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  • Rautiola, Juhana (2015)
    Renal cell carcinoma (RCC) is the seventh most common cancer in men and the ninth most common in women and it comprises 2-3% of all malignant tumors in adults. In its localized form, the five-year survival is almost 60%. However, in metastatic renal cell carcinoma (mRCC), the five-year survival has historically remained under 10%. Renal cell carcinoma is highly tolerant to traditional chemotherapy and and recent treatment has instead relied on inhibiting the vascular endothelial growth factor (VEGF) and mammalian target of rapamycin (mTOR) signaling pathways. Since 2005, seven new agents have been approved for the treatment of metastatic RCC. Despite the increased knowledge of RCC, the molecular mechanisms behind acquired and intrinsic resistance to treatment remain largely unknown. Our results demonstrate that hypertension is a strong predictor of good prognosis and response to sunitinib treatment. In addition to hypertension we identified neutropenia and thrombocytopenia as predictive markers of good prognosis. A novel, triple biomarker profile consisting of these adverse events strongly predicts improved survival and outcome in mRCC patients. Results from our investigation of sequential treatment suggest that a newer VEGF receptor inhibitor, pazopanib, has clinically meaningful activity in a patient population where mRCC has progressed through sunitinib treatment and in patients who do not tolerate sunitinib. Encouraged by the fuller understanding of tumor biology, we then investigated the role of an endothelial-cell derived growth factor, angiopoietin-2 (Ang2), on patients outcome. High expression of Ang2 was demonstrated to correlate with a high vessel density and to predict a better response to sunitinib-based first-line therapy.
  • Lindh, Erika (Hansaprint, 2015)
    Influenza A virus (IAV) is a significant zoonotic pathogen, with a diverse range of subtypes infecting both humans and birds. NDV is the causative agent of Newcastle Disease, a significant infectious disease of poultry. While wild waterfowl, the natural host, remains apparently healthy during the even frequent infections with these viruses, poultry is susceptible for mild to severe disease. Both viruses have a heavy impact on the poultry industry by causing outbreaks with significant economical losses. Low pathogenic IAV and NDV viruses are efficiently transmitted in host populations and have the ability to mutate and become virulent. The segmented genome of IAV also allows it to reassort during dual infections, giving rise to novel viruses with unpredictable pathogenic properties including the potential to transmit to humans and other mammals. Extensive surveillance studies of IAV and NDV during the past decades have revealed their wide global distribution and host species range. Previous surveillance studies have yielded limited information about avian IAV in Finland and so far, NDV has been the causative agent of varying outbreaks in Finland, for example in poultry, a zoo and domestic pigeons. This project was initiated for surveillance purposes, to gather information about prevalence and subtype/genotype distribution of IAVs and NDVs and to assess the potential presence of highly pathogenic strains in wild waterfowl in Finland. Through annual sampling and screening of wild waterfowl, 875 birds were screened during the years 2006-2010 and 2014. The birds represent mainly young, hunted ducks from local breeding areas. We detected altogether 76 IAVs (in 8.7%) and 39 NDVs (in 5.5%). From most of the samples (75%) the viruses were successfully propagated in embryonated chicken eggs and partially sequenced for phylogenetic and pathogenicity analyses. Importantly, while no highly pathogenic strains were encountered, several of the wild waterfowl derived viruses detected in Finland were phylogenetically closely related to viruses detected during outbreaks in Finland and elsewhere in Europe. These viruses includ IAV subtypes H5N2, H7N3, H9N2 as well as NDVs. An unexpectedly high subtype dominance of H3N8 was recorded each year, counting for over 60% of all the subtyped IAVs. Genetic characterization of these viruses demonstrated high sequence homology, even between temporally separated viruses and suggests local perpetuation of the viruses. The past decades have witnessed an increase in the incidence of IAV outbreaks in poultry in Europe and zoonotic transmission of a growing number of subtypes. Surveillance data of IAV in the natural host is important for designing efficient national policies aiming to control the transmission of IAV to susceptible populations and for risk assessment. Our results also show that the northern breeding sites have implications on a European scale.
  • Hankaniemi, Minna (2012)
    Bone morphogenetic protein 15 (BMP15) and growth differentiation factor 9 (GDF9) are growth factors expressed by oocytes and are among the key regulators transmitting the signaling between the oocyte and the surrounding granulosa cells. Genetic studies have shown that these factors are essential regulators of normal fertility in mammals. BMP15 gene deficient mice exhibit a mild reduction in fertility, whereas sheep homozygous for a mutant form of BMP15 are sterile. In the case of GDF9, knockout mice and homozygous mutant sheep are also sterile the ovaries being non functional due to an early block in folliculogenesis. In humans, aberrant expression of these factors has been involved in premature ovarian failure (POF), polycystic ovary syndrome (PCOS) or dizygotic twinning. ----- This thesis was aimed to define the signaling and activation of human BMP15 (hBMP15) and GDF9 (hGDF9) proteins. The lack of bioactive hGDF9 had restricted the in vitro studies previously and the first aim of this thesis was to produce and purify bioactive hGDF9 protein. We found that hGDF9 is secreted in a latent form. Although mouse GDF9 (mGDF9) and hBMP15 are produced in active form their bioactivity is lost when a C-terminal affinity purification tag is added. We found also, that an N-terminal epitope tag does not have a deleterious effect on the bioactivity of mGDF9 or hBMP15 and they can be purified using immobilized metal affinity (IMAC) and high performance liquid chromatography approaches. A purified wild type hGDF9 (hGDF9wt) mature region became available to us during this thesis project and we found that it is a highly bioactive and stable protein activating the Smad3/4 signaling pathway. Subsequently we produced and purified a bioactive wild type hBMP15 (hBMP15wt) form, which was highly bioactive and activated the Smad1/5/8 signaling pathway in human granulosa luteal (hGL) cells. Physico-chemical characterization of BMP15 revealed that it consists of P16 and P17 forms, where P16 seems to be phosphorylated and P17 glycosylated. ------ It has been shown previously that a recombinant BMP type II receptor ectodomain Fc fusion protein (BMPR2ecd Fc) is able to inhibit the actions of BMP15 and GDF9 in vitro. For studying the effect of BMPR2ecd Fc on mouse folliculogenesis, we produced and purified this protein. These in vivo studies demonstrated that by administering the BMPR2ecd-Fc protein, we were able to dose-dependently modulate ovarian folliculogenesis in mice. -------- In order to determine the cell surface receptor binding components for human BMP15, we engineered and produced a covalent dimer of hBMP15 and labeled it radioactively. We found that it behaves biologically in a similar manner as the hBMP15wt. Human BMP15 strongly bound to BMPR1B and BMPR2 on the COV434 cell surface, an association which was further verified in COS-7 cells by overexpression of these receptors. In conclusion, we have for the first time produced and characterized recombinant human GDF9 and BMP15 protein in native form and thus developed valuable tools for studies aiming at understanding their role in human reproductive medicine. The production and purification methods developed during this thesis are widely applicable for the isolation and characterization of comparable oocyte growth factors in other species and thus have wide applicability in reproductive biology studies in the future.
