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(2022)Kv7.1 is a potassium ion channel comprised of the KCNQ1 protein, which can coassemble with distinct β-subunits modulating the channel functions in different tissues. In 2017, Raivio’s group (from the University of Helsinki) found two missense mutations in the KCNQ1 gene, p.(Arg116Leu) and p.(Pro369Leu), responsible for causing pituitary hormone deficiency and maternally inherited gingival fibromatosis. The facial features and bone structure pointed to a cranial neural crest (CNC)-derived phenotype caused by an alteration in the potassium channel balance, given that these cells form the bone and cartilage of the cranial zone. To understand the implication of the CNC in the KCNQ1 syndrome, I attempted to replicate the CNC differentiation protocol of Suga and Furue (2019) with the aim of obtaining cranial neural crest cells (CNCCs). This would enable future generation of a KCNQ1-related disease model. The differentiation process was carried out thrice, and two BMP4 concentrations (10 and 100 ng/ml) were assayed. The differentiated cells exhibited a CNC-like morphology as well as upregulation of the marker genes (TFAP2A, SOX10, DLX1, MSX1, and DLX2) associated to this cell lineage. However, the gene expression was low according to the qRT-PCR Ct values, which were in most cases higher than 30. Additionally, no differences were found between the two BMP4 treatments. Furthermore, the cells did not express KCNQ1, and thus the impact of the two KCNQ1 mutations was not investigated under this protocol. In conclusion, the protocol had a low efficiency in the generation of CNCCs that was not improved by increasing the BMP4 concentration. Further optimization of the protocol, such as the BMP4 concentration or the cell density of the culture, will be needed to improve its efficiency and obtain an adequate disease model.
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(2022)Digitaalisia teknologioita hyödynnetään lisääntyvästi kansalaisosallistumisen vahvistamisessa sekä vahvan ja osallistuvan demokratian toteuttamisessa. Tämän niin sanotun e-osallistumisen tai digitaalisen osallistumisen päämääränä on aktivoida kansalaisia ja madaltaa osallistumiskynnystä sekä rohkaista heitä keskusteluun julkisen hallinnon kanssa. Teknologiset ratkaisut lupaavat tiiviimmän ja reaaliaikaisenkin keskusteluyhteyden hallinnon ja kansalaisten välillä. E-osallistuminen koetaan myös ratkaisuna maaseuduille tyypillisten niukkenevien resurssien ja pitkien etäisyyksien haasteisiin. Kiinnostus e-osallistumiseen on tutkimusten mukaan kuitenkin vähäistä. Osallistumista tukevia teknologisia ratkaisuja olisi tarjolla, mutta niiden hyödyntämiseen ei olla innostuttu kunnissa. Tässä tutkimuksessa kartoitetaan, kuinka kuntaorganisaation viranhaltijat kokevat kuntalaisten osallistumisen sekä millaisia haasteita ja mahdollisuuksia he liittävät e-osallistumisen teknologisiin ratkaisuihin. Tutkimusaineisto koostuu 11 teemahaastattelusta, jotka kerättiin Mikkelin kaupungin eri palvelualueita edustavilta viranhaltijoilta. Haastattelupuhe on sosiaalisesti rakentunutta ja aineistoa tarkasteltiin teoriasidonnaisesti kehysanalyyttisella otteella. Analyysissa selvitettiin keskeiset osallistumiseen liittyvät kehystämistavat viranhaltijoiden näkökulmasta. Haastateltavat puhuivat osallistumisesta velvollisuuksien kehyksessä ja käytäntöjen kehyksessä. Vaikka osallistumista pidettiin tärkeänä, ristivetoa esiintyi velvollisuuksien (kuntalaki ja kuntalaisten osallistumismahdollisuudet) ja käytäntöjen (nykyiset toimintakulttuurit ja niukat resurssit) välillä. Edustuksellisen päätöksenteon ja kuntalaisten osallistumisen suhde osoittautui kitkaiseksi. Myös kuntalaisten näkökulmaa ja mielekkäitä osallistumisen tapoja olisi tulosten mukaan kehitettävä. Lisäksi kuntaorganisaatioiden nykyiset toimintakulttuurit kaipaavat osallistumisen huomioon ottavaa uudistamista ja riittävän resursoinnin varmistamisen.
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(2020)As biodiversity is being lost worldwide at an accelerating rate due to anthropogenic activities, the frequency and severity of many infectious diseases has been observed to increase. Together these patterns have brought forth an urgent need to understand the possible linkages between biodiversity and disease risk. Two contradicting hypotheses have been proposed to explain the diversity-disease relationship. The dilution effect hypothesis suggests that increasing host community species diversity ‘dilutes’ disease risk, whereas the amplification effect hypothesis predicts disease risk to increase with increasing diversity. Even though most of the studies support the dilution effect, there remains an intensive debate regarding the generality of this effect. As most of the research efforts to understand the relationship between diversity and disease have focused on animals and crop plants or have been carried out experimentally, one of the research gaps is how relevant the dilution effect is in wild plant communities. In nature, plants and their diseases are affected simultaneously by multiple abiotic and biotic environmental factors that might confound or supersede the effects of diversity. It is also poorly understood, whether we might expect dilution effects to occur not only on diversity gradients driven by anthropogenic diversity loss, but also on natural diversity gradients. To study the possible association between host community species diversity and disease risk in the wild and to test whether this association could be detected after accounting for the effects of abiotic factors, I surveyed grassland vascular plant communities for their species diversity and foliar disease symptoms along a natural diversity gradient driven by elevation. I also recorded data on the mean soil surface temperature in the surveyed plant communities and used structural equation modelling to differentiate and compare the effects of biotic and abiotic variables on disease risk. The data were collected on Mount Calanda in the Swiss Alps during summer 2019. In this thesis I show that host community species diversity and disease risk are negatively associated with each other along a natural diversity gradient driven by elevation. Furthermore, this negative effect can be detected even after accounting for the effects of elevation and mean soil surface temperature on disease. Together the results support the occurrence and the ecological relevance of the dilution effect in wild plant communities along natural diversity gradients and suggest that diversity might protect wild plant communities from increased disease risk. Future studies should aim to identify the exact mechanisms of the association to help us better understand when and where we might expect dilution effects to occur in the wild. This knowledge can be used to predict how epidemics, that affect the well-being of ecosystems, humans and wildlife, are born in the changing world.
