Lääketieteellinen tiedekunta

 

Recent Submissions

  • Robciuc, Alexandra (Helsingin yliopisto, 2017)
    The cornea is the first optical element of the eye and, together with the eyelids, eye socket, tears, and sclera, shares an important part in ocular protection. It is a thin, transparent, avascular tissue with a rigorous layered structure. The continuous contact with the outside environment exposes the ocular surface tissues, such as the corneal epithelium, to pathogens, mechanical traumas, irritants, toxins, allergens, or radiation from the sun. The cellular stress response represents an adaptive reaction to environmental stimuli and defines the health-state of the tissue, the absence/presence of clinical manifestations. We have aimed in this thesis project to study the stress response of the corneal epithelium to environmental stimuli and to determine its contribution to ocular surface diseases such as climatic droplet keratopathy (CDK), infection, or dry eye disease. A cell culture model of the corneal epithelium was exposed to environmental stress – UV radiation, LPS, or hyperosmolarity (HO) – to identify macromolecular alterations: mRNA expression, protein localization, enzyme activation, lipid conversions. CDK is a degenerative disease of the cornea with increased prevalence in warm, dry climate. Examination of corneal tissue and tears from patients with CDK suggested an involvement of metalloproteinases (MMPs) in the disease-associated tissue degradation. Our cellular model helped reveal the connexion between UV radiation and the unbalanced secretion of gelatinases (MMP-2 and MMP-9) and thus explain in part the pathogenesis of this rare disease. The evaluation of the inflammatory response to UV, initially, and then to LPS, or HO, highlighted IL-8 secretion as an acute stress marker and followed throughout the studies. Human corneal epithelial cells were found also to release lipid-modifying enzymes into the cell culture medium as a response to stress. Of particular importance to us were the enzymes of the sphingolipid metabolism, a lipid signalling pathway of great importance in the stress response. These enzymes were released as part of cell-derived extracellular vesicles, the vesicle-lipids, however, were the mediators of a significant decrease in IL-8 levels. The same sphingolipid enzymes appeared responsible for the intracellular response to HO, controlling the IL-8 production but also the stress-induced neutral lipid loading. We have therefore succeeded to establish a causative link between UV radiation and tissue degeneration in CDK, to determine the role of the sphingolipid signalling pathway in ocular surface stress and to discover more about HO consequences in the corneal epithelium. The stress response at the ocular surface is a thin balance between tissue protection and maintenance of function. Inflammation represents one of the most relevant clinical signs of distress and we aimed to identify targets for therapies that seek to restore tissue homeostasis.
  • Pyykönen, Aura (Helsingin yliopisto, 2017)
    Optimal care of pregnant women and newborn babies has an immeasurable effect on the well-being of a family, but it also significantly impacts society overall. In Finland, the quality of obstetric and perinatal care is high, but in order to further improve it, sensitive and commonly agreed quality indicators are required. The aim of the thesis was to validate different potential quality indicators for obstetric care, and to assess differences between different hospital size-categories in Finland. To facilitate reliable comparisons for benchmarking, we evaluated the usability of the Robson classification, a system for grouping the parturient population. This was done as an international comparison between the Nordic countries. The thesis also aims to contribute to more unified and high-quality obstetric care by examining the optimal time of labor induction in prolonged pregnancies. This thesis is based on data from the medical birth registries in Finland and in the other Nordic countries. The findings of this thesis indicate true differences in the obstetric management and treatment culture between the Nordic countries and between different-sized birth units in Finland. However, it should be acknowledged that part of the differences may be explained by confounding background factors. The results enforce the existing evidence on the effect of birth centralization on neonatal outcomes, e.g. lower neonatal mortality in large units, but indicate a higher risk for instrumental delivery. Obstetric trauma, a potentially preventable complication of vaginal delivery, showed substantial size-dependent variation between the Finnish birth units; the risk was the lowest in mid-sized birth units. The Robson classification proved to bed usable in the Nordic setting, facilitating more accurate comparisons, but comprehensive interpretations require experience and expertise. In our study on the effect of labor induction in prolonged pregnancy, we found an increased risk for Cesarean section around 41 gestational weeks, but no longer when the gestational age approached 42 weeks. From a neonatal perspective, labor induction resulted in a decreased risk for meconium aspiration syndrome, but only when the intervention took place before 41+5 weeks. Labor induction did not have a significant effect on neonatal mortality. International and inter-unit benchmarking pinpoints the areas needing improvement; e.g. prevention of obstetric trauma or induction protocols among nulliparous women. Some indicators, such as Cesarean section rates within Robson groups, show potential for readily directing obstetric practices, and are therefore of special interest. Like our neighboring countries Denmark and Sweden, we should establish a national quality monitoring program for obstetric care in Finland, to ensure its high-quality care also in the future.