  • Isohanni, Pirjo (2010)
    Childhood-onset mitochondrial diseases comprise a heterogeneous group of disorders, which may manifest with almost any symptom and affect any tissue or organ. Due to challenging diagnostics, most children still lack a specific aetiological diagnosis. The aim of this thesis was to find molecular causes for childhood-onset mitochondrial disorders in Finland. We identified the underlying cause for 25 children, and found three new diseases, which had not been diagnosed in Finland before. These diseases caused severe progressive infantile-onset encephalomyopathies, and were due to defects in mitochondrial DNA (mtDNA) maintenance. Furthermore, the thesis provides the molecular background of Finnish patients with ‘leukoencephalopathy with brain stem and spinal cord involvement and elevated brain lactate’ (LBSL). A new phenotype was identified to be due to mutations in Twinkle, resembling ‘infantile onset spinocerebellar ataxia’ (IOSCA). These mutations caused mtDNA depletion in the liver, thus confirming the essential role of Twinkle in mtDNA maintenance, and expanding the molecular background of mtDNA depletion syndromes. The major aetiology for infantile mitochondrial myopathy in Finland was discovered to be due to mutations in thymidine kinase 2 (TK2). A novel mutation with Finnish ancestry was identified, and a genotype-phenotype correlation with mutation-specific distribution of tissue involvement was found, thus proving that deficient TK2 may cause multi-tissue depletion and impair neuronal function. This work established the molecular diagnosis and advanced the knowledge of phenotypes among paediatric patients with polymerase gamma (POLG) mutations. The patients showed severe early-onset encephalopathy with intractable epilepsy. POLG mutations are not a prevalent cause of children’s ataxias, although ataxia is a major presenting symptom among adults. Our findings indicate that POLG mutations should be investigated even if typical MRI, histochemical or biochemical abnormalities are lacking. LBSL patients showed considerable variation in phenotype despite identical mutations. A common, most likely European, ancestry, and a relative high carrier frequency of these mutations in Finland were discovered; suggesting that LBSL may be a quite common leukoencephalopathy in other populations as well. The results suggest that MRI findings are so unique that the diagnosis of LBSL is possible to make without genetic studies. This thesis work has resulted in identification of new mitochondrial disorders in Finland, enhancing the understanding of the clinical variability and the importance of tissue-specificity of these disorders. In addition to providing specific diagnosis to the patients, these findings give light to the underlying pathogenetic mechanisms of childhood-onset mitochondrial disorders.
  • Päiväniemi, Outi (2011)
    Chronic rejection in the form of obliterative bronchiolitis (OB) is the major cause of death 5 years after lung transplantation. The exact mechanism of OB remains unclear. This study focused on the role of cyclo-oxygenase (COX) -2, tenascin, and C-reactive protein (CRP) expression, and the occurrence of ingraft chimerism (= cells from two genetically distinct individuals in a same individual) in post-transplant OB development. In our porcine model, OB developed invariably in allografts, while autografts stayed patent. The histological changes were similar to those seen in human OB. In order to delay or prevent obliteration, animals were medicated according to certain protocol. In the beginning of the bronchial allograft reaction, COX-2 induction occurred in airway epithelial cells prior to luminal obliteration. COX-2 expression in macrophages and fibroblasts paralleled the onset of inflammation and fibroblast proliferation. This study demonstrated for the first time, that COX-2 expression is associated with the early stage of post- transplant obliterative airway disease. Tenascin expression in the respiratory epithelium appeared to be predictive of histologic features observed in human OB, and influx of immune cells. Expression in the bronchial wall and in the early obliterative lesions coincided with the onset of onset of fibroblast and inflammatory cell proliferation in the early stage of OB and was predictive of further influx of inflammatory and immune cells. CRP expression in the bronchial wall coincided with the remodelling process. High grade of bronchial wall CRP staining intensity predicted inflammation, accelerated fibroproliferation, and luminal obliteration, which are all features of OB. In the early obliterative plaque, majority of cells expressed CRP, but in mature, collagen-rich plaque, expression declined. Local CRP expression might be a response to inflammation and it might promote the development of OB. Early appearance of chimeric (= recipient-derived) cells in the graft airway epithelium predicted epithelial cell injury and obliteration of the bronchial lumen, which both are features of OB. Chimeric cells appeared in the airway epithelium after repair following transplantation-induced ischemic injury. Ingraft chimerism might be a mechanism to repair alloimmune-mediated tissue injury and to protect allografts from rejection after transplantation. The results of this study indicate, that COX-2, tenascin, CRP, and ingraft chimerism have a role in OB development. These findings increase the understanding of the mechanisms of OB, which may be beneficial in further development of diagnostic options.
