Browsing by Subject "genetiikka"

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  • Översti, Sanni (Helsingin yliopisto, 2021)
    A mitochondrion is a cytoplasmic organelle responsible for the energy production of the eukaryotic cells. Mitochondria contain their own genome, mitochondrial DNA (mtDNA), which is a double-stranded circular molecule. Due to mitochondria’s essential role in metabolism, a cell can contain hundreds of thousands of copies of mitochondrial DNA, depending upon the cell’s energy requirements. In mammals, mtDNA is generally maternally inherited, meaning that it is transmitted from a mother to all of her descendants. Although mtDNA constitutes only a small fraction of the cell genome, it has several qualities which make it widely used in population genetic studies such as uniparental inheritance, and the fact that the mitochondrial genome does not recombine. Moreover, mtDNA has a mutation rate ten times higher than that of the nuclear genome and therefore allows us to trace back matrilineal lineages through generations and subsequently make inferences about maternal ancestors. The human mtDNA sequence consists of approximately 16,570 base pairs and also contains both a coding region and a non-coding control region, the latter constituting around 7% of the whole mtDNA genome. Since mtDNA does not undergo recombination, an individual’s mitochondrial haplotype can be determined simply by the direct sequencing of target amplicons. Haplotypes containing certain defining variants are considered to be descendants of a common ancestor and are classified into haplogroups. The geographical distribution of haplogroups among contemporary populations is well-known – for instance, the majority of Europeans exhibit mitochondrial lineages H, U, J, K, T and V. Ancient DNA research has uncovered that lineage U was already highly prevalent among the earliest hunter-gatherer settlers of Europe, whereas the gradual spread of agriculture from the Near East that started approximately 10,000 years ago brought along new people and hence also novel mitochondrial lineages (H, J, K and T). Previous studies have stated that compared with other European populations, contemporary Finns do not seem to be an exception in terms of mitochondrial genome pool. This is rather surprising, since other genetic markers have revealed that contemporary Finns are characterized by a strong Eastern genetic influence and are distinguishable from other Europeans. Moreover, the evident East-West distinction within Finland, apparent in Y-chromosomes and autosomes, has not been previously identified in the mtDNA. This thesis outlines the mitochondrial DNA variation among present-day Finns in a Bayesian framework. The aims were to evaluate if Finns display a homogeneous geographical distribution of haplogroups and if the mtDNA composition of Finns resembles that of other European populations, as previously suggested. While no spatial differences have previously been detected in the mitochondrial haplogroup frequencies within Finland, a clear geographical distinction arose when clustering haplogroups into ‘hunter-gatherer’ (U and V) and ‘farmer’ associated lineages (H, J, K and T). Whereas the farmer related haplogroups were notably more common in Southwestern Finland, the hunter-gatherer lineages had higher densities in the Northeastern parts compared to the Southwest. Furthermore, utilization of the complete mitochondrial genomes allowed for reassessing the Finnish mtDNA pool on a larger scale. One third of the subhaplogroups in Finland today were characteristic only of Finns, i.e. these lineages were virtually absent from other populations. When further partitioning the Finnish samples based on their inclusion in ‘local’ and ‘non-local’ lineages, two notably different demographic trajectories were obtained. The population history for Finn-characteristic lineages was more in accordance with what is known through other data types, such as Y-chromosomal and archaeological data. In general, the observed geographical within-country deviation in the Finnish mtDNA pool and the high proportion of Finn-characteristic lineages reflected the signals reported from other genetic markers. Alongside Finnish mtDNA, this thesis explores the molecular rate variation among the different subhaplogroups of lineage U. Unexpectedly, a noteworthy discrepancy emerged from the tip calibrated phylogenies: haplogroup U5b had a notably lower substitution rate when compared to U2, U4 and U5a. This lineage-specificity in the rates most likely arose, at least to some extent, from differences in past population dynamics. In particular, U4 and U5a have been associated with the rapid population expansion which occurred during the Bronze Age, whereas the frequency of U5b has remained rather stable. Subsequently, the observed rate of deviation influences the divergence estimates for subhaplogroups, suggesting that U5b emerged considerably earlier than U5a. Since molecular rates are fundamental to several population genetic analyses and the timing of divergence and demographic events relies heavily on the rate used, more attention should be paid to the interlineage molecular rate variation. The results of this thesis demonstrate not only the importance of using complete mtDNA genomes and the appropriate molecular rate, but also the relevance of approaching the data from new angles when assessing the demographic past of mitochondrial lineages.
  • Tegelberg, Saara (Helsingin yliopisto, 2013)
    Progressive myoclonus epilepsy of Unverricht-Lundborg type (EPM1) is an autosomal recessively inherited neurodegenerative disorder caused by mutations in the cystatin B (CSTB) gene. Although the gene behind the disease has long been known, the exact physiological function of CSTB and the molecular pathogenesis in EPM are largely unknown. In my thesis, I have characterized the developmental and degenerative processes in the brains of young, pre-symptomatic and fully symptomatic Cstb-deficient mice. This mouse is an excellent model for EPM1, as its phenotype closely resembles the human disease. To study the spatiotemporal disease progression, we conducted systematic analyses of changes in neurons and glia in Cstb-deficient brains. We saw significant microgliosis already in two-week-old Cstb-deficient animals, before the appearance of myoclonus at 1 month of age. Early microgliosis was followed by astroglial activation along with progressive neuron loss and brain atrophy. The phenotype of activated microglial cells changed during disease progression. To characterize molecular alterations in Cstb-deficient mouse brains, we performed a gene expression profiling. As the cerebellum shows the earliest and most prominent pathological changes, we focused the gene expression analyses on cerebellum of young Cstb-deficient animals. The results implied disturbances in pathways related to synaptic development, structure and function. Immunohisto¬chemical, electrophysiological and ligand binding studies suggested the involvement of especially GABAergic synapses. Data also showed alterations in genes related to the cytoskeleton and intracellular transport, along with pathways implying activation of immune defence. To shed light on the neuron death seen in the brain of Cstb-deficient mouse, we studied the effect of oxidative stress on Cstb-deficient neurons. As CSTB is an inhibitor of cysteine cathepsins and thus is proposed to protect cells from adverse cathepsin functions, we studied also the role of cathepsins in oxidative stress. The results show that oxidative insult increases the expression of both CSTB and cathepsin B. Cstb-deficient cells are sensitized to oxidative stress-induced damage and cell death, which is at least partially mediated through increased cathepsin B activity. These findings in Cstb-deficient mouse suggest that CSTB has an important role in protecting normal cellular integrity and functions along with affecting the development and function of synaptic connections. Our results also imply that microglia have a central role in nervous system damage already before the appearance of symptoms of EPM1. The results of my thesis study have guided the research of our group into new fields by revealing the importance of microglia in the early stages of EPM1. This study also shows that CSTB-deficiency leads not only to neurodegeneration per se, but also results in developmental defects whose consequences manifest later in EPM1 disease progression.