  • Meinilä, Jelena (Helsingin yliopisto, 2017)
    Gestational diabetes mellitus (GDM) is increasing globally and it causes pregnancy complications and later type 2 diabetes (T2D) for the mother and offspring. Studies of the association between diet in Nordic populations and GDM, as well as the effect of observed dietary change on the risk is scanty. The thesis was based on data from the Finnish Gestational Diabetes Prevention Study RADIEL, a randomized controlled trial with diet and physical-activity counselling. The participants were either obese (body mass index, BMI ≥30 kg/m2) or had a history of GDM, and they were recruited either before pregnancy or at early pregnancy. Based on 3-day diet records, at the 1st trimester of pregnancy the pregnant women had fat intake of 33 (standard deviation (SD) 6) % from total energy (E%), intake of saturated fatty acids higher than recommended (12, SD 3 E%), and low intake of carbohydrate (46, SD 6 E%). Average intakes of vitamins D (mean 7 µg, SD 4) and A (724 µg, SD 357), folate (282 µg, SD 85), and iron (12 mg, SD 3) from food sources were below the Nordic Nutrition Recommendations (NNR), but mean total intakes (from foods and supplements), excluding vitamin A, were above the recommended lower level. The proportion of users of any dietary supplements was 77%. The purpose of developing a diet quality index (Healthy Food Intake Index) was to study the level of adherence to the NNR in pregnant women at high risk of GDM. The 11 components of the HFII reflected the food guidelines of the NNR, intakes of relevant nutrients, and characteristics known to vary with diet quality. The HFII showed reproducibility. High scores in the HFII, and thus, high adherence to the NNR at 1st trimester was associated with 2nd trimester’s lower glucose concentrations 2 hours after 75 g oral glucose tolerance test. Dietary changes towards the food guidelines of the NNR during pregnancy were associated with a lower risk of GDM. A diet adherent to the NNR may lower the risk of GDM in high-risk women. This highlights the need for adequate NNR-based dietary intervention in early pregnancy of obese women and women with a history of GDM. Guidance should emphasize importance of quality of fats and safe sources of vitamin A. Vitamin A status of Finnish pregnant women warrants further investigations. With minor adjustments, the HFII is a promising instrument for maternity clinics for quick screening of pregnant women’s diet quality.
  • Kontro, Mika (Helsingin yliopisto, 2017)
    Direct translation of cancer-specific genomic information into effective personalized therapies for acute leukemias has proven to be difficult. The aim of this study was to utilize novel tools for detailed characterization of genomic, transcriptomic, and functional aberrations in acute leukemia to gain understanding of disease pathology and guide individualized therapies. The main methods used were ex vivo drug sensitivity and resistance testing (DSRT), exome and transcriptome (RNA) sequencing, all of which were facilitated by extensive biobanking. In study I, we developed DSRT platform performed on primary leukemic cells ex vivo for identification of effective patient-specific drugs. Exome and RNA sequencing of serial samples were used for molecular characterization and to understand mechanisms of acquired resistance. Although AML samples exhibited unique DSRT profiles, responses could be clustered in five distinctive groups. Individualized treatment of refractory patients with DSRT-guided therapy resulted in meaningful clinical responses in 3/7 patients. In study II, we studied molecular drivers for relapsed T-cell acute lymphoblastic leukemia (T-ALL) and found novel STAT5B mutations. Functional studies demonstrated these mutations to enhance the transcriptional activity and to induce constitutive phosphorylation of STAT5B. The mutated blasts showed elevated BCL-XL expression and sensitivity to the pan-BCL-2 inhibitor navitoclax. Targeted sequencing revealed activating STAT5B mutations in 6/68 patients. In study III, we first evaluated ex vivo BCL-2 inhibitor sensitivity of AML cells, then we systematically assessed whether these responses correlated to specific mutations or gene expression signatures. BCL-2 inhibitor sensitivity associated with mutations in IDH1/IDH2 and WT1, as well as with aberrations in chromatin modifiers. Importantly, overexpression of a specific set of HOX genes predicted highly selective responses to BCL-2 inhibition. In study IV, we developed a national hematological biobank to allow researchers to access high-quality samples with accompanying clinical data. The samples from three collection time-points—diagnosis, potential remission, and relapse—are available. For this study, we evaluated the quality of stored samples and demonstrated that extracted DNA and RNA remain usable for demanding down-stream experiments.