  • Niinistö, Sari (Unigrafia, 2014)
    The aim of the present study was to evaluate the associations between the maternal intakes of fatty acids, vitamin D, and their respective dietary sources during pregnancy or lactation, as well as the child s fatty acid status, and the risk of type 1 diabetes. The present study is a prospective cohort belonging to the nutrition part of the Finnish Type 1 Diabetes Prediction and Prevention Study (DIPP). The cohort comprised children at increased genetic risk for type 1 diabetes (n=6069) born at the Oulu and Tampere University Hospitals between the years 1997 and 2004. Maternal diet was assessed with a validated food frequency questionnaire during the 8th month of pregnancy and the 3rd month of lactation. The children were monitored for the appearance of autoantibodies associated with type 1 diabetes and for the development of clinical type 1 diabetes, at intervals between 3 and 12 months. The associations between the maternal intakes of fatty acids, vitamin D, as well as their respective food sources and the endpoints, were analyzed by the piecewise linear log-hazard survival model and Cox proportional hazard analyses. The associations between the children s serum fatty acid composition and the risk of type 1 diabetes were studied in a nested case-control study design that included 108 children with preclinical type 1 diabetes and 216 matched controls. The total serum fatty acid composition was analyzed by gas chromatography from samples that were taken from the children between the ages of 1 and 6 years. Conditional logistic regression was applied in the statistical analysis. After adjustment for putative confounders, neither the maternal intake of fatty acids during pregnancy and lactation nor the intake of vitamin D during pregnancy, showed any association with the development of type 1 diabetes. The maternal use of fresh milk and cheese, and the intake of protein from fresh milk during pregnancy, were associated with a lower risk of clinical type 1 diabetes. The maternal total consumption of red meat and meat products, particularly processed meat, during lactation was associated with an increased risk of type 1 diabetes. The child s serum profile of milk-associated fatty acids at, or closest to, the time of seroconversion, was associated with a higher risk of preclinical type 1 diabetes. Findings give one of the first indications that prenatal exposure to certain dairy foods may affect immune regulation, and our results on the milk-associated fatty acid biomarkers confirm the previous findings that the child s own direct cow s milk exposure is associated with autoimmunity in children with increased genetic risk of type 1 diabetes. These results thus indicate that milk and meat products in particular, are interesting topics for future studies on potential nutritional risk and protective factors for type 1 diabetes.
  • Saarinen, Antti (2009)
    Data on the influence of unilateral vocal fold paralysis on breathing, especially other than information obtained by spirometry, are relatively scarce. Even less is known about the effect of its treatment by vocal fold medialization. Consequently, there was a need to study the issue by combining multiple instruments capable of assessing airflow dynamics and voice. This need was emphasized by a recently developed medialization technique, autologous fascia injection; its effects on breathing have not previously been investigated. A cohort of ten patients with unilateral vocal fold paralysis was studied before and after autologous fascia injection by using flow-volume spirometry, body plethysmography and acoustic analysis of breathing and voice. Preoperative results were compared with those of ten healthy controls. A second cohort of 11 subjects with unilateral vocal fold paralysis was studied pre- and postoperatively by using flow-volume spirometry, impulse oscillometry, acoustic analysis of voice, voice handicap index and subjective assessment of dyspnoea. Preoperative peak inspiratory flow and specific airway conductance were significantly lower and airway resistance was significantly higher in the patients than in the healthy controls (78% vs. 107%, 73% vs. 116% and 182% vs. 125% of predicted; p = 0.004, p = 0.004 and p = 0.026, respectively). Patients had a higher root mean square of spectral power of tracheal sounds than controls, and three of them had wheezes as opposed to no wheezing in healthy subjects. Autologous fascia injection significantly improved acoustic parameters of the voice in both cohorts and voice handicap index in the latter cohort, indicating that this procedure successfully improved voice in unilateral vocal fold paralysis. Peak inspiratory flow decreased significantly as a consequence of this procedure (from 4.54 ± 1.68 l to 4.21 ± 1.26 l, p = 0.03, in pooled data of both cohorts), but no change occurred in the other variables of flow-volume spirometry, body-plethysmography and impulse oscillometry. Eight of the ten patients studied by acoustic analysis of breathing had wheezes after vocal fold medialization compared with only three patients before the procedure, and the numbers of wheezes per recorded inspirium and expirium increased significantly (from 0.02 to 0.42 and from 0.03 to 0.36; p = 0.028 and p = 0.043, respectively). In conclusion, unilateral vocal fold paralysis was observed to disturb forced breathing and also to cause some signs of disturbed tidal breathing. Findings of flow volume spirometry were consistent with variable extra-thoracic obstruction. Vocal fold medialization by autologous fascia injection improved the quality of the voice in patients with unilateral vocal fold paralysis, but also decreased peak inspiratory flow and induced wheezing during tidal breathing. However, these airflow changes did not appear to cause significant symptoms in patients.
