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  • Kallijärvi, Jukka (Helsingin yliopisto, 2006)
  • Salmi, Heli-Maria (Helsingin yliopisto, 2012)
    Normal human metabolism is a well-coordinated process involving the whole organism. It can be disturbed in true inborn errors of metabolism or, alternatively, in acquired situations where inappropriate nutrition and impaired energy production compromise normal metabolic control. These situations may share common pathophysiological mechanisms and metabolic markers. Their closer knowledge could provide new approaches for treatment or diagnosis. This thesis had two objectives, both dealing with biochemical changes in situations where significant metabolic perturbations occur. (1) To study thiol metabolism in children with several inborn errors of metabolism (n = 36) and in cell culture models, where similar metabolic conditions are created. Thiol levels and thiol redox status, serving multiple metabolic and regulatory purposes, could be affected in inborn errors of metabolism following inappropriate nutrition and compromised energy metabolism. Changes in thiol status could have an important role in the development of complications and even offer therapeutic potential in these diseases, as thiol levels can be influenced by diet or medication. (2) To investigate end products of metabolism in infants with cow s milk allergy (CMA; n = 35) as metabolic markers of inadequate nutrition in early childhood and altered intestinal microbial metabolism. Eventually, these changes could provide a novel diagnostic tool. Plasma and erythrocyte non-protein thiols (glutathione and cysteine) as well as thiols in cultured human fibroblasts, HEPG2 and 293T cells were measured with a liquid chromatography based method. Mass spectrometry was used for quantification of urinary excretion of end products of metabolism in CMA patients. Activities of enzymes related to thiol metabolism in human erythrocytes were studied in spectrophotometric assays. Patients with inborn errors of energy and nutrient metabolism had altered levels of plasma non-protein thiols glutathione and cysteine. Their plasma thiol redox status was indicative of oxidative stress, which thus seems to play a role in the pathophysiology of complications of these diseases. With further research, the changes in thiol status could have therapeutic implications. CMA is associated with measurable changes in urinary levels of end products of metabolism, which may be seen as markers of inadequate nutrition or altered intestinal bacterial metabolism. If confirmed in future studies, these changes could provide an innovative new approach to the diagnosis of CMA.
  • Edgren, Henrik (Helsingin yliopisto, 2012)
    Cancer is a disease characterized by the continuous accumulation of somatic cellular aberrations, whether in the DNA or epigenetic changes. During the last decade, many different techniques, including high resolution microarrays as well as exome- and whole genome sequencing, have been developed to comprehensively characterize these changes in cancer cells. In parallel with the development of laboratory techniques, a large variety of bioinformatic methods to analyze data from these have been developed. However, many of these concentrate on the analysis of data from only one laboratory technique, while it is becoming clear that advances in cancer research increasingly depend on integration of multiple different data types for the same tumors. Simultaneously, the recent explosive growth in sequencing data requires the development of new analythical methods. The aim of this thesis was to further characterize the genomic changes in breast cancer, with an emphasis on the development and application of bioinformatic methods to analyze and integrate data from different high throughput analysis techniques. In the first part of this work, the Gene Identification by Nonsense Inhibition method was applied to identify potential tumor suppressor genes in breast cancer. The integration of steady state gene expression, transcript stabilization and array comparative genomic hybridization data for six breast cancer cell lines led to the identification of a nonsense mutation in the RIC8A gene located at 11p15, a region deleted in ~15% of breast tumors. Taken together, our results suggest loss of RIC8A expression may be important in a subgroup of aggressive breast cancers. In study II, we developed a bioinformatic method for highly specific fusion gene identification from paired-end RNA-seq data. Application of the bioinformatic pipeline to data from four breast cancer cell lines led to the identification of 24 novel and three previously published fusion genes, with 95% specificity. In addition to showing that fusion genes are more prevalent in breast cancer than previously thought, several biological characteristics of fusion genes were identified. Most prominently, fusion genes were frequently associated with DNA copy number transitions, particularly high level amplifications, suggesting that most of them are not generated by balanced rearrangements. siRNA knock-down studies furthermore provided evidence for the functional importance of the VAPB-IKZF3 fusion gene in the BT-474 cell line. In the final study, we used aCGH to characterize the size distribution of the ERBB2 amplicon across 71 amplicon carrying tumors and 10 cell lines. To study the possible contribution to cancer of other coamplified genes in the amplicon, 23 genes amplified in 60% of tumors were selected for siRNA screening in two trastuzumab sensitive, two insensitive and one control cell line. In addition to single gene siRNA silencing experiments, five amplicon genes were knocked-down together with ERBB2 to identify synergistic effects. Our results suggest that cancer cells may be dependent on a number of genes in the ERBB2 amplicon besides the primary driver oncogene, a phenomenon termed non-oncogene addiction.