  • Salo, Emma (Helsingin yliopisto, 2017)
    This thesis investigated brain activity with functional magnetic resonance imaging (fMRI) during selective and divided attention. Selective attention refers to the ability to selectively attend and process certain stimuli while ignoring others. Divided attention is needed, when two or more tasks requiring selective attention are performed in parallel. However, attention is easily caught by sudden changes in the environment. Brain activity and task performance during distracted and undistracted attention was also compared. The results showed that selective attention to auditory or visual modality enhanced activity in the auditory and visual cortices, respectively. However, the results also implied that the effects of selective attention may be influenced by task-irrelevant variation of the stimuli. The present thesis also showed that during divided attention, when two cognitively demanding tasks are performed in parallel, brain activity is enhanced in cortical areas in relation to the two tasks performed one at a time. Such activity enhancements specifically associated with divided attention were found in left middle frontal cortex during several different divided attention tasks. This suggests that division of attention requires specific, higher-level cognitive processes not needed in other attention-engaging tasks. However, not all dual tasks were associated with frontal activity enhancements. Complex selective attention tasks performed in parallel with another selective attention task may influence the higher-level cognitive processes or integration of two tasks needed during dual tasking.
  • Tenca, Andrea (Helsingin yliopisto, 2017)
    BACKGROUND AND AIM This thesis includes two studies conducted in a paediatric and two studies conducted in an adult primary sclerosing cholangitis (PSC) population. The common denominator was endoscopic retrograde cholangiography (ERC) with brush cytology that was performed in all patients. The aims were to: i) identify the possible environmental risk factors (Study I) and report the long-term outcome (Study II) of paediatric-onset PSC, ii) compare ERC and magnetic resonance imaging with cholangiopancreatography (MRI-MRCP) in the evaluation of disease activity and severity of patients with PSC (Study III) and evaluate the role of ERC with brush cytology as screening for cholangiocarcinoma (CC) in patients with PSC (Study IV). MATERIAL AND METHODS PSC was diagnosed, followed-up (or both) in Helsinki University Hospital (HUH). Study I: 71 patients with a new diagnosis of paediatric-onset (age < 16 years) PSC, autoimmune hepatitis (AIH) or PSC-AIH (togheter autoimmune liver diseases or AILD) between 1985-2011. Two control groups were used: 1) 91 IBD patients matched for gender and age, collected from the IBD Population Registry at HUH and 2) 716 healthy subjects matched for gender, age and also place of birth at the time of AILD diagnosis, collected from the Population Registry Centre. A questionnaire of 22 items was administered. Study II: 41 patients with a new diagnosis of paediatric-onset PSC between 1993-2011. Study III: 48 patients with PSC who underwent ERC and MRI-MRCP within + 3 months for the diagnosis or the follow-up of the disease. Study IV: 261 patients with a new diagnosis of PSC (age > 18 years) between 1 January 2006 and 31 October 2011. All cholangiographic images were scored according to the modified Amsterdam PSC score. RESULTS Study I: In multivariate analysis, children ‘living with a cat in a block of flats’ had a higher risk (OR 3.6; 95% CI: 1.2-10.8) of having AILD than healthy controls, but not IBD controls. Study II: At the end of follow-up (9 years, range 2-20 years) all children were alive and no malignancy occurred. 29/33 (88%) were not transplanted; 26/29 (78%) were not cirrhotic and 3/29 (10%) were cirrhotic. 4/33 (12%) were transplanted after a median of 7.5 years; no PSC recurrence in the graft occurred. Study III: MRCP and ERC scores for IHBD were associated with alkaline phosphatase (p = 0.016 and p = 0.018, respectively) and CA19-9 level (p < 0.001 and p = 0.030, respectively); MRCP score for EHBD was also associated with CA19-9 level (p = 0.021). Finally, peribiliary enhancement detected on MRI correlated with cytology findings for both IHBD (Spearman’s rho = 0.322, SE: 0.095, p = 0.022) and EHBD (Spearman’s rho = 0.319, SE: 0.113, p = 0.025, respectively). Study IV: Most of the patients were asymptomatic (211/261; 80.8%) and had only mild changes on cholangiography (149/261; 57.1%) at time of first ERC. Follow-up was completed in 249/261 (95%). CC developed in 7 patients and biliary dysplasia in 8 patients; brush cytology was suspicious or malignant in 8 patients at time of PSC diagnosis. Advanced EHBD cholangiographic changes (HR: 1.7; 95% CI: 1.2-2.3) and alanine aminotransferase (HR: 14.2; 95% CI: 1.9-106.4) were associated with increased risk of biliary neoplasia. CONCLUSIONS An unidentified environmental risk factor (i.e., microbial) especially associated with cats may increase the risk of PSC in children. The clinical course and outcome of paediatric-onset PSC seems to be good until adulthood with a high survival rate, with no occurrence of malignancy and LT required in only a minority of patients. MRI-MRCP use in PSC follow-up seems to be low. In this respect, ERC with brush cytology is a good screening tool for detection of biliary dysplasia or neoplasia (or both) in patients with PSC. Advanced extrahepatic disease and alanine aminotransferase elevation may predict the occurrence of CC.