  • Grimaldi-Toriz, Sharon (Helsingin yliopisto, 2009)
    The aim of this study was to measure seasonal variation in mood and behaviour. The dual vulnerability and latitude effect hypothesis, the risk of increased appetite, weight and other seasonal symptoms to develop metabolic syndrome, and perception of low illumination in quality of life and mental well-being were assessed. These variations are prevalent in persons who live in high latitudes and need balancing of metabolic processes to adapt to environmental changes due to seasons. A randomized sample of 8028 adults aged 30 and over (55% women) participated in an epidemiological health examination study, The Health 2000, applying the probability proportional to population size method for a range of socio-demographic characteristics. They were present in a face-to-face interview at home and health status examination. The questionnaires included the modified versions of the Seasonal Pattern Assessment Questionnaire (SPAQ) and Beck Depression Inventory (BDI), the Health Related Quality of Life (HRQoL) instrument 15D, and the General Health Questionnaire (GHQ). The structured and computerized Munich Composite International Diagnostic Interview (M-CIDI) as part of the interview was used to assess diagnoses of mental disorders, and, the National Cholesterol Education Program Adult Treatment Panel III (NCEP-ATPIII) criteria were assessed using all the available information to detect metabolic syndrome. A key finding was that 85% of this nationwide representative sample had seasonal variation in mood and behaviour. Approximately 9% of the study population presented combined seasonal and depressive symptoms with a significant association between their scores, and 2.6% had symptoms that corresponded to Seasonal Affective Disorder (SAD) in severity. Seasonal variations in weight and appetite are two important components that increase the risk of metabolic syndrome. Other factors such as waist circumference and major depressive disorder contributed to the metabolic syndrome as well. Persons reported of having seasonal symptoms were associated with a poorer quality of life and compromised mental well-being, especially if indoors illumination at home and/or at work was experienced as being low. Seasonal and circadian misalignments are suggested to associate with metabolic disorders, and could be remarked if individuals perceive low illumination levels at home and/or at work that affect the health-related quality of life and mental well-being. Keywords: depression, health-related quality of life, illumination, latitude, mental well-being, metabolic syndrome, seasonal variation, winter.
  • 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.
  • Ekholm, Marja (Helsinki University Printing House, 2006)
    The purpose of this series of studies was to evaluate the biocompatibility of poly (ortho) ester (POE), copolymer of ε-caprolactone and D,L-lactide [P (ε-CL/DL-LA)] and the composite of P(ε-CL/DL-LA) and tricalciumphosphate (TCP) as bone filling material in bone defects. Tissue reactions and resorption times of two solid POE-implants (POE 140 and POE 46) with different methods of sterilization (gamma- and ethylene oxide sterilization), P(ε-CL/DL-LA)(40/60 w/w) in paste form and 50/50 w/w composite of 40/60 w/w P(ε-CL/DL-LA) and TCP and 27/73 w/w composite of 60/40 w/w P(ε-CL/DL-LA) and TCP were examined in experimental animals. The follow-up times were from one week to 52 weeks. The bone samples were evaluated histologically and the soft tissue samples histologically, immunohistochemically and electronmicroscopically. The results showed that the resorption time of gamma sterilized POE 140 was eight weeks and ethylene oxide sterilized POE 140 13 weeks in bone. The resorption time of POE 46 was more than 24 weeks. The gamma sterilized rods started to erode from the surface faster than ethylene oxide sterilized rods for both POEs. Inflammation in bone was from slight to moderate with POE 140 and moderate with POE 46. No highly fluorescent layer of tenascin or fibronectin was found in the soft tissue. Bone healing at the sites of implantation was slower than at control sites with the copolymer in small bone defects. The resorption time for the copolymer was over one year. Inflammation in bone was mostly moderate. Bone healing at the sites of implantation was also slower than at the control sites with the composite in small and large mandibular bone defects. Bone formation had ceased at both sites by the end of follow-up in large mandibular bone defects. The ultrastructure of the connective tissue was normal during the period of observation. It can be concluded that the method of sterilization influenced the resorption time of both POEs. Gamma sterilized POE 140 could have been suitable material for filling small bone defects, whereas the degradation times of solid EO-sterilized POE 140 and POE 46 were too slow to be considered as bone filling material. Solid material is difficult to contour, which can be considered as a disadvantage. The composites were excellent to handle, but the degradation time of the polymer and the composites were too slow. Therefore, the copolymer and the composite can not be recommended as bone filling material.
  • Sammalkorpi, Heli (2008)
    Colorectal cancer (CRC) is the third most common cancer in Finland. Of all CRC tumors, 15% display microsatellite-instability (MSI) caused by defective cellular mismatch repair. Cells displaying MSI accumulate a high number of mutations genome-wide, especially in short repeat areas, microsatellites. When targeting genes essential for cell growth or death, MSI can promote tumorigenesis. In non-coding areas, microsatellite mutations are generally considered as passenger events. Since the discovery of MSI and its linkage to cancer, more that 200 genes have been investigated for a role in MSI tumorigenesis. Although various criteria have been suggested for MSI target gene identification, the challenge has been to distinguish driver mutations from passenger mutations. This study aimed to clarify these key issues in the research field of MSI cancer. Prior to this, background mutation rate in MSI cancer has not been studied in a large-scale. We investigated the background mutation rate in MSI CRC by analyzing the spectrum of microsatellite mutations in non-coding areas. First, semenogelin I was studied for a possible role in MSI carcinogenesis. The intronic T9 repeat of semenogelin I was frequently mutated but no evidence for selection during tumorigenesis was obtained. Second, a sequencing approach was utilized to evaluate the general background mutation rate in MSI CRC. Both intronic and intergenic repeats harbored extremely high mutation rates of ≤ 87% and intergenic repeats were more unstable than the intronic repeats. As mutation rates of presumably neutral microsatellites can be high in MSI CRC in the absence of apparent selection pressure, high mutation frequency alone is not sufficient evidence for identification of driver MSI target genes. Next, an unbiased approach was designed to identify the mutatome of MSI CRC. By combining expression array data and a database search we identified novel genes possibly related to MSI CRC carcinogenesis. One of the genes was studied further. In the functional analysis this gene was observed to cause an abnormal cancer-prone cellular phenotype, possibly through altered responses to DNA damage. In our recent study, smooth muscle myosin heavy chain 11 (MYH11) was identified as a novel MSI CRC gene. Additionally, MYH11 has a well established role in acute myeloid leukemia (AML) through an oncogenic fusion protein CBFB-MYH11. We investigated further the role of MYH11 in AML by sequencing. Three novel missense variants of MYH11 were identified. None of the variants were present in the population-based control material. One of the identified variants, V71A, lies in the N-terminal SH3-like domain of MYH11 of unknown function. The other two variants, K1059E and R1792Q are located in the coil-coiled myosin rod essential for the regulation and filament formation of MYH11. The variant K1059E lies in the close proximity of the K1044N that has been functionally assessed in our earlier work of CRC and has been reported to cause total loss of MYH11 protein regulation. As the functional significance of the three novel variants examined in this work remains unknown, future studies should clarify the further role of MYH11 in AML leukaemogenesis and in other malignancies.
