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Recently, our research group together with the Hematology Research Unit Helsinki, found that 31 of 77 patients (40%) with T-cell large granular lymphocytic leukemia have somatic point mutations in the Src Homology 2 (SH2) domain of the STAT3 gene. LGL leukemia is a rare and indolent disease characterized by the clonal expansion of large granular lymphocytes of unknown etiology. The aim of this master's thesis study was to elucidate whether the identified Y640F and D661V mutations can cause hyperactivity of the STAT3 protein and excessive proliferation of T-cells. STAT3 is a transcription factor that is known to have a key role in cell proliferation and apoptosis, and which is activated by the phosphorylation of receptor-associated kinases. The phosphorylation of tyrosine residue 705 in STAT3 induces dimerization and localization of the STAT3 dimer to the nucleus. In several cancers STAT3 protein has been reported to be constitutively active. Furthermore, previously published data strongly support our hypothesis that the mutations identified in LGL patients might cause STAT3 to be hyperactive resulting in inhibition of apoptotic pathways in cytotoxic T cells. In order to evaluate the function of the novel mutations, expression constructs of STAT3 containing the D661V and Y640F mutations were generated. In addition, lentiviral vectors were produced to establish a T-cell line (Jurkat) with stable expression of mutant STAT3. After the successful generation of STAT3 constructs and cell line models, several functional assays were performed. Transcriptional activity of STAT3 was measured by luciferase reporter assay and immunocytochemistry was used to determine whether the mutations promote nuclear localization of STAT3. STAT3 phosphorylation was examined by immunoblotting. In addition, quantitative RT-PCR was used to detect differential expression of five STAT3 target genes from patient samples. The results, particularly the luciferase reporter assay, indicated a significant difference between the mutant and wild type STAT3 providing strong evidence that the STAT3 mutants are transcriptionally more active. The localization assay, imaged by fluorescence microscopy, showed more STAT3 D661V and Y640F protein present in the nucleus when compared to wild type STAT3. However, the proliferation rate of mutant STAT3 expressing Jurkat cells was not increased. In addition, STAT3 target gene expression levels of two patient samples did not show large differences when compared to healthy LGL cells. As a result of these findings, it can be strongly hypothesized that aberrant STAT3 signaling underlies the cause of T-cell LGL leukemia. Understanding the molecular basis of LGL leukemia is important in order to develop diagnostic and therapeutic strategies for patients suffering from the disease. Since constitutively active STAT3 is common amongst many cancers and autoimmune disorders, activating mutations could possibly be found in these diseases. More careful sequencing studies of STAT3 upstream molecules are warranted as well, and will be performed in the future from leukemic LGL samples.

There is a need to find more sustainable sources of protein due to increasing demand and population growth. Dry milling and air classification yields oat protein concentrate (OPC, 45.5% protein) from defatted oat kernels. However, applications of oat protein in foods require improved technological functionalities. The literature review of this study discusses the chemical composition of oats, and modifications of plant protein through thermal treatment, microfluidizer, and high hydrostatic pressure (HHP). The experimental work aimed to understand the effect of heat treatment (40, 60, 80 °C), microfluidization (50 MPa), and high hydrostatic pressure (HPP, 300 and 600 MPa) on the applicability of OPC in semi-solid food matrices. The treatments were performed on OPC suspensions with 2.4, 4.6, 8.5, 12, or 20 % protein. Changes in particle size, protein solubility, surface hydrophobicity, and rheological properties were measured. Stability of OPC dispersion was improved by microfluidizer treatment while HHP at 600 MPa accelerated sedimentation. Viscosity increased significantly by thermal treatment at 80 °C and HHP at 600 MPa. Gel formation occurred at 12 and 20% protein concentration with 80 °C heat treatment. Highest increase in particle size was shown by HHP at 600 MPa resulting in volume mean diameter, d4,3 of 32-38 µm. On the other hand, microfluidizer reduced d4,3 to 3.3-3.4 µm. Protein solubility increased from 13 to 23% by increasing protein concentration from 2.4% to 20%. Protein solubility was improved by 1-1.7% at 2.4-8.5% protein concentration by thermal treatment at 60 °C, homogenization, and HHP at 300 MPa. Protein solubility was reduced by HHP at 600 MPa by 1.4-1.5 %. Protein surface hydrophobicity was increased by thermal treatment at 60 and 80 °C and HHP. Microfluidizer showed slight increase in solubility at 4.6% protein concentration (1.1-1.6 %) and lowered surface hydrophobicity at 8.5% protein concentration. Thermal treatment showed best result in achieving gel-like structure, whereas microfluidization showed potential for stabilising dispersion.