  • Känsäkoski, Johanna (Helsingin yliopisto, 2017)
    Sexual differentiation and pubertal development are complex processes whose disruption leads to the abnormal development of primary and/or secondary sexual characteristics. Although mutations in several genes have been implicated in these disorders, the majority of the patients still lack a molecular genetic diagnosis. The aim of this thesis work was to identify genetic defects underlying complete androgen insensitivity syndrome (CAIS), congenital hypogonadotropic hypogonadism (CHH) and gonadotropin-dependent precocious puberty (GDPP), in Finnish and Danish patients. The genetic cause of CAIS was investigated in two siblings without identified mutations in the androgen receptor gene (AR). Whole-genome sequencing and AR cDNA analysis revealed a deep intronic mutation that led to abnormal splicing of AR mRNA and undetectable amount of AR protein in patient fibroblasts. The genetic causes of both extremes of pubertal variation, GDPP and CHH, were investigated in Danish patients. Twenty-nine Danish girls with GDPP were screened for mutations in MKRN3, which was recently identified as a regulator of pubertal onset, and one girl was found to have a mutation in this gene. Forty-one Danish CHH patients were screened for mutations in the CHH genes ANOS1, FGFR1, FGF8, PROK2, PROKR2, GNRHR, TAC3, TACR3, and KISS1R. Additionally, CHD7 was screened in two patients with hearing loss. Twelve patients were found to have a conclusive mutation; either in FGFR1 (5), ANOS1 (4), GNRHR (1), or CHD7 (2). SEMA3A and SEMA7A, two candidate genes of CHH, were screened in fifty Finnish CHH patients. Three SEMA3A variants and two SEMA7A variants were identified in the patients, but the identified variants do not seem to be sufficient to cause CHH alone. In conclusion, the intronic AR mutation is the first reported case of pseudoexon activation leading to CAIS demonstrating the importance of AR cDNA analysis in AIS patients without a molecular genetic diagnosis. Mutations in MKRN3 underlie GDPP in Denmark, although they are not very common in sporadic cases. FGFR1, ANOS1, GNRHR, and CHD7 mutations were found to underlie CHH in the Danish patients, but the majority still remain without a molecular genetic diagnosis. Finally, mutations in SEMA3A and SEMA7A do not seem to contribute significantly to CHH, and it remains to be seen whether mutations in these genes cause CHH in humans.
  • Sagalajev, Boriss (Helsingin yliopisto, 2017)
    Despite extensive research on the mechanisms of nociceptive pain, little is known about the processes that lead to neuropathic pain development. As one of the most severe and drug-resistant forms of chronic pain, neuropathic pain represents a major burden for patients, their families, and society. Therefore, new insights into the pathophysiology of neuropathic pain are needed. The amygdala represents a complex of brain nuclei responsible for mediating negative emotions, such as fear. Moreover, maladaptive changes in the amygdala contribute to the development of chronic pain and its comorbidities, such as anxiety and depression. However, little is known about the mechanisms for amygdala-mediated pain hypersensitivity in neuropathy. We therefore investigated descending pathways engaged by the amygdala for modulation of spinal nociception in neuropathic rats. In particular, we investigated the contribution of oxidative stress in the amygdala on the development of neuropathic pain. Oxidative stress is characterized by overproduction and poor detoxification of reactive oxygen species (ROS). ROS act as endogenous agonists of transient receptor potential (TRP) channels, which account for transduction of noxious stimulation to a nociceptive signal in peripheral nerves and for nociceptive transmission in the spinal cord. We therefore assessed how blocking TRP channels and detoxification of ROS with antioxidants in the amygdala influence pain hypersensitivity and pain affect in neuropathic rats. Furthermore, we investigated whether electrical cortical stimulation can reverse changes associated with pronociceptive activity of the amygdala. In particular, we examined descending pathways recruited by stimulation of the secondary somatosensory cortex (S2), which has proven to be an efficacious site for attenuation of drug-resistant neuropathic pain by transcranial magnetic stimulation (TMS) in patients. In the present series of studies, we demonstrate that N-methyl-D-aspartate (NMDA), TPRPA1, and TRPC4/5 channels in the amygdala contribute to the development of neuropathic pain and that amygdaloid treatment with antioxidants can attenuate this pain. Furthermore, we provide evidence that S2 stimulation suppresses spinal nociception in neuropathic animals with hypersensitivity, but not in animals without hypersensitivity or in control animals. Medullospinal serotonergic pathways acting on the spinal 5-HT1A receptor underlie the descending pain inhibition in neuropathic animals both following treatment of the amygdala with antioxidants or TRPA1 antagonists and following S2 cortex stimulation.
