaasti tupakointi ja toistuva laihduttaminen liittyvät toisiinsa suomalaisilla ja onko tämä yhteys erilainen eri ikäryhmissä ja sukupuolilla. Työ perustuu kolmeen laajaan kyselyaineistoon: Nuorten Kaksosten Terveystutkimuksen (englanniksi FinnTwin16) aineistossa on seurattu 1975-79 syntyneitä kaksosia 16, 17, 18 ja 24 vuoden ikäisinä (N=5563). Suomen kaksoskohortin aineisto (N= 12 793) on kerätty vuonna 1990 samaa sukupuolta olevilta, vuosina 1930-57 syntyneiltä kaksosilta. Entisten huippu-urheilijoiden (N=1838) ja heille kaltaistettujen verrokkien (N=834) seurantatutkimuksessa tiedot on kerätty vuosina 1985, 1995 ja 2001. Pituus, paino ja tupakointi on kysytty kaikissa kyselyissä. Kaksoset vastasivat laihdutuskäyttäytymistä koskeviin kysymyksiin. Urheilijoiden laihdutuskäyttäytyminen pääteltiin lajin perusteella, sillä toistuvan laihduttamisen tiedetään olevan yleistä painoluokissa urheilevilla urheilijoilla (esim.painijat, nyrkkeilijät). Nuoruusiän tupakointi ennusti vyötärölihavuutta molemmilla sukupuolilla ja lisäksi ylipainoisuutta naisilla. Toistuva laihduttaminen oli yhteydessä myöhempään painonnousuun ja lihavuuteen miehillä. Lisäksi toistuvan laihduttamisen ja tupakoinnin todettiin liittyvän toisiinsa nuorilla aikuisilla. Vanhemmissa ikäluokissa miehet, jotka tupakoivat, laihduttivat harvemmin kuin tupakoimattomat. Lihavuuteen ja vyötärölihavuuteen liittyvän oheissairastavuuden ennaltaehkäisyssä tupakoinnin ja toistuvan laihduttamisen vähentäminen saattavat olla aiemmin luultua tehokkaampia keinoja.

Early growth and adult health: focus on blood pressure, glucose tolerance status and body composition

Theory of developmental origins of adult health and disease proposes that experiences during critical periods of early development may have consequences on health throughout a lifespan. Thesis studies aimed to characterize associations between early growth and some components of the metabolic syndrome cluster. Participants belong to two epidemiological cohorts with data on birth measurements and, for the younger cohort, on serial recordings of weight and height during childhood. They were born as singletons between 1924-33 and 1934-44 in the Helsinki University Central Hospital, and 500 and 2003 of them, respectively, attended clinical studies at the age of 65-75 and 56-70 years, respectively. In the 65-75 year old men and women, the well-known inverse relationship between birth weight and systolic blood pressure (SBP) was confined to people who had established hypertension. Among them a 1-kg increase in birth weight was associated with a 6.4-mmHg (95% CI: 1.0 to 11.9) decrease in SBP. This relationship was further confined to people with the prevailing Pro12Pro polymorphism of the peroxisome proliferator-activated receptor-γ2 (PPARγ2) gene. People with low birth weight were more likely to receive angiotensin-converting enzyme inhibitors/angiotensin-receptor blockers (ACEI/ARB, p=0.03), and, again, this relationship was confined to the carriers of the Pro12Pro (p=0.01 for interaction). These results suggest that the inverse association between birth weight and systolic BP becomes focused in hypertensive people because pathological features of BP regulation, associated with slow fetal growth, become self-perpetuating in adult life. Insulin resistance of the Pro12Pro carriers with low birth weight may interact with the renin-angiotensin system leading to raised BP levels. Habitual physical activity protected men and women who were small at birth, and thus at increased risk for the development of type 2 diabetes, against glucose intolerance more strongly. Among subjects with birth weight ≤3000 g, the odds ratio (OR) for glucose intolerance was 5.2 (95% CI: 2.1 to 13) in those who exercised less than 3 times per week compared to regular exercisers; in those who scored their exercise light compared with moderate exercisers (defined as comparable to brisk walking) the OR was 3.5 (1.5 to 8.2). In the 56-70 year old men a 1 kg increase in birth weight corresponded to a 4.1 kg (95% CI: 3.1 to 5.1) and in women to a 2.9 kg (2.1 to 3.6) increase in adult lean mass. Rapid gain in body mass index (BMI), i.e. crossing from an original BMI percentile to a higher one, before the age of 2 years increased adult lean mass index (LMI, lean mass/height squared) without excess fat accumulation whereas rapid gain in BMI during later childhood, despite the concurrent rise in LMI, resulted in a relatively higher increase in adult body fat mass. These findings illustrate how genes, the environment and their interactions, early growth patterns, and adult lifestyle modify adult health risks which originate from early life.