  • Lahdelma, Liisa (Helsingin yliopisto, 2009)
    Clozapine is the most effective drug in treating therapy-resistant schizophrenia and may even be superior to all other antipsychotics. However, its use is limited by a high incidence (approximately 0.8%) of a severe hematological side effect, agranulocytosis. The exact molecular mechanism(s) of clozapine-induced agranulocytosis is still unknown. We investigated the mechanisms behind responsiveness to clozapine therapy and the risk of developing agranulocytosis by performing an HLA (human leukocyte antigens) association study in patients with schizophrenia. The first group comprised patients defined by responsiveness to first-generation antipsychotics (FGAs) (n= 19). The second group was defined by a lack of response to FGAs but responsiveness to clozapine (n=19). The third group of patients had a history of clozapine-induced granulocytopenia or agranulocytosis (n=26). Finnish healthy blood donors served as controls (n= 120). We found a significantly increased frequency of HLA-A1 among patients who were refractory to FGAs but responsive to clozapine. We also found that the frequency of HLA-A1 was low in patients with clozapine-induced neutropenia or agranulocytosis. These results suggest that HLA-A1 may predict a good therapeutic outcome and a low risk of agranulocytosis and therefore HLA typing may aid in the selection of patients for clozapine therapy. Furthermore, in a subgroup of schizophrenia, HLA-A1 may be in linkage disequilibrium with some vulnerability genes in the MHC (major histocompatibility complex) region on chromosome 6. These genes could be involved in antipsychotic drug response and clozapine-induced agranulocytosis. In addition, we investigated the effect of clozapine on gene expression in granulocytes by performing a microarray analysis on blood leukocytes of 8 schizophrenic patients who had started clozapine therapy for the first time. We identified an altered expression in 4 genes implicated in the maturation or apoptosis of granulocytes: MPO (myeloperoxidase precursor), MNDA (myeloid cell nuclear differentiation antigen), FLT3LG (Fms-related tyrosine kinase 3 ligand) and ITGAL (antigen CD11A, lymphocyte function-associated antigen 1). The altered expression of these genes following clozapine administration may suggest their involvement in clozapine-induced agranulocytosis. Finally, we investigated whether or not normal human bone marrow mesenchymal stromal cells (MSC) are sensitive to clozapine. We treated cultures of human MSCs and human skin fibroblasts with 10 µM of unmodified clozapine and with clozapine bioactivated by oxidation. We found that, independent of bioactivation, clozapine was cytotoxic to MSCs in primary culture, whereas clozapine at the same concentration stimulated the growth of human fibroblasts. This suggests that direct cytotoxicity to MSCs is one possible mechanism by which clozapine induces agranulocytosis.