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(2024)Dispersal is a vital life-history trait in terms of species survival, reproduction, and persistence, among others. We know that temporal and spatial differences in the environment alter the risks and rewards involved in dispersal, thus influencing dispersal behaviour and ability. Less known, however, is how exactly seasonal shifts and land use together affect dispersal ability. Butterflies are a well-known and ecologically important study group whose traits are majorly influenced by environmental conditions. Here, I study how the dispersal ability of the green veined white butterfly (Pieris napi) is influenced by when they fly, as well as land use in their local habitat. I collected butterflies of two subsequent generations during the late spring and summer of 2023 in the Åland archipelago (Finland). I then measured the flight metabolic rates and forewing lengths of these butterflies to estimate their dispersal ability. The results described here indicate that P. napi females flying in late spring and early summer are more capable of dispersing than those flying later in the summer. Land use had seemingly no effect on P. napi dispersal ability. Identifying the factors that influence dispersal is key in understanding and securing the future for species in a changing world.
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(2024)The S209F variant of the Abelson Interactor family member 3 (ABI3) gene has emerged as a risk factor for late-onset Alzheimer’s Disease (LOAD). The ABI3 protein is functionally related to the WAVE Regulatory Complex (WRC) participating in the control of cytoskeletal processes favoring either filopodia for chemotaxis or pseudopodia for phagocytosis. The S209F coding variant is thought to impair phosphorylation of the ABI3 protein leading to dysfunctional association with WRC. In the brain, the ABI3 gene is mainly expressed by microglia, macrophages representing the resident immune cells of the brain. Despite some research about the variant based on rodent models and reporting sometimes contrasting results, the role of the ABI3 S209F variant in AD remains poorly understood. Here, human-induced pluripotent stem cells (h-iPSCs) reprogrammed from fibroblasts of controls and variant carriers are sequenced to ensure retention of the original phenotype upon reprogramming. H-iPSCs are differentiated into microglia (iPSC-derived microglia, iMGL) following an established protocol. Morphological changes and microglia-specific gene expression partially show that iMGL between days 31 and 38 of differentiation in vitro can be considered mature. To assess the functional properties of microglia, cytokines/chemokines production, cathepsin gene expression, lysosomal activity, and Apolipoprotein E (ApoE) protein levels are measured. It is found that S209F microglia downregulate CCL5/RANTES and upregulate cathepsins B and L (CTSB and CTSL) upon LPS+IFNg stimulation which may lead to motility, migratory and endo-lysosomal dysfunctions. Lysosomal activity is found to positively correlate with CD163, but not with either CTSB or CTSL expression. ApoE protein levels show an upregulation trend in S209F microglia which may indicate modifications in lipid metabolism. Metabolic assessment based on mitochondrial respiration and glycolysis does not show any difference between S209F and control microglia, but ABI3 knock-out (KO) shows glycolysis dysfunctions. Overall, this study offers some hints into the mechanisms that make the ABI3 S209F variant a risk factor for AD pointing at the need to investigate microglia motility and migration focusing on pathologically relevant protein aggregates and their clearance and with particular attention to phagocytosis and endo-lysosomal pathway.
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(2024)Glioblastoma is the most aggressive and lethal tumor in the central nervous system. One of the main challenges of current treatment reside in its high intratumoral heterogeneity. Within one tumor, there could be several glioblastoma subtypes which harbor distinct molecular signatures and associated with different clinical properties. For instance, mesenchymal-like glioblastoma patients have the shortest survival time and this subtype is highly resistant to radiation. The current subtype classification heavily relies on transcriptomic analysis, which is not available for every patient due to its high cost. Here, I utilized a polycation reagent to treat neural-progenitor-like and mesenchymal-like glioblastoma cells with increasing concentrations and evaluate their cell viability using luminescence-based CellTiter-GLO assay. I found that neural-progenitor-like glioblastomas cell growth was significantly inhibited at intermediate concentrations of the polymer, while mesenchymal-like glioblastomas were less affected. With polymer concentration increasing, the inhibition effect displays a dose-dependent trend. My results demonstrated that neural-progenitor-like glioblastomas are more sensitive to the polymer than mesenchymal-like ones. The different sensitivity towards the polymer between these two subtypes suggests polymer could be used as a potential reagent to distinguish glioblastoma subtypes.
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(2023)Macrozooplankton is an understudied size class of plankton in the Arctic, but even though species composition is similar around the Svalbard archipelago, the relative composition can act as a proxy for climate change. This study investigated if the fjords around Svalbard are similar in species composition and if any differences could be explained. 11 fjords were sampled with a focus on common and indicator species in terms of abundance, relative composition and length distribution. I found low presence of T.libellula and high abundances of T.abyssorum and T.inermis. T.abyssorum was the only species whos abundance was significantly correlated to water mass (Atlantic water), but T.libellula, T.abyssorum, T.inermis, T.longicaudata and A.digitale all showed significant differences in length distribution. This study provides further understanding of species composition in the different high Arctic fjords around Svalbard.