Mast cells and endothelial erosion: Implications for human atherothrombotic disease

More than 40% of all deaths in Finland are caused by atherosclerosis. The complications of atherosclerosis are due to either detachment of the luminal endothelium (erosion) or rupture of the fibrous cap of an atherosclerotic plaque (rupture). As a result, a thrombus is formed at the site of the intimal lesion. Indeed, erosions cause roughly 40% of sudden atherothrombotic deaths and 25% of all atherothrombotic deaths. Erosions are overrepresented in young subjects, diabetics, smokers and women. This dissertation focuses on endothelial erosion. Endothelial erosions were studied in the context of arterial grafting and vascular inflammation. Special attention was given to the role of intimal mast cells and the methodological viewpoints of reliable identification of endothelial erosions. Mast cells are inflammatory cells mostly known for their ability to cause allergic symptoms. In addition to occurring in skin and mucosal surfaces, mast cells are abundant in arterial intima and adventitia. In this study, mast cells were found to associate with endothelial erosions in non-lesional and atherosclerotic human coronary arteries. Thus, mast cells may participate in atherogenesis at the initial phases of the disease process already. We also showed that the mast cell proteases tryptase, chymase, and cathepsin G are all capable of cleaving molecules essential for endothelial cell-to-cell and cell-to-extracellular matrix interactions, such as VE-cadherin and fibronectin. Symptom-causing carotid plaques were found to contain more inflammatory cells, especially mast cells, than non-symptom-causing plaques. Furthermore, the atherogenic serum lipid profile and the degree of carotid stenosis turned out to correlate with the density of carotid plaque mast cells. Apoptotic and proliferating cells were more abundant in non-symptom causing plaques (active renewal of endothelial cells), but erosions were larger in symptom-causing plaques (capacity of endothelial regeneration exceeded). The process of identifying endothelial erosions with immunostainings has been ambiguous, since both endothelial cells and platelets express largely the same antigens. This may have caused inaccurate interpretations of the presence of endothelial erosion. In the last substudy of this thesis we developed a double immunostaining method for simultaneous identification of endothelial cells and platelets. This method enables more reliable identification of endothelial erosions.

