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  • Joensuu, Emmi (Helsingin yliopisto, 2015)
    Cancer usually arises through mutational changes in the genome but also epigenetic changes can contribute to tumorigenesis. In this research we studied both sporadically occurring and familial colorectal, endometrial and gastric tumors. Sporadic tumors were divided into separate categories depending on the microsatellite instability status of the tumor. In addition to sporadic tumors we studied tumors from patients with different cancer syndromes: Lynch syndrome, Familial colorectal cancer type X and Familial site-specific endometrial cancer. Lynch syndrome patients have a predisposing germline mutation in one of the mismatch repair genes (MLH1, MSH2 or MSH6) and the tumors are typically microsatellite unstable. Despite the extensive research efforts, the genetic or epigenetic background of the other studied syndromes is not known and remains to be molecularly characterized. We therefore explored the possible epigenetic basis of cancer susceptibility in these syndromes. First we studied the promoter methylation of 24 established tumor suppressor genes. Hypermethylation patterns were found to be characteristic of each tissue and diversely dependent on the microsatellite instability status of the tumor, or family category. The CpG island methylator phenotype (CIMP) in which multiple loci are silenced by promoter methylation, was most evident in sporadic microsatellite unstable tumors (P less than 0.001) and was present in 38% of all of the studied colorectal, 19% of endometrial and 29% of gastric tumors. In these tumors the CIMP phenotype can contribute to the genomic instability and the progression of cancer. In addition, despite being microsatellite stable, 50% of Familial colorectal cancer type X tumors displayed the CIMP phenotype. Our results of global hypomethylation confirm that tumors have significantly lower methylation levels compared to normal tissues in most of the studied patient groups (P less than 0.05) and that the hypomethylation levels depend significantly on the microsatellite instability status of the tumors (P = 0.042 for colorectal and P = 0.018 for gastric tumors). The significant decrease in the methylation levels, observed especially in the normal tissues of Familial colorectal cancer type X patients, could function as a premalignant field defect, where a large area of tissue is affected by carcinogenic alteration, and hence promote cancer development by facilitating the accumulation of other lesions such as genetic mutations or other epigenetic changes in the affected areas. After the characterization of different DNA methylation aberrations in distinct tumor categories, we studied the possible mechanisms behind the observed methylation changes. We evaluated the association of the expression of DNA methyltransferases DNMT1 and DNMT3B and histone methyltransferase EZH2 with CIMP+ phenotype and global hypomethylation patterns. Compared to the normal tissues, all the studied methyltransferases were significantly overexpressed in colorectal tumors (P less than 0.001) and DNMT3B also in endometrial tumors (P less than 0.001). EZH2 overexpression was shown to associate with CIMP+ phenotype especially in sporadic colorectal tumors and the finding was statistically significant (P = 0.003). The overall aim of this research was to elucidate epigenetic mechanisms in cancer, including cancers of different organs and also different familial cancers. Available information on the epigenetic events of cancers is increasing and although the topic is under continuous study, our understanding of it is still limited. New knowledge in the field can increase the understanding of the basic tumorigenic mechanisms and thereby facilitate more specific and earlier diagnosis and treatment of different types of cancer. Also the potential reversibility of epigenetic states offers interesting possibilities for drug development.
  • Ropponen, Jussi (Helsingin yliopisto, 2015)
    Chronic lung allograft dysfunction (CLAD) is the major life-limiting factor after lung transplantation and bronchiolitis obliterans syndrome (BOS) is the most common subtype and best-characterized form of CLAD. Pathologically, BOS presents as obliterative bronchiolitis (OB) and it is characterized by peribronchial inflammation, epithelial damage, and obliteration of small and medium-sized bronchioli by fibrotic plaques. BOS is the leading cause mortality after the first post-operative year. In this study, we hypothesized that inhibiting innate immune activation through different pathways influences the development of experimental OB. To test our hypothesis, we investigated different factors and pathways leading to experimental OB using both rat and mouse heterotopic tracheal allograft models. In the presence of alloantigens, early ischemic injury induced both innate and adaptive immune responses followed by Th17 activation and afterwards by a sustained Th1 immune response. This was accompanied by infiltration of the allograft with proinflammatory effector cells and lead to progressive fibroproliferation and total tracheal occlusion. Interestingly, recipient treatment with simvastatin, a cholesterol-lowering drug with lipid-independent immunomodulatory properties, enhanced early epithelial recovery after transplantation in the allografts. It also inhibited the infiltration of inflammatory cells and the expression of lymphocyte chemokines and proinflammatory cytokine mRNA. Most importantly, simvastatin inhibited the development of experimental OB in the absence of other immunosuppression. The cellular responses to hypoxia are regulated by transcription factors called hypoxia inducible factors (HIFs). HIF-1 is a principle regulator of hypoxic adaptation. In this study, we found that continuous HIF-1 expression in myeloid cells improved epithelial recovery, reduced inflammatory cell accumulation, and increased regulatory FoxP3 mRNA expression in mouse tracheal allografts. Importantly, these effects led to better preservation of tracheal epithelium and a decrease in the development of experimental OB suggesting a protective role of HIF-1 in this constellation.
  • Koivunen, Riku-Jaakko (Helsingin yliopisto, 2015)
    Intracerebral hemorrhage (ICH) is a devastating form of stroke. Its common risk factors include hypertension and smoking, but different underlying causes are numerous. Knowledge regarding clinical characteristics and outcome of young ICH patients, encompassing 10% of all ICH patients, is limited. The aims of this study were to define the risk factors, etiologic distribution, clinical picture, medical complications suffered, and prognosis of ICH at young age. We collected detailed clinical, radiological, mortality and follow-up data on all consecutive patients between 16 and 49 years of age with first-ever ICH treated at the Helsinki University Hospital (HUH) between 2000 and 2010. Results concerning the early course of ICH were compared to a series of ICH patients aged >49 years treated in HUH between 2005 and 2010. Median age was 42 years (interquartile range 34-47) and males comprised 59.5% of the 336 patients included. Annual incidence of ICH was 4.9 (95% confidence interval 4.5-5.3) per 100 000. The most prevalent risk factors were hypertension (29.8%) and smoking (22.3%). Compared to older ICH patients (n=921) hypertensive microangiopathy was less common (25.0% vs. 34.3%, P=0.002) and structural lesions more common (25.0% vs. 4.9%, P<0.001) assumed etiologies of ICH. The cause remained elusive in 32.1% of all young patients, and in 22.5% of those who underwent MRI and any angiography (n=89, P=0.023). Three-month mortality rate was lower among young patients compared to older ones, (17.0% vs. 32.7%, p<0.001). Hematoma volumes were similar across all ages (p=0.324) and it independently predicted mortality in older patients, but not in the young. More severe stroke initially, measured by the National Institutes of Health Stroke Scale (NIHSS) score, infratentorial hematoma location, hydrocephalus, herniation, and multiple hemorrhages associated with increased 3-month mortality. When adjusted for these factors as well as demographics, ICH volume, and the underlying cause, we found that surgical evacuation was associated with lower mortality (odds ratio 0.06; 95% confidence interval 0.02-0.21, P<0.001). In propensity-score matched analysis, case-fatality rates were three-fold in those treated conservatively (27.5% vs. 7.8%, P<0.001). The most common medical complications included hyperglycemia (51%), hyponatremia (45%), hypopotassemia (32%), and infections (28%). Hyperglycemia was the only single complication independently associated with increased mortality (5.90, 2.25-15.48, P<0.001). However, three or more concomitant complications also associated with increased mortality (7.76, 1.42-42.49, P=0.018). Among the 268 one-month survivors, 1-year survival was 98.1% (95% confidence interval 96.2-100%), 5-year survival 93.2% (89.3-97.1%), and 10-year survival 88.8% (84.9-92.7%), with male gender (3.36, 1.28-8.80, P=0.014) and diabetes (2.64, 1.01-6.89, P=0.047) being associated with mortality. Unfavorable outcome (modified Rankin Scale score 2-5) emerged in 49%, and was independently predicted by higher age (1.09 per one year, 1.03-1.15, P=0.002) stroke severity (1.17 per one NIHSS score point, 1.08-1.27, P<0.001), and intraventricular extension of hemorrhage (3.26, 1.11-9.55, P=0.031). PSD was present among one out of four survivors of ICH at young age. Since only one out of ten currently used antidepressants, treatment of depression appears as an unmet need in young ICH survivors. In summary, prevention and treatment of cardiovascular risk factors are vital in ICH prevention among young adults. Comprehensive diagnostic work-up and imaging are essential in identifying the underlying cause of ICH. The young seem to survive ICH better than the elderly, particularly if surgical hematoma evacuation is pursued. A holistic approach to prevent and treat associated complications, specifically hyperglycemia, is vital in regard of survival. Only half of the survivors reach favorable functional outcome. Therefore, more effective measures of rehabilitation and mental health must be developed to improve the quality of life of this patient population.