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(2023)Morphological features are considered as markers of microglial functionality, and they show regional heterogeneity in the brain. Recently the sleep-wake cycle was shown to affect microglial morphology in mice and correlate with cortical sleep slow wave activity (SWA). Microglial sizes and ramification increased during the dark period and decreased during the light period in cerebral areas associated with SWA, suggesting that neuronal activation could be affecting microglial morphology through SWA. I studied microglia in the hindbrain areas with and without functional connection to SWA to further investigate the association between SWA and alterations in morphology, and to investigate if there are differences in microglial morphology and their diurnal alterations in brain regions other than those commonly investigated. I examined three hindbrain areas (cerebellar cortex (CC), deep cerebellar nucleus (DCN) and medial vestibular nucleus (MVN)) and somatosensory cortex (SC) of mice (n=15) at two timepoints: 6 hours after the light onset (high SWA) and offset (low SWA). My aims were to answer if there are morphological differences in microglia between 1) the four brain areas at both timepoints and 2) between the two timepoints in each brain area. My hypotheses were that CC and DCN which have functional connections to cortical SWA, would show similar diurnal morphology alterations as demonstrated in the cerebral areas, and MVN that has no known cortical SWA connection, would lack significant alterations. As microglia are heterogenous throughout brain, I expected microglia to differ between different brain areas, especially the hindbrain and the SC. I found that microglial morphologies significantly differed between the hindbrain and the cortex, while the hindbrain areas were more similar in morphology. Moreover, the brain areas demonstrated diurnal morphology alterations of microglia with varying extent: CC and DCN microglial morphology did not correlate with SWA as clearly as SC did, and interestingly, morphological features of MVN microglia showed a pattern opposite to other areas, microglia being larger during the light period than the dark period. These results highlight the importance of the diurnal time to microglial morphology and the heterogeneity of microglia between different brain regions.
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(2020)The diversity of different neuronal types lays the foundation for different functions in the brain. The development of different subpopulations and special features of neurons in the central nervous system are still partly unknown. Finding answers to these developmental issues could help in the process of characterisation of cell types and mapping of neuronal networks between the brainstem nuclei in the brain. Previous studies have shown that a ventrolateral neuroepithelial domain in the anterior hindbrain, rV2, produces excitatory (glutamatergic) and inhibitory (GABAergic) neurons, which are related to monoaminergic nuclei in the brainstem (Lahti et al., 2016). In this master’s thesis project, the development of a subpopulation of neurons expressing Gsc2 transcription factor in the interpeduncular nucleus was studied. This project was based on single-cell RNA sequencing results conducted in E13.5 mice. Predicted by single-cell RNA sequencing results, Gsc2 expressing cells are GABAergic interneurons and originate from the rV2 domain of the rhombomere 1 region in the hindbrain. Co-expression pattern with another transcription factor Sall3 with Gsc2 during development was also addressed in the study. Furthermore, the role of Notch signalling in the binary cell fate decision between GABAergic and the glutamatergic fate of rV2 neurons was investigated. Validation of single-cell RNA sequencing results was performed using in situ hybridisation and immunohistochemistry methods with mice embryos at the age of E12.5 and E15.5. This study verified previously shown origin of Gsc2 expressing cells to the rhombomere 1 region and in addition, showed that Gsc2 expressing cells are GABAergic. Co-expression pattern of Gsc2 with Sall3 neither in the rV2 domain nor in the interpeduncular nucleus was seen in our results. In the rV2 domain, the depletion of Notch signalling decreased the expression of differentiating GABAergic neurons. This indicates that Notch has a role in GABAergic neurotransmitter identity during the development of brainstem neurons in mice. Based on our results, Gsc2 could be used as a lineage marker for GABAergic interneurons originating from the rhombomere 1 region and as a marker for a subpopulation of the interpeduncular nucleus. Furthermore, results from the role of Notch signalling could help in discovering the mechanisms related to the determination of neurotransmitter identity in rV2 neurons. Further investigations, in different developmental time points and with additional markers, are needed to verify these results.
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(2019)Background- Colorectal cancer (CRC) is the third most common epithelial carcinoma. There is an increased risk of colorectal cancer in people with longstanding inflammation in the large intestine, including individuals with ulcerative colitis (UC). Epigenetic changes in CRC such as aberrant DNA methylation alterations are common changes in human cancer. The aim of this study is to identify the DNA methylation alterations of selected inflammation related genes in UC-CRC vs. Lynch syndrome (LS). Method- DNA was extracted from archival tissue specimens from normal and tumor samples from UC-CRC (n= 31), and LS-CRC (n=29). Methylation-specific multiple ligation-dependent probe amplification (MS-MLPA) assays were used to detect CIMP status and CpG promoter methylation status of seven inflammation related genes. Microsatellite instability analysis was carried out using two mononucleotide repeat markers BAT25 and BAT26. Results- Increased hypermethylation frequencies in carcinoma vs. normal colonic mucosa were detected for all the inflammatory marker genes in specimens of UC-CRC patients. Statistically significant differences for methylation frequencies were observed in the NTSR1 gene (p value =0.008) and SOCS2 gene (p value =0.04) in specimens of UC-CRC patients. NTSR1 gene showed significantly increased methylation of normal colonic mucosae from UC-CRC vs. LS patients (p value=0.01). Conclusion- UC-CRC and LS tumor specimens revealed varying frequencies of hypermethylation in all the inflammatory genes. Methylation of the NTSR1 in the normal colonic mucosa suggests a possible field defect in UC-CRC, and could thus be used as an early biomarker to detect increased UC-CRC risk in non-neoplastic epithelium.