  • Nevalainen, Maarit (Unigrafia, 2014)
    The aim of this study was to investigate medical students and primary care physicians feelings of uncertainty, experiences of medical errors and ways of coping with them, and attitudes towards the career of a GP. In Study I the learning diaries (n=79) and writings on specific themes (n=94) of 22 students during their first clinical year were analysed by thematic content analysis. The topics uncertainty and medical errors were studied in more detail. Studies II and III are based on the responses of 5th-year students to a cross-sectional survey (Study II, n=307 and Study III, n=309, response rate 86%) during their main course in general practice. Study IV concerned younger (n=85) and experienced physicians (n=80) responses to an electronic survey related to uncertainty and experiences of medical errors (response rate 68%). Factors predisposing physicians to medical errors and means to avoid making them were assessed. In the second, third and fourth studies the responses were cross-tabulated. The variables are presented as means with standard deviations and percentages. Comparisons of categorial variables were carried out by using the X2 test or Fischer s exact test, comparisons of non-normally distributed continuous variables were made by using the Mann-Whitney U-test. (P values less than 0.05 were considered significant.) 95% CIs are presented for the most important results. Logistic regression analysis was used to explore which factors predicted good tolerance of uncertainty among GPs. A developmental path among the 3rd- to 4th-year medical students was discovered regarding their experiences of uncertainty and fears medical errors. All students wrote about several aspects of uncertainty. They also feared mistakes. However, they started gradually to be more confident in coping with responsibility. The 5th-year medical students were divided into two groups based on their tolerance of uncertainty and fear of medical errors. Those who tolerated uncertainty quite well or well (n=240, 78%) were older, more often males, and had been working for a longer time as locum doctors than those tolerating uncertainty poorly (n=67, 22%). In the latter group, 100% were afraid of medical errors, in the former group the figure was 86%. Those tolerating uncertainty poorly felt more often that a GP s work is too challenging and difficult and involves too much responsibility. About 3/4 of the fifth-year medical students considered the work of a GP positively versatile and challenging, but about 2/3 of them considered it to be too hasty and pressing. The females thought that a GP s work is too lonely. The males considered that a GP deals too much with non-medical problems and the work may be too routine and tedious. According to the majority of the students (82%) the most important aim of a GP s work is to identify serious diseases and refer such patients to specialized care. In the fourth study 6% of the younger and 1% of the experienced physicians tolerated uncertainty poorly. The younger physicians feared medical errors more (70% vs. 48%) and more often admitted having made an error (84% vs. 69%) compared to the experienced ones (48%). The experienced physicians apologized more willingly to their patients for an error. The younger physicians were more prone to use electronic databases and consult on site. Tolerance of uncertainty seems to develop gradually during the medical studies and this development also continues after graduation. Students are already aware of the possibility of a medical error. Almost all of them are afraid of them. Also half of the experienced physicians admitted to feeling some fear. The attitudes of fifth-year medical students reflect their experiences of general practice.
  • Söderlund, Sanni (2011)
    AIMS An independent, powerful coronary heart disease (CHD) predictor is a low level of high-density lipoprotein cholesterol (HDL-C). Discoidal preβ-HDL particles and large HDL2 particles are the primary cholesterol acceptors in reverse cholesterol transport, a key anti-atherogenic HDL mechanism. The quality of HDL subspecies may provide better markers of HDL functionality than does HDL-C alone. We aimed I) to study whether alterations in the HDL subspecies profile exist in low-HDL-C subjects II) to explore the relationship of any changes in HDL subspecies profile in relation to atherosclerosis and metabolic syndrome; III) to elucidate the impact of genetics and acquired obesity on HDL subspecies distribution. SUBJECTS The study consisted of 3 cohorts: A) Finnish families with low HDL-C and premature CHD (Study I: 67 subjects with familial low HDL-C and 64 controls; Study II: 83 subjects with familial low HDL-C, 65 family members with normal HDL-C, and 133 controls); B) a cohort of 113 low- and 133 high-HDL-C subjects from the Health 2000 Health Examination Survey carried out in Finland (Study III); and C) a Finnish cohort of healthy young adult twins (52 monozygotic and 89 dizygotic pairs) (Study IV). RESULTS AND CONCLUSIONS The subjects with familial low HDL-C had a lower preβ-HDL concentration than did controls, and the low-HDL-C subjects displayed a dramatic reduction (50-70%) in the proportion of large HDL2b particles. The subjects with familial low HDL-C had increased carotid atherosclerosis measured as intima-media-thickness (IMT), and HDL2b particles correlated negatively with IMT. The reduction in both key cholesterol acceptors, preβ-HDL and HDL2 particles, supports the concept of impaired reverse cholesterol transport contributing to the higher CHD risk in low-HDL-C subjects. The family members with normal HDL-C and the young adult twins with acquired obesity showed a reduction in large HDL2 particles and an increase in small HDL3 particles, which may be the first changes leading to the lowering of HDL-C. The low-HDL-C subjects had a higher serum apolipoprotein E (apoE) concentration, which correlated positively with the metabolic syndrome components (waist circumference, TG, and glucose), highlighting the need for a better understanding of apoE metabolism in human atherosclerosis. In the twin study, the increase in small HDL3b particles was associated with obesity independent of genetic effects. The heritability estimate, of 73% for HDL-C and 46 to 63% for HDL subspecies, however, demonstrated a strong genetic influence. These results suggest that the relationship between obesity and lipoproteins depends on different elements in each subject. Finally, instead of merely elevating HDL-C, large HDL2 particles and discoidal preβ-HDL particles may provide beneficial targets for HDL-targeted therapy.