  • Gluschkoff, Kia (Helsingin yliopisto, 2017)
    Occupational stress in teaching concerns not only teachers; it also impacts on wider social contexts. It may negatively affect teachers’ health, but also indirectly influence students’ health and their academic achievements. Yet, despite the challenges teacher stress and health problems pose for society, little research has examined and compared the relevance of different psychosocial work characteristics in predicting poor teacher health, or explored mechanisms that explain or moderate these associations in the teaching profession. This thesis examined the associations between different psychosocial work characteristics and health-related outcomes among Finnish teachers, as well as the potential explanatory (i.e., mediating) and moderating mechanisms for these associations. The data were gathered via self-report questionnaires and included a cross-sectional sample of primary school teachers from the Helsinki metropolitan area of Finland and a longitudinal sample of teachers in primary or secondary education from the prospective Finnish Public Sector study. Psychosocial work characteristics involved job strain, effort-reward imbalance, organizational injustice, and teacher-targeted violence. The health-related outcomes potentially associated with psychosocial work characteristics included depressive symptoms, burnout, and sleep problems. In addition, the extent to which different aspects of recovery explained the associations and the moderating role of organizational justice were examined. Psychosocial work characteristics in terms of effort-reward imbalance and, to a lesser extent, with regard to job strain, were found to be relevant predictors of poor health in teaching. Effort-reward imbalance was associated with higher levels of burnout and, compared with job strain and organizational injustice, this was the most important predictor of depressive symptoms. Job strain was associated particularly with impaired sleep. Although organizational injustice did not seem to be a major predictor of poor health, high organizational justice represented a valuable resource in the teachers’ psychosocial work environment. Encountering violence at work had the most pronounced effect on sleep among teachers working in relatively unjust conditions, whereas the sleep of those perceiving high organizational justice was not affected by violence. Some of the effects of psychosocial work characteristics on health were mediated through aspects of recovery; namely, through sleep and recovery experiences during leisure time. Non-restorative sleep partially explained both the association of job strain with depressive symptoms and the association of effort-reward imbalance with depressive symptoms and overall burnout score. Furthermore, the association between effort-reward imbalance and burnout was partially mediated through poor relaxation during leisure time. The indirect effects were relatively weak, suggesting that although poor recovery may partly mediate the association between psychosocial work characteristics and health-related outcomes in teaching, it does not play a major role in the process. For teachers in basic education, reducing the demanding aspects of the psychosocial work environment and increasing the rewarding elements, such as the respect and support they receive, may be important in occupational stress prevention. Furthermore, although preventive measures against teacher-targeted violence should be prioritized, resources aimed at promoting organizational justice in schools may also mitigate the adverse consequences of teacher victimization. Although improving teachers’ psychosocial work environment is probably the most important means of supporting their health, interventions that help teachers unwind after working hours and reduce sleep problems may further complement workplace development programs.
  • Holstila, Ansku (Helsingin yliopisto, 2017)
    Physical inactivity is a significant risk factor for non-communicable diseases. It has also been associated with a decline in functioning and a higher risk of work disability. However, there is limited evidence concerning the causes and consequences of changes in physical activity. Most Finnish adults of working age do not meet the recommendations for health-enhancing physical activity, and people tend to become less physically active as they age. Increasing activity levels among older age groups could enhance functioning and work ability among the ageing population. The aim of this thesis was, first, to examine how changes in physical activity are associated with subsequent health functioning, sickness absence and disability retirement. The physical and mental health functioning and sickness absence attributable to musculoskeletal and mental causes were examined separately. Second, the intention was to investigate how physical activity changes after the transition to statutory retirement and during post-retirement years. The research was part of the Helsinki Health Study being carried out at the Department of Public Health, University of Helsinki. The baseline surveys were conducted in 2000-2002 (N=8,960, response rate 67%) among employees of the City of Helsinki aged 40-60. The employees who responded to the baseline survey were followed up in two later surveys, meanwhile the cohort aged and some of the employees retired. The phase-2 follow-up survey was conducted in 2007 (N=7,332, response rate 83%) and phase 3 in 2012 (N=6,814, response rate 79%). The survey data were linked with register data on sickness absence from the Social Insurance Institution of Finland, and on disability retirement from the Finnish Centre for Pensions among those who consented to the register linkage (N=6,606). The register data on sickness absence and disability retirement includes medically confirmed diagnoses. Sickness absence periods were followed up from phase 2 until 2009, and disability retirement from phase 2 until 2013. Increased physical activity was associated with better physical health functioning and decreased activity with worse physical health functioning. There were fewer associations between changes in physical activity and mental health functioning. Increases in physical activity were associated with a lower risk of sickness absence. Vigorous physical activity was especially beneficial for physical health functioning and contributed to a lower risk of sickness absence attributable to musculoskeletal diseases. In contrast, a higher intensity of physical activity had less of an effect on mental functioning and sickness absence attributable to mental causes. In some cases, moderate-intensity physical activity was more beneficial to mental health functioning than higher-intensity activity. In addition, adopting vigorous physical activity was associated with a lower risk of disability retirement, and decreasing the intensity from vigorous to moderate or low was associated with a higher risk. Physical activity increased after the transition to statutory retirement, but declined a few years after retirement. Given the results of this study, ageing employees and retirees engaging in a low level of physical activity should be encouraged to increase the level. Vigorous activity could also be promoted, at least among healthy individuals. The transition to statutory retirement is a good opportunity to promote physical activity and thereby facilitate a change for the better. It is also important to support the maintenance of physical activity in the years following the transition to retirement.  
