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  • Wang, Feng (Helsingin yliopisto, 2013)
    Hydrophobic steroid hormone derivatives, carried in plasma exclusively by lipoproteins, constitute a unique hormone family. They are hormonally inactive and must be liberated from their fatty acyl partner before their physiological response. 17β-estradiol (E2) fatty acyl esters are reportedly enriched in adipose tissue, which has the enzyme machinery necessary for synthesis of estrogen from precursor steroids. The aims of the thesis were to explore the enzymes responsible for steroid-ester hydrolysis and to quantify dehydroepiandrosterone (DHEA) and E2 fatty acyl esters in adipose tissue and serum both in men and in women. [3H]DHEA-ester hydrolysis was investigated by use of lysosomal acid lipase (LAL)-depleted HeLa cells and Wolman fibroblasts. LDL-associated [3H]DHEA fatty acyl esters were partially taken up in cells via LDL receptor or LDL receptor-related receptors, after which, they were hydrolyzed and further metabolized to [3H]-5α-androstanedione and [3H]androstenedione that were secreted by the cells. LAL displayed a partial role in the hydrolysis of [3H]DHEA fatty acyl esters. DHEA fatty acyl esters were undetectable in adipose tissue, although 32 to 178 pmol/g of free DHEA was quantified. DHEA fatty acyl ester concentrations in serum ranged from 0.5 to 2.8 pmol/ml, corresponding to 5 to 15% of total DHEA. E2 fatty acyl esters in subcutaneous adipose tissue from the human breast may function as a reservoir for free E2, and conversion process to free E2 was partly dependent on hormone-sensitive lipase (HSL) function. In obese men, E2 fatty acyl ester concentrations in adipose tissues were similar to those of E2, but in obese women ester derivatives were significantly lower than E2. In obese subjects, E2 fatty acyl ester levels in adipose tissue significantly correlated with serum levels whereas in case of E2 this was not the case. Compared to obese men, E2 levels in subcutaneous adipose tissue in obese women were higher, along with higher relative mRNA expression of steroid sulfatase and 17ß-hydroxysteroid dehydrogenases 1, 7, and 12. In summary, DHEA fatty acyl ester concentrations in serum were attributable to 5 to 15 % of total DHEA. Circulating DHEA fatty acyl esters bound to LDL may enter target cells, and after their hydrolysis step to free DHEA it can be metabolized to other active steroids. LAL may partially participate in the hydrolysis of DHEA esters. In addition, subcutaneous adipose tissue from human breast had E2 fatty acyl esterifying and hydrolysing activity, and the hydrolysis was in part dependent on HSL activity. Production of E2 from a large adipose mass was not reflected by increased circulating E2 concentrations in severely obese men or women. Adipose tissue may, however, contribute to serum E2 fatty acyl ester concentrations.
  • 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.
  • Louhimo, Riku (Helsingin yliopisto, 2015)
    Cancer is one of the leading causes of death in industrialized nations and its incidence is steadily increasing due to population aging. Cancer constitutes a group of diseases characterized by unwanted cellular growth which results from random genomic alterations and environmental exposure. Diverse genomic and epigenomic alterations separately and jointly regulate gene expression and stimulate and support neoplastic growth. More effective treatment, earlier and more accurate diagnosis, and improved management of cancer are important for public health and well-being. Technological improvements in data measurement, storing and transport capability are transforming cancer research to a data-intensive field. The large increases in the quality and quantity of data for the analysis and interpretation of experiments has made employing computational and statistical tools necessary. Data integration - the combination of different types of measurement data - is a valuable computational tool for cancer research because data integration improves the interpretability of data-driven analytics and can thereby provide novel prognostic markers and drug targets. I have developed two computational data integration tools for large-scale genomic data and a simulator framework for testing a specific type of data integration algorithm. The first computational method, CNAmet, enhances the interpretation of genomic analysis results by integrating three data levels: gene expression, copy-number alteration, and DNA methylation. The second computational method, GOPredict, uses a knowledge discovery approach to prioritize drugs for patient cohorts thereby stratifying patients into potentitally drug-sensitive subgroups. Using the simulator framework, we are able to compare the performance of integration algorithms which integrate gene copy-number data with gene expression data to find putative cancer genes. Our experimental results indicate in simulated, cell line, and primary tumor data that well-performing integration algorithms for gene copy-number and expression data use and process genomic data appropriately. Applying these methods to diffuse large B-cell lymphoma, integrative analysis of copy-number and expression data helps to uncover a gene with putative prognostic utility. Furthermore, analysis of glioblastoma brain cancer data with CNAmet suggests that a number of known cancer genes, including the epidermal growth factor receptor, are highly expressed due to co-occuring alterations in their promoter DNA methylation and copy-number. Finally, integration of publicly available molecular and literature data with GOPredict suggests that treating patients with FGFR inhibitors in breast cancer and CDK inhibitors in ovarian cancer could support standard drug therapies. Collectively, the methods developed here and their application to varied molecular cancer data sets illustrates the benefits of data integration in cancer genomics.