  • Salonen, Jonna (Helsingin yliopisto, 2009)
    Germ cell tumors occur both in the gonads of both sexes and in extra-gonadal sites during adoles-cence and early adulthood. Malignant ovarian germ cell tumors are rare neoplasms accounting for less than 5% of all cases of ovarian malignancy. In contrast, testicular cancer is the most common malignancy among young males. Most of patients survive the disease. Prognostic factors of gonadal germ cell tumors include histology, clinical stage, size of the primary tumor and residua, and levels of tumor markers. Germ cell tumors include heterogeneous histological subgroups. The most common subgroup includes germinomas (ovarian dysgerminoma and testicular seminoma); other subgroups are yolk sac tumors, embryonal carcinomas, immature teratomas and mixed tumors. The origin of germ cell tumors is most likely primordial germ cells. Factors behind germ cell tumor development and differentiation are still poorly known. The purpose of this study was to define novel diagnostic and prognostic factors for malignant gonadal germ cell tumors. In addition, the aim was to shed further light into the molecular mechanisms regulating gonadal germ cell tumorigenesis and differentiation by studying the roles of GATA transcription factors, pluripotent factors Oct-3/4 and AP-2γ, and estrogen receptors. This study revealed the prognostic value of CA-125 in malignant ovarian germ cell tumors. In addition advanced age and residual tumor had more adverse outcome. Several novel markers for histological diagnosis were defined. In the fetal development transcription factor GATA-4 was expressed in early fetal gonocytes and in testicular carcinoma precursor cells. In addition, GATA-4 was expressed in both gonadal germinomas, thus it may play a role in the development and differentiation of the germinoma tumor subtype. Pluripotent factors Oct-3/4 and AP-2γ were expressed in dysgerminomas, thus they could be used in the differential diagnosis of the germ cell tumors. Malignant ovarian germ cell tumors expressed estrogen receptors and their co-regulator SNURF. In addition, estrogen receptor expression was up-regulated by estradiol stimulation. Thus, gonadal steroid hormone burst in puberty may play a role in germ cell tumor development in the ovary. This study shed further light in to the molecular pathology of malignant gonadal germ cell tumors. In addition, some novel diagnostic and prognostic factors were defined. This data may be used in the differential diagnosis of germ cell tumor patients.
  • Okkonen, Marjatta (Helsingin yliopisto, 2012)
    Acute respiratory failure (ARF) is the most common type of organ failure leading to the need for intensive care. It is often secondary to acute lung injury (ALI) and its more severe form, acute respiratory distress syndrome (ARDS). ARF, and especially ALI and ARDS, cause increased morbidity, and mortality rates remain high (up to 40%). These disorders are characterised by inflammatory reaction and tissue damage. In some cases, inflammation continues and leads to an overwhelming repair process with ongoing fibrosis, accompanied by organ dysfunction and eventually a loss of function. Measuring the magnitude of the inflammation, and the repair process, would theoretically offer information concerning outcome. Early identification of patients whose disease process is likely to proceed unfavourably, would help clinicians to optimise their treatment. The aim of this study was to evaluate the epidemiology of ARF, its treatment, and outcome in Finland, with special interest in biomarkers, and their value in the prediction of mortality. Altogether, 958 adult patients treated with ventilatory support were prospectively included in this study during an eight week period in 2007 in 25 intensive care units. Plasma aminoterminal pro-brain natriuretic peptide (NT-pro-BNP) was assessed in 602 patients, and plasma cell-free DNA in 580 patients, to evaluate their prognostic value in ARF. Markers of collagen metabolism were studied in longitudinal serum samples in 68 patients in order to evaluate their evolution in ARF and the association to multiple organ dysfunction (MOD). Ventilatory support was used in 39% of all ICU patients. The estimated incidence of ARF was 149.5/100 000 per year. Median tidal volumes used were higher than recommended. Overall mortality at 90 days was 31%. Plasma NT-pro-BNP and cell-free DNA were highly increased in the majority of patients. Both markers were independent predictors of 90-day mortality, but their discriminative power was at most moderate when used separately. The mortality was highest in those patients, in whom both biomarkers were over their separate cut-off values. Thus, combined use of these biomarkers may increase their clinical value in the mortality prediction. The markers of collagen metabolism changed significantly over time in surviving patients. None of these markers did associate with MOD in these patients.