  • Vlachopoulou, Efthymia (2015)
    The Human Leukocyte Antigen (HLA) region is located on chromosome 6 (6p21.3) and its main function is to regulate the immune system. This region has been associated with several autoimmune and other inflammatory conditions. However, many aspects of these associations remain unknown due to high polymorphism, high linkage disequilibrium (LD) and high gene density of this region that increase the complexity and create computational challenges. In this Thesis, the aim is to investigate the HLA haplotype variation in the Finnish population, to evaluate the HLA imputation using two of the existed softwares and to analyze the HLA association with Acute Coronary Syndrome (ACS). The main question in this study is whether the HLA region varies in the Finnish population compared to other populations and how this variation can influence the results of further analyses. The HLA haplotypes showed great variation in the Finnish population compared to other European populations due to different linkage disequilibrium (LD) structures and this highlights the diversity of the HLA haplotypes among different populations. The most common European haplotype HLA-A*01~HLA-B*08~HLA-DRB1*03 shows a lower frequency in Finland, whereas the enrichment of the haplotype HLA-A*03~HLA-B*35~HLA-DRB1*01 was observed (I). Furthermore, the poor imputation results of the HLA-DRB1 gene enhance the power of the previously mentioned arguments (from I) due to the inaccurate imputation from other European populations. The study emphasizes the need for population-specific reference materials for a successful imputation (II). The diversity of HLA alleles, the variability in the frequencies of Single Nucleotide Polymorphisms (SNP) among populations, the difference of the rare HLA alleles appearing in various populations and the extremely high amount of combinations of HLA alleles (due to high polymorphism) contribute to the low success rates of imputation. The association (OR = 4.43, 95% CI = 3.57-5.50) of a novel BTNL2;HLA-DRA;HLA-DRB1*01-haplotype with the risk of Acute Coronary Syndrome (ACS) was identified (III). The analysis showed the implication of the HLA-DRB1*01 allele and the BTNL2 gene to the inflammation process. These two genes affect the antigen presentation and its immune response. Overall, the associated haplotype appeared approximately in 15% of ACS patients and in 5% of healthy individuals. Furthermore, the survival analysis shows that the HLA-DRB1*01 allele does not affect the survival time after an ACS event. Our results provide more insight into the detailed patterns of the haplotypes in the Finnish population, into the success of the imputation approaches and into the HLA association with ACS. Genetic diversity is depicted on these haplotypes which might lead to different immune responses. These findings help future development of appropriate statistical tools for the HLA region.
  • Anttila, Timo Verneri (Helsingin yliopisto, 2010)
    Migraine is the common cause of chronic episodic headache, affecting 12%-15% of the Caucasian population (41 million Europeans and some half a million Finns), and causes considerable loss of quality of life to its sufferers, as well as being linked to increased risk for a wide range of conditions, from depression to stroke. Migraine is the 19th most severe disease in terms of disability-adjusted life years, and 9th among women. It is characterized by attacks of headache accompanied by sensitivity to external stimuli lasting 4-72 hours, and in a third of cases by neurological aura symptoms, such as loss of vision, speech or muscle function. The underlying pathophysiology, including what triggers migraine attacks and why they occur in the first place, is largely unknown. The aim of this study was to identify genetic factors associated with the hereditary susceptibility to migraine, in order to gain a better understanding of migraine mechanisms. In this thesis, we report the results of genetic linkage and association analyses on a Finnish migraine patient collection as well as migraineurs from Australia, Denmark, Germany, Iceland and the Netherlands. Altogether we studied genetic information of nearly 7,000 migraine patients and over 50,000 population-matched controls. We also developed a new migraine analysis method called the trait component analysis, which is based on individual patient responses instead of the clinical diagnosis. Using this method, we detected a number of new genetic loci for migraine, including on chromosome 17p13 (HLOD 4.65) and 10q22-q23 (female-specific HLOD 7.68) with significant evidence of linkage, along with five other loci (2p12, 8q12, 4q28-q31, 18q12-q22, and Xp22) detected with suggestive evidence of linkage. The 10q22-q23 locus was the first genetic finding in migraine to show linkage to the same locus and markers in multiple populations, with consistent detection in six different scans. Traditionally, ion channels have been thought to play a role in migraine susceptibility, but we were able to exclude any significant role for common variants in a candidate gene study of 155 ion transport genes. This was followed up by the first genome-wide association study in migraine, conducted on 2,748 migraine patients and 10,747 matched controls followed by a replication in 3,209 patients and 40,062 controls. In this study, we found interesting results with genome-wide significance, providing targets for future genetic and functional studies. Overall, we found several promising genetic loci for migraine providing a promising base for future studies in migraine.