  • Hemminki, Otto (Helsingin yliopisto, 2015)
    In 2012 WHO announced cancer as the leading cause of death. Every day 20 000 people die due to cancer, and the rate is estimated to double before year 2030. While treatments have progressed, there are still few good treatment options for advanced cancer. Thus, there is an urgent need for new treatments. Immunotherapy with gene modified oncolytic adenoviruses provides novel promising means of treating cancer. These treatments incorporate two basic concepts. Firstly, adenoviruses are modified so that they replicate only in cancer cells, which makes the treatments safer. Secondly, the virus induced cancer cell oncolysis elicits a danger signal that awakens the immune system to fight the cancer. Viruses can be further armed with different genes that can stimulate the immune system even more. Most of these oncolytic viruses are based on adenovirus serotype 5, as indicated in thousands of publications. However, the primary receptor for serotype 5 is down-regulated in advanced cancer. On the contrary, adenovirus serotype 3 receptor is known to be abundant in advanced cancer making it an interesting subject of research. While a different serotype per see offers an alternative immune response, serotype 3 incorporates also other interesting features that might further potentiate its utility. Our first goal was to create serotype 3 based oncolytic adenoviruses for the treatment of human cancer. The goal was achieved, making this virus, to our knowledge, the first non- adenovirus 5 based oncolytic adenovirus in the world used in humans. The publications, study I and II, are now part of this thesis. The virus was designed to have a human telomerase reverse transcriptase (hTERT) promoter diverting the replication of the virus into cancer cells. This virus, Ad3-hTERT-E1A, was successfully cloned, rescued and produced in large scale, which was followed by rigorous preclinical testing of the virus. Rigorous preclinical testing of the virus followed. Several in vitro and in vivo experiments were performed, including sequencing, qPCR, electron microscopy and neutralizing antibody assays, while the most convincing data was gained from the cell cultures and the animal models. We found the serotype 3 effective in all major cancer types in vitro. In vivo, the serotype 3 virus was found at least as potent as serotype 5 based control viruses in several murine models of human cancer. Before clinical treatments, biodistribution and toxicity experiments were performed. In toxicity studies, adenovirus 3 was found less toxic than the serotype 5 based control viruses in an immune competent murine model. The histology of all major organs and basic blood values were analyzed. The preclinical data suggested strong efficacy with good safety. In study II, we publish the data of the first 25 patients treated with the Ad3-hTERT-E1A virus. All patients had advanced solid tumors refractory to standard therapies. Th e safety of the treatment was good with up to 4x10 12 virus particles given intravenously and/or intratumorally. Overall, all patients experienced mild (grade 1-2) self-limiting flu-like adverse events. No severe adverse events were noted attributable to the treatments. After treatment, many patients showed signs of efficacy. Of the 15 patients with elevated tumor markers before the treatment, 73% responded with a decrease or no change in the markers. Even a few complete responses were reported, while some patients also showed a clear decrease in the tumor mass according to imaging. Also the clinical data suggested strong efficacy with good safety, proposing a need for a randomized study. Our next goal was to evaluate better ways in finding treatment responders, as size based computed tomography (CT) is known to be suboptimal in evaluating immunotherapeutics where initial swelling of the tumor due to the immune response is common. In study III, we examined the ability of magnetic resonance imaging (MRI) and spectroscopy (MRS) in immunocompetent Syrian hamsters. T2 weighed MRI seemed encouraging in finding responding hamsters as soon as two days after treatment. Similar findings were noted with a patient responding to oncolytic treatments. MRS of taurine, choline and unsaturated fatty acids were found to be promising metabolites when evaluating responders after oncolytic immunotherapy. These results propose MRI and MRS as potential methods in evaluating responding patients. T2 weighed MRI is already widely used in the clinics, thus a clinical trial should be easy to implement. In study IV, we evaluated the first 16 patients treated with a quadruple modified oncolytic serotype 5 adenovirus. Th e fiber knob of this virus is from serotype 3, while the virus also produces an immunostimulatory GM-CSF molecule. Th e two other modifications restrict the replication to cancer cells. The safety profile of the virus resembled that of the oncolytic serotype 3 virus, and also numerous signs of effi cacy were noted. Immunological studies indicated activation of the immune system in responding patients. Rationale for a randomized study exists also for this virus.
  • He, Liang (Helsingin yliopisto, 2015)
    Statistical inference of genome-wide association studies (GWAS) on a variety of epidemiological phenotypes with special focus on rare variants (RVs) is challenging. In this thesis, new statistical models for detecting RV association have been developed under the hierarchical Bayesian framework. Special attention with respect to appropriate statistical inference has been given to the case of incorporation of sequencing error information, family-based models and detection of interaction effects in twin data. Estimation of the three new statistical models proposed in this thesis have been implemented using Markov chain Monte Carlo (MCMC) methods and their statistical properties have been evaluated through simulations. Their performances have been compared with other generally used statistical methods. In effect, through hierarchical Bayesian modeling that incorporates more complex settings, some of these models are shown to be superior to the other methods in certain scenarios in terms of statistical power and robustness in identifying RV association. All models have also been applied to real data analyses to detect RVs significantly associated with continuous phenotypes such as systolic blood pressure and low-density lipoprotein cholesterol level. Some of our results confirm previous findings and others provide novel evidence of the involvement of RVs in these complex phenotypes that are missed by other methods such as SKAT and Granvil. Additionally, we focus on applying a time-to-event model with a kinship matrix to GWAS on transitions between different smoking stages to improve our understanding of the genetic architecture underlying smoking behavior. We employ a Cox model with multivariate normal random effects to deal with correlated time-to-event phenotypes. Our results provide novel evidence for supporting the hypothesis that complex neurotransmitter networks are involved in initiation of smoking behavior.