Perspectives on prescribing in nursing homes in Helsinki

Prescribing for older patients is challenging. The prevalence of diseases increases with advancing age and causes extensive drug use. Impairments in cognitive, sensory, social and physical functioning, multimorbidity and comorbidities, as well as age-related changes in pharmacokinetics and pharmacodynamics all add to the complexity of prescribing. This study is a cross-sectional assessment of all long-term residents aged ≥ 65 years in all nursing homes in Helsinki, Finland. The residents’ health status was assessed and data on their demographic factors, health and medications were collected from their medical records in February 2003. This study assesses some essential issues in prescribing for older people: psychotropic drugs (Paper I), laxatives (Paper II), vitamin D and calcium supplements (Paper III), potentially inappropriate drugs for older adults (PIDs) and drug-drug interactions (DDIs)(Paper IV), as well as prescribing in public and private nursing homes. A resident was classified as a medication user if his or her medication record indicated a regular sequence for its dosage. Others were classified as non-users. Mini Nutritional Assessment (MNA) was used to assess residents’ nutritional status, Beers 2003 criteria to assess the use of PIDs, and the Swedish, Finnish, INteraction X-referencing database (SFINX) to evaluate their exposure to DDIs. Of all nursing home residents in Helsinki, 82% (n=1987) participated in studies I, II, and IV and 87% (n=2114) participated in the study III. The residents’ mean age was 84 years, 81% were female, and 70% were diagnosed with dementia. The mean number of drugs was 7.9 per resident; 40% of the residents used ≥ 9 drugs per day, and were thus exposed to polypharmacy. Eighty percent of the residents received psychotropics; 43% received antipsychotics, and 45% used antidepressants. Anxiolytics were prescribed to 26%, and hypnotics to 28% of the residents. Of those residents diagnosed with dementia, 11% received antidementia drugs. Fifty five percent of the residents used laxatives regularly. In multivariate analysis, those factors associated with regular laxative use were advanced age, immobility, poor nutritional status, chewing problems, Parkinson’s disease, and a high number of drugs. Eating snacks between meals was associated with lower risk for laxative use. Of all participants, 33% received vitamin D supplementation, 28% received calcium supplementation, and 20% received both vitamin D and calcium. The dosage of vitamin D was rather low: 21% received vitamin D 400 IU (10 µg) or more, and only 4% received 800 IU (20 µg) or more. In multivariate analysis, residents who received vitamin D supplementation enjoyed better nutritional status, ate snacks between meals, suffered no constipation, and received regular weight monitoring. Those residents receiving PIDs (34% of all residents) more often used psychotropic medication and were more often exposed to polypharmacy than residents receiving no PIDs. Residents receiving PIDs were less often diagnosed with dementia than were residents receiving no PIDs. The three most prevalent PIDs were short-acting benzodiazepine in greater dosages than recommended, hydroxyzine, and nitrofurantoin. These three drugs accounted for nearly 77% of all PID use. Of all residents, less than 5% were susceptible to a clinically significant DDI. The most common DDIs were related to the use of potassium-sparing diuretics, carbamazepine, and codeine. Residents exposed to potential DDIs were younger, had more often suffered a previous stroke, more often used psychotropics, and were more often exposed to PIDs and polypharmacy than were residents not exposed to DDIs. Residents in private nursing homes were less often exposed to polypharmacy than were residents in public nursing homes. Long-term residents in nursing homes in Helsinki use, on average, nearly eight drugs daily. The use of psychotropic drugs in our study was notably more common than in international studies. The prevalence of laxatives equaled other prior international studies. Regardless of the known benefit and recommendation of vitamin D supplementation for elderly residing mostly indoors, the proportion of nursing home residents receiving vitamin D and calcium was surprisingly low. The use of PIDs was common among nursing home residents. PIDs increased the likelihood of DDIs. However, DDIs did not seem a major concern among the nursing home population. Monitoring PIDs and potential drug interactions could improve the quality of prescribing.

Physical activity, fitness, abdominal obesity, and cardiovascular risk factors in Finnish men and women : The National FINRISK 2002 Study