  • Talling, Maria Louise (2009)
    Juvenile neuronal ceroid lipofuscinosis (JNCL) is one of the most common neurodegenerative diseases in childhood. Its clinical onset, with visual failure as the first sign, is between the ages of 4 to 8 years. During the disease progress, epilepsy, motor symptoms, cognitive decline, and psychiatric symptoms become apparent. It leads to premature death between ages 15 and 30. Treatment consists of symptomatic drug administration and various forms of rehabilitation, but to date, no curative treatment exists. To gain a more comprehensive picture of psychiatric problems, symptoms were evaluated by the Child Behavior Checklist, the Teacher Report Form, and the Children s Depression Inventory. The JNCL patients had a great number of severe psychiatric symptoms, with wide inter-individual variability. The most common symptoms were social, thought, attention, and sleep problems, somatic complaints, and aggressive behaviour. Patients with psychotropic treatment had more problems than did those without psychotropic treatment, and female patients had more problems than did males. Between 10 and 20% of the patients reported depressive symptoms. In a 5-year follow-up, [123I]β-CIT SPECT and MRI revealed a tendency of decreasing serotonin transporter (SERT) availability and progressive brain atrophy. The correlation between changes in midbrain SERT and total brain volume was positive; no correlation appeared between SERT or brain atrophy and depressive symptoms. Thus, it seems likely that the low SERT availability is associated with progressive brain atrophy; it may also predispose towards depression, however. An open survey of psychotropic drugs and their efficacy was performed on JNCL patients in Finland. The most commonly used psychotropic drugs were the antidepressant citalopram and the antipsychotic risperidone. Their efficacy was good or satisfactory in the majority of cases and they seemed well tolerated. Quetiapine had a marked effect on one patient with a history of severe psychotic symptoms. Glutamate decarboxylase 65 autoantibodies (GAD65ab), found in JNCL patients, indicate that an immunomediated reaction against GAD or GABAergic neurons may play a part in the underlying pathogenetic mechanism. GAD65ab s also appeared in the serum of all eight JNCL patients included and intermittent corticosteroid therapy was initiated in all cases. After one year, the GAD65ab s had disappeared in the two oldest patients, who experienced an improvement in motor symptoms and alertness associated with their prednisolone therapy. Two younger patients experienced a significant IQ increase, but no change in GADab s. A randomized study with longer follow-up time is needed, however, to clarify the effect of prednisolone on disease progression.
  • Salminen, Minna (Yliopistopaino, 2006)
    Lactobacilli are gram positive rods, which belong to normal oropharyngeal, gastrointestinal and urogenital flora. They are widely used in food industry and as food additives. Although their virulence is presumed to be very low, opportunistic bacteremic infections, especially in immunocompromised hosts, have been detected. In the present study, the possible effects of increasing probiotic use of Lactobacillus rhamnosus GG (LGG) on the occurrence of bacteremia due to lactobacilli was evaluated on population level. In Finland, 90 Lactobacillus bacteremia cases were reported to the National Infectious Disease Register maintained by National Public Health Institute, during 1995-2000. Their proportion of all bacteremia cases was on average 0.24%, corresponding to 0.3 cases/100 000 inhabitants annually. In the Helsinki University Central Hospital district the corresponding proportion of all bacteremia cases was observed during 1990-2000. Despite LGG intake increased six folded no increasing trend in the occurrence of lactobacilli bacteremia was seen. A total of 85 Lactobacillus sp. blood isolates collected from different human bacteremic cases were characterised and compared with the commercial probiotic LGG strain. In species characterisation 46 L. rhamnosus strains, 12 L. fermentum and L. casei strains each, three each of L. gasseri, L. salivarius and L. jensenii species, two L. curvatus, and one each of L. plantarum, L. sakei, L. zeae and L. reuteri species were detected. Nearly half of the L. rhamnosus findings turned out to be indistinguishable from the probiotic LGG strain. Common predisposing factors to Lactobacillus bacteremia were immunosuppression, prior prolonged hospitalisation and prior surgical interventions. Severe or fatal comorbidities were found in 82% of the patients. Mortality at one month was 26% and severe underlying diseases were a significant predictor of death (OR 15.8). Antimicrobial susceptibility of Lactobacillus strains was species dependent. The Lactobacillus isolates were generally susceptible to imipenem, piperacillin-tazobactam, clindamycin and erythromycin, whereas all other than L. gasseri and L. jensenii species were not at all susceptible to vancomycin. The susceptibility to cephalosporin varied greatly even within species why they might not be recommended for treatment of Lactobacillus infections. The effect and safety of probiotic LGG preparation in amelioration of gastric symptoms and diarrhea in HIV-infected patients was evaluated. No significant differences in gastrointestinal symptoms or diarrhoea in LGG treated patients compared to placebo could be found. LGG was well tolerated with no adverse effects including bacteremic outbreaks could be observed. The use of probiotic LGG can be regarded safe in this immunocompromised patient group.
  • Leskelä, Ulla (2008)
    The study is part of a research project of 269 psychiatric patients with major depression, Vantaa Depression Study, in the Department of Mental Health and Alcohol Research of the National Public Health Institute and the Department of Psychiatry of the Peijas Medical Care District. The aim was to study at the onset of MDE psychosocial differences in subgroups of patients and clustering of events into time before depression and its prodromal phase, to study whether more severe life events and less social support predict poorer outcome in all patients, but most among those currently in partial remission, whether social support declines as a consequence of time spent in MDE, is sensitive to improvement, and whether social support is influenced by neuroticism and extraversion. After screening, a semistructured interview (SCAN, version 2.0) was used for the presence of DSM-IV MDE, and other psychiatric diagnoses. Life events and social support were studied with semistructured methods (IRLE, Paykel 1983; IMSR, Brugha et al. 1987), perceived social support and neuroticism/extraversion with questionnaires (PSSS-R, Blumenthal et al. 1987; EPI, Eysenck and Eysenck 1964) at baseline, 6 and 18 months. At the onset of depression life events were common. No major differences between subgroups of patients were found; the younger had more events, whereas those with comorbid alcoholism and personality disorders perceived less support. Although events were distributed evenly between the time before depression, the prodromal phase and the index MDE, two thirds of the patients attributed their depression to some life event. Adversities and poor perceived support influenced the outcome of all psychiatric patients, most in the subgroup of full remission. In the partial remission group, the impact of severe events and in the MDE, perceived support was important. Low objective and subjective support were predicted by longer time spent in MDE. Along with improvement subjective support improved. Neuroticism and extraversion were associated with the size of social network and perceived support and predicted change of perceived support. In conclusion, adversities were common in all phases of depression. They may thus have many roles; before depression they may precipitate it, in the prodromal phase worsen symptoms, and during the MDE, the outcome of depression. Patients often attributed their depression to a life event. Psychosocial subgroup differences were quite small. Perceived support predicted the outcome of depression, and time spent in MDE objective and subjective support. Neuroticism and extraversion may modify the level and change particularly in perceived support, thereby indirectly effecting vulnerability to depression.