Lynch syndrome (LS) is the most common cancer predisposition disease caused by dominantly inherited pathogenic variant (PV) of a mismatch repair (MMR) gene leading to a defective gene allele. The four major MMR genes encode MMR proteins – MSH2, MSH6, MLH1 ja PMS2 – that participate in the proofreading and repairing of the daughter strand for mismatches after every replication. The inherited PVs predispose to cancer development as only one somatic allele loss is required for biallelic loss according to the Knudson’s “two-hit” hypothesis. The biallelic loss of an MMR-gene leads to disrupted protein function altering the MMR process. When mismatches are left unrepaired, genomic instability is caused, which can eventually lead to tumorigenesis. Especially, the risk of colorectal cancer (CRC) and endometrial cancer (EC) is increased in LS. The predisposition syndrome, LS, is important to detect as early as possible to decrease the risk of cancer by prevention and surveillance. The MMR genes and their defects vary in their consequences to the repair process considerably, and thus, it is crucial to know the different characteristics and functional effects of them when estimating the level of cancer risk. Variants of uncertain significance (VUS) are especially prevalent among LS variants. More information about their impact to the disease can be acquired by in vitro and in silico methods, for instance. The main goal of the efforts for early detection and prevention is to reduce cancer morbidity and mortality. In this thesis, the pathogenicities of MSH2 and MSH6 variants were studied with DiagMMR assay, which has been developed for studying the protein function of these genes. In addition to the traditional agarose gel electrophoresis (AGE), the samples were also analyzed by a fragment analyzer, Labchip, that bases its function on capillary electrophoresis. This way the MMR detection efficiency of the methods could be compared. Samples were collected as skin biopsies from controls and LS patients with known MMR gene variants by Helsinki University Central Hospital (HUCH). InSiGHT database, that collects the different MMR-gene variants and their pathogenicity classification, was used to ensure that different kinds of variations, both pathogenic (class 5) and currently internationally unlisted variants, were analysed. The skin samples were cultured to acquire primary fibroblasts for nuclear protein extraction. The level of pathogenicity was revealed by MMR-protein activity when substrate DNA with a mismatch was added to the extract. Then, restriction enzymes were used for producing fragments of different lengths, depending on the repair action, and the MMR efficiency was visualized by both electrophoretic methods. Additionally, MAPP-MMR tool was used for studying the MSH2 mismatch variants in silico. By comparing the results from these two methods, we show that the more quantitative Labchip brings diagnostic value to DiagMMR suggesting 100% specificity (n=10) and 90,9% (n=11) sensitivity in reference to the variant information. For example, MSH6 c.3103C>T, which is listed as pathogenic in InSiGHT, was more consistent in giving a MMR deficient (dMMR) result with Labchip. Difference in the functional detection could be seen particularly with the MSH6 variants, but the differences were less notable when Labchip results were compared to the previous interpretations of the samples made based on the validated DiagMMR protocol. With the unlisted MSH6 variants, c.3139dupT was detected as dMMR by Labchip which was in unison with the previous interpretation. Another one, MSH6 c.551delA, was seen as MMR proficient (pMMR) in all the results by both the methods, and with the previous interpretation being unclear, which highlights the importance of further testing of this variant. There was also one unlisted variant (c.1805T>C) among MSH2 for which we got uniform dMMR results in two patients. The high MAPP-MMR score (25.150) for the MSH2 p.Leu602Pro amino acid change also supported the evidence gained of the pathogenic nature of this variant. As a conclusion, DiagMMR can be used reliably for MMR efficiency analysis, especially when performed together with a more quantitative analysis method.