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(2024)Climate change and overexploitation are an increasing threat in marine ecosystems as human activities have risen over the past decades. One of the main tools for protecting biodiversity and ecosystem functions is the designation and implementation of Marine Protected Areas (MPAs). Despite their potential for conservation, recent assessments question the effectiveness of MPAs to conserve ecologically resilient ecosystems. In this thesis, I aim to assess the effectiveness of MPAs in Finland using long-term phytoplankton data, which have been collected for almost 60 years (1960-2020) in the Finnish territorial waters from over 700 sites and contain 991 different phytoplankton taxa. Phytoplankton is the basis of marine food networks and thus shifts in phytoplankton communities can affect higher trophic levels. I compared the total biomass, species richness and species diversity of the phytoplankton communities between protected and unprotected sites. To control for the confounding effects of environmental variables that affect phytoplankton biomass and distribution, and to isolate the effect of protection, I employed nonparametric distance matching; essentially, this process selects comparable pairs of samples from protected and unprotected sites. I found that even though there was a general increase in species diversity with time, protection had no effect on phytoplankton biomass, species richness or species diversity. The size and age of the MPAs did not affect the protection outcomes. However, strictly protected MPAs harbored lower biomass compared to unprotected sites, which could indicate that MPAs can help reduce the effects of eutrophication. These results suggest that MPAs in Finland still have room to be improved and there is a need to accurately assess their effectiveness in order to successfully safeguard biodiversity. Considering the phytoplankton component during the designation and planning of the MPA network in Finland could be valuable for marine conservation.
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(2011)Pro gradu tutkielmani aiheena oli tutkia koirien (Canis familiaris) saalistushalukkuutta ja persoonallisuutta sekä näiden mahdollista yhteyttä dopamiinireseptori D4 –geeniin (myöh. DRD4-geeni). DRD4-geeni ilmenee lähinnä hermosoluissa ja se vaikuttaa erityisesti aivojen limbisen järjestelmän toimintaan. Limbinen järjestelmä säätelee tunnetiloja ja tunteisiin liittyviä autonomisia toimintoja sekä motivaatiota eli samoja asioita, joihin dopamiinin tiedetään vaikuttavan. Dopamiini vaikuttaa elimistöön sitoutumalla esimerkiksi aivojen limbisen alueen dopamiinireseptoreihin. Dopamiinin on havaittu liittyvän assosiatiiviseen oppimiseen ja DRD4-geenin puolestaan mm. aktiivisuuteen ja impulsiivisuuteen sekä uteliaisuuteen mistä syystä se valittiinkin kandidaattigeeniksemme tutkimukseen. Koira puolestaan sopii erityisen hyvin mallilajiksi kandidaattigeenitutkimukseen, koska koirarodut ovat geneettisiä isolaatteja ja eroavat toisistaan niin rakenteensa kuin käyttäytymisensäkin puolesta. Kytkentäepätasapaino eli alleeliassosiaatio on koirissa huomattavasti voimakkaampaa kuin ihmisissä ja siten koiralta on merkittävästi helpompaa löytää mm. tautigeenejä. Koirilla kytkentäepätasapaino yltää noin 2 Mb laajuudelle, kun vastaava luku ihmisellä on vain noin 0,28 Mb. Tutkimuksen 405 suursnautserin ja saksanpaimenkoiran saalistushalukkuus testattiin koiran luonteenkuvauksesta eli MH-testistä tutulla viehetestillä. Viehetestissä koiran edestä yllättäen "pakenee" saalista muistuttava mutkittelevasti kulkeva kankainen viehe. Koirien suoritukset viehetestissä videoitiin ja suoritukset luokiteltiin neljään luokkaan äärimmäisen saalistushalukkaasta saalistushaluttomaan. Muita persoonallisuuspiirteitä tutkittiin koirien omistajien täyttämien luonnelomakkeiden avulla. Kaikista viehetestillä testatuista koirista otettiin verinäytteet, joista eristettiin DNA. Saalistustestissä äärimmäisen saalistushalukkaat ja täysin saalistushaluttomat saksanpaimenkoirat (N=44) valittiin DRD4-geenin alleeliassosiaatiotutkimukseen. Tutkittavaa geenialuetta monistettiin käyttämällä alleelispesifejä alukkeita PCR-reaktiossa. Tutkimuksen koirista löydettiin DRD4-geenin alleeleja 2 ja 3a. Luonnelomakeaineiston taustalta paljastui faktorianalyysin avulla neljä faktoria: sosiaalisuus ihmisiä kohtaan, leikkisyys ja aktiivisuus, aggressiivisuus koiria kohtaan sekä rohkeus. Näistä ensimmäinen assosioitui merkittävästi alleelin 3a kanssa. Koirat jotka kantoivat tätä alleelia olivat vähemmän aggressiivisia ihmisiä kohtaan kuin koirat, joilla 3a-alleelia ei ollut. Saalistushalukkuuteen kumpikaan alleeleista eikä mikään genotyypeistä assosioitunut. Saalistushalukkuutta kokonaisaineistossa selitti parhaiten leikkisyys ja aktiivisuus. Rodut erosivat toisistaan ihmisiin ja koiriin suuntautuvalta aggressiivisuudeltaan siten, että saksanpaimenkoirat olivat suursnautsereita aggressiivisempia.