  • Helin, Tuukka (Helsingin yliopisto, 2017)
    Direct oral anticoagulants (DOACs), the thrombin inhibitor dabigatran and the anti-Xa inhibitors rivaroxaban and apixaban are are simple to use as standard dosing is recommended and no routine monitoring of coagulation is advocated. However, in certain clinical situations, e.g. acute thrombosis or bleeding, emergency surgery and renal or hepatic failure, assessing anticoagulant bioactivity is essential. The aims of this study were to firstly assess the effects of these DOACs in commonly used coagulation screening tests (PT, INR and APTT) using both spiked samples and samples collected from patients on treatment. Secondly, the availability of specific methods for drug effect assessment, namely drug-calibrated thrombin time (dabigatran) and anti-Xa (rivaroxaban, apixaban) assays were assessed in laboratory surveys. Thirdly, the effects of DOACs on these specific assays were explored using patient samples. Finally, the effect of DOACs on the global coagulation assay thrombin generation (TG) was assessed in patients using dabigatran, rivaroxaban or apixaban. The spiked sample laboratory surveys included a total of 86 European laboratories. The effects of all DOACs on INR were modest, but large reagent variability was noted in the responses (p<0.001). In APTT, rivaroxaban and apixaban prolonged the APTT only modestly, but with dabigatran there was a clear prolongation. Only about a fourth of the laboratories were able to provide more specific methods, TT or anti-Xa assays for drug quantification. Dabigatran effects in patient samples were studied 241 unselected patient samples. The PT was rarely prolonged and the APTT was in curvilinear relationship (R2 = 0.71) with dabigatran. However, the sensitivity effects among patients varied, with some patients having a normal APTT at dabigatran levels up to 160 ng/mL. Several specific assays, (diluted thrombin time, ecarin clotting assay and anti-IIa assay) accurately quantified dabigatran concentrations. In TG, a paradoxical increase was observed in the endogenous thrombin potential (ETP) and Peak TG, while lag time also prolonged with increasing concentrations of dabigatran. Rivaroxaban and apixaban effects on coagulation in patient samples were further assessed in a well-characterised group of patients using these drugs for thromboprophylaxis after orthopaedic surgery. Rivaroxaban peak drug levels were higher and trough levels lower than with apixaban, reflecting the once daily vs twice daily dosing of the drugs. In TG, the ETP response at peak drug levels was strong with rivaroxaban, whereas with the trough drug levels, the TG parameters were close to the baseline levels. With apixaban the responses were more stable. The RVVT, used as a qualitative test, detected prolongation only at rivaroxaban peak levels. In conclusion, DOACs present significant challenges in the clinical laboratory, as a wide variety of reagents are used in coagulation assays, and the drug responses also vary widely between individual patients. Many factors other than drug concentration influence the responses in coagulation assays and TG. In clinical practice, care and diligence is required when assessing the effects of DOACs on patients.