  • Lotsari-Salomaa, Johanna (2015)
    Lynch syndrome (LS) is the most common hereditary colon cancer syndrome with germline mutated DNA mismatch repair (MMR) genes predisposing to early onset colon tumors and various extracolonic tumors. Microsatellite instability (MSI) is a hallmark of the Lynch syndrome, but 15% of sporadic tumors show instability of repeated sequences, too. Endometrial carcinoma is the most common LS related extra-colonial tumor. Whether or not breast carcinoma belongs to the Lynch syndrome tumor spectrum has been under debate because of controversial study results. Epigenetic mechanisms regulate gene expression without altering the DNA sequence. DNA methylation is a well-established epigenetic modification. Colon, endometrial and breast carcinomas and endometrial hyperplasia from 256 Finnish Lynch syndrome families were examined for methylation changes as well as several types of genetic alterations. In addition, Finnish sporadic breast tumors with similar hormone receptor statuses to LS breast carcinomas were analyzed. Sporadic colon tumors with microsatellite unstable (MSI) and microsatellite stable (MSS) subgroups from Finnish and Australian populations were also investigated. The first aim was to investigate whether or not breast tumors belong to the Lynch syndrome tumor spectrum by studying the MMR status and the methylation profiles of several tumor suppressor genes. The second aim was to identify epigenetically regulated miRNA genes relevant for colon and endometrial tumorigenesis. The third aim was to examine tumorigenic mechanisms in the minor subset of colon carcinomas with inactive Wnt signaling. The candidate genes identified in the second and third studies were examined for promoter methylation in patient specimens. Breast carcinomas from LS mutation carriers showed significantly higher frequencies of MSI(35%) and loss of MMR protein expression (65%) compared to breast tumors from proven noncarriers (0%) and sporadic cases (0%). This suggested that breast carcinoma is a likely LS spectrum tumor. MMR deficient breast tumors were diagnosed at age 53. RASSF1, APC, CDH13, and GSTP1 genes were often methylated in LS breast tumors, the same as in tumors from several other organs from LS mutation carriers. The CDKN2B gene showed breast cancer specific methylation. A breast tumor may be the first sign of LS, and therefore female MMR mutation carriers should participate in mammographic surveillance. Novel associations between promoter methylation of tumor-suppressive miRNAs or protein coding tumor suppressor genes and clinical subtypes were observed. miR-132 was frequently hypermethylated in sporadic Finnish and Australian MSI tumors compared to paired normal mucosa. miR-132 hypermethylation correlated with traditional (so called CpG island methylator phenotype; CIMP) markers and were associated with the female gender, later onset, and tumors located in the proximal colon. miR-129-2 hypermethylation was seen in advanced endometrial hyperplasia and may be an early marker for endometrial tumorigenesis. The CMTM3, DGKI, and OPCML genes showed frequent hypermethylation in LS and sporadic colon tumors. The EPCAM and KLK10 genes had LS specific methylation. Colon tumors with inactive Wnt signaling were found to constitute a group of tumors with varying features of chromosomal instability and epigenetic dysregulation.
  • Rautila, Osma (Helsingin yliopisto, 2021)
    Suomalainen perinnöllinen amyloidoosi on kromosomissa 9 vallitsevasti periytyvä systeeminen amyloidoosi, joka tunnetaan myös nimillä Meretojan ja Kymenlaakson tauti. Taudin kuvasi vuonna 1969 Jouko Meretoja, joka arveli potilaiden yhteisen esi-isän olevan lähtöisin 1300- luvulta Kymenlaaksosta. Taudin aiheuttaa pistemutaatio c.640G > A p.D187N gelsoliini-geenissä. Hiljattain osoitettiin SNP-siru genotyypitykseen perustuen, että suomalaisilla potilailla on todennäköisesti yhteinen esi-isä. Tutkimuksen tavoitteena on arvioida, kuinka monen sukupolven päässä sijaitsee suomalaisten gelsoliiniamyloidoosipotilaiden viimeisin yhteinen esi-isä. Toisena tavoitteena on arvioida uuden perustajahaplotyypin säilymiseen perustuvan iänmääritysmenetelmän toimivuutta. Verifioiduksi kontrollimenetelmäksi tämän rinnalle valittiin laajasti iänmääritykseen käytetty alleeliassosiaation hajoamiseen perustuva menetelmä geenin ympärillä sijaitsevia polymorfisia toistojaksoja hyödyntäen. Polymorfisiin toistojaksoihin perustuva menetelmä arvioi etäisyyden yhteiseen esi-isään olevan noin 31 sukupolvea eli noin 775 vuotta. Tämä ei ollut linjassa perustajahaplotyypin säilymiseen nojaavan menetelmän kanssa, joka arvioi etäisyyden yhteiseen esi-isään olevan noin 79 sukupolven päässä.