Physical inactivity, low cardiorespiratory fitness, and abdominal obesity are direct and mediating risk factors for cardiovascular disease (CVD). The results of recent studies suggest that individuals with higher levels of physical activity or cardiorespiratory fitness have lower CVD and all-cause mortality than those with lower activity or fitness levels regardless of their level of obesity. The interrelationships of physical activity, fitness, and abdominal obesity with cardiovascular risk factors have not been studied in detail. The aim of this study was to investigate the associations of different types of leisure time physical activity and aerobic fitness with cardiovascular risk factors in a large population of Finnish adults. In addition, a novel aerobic fitness test was implemented and the distribution of aerobic fitness was explored in men and women across age groups. The interrelationships of physical activity, aerobic fitness and abdominal obesity were examined in relation to cardiovascular risk factors. This study was part of the National FINRISK Study 2002, which monitors cardiovascular risk factors in a Finnish adult population. The sample comprised 13 437 men and women aged 25 to 74 years and was drawn from the Population Register as a stratified random sample according to 10-year age groups, gender and area. A separate physical activity study included 9179 subjects, of whom 5 980 participated (65%) in the study. At the study site, weight, height, waist and hip circumferences, and blood pressure were measured, a blood sample was drawn, and an aerobic fitness test was performed. The fitness test estimated maximal oxygen uptake (VO2max) and was based on a non-exercise method by using a heart rate monitor at rest. Waist-to-hip ratio (WHR) was calculated by dividing waist circumference with hip circumference and was used as a measure of abdominal obesity. Participants filled in a questionnaire on health behavior, a history of diseases, and current health status, and a detailed 12-month leisure time physical activity recall. Based on the recall data, relative energy expenditure was calculated using metabolic equivalents, and physical activity was divided into conditioning, non-conditioning, and commuting physical activity. Participants aged 45 to 74 years were later invited to take part in a 2-hour oral glucose tolerance test with fasting insulin and glucose measurements. Based on the oral glucose tolerance test, undiagnosed impaired glucose tolerance and type 2 diabetes were defined. The estimated aerobic fitness was lower among women and decreased with age. A higher estimated aerobic fitness and a lower WHR were independently associated with lower systolic and diastolic blood pressure, lower total cholesterol and triglyceride levels, and with higher high-density lipoprotein (HDL) cholesterol and HDL to total cholesterol ratio. The associations of the estimated aerobic fitness with diastolic blood pressure, triglycerides, and HDL to total cholesterol ratio were stronger in men with a higher WHR. High levels of conditioning and non-conditioning physical activity were associated with lower high-sensitivity C-reactive protein (CRP) levels. High levels of conditioning and overall physical activities were associated with lower insulin and glucose levels. The associations were stronger among women than men. A better self-rated physical fitness was associated with a higher estimated aerobic fitness, lower CRP levels, and lower insulin and glucose levels in men and women. In each WHR third, the risk of impaired glucose tolerance and type 2 diabetes was higher among physically inactive individuals who did not undertake at least 30 minutes of moderate-intensity physical activity on five days per week. These cross-sectional data show that higher levels of estimated aerobic fitness and regular leisure time physical activity are associated with a favorable cardiovascular risk factor profile and that these associations are present at all levels of abdominal obesity. Most of the associations followed a dose-response manner, suggesting that already low levels of physical activity or fitness are beneficial to health and that larger improvements in risk factor levels may be gained from higher activity and fitness levels. The present findings support the recommendation to engage regularly in leisure time physical activity, to pursue a high level of aerobic fitness, and to prevent abdominal obesity.

Implications of co-twin dependence for twins' social interactions, mental health and alcohol use : A follow-up study of Finnish twins from adolescence to early adulthood

Objective: The aim of the present study was to examine co-twin dependence and its impact on twins' social contacts, leisure-time activities and psycho-emotional well-being. The role of co-twin dependence was also examined as a moderator of genetic and environmental influences on alcohol use in adolescence and in early adulthood. Methods: The present report is based on the Finnish Twin Cohort Study (FinnTwin16), a population-based study of five consecutive birth cohorts of Finnish twins born in the years 1975-1979. Baseline assessments were collected through mailed questionnaires, within two months of the twins' sixteenth birthday yielding replies from 5563 twin individuals. All respondent twins were sent follow-up questionnaires at ages of 17, 18½, and in early adulthood, when twins were 22-27 years old. Measures: The questionnaires included a survey of health habits and attitudes, a symptom checklist and questions about twins' relationships with parents, peers and co-twin. Measures used were twins' self-reports of their own dependence and their co-twin's dependence at age 16, reports of twins' leisure-time activities and social contacts, alcohol use, psychological distress and somatic symptoms both in adolescence and in early adulthood. Results: In the present study 25.6% of twins reported dependence on their co-twin. There were gender and zygosity differences in dependence, females and MZ twins were more likely to report dependence than males and DZ twins. Co-twin dependence can be viewed on one hand as an individual characteristic, but on the other hand as a pattern of dyadic interaction that is mutually regulated and reciprocal. Most of the twins (80.7%) were either concordantly co-twin dependent or concordantly co-twin independent. The associations of co-twin dependence with twins' social interactions and psycho-emotional characteristics were relatively consistent both in adolescence and in early adulthood. Dependence was related to higher contact frequency and a higher proportion of shared leisure-time activities between twin siblings at the baseline and the follow-up. Additionally co-twin dependence was associated with elevated levels of psycho-emotional distress and somatic complaints, especially in adolescence. In the framework of gene-environment interaction, these results suggest that the genetic contribution to individual differences in drinking patterns is dependent on the nature of the pair-wise relationship of twin siblings. Conclusions: The results of this study indicate that co-twin dependence is a genuine feature of the co-twin relationship and shows the importance of studying the impact of various features of co-twin relationships on individual twins' social and psycho-emotional life and well-being. Our study also offers evidence that differences in inter-personal relationships contribute to the effects of genetic propensities.