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(2005)1-Bentsyylipiperatsiini (BZP) on viime vuosina laittomille markkinoille ilmaantunut ns. design-huume, jonka on todettu muistuttavan vaikutuksiltaan amfetamiinia. Sen myynti tapahtuu tyypillisesti tanssiklubeilla ja Internetin välityksellä. Esimerkiksi Yhdysvalloissa BZP on luokiteltu laittomaksi, mutta suurimmassa osassa maailmaa aine on huumausainelainsäädännön ulkopuolella. Suomesta ainetta takavarikoitiin neljästi vuosina 2002-04. Tässä tutkimuksessa selvitetään ehdollistetun paikkahakuisuuskokeen avulla, onko BZP:lla palkitsevia ominaisuuksia. Kokeessa koe-eläimet opetetaan yhdistämään tutkittavan aineen vaikutukset tiettyyn ympäristöön, ja mikäli eläin alkaa tämän jälkeen suosia ympäristöä, eli aine aiheuttaa paikkahakuisuutta, tulkitaan aineella olevan palkitsevia ominaisuuksia. Palkitsevien ominaisuuksien katsotaan heijastavan aineen riippuvuuspotentiaalia, sillä useimpien ihmisten väärinkäyttämien aineiden on todettu aiheuttavan ehdollistettua paikkahakuisuutta. BZP:n todettiin aiheuttavan rotille ehdollistettua paikkahakuisuutta annosriippuvaisesti, minkä jälkeen selvitettiin reseptorisalpaajien avulla dopamiinireseptorien merkitystä paikkahakuisuuden synnylle. Dopamiini-1-, eli D1-reseptorisalpaus esti paikkahakuisuuden, kun taas D2-salpauksella ei ollut siihen vaikutusta. Kokeiden perusteella BZP:lla on palkitsevia vaikutuksia, jotka välittyvät D1-, mutteivät D2-reseptorien kautta. Tulokset viittaavat BZP:lla olevan väärinkäyttö- ja riippuvuuspotentiaalia, minkä vuoksi olisi perusteltua, että se kuuluisi lainsäädännöllisesti samaan kategoriaan mm. amfetamiinin kanssa.
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Drone-kuvien käyttö haurujen (Fucus spp.) kartoitukseen ja monitorointiin itäisellä Suomenlahdella (2023)Tämä opinnäytetyö käsittelee uuden ympäristöseurantamenetelmän kehitystyötä. Hauruja (Fucus spp.) on seurattu Suomen rannikolla jo pitkään, mutta nykyisiin kartoitus- ja seurantamenetelmiin liittyy haasteita ja epätarkkuutta. Hauruvaltaiset elinympäristöt ovat uudessa uhanalaisuusarvioinnissa määritelty erittäin uhanalaisiksi. Tarvetta uusille seurantamenetelmille siis on. Drone-teknologia on kehittynyt ja halventunut viime vuosina, mikä on avannut uusia mahdollisuuksia hyödyntää kaukokartoitusta. Edulliset valokuvaamiseen tarkoitetut dronet mahdollistavat sekä ajallisesti että paikallisesti erittäin tarkkojen ilmakuvien tuottamisen. Näitä on maailmalla käytetty menestyksellisesti myös rannikon kartoituksessa ja seurannassa. Suomessa drone-kuvia ei kirjallisuuden perusteella ole käytetty makrolevien kartoitukseen tai seurantaan. Tämän tutkimuksen tarkoitus on selvittää, sopivatko tällaiset drone-kuvat hauruyhteisöjen kartoitukseen ja monitorointiin itäisellä Suomenlahdella sekä tuottaa tietoa menetelmän jatkokehitystä ajatellen. Kuvasin 22 merenranta-alueen reunaympäristön alkukesällä 2020. Kuvatuista alueista valitsin kahdeksan jatkotutkimuksiin, jotka kuvattiin uudelleen syksyllä 2020. Tällöin niiden alueelta kerättiin kuvatulkinnan tueksi 263 tukipistettä, joilta mitattiin veden syvyys ja arvioitiin haurujen esiintyvyys Drop-kuvausmenetelmällä. Rajasin ilmakuvilta näkyvät haurualueet ja vertasin niitä tukipisteaineistoon. Havaitsin, että 80 % havainnoista tieto luokittui oikein. Cohenin kappa -testin perusteella kohteiden välillä sekä eri syvyyksillä olevilla kasvustoilla oli suuria eroja havaintojen luotettavuudessa. Syvemmällä kuin 2,5 metrissä olevien haurukasvustojen erottaminen onnistui huomattavasti huonommin. Myös matalimman kasvuston (0 – 0,5 m) erottaminen oli hankalaa. Tulokset ovat lupaavat, mutta niiden perusteella menetelmää ei kuitenkaan vielä voi todeta toimivaksi, vaan lisätutkimuksia tarvitaan. Tämän tutkimuksen perusteella kehitystyötä kannattaa jatkaa käyttämällä tarkkaa paikkatietoa ilmakuvien ja tukipisteaineiston tuottamiseen sekä uusia kuvantamismenetelmiä, jotka kykenevät tuottamaan tavanomaista RGB-kameraa paremman erottelukyvyn. Drone- ja meritöiden turvallisuuden parantamiseksi tehtiin riskienhallintaprosessi, jossa tunnistin, analysoin sekä raportoin töihin liittyvät merkittävimmät riskit.
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(2020)Multiple myeloma (MM) is a heterogeneous plasma cell cancer that results from the excessive proliferation of mutated B cells in the bone marrow and the accumulation of ineffective antibodies, monoclonal proteins, in the blood. Despite recent advances in research and novel therapeutics, MM remains incurable, mainly due to the mechanisms underlying disease progression and drug resistance. Therefore, novel biomarkers and therapeutics for the treatment of relapsed and refractory MM are urgently needed. MicroRNAs (miRNAs), short non-coding RNA molecules that play a key role in post-transcriptional gene regulation, have been found to be associated with different hallmarks of MM. Previous studies have indicated that abnormally functioning miRNA-mediated gene regulation followed by oncogene activation and tumor suppressor gene silencing results in drastic alterations in cell proliferation, apoptosis, growth, and metabolism. These changes in cellular functions have been indicated to be associated with the pathogenesis, progression, and formation of drug resistance in MM. Therefore, the role and potential of miRNAs to act as biomarkers to predict MM progression and drug sensitivity should be further investigated to ultimately improve the survival rates of patients. The aim of this master’s thesis was to investigate the relationships between drug sensitivity, disease progression and miRNA regulation in MM patients. Bioinformatically predicted miRNAs identified to be associated with sensitivity to panobinostat, a novel histone deacetylase inhibitor, and MM progression were validated in MM patient samples by using real-time quantitative reverse transcription PCR (RT-qPCR). In addition, the specific gene targets of miRNAs involved in the regulation of drug responses and MM progression were predicted by identifying statistically significant, negatively correlated interactions between the miRNA and RNA sequencing data of 45 MM patients in pairwise comparative correlation analysis. Finally, the predicted miRNA targets genes were validated in MM patient samples using RT-qPCR. Based on the bioinformatic analyses and RT-qPCR validation, mir-424 expression was significantly increased in relapsed MM patients as compared to respective patient samples taken at diagnosis, suggesting a potential role of mir-424 in MM progression. Similarly, mir-4433b expression was significantly elevated in panobinostat-resistant patients compared to sensitive patients, suggesting a potential effect of mir-4433b on the regulation of panobinostat drug response in MM patients. In addition, the RT-qPCR validation demonstrated that the disease progression and drug sensitivity associated mir-92b, mir-363 and mir-221, would potentially regulate the expression of FGF2, MFF, and TMEM248, respectively, providing novel insights into the functional roles of miRNAs in MM pathways.