  • Mattila, Juha-Pekka (Helsingin yliopisto, 2009)
    The present study aims to elucidate the modifications in the structure and functionality of the phospholipid matrix of biological membranes brought about by free radical-mediated oxidative damage of its molecular constituents. To this end, the surface properties of two oxidatively modified phospholipids bearing an aldehyde or carboxyl function at the end of truncated sn-2 acyl chain were studied using a Langmuir balance. The results obtained reveal both oxidized species to have a significant impact on the structural dynamics of phospholipid monolayers, as illustrated by the progressive changes in force-area isotherms with increasing mole fraction of the oxidized lipid component. Moreover, surface potential measurements revealed considerable modifications in the electric properties of oxidized phospholipid containing monolayers during film compression, suggesting a packing state-controlled reorientation of the intramolecular electric dipoles of the lipid headgroups and acyl chains. Based on the above findings, a model describing the conformational state of oxidized phospholipid molecules in biological membranes is proposed, involving the protrusion of the acyl chains bearing the polar functional groups out from the hydrocarbon phase to the surrounding aqueous medium. Oxidative modifications alter profoundly the physicochemical properties of unsaturated phospholipids and are therefore readily anticipated to have important implications for their interactions with membrane-associating molecules. Along these lines, the carboxyl group bearing lipid was observed to bind avidly the peripheral membrane protein cytochrome c. The binding was reversed following increase in ionic strength or addition of polyanionic ATP, thus suggesting it to be driven by electrostatic interactions between cationic residues of the protein and the deprotonated lipid carboxyl exposed to the aqueous phase. The presence of aldehyde function bearing oxidized phospholipid was observed to enhance the intercalation of four antimicrobial peptides into phospholipid monolayers and liposomal bilayers. Partitioning of the peptides to monolayers was markedly attenuated by the aldehyde scavenger methoxyamine, revealing it to be mediated by the carbonyl moiety possibly through efficient hydrogen bonding or, alternatively, formation of covalent adduct in form of a Schiff base between the lipid aldehydes and primary amine groups of the peptide molecules. Lastly, both oxidized phospholipid species were observed to bind with high affinity three small membrane-partitioning therapeutic agents, viz. chlorpromazine, haloperidol, and doxorubicin. In conclusion, the results of studies conducted using biomimetic model systems support the notion that oxidative damage influences the molecular architecture as well as the bulk physicochemical properties of phospholipid membranes. Further, common polar functional groups carried by phospholipids subjected to oxidation were observed to act as molecular binding sites at the lipid-water interface. It is thus plausible that oxidized phospholipid species may elicit cellular level effects by modulating integration of various membrane-embedded and surface-associated proteins and peptides, whose conformational state, oligomerization, and functionality is known to be controlled by highly specific lipid-protein interactions and proper physical state of the membrane environment.
  • Durukan Tolvanen, Aysan (Helsingin yliopisto, 2014)
    Acute ischemic stroke is a devastating disease leaving more than half of its victims disabled and causing nearly 5% of all deaths worldwide. In large ischemic strokes, a major cause of death is brain edema, which follows blood-brain barrier (BBB) leakage. The BBB ensures brain homeostasis in health and disease by limiting the entry of harmful blood-borne substances into the brain parenchyma. With a leaky BBB, the brain becomes devoid of protection from detrimental components of the circulating blood. The BBB leakage in animal models of ischemia reperfusion has long been considered to be biphasic; however, a considerable amount of discrepancies exist among the studies. Knowing exact temporal changes of the BBB permeability (BBBP) is important for the management of stroke patients. When the BBB is open, BBBP alleviating therapies would be effective, neuroprotective or neurorestorative drugs would be introduced, and if the BBB is closed these drugs would not enter the brain. Practical and reliable biomarkers of BBBP status are needed. Stanniocalcins (STCs) are widely expressed in the brain and STC-1 expression is elevated in pathologies, such as hypoxia and focal ischemia. Recent data suggest a neuroprotective role for STC-1 especially trough hypoxic preconditioning (HPC). No previous data associate STC-1 and the BBB. We systematically evaluated disruption of the BBB following ischemia-reperfusion in a rat model of transient focal ischemia via suture occlusion of the middle cerebral artery for 90 min. Firstly (I, II), animals were allocated to 15 groups after reperfusion (25 min to 5 weeks). Secondly (III), a group of animals were evaluated repeatedly from 2 h to 1 week after reperfusion. BBBP to both small (gadolinium) (I, II, III), and large (Evans blue) (I) molecules were quantified by magnetic resonance imaging and fluorescence, respectively. Lastly, the contribution of STC-1 to HPC and the BBB was explored using STC-1 deficient mice (STC-/-). (I, II, III) After transient ischemia, the BBB leakage was continuous. Leakage to Evans Blue persisted up to 3 weeks and to gadolinium up to 5 weeks. Evans blue leakage slightly decreased at 36 and 72 h, gadolinium leakage was lesser at 25 min, 3 and 4 weeks. (IV) In STC-/- mice, HPC was effective in reducing lesion size, but these mice scored worse than wild type littermates. BBBP to Evans blue was not increased in STC-/- mice; neither under normal conditions, nor after hypoxia. To conclude, transient focal ischemia in rats triggers a continuous BBB leakage lasting for several weeks. Until the final closure of the BBB, no earlier transient closure occurs. This finding indicates a long therapeutic window opportunity in respect to BBB passage of drugs to treat stroke. BBBP imaging method used in these studies may be easily translated to clinics. STC-1 is not obligatory for hypoxic preconditioning and is not a determining component of the BBB. Yet, STC-1 is important for preservation of neurological function after transient ischemia.