Statistical analysis of the associations of hereditary factors with the risk of Type 1 diabetes and Diabetic Nephropathy based on the familial data collected through ascertaiment from population-based registers

In genetic epidemiology, population-based disease registries are commonly used to collect genotype or other risk factor information concerning affected subjects and their relatives. This work presents two new approaches for the statistical inference of ascertained data: a conditional and full likelihood approaches for the disease with variable age at onset phenotype using familial data obtained from population-based registry of incident cases. The aim is to obtain statistically reliable estimates of the general population parameters. The statistical analysis of familial data with variable age at onset becomes more complicated when some of the study subjects are non-susceptible, that is to say these subjects never get the disease. A statistical model for a variable age at onset with long-term survivors is proposed for studies of familial aggregation, using latent variable approach, as well as for prospective studies of genetic association studies with candidate genes. In addition, we explore the possibility of a genetic explanation of the observed increase in the incidence of Type 1 diabetes (T1D) in Finland in recent decades and the hypothesis of non-Mendelian transmission of T1D associated genes. Both classical and Bayesian statistical inference were used in the modelling and estimation. Despite the fact that this work contains five studies with different statistical models, they all concern data obtained from nationwide registries of T1D and genetics of T1D. In the analyses of T1D data, non-Mendelian transmission of T1D susceptibility alleles was not observed. In addition, non-Mendelian transmission of T1D susceptibility genes did not make a plausible explanation for the increase in T1D incidence in Finland. Instead, the Human Leucocyte Antigen associations with T1D were confirmed in the population-based analysis, which combines T1D registry information, reference sample of healthy subjects and birth cohort information of the Finnish population. Finally, a substantial familial variation in the susceptibility of T1D nephropathy was observed. The presented studies show the benefits of sophisticated statistical modelling to explore risk factors for complex diseases.

Palladin, a novel microfilament protein

Palladin is a novel actin microfilament associated protein, which together with myotilin and myopalladin forms a novel cytoskeletal IgC2 domain protein family. Whereas the expression of myotilin and myopalladin is limited mainly to striated muscle, palladin is widely expressed in both epithelial and mesenchymal tissues, including heart and the nervous system. Palladin has a complex genetic structure and it is expressed as several different sized and structured splice variants, which also display differences in their expression pattern and interactions. In muscle cells, all the family members localize to the sarcomeric Z-disc, and in non-muscle cells palladin also localizes to the stress-fiber-dense regions, lamellipodia, podosomes and focal adhesions. A common feature of this protein family is the binding to α-actinin, but other interactions are mostly unique to each member. Palladin has been shown to interact with several proteins, including VASP, profilin, Eps8, LASP-1 and LPP. Its domain structure, lack of enzymatic activity and multiple interactions define it as a molecular scaffolding protein, which links together proteins with different functional modalities into large complexes. Palladin has an important role in cytoskeletal regulation, particularly in stress fiber formation and stabilization. This assumption is supported by several experimental results. First, over-expression of palladin in non-muscle cells results in rapid reorganization of the actin cytoskeleton and formation of thick actin bundles. Second, the knock-down of palladin with anti-sense and siRNA techniques or knock-out by genetic methods leads to defective stress fiber formation. Furthermore, palladin is usually up-regulated in situations requiring a highly organized cytoskeleton, such as differentiation of dendritic cells, trophoblasts and myofibroblasts, and activation of astrocytes during glial scar formation. The protein family members have also direct disease linkages; myotilin missense mutations are the cause of LGMD1A and myofibrillar myopathy. Palladin mutations and polymorphisms, on the other hand, have been linked to hereditary pancreatic cancer and myocardial infarction, respectively. In this study we set out to characterize human palladin. We identified several palladin isoforms, studied their tissue distribution and sub-cellular localization. Four novel interaction partners were identified; ezrin, ArgBP2, SPIN90 and Src-kinase.The previously identified interaction between palladin and α-actinin was also characterized in detail. All the identified new binding partners are actin cytoskeleton associated proteins; ezrin links the plasma membrane to the cytoskeleton, ArgBP2 and SPIN90 localize, among other structures, to the lamellipodia and in cardiomyocytes to the Z-disc. Src is a transforming tyrosine kinase, which besides its role in oncogenesis has also important cytoskeletal associations. We also studied palladin in myofibroblasts, which are specialized cells involved in diverse physiological and pathological processes, such as wound healing and tissue fibrosis. We demonstrated that palladin is up-regulated during the differentiation of myofibroblasts in an isoform specific manner, and that this up-regulation is induced by TGF-β via activation of both the SMAD and MAPK signalling cascades. In summary, the results presented here describe the initial characterization of human palladin and offer a basis for further studies.