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(2020)Upregulation of specific helpful proteins represents a possible method for preventing or treating human diseases. Endogenous upregulation (knockup) is the increase of a gene's expression only in cells in which it is already expressed, thus avoiding physiologically abnormal spatiotemporal patterning. A gene's three prime untranslated region (3′UTR) affects protein expression through stability regulation of RNA already transcribed, which suggests 3′UTR modification as a viable route for endogenous upregulation. Mammalian model organisms can be generated in order to test the effects of different 3′UTR modifications, but at great cost of time, effort, and money. If able to predict in advance with an in vitro assay whether an in vivo modification would cause a desirable or undesirable change, these costs could be substantially reduced. In this thesis project, an in vitro assay was used to compare the protein expression influence of twenty neurodegeneration-relevant mouse genes' 3′UTRs to that of a flip-excision cassette (flex-cassette) previously used for in vivo conditional knockup. The assay used was the Promega Dual-Luciferase Reporter Assay, in which plasmids expressing Renilla and Firefly luciferase as reporter and internal control are co-transfected into in vitro cells, then each luciferase's expression measured with its respective substrate and a luminometer. Transfections were carried out in three-well replicates and on multiple days. The aims of the project were the evaluation of the assay's ability to predict in vivo results, the suggestion of 3′UTRs which could be upregulated in vivo by the conditional knockup flex-cassette, and the identification of any trends in 3′UTR-based protein expression influence according to gene function. A number of gene 3′UTRs were identified which were either candidates for flex-cassette upregulation or candidates for use in the flex-cassette to upregulate other genes. However, the flex-cassette's in vitro results were only partially consistent with its previous in vivo results. Specifically, the lox sites in the flex-cassette was observed to lower expression level to a degree not observed in vivo. Additionally, in the course of the project a number of possible workflow improvements were identified, for which suggestions have been made in the text. As such, this in vitro approach requires further study in order to determine suitability for prediction of in vivo 3′UTR behaviour.
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(2014)The purpose of this thesis was to examine the diversity and species composition of dung beetle (Scarabaeinae) communities in degraded rainforest landscapes in southeastern Madagascar. Several studies elsewhere in the world have revealed that forest-dwelling dung beetle communities and especially large species suffer from forest degradation and fragmentation by decreased species diversity. The most important factors affecting community structure of forest-dwelling dung beetles are habitat area, connectivity and vegetation quality i.e. microclimate. The hypothesis of this study was that the situation is the same in Madagascar. As dung beetles provide several important ecosystem services, like nutrient cycling and bioturbation, loss of dung beetle diversity imposes a secondary threat to the extraordinary nature of Madagascar by decreasing the regeneration ability of vegetation. Material for the study was collected in forest fragments of different size and quality between two areas of primary tropical rainforest – Ranomafana National Park and Vatovavy mountain – in November and December 2011 and January 2012. The sampling was conducted by transects of 30–60 fish- or carrion-baited pitfall traps which were set up in 55 localities in the study area. In each locality, several variables were measured to describe the vegetation and microclimatic conditions. The variables included temperature, humidity, estimate of vegetation quality by 6 observation-based classes, vegetation density, hights of three clearly visible vegetation layers, altitude and slope steepness. In addition, connectivities were measured for the localities using GIS and a satellite image –based vegetation classification. In order to demonstrate the differences between certain localities the study sites were divided into seven zones in terms of their distance from the Ranomafana National Park, average connectivity of the transects and elevation. Altogether 4,199 individuals belonging to 24 species were collected. Six of the species are currently under identification process in the Paris Museum of Natural History. According to the preliminary results they include two species new to science. Largest numbers of species were collected from good quality fragments between Ranomafana and Vatovavy. Also, a clear transition zone in species composition was detected a few kilometers west from Vatovavy, where altitude changes sharply. The study reveals that the species assemblages in the forest fragments and degraded forest areas are surprisingly species rich. This may, however, be partly because of extinction debt, and many of the still surviving species may soon die out due to restricted dispersal possibilities. Connectivity and vegetation quality were shown to have an effect on Canthonini species richness, with less species in less connected areas and lower vegetation quality. Vegetation quality was also shown to have an impact on the proportions of species with different body length: more small and medium-sized (< 8 mm) species were found in fragments where vegetation was more degraded. In addition to revealing how rainforest fragmentation and degradation affect local communities, the study gives interesting information about the distribution of certain species of Epilissus (Scarabaeinae: Canthonini). It has been known before that four species of Epilissus show elevational differentiation in their occurrence in Ranomafana. In this study, two more species of the same genus, E. prasinus and E. emmae obscurpennis, were shown to continue this pattern in lower elevations near Vatovavy mountain, about 50 kilometers east of Ranomafana.