  • Raevuori, Anu (2009)
    The aim of this study was to deepen the understanding of eating disorders, body image dissatisfaction and related traits in males by examining the epidemiology and genetic epidemiology of these conditions in representative population-based twin samples. The sample of Study I included adolescent twins from FinnTwin12 cohorts born 1983 87 and assessed by a questionnaire at ages 14 y (N=2070 boys, N=2062 girls) and 17 y (N=1857 boys, N=1984 girls). Samples of Studies II-V consisted of young adult twins born 1974-79 from FinnTwin16 cohorts (Study II N=1245 men, Study III N=724 men, Study IV N=2122 men, Study V N=2426 women and N=1962 men), who were assessed by a questionnaire at the age 22-28 y. In addition, 49 men and 526 women were assessed by a diagnostic interview. The overall response rates for both twin cohorts in all studies were 80-90%. In boys, mainly genetic factors (82%, 95% confidence interval [CI] 72-92) explained the covariation of self-esteem between the ages 14 y and 17 y, whereas in girls, environmental factors (69%, 95% CI 43-93) were the largest contributors. Of young men, 30% experienced high muscle dissatisfaction, while 12% used or had used muscle building supplements and/or anabolic steroids on a regular basis. Muscle dissatisfaction exhibited a robust association with the indicators of mental distress and a genetic component (42%, 95% CI 23-59) for its liability in this population was found. The variation of muscle-building substance use was primarily explained by the environmental factors. The incidence rate of anorexia nervosa in males for the age of 10-24 y was 15.7 (95% CI 6.6-37.8) per 100 000 person-years, and its lifetime prevalence by the young adulthood was 0.24% (95% CI 0.03-0.44). All detected probands with anorexia nervosa had recovered from eating disorders, but suffered from substantial psychiatric comorbidity, which manifested also in their co-twins. Additionally, male co-twins of the probands displayed significant dissatisfaction with body musculature, a male-specific feature of body dysmorphic disorder. All probands were from twin pairs discordant for eating disorders. Of the five male probands with anorexia nervosa, only one was from an opposite-sex twin pair. Among women from the opposite-sex pairs, the prevalence of DSM-IV or broad anorexia nervosa was no significantly different compared to that of the women from monozygotic pairs or from dizygotic same-sex pairs. The prevalence of DSM-IV or broad bulimia nervosa did not differ in opposite- versus same-sex female twin individuals either. In both sexes, the overall profile of indicators on eating disorders was rather similar between individuals from opposite-sex and same-sex pairs. In adolescence, development of self-esteem was differently regulated in boys compared to girls: this finding may have far-reaching implications on the etiology of sex discrepancy of internalizing and externalizing disorders. In young men, muscle dissatisfaction and muscle building supplement/steroid use were relatively common. Muscle dissatisfaction was associated with marked psychological distress such as symptoms of depression and disordered eating. Both genetic and environmental factors explained muscle dissatisfaction in the population, but environmental factors appeared to best explain the use of muscle-building substances. In this study, anorexia nervosa in boys and young men from the general population was more common, transient and accompanied by more substantial co-morbidity than previously thought. Co-twins of the probands with anorexia nervosa displayed significant psychopathology such as male specific symptoms of body dysmorphic disorder, but none of them had had an eating disorder: taken together, these traits are suggestive for an endophenotype of anorexia nervosa in males. Little evidence was found on that the risk for anorexia nervosa, bulimia nervosa, disordered eating or body dissatisfaction were associated with twin zygosity. Thus, it is unlikely that in utero femininization, masculinization or postnatal socialization according to the sex of the co-twin have a major influence on the later development of eating disorders or related traits.
  • Tyynismaa, Henna (Yliopistopaino, 2007)
    Mitochondrial diseases are caused by disturbances of the energy metabolism. The disorders range from severe childhood neurological diseases to muscle diseases of adults. Recently, mitochondrial dysfunction has also been found in Parkinson s disease, diabetes, certain types of cancer and premature aging. Mitochondria are the power plants of the cell but they also participate in the regulation of cell growth, signaling and cell death. Mitochondria have their own genetic material, mtDNA, which contains the genetic instructions for cellular respiration. Single cell may host thousands of mitochondria and several mtDNA molecules may reside inside single mitochondrion. All proteins needed for mtDNA maintenance are, however, encoded by the nuclear genome, and therefore, mutations of the corresponding genes can also cause mitochondrial disease. We have here studied the function of mitochondrial helicase Twinkle. Our research group has previously identified nuclear Twinkle gene mutations underlying an inherited adult-onset disorder, progressive external ophthalmoplegia (PEO). Characteristic for the PEO disease is the accumulation of multiple mtDNA deletions in tissues such as the muscle and brain. In this study, we have shown that Twinkle helicase is essential for mtDNA maintenance and that it is capable of regulating mtDNA copy number. Our results support the role of Twinkle as the mtDNA replication helicase. No cure is available for mitochondrial disease. Good disease models are needed for studies of the cause of disease and its progression and for treatment trials. Such disease model, which replicates the key features of the PEO disease, has been generated in this study. The model allows for careful inspection of how Twinkle mutations lead to mtDNA deletions and further causes the PEO disease. This model will be utilized in a range of studies addressing the delay of the disease onset and progression and in subsequent treatment trials. In conclusion, in this thesis fundamental knowledge of the function of the mitochondrial helicase Twinkle was gained. In addition, the first model for adult-onset mitochondrial disease was generated.