  • Jernman, Juha (Helsingin yliopisto, 2015)
    Neuroendocrine tumors of the rectum were regarded as benign, when Oberndorfer originally described the entity in 1907. Later, he acknowledged that some neuroendocrine tumors (or carcinoids, the term at that time) behave in a more aggressive manner, and a few of them even had the potential to metastasize with poor outcome. In the novel World Health Organization (WHO) classification launched in 2010, all neuroendocrine tumors of the gastrointestinal (GI) tract are malignant. In this classification, tumors of every part of the GI tract are graded uniformly according to proliferation index and mitotic frequency, whereas the TNM-classification (tumor, node, metastasis) is specific for each site. Around 10% of gastroenteropancreatic neuroendocrine tumors (GEP-NETs) occur in the rectum. The tumor series comprised 73 rectal NETs, with the main objective being to study the prognostic value of the WHO 2010 classification in rectal NETs: additionally, as the WHO classification has been used for a rather short time, tumor markers were tested to find a good, reliable prognostic tool. The WHO 2010 had excellent prognostic significance; none of the G1-NETs (grade 1) metastasized, whereas G2-NETs were often disseminated, some of them at initial presentation. Metastatic NETs have a poor prognosis. Cell-cycle antigen cyclin A also correlated with prognosis, and G2-NETs with high cyclin A expression were all metastatic. Transcription factor prospero homeobox 1 (PROX1) was immunohistochemically positive in a significant proportion of rectal NETs, and showed a correlation with metastatic potential and survival. It was also possible to conclude that the novel stem cell-associated factor HES77 (human embryonic stem cell factor 77) correlated well with rectal NETs metastatic potential and prognosis. These results support the validity of the WHO 2010 classification in rectal NETs. In view of this study, for patients with a rectal G1-NET, one follow-up endoscopy to exclude local recurrence might suffice. Intensive follow-up does not seem indicated, as metastatic potential is very low. As to G2-NETs, a thorough work-up is recommended, since most of these tumors disseminate eventually, some after several years, and a standard 5-year follow-up may not suffice. PROX1-positivity suggests that colorectal adenocarcinoma and rectal NET may, to some extent, share the same pathway in oncogenesis, which could lead to future therapeutic applications.
  • Luostarinen, Teemu (Helsingin yliopisto, 2015)
    The main goals of neuroanesthesia are to maintain adequate cerebral perfusion pressure (CPP) and, consequently, cerebral blood flow (CBF) to guarantee sufficient oxygenation of the brain and to provide good surgical conditions for the neurosurgeon. This thesis aimed to examine critical aspects of neuroanesthesia with regard to CBF, CPP, blood coagulation, and transfusion of blood products. In our material the requirement of intraoperative blood product transfusion was low during surgery for ruptured arterial aneurysm. Intraoperative red blood cell transfusion (RBCT) seems to be preemptive in nature according to the hemoglobin levels prior to transfusion. RBCT is associated with intraoperative rupture of an aneurysm. Intraoperative RBCT may itself worsen SAH patients neurological outcome. In the event of sudden intraoperative rupture of an aneurysm, adenosine-induced asystole can be used to stop the bleeding and facilitate clipping of the aneurysm. Early infusion of fresh frozen plasma instead of crystalloids should be considered to compensate for expected excess bleeding in neurosurgery to preserve normal coagulation capacity. The potentially less harmful effect of hypertonic saline (HS), relative to mannitol, on blood coagulation may shift the decision towards HS when choosing an optimal solution for treatment of elevated ICP or brain swelling, at least when excess bleeding occurs. However, the clinical relevance of this finding remains unclear and warrants further study. Reliability of end-tidal concentration of carbon dioxide (EtCO2) as an estimate of arterial carbon dioxide partial pressure (PaCO2) after anesthesia induction is not adequate, as seen in the correlation between a decrease in mean arterial pressure and EtCO2-PaCO2 difference in our study. Optimal ventilation after induction of anesthesia should be confirmed by arterial blood gas analysis in patients undergoing neurosurgery to prevent a potentially harmful increase in PaCO2, and consequently, in CBF. Anesthesia in both sitting and prone positions is associated with changes in blood pressure and cardiac function. However, preemptive goal-directed therapy with either Ringer s acetate or hydroxyethyl starch (HES) solutions before positioning enables a stable hemodynamic state during neurosurgery in both positions. The fluid requirement did not differ between the two positions. Slightly less HES is needed to achieve comparable hemodynamics, but is it questionable whether this advantage outweighs the recent concerns regarding colloid safety.
  • Airaksinen, Katja (Helsingin yliopisto, 2015)
    The aims of this PhD research were to study the feasibility of magnetoencephalography (MEG) measurements in patients with advanced Parkinson's disease (PD) treated with deep brain stimulation (DBS) of the subthalamic nucleus (STN), and to cast light onto the effects of DBS on the electrical activity of the brain. The sensitivity of MEG to magnetic fields makes the measurements of patients using electrical devices challenging because of confounding artifacts. One aim was to investigate how an artifact suppression method called spatiotemporal signal space separation (tSSS) works in the suppression of DBS artifacts. The measured MEG signals included evoked responses, spontaneous brain activity and cortico-muscular coherence (CMC) in DBS-treated PD patients. A total of 24 PD patients were included in the studies. The effect of STN-DBS was measured with DBS first being on and then turned off. CMC and corticokinematic coherence (CKC) were studied in 10 healthy subjects. The motor task for CMC and CKC was to hold the dorsiflexion of the wrist. For CMC recordings, MEG was recorded simultaneously with surface EMG over the activated extensor carpi radialis muscle. The healthy volunteers had an accelerometer sensor attached to their index fingernail for the CKC measurements. The DBS artifact reduction by tSSS enabled a reliable MEG signal analysis for most patients. The effects of DBS on the brain's electrical activity varied considerably between patients. AEF N100m amplitude increased in the right hemisphere for ipsilateral stimulation during DBS. When the DBS was on and eyes open, the source strength of the sensorimotor spontaneous MEG in the 6 10 Hz and 12 20 Hz bands correlated with the UPDRS rigidity measures. No systematic increase in CMC in the 13 25 Hz band during DBS was observed. The patients that had CMC also had the best UPDRS motor scores. The source locations of CMC and CKC in healthy subjects were overlapping and their spectral profiles resembled each other. tSSS is useful in the artifact suppression in the MEG measurements of DBS patients. Despite this, the MEG measurements of DBS patients are still challenging and need exact planning of the experimental setups and caution in the data analysis. DBS modifies cortical electrical activity, and some of these modifications correlate with the clinical improvement in PD. The accelerometer device may turn out to be a good alternative to EMG for coherence calculations even during hold tasks.