  • Parkkonen, Olavi (Helsingin yliopisto, 2017)
    Takotsubo cardiomyopathy (TTC) is an acute cardiac condition resembling in symptoms acute coronary syndrome (ACS), but without obstructive coronary artery disease. TTC develops almost solely in post-menopausal women and usually after preceding stress. Of all patients with ACS symptoms, TTC incidence is 2%. Due to similar symptoms and findings, differential diagnosis requires coronary angiography (CAG). The pathophysiology of TTC is unknown. Even though the accumulated evidence suggests a causative role for a catecholamine surge, other theories exist. Aborted myocardial infarction (MI) produces similar electrocardiography (ECG) and biochemical findings as in TTC. In such cases, because of non-stenotic coronary artery plaques, a dissolved coronary thrombus might show no any signs in the CAG, which could lead to an assumption of non-atherothrombotic etiology for the heart attack. In ACS, altered levels of proteolytic enzyme called matrix metalloproteinase 8 (MMP-8), and its inhibitor, the tissue inhibitor of matrix metalloproteinase 1 (TIMP-1), associate to plaque rupture. Their direct comparison between ACS and TTC remains unknown. The purpose of this thesis is to test whether either of two non-invasive methods: the ECG (I) or MMP-8 and TIMP-1 (III), could differentiate TTC from ACS. We also set out to find whether, after all, what is responsible for the TTC is transient thrombosis in the coronary circulation (II). Both our prospective and retrospective collection of patients resulted in 92 TTC cases. The demographics of our material were similar to those of other TTC reports worldwide. In the ECG study (I), a review and comparison of 57 TTC and ACS acute ECGs resulted in criteria differentiating the two with 63% sensitivity and 93% specificity. In cases of suspected ST-elevation myocardial infarction, such accuracy is insufficient for a decision against coronary intervention. Blood samples from 45 TTC patients and matching numbers of ACS patients and controls were analyzed for coagulation markers. Despite the similar acute-phase reaction in TTC and ACS patients, the TTC d-dimer levels matched those of controls, and were lower and less frequently above reference level than with those in ACS. The blood samples were further analyzed for MMP-8 and TIMP-1 levels showing, on admission, better differentiation between TTC and ACS by TIMP-1 than by troponin T (TnT). In TTC, low ejection fraction (EF) correlated with low MMP-8/TIMP-1 ratio. In conclusion, ECG lacked the ability to differentiate TTC from ACS that would have allowed avoidance of invasive diagnostics. Secondly, the coagulation results supported catecholamine and argued against the thrombosis theory. Finally, TIMP-1 emerged as a potential future biomarker in differentiation between ACS and TTC. Furthermore, in some TTC patients MMP-8 and TIMP-1 levels may explain more severe left-ventricle (LV) impairment.
  • Lorey, Martina (Helsingin yliopisto, 2017)
    The innate immune system is the first line of defence against microbial infections. Macrophages are the key cells of the innate immune system. Activated macrophages release inflammatory mediators through conventional and extracellular vesicle (EV)-mediated protein secretion. These protein secretion pathways are major components of intercellular communication in the immune system. Protein secretion is activated by inflammasomes in response to microbial components and endogenous danger signals. Inflammasomes are molecular platforms of the innate immune system that are critical to both local and systemic inflammation: they are involved in the pathogenesis of immune-mediated inflammatory diseases. Inflammasomes can be activated by both microbial stimuli as well as by endogenous danger signal molecules. The secretome of the cell is the pattern of all secreted molecules at a given time; it can provide important insights about the cell’s status as well as information about intercellular communication. The secretome can be studied by mass spectrometry (MS)-based proteomics; this technique provides a system-level overview of the current state of the cell. While traditional assays like ELISA or Luminex can quantify selected proteins in low pg/mL levels, they can only analyse a limited number of proteins. Proteomics offers the same sensitivity but supplemented with the capability of multiplex quantitative analysis of thousands of proteins in the same sample. The main goal of this work was to unravel the innate immune response of human macrophages activated by microbial stimuli with a focus on protein secretion using proteomics combined with bioinformatics and functional studies. The microbial stimuli used included Influenza A viruses (IAVs) and lipopolysaccharide (LPS), the cell wall components of Gram-negative bacteria. Both IAVs and Gram-negative bacteria are important human pathogens associated with aggressive infections. Influenza A virus and LPS are known to activate canonical NLRP3 inflammasome and non-canonical caspase-4/5 inflammasome, respectively. Inflammasome activation is followed by protein secretion which was studied in detail, with a special emphasis on unconventional protein secretion. In both studies, mediators of inflammation were released through EVs in response to influenza A virus infection and intracellular LPS stimulation. These included many danger signal proteins, cytokines and components of translational machinery. Our system-level characterization using modern mass spectrometry-based proteomics approach provides novel information how macrophages communicate to other cells through protein secretion. The results suggest that unconventional protein secretion is an essential part of the innate immune response to different activation stimuli.