  • Hytönen, Marjo (Helsingin yliopisto, 2013)
    Since the sequencing of the Canis lupus familiaris genome the dog has become a powerful tool for scientists. Selective breeding has created more than 400 different breeds each representing genetic isolates with breed-specific morphological and behavioral characteristics. Unique population history, available genealogical records, veterinary diagnostics and novel genomic tools greatly facilitate gene mapping studies in dogs. Given that over 600 genetic disorders have been described in dogs and that most of them are similar to human conditions, dogs have emerged as a clinically relevant model for human inherited disorders. This study explores the genetics of three different inherited developmental defects in dogs, caudal dysplasia, ectodermal dysplasia, and mucopolysaccharidosis VII, which all have counterparts in human. In this study, various clinical and pathological techniques have been used to characterize the phenotypes, and genetic methods such as genome-wide association studies and next-generation sequencing to resolve the genetics of the diseases. Moreover, functional studies in mice have been performed to explore the molecular role during embryonic development. The discoveries made here have established the affected breeds as models to further explore disease mechanisms and therapeutic methods, identified new disease pathways, and offered novel approaches for further developmental studies. Furthermore, this work has enabled the development of genetic tests for breeding purposes. Three different phenotypes have been investigated in this study. First, we studied genetics of caudal dysplasia, which in its mildest form is presenting as short-tail phenotype in dogs. A mutation in T (brachyury homolog) was earlier identified to cause this phenotype in Pembroke Welsh Corgis. Homozygous mutations of T in mouse result in severe caudal dysplasia and embryonic lethality suggesting an essential role for the T gene during mammalian development. The presence of the documented T mutation, c.189C>G, was investigated in 23 other breeds demonstrating that short-tailed dogs from 17 breeds were heterozygous for the mutation that associated completely with the phenotype. The homozygous mutation was suggested to be lethal, as no dogs homozygous for the mutation were found and an approximately 30% decrease was seen in the size of Swedish Vallhund litters when both parents were short-tailed. However, short-tailed dogs were found from six breeds that did not carry the known substitution or any other mutations in the T coding regions and therefore other genetic factors are yet to be discovered that affect the development of the posterior mesodermal region. The short-tailed dogs which do not have T mutation will serve as models in future studies to identify possible novel genetic factors for caudal dysplasia and related medical conditions. Second, a new gene was identified for a hairless phenotype and some of its upstream regulators were characterized. Hairless dog breeds show a breed characteristic which is in clinical terms an ectodermal dysplasia. In this study, the causative mutation for canine ectodermal dysplasia (CED) was sought and subsequently the function of this novel gene in the ectodermal organ development was explored. Accordingly, a genome-wide association study was performed and CED was mapped to dog chromosome 17. Haplotype association testing revealed a 160-kb haplotype, which was fine-mapped using three different breeds. The causative genetic mutation for CED was identified as a 7-bp duplication producing a frameshift and premature stop codon in a previously uncharacterized canine gene forkhead box protein I3 (FOXI3). The study provided a novel gene focus to aid research into ectodermal development. Therefore, a detailed expression pattern of murine Foxi3 during the development of the ectodermal organs was constructed and a series of tissue culture experiments and expression analyses with mouse embryos were performed to assess the function of Foxi3 in mammalian embryogenesis. The results suggest that Foxi3 regulates hair follicle and tooth formation as well as the development of mammary and salivary gland, nail, and eye. Ectodysplasin and activin A were identified as upstream regulators of Foxi3. Third, Brazilian Terriers with severe skeletal defects at early puppyhood were identified through information provided by breeders. Subsequently, a major aim of this work was to describe the clinical and pathological features of the syndrome and to identify its genetic cause. Clinicopathological examinations and pedigree analysis demonstrated that the affected puppies had a recessive spondyloepihyseal dysplasia. The disease locus was mapped to chromosome 6 and a mutation leading to pathogenic p.P289L change in a conserved functional domain of β-glucuronidase (GUSB) was identified. Elevated glycosaminoglycans were detected in urine and only a residual β-glucuronidase activity was observed in the serum of the affected dogs, which confirmed the pathogenity of the mutation. GUSB defects result in mucopolysaccharidosis VII (MPS VII) in several species and thus the mutation defined the syndrome as MPS VII in Brazilian Terriers. Overall, this study illustrates how unique morphological diversity and enriched genetic alterations in closed populations can be efficiently harnessed to gain new insights into developmental biology across species. For example, the identification of the CED mutation in FOXI3 revealed a completely novel gene with a previously unknown essential function in ectodermal development. This work has established several novel large animal models to further explore disease mechanisms and to develop therapeutic methods. Moreover, several new DNA tests have been developed for different breeds of dogs to eradicate or, to control better, the conditions through improved breeding plans. This will improve the welfare of our beloved pets.