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.

Development of regulatory T cells in the human thymus

The role of the immune system is to protect an organism against pathogens while maintaining tolerance against self. T cells are an essential component of the immune system and they develop in the thymus. The AIRE (autoimmune regulator) gene product plays an important role in T cell development, as it promotes expression of peripheral tissue antigens in the thymus. Developing T cells, thymocytes, which recognize self-antigens with high affinity are deleted. However, this deletion process is not perfect and not all autoreactive T cells are destroyed. When the distinction between self and non-self fails, tolerance breaks and the immune system attacks the host s own tissues. This results in autoimmunity. Regulatory T cells contribute to the maintenance of self-tolerance. They can actively suppress the function of autoreactive cells. Several populations of cells with regulatory properties have been described, but the best characterized population is the natural regulatory T cells (Treg cells), which develop in the thymus and express the transcription factor FOXP3. The thymic development of Treg cells in humans is the subject of this thesis. Thymocytes at different developmental stages were analyzed using flow cytometry. The CD4-CD8- double-negative (DN) thymocytes are the earliest T cell precursors in the T cell lineage. My results show that the Treg cell marker FOXP3 is up-regulated already in a subset of these DN thymocytes. FOXP3+ cells were also found among the more mature CD4+CD8+ double-positive (DP) cells and among the CD4+ and CD8+ single-positive (SP) thymocytes. The different developmental stages of the FOXP3+ thymocytes were isolated and their gene expression examined by quantitative PCR. T cell receptor (TCR) repertoire analysis was used to compare these different thymocyte populations. My data show that in humans commitment to the Treg cell lineage is an early event and suggest that the development of Treg cells follows a linear developmental pathway, FOXP3+ DN precursors evolving through the DP stage to become mature CD4+ Treg cells. Most T cells have only one kind of TCR on their cell surface, but a small fraction of cells expresses two different TCRs. My results show that the expression of two different TCRs is enriched among Treg cells. Furthermore, both receptors were capable of transmitting signals when bound by a ligand. By extrapolating flow cytometric data, it was estimated that the majority of peripheral blood Treg cells are indeed dual-specific. The high frequency of dual-specific cells among human Treg cells suggests that dual-specificity has a role in directing these cells to the Treg cell lineage. It is known that both genetic predisposition and environmental factors influence the development of autoimmunity. It is also known that the dysfunction or absence of Treg cells leads to the development of autoimmune manifestations. APECED (autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy) is a rare monogenic autoimmune disease, caused by mutations in the AIRE gene. In the absence of AIRE gene product, deletion of self-specific T cells is presumably disturbed and autoreactive T cells escape to the periphery. I examined whether Treg cells are also affected in APECED. I found that the frequency of FOXP3+ Treg cells and the level of FOXP3 expression were significantly lower in APECED patients than in controls. Additionally, when studied in cell cultures, the suppressive capacity of the patients' Treg cells was impaired. Additionally, repertoire analysis showed that the TCR repertoire of Treg cells was altered. These results suggest that AIRE contributes to the development of Treg cells in humans and the selection of Treg cells is impaired in APECED patients. In conclusion, my thesis elucidates the developmental pathway of Treg cells in humans. The differentiation of Tregs begins early during thymic development and both the cells dual-specificity and AIRE probably affect the final commitment of Treg cells.