  • Polinati, Padmini (Unigrafia, 2015)
    Mitochondrial diseases are generally caused by genetic variants that may affect cell function during the process of energy generation: right from the start of protein translocation to the fatty acid degradation by beta-oxidation (β-oxidation). The main objective of this PhD thesis is to study genetic variants that cause mitochondrial diseases and also to understand the disease pathogenesis of a known disease using the induced pluripotent stem cell (iPSC) method, a revolutionary approach in regenerative medicine. In the first study, we carried out a long-term follow up of six metabolic diseased patients and subsequently we performed a carrier frequency study of the identified carnitine palmitoyl transferase 1A (CPT1A) gene variant in the Finnish population. We identified a novel homozygous variant c.1364A>C (p.Lys455Thr) in exon 12 of the CPT1A gene. No carriers of the variant c.1364A>C were detected upon minisequencing of 150 control samples but the allele frequency of CPT1A variant in global population is 0.0002142 (ExAC Browser) whereas in the Finnish population (6614 allele number) the frequency is 0.001966. The identified variant was predicted to cause improper folding of the CPT1A protein, which leads to its degradation. All patients were treated with a high-carbohydrate and a low fat diet. In the second study, we focused on the human DnaJ (Hsp40 homolog) subfamily C, member 19 (DNAJC19) deficiency. Our studies showed that it causes early onset dilated cardiomyopathy syndrome (DCMA). This is the first report of a genetic defect in the mitochondrial protein, DNAJC19, outside of the Canadian Dariusleut Hutterite population. This defect is characterized by an unusual aetiology for an early onset recessively inherited dilated cardiomyopathy that is associated with ataxia and male genital anomalies. Sequencing of the human DNAJC19 gene revealed a homozygous single nucleotide (A) deletion in exon 6 that cause a frameshift and lead to the premature termination of the protein. In the third study, the pathogenesis of retinopathy in long-chain acyl-CoA dehydrogenase deficiency (LCHADD) was studied using iPSC technology. Retinopathy is an unusual manifestation of LCHADD, as mitochondrial fatty acid β-oxidation (FAβO) has not been considered to play a major role in the metabolism of the retina. Among all defects of mitochondrial FAβO, only long-chain acyl-CoA dehydrogenase (LCHAD) and mitochondrial trifunctional protein (TFP) deficiencies have developed pigmentary retinopathy and peripheral neuropathy. We elucidated how a genetic variant in the FAβO cycle can disrupt the retinal pigment epithelium (RPE) that can eventually lead to blindness. In addition, we developed a new in vitro cell model; iPSC clones were generated from LCHADD patient fibroblasts and further differentiated into RPE cells. Several changes were observed in patient RPE cells such as decreased cell size, lower pigmentation and irregular pattern of morphology. Electron microscopy analysis showed an accumulation of a few melanosomes, more melanolysosomes, and large sized lipid droplets in patient RPE cells. Furthermore, increased levels of triglycerides in patient RPE cells were observed upon mass spectrometric analysis. We concluded that all these changes had contributed to the disruption of the RPE layer that leads to blindness in LCHAD deficiency patients. Finally, the research done for this thesis succeeded in identifying novel variants in CPT1A and DNAJC19 genes in Finnish patients. Our long-term follow up studies on CPT1A deficiency can help patients in better diagnosis, which further helps clinicians to identify the genetic cause. We also found a novel phenotype with DNAJC19 deficiency. Further we established the groundwork to understand the pathogenesis of retinopathy in LCHADD patients using an advanced method that helps to study in depth pathogenesis mechanism.
  • Kriikku, Pirkko (Helsingin yliopisto, 2015)
    Driving under the influence of drugs (DUID) can adversely affect driving skills in numerous ways and put lives at risk. Legal approaches to DUID vary considerably from country to country, even within Europe, and, in the last decades the emergence of new psychoactive substances (NPS) has further complicated the scene. DUID is an unlawful act if the substance taken is banned or impairs driving. The latter is hard to define and prove, putting pressure on governments to ban NPS as quickly as possible in order to protect the public by facilitating enforcement of DUID laws. However, banning requires knowledge on several aspects of NPS such as prevalence, pharmacology, abuse potential and toxicity. Up-to-date, evidence-based information on NPS is needed by legislators, toxicologists, clinicians, and other health care professionals. Such information would enable potential drug users and the public to be more aware of the risks associated with illicit use of NPS. This study aimed to add to the knowledge of the NPS most relevant in Finland. In this thesis, the prevalence, blood concentrations in drivers and in post-mortem cases, and demographic details of 3,4-methylenedioxypyrovalerone (MDPV) and desoxypipradrol (2-DPMP), were investigated. Changes in prevalence and other characteristics of MDPV were monitored over a time span covering a period before its banning as well as a few years after banning. Phenazepam, a Russian therapeutic benzodiazepine now illegal in Finland, was studied by examining both DUID and post-mortem cases. The use by apprehended drivers of pregabalin, a prescription anticonvulsant with therapeutic indications for neuropathic pain, partial seizures and generalised anxiety disorder, was also studied. The results of this study showed that DUID cases provide a valuable source of information on NPS prevalence and user profiles. However, little specific information could be gained about the impact on driving performance and health risks of NPS mainly due to the fact that NPS were usually used together with a spectrum of other psychoactive substances. It could, however, be concluded that all of the studied NPS were frequently detected in the samples collected from apprehended drivers and, in the case of MDPV, the prevalence changed with time. The number of MDPV-positive cases among apprehended drivers decreased by 51.1% after the drug was banned. The concentrations of NPS found in DUID cases were within the range anticipated to produce significant adverse effects on driving performance, or, in some cases, in the range found in post-mortem cases where the drug may have contributed to the fatality. The presence of the medicinal drug, pregabalin, was found to be connected to abuse rather than appropriate medical use since it was in most cases found in concentrations higher than those recommended for therapeutic use and together with illegal drugs such as amphetamine or cannabis. In post-mortem cases positive for MDPV, the prevalence of suicide was much greater than in fatalities related to other drugs. Three independent registries, namely the DUID toxicology data, the post-mortem toxicology database, and court documents, were examined to gain novel information on the characteristics of NPS use and those abusing them. The large number of cases studied produced information on concentration ranges associated with abuse of the studied substances.