  • Koskela, Elli (Helsingfors universitet, 2009)
    Strawberry (Fragaria × ananassa) is the most important berry crop cultivated in Finland. Due to the species' economic importance, there is a national breeding programme aimed at extending the cropping season from the current one month to up to three months. This could be achieved by growing cultivars which would initiate flowers throughout the summer months, without the requirement of a period of short days as is the case with currently grown cultivars. The cultivated strawberry is an octoploid and therefore has complex patterns of inheritance. It is desirable to study the genetic mechanisms of flowering in the closely related but diploid species F. vesca (L). In the diploid Fragaria, a mutation in a single locus, namely the SEASONAL FLOWERING LOCUS (Sfl), changes the flowering phenotype from seasonal to perpetual flowering. There is also an array of genetic tools available for F. vesca, which facilitate genetic studies at molecular level. Experiments described here aimed at elucidating the identity of the gene which confers perpetual flowering in F. vesca by exploring the flowering characteristics and genotypes of five F2 populations (crosses between seasonal × perpetual flowering cultivars). The study took advantage of a genetic map for diploid Fragaria, publicly available EST and genomic Fragaria sequences and a recently developed BAC library. Sequence information was used for designing gene–specific primers for a host of flowering–related candidate genes, which were subsequently mapped on the diploid Fragaria genetic map. BAC library was screened with molecular markers supposedly located close to the Sfl, with the aim of positionally cloning the Sfl. Segregation of flowering phenotypes in the five F2 populations showed, that the Sfl indeed controls flowering in all the tested cultivars. A genetic map was constructed of the chromosome with the Sfl, and a positional cloning attempt was initiated with the closest flanking markers. 45 gene–specific primers pairs were designed for 21 flowering–related genes, and eight genes were successfully mapped on the diploid Fragaria map. One of the mapped genes, namely PRR7, located very close to the Sfl, and is a potential candidate for the gene that has evaded identification so far.
  • Salmela, Elina (Helsingin yliopisto, 2012)
    The genetic structure of populations is a potential source of population history information and an important factor in gene mapping studies. The main aim of this thesis was to study the population structure in Finland and Sweden using, for the first time, genome-wide data from thousands of single nucleotide polymorphism (SNP) markers. Furthermore, this thesis introduced a novel gene mapping approach, subpopulation difference scanning (SDS), and tested its theoretical applicability in the Finnish population. The study subjects included 280 Finnish and 1525 Swedish individuals, and genotypes from Russian, German, British and other populations served as reference data. The results revealed that the Finns differed clearly from central Europeans. Within Finland, the genetic difference between eastern and western Finns was striking. The Finns, particularly eastern Finns, also showed reduced genetic diversity as well as an increased genetic affinity to East Asian reference populations. In Sweden, the overall population structure seemed clinal and lacked strong borders. The population in southern Sweden was relatively homogeneous and genetically close to the Germans and British, while the northern subpopulations differed from the south and also from each other. Overall, these results are congruent with earlier observations from smaller numbers of markers and with population history, particularly the small population sizes that have led to genetic drift. The genetic substructure within Finland could prove useful in mapping genes for diseases that show regional incidence differences. Because a gene underlying an incidence difference must itself harbor a frequency difference, the SDS approach proposes that such genes could be mapped by focusing further analyses on genome areas that show a (sufficient) difference between the high- and low-incidence subpopulations. Population simulations demonstrated that the SDS approach may work for cardiovascular diseases, which have an east-west difference within Finland. In summary, the results of this thesis emphasize the capacity of genome-wide SNP data to detect patterns of population structure, also in populations that have often been assumed homogeneous, such as the Finns and Swedes. Obviously, knowledge of genome-wide population structure has immediate relevance also to studies focusing on diseases and other important phenotypes.
  • Peuralinna, Terhi (Helsingin yliopisto, 2015)
    This thesis project aimed to study the genetic background of common neurodegenerative diseases such as Alzheimer's disease (AD), dementia with Lewy bodies (DLB) and amyotrophic lateral sclerosis (ALS) in the Finnish population. In the first study, we analysed the role of the amyloid precursor protein (APP) and apolipoprotein E (APOE) genes in AD neuropathology. Mutations in the APP gene cause early-onset familial AD and there is some evidence that APP gene variants would play a role in late-onset AD, too. We genotyped more than 50 common variations in the APP gene and sequenced the APP promoter area to detect rare variations in the area, both known and new. In the Finnish elderly population we did not find any of the APP variations to clearly associate with neuropathologically diagnosed AD or with any of the neuropathological features of AD such as cortical β-amyloid, cerebrovascular β-amyloid or neurofibrillary tangles. In addition, the APOEε4 results were updated, using the whole Vantaa85+ cohort. APOE ε4 is currently the strongest known risk factor for late-onset AD. We found a very strong association of APOE ε4 to all neuropathological features of AD. In the second study, we investigated how the common variants in the α-synuclein (SNCA) gene affect different brain pathologies. -synuclein is the main component of the Lewy bodies, which are the pathological hallmarks of PD and DLB, and which are sometimes found also in AD. We genotyped 11 SNPs from the SNCA gene region. We found no association with the cortical β-amyloid and only a faint one with Lewy related pathology. However, we found an association with neurofibrillary tangles. The SNP (rs2572324) associated with a p=0.004 after the Bonferroni correction. The two-locus analysis suggest an independent effect of APOE ε4 (OR=8.67) and rs2572324 (OR=3.36). In the subjects with both risk factors the effect was almost multiplicatively increased (OR=23.3). 35 subjects out of 38 with both risk factors had severe tau-pathology. These results demonstrated the first evidence for a role of genetic variation in SNCA in tau pathology. Moreover, β-amyloid pathology was not associated with the SNCA variants, demonstrating a dissection of genetic effect on the two principal pathological features of AD. In the third study, we performed a whole-genome genotyping using Finnish ALS samples. The Vantaa85+ study subjects were used as controls. This was the first ALS genome-wide association study to find a genome-wide significant association. The location associated with familial ALS was on the chromosome 9p21, which had been noticed before, but the area had been much larger. The area has also been associated with frontotemporal dementia (FTD). Now we managed to define the area with a 42 SNP haplotype. This considerably reduced the area of interest (down to 232 kb) and increased the possibility to find the mutation behind the disease. This haplotype was found in around 40% of the Finnish familial ALS cases and likely explains the high rate of ALS as well as FTD in Finnish population. In the fourth study, we performed a genome-wide association study of DLB in the Vantaa85+ cohort and found two novel areas to be associated with DLB: The c2p21 location with 9 SNP haplotype (p=5.2x10-7) and the c6p21 location with 6 SNP haplotype (p=1.3x10-7). The c6p21 was significant at the genome-wide level. The c2orf21 haplotype has two genes on its area, c2orf72 and SPTBN1. SPTBN1 is the candidate gene since it encodes beta-spectrin, a component of Lewy bodies, while c2orf72 is barely expressed in the brain. The c6p21 associated haplotype block is located in the HLA region and includes HLA-DPB1 and -DPA1 genes. These studies show that the Finnish population is well-suited to study the genetics of neurodegeneration. Our results showed that neuropathologically defined parameters and diagnoses are strongly associated with genetic risk factors, even with relatively low numbers of samples. Hence, phenotypic precision (pathology) is an important element of the statistical power of a study.