Objectives. Functioning impairments as an outcome for severe mental disorders have been well established. There is also growing evidence for early functioning impairment being a vulnerability marker for psychotic disorders. Recent findings suggest that especially early social functioning impairment may predict later psychotic disorders. However previous studies are few and they are limited to pre-selected psychosis risk samples. In addition, little is known about early functioning in other severe mental disorders. The aim of this study is to determine the level of functioning in adolescents at entry to general psychiatric services and to examine whether early functioning impairment predicts later psychosis or psychiatric hospitalizations directly, and when controlling for the effect of psychosis risk symptoms. Methods. This study is a part of the larger Helsinki Prodromal Study conducted in the National Institute for Health and Welfare. The sample consists of 154 treatment-seeking adolescents aged 15–18 years, recruited at the first visit to psychiatric services in Helsinki. Semi-structured psychiatric interviews were conducted for all participants at baseline. Functioning was assessed with validated and reliable functioning measures for social (Global functioning: social) and role functioning (Global functioning: role) and an experimental functioning measure for psychological functioning (Global functioning: psychological). Information about hospitalizations was extracted from the Finnish hospital discharge register during a 2.8–8.9 year follow-up until the end of 2011. A Cox regression survival analysis was used to examine whether functioning domains predicted later hospitalization for psychosis or psychiatric hospitalizations overall. Positive and general psychosis risk symptoms as well as psychosis risk status were controlled for the analyses. Results and conclusions. The functioning of adolescents at entry to psychiatric services was mildly impaired and the impairment was greater for individuals in psychosis risk and those with a mental disorder diagnosis. 7 transitions to psychosis and 26 psychiatric hospital admissions emerged during the follow-up. The level of psychological functioning was the only significant predictor of increased risk of hospitalizations whereas the level of positive symptoms was the only significant predictor of increased risk of psychosis. Neither social nor role functioning predicted the outcomes. The results concerning social functioning are in contrast with previous finding, which may be attributable to the low incidence of psychosis in the sample or the treatment received by the participants. Overall the results emphasize the importance of interventions for early symptoms to prevent these severe outcomes.

Tutkielman tavoitteena on selvittää, mitä erilaisia funktioita koodinvaihdolla on tansanialaisissa blogeissa. Aineisto koostuu 55 blogikirjoituksesta, jotka on kerätty viidestä eri blogista aikaväliltä 2014–2016. Tutkielmassa tutkitaan koodinvaihtoa swahilin ja englannin välillä. Tutkimuksessa tarkastellaan koodinvaihdon funktioita tansanialaisissa blogeissa ja verrataan niitä Gumperzin esittämiin funktioihin, joita esiintyy konversationaalisessa koodinvaihdossa. Hypoteesina on, että koodinvaihdon funktiot blogeissa ja puheessa ovat hyvin saman tyyppisiä, sillä kielenkäyttö blogikirjoituksissa on spontaania ja puheenomaista. Tutkielman teoriaosuudessa käsitellään koodinvaihdon terminologiaa ja tutustutaan erilaisiin lähestymistapoihin koodinvaihdon tutkimuksessa. Teoriaosuudessa perehdytään myös Gumperzin koodinvaihdon funktioihin, joita sovelletaan aineistoa analysoidessa. Lisäksi tutustutaan kielenkäyttöön internetissä ja erityisesti blogeissa. Tutkimuksessa havaitaan, että viisi kuudesta Gumperzin konversationaalisen koodinvaihdon funktiosta esiintyy myös aineistona käytetyissä blogikirjoituksissa. Näiden lisäksi aineistosta löydetään myös neljä uutta funktiota. Lisäksi analyysissa käsitellään yleisiä havaintoja koodinvaihdosta blogeissa sekä esitellään funktioiden lisäksi muita syitä koodinvaihdon esiintymiselle tansanialaisissa blogeissa. Tutkimuksen tulokset osoittavat, että koodinvaihto puheessa ja blogeissa on monin tavoin samanlaista, mutta myös eroja näiden kahden genren väliltä löytyy. Ainoastaan yhtä Gumperzin esittämistä funktioista ei esiinny blogikirjoituksissa blogien ja puheen eriävien ominaisuuksien vuoksi. Aineistosta löydettävät uudet funktiot ovat: tärkeiden seikkojen korostaminen (emphasis), lyhempien sanojen ja rakenteiden suosiminen (speech economy), tiettyyn ryhmään kuulumisen ilmaiseminen (expressing fellowship) ja tunteiden ilmaiseminen (expressing emotions).

