Semanttisen webin ontologisen tekstiilikäsitteistön kehittäminen ja liittäminen museoiden luettelointitietoihin

DEVELOPING A TEXTILE ONTOLOGY FOR THE SEMANTIC WEB AND CONNECTING IT TO MUSEUM CATALOGING DATA The goal of the Semantic Web is to share concept-based information in a versatile way on the Internet. This is achievable using formal data structures called ontologies. The goal of this re-search is to increase the usability of museum cataloging data in information retrieval. The work is interdisciplinary, involving craft science, terminology science, computer science, and museology. In the first part of the dissertation an ontology of concepts of textiles, garments, and accessories is developed for museum cataloging work. The ontology work was done with the help of thesauri, vocabularies, research reports, and standards. The basis of the ontology development was the Museoalan asiasanasto MASA, a thesaurus for museum cataloging work which has been enriched by other vocabularies. Concepts and terms concerning the research object, as well as the material names of textiles, costumes, and accessories, were focused on. The research method was terminological concept analysis complemented by an ontological view of the Semantic Web. The concept structure was based on the hierarchical generic relation. Attention was also paid to other relations between terms and concepts, and between concepts themselves. Altogether 977 concept classes were created. Issues including how to choose and name concepts for the ontology hierarchy and how deep and broad the hierarchy could be are discussed from the viewpoint of the ontology developer and museum cataloger. The second part of the dissertation analyzes why some of the cataloged terms did not match with the developed textile ontology. This problem is significant because it prevents automatic ontological content integration of the cataloged data on the Semantic Web. The research datasets, i.e. the cataloged museum data on textile collections, came from three museums: Espoo City Museum, Lahti City Museum and The National Museum of Finland. The data included 1803 textile, costume, and accessory objects. Unmatched object and textile material names were analyzed. In the case of the object names six categories (475 cases), and of the material names eight categories (423 cases), were found where automatic annotation was not possible. The most common explanation was that the cataloged field was filled with a long sentence comprised of many terms. Sometimes in the compound term, the object name and material, or the name and the way of usage, were combined. As well, numeric values in the material name cataloging field prevented annotation and so did the absence of a corresponding concept in the ontology. Ready-made drop-down lists of materials used in one cataloging system facilitated the annotation. In the case of naming objects and materials, one should use terms in basic form without attributes. The developed textile ontology has been applied in two cultural portals, MuseumFinland and Culturesampo, where one can search for and browse information based on cataloged data using integrated ontologies in an interoperable way. The textile ontology is also part of the national FinnONTO ontology infrastructure. Keywords: annotation, concept, concept analysis, cataloging, museum collection, ontology, Semantic Web, textile collection, textile material

Verkkokurssilla : Opiskelijoiden kokemuksia toiminnasta yhteisöllisessä oppimisprosessissa Fle2-ympäristössä ja kokemusten näkyminen toimintaan osallistumisessa

The study examines one case of students' experiences from the activity in a collaborative learning process in a networked learning environment, and explores whether or not the experiences explain the participation or lack of participation in the activities. As a research task the students' experiences in the database of the networked learning environment, participating in its construction, and the ways of working needed to build the database, were examined. To contrast the students' experiences, their actual participation in the building of the database was clarified. Based on actual participation, groups more active and more passive than average were separated, and their experiences were compared to each other. The research material was collected from the course Cognitive and Creative Processes, which was offered to studentsof the Department of Textile Teacher Education in University of Helsinki, and students of the Departments of Teacher Education Units giving textile education in Turku, Rauma and Savonlinna in the beginning of 2001. In this course, creativity was examined from a psychological and sosiocultural context with the aim of realizing a collaborative progressive inquiry process. The course was held in a network-based Future Learning Environment (Fle 2) except for the starting lecture and training the use of the learning environment. This study analyzed the learning diaries that the students had sent to the tutor once a week for four weeks, and the final thoughts written into the database of the learning environment. Content analysis was applied as the research method. The case was enriched from another point of view by examining the messages the students had written into the learning environment with the social network analysis. The theoretical base of the study looks at the research of computer-supported collaborative learning, the conceptions of learning as a process of participation and knowledge building, and the possibilities and limitations of network-based learning environments. The research results show, that both using the network-based learning environment and collaborative ways of studying were new to the students. The students were positively surprised by the feedback and support provided by the community. On the other hand, they also experienced problems with facelessness and managing the information in the learning environment. The active students seemed to be more ready for a progressive inquiry process. It can be seen from their attitudes and actions that they have strived to participate actively and invested into the process both from their own and the community's point of view. The more passive students reported their actions to get credits and they had a harder time of perceiving the thoughts presented in the net as common progression. When arranging similar courses in the future, attention should be paid to how to get the students to act in ways necessary for knowledge building, and different from more traditional ways of studying. The difficulties of students used to traditional studying methods to adapt to collaborative knowledge building were evident on the course Cognitive and Creative Processes. Keywords: computer supported collaborative learning, knowledge building, progressive Inquiry, participation