  • Jallinoja, Piia (2002)
    The present study focuses on the introduction both of genetic screening and testing and of the related counselling in Finnish health care during the 1990s, i.e. clinical genetics. In addition, the formation of medical ethics as it pertains to clinical genetics is analysed from 1970 onwards. The study consists of five original publications and a summary. The material consists of data collected by means of questionnaires (conducted among lay people, public health nurses and midwives), semistructured interviews (of physicians, public health nurses, midwives and psychologists) and text material collected in connection with four genetic screening and counselling projects carried out in Finland. In general, Finns approve of gene tests, but this general approval is combined with disapproval of certain aspects and prospects of such tests. Furthermore, clinical genetics creates agonizing situations where all options have undesirable consequences. Abortion because of a disorder of the fetus is an especially perplexing issue reflecting the ambivalent status of the fetus. Within the field of medicine, problems related to the increasing number of applications of clinical genetics have been addressed to an increasing extent by developing ethical guidelines and recommendations, by establishing ethics committees and by encouraging public discussion on ethics. In the guidelines, voluntariness and autonomy are repeated principles, and they are also widely accepted principles among lay people and professional groups. Choice may even be regarded as an imperative. What is feared is its opposite, concretized in the fear of eugenics. The situation is paradoxical, however, and characteristic of modern societies: On the on hand there are constant demands for more ethical rules - on the other hand these rules cannot but remain superficial, since they have to be open and flexible in the face of new scientific discoveries and individuals' life situations. A contradiction exists between modern ethics setting the rules for advancing genetic technologies and the real-life situations of clients seeking genetic screening and counselling. Individuals are at the same time struggling to form their own choice and influenced by a plethora of expectations, interests, medical determinants and technological conditions.
  • Borze, Ioana (Helsingin yliopisto, 2011)
    Myeloproliferative neoplasms (MPN) and myelodysplastic syndromes (MDS) are a heterogeneous group of clonal hematopoietic disorders whose etiology and molecular pathogenesis are poorly understood. During the past decade, enormous developments in microarray technology and bioinformatics methods have made it possible to mine novel molecular alterations in a large number of malignancies, including MPN and MDS, which has facilitated the detection of new prognostic, predictive and therapeutic biomarkers for disease stratification. By applying novel microarray techniques, we profiled copy number alterations and microRNA (miRNA) expression changes in bone marrow aspirate and blood samples. In addition, we set up and validated an miRNA expression test for bone marrow core biopsies in order to utilize the large archive material available in many laboratories. We also tested JAK2 mutation status and compare it with the in vitro growth pattern of hematologic progenitors cells. In the study focusing on 100 MPN cases, we detected a Janus kinase 2 (JAK2) mutation in 71 cases. We observed spontaneous erythroid colony growth in all mutation-positive cases in addition to nine mutation negative cases. Interestingly, seven JAK2V167F negative ET cases showed spontaneous megakaryocyte colony formation, one case of which also harbored a myeloproliferative leukemia virus oncogene (MPL) mutation. We studied copy number alterations in 35 MPN and 37 MDS cases by using oligonucleotide-based array comparative hybridization (array CGH). Only one essential thrombocythemia (ET) case presented copy number alterations in chromosomes 1q and 13q. In contrast, MDS cases were characterized by numerous novel cryptic chromosomal aberrations with the most common copy number losses at 5q21.3q33.1 and 7q22.1q33, while the most common copy number gain was trisomy 8. As for the study of the bone marrow core biopsy samples, we showed that even though these samples were embedded in paraffin and underwent decalcification, they were reliable sources of miRNA and suitable for array expression analysis. Further, when studying the miRNA expression profiles of the 19 MDS cases, we found that, compared to controls, two miRNAs (one human Epstein-Barr virus (miR-BART13) miRNA and one human (has-miR-671-5p) miRNA) were downregulated, whereas two other miRNAs (hsa-miR-720 and hsa-miR-21) were upregulated. However, we could find no correlation between copy number alterations and microRNA expression when integrating these two data. This thesis brings to light new information about genomic changes implicated in the development of MPN and MDS, and also underlines the power of applying genome-wide array screening techniques in neoplasias. Rapid advances in molecular techniques and the integration of different genomic data will enable the discovery of the biological contexts of many complex disorders, including myeloid neoplasias.