In this thesis we examine the properties of Wigner matrices. We will give proofs for two fundamental limit theorems of random Hermitian matrices. One of them is Wigner's semicircular law which states that the distribution of the eigenvalues approaches the Wigner's semicircular distribution when the size of the matrix increases. The other, Bai-Yin theorem, tells that the operator norm of such n × n matrix is almost surely (2+o(1))\sqrt{n}. In Chapter 1 we begin by introducing Wigner matrices and deduce the proper normalizing factor. We will also consider what does it mean for the distribution of the eigenvalues to converge by demanding that the empirical spectral distribution should converge almost surely in weak topology. In Chapter 2 we introduce the Stieltjes transform, a useful tool for finding the limit measure. We prove some basic properties and prove that the weak convergence of measures is equivalent to the convergence of their Stieltjes transforms. In Chapter 3 we prove the Wigner's semicircular law. The proof is based on using the Stieltjes transform, and contains several steps. First we find the pointwise expectation of the Stieltjes transform by deriving a polynomial equation for it. While deriving this equation we use several techniques and theorems from both probability theory and linear algebra, covered in the appropriate Appendix. After deducing that the expectation converges, we see quite straightforwardly that the pointwise limit is in fact almost sure. From this we deduce that the empirical spectral distribution of the Wigner matrix converges to the Wigner's semicircular distribution. We begin Chapter 4 by concluding the lower bound in Bai-Yin theorem directly by the semicircular law. For upper bound, we can split the matrix into three parts and use triangle inequality. The diagonal part is easily seen to grow slower than \sqrt{n}. The part with smaller elements is estimated using even moments and combinatorics to see that it has the wanted upper bound. Finally the part containing all large elements is almost surely sparse, and therefore its operator norm grows slower than \sqrt{n}.

The thesis gives an overview of indoor air pollution sources, health effects, and the detection of the fungal markers in indoor environment. In the experimental part of the thesis, solid phase extraction-liquid chromatography-tandem mass spectrometry method was developed for the quantitative analysis of 23 fungal secondary metabolites. The method was used to analyze the condensate water in indoor air from two sick houses in the Southern part of Finland. Furthermore, the practice was used to find other possible fungal marker compounds. The concerns of the indoor air pollution have been steadily increasing for the past couple decades since the prevalence of the irritative symptoms, such as skin, eye and respiratory track problems, that have been increasing without any clear reasoning. These symptoms are therefore being referred as building related symptoms since no other functionality have been discovered to be responsible for those. The general sources that could be linked to the cause of these symptoms are comfort parameters, material and burning related sources, and microbial metabolism products. From the microbes, the fungal species exhibit biggest concerns related to the indoor air quality since the growth of the fungus in the suitable conditions can lead to sizeable emissions of the particles and secondary metabolites which might be toxic, irritating or otherwise unhealthy. The traditional way of detecting the fungal contamination has required highly trained professional who investigates the interiors of the suspected contaminated house with the help of a moisture detector. When the contaminated spot is detected, the expert would conduct a visual inspection, and take a sample of the fungal growth for the microscopic analysis. Since then, methods focusing on the detection of the fungal particles and compounds have been developed. These include the collection of the bioaerosols and the detection of the volatile organic compounds originating from the fungal metabolism. However, there are challenges and limitations on these methods such as dependencies on time, low concentrations, and other pollution sources having similar chemical profiles than the fungal sources. Therefore, diverse ways to analyze the chemical profile of the sick houses is needed. Consequently, in the experimental part of this thesis, the method was developed to analyze fungal secondary metabolites from the condensate of the indoor air. The investigation consisted of the two fungal contaminated sick houses and from the one reference laboratory air sample. The results revealed twelve compounds to be elevated in the indoor air compared to the outdoor air and seven of these compounds were not detected in the reference sample. From these seven compounds, the studied amino acids and caffeine were the most promising fungal marker compounds since these compounds had consistently larger concentrations indoors than outdoors. Furthermore, the precursor ion scan revealed nine more potential marker compounds.