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(2020)Finland är ett skogsrikt land, men de existerande skogarna utgör dock kvalitativt enformiga habitat och det råder brist på varierade habitat i form av till exempel blandskog, skogar i naturligt tillstånd och äldre skog. Enligt tidigare studier utgör äldre skogar viktiga habitat för många skogslevande arter och mångfalden i skogarna minskar som en följd av habitatförstörelse. Vidare påvisar forskning att skogarna i Finland är ojämnt skyddade och merparten av skogsskyddet förekommer i norra Finland på statlig mark. Forskning påvisar att dagens moderna skogsbruksmetoder bidrar till att våra skogar är artfattiga och till de största hoten mot mångfalden i våra skogar klassas skogsbruket samt klimatförändringen. Att skogslandskapet i Finland utarmas är ett stort problem i och med att mångfald ökar ett ekosystems beständighet mot yttre störningar. För att åtgärda förlusten av biologisk mångfald har genvägar eller verktyg, med vars hjälp större arealer och flera arter samtidigt kan skyddas tagits fram. Till de här verktygen hör konceptet paraplyarter och en paraplyart är kortfattat en art med vars hjälp flera andra arter kan skyddas och vars förekomst indikerar på att ett lokalt habitat i närheten av paraplyarten är av hög kvalitet. Paraplyarter används explicit för att skydda habitat av hög kvalitet till exempel på områden där biodiversiteten är hög. Tidigare studier har visat att stora dagrovfåglar vanligen är effektiva paraplyarter i och med att deras boplatser kan associeras med en hög biodiversitet. Syftet med den här undersökningen är att klargöra huruvida skogslandskap i närheten av duvhökens (Accipiter gentilis) boplatser uppvisar en högre förekomst av vissa arter jämfört med kontrollområden. Kontrollområdena med vilka boplatserna jämfördes var av två slag: (a) äkta skogskontroller och (b) slumpmässigt utvalda skogskontroller. De äkta skogskontrollerna utgjordes av områden som kvalitativt motsvarade skogen intill duvhöksbon, medan de slumpmässigt utvalda skogskontrollerna utgjordes av skog av vilken typ som helst. För vart och ett av de här områdena undersöktes förekomsten av flygekorre (Pteromys volans) och blåbär (Vaccinium myrtillus). Förekomst av flygekorre karterades på basen av spillningsfynd och blåbärsrisets riklighet uppskattades via analyser av fotografier tagna över fältskiktet. Det insamlade materialet för båda arterna analyserades statistiskt med hjälp av LME - modeller. Resultaten påvisade att flygekorre förekom rikligare vid duvhökens boplatser än i de två kontrollskogarna. Flygekorrens habitatpreferenser överlappar till stor del med duvhökens och mina resultat överensstämmer med tidigare forskning som har påvisat att flygekorren har en nytta av duvhökens närvaro i form av skydd mot predation från nattaktiva rovfåglar. Flygekorren prefererar således samma skogstyper som duvhöken och intressant nog verkar duvhökens närvaro i sig vara viktigare för flygekorren än själva skogstypen. Enligt resultaten från avhandlingen är det dock inte heller uteslutet att en annan art/egenskap tillsammans med duvhöken fungerade som en paraplyart för flygekorren. Blåbärsris däremot uppvisade en rikligare förekomst vid äkta skogskontroller och duvhöksskogarna var troligen överlag för lummiga för blåbärets trivsel, men blåbärsris förekom dock rikligt vid 50 m från boplatserna. Således kan slutsatsen dras att flygekorren kan skyddas i fall av att skogslandskap med duvhöksbon skyddas. Duvhöken kan även ses som ett verktyg för naturskydd eller som en indikator för skogslandskap med hög mångfald, i vilka flygekorre samt en del blåbärsris förekommer. Om vi i framtiden vill ha flygekorre och blåbär i våra skogar bör därmed äldre skogar bevaras i och med resultaten påvisar att nämnda arter inte verkar trivas i moderna ekonomiskogar. En tillämpning av resultaten kunde vara att bruka paraplyarter och förekomst av nyckelbiotoper för naturskydd simultant.