  • Kallio, Niko (Helsingin yliopisto, 2020)
    Genomiikka keskittyy geenien ja niiden ilmentymisen systemaattiseen analyysiin ja genomin tutkimus on yhdistänyt tuhansia geneettisiä alueita yleisiin sairauksiin ja ominaisuuksiin. Perimä ja ympäristötekijät vaikuttavat yhdessä sairauksien syntyyn ja siksi perinnöllisten tekijöiden tarkka tuntemus on tärkeää. Perimätiedon avulla voidaan kehittää uusia ja parantaa olemassaolevia työkaluja ennustavaan, ennaltaehkäisevään, henkilökohtaiseen ja osallistavaan lääketieteeseen (P4-lääketiede). Tässä tutkimuksessa keskityimme laskimotukosriskiin ja pyrimme arvioimaan onko jo julkaistuilla monigeenisillä riskisummilla ennustavaa arvoa suomalaisessa väestössä ja pohdimme voidaanko tietoa hyödyntää P4-lääketieteessä. Käytimme FINRISK-aineistoa, jossa oli 255 laskimotukokseen sairastuneen ja 20160 sairastumattoman näytteet ja arvioimme elinaika-analyysillä kahden monigeenisen riskisumman ennustearviota suhteessa laskimotukosriskiin. Tutkimuksen tuloksena oli, että monigeenisillä riskisummilla ei saatu lisäarvoa kun otettiin huomioon yksittäinen vaikutukseltaan voimakas mutaatio (FV Leiden -mutaatio). Kun halutaan käyttää monigeenisiä riskisummia työkaluna kliinisessä työssä pitää niiden arvo ja vaikutus tuntea tarkasti. Tutkimus osoittaa että tähän tutkimukseen valikoiduilla monigeneenisillä riskisummilla ei ole toivottua ennustearvoa suomalaisessa väestössä ja suomalaiseen väestöön paremmin sopivat monigeeniset riskisummat pitää luoda.
  • Forsström, Linda (Helsingin yliopisto, 2012)
    ABSTRACT Idiopathic developmental disorders (DDs) affect ~1% of the population worldwide. This being a considerable amount, efforts are being made to elucidate the disease mechanisms. One or several genetic factors cause 30-40% of DDs, and only 10% are caused by environmental factors. The remaining 50% of DD patients go undiagnosed, mostly due to a lack of diagnostic techniques. The cause in most undiagnosed cases is though to be a genetic factor or a combination of genetic and environmental factors. Despite the surge of new technologies entering the market, their implementation into diagnostic laboratories is hampered by costs, lack of information about the expected diagnostic yield, and the wide range of selection. This study evaluates new microarray methods in diagnosing idiopathic DDs, providing information about their added diagnostic value. Study I analysed 150 patients by array comparative genomic hybridization (array CGH, 44K and 244K), with a subsequent 18% diagnostic yield. These results are supported by other studies, indicating an enourmous added diagnostic value of array CGH, compared with conventional cytogenetic analysis. Nevertheless, 80% of the patients remained undiagnosed in Study I. In an effort to diagnose more patients, in Study IV the resolution was increased from 8.9 Kb of the 244K CGH array to 0.7 Kb, by using a single-nucleotide polymorphism (SNP) array. However, no additional pathogenic changes were detected in the 35 patients assessed, and thus, for diagnostic purposes, an array platform with ca 9 Kb resolution appears adequate. The recent vast increase in reports of detected aberrations and associated phenotypes has enabled characterization of several new syndromes first based on a common aberration and thereafter by delineation of common clinical characteristics. In Study II, a familial deletion at 9q22.2q22.32 with variable penetrance was described. Despite several reports of aberrations in the adjacent area at 9q associated with Gorlin syndrome, the patients in this family had a unique phenotype and did not present with the syndrome. In Study III, a familial duplication of chromosome 6p22.2 was described. The duplication caused increased expression of an important enzyme of the γ-aminobutyric acid (GABA) degradation pathway, causing oxidative stress of the brain, and thus, very likely, the mild mental retardation of these patients. These two case studies attempted to pinpoint candidate genes and to resolve the pathogenic mechanism causing the clinical characteristics of the patients. Presenting rare genetic and clinical findings to the international science and medical community enables interpretation of similar findings in other patients. The added value of molecular karyotyping in patients with idiopathic DD is evident. As a first line of testing, arrays with a median resolution of at least 9 Kb should be considered and further characterization of detected aberrations undertaken when possible. Diagnostic whole-exome sequencing may be the best option for patients who remain undiagnosed after high-resolution array analysis.
  • Tadji, Ahmed Abdessalam (Helsingin yliopisto, 2021)
    Tutkimuksen tavoitteena on selvittää suomalaisten juveniiliglaukoomaa sairastavien potilaiden genetiikkaa. Juveniiliglaukooma on 4-40-vuotiaana diagnosoitu primaarisen avokulmaglaukooman harvinainen alatyyppi, jonka kliininen kuva on usein hankalampi kuin tavanomaisen avokulmaglaukooman. Avokulmaglaukooma on silmän verkkokalvon hermorakenteita rappeuttava krooninen sairaus, jota yleensä tavataan ikääntyneellä väestöllä. Juveniiliglaukoomassa keskeiset tautimuutokset -ja löydökset ovat pääosin yhtenevät avokulmaglaukooman kanssa, mutta ne alkavat nuoremmalla iällä. Juveniiliglaukooman periytyvyys on huomattavasti merkittävämpää kuin primaarisen avokulmaglaukooman. Aikaisemmissa tutkimuksissa juveniiliglaukoomaan on yhdistetty erityisesti MYOC-geeni, jonka patogeeniset variantit eli tautia aiheuttavat muutokset ovat tärkein