Pre-eklampsia on istukkaperäinen raskaudenajan häiriö, joka laskee äidin ja lapsen selviytymistodennäköisyyttä raskaudesta. Taudille tyypillisiä oireita ovat odottavan äidin korkea verenpaine ja tämän virtsan proteiinipitoisuus. Monitekijäisen taudin syyt tunnetaan vielä heikosti. Taudin perinnöllisyystutkimuksessa yksi kiinnostava kohde on ollut verisuonten kasvutekijäreseptoria koodaava geeni FLT1, jonka tuottaman proteiinin s-Flt1:n korkeat pitoisuudet assosioituvat tautiin. Geenin läheisyydestä tunnetaan yhden nukleotidin polymorfismi, jonka tietty muoto sitoo p53-proteiinia tehokkaasti. p53:n ilmentymisen tiedetään olevan koholla pre-eklampsiassa. Tutkimuksessa selvitettiin tapaus-verrokkitutkimuksena joukolta pre-eklamptikkoja ja näiden terveiltä verrokeilta genotyypit tutkittavan polymorfismin kohdalta perimän ja sairastavuuden mahdollisen yhteyden varalta. Tutkittavien seeruminäytteistä eristettyä perimäainesta monistettiin PCR-tekniikalla, ja DNA sekvensoitiin tarkan emäsjärjestyksen selvittämiseksi. Genotyyppien ja sairastavuuden sekä sairauden kannalta oleellisten kliinisten muuttujien mahdollisesta yhteydestä suoritettiin assosiaatioanalyysi Fisherin tarkkaa testiä käyttäen. Tuloksista ei löytynyt tilastollisesti merkitseviä assosiaatioita lukuun ottamatta s-Flt1-tasoja, joiden kohdalla kynnysarvon alittavien tasojen ja T-alleelin välillä nähtiin pieni viitteellinen yhteys sekä lasten että äitien kohdalla. Tulokset ovat ristiriidassa aiempien tutkimusten pohjalta luodun tutkimushypoteesin kanssa. Tutkimus aiheesta on vielä varhaista, mikä vaikeuttaa tulosten suhteuttamista.

Tässä tutkielmassa selvitetään, kuinka venäläiset kuluttajat suhtautuvat funktionaalisiin elintarvikkeisiin. Funktionaaliset elintarvikkeet ovat Venäjällä verrattain uusi ilmiö ja kuluttajien asenteet tuotteita kohtaan yhä vakiintumattomia. Tutkimuksen kannalta keskeisiä aiheita ovat kuluttajien asenteet teknologisesti muokattuja elintarvikkeita kohtaan, elintarvikkeen terveysväitteen merkitys tuotteen lisäarvon muodostumisen kannalta sekä venäläisten kuluttajien yleinen suhtautuminen terveysväittämällä varustettuja elintarvikkeita kohtaan. Suhtautumista selvitettiin venäläisille kuluttajille suunnattuna internet-kyselynä. Tutkielman aineisto koostuu 159 vastauksesta, jotka analysoitiin tilastollisia työkaluja apuna käyttäen. Tutkimuksen tulosten perusteella voidaan todeta, että Venäjällä ollaan hyvin kiinnostuneita funktionaalisista elintarvikkeista, mutta käsite ”funktionaalinen elintarvike” ei ole kuluttajille juuri itsessään tuttu. Saatujen tulosten mukaan suurin osa kuluttajien funktionaalisiin elintarvikkeisiin liittyvistä epäilystä kohdistuu hyvin periaatteellisiin kysymyksiin, esimerkiksi uusi valmistusteknologia saattaa saada valintatilanteessa merkittävää painoarvoa. Funktionaalisten elintarvikkeiden omaksumisessa korostuu kuluttajan luottamus