  • Pussila, Marjaana (Helsingin yliopisto, 2017)
    Colorectal cancer (CRC) is the second most common cause of cancer-related deaths in the Western world and interactions between genetic and environmental factors, including diet, are suggested to play a critical role in its etiology. Yet, the mechanisms by which diet impacts colorectal tumorigenesis remain largely unknown. Colorectal cancer evolves as a multistep process, which requires a series of genetic and epigenetic alterations in growth regulatory genes. The process is accelerated in individuals with inherited cancer predisposition such as Lynch syndrome (LS) which is one of the most common inherited cancer susceptibility syndromes and caused by inherited mutation in one of the DNA mismatch repair (MMR) genes. CRC is thought to develop via the so called adenoma-carcinoma sequence. However, the early events that occur in colon mucosa prior to polyp formation remain unknown. The research presented here investigates the gene expression changes arising in histologically normal colonic mucosa as putative cancer-preceding events available for early detection. This was achieved by pursuing a long-term feeding experiment in the mouse model for human Lynch syndrome (Mlh1+/-), and the wild type (Mlh1+/+) littermates, fed with either Western-style (WD) diet or healthy AIN-93G control diet. Carcinomas developed mainly in WD fed mice. WD also accelerated the progression of carcinogenesis. In the first study, the expression of 94 growth-regulatory genes previously linked to human CRC was studied for 5 weeks and 12 months old mice. Promoter CpG island methylation status was also studied for the genes which showed reduced expression. In mice fed for 12 months with WD, proximal colon mucosa, the predominant site of cancer formation in LS, exhibited a significant expression decrease in several tumor suppressor genes. Furthermore, a reduced mRNA expression was accompanied by an increased CpG dinucleotide promoter methylation of the respective genes suggesting a cause for the mRNA down regulation. The strongest expression decrease together with a significant increase in its promoter methylation was seen in Dkk1, an antagonist of the canonical Wnt signaling pathway. Furthermore, the inactivation of Dkk1 seemed to predispose to neoplasias in the proximal colon. The findings suggest that the inactivation of Dkk1 is a prominent early marker for colon oncogenesis. In study 2 the aim was to comprehensively clarify the role of Mlh1 expression during colon tumorigenesis, which is usually associated with Lynch syndrome and MSI. Here, the same mouse model and diets were used to study cancer-preceding expression changes in the colon mucosa of 12 and 18-month-old mice. The Mlh1 protein expression and MSI status were studied in the colon carcinomas, and the effect of inherited predisposition (Mlh1+/) and Western-style diet on those. CRC development always includes a lack of genomic integrity and the different types of genomic instabilities, such as chromosomal instability and MSI are thought to reflect distinct cancer initiating mechanisms. In the present study neither wildtype Mlh1+/+ nor heterozygote Mlh1+/- mice lacked the Mlh1 protein or showed MSI in their CRCs, while Mlh1 RNA expression was already significantly decreased in their normal mucosa. Instead, CRC mice showed a distinct expression profile with shortage of Mlh1 and several other chromosomal segregation gene-specific transcripts in mucosa and aberrant mitosis in tumors. The genome wide expression profiling experiment demonstrated that cancer-preceding changes are already seen in histologically normal colon mucosa and a that decreased expression of Mlh1 together with other chromosomal segregation genes may form a field-defect in mucosa and trigger MMR-proficient, chromosomally unstable CRC.
  • Jiang, Ping (Helsingin yliopisto, 2017)
    The high-level cognitive abilities such as attention and working memory develop throughout childhood and adolescence. Neuroimaging studies have shown that several brain regions including areas in the prefrontal (PFC) and parietal cortices play an important role in cognitive control. The prolonged maturation of the PFC and related networks may underlie the immature cognitive control abilities in children. In this thesis, functional magnetic resonance imaging and 1-back WM tasks were used to investigate 1) top-down modulation of brain activity in cortical areas related to visual information processing and 2) functional connectivity of resting state and task-related brain networks in healthy 7-11-year-old children and young adults. The tasks required the subjects to attend to either face or scene stimuli and to ignore distracting scene or face images, respectively. Studies I and II of this thesis found weaker or otherwise immature top-down modulation of the face processing-related visual association cortices that could partially be explained by the observed weaker functional connectivity between the PFC and the visual association cortex in the typically developing 7–11-year-old children compared to the young adults. Moreover, there were age-dependent differences in the recruitment of the PFC during visual working memory tasks. These age-dependent differences between the two groups are in line with the observed differences in the performance of the working memory tasks that was poorer in children than young adults. Study III showed that the 7-11-year-old children have already established an adult-like pattern of resting state networks, but especially during task performance, the functional connectivity within and between these networks differed from that in young adults. The group differences observed in the brain activation and functional connectivity are likely partly related to the morphological developmental state of the grey- and white matter in the 7–11-year-old children (i.e., the ongoing synaptic pruning and myelination of axons that continue up to young adulthood). The findings of this thesis conform to the suggestion that during development, the function of brain regions, especially the PFC, and the functional connectivity of brain networks, undergo dynamic changes, and that the same cognitive function may rely on different brain networks at different ages.