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(2017)Tausta ja tavoitteet: D-vitamiinia sitova proteiini (DBP) kuljettaa D-vitamiinia ja sen aineenvaihduntatuotteita, esim. 25-hydroksi-D-vitamiinia (25(OH)D), verenkierrossa. Geneettinen muuntelu GC-geenissä, joka koodaa DBP:tä, on yhdistetty 25(OH)D-konsentraatioiden vaihteluun. Tavoitteena olikin tutkia GC:n geneettisten tekijöiden vaikutusta 25(OH)D-konsentraatioon suomalaisilla vastasyntyneillä. Lisäksi tutkittiin GC:ssä esiintyvän muuntelun aiheuttamia eroja lapsen kasvussa raskauden aikana. Menetelmät: GC:n SNP:ssä, eli yksittäisen nukleotidin vaihdoksissa, esiintyvää polymorfiaa ja raskauden aikaisen 25(OH)D-konsentraation ja napaveren 25(OH)D-konsentraation suhdetta arvioitiin 933 valkoihoisella vastasyntyneellä pojalla (468) ja tytöllä (465). 25(OH)D-konsentraatio määritettiin raskausviikoilta 6-13 ja napaverestä. Äidin D-vitamiinin saanti ravintolisistä määritettiin ruuankäytön frekvenssikyselylomakkeen avulla kahdelta viimeiseltä raskauskuukaudelta. GC:n kolme SNP:tä genotyypitettiin Taq-polymeraasin aktiivisuuteen perustuvan qPCR:n avulla (Bio-Rad, CFX384 C1000 Touch™ Real-Time PCR Detection System, USA): rs4588, rs7041 ja rs705124. Genotyyppien rs4588 ja rs7041 yhdistelmästä muodostettiin kuusi tunnettua diplotyyppiä ja kaikista kolmesta genotyypistä edelleen haplotyypit. Eroja geno-, diplo- ja haplotyyppien välillä 25(OH)D-konsentraatioissa ja kasvussa testattiin kovarianssianalyysin (ANCOVA) avulla käyttämällä relevantteja kovariaatteja. Tulokset: SNP rs4588 ja rs7041 geno- ja diplotyyppien välillä löydettiin tilastollisesti merkitsevä ero 25(OH)D-konsentraatioissa (keskiarvo (nmol/l) ± keskivirhe). Napaveren 25(OH)D-konsentraatio oli alhaisin geno- ja diplotyypillä GC 2/2 (77.8 ± 4.3, p=0.010, p=0.028, ANCOVA) ja korkein diplotyypillä GC 1F/1S (93.8 ± 2.0). Haplotyypillä ATG oli alhaisin napaveren 25(OH)D-konsentraatio (77.9 ± 3.8, p=0.016, ANCOVA). Raskauden ajan 25(OH)D-konsentraatiossa ei löytynyt eroja geno-, diplo- tai haplotyyppien välillä (ANCOVA). Genotyypin rs7041 sekä diplotyyppien ja päänympäryksen välilllä havaittiin ero (p=0.005, p=0.002, ANOVA), mutta geneettisen muuntelun ja syntymäpainon ja –pituuden välillä ei löytynyt eroja. Kun valittiin relevantit kovariaatit, niin genotyyppien rs7041 sekä diplotyyppien ja päänympäryksen välillä ei löytynyt enää tilastollisesti merkitseviä eroja. Yhteenveto: Tulokset osoittavat, että DBP:tä koodaavan geenin geneettisellä muuntelulla ja napaveren 25(OH)D-konsentraatiolla olisi vaikutusta jo vastasyntyneiden D-vitamiinitilaan ja aineenvaihduntaan. Saatujen tulosten perusteella GC:n geneettinen muuntelu ei kuitenkaan ollut yhteydessä lapsen kasvuun. Tulevaisuudessa GC:n SNP:n vaikutusta 25(OH)D-konsentraatioon voisi tarkastella syntymän jälkeisten vuosien aikana.
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(2021)Post-transcriptional modifications (PTMs) in RNA are present in all known RNA species and conserved in all kingdoms of life. Transfer RNA (tRNA) has been shown to have numerous conserved modifications, which exemplifies the importance of modifications having impact on the structure of the tRNA and its function as carrier of the amino acids. Ribosomal RNAs (rRNA) are universally modified as well, and modifications are situated at functionally important spots of the ribosome. Given the fact that types and sites of modifications are conserved, it is likely that these modifications have been selected for and that they optimize the ribosomal structure and functions. Stress, such as temperature or infection by a pathogen, is known to change the presence or abundance of modifications in RNA molecules and thereby affect translation efficacy. In line with that, this master’s thesis project sought to gain insight into the dynamics of PTMs in tRNA and rRNA upon oxidative stress, with the goal of utilizing recently optimized UPLC/MS method for identifying modified ribonucleosides. As the specific aim of the thesis was to estimate the change in PTMs in tRNA and rRNA in response to oxidative stress with 0.5 mM and 2 mM hydrogen peroxide H2O2, 3 immediate goals were: (i) to isolate total tRNA from yeast grown in stress conditions, (ii) to isolate rRNA from yeast 80S ribosomes, and (iii) to identify present modifications using mass spectrometry. Yeast was cultured in presence of H2O2 as a stressor in mentioned concentrations, and both treatments considered showed a difference in survival when compared to the control. Rough cell concentration estimates (OD600) did not show the effect of the stressor on cell survival clearly, but when number of viable cells per mL was estimated, it was clear that growth of the stressed yeast cultures was hindered 2 hours after exposure to H2O2 but recovered during the 24 hours. Firstly, using UPLC/MS analysis, 29 modifications were identified in tRNA from control and H2O2 treated yeast. Most identified modifications showed no change in abundance in treatments, which is to be verified with additional replicates. However, distinct dynamics of stress-related change was found for several modifications, revealing additional modifications that may play a role in stress related modificome reprogramming to the previously known signature modifications of oxidative stress. It was expected that recovery of culture growth after 24 hours may be accompanied with modification level recovery. However, that was not demonstrated here as downregulation at 2 hours followed by upregulation at 24 hours was seen for 2-methylthio-N6-methyladenosine, N4-acetylcytidine and 5-methoxycarbonylmethyl-2-thiouridine, and the reverse was shown for N4-methylcytidine. Upregulation in both time points was also shown here for some modifications. Taken together, these results confirm a complex and dynamic control of tRNA modifications in cellular survival responses. Modifications found to be affected by oxidative stress are most frequently located on the wobble position 34 and anticodon loop position 37, so it is expected that changes in their modification levels could directly affect the tRNA function in translation, making them a specific target for future research. Secondly, modifications in rRNA from control yeast cultures were identified, such as expected methylations of all 4 canonical nucleosides. However, further analysis will be needed to confirm the other identified modifications, due to the potential mRNA and tRNA contamination. Optimizing the method for rRNA modifications identifications by acquiring more modified nucleosides specific for the rRNA to use as standards in the analysis, analyzing rRNA types separately and using tandem mass spectrometry would enable getting a deeper understanding of which modifications are present and where they are positioned. Finally, it would enable reliable identification of the signals of novel modifications present in rRNA, such as the tRNA modification 5-carbamoylmethyluridine signal found here. In conclusion, this thesis work lays the foundation to study the evolutionary conserved function of PTM changes during stress as modulators of translation, using the methodological approaches discussed in-depth within the thesis, primarily to confirm the intriguing results found here.
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