  • Mäkinen, Mauno (Helsingin yliopisto, 2015)
    The psychological influence of obesity on health is less clear than the physical impacts. Further follow-up studies are needed to examine causality/directionality. To investigate psychiatric disorders in excess-weight adolescents, more studies with diagnostic interviews are required, because the results of previous results have been contradictory, probably reflecting methodological differences. The aim of the study presented in this dissertation was to examine psychological well-being and psychiatric disorders linked to overweight and obesity in a general mid-adolescent population. The study was performed from 2003 to 2005, involving girls and boys aged approximately 14.5 years attending secondary schools in Helsinki. The students completed self-assessments surveying self-esteem (RSES), their thoughts and ideas concerning eating behaviors (EDI), as well as their lifestyle. Both measured and self-reported weights and heights were recorded (Study I sample: 650 girls, 693 boys; Study II sample: 614 girls, 651 boys). A subgroup of adolescents (Study III subsample: 86 girls, 96 boys) was diagnosed by an adolescent psychiatrist using a structured diagnostic instrument (K-SADS-PL). Furthermore, a subgroup (Study IV follow-up subsample: 78 girls, 88 boys) was followed up for one year and completed a questionnaire assessing the self-image (OSIQ-R) both at baseline and on follow-up. In summary, psychological well-being was good in most of the overweight and obese adolescents. However, the excess-weight adolescents significantly more often revealed body dissatisfaction and other symptoms related to eating disorders (p < 0.001) and abnormal dietary behavior (p < 0.001) than their normal-weight peers. Adolescents with abnormal eating behavior reported significantly greater body dissatisfaction than those with normal eating behavior (p < 0.001). The excess-weight adolescents significantly more seldom reported experiences of dating than their normal-weight peers (p < 0.001). The boys with excess weight exercised significantly more seldom than their normal-weight peers (p < 0.001). The prevalence of lifetime and current psychiatric disorders did not significantly differ between the excess-weight and normal-weight adolescents. The prevalence of one or more current psychiatric disorders was 13.2% among adolescents with excess-weight. The self-image of girls with normal weight developed intensively during the one-year follow-up period compared to girls with excess weight (p < 0.024). In component scales, the difference in change scores was largest in sexuality (p = 0.018) and vocational attitudes (p = 0.041).
  • Hellsten, Kati Susanna (Helsingin yliopisto, 2015)
    GABAergic inhibition is crucial for regulation of neuronal excitability and hence for optimal function of the nervous system. The GABAA receptor forms a pentameric ligand-gated anion channel. The subunit combinations of the receptors define their pharmacological and electrophysiological properties. The individual subunits display a distinct but often widespread distribution throughout the nervous system. GABAA receptor-mediated inhibition can be divided to fast and transient synaptic inhibition and to background (tonic) inhibition that is mediated by extrasynaptic GABAA receptors. The synaptic inhibition regulates the rhythmic activities in neuronal networks. Benzodiazepines, interacting with an allosteric binding site of GABAA receptors typically concentrated at synaptic locations, are anxiolytic drugs, but their long-term use evokes severe side-effects. The functional role of extrasynaptic GABAA receptor-mediated inhibition is not yet well understood. Expanding the understanding of local role of GABAA receptors and neuronal circuits mediating anxiolysis is relevant for the development of more selective and safe treatment for anxiety disorders. Studying the properties of extrasynaptic GABAA receptors may help to understand their physiological relevance and role in psychiatric and neurological disorders. The main objectives of this thesis were to study the local expression of benzodiazepine-sensitive GABAA receptors in human locus coeruleus (LC), the pharmacology of extrasynaptic GABAA receptors in vivo (in mice) and in vitro and the brain structures mediating acute anxiolytic responses in a transgenic mouse model as a consequence of enhanced tonic inhibition in specific forebrain structures. Firstly, the present in situ hybridization and receptor autoradiography studies in human LC revealed benzodiazepine binding sites indicating that LC may directly mediate the sedative and/or anxiolytic effects of benzodiazepines in humans. Previously, contradictory reports had been published in regard to the benzodiazepine binding sites in the rodent LC suggesting a potential species difference in the direct sites of action of benzodiazepines. Secondly, the behavioral tests showed that gaboxadol, a GABA site agonist, acts preferentially via extrasynaptic α6β/γ2 receptors in vivo as the transgenic mice overexpressing these receptors were significantly more sensitive to the anxiolytic and hypnotic effects of gaboxadol than the wild-type mice. Electrophysiological recordings on recombinant receptors revealed that GABA is a partial agonist as compared to gaboxadol at α6β3, α6β3δ and α4β3δ receptors but not at α6β3γ2 receptors. Gaboxadol and GABA displayed also different receptor desensitization and deactivation kinetics on these receptors. Thirdly, by increasing tonic inhibition in specific forebrain structures, the anxiolytic dose of gaboxadol increased c-Fos expression in the transgenic mouse model especially in the limbic areas, such as the cingulate cortex, septal nuclei, central extended amygdala and basolateral nucleus of amygdala. In addition to demonstrating how neuronal excitability can be altered in different brain regions as a consequence of enhanced tonic inhibition, this result suggests that a widespread neuronal inhibition, as typically associated with benzodiazepines, may not be the exclusive mechanism of acute anxiolysis.
  • Kotilainen, Hannele (Helsingin yliopisto, 2015)
    Introduction Non-tuberculous mycobacteria (NTM) are ubiquitous and they have become more common than M. tuberculosis (MTB). Most infections due to NTM are pulmonary and cutaneous, thereafter disseminated infections and lymphadenitis. Mycobacterium avium complex (MAC) is entailing for more than half of clinical cases. Chronic pulmonary patients and non-smoking healthy elderly females are the main risk groups for pulmonary NTM infections and others have been opportunistic NTM among in immunosuppressive patients and in families, rare genetic defects with disseminated NTM. Host defence role and immunological defects behind pulmonary NTM infections have not been found except a local defect in nitric oxide production and ciliary beat of airway epithelium. In present studies, the clinical symptoms and smoking as a risk factor for NTM infection and the prognostic value of The American Thoracic Society (ATS) criteria have been evaluated. Prognosis and clinical findings of patients infected with MAC has been compared to patients with other NTM infections. The clinical signs suggested immunological deficiency, therefore we investigated an association between deficiencies of complement components C4A or C4B and NTM and MTB patients in Finland. Study population. The studies I-II were based on 120 adult non-HIV patients with at least one NTM isolation retrospectively collected data in 1990 1998 followed at least four years. The patients were classified as smokers or never smokers and according to fulfilment of ATS 2007 criteria. In study III, 167 adult non-HIV patients with at least one positive NTM classified as MAC or other NTM and according to ATS 2007 criteria including patients from Studies I-II and IV with the median follow-up time 7.0 y. Study IV based on prospectively collected data and blood samples on HIV-negative 50 adult NTM and 31 MTB patients in the area of Helsinki University Central Hospital in 2004- 2009. The blood samples analyzed for C4A and C4B genotype. Controls were comprised of 149 healthy, unselected Finnish people. Results. Nearly half of patients with a NTM isolation had never smoked. Among non-smokers, 72 % were female, instead among smokers, only 30% were female. MAC comprised a majority of isolates among non-smokers compared to smokers (72% vs. 41%). Symptoms had started within a year of positive NTM isolation in 48% of patients. Smokers had higher risk of mortality than non-smokers but no difference was observed after adjusting for underlying diseases. The 2007 ATS criteria were fulfilled by half of patients. ATS positive cases were, more often, female with less fatal underlying diseases as compared to ATS-negative cases. No significant difference was seen in median survival time or symptoms between ATS-positive and -negative cases. ATS criteria fulfillment was a weak prognostic marker. MAC was isolated in 59% of overall 167 cases and MAC patients were significantly more often female (70 % vs. 34 %), and presented significantly fewer fatal underlying diseases than patients with other NTM (23 % vs. 47 %).The other NTM patients had suffered from symptoms less than a year as compared to MAC patients (54% vs. 34%). Pulmonary MAC patients had significantly lower risk of death and significantly longer survival time than other pulmonary NTM patients (13.0 years vs. 4.6 years). Serious underlying diseases and age were the significant predictors of mortality, female sex was a predictor of survival. Finnish NTM patients (72%) had significantly more frequent C4 deficiencies (C4A or C4B) as compared to unselected healthy controls (56%) and MTB (35%). C4 deficiencies were significantly common in female NTM patients (81%) as compared to female controls (55%). Conclusion. Risk factors for NTM infection were different for smokers and non-smokers. ATS 2007 criteria had a poor prognostic value in finding patients with risk of fatal outcome. Patients with MAC had a longer survival than patients with other NTM. Obviously symptom onset suggests a fairly rapid disease course. Complement C4 deficiency was found to be a plausible risk factor for NTM infection, especially among elderly female patients.