  • Lorenzetti, Fulvio (Helsingin yliopisto, 2001)
  • Jauhiainen, Tiina (Helsingin yliopisto, 2007)
    The purpose of the present study was to evaluate the effects of Lactobacillus helveticus fermented milk (peptide milk) containing the casein-derived tripeptides Isoleucyl-prolyl-proline (Ile-Pro-Pro) and Valyl-prolyl-proline (Val-Pro-Pro) on blood pressure and vascular function in hypertensive subjects. The peptide milk lowered systolic and diastolic blood pressure in long-term use in hypertensive subjects when blood pressure was measured by using 24-hour ambulatory blood pressure measurement (ABPM). The blood pressure lowering effect was seen with the dose of 50 mg of tripeptides, and a tendency for lowering blood pressure was also observed when the dose was 5 mg. No adverse effects compared to the placebo group were reported or detected in laboratory analysis. The effect of the peptide milk on arterial stiffness was shown using two different methods, the ambulatory arterial stiffness index (AASI) and pulse wave analysis (PWA). According to the AASI, arterial stiffness was significantly reduced in the peptide milk group compared to the baseline level, but the difference was not significant compared to the placebo group. PWA showed that the peptide milk reduced arterial stiffness significantly compared to the placebo group. Endothelium-independent relaxation (nitroglycerin) and endothelium-dependent relaxation (salbutamol) did not differ between the groups. The blood pressure lowering mechanisms of the tripeptides and the kinetics of Ile-Pro-Pro were investigated using spontaneously hypertensive rats (SHR) and Sprague-Dawley rats. Previous studies have suggested that the blood pressure lowering effect of the tripeptides Ile-Pro-Pro and Val-Pro-Pro is based on angiotensin-converting enzyme inhibition, but the present findings did not agree with these previous studies. It was shown in SHR that calcium, potassium and magnesium may also have an important role in attenuating the development of hypertension as part of the peptide milk effect. In addition, the present study suggests indirectly that improved endothelial nitric oxide release capacity is not the mechanism by which peptide milk mediates its favourable circulatory effects. The kinetics of Ile-Pro-Pro were studied using adult Sprague-Dawley rats. The results showed that orally administered Ile-Pro-Pro is absorbed at least partly intact from the gastrointestinal tract. Radiolabelled Ile-Pro-Pro was distributed in different tissues and considerable radioactivity levels were found in tissues related to the renin-angiotensin system (RAS), adrenals, aorta and kidneys. Ile-Pro-Pro does not bind to plasma proteins, and therefore it is possible that its blood pressure lowering effect is mediated by free Ile-Pro-Pro. In conclusion, consumption of the peptide milk lowers blood pressure and reduces arterial stiffness in hypertensive subjects. Ile-Pro-Pro can be absorbed partly intact from the gastrointestinal tract and might accumulate in tissues related to the RAS. The precise blood pressure lowering mechanism of peptide milk remains to be studied.