yksittäinen tautia aiheuttava geenimuutosten ryhmä. Glaukooma on maailmanlaajuisesti yleisin sokeutta aiheuttava silmäsairaus, eikä sen syntymekanismeja vuosikymmenien ajan tutkimustyöstä huolimatta vieläkään tunneta tarkasti. Geneettisen tutkimustyön merkitys sairauksien - mukaan lukien juveniiliglaukooman - syvällisemmässä ymmärtämisessä on korostunut viime vuosina. HUS:n Silmätautien klinikan vuosien 2014-2018 glaukoomapoliklinikan käynneistä on kerätty 65 potilasta, joilla on juveniiliglaukooma. Heistä on sekvensoinnilla määritetty tähän asti tunnetut tautiin yhdistetyt geenit. MYOC-geenin patogeeninen variantti löytyi kuudelta tutkittavalta (9,7%). Kirjallisuudessa tähän asti MYOC-geenin mutaatiot ovat selittäneet 6-36% potilaiden juveniiliglaukoomista, mihin tämänkin tutkimuksen löydös täsmää. Lisäksi yhdellä tutkittavalla todettiin sporadinen LTBP2-geenin patogeeninen variantti, ja kahdella keskenään lähisukua olevalla todettin FOXC1-geenin patogeeninen variantti. Tunnetut geenit selittävät yhteensä 12,9% nuoruustyypin glaukoomasta Suomessa. (186 sanaa)
  • Törmälä, Ritva (University of Helsinki, 1997)
  • Koskenranta, Anna (Helsingin yliopisto, 2020)
    Hammaspuutokset ovat yleisin kraniofakiaalinen anomalia. Ne voivat esiintyä sekä erillisinä anomalioina että oireyhtymien yhteydessä. Niiden vaikutus elämänlaatuun on negatiivinen muun muassa heikentyneen suunterveyden ja sosiaalisten suhteiden vaikeutumisen kautta. Pekka Niemisen tutkimusryhmä Helsingin yliopistosta löysi hammaspuutospotilaiden perimätutkimuksissa kaksi uutta mahdollisesti hammaspuutoksia aiheuttavaa geenimutaatiota geenissä RNF43. RNF43-proteiini on normaalisti toimiessaan osa hampaiden kehitykseen ja kasvuun vaikuttavan WNT-viestinnän negatiivista takaisinkytkentää. Mutaatioiden arvellaan häiritsevän hampaiden kehitystä WNT-viestinnän aktiivisuuden muuntelemisen kautta. Tutkimuksessamme tutkimme mutaatioiden vaikutusta RNF43-proteiinin määrään ja sijoittumiseen solussa ja pyrimme näin todistamaan mutaatioiden patogeenisyyden. Käytimme tutkimuksessa HEK293-soluja. Transfektoimme niitä villityypin ja kahden eri mutaation plasmideilla, jotka sisälsivät myös GFP-proteiinin. Värjäsimme niitä membraani- ja tumaväriaineilla, jotta pystyimme hahmottamaan GFP:n ja siten proteiinin sijaintia solussa fluoresenssi- ja konfokaalimikroskooppien avustuksella. Mutaatioilla ei havaittu olevan vaikutusta RNF43:n tuotannon määrään tai sijoittumiseen soluissa, emmekä näin ollen voineet tässä tutkimuksessa todentaa niiden patogeenisyyttä. Veimme tutkimuksellamme kuitenkin kyseisten mutaatioiden ja hammaspuutosten genetiikan tutkintaa eteenpäin. Tästä tutkimuksesta saatujen havaintojen pohjalta voidaan RNF43-geenin mutaatioiden tutkimusta jatkaa muilla menetelmillä.
  • Ainasoja, Miia (Helsingin yliopisto, 2008)
    Plants produce a diversity of secondary metabolites, i.e., low-molecular-weight compounds that have primarily ecological functions in plants. The flavonoid pathway is one of the most studied biosynthetic pathways in plants. In order to understand biosynthetic pathways fully, it is necessary to isolate and purify the enzymes of the pathways to study individual steps and to study the regulatory genes of the pathways. Chalcone synthases are key enzymes in the formation of several groups of flavonoids, including anthocyanins. In this study, a new chalcone synthase enzyme (GCHS4), which may be one of the main contributors to flower colour, was characterised from the ornamental plant Gerbera hybrida. In addition, four chalcone synthase-like genes and enzymes (GCHS17, GCHS17b, GCHS26 and GCHS26b) were studied. Spatial expression of the polyketide synthase gene family in gerbera was also analysed with quantitative RT-PCR from 12 tissues, including several developmental stages and flower types. A previously identified MYB transcription factor from gerbera, GMYB10, which regulates the anthocyanin pathway, was transferred to gerbera and the phenotypes were analysed. Total anthocyanin content and anthocyanidin profiles of control and transgenic samples were compared spectrophotometrically and with HPLC. The overexpression of GMYB10 alone was able to change anthocyanin pigmentation: cyanidin pigmentation was induced and pelargonidin pigmentation was increased. The gerbera 9K cDNA microarray was used to compare the gene expression profiles of transgenic tissues against the corresponding control tissues to reveal putative target genes for GMYB10. GMYB10 overexpression affected the expression of both early and late biosynthetic genes in anthocyanin-accumulating transgenic tissues, including the newly isolated gene GCHS4. Two new MYB domain factors, named as GMYB11 and GMYB12, were also upregulated. Gene transfer is not only a powerful tool for basic research, but also for plant breeding. However, crop improvement by genetic modification (GM) remains controversial, at least in Europe. Many of the concerns relating to both human health and to ecological impacts relate to changes in the secondary metabolites of GM crops. In the second part of this study, qualitative and quantitative differences in cytotoxicity and metabolic fingerprints between 225 genetically modified Gerbera hybrida lines and 42 non-GM Gerbera varieties were compared. There was no evidence for any major qualitative and quantitative changes between the GM lines and non-GM varieties. The developed cell viability assays offer also a model scheme for cell-based cytotoxicity screening of a large variety of GM plants in standardized conditions.