niin tieteelliseen tutkimukseen, tuotteiden valmistajiin kuin tuotteiden markkinoijiin. Tulosten mukaan kuluttajat harvemmin luottavat markkinointikampanjoihin, joten erityisesti asiantuntijoiden rooli tiedonvälittäjänä on tärkeä. Terveydenhuollon ammattilaisilla nähdään olevan olennainen rooli kuluttajien tietoisuuden kasvattamisessa Venäjällä, sillä elintarviketeollisuuden yritykset koetaan liian puolueellisina todenmukaisen informaation levittämiseen. Lisäksi tulokset osoittavat, että terveysväitteen nähdään tuovan tuotteelle lisäarvoa. Motivaatio tuotteen käytölle syntyy kuitenkin ainoastaan siinä tapauksessa, mikäli kuluttaja ymmärtää terveysväitteen ja tuotteen terveysvaikutus tuntuu itselle merkitykselliseltä. Venäjällä suomalaisia elintarvikkeita pidetään saatujen tulosten perusteella korkealaatuisina, ja tuotteen suomalainen alkuperä lisää kuluttajien luottamusta funktionaalisen elintarvikkeen toimivuuteen. Hyvin Suomessa menestyneet funktionaaliset elintarvikkeet luovat uskoa alaan kohdistuville suurille odotuksille, ja saadut tutkimustulokset lisäävät toivoa menestystarinoihin myös Venäjällä.

Kultakomplekseja on katalyysikemiassa perinteisesti käytetty aktivoimaan alkyynien, alleenien ja alkeenien π-sidoksia nukleofiilistä additiota varten. Kultakomplekseja, joissa on halogeeniligandeja, kutsutaan yleensä esikatalyyteiksi. Näistä voidaan muodostaa aktiivinen katalyytti luomalla kultaioniin vapaa koordinointipaikka vaihtamalla vahvasti koordinoiva halogeeni-anioni ei-koordinoivaan anioniin. Ligandeissa mahdollisesti esiintyvän funktionaalisen osan tarkoituksena on korvata vastaionien käyttö koordinoitumalla heikosti kultaan. Hypoteesin mukaan funktionaalinen ryhmä kykenisi spontaanisti irtautumaan kultaytimestä ja näin ollen luomaan vapaan koord inointipaikan katalysoitavalle substraatille. Kirjallisuuskatsauksessa tarkastellaan alkuun organokultakompleksien toimintaa homogeenisessä katalyysissä sekä vastaionien merkitystä katalyyttiseen aktiivisuuteen. Tämän jälkeen tarkastellaan syvemmin funktionaalisten NHC-kultakompleksien ominaisuuksia, kuten niiden funktionaalisen osan kykyä koordinoitua kultaytimeen katalyyttien aktivoimiseksi. Samalla tarkastellaan myös missä reaktioissa ja minkälaisella menestyksellä funktionalisoituja NHC-kultakomplekseja on käytetty. Muutamassa tapauksessa kultakomplekseja onnistuttiin aktivoimaan NHC-ligandien funktionaalisuuden ansiosta. Kokeellisessa osuudessa syntetisoitiin ksyleeni- ja pyridiinipohjaisilla linkkereillä sekä suojatulla amiinifunktionaalisuudella varustettuja NHC-ligandeja. Vastaavien kulta(I)- ja kulta(III)- kompleksien katalyyttistä aktiivisuutta tutkittiin alkynyylifuraanin toisiintumisreaktiossa, terminaalisen alkynyylihapon ja propargyyliamidin syklisointireaktioissa sekä imiinisynteesissä. Valmistetut katalyytit osoittivat vaihtelevaa aktiivisuutta eri reaktioissa.