  • Lahti-Koski, Marjaana (Helsingin yliopisto, 2001)
  • Kilpinen-Loisa, Päivi (Helsingin yliopisto, 2010)
    Osteoporosis is not only a disease of the elderly, but is increasingly diagnosed in chronically ill children. Children with severe motor disabilities, such as cerebral palsy (CP), have many risk factors for osteoporosis. Adults with intellectual disability (ID) are also prone to low bone mineral density (BMD) and increased fractures. This study was carried out to identify risk factors for low BMD and osteoporosis in children with severe motor disability and in adults with ID. In this study 59 children with severe motor disability, ranging in age from 5 to 16 years were evaluated. Lumbar spine BMD was measured with dual-energy x-ray absorptiometry. BMD values were corrected for bone size by calculating bone mineral apparent density (BMAD), and for bone age. The values were transformed into Z-scores by comparison with normative data. Spinal radiographs were assessed for vertebral morphology. Blood samples were obtained for biochemical parameters. Parents were requested to keep a food diary for three days. The median daily energy and nutrient intakes were calculated. Fractures were common; 17% of the children had sustained peripheral fractures and 25% had compression fractures. BMD was low in children; the median spinal BMAD Z-score was -1.0 (range -5.0 – +2.0) and the BMAD Z-score <-2.0 in 20% of the children. Low BMAD Z-score and hypercalciuria were significant risk factors for fractures. In children with motor disability, calcium intakes were sufficient, while total energy and vitamin D intakes were not. In the vitamin D intervention studies, 44 children and adolescents with severe motor disability and 138 adults with ID were studied. After baseline blood samples, the children were divided into two groups; those in the treatment group received 1000 IU peroral vitamin D3 five days a week for 10 weeks, and subjects in the control group continued with their normal diet. Adults with ID were allocated to receive either 800 IU peroral vitamin D3 daily for six months or a single intramuscular injection of 150 000 IU D3. Blood samples were obtained at baseline and after treatment. Serum concentrations of 25-OH-vitamin D (S-25-OHD) were low in all subgroups before vitamin D intervention: in almost 60% of children and in 77% of adults the S-25-OHD concentration was below 50 nmol/L, indicating vitamin D insufficiency. After vitamin D intervention, 19% of children and 42% adults who received vitamin D perorally and 12% of adults who received vitamin D intramuscularly had optimal S-25-OHD (>80 nmol/L). This study demonstrated that low BMD and peripheral and spinal fractures are common in children with severe motor disabilities. Vitamin D status was suboptimal in the majority of children with motor disability and adults with ID. Vitamin D insufficiency can be corrected with vitamin D supplements; the peroral dose should be at least 800 IU per day.
  • Valta, Helena (Helsingin yliopisto, 2009)
    Osteoporosis is a skeletal disorder characterized by compromised bone strength that predisposes to increased fracture risk. Childhood and adolescence are critical periods for bone mass gain. Peak bone mass is mostly acquired by the age of 18 years and is an important determinant of adult bone health and lifetime risk for fractures. Medications, especially glucocorticoids (GCs), chronic inflammation, decreased physical activity, hormonal deficiencies, delayed puberty, and poor nutrition may predispose children and adolescents with a chronic disease to impaired bone health. In this work, we studied overall bone health, the incidence and prevalence of fractures in children and adolescents who were treated for juvenile idiopathic arthritis (JIA) or had undergone solid organ transplantation. The first study cohort included 62 patients diagnosed with JIA and treated with GCs. The epidemiology of fractures after transplantation was investigated in 196 patients and a more detailed analysis of bone health determinants was performed on 40 liver (LTx) and 106 renal (RTx) transplantation patients. Bone mineral density (BMD) and vertebral morphology were assessed by dual-energy x-ray absorptiometry. Standard radiographs were obtained to detect vertebral fractures and to determine bone age; BMD values were adjusted for skeletal maturity. Our study showed that median BMD values were subnormal in all patient cohorts. The values were highest in patients with JIA and lowest in patients with LTx. Age at transplantation influenced BMD values in LTx but not RTx patients; BMD values were higher in patients who had LTx before the age of two years. BMD was lowest during the immediate posttransplantation years and increased subnormally during puberty. Delayed skeletal maturation was common in all patient groups. The prevalence of vertebral fractures ranged from 10% to 19% in the cohorts. Most of the fractures were asymptomatic and diagnosed only at screening. Vertebral fractures were most common in LTx patients. Vitamin D deficiency was common in all patient groups, and only 3% of patients with JIA and 25% of transplantation patients were considered to have adequate serum vitamin D levels. The total cumulative weight-adjusted dose of GC was not associated with BMD values in JIA or LTx patients. The combination of female gender and age over 15 years, parathyroid hormone concentration over 100 ng/L, and cumulative weight-adjusted methylprednisolone dose over 150 mg/kg during the three preceding years were found to be important predictors for low lumbar spine BMD in RTx patients. Based on the high prevalence of osteoporosis in the study cohorts more efforts should be put to prevention and early diagnosis of osteoporosis in these pediatric patients.