  • Rämö, Joel (Helsingfors universitet, 2016)
    Familial combined hyperlipidemia (FCH) is a complex and common familial dyslipidemia characterized by elevated total cholesterol and/or triglyceride levels with over five-fold risk of coronary heart disease. The genetic architecture and contribution of rare Mendelian and common variants to FCH susceptibility is unknown. In 53 Finnish FCH families, we genotyped and imputed nine million variants in 715 family members with DNA available. We studied the enrichment of variants previously implicated with monogenic dyslipidemias and/or lipid levels in the general population by comparing allele frequencies between the FCH families and population samples. We also constructed weighted polygenic scores using 212 lipid-associated SNPs and estimated the relative contributions of Mendelian variants and polygenic scores to the risk of FCH in the families. We identified, across the whole allele frequency spectrum, an enrichment of variants known to elevate, and a deficiency of variants known to lower LDL-C and/or TG levels among both probands and affected FCH individuals. The score based on TG associated SNPs was particularly high among affected individuals compared to non-affected family members. Out of 234 affected FCH individuals across the families, seven (3%) carried Mendelian variants and 83 (35%) showed high accumulation of either known LDL-C or TG elevating variants by having either polygenic score over the 90th percentile in the population. The positive predictive value of high score was much higher for affected FCH individuals than for similar sporadic cases in the population. FCH is highly polygenic, supporting the hypothesis that variants across the whole allele frequency spectrum contribute to this complex familial trait. Polygenic SNP panels improve identification of individuals affected with FCH, but their clinical utility remains to be defined.
  • Tomppo, Liisa (Helsingin yliopisto, 2010)
    Schizophrenia is a severe psychotic disorder affecting 0.5-1 % of the population. The disorder is characterized by hallucinations; delusions; disorganized behavior and speech; avolition; anhedonia; flattened affect and cognitive deficits. The etiology of the disorder is complex with evidence for multiple genes contributing to the onset of the disorder along with environmental factors. DISC1 is one of the most promising candidate genes for schizophrenia. It codes for a protein which takes part in numerous molecular interactions along several pathways. This network, termed as the DISC1 pathway, is evidently important for the development and maturation of the central nervous system from the embryo until young adulthood. Disruption at these pathways is thought to predispose schizophrenia. In the present study, we have studied the DISC1 pathway in the etiology of schizophrenia in the Finnish population. We have utilized large Finnish samples; the schizophrenia family sample where DISC1 was originally shown to associate with schizophrenia and the Northern Finland birth cohort 1966 (NFBC66). Several DISC1 binding partners displayed evidence for association in the family sample along with DISC1. Through a genome-wide linkage study, we found a significant linkage signal to a locus where a DISC1 binding partner NDE1 is located at the carriers of a certain DISC1 risk variant. In a follow-up study, genetic markers in NDE1 displayed significant evidence for association with schizophrenia. Further exploration of association between 11 genes of the DISC1 pathway and schizophrenia led to recognition of novel variants in NDEL1, PDE4B and PDE4D that significantly either increased or decreased the risk for schizophrenia. Further, we found evidence that DISC1 itself has a significant role in the human mental functioning even in the healthy population. Variants in DISC1 had a significant effect on anhedonia which is a trait present at everybody but is in its severe form one of the main symptoms of schizophrenia and correlates with the risk of developing the disorder. Further, utilizing genome-wide marker data, we recognized three genes; MIR620; CCDC141 and LCT; that are closely related to the DISC1 pathway but which effects on anhedonia were observable only at the individuals who carried these specific DISC1 variants. Our findings significantly add up to the previous evidence for the involvement of DISC1 and the DISC1 pathway in the etiology of schizophrenia and psychosis. Our results support the concept of a number of DISC1 pathway related genes contributing in the etiology of schizophrenia along with DISC1 and provide new candidates for the studies of schizophrenia. Our findings also significantly increase the importance of DISC1 itself as having a role in psychological functioning in the general population.
  • Anderson, Heidi (Helsingin yliopisto, 2008)
    New blood cells are continuously provided by self-renewing multipotent hematopoietic stem cells (HSC). The capacity of HSCs to regenerate the hematopoietic system is utilized in the treatment of patients with hematological malignancies. HSCs can be enriched using an antibody-based recognition of CD34 or CD133 glycoproteins on the cell surface. The CD133+ and CD34+ cells may have partly different roles in hematopoiesis. Furthermore, each cell has a glycome typical for that cell type. Knowledge of HSC glycobiology can be used to design therapeutic cells with improved cell proliferation or homing properties. The present studies characterize the global gene expression profile of human cord blood-derived CD133+ and CD34+ cells, and demonstrate the differences between CD133+ and CD34+ cell populations that may have an impact in transplantation when CD133+ and CD34+ selected cells are used. In addition, these studies unravel the glycome profile of primitive hematopoietic cells and reveal the transcriptional regulation of N-glycan biosynthesis in CD133+ and CD34+ cells. The gene expression profile of CD133+ cells represents 690 differentially expressed transcripts between CD133+ cells and CD133- cells. CD34+ cells have 620 transcripts differentially expressed when compared to CD34- cells. The integrated CD133+/CD34+ cell gene expression profiles proffer novel transcripts to specify HSCs. Furthermore, the differences between the gene expression profiles of CD133+ and CD34+ cells indicate differences in the transcriptional regulation of CD133+ and CD34+ cells. CD133+ cells express a lower number of hematopoietic lineage differentiation marker genes than CD34+ cells. The expression profiles suggest a more primitive nature of CD133+ cells. Moreover, CD133+ cells have characteristic glycome that differ from the glycome of CD133- cells. High mannose-type and biantennary complex-type N-glycans are enriched in CD133+ cells. N-glycosylation-related gene expression pattern of CD133+ cells identify the key genes regulating the CD133+ cell-specific glycosylation including the overexpression of MGAT2 and underexpression of MGAT4. The putative role of MAN1C1 in the increase of unprocessed high mannose-type N-glycans in CD133+ cells is also discussed. These studies provide new information on the characteristics of HSCs. Improved understanding of HSC biology can be used to design therapeutic cells with improved cell proliferation and homing properties. As a result, HSC engineering could further their clinical use.