  • 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.
  • Markula-Patjas, Kati (Helsingin yliopisto, 2015)
    Children with juvenile idiopathic arthritis (JIA) are predisposed to compromised bone health and alterations in body composition because of chronic inflammation, nutritional and hormonal disturbances, limited physical activity and glucocorticoid (GC) therapy. Compromised bone health may present as pathological vertebral compression fractures, but data on their prevalence and risk factors in children are limited. Excess fat, and especially adipose tissue-derived adipokines leptin and adiponectin, may also contribute to impaired bone health. Furthermore, adipokines modulate immunity and inflammation in adults with rheumatic diseases, but their role in JIA has not been explored. We evaluated bone health in patients with severe JIA and investigated body composition and adipokines and their contribution to bone health and disease activity in JIA. We recruited two cohorts of patients for cross-sectional studies. The Severe JIA Cohort comprised 50 patients with severe polyarticular or systemic JIA. The GC-treated Cohort included 50 patients with JIA with mostly mild to moderate disease severity and at least three months' exposure to systemic GC. The results were compared with those of sex-and age-matched healthy controls. The study protocol included clinical and laboratory assessments, evaluation of bone mineral density (BMD) and body composition by dual-energy X-ray absorptiometry (DXA), spinal radiography and spinal magnetic resonance imaging (MRI). Spinal radiography showed vertebral compression fractures in 22% of the patients with severe JIA. Patients with fractures had higher weight-adjusted cumulative GC dose, higher disease activity and higher body mass index than those without fractures. Bone age-corrected BMD Z-scores for lumbar spine and whole body were similar between those with and without fractures. On spinal MRI, altogether 28% of patients with severe JIA showed vertebral fractures and several other vertebral changes, including end plate irregularities in 26%, anterior vertebral corner lesions in 16% and disc changes in 46%. Based on concentrations of bone turnover markers, the patients with severe JIA had increased bone resorption, but normal bone formation. Further, patients with severe JIA had increased body adiposity, and their serum leptin was increased even independently of fat mass. Leptin showed an inverse association with bone turnover markers in patients, while in controls the association was dependent on fat mass. In the GC-treated Cohort, fat mass, lean mass and serum leptin and adiponectin were similar to those of controls, but patients had slightly lower BMD values than controls. Those patients with lumbar spine BMD Z-score -1.0 tended to have higher serum leptin values than those with higher BMD Z-scores, but in regression analysis leptin was not associated with BMD. Adipokines did not correlate with current disease activity in either patient cohort. Patients with severe JIA have compromised bone health based on high prevalence of compression fractures. Risk factors include high GC exposure, high disease activity and high body mass index. BMD, as measured by DXA, is unable to differentiate between those with and without compression fractures. According to spinal MRI findings, patients with severe JIA have, besides compression fractures, several other changes involving intervertebral discs and vertebral end plates; the clinical relevance of these remains uncertain. Patients with severe JIA are prone to high adiposity, whereas those with less severe disease have normal body composition despite previous GC exposure. Leptin may negatively contribute to bone metabolism in severe JIA, but larger and longitudinal studies are needed to prove causality and to evaluate whether these preliminary findings are generalizable to other JIA groups. We did not observe a correlation between leptin or adiponectin and disease activity in either JIA cohort. The possible role of adipokines as a modulator of immunity and inflammation in JIA remains to be evaluated.
  • 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.