  • Helenius, Mikko (Helsingin yliopisto, 2015)
    Irreversible vascular remodeling has a central role in a variety of pathophysiological conditions including pulmonary arterial hypertension (PAH). Hypoxia and inflammation are prominent features in PAH, along with hyperplasia and hypertrophy of vessel wall layers. Although, endothelial cell (EC) dysfunction is thought to drive the multiple forms of vascular remodeling, the origins of this phenomenon are poorly understood. Extracellular ATP and its metabolites are important regulators of vascular tone, permeability, and homeostasis. Yet little is known about their role in pathological vascular remodeling. By using chronic hypoxia and PAH animal models as well as human PAH patient samples, this study was undertaken to evaluate the catalytic activities and expression levels of nucleoside triphosphate diphosphohydrolase-1 (NTPDase1, otherwise known as CD39) and other purine-converting ectoenzymes with a primary focus on vascular EC. For this purpose we employed thin-layer chromatographic enzyme assays with 3H-labelled nucleotide substrates, in combination with various immunoassays and qPCR. In addition we have developed a highly sensitive assay for simultaneous sensing of extracellular ATP and its metabolites and also a novel method for measuring CD39 activity in modeled to in vivo conditions. In functional assays, cells or animals were stimulated through purine signaling pathways and proliferation, apoptosis, permeability, and DNA damage were assayed. Our results clearly demonstrated that the activity of CD39 was downregulated in chronic hypoxia, monocrotaline induced animal models of PAH and in human PAH patients. Attenuated enzyme activities could create a niche in the vasculature where ATP levels were increased and adenosine levels were decreased. Even a small increase in ATP concentration was enough to induce an apoptosis-resistant, hyper-proliferative, and DNA-damageresistant phenotype in ECs of pulmonary origin. The observed effects were at least partly dependent on P2Y11 receptor activation. In addition, we found that low ATP concentrations could induce pulmonary smooth muscle cell proliferation and migration. Interestingly, we found that small apelin peptide could directly restore the downregulated CD39 activity. This study implies that purinergic signaling, ATP mediated cell activation in particular, plays a truly significant role in pathological vascular remodeling, and that it could be used as a therapeutic target. Moreover, purinergic signaling pathways could be used before vascular injury to precondition EC against irradiation or chemotherapy induced DNA damage.
  • Laitakari, Elina (Helsingin yliopisto, 2015)
    Background: In recent decades, the total number of burn injuries has decreased globally, while the number of childhood burns has been increasing. Infant burn victims are a specific group because they are highly dependent on parents or caregivers. Aim of the study: The aim of this study was to clarify special characteristics and clinical outcome of infant burns. The purpose of this study was to determine the incidence and trends of inpatient and outpatient treatments for burn-injured infants. Patients and methods: Burn victims younger than 1 year born 1990 2010 were identified from the National Institute of Health and Welfare (THL) and the Statistics Finland registers. The analysis consisted of the incidence of burn injury, trends of treatment, and risk factors for burns. Between 2005 and 2009 126 burn victims aged less than 1 year were treated in the Children s Hospital, Helsinki, Finland. Hospital electronic databases were searched to clarify detailed information and patients treated as inpatients and outpatients were analyzed separately. The HRQoL of the 126 burn-injured children was evaluated with a validated, standardized 17D questionnaire 5 to 9 years after the burn injury. Results: Between 1990 and 2011, 1842 infant burn victims were treated at Finnish hospitals. The annual overall incidence per 1000 persons rose from 0.77 to 2.04, and the incidence of outpatient-treated burns increased from 1.11 to 1.67. Based on the analysis, the risk factors for infant burn injuries are male gender, being firstborn, the mother s young age, and low socioeconomic status. During the 5-year study period 20 burn injured infants were treated as inpatients and 106 as outpatients in the Children s Hospital. Girls outnumbered boys in the inpatient-treated group, and 45% were younger than 6 months. The final mean TBSA was 8.5% (from 0.5 to 40%). In the outpatient group, 52% were boys, and 57% were aged 9 to 12 months. The final mean TBSA burned was 1.4% (range from 0.5 to 7%). Caregiver eyewitnessed the accident in 66% of the cases, and 44% of the burns were on the hands. Most burns occurred at home, and two thirds of the burns were scalds caused by hot liquids. The respondents (n=44, 35%) appeared not to suffer from long-term consequences of the burn injury, and the respondents and non-respondents proved to be similar in terms of age, injury severity, and treatment given. Conclusions: The number of infant burns increased in Finland during the study period. Two thirds of infant burns were scalds caused by hot liquids, and most of these were preventable. Firstborn boys aged 9 to 12 months of young mothers with low socioeconomic status are at greater risk for burn injuries. The long-term HRQoL of burn-injured children was comparable to that of the controls.