  • Lauttia, Susanna (Helsingin yliopisto, 2014)
    Tumor is more than a mass of transformed cells. Tumor cells are embedded in the supporting tumor stroma, that is composed of cellular and non-cellular components. The cellular components include vascular cells, bone marrow-derived cells (BMDCs), and fibroblasts. In concert, tumor cells and stromal cells secrete proteases, cytokines, chemokines, and growth factors that modulate the tumor behavior. The mechanisms how stromal cells contribute to the tumor growth and tumor angiogenesis are still incompletely understood. This thesis aimed to study the role of BMDCs in angiogenesis and cancer. First, the role of BMDCs as a source of vascular endothelium was examined. The second study addressed the effects of an EGFR inhibitor, gefitinib, on the vascular cells and BMDCs in an EGFR-deficient B16 mouse melanoma tumor model. In the third study, chemokine-like prokineticins, that may have both angiogenic and immunomodulatory properties, were examined in a virus-associated human skin cancer type, Merkel cell carcinoma (MCC). BMDCs did not incorporate into the vascular endothelium in any of the experimental models, suggesting that the pre-existing vasculature is the main source of endothelium during angiogenesis. The potential of the BMDCs to differentiate into vascular endothelium may require specific molecular microenvironments, which needs further analysis. Treatment of B16 tumors with the EGFR inhibitor gefitinib reduced the pericyte number and coverage in the small CD31+ capillaries and the numbers of perivascular BMDCs suggesting that gefitinib treatment might have vascular and stromal effects in some tumors. The identity of the perivascular BMDCs that responded to treatment is not currently known, but the intimate perivascular location of the affected cells proposes that these cells might contribute to tumor angiogenesis via paracrine mechanisms. In MCC, higher than median tumor prokineticin-2 (PROK2) mRNA content was strongly associated with the presence of Merkel cell polyomavirus (MCPyV) DNA, viral large T antigen expression, higher than median number of tumor infiltrating CD68+ and CD163+ macrophages, and with favorable survival. The presence of prokineticin-1 (PROK1) mRNA in the tumor was associated with absence of MCPyV DNA and tended to be associated with poor survival. Neither PROK1 nor PROK2 mRNA content was associated with the tumor microvascular density. Taken together, these findings support the immunomodulatory role of prokineticins and suggest that prokineticins are involved in mediating the immune response in MCC, but their role in tumor angiogenesis in MCC requires further evaluation.
  • Kananen, Kristiina (Helsingin yliopisto, 2005)
  • Sormaala, Markus (Helsingin yliopisto, 2006)
    Bone stress injuries of the foot have been known for more than 150 years. For a century, their primary diagnostic imaging tool has been radiography. However, currently the golden standard for establishing the diagnosis of stress injuries is magnetic resonance imaging (MRI). Although the injury type has been fairly well documented in the earlier literature, little information is available on the healing of stress injuries located in e.g. the talus and calcaneus. The current study retrospectively evaluated the stress injuries of the foot and ankle treated at the Central Military Hospital over a period of eight years in patients who underwent MRI for stress injury of the foot. The imaging studies of the patients were reevaluated to determine the exact nature of the stress injury. Moreover, the hospital records of the patients were reviewed to determine the healing of stress injuries of the talus and calcaneus. Patients with a stress fracture in the talus were recalled for a follow-up examination and MRI scan one to six years after the initial injury to determine if the fracture had completely healed, clinically and radiologically. The bone stress injuries of the foot were found to affect more than one bone in a majority of the cases. The talus and the calcaneus were the bones most commonly affected. In the talus, the most common site for the injuries was the head of the bone, and in the calcaneus, the posterior part of the bone. The injuries in these bones were associated with injuries in the surrounding bones. Stress injuries in the calcaneus seemed to heal well. No complications were seen in the primary healing process. The patients were, however, sometimes compelled to refrain from physical training for up to months. In the talus, minor degenerative findings of the articular surface were seen in half of the patients who participated in a follow-up MRI scan and radiographs taken one to six years after the initial injury. Half of the patients also reported minor exercise related symptoms in the follow-up. The symptoms were, however, not noticeable in everyday life.