  • Medvedovsky, Mordekhay (Helsingin yliopisto, 2015)
    During the last years magnetoencephalography (MEG), has become an important part of the pre-surgical epilepsy workup. Interictal activity is usually recorded. Nevertheless, the technological advances now enable ictal MEG recordings as well. The records of 26 pharmaco-resistant focal epilepsy patients, who underwent ictal MEG and epilepsy surgery, were reviewed. In 12 patients prediction of ictal onset zone (IOZ) localization by ictal and interictal MEG was compared with ictal intracranial EEG (icEEG). On the lobar surface level the sensitivity of ictal MEG in IOZ location was 0.71 and the specificity 0.73. The sensitivity of the interictal MEG was 0.40 and specificity 0.77. The records of 34 operated epilepsy patients with focal cortical dysplasia (FCD) were retrospectively evaluated. The resected proportion of the source cluster related to interictal MEG was evaluated in respect to postoperative seizure outcome. 17 out of 34 patients with FCD (50%) achieved seizure freedom. The seizure outcome was similar in patients with MR-invisible and MR-visible FCD. With MEG source clusters and favorable seizure outcome (Engel class I and II) the proportion of the cluster volume resection was 49% - significantly higher (p=0.02) than with MEG clusters but unfavorable outcome (5.5% of cluster volume resection). Median nerve somatosensory evoked MEG responses were processed by movement compensation based on signal space separation (MC-SSS) and on spatio-temporal signal space separation (MC-tSSS). MEG was recorded in standard and deviant head positions. With up to 5 cm head displacement, MC-SSS decreased the mean localization error from 3.97 to 2.13 cm, but increased noise of planar gradiometers from 3.4 to 5.3 fT/cm. MC-tSSS reduced noise from 3.4 to 2.8 fT/cm and reduced the mean localization error from 3.91 to 0.89 cm. The MEG data containing speech-related artifacts and data containing alpha rhythm were processed by tSSS with different correlation limits. The speech artifact was progressively suppressed with the decreasing tSSS correlation limit. The optimal artifact suppression was achieved at correlation of 0.8. The randomly distributed source current (RDCS), and auditory and somatosensory evoked fields (AEFs and SEFs) were simulated. The information was calculated employing Shannon's theory of communication for a standard 306-sensor MEG device and for a virtual MEG helmet (VMH), which was constructed based on simulated MEG measurements in different head positions. With the simulation of 360 recorded events using RDCS model the maximum Shannon's number was 989 for single head position in standard MEG array and 1272 in VMH (28.6% additional information). With AEFs the additional contribution of VMH was 12.6% and with SEFs only 1.1%. To conclude, ictal MEG predicts IOZ location with higher sensitivity than interictal MEG. Resection of larger proportion of the MEG source cluster in patients with FCD is associated with a better seizure outcome, however, complete resection of MEG source cluster is often not required for achievement of favorable seizure outcome. The seizure outcome is similar in patients having MR-positive and MR-negative FCD. MC-tSSS decreases the source localization error to less than 1 cm, when the head is displaced up to 5 cm; however, it is reasonable to limit use of movement compensation for no more than 3-cm head displacement to keep the head inside sensor helmet. The optimization of the tSSS correlation limit to about 0.8 can improve the artifact suppression in MEG without substantial change of brain signals. MEG recording of the same brain activity in different head positions with subsequent construction of VMH can improve the information content of the data.
  • Ahmed Haji Omar, Abdirisak (Helsingin yliopisto, 2015)
    Oral (OSCC) and cutaneous (CSCC) squamous cell carcinomas are epithelial neoplasms, which are both derived from keratinocyte cells. However, the etiology and prognosis of OSCC and CSCC are different. The main etiological factors behind OSCC are tobacco smoke and alcohol consumption, and for CSCC it is UV-radiation. OSCC has poorer prognosis than CSCC. In order to be invasive, cancer cells have to pass various barriers. They have to disrupt cell-to-cell junctions, penetrate basement membranes, and invade connective tissue. The pattern of invasion of tumors varies strikingly. Some invade in large border fronts while others invade in single cell manner. Expression of the transmembrane proteins, E-cadherin and syndecan-1, in cell membrane are lost during tumor invasion, and therefore loosening cell adhesion. Matrix metalloproteinases (MMP) are tissue proteinases, which have a proteolytic role in various physiological events and during tumor progression MMPs are capable of degrading extracellular matrix (ECM) proteins but also have an immunomodulatory role. Toll-like receptors (TLRs) are part of the innate immunity and can recognize exogenous pathogen associated molecular patterns or endogenous damage associated molecular patterns. Cancer cells may use TLRs to induce tumor-promoting inflammation. The aim of the study was to examine possible cellular and molecular differences between OSCC and CSCC explaining their different behaviors as cancers despite having cellular similarities. The study included 36 OSCC and 27 CSCCs from patients with early stage histological risk assessment (HRA) model and histological risk assessment score (HRS). We performed immunohistochemical staining for E-cadherin, Snail (Snail1), Syndecans (1 and 2), MMPs (7, 8 and 9) and TLRs (4 and 5). In vitro, with oral and cutaneous cell lines the effect of TLR-5 ligand flagellin on proliferation, migration and invasion was studied. OSCC patients had worse disease-specific survival than CSCC patients and this correlated with the invasion depths of the OSCC tumors. OSCC had a more severe histological pattern of invasion than CSCC. E-cadherin and Syndecan-1 expression decreased in the invasive front of OSCCs and CSCCs. Syndecan-1 expression in the tumor stroma was higher in OSCC than in CSCC in tumors with invasion depth over 4 mm. MMP-7 was mainly expressed in the invasive front of OSCC and CSCC and was stronger in OSCC than in CSCC. MMP-8 and MMP-9 were mainly expressed in the peritumoral inflammatory cells. TLR-5 expression was stronger in OSCC than in CSCC. In vitro, TLR-5 ligand flagellin increased proliferation, migration and invasion of less aggressive oral and cutaneous cell lines, but failed to do so with the most aggressive oral cancer cell line. In conclusion, the OSCC patients of this study had poorer disease specific survival than CSCC patients. Increased stromal syndecan-1 expression in OSCC, MMP-7 expression in the invasive front of the tumor, and MMP-9 expression in inflammatory cells could partly explain the differences in survival between OSCC and CSCC.
  • Keltanen, Terhi (Helsingin yliopisto, 2015)
    Postmortem (PM) biochemistry is utilized in cause of death (CoD) investigations. The challenges in PM biochemistry are caused mainly by the PM changes. Analytes may be degraded or denatured leading to methodological difficulties. In addition, the re-distribution may cause falsely elevated values when compared to clinical samples. Analytes may also be consumed or produced by microbial metabolism. The sample matrix is often different from that in clinical biochemistry and the reference levels need be obtained through research. The actual death process and agonal period can also lead to unique changes in the analyte concentrations. Despite these challenges, with a sufficient number of samples and adequate research it is possible to optimize biochemical methods for PM samples and obtain reference values for certain conditions. The biochemical results can provide supporting information in cases where the death is caused or associated with metabolic disturbances for instance in diabetes mellitus (DM) or alcohol abuse. Hyperglycemia and ketoacidosis in diabetes can be fatal if not treated. The macroscopical, histological and toxicological findings in autopsy may be scarce or even absent. Determination of vitreous humor (VH) glucose and ketone bodies are very informative in these cases. As ketoacidosis can be present also after heavy alcohol use, the determination of glycemic status prior to death is essential in distinguishing between these two conditions. Glycated hemoglobin (HbA1c) provides information on the glycemic balance in previous weeks. In our studies, we have assessed the results, methods and interpretation provided by analysis of glucose, lactate, ketone bodies, HbA1c and C-reactive protein (CRP) in PM samples. Optimization has been performed in routine casework according to our findings providing more accurate information to the medico-legal pathologists in their CoD investigation work. In addition, our results have provided some insights into the pathology of severe diabetic emergencies as well as to ketoacidosis due to alcohol. According to our results, we recommend that in DM and alcohol abuse related deaths VH glucose, total ketone bodies or beta-hydroxy butyrate and blood HbA1c should always be analyzed. Lactate levels may provide additional information, when the results are interpreted considering the PM interval (PMI). These analyses are recommended also in cases where no suspicion of metabolic disturbances exists, but the findings in autopsy are scarce. The collaboration between the medico-legal pathologists and the laboratory is very important for the development and understanding of PM biochemistry and to include novel analyses to support the CoD investigation. The CoD determination is very important not only on the individual level, but also nationally, since Finnish Statistics collects the CoD data, which can be than further utilized in recommendations concerning health and primary prevention.
  • Zhang, Yanlei (Helsingin yliopisto, 2015)
    Type 2 diabetes occurs as a consequence of aging, family history, physical inactivity, unhealthy diet and obesity and it is an increasing public health problem worldwide. The condition is associated with high rates of mortality from co-morbid cardiovascular diseases and poor health-related quality of life (HRQoL). A large proportion of individuals with type 2 diabetes are not diagnosed for up to a decade after onset unless a screening programme has been implemented. The estimated prevalence of undiagnosed diabetes in China accounts for about 60-70% of the diabetes population. Diabetes education that is targeted at the general population is the key to increase public knowledge and awareness and is a fundamental building block for addressing the diabetes epidemic. Screening for identifying undiagnosed diabetes could lead to earlier identification and intervention, and postpone or prevent the onset of diabetes and its complications. However, there is a paucity of study on the impact of education and screening programmes on HRQoL, lifestyle modification of the targeted population, and the cost-effectiveness of such programmes. The objectives of this study were to investigate 1) the cost and effectiveness of a population-based education programme to increase public knowledge and awareness of diabetes; 2) the cost-effectiveness of two different screening tools for undiagnosed diabetes; 3) impact of type 2 diabetes and its complications on individuals HRQoL; 4) impact of a screening programme on individuals overall HRQoL, depression dimension and lifestyle modification. This study was based on data analyses of two population-based diabetes surveys conducted in 2006 (Survey A) and 2009 (Survey C), and a dataset of diabetes high-risk population identified through the Qingdao Diabetes Prevention Program (QD-DPP) between 2007 and 2010 in Qingdao, China (Survey B). The same stratified, random cluster sampling method was used in Surveys A and C to recruit a representative sample of the general population who had lived in Qingdao city for at least five years. A total of 5355 individuals in Survey A and 5110 in Survey C aged 35-74 years participated in the surveys. A total of 3108 rural participants who did not have diabetes at baseline in 2009 were invited for a re-examination and 1782 individuals attended the follow-up survey. A standard 2h 75g oral glucose tolerance test was administrated to all participants in both surveys. Diabetes education and health promotion information and activities under the framework of QD-DPP were given via printed and audio-visual media, the internet, free distribution of information booklets and diabetes risk score (DRS) flyers that targeted the entire population of 1.94 million who lived in the intervention areas. An adult with a DRS more or as much as 14 was considered at high-risk for diabetes and invited to a nearby community clinic for a free capillary blood glucose test. The proportions of participants who correctly marked obesity, physical inactivity and positive family history of diabetes as the risk factors of diabetes were doubled in both urban and rural populations, irrespective of age and gender during the QD-DPP education campaign period. The cheapest way to inform 1000 individuals about type 2 diabetes was to distribute DRS flyers (¥54, ¥ = Chinese yuan), followed by the newspapers articles (¥77), booklets (¥313) and by radio programmes (¥375) (1 ≈7¥, for the year 2015). The fasting capillary glucose (FCG) test and Chinese DRS questionnaire were used as the first-line screening tools and these were evaluated for detecting undiagnosed diabetes in primary care settings. The sensitivity of FCG and DRS was 65.1% and 65.8%, whereas their respective specificity was 72.4% and 55.2%. The costs per undiagnosed diabetes identified at the optimal cut-off values of 6.1 mmol/l for FCG and 14 for DRS were ¥674 and ¥844, respectively. The area under curve (AUC) was higher for FCG than for DRS (75.3% vs. 63.7%, p less than 0.001). People with previously known type 2 diabetes reported that the symptomatic comorbidities had a strong negative impact on HRQoL; no significant difference was detected between people without diabetes and with newly diagnosed diabetes. The screening and labelling as pre-diabetes or normoglycaemia had no adverse impact on the participants overall HRQoL and depression. An improvement in lifestyle as measured by the frequency of physical activity and vegetable intake was observed at 3 years post screening in both groups. In conclusion, the QD-DPP education campaign efficiently increased public knowledge and awareness of diabetes. The DRS questionnaire is a simple, non-invasive and reliable first-line screening tool to identify undiagnosed diabetes at primary care settings. The diabetes screening programme in Qingdao generated positive changes towards a healthy lifestyle and did not result in any harm to the participants.
  • Pemovska, Tea (Helsingin yliopisto, 2015)
    Adult acute myeloid leukemia (AML) effectively illustrates the challenges of contemporary cancer drug discovery and development, as molecularly targeted therapies have not yet been translated in clinical practice. In fact the standard therapy (cytarabine and an anthracycline) for AML has not been changed in over 40 years. As a consequence, outcome remains poor with overall survival of 30-40%. The genetic alterations that are associated with AML have been mapped, but the underlying disease mechanism is poorly defined due to large inter-patient heterogeneity. In contrast, chronic myeloid leukemia (CML) is strictly driven by BCR-ABL1 and drugs targeting the ABL1 kinase activity have paved the way for oncoprotein targeting drugs in the treatment of cancer. In CML the main clinical challenge is instead the emergence of resistance to ABL1-directed therapy. This resistance typically occurs through point mutations in the kinase domain of ABL1 such as the clinically challenging T315I mutation. Hence, in both leukemia types there is an unmet need for novel therapeutic strategies. This study focused on development and implementation of an Individualized Systems Medicine (ISM) platform to identify novel therapeutic strategies for leukemia patients. The ISM strategy incorporated functional ex vivo drug sensitivity and resistance testing (DSRT), deep molecular profiling and clinical information to facilitate identification of personalized therapy approaches. A large number of approved and investigational anti-cancer compounds were tested and individualized selective responses were quantified with drug sensitivity scores (DSS). RNA and exome sequencing data was used to identify genetic alterations that enabled associating drug sensitivities with genetic alterations and biomarkers. The DSRT approach enabled functional taxonomy of AML patient samples based on drug responses, provided insights into disease biology, and identified effective drugs and drug combinations for individual patients and thus facilitated drug repurposing. In addition, integration of DSRT and molecular data identified phenotype to genotype links that has a potential for rapid translation of results. Clinical implementation of ISM data was plausible in approximately 80% of relapsed and refractory AML patient cases, and meaningful and evaluable responses were achieved in approximately 40% of cases. Notably, emergence of in vivo resistance to targeted therapy was mirrored in the DSRT profile of the relapsed samples, highlighting a solid correlation between ex vivo and in vivo drug responses. Finally, this study identified a number of kinase inhibitors that can be repositioned for specific subtypes of AML and CML, such as dasatinib in combination with a FLT3 inhibitor for AML patients with FLT3-ITD mutations and axitinib for patients with BCR-ABL1(T315I)-driven leukemia. The results of this thesis demonstrate how unbiased drug sensitivity profiling of patient-derived cancer cells is a powerful way to discern unforeseen drug-disease and drug-target links with clinical implications and provides a workable concept that can be implemented in routine clinical care of cancer patients in the future.