Of tolerance in mice and men : studies in APECED and Aire

Autoimmune diseases affect 5 % of the population and come in many forms, such as diabetes, rheumatoid arthritis and MS. However, how and why autoimmune diseases arise are not yet fully resolved. In this thesis, the onset of autoimmunity was investigated using both patient samples and a mouse model of autoimmunity. Autoimmune diseases are usually complex, due to a number of different causative genes and environmental factors. However, a few monogenic autoimmune diseases have been described, which are caused by mutations in only one gene per disease. One of such disease is called APECED (autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy) and is enriched in the Finnish population. The causative gene behind APECED is named AIRE from AutoImmune REgulator. How malfunction of just one gene product can cause the multitude of disease components found in APECED is not yet resolved. This thesis sought out to find out more about the functions of AIRE, in order to reveal why APECED and other autoimmune diseases arise and what goes wrong? Usually, immune cells are taught to distinguish between self and non-self during their development. That way, immune cells can fight off bacteria and microbes while leaving the tissues and organs of the host organism itself unharmed. In APECED, the development of immune cells called αβ T cells is incomplete. The cells are not able to fully distinguish between self and non-self. This leads to autodestruction of self tissues and autoimmune disease. One of the achievements of this thesis was the finding that the development of another set of T cells called γδ T cells is not affected by AIRE in mice or in men. Instead, we found that another type of immune cell important in tolerance, called the dendritic cell is defective in APECED patients and is not able to respond to microbial stimulus in a normal fashion. Finally, we studied Aire-deficient mice and found that autoantibodies expressed in the mice were not targeted against the same molecules as those found in APECED patients. This indicates differences in the autoimmune pathology in mice and men. More work is still required before we understand the mechanisms of tolerance and autoimmunity well enough to be able to cure APECED, let alone the more complex autoimmune diseases. Yet altogether, the findings of this thesis work bring us one step closer to finding out why and how APECED and common autoimmune diseases arise.

Development of regulatory T cells in the human thymus

The role of the immune system is to protect an organism against pathogens while maintaining tolerance against self. T cells are an essential component of the immune system and they develop in the thymus. The AIRE (autoimmune regulator) gene product plays an important role in T cell development, as it promotes expression of peripheral tissue antigens in the thymus. Developing T cells, thymocytes, which recognize self-antigens with high affinity are deleted. However, this deletion process is not perfect and not all autoreactive T cells are destroyed. When the distinction between self and non-self fails, tolerance breaks and the immune system attacks the host s own tissues. This results in autoimmunity. Regulatory T cells contribute to the maintenance of self-tolerance. They can actively suppress the function of autoreactive cells. Several populations of cells with regulatory properties have been described, but the best characterized population is the natural regulatory T cells (Treg cells), which develop in the thymus and express the transcription factor FOXP3. The thymic development of Treg cells in humans is the subject of this thesis. Thymocytes at different developmental stages were analyzed using flow cytometry. The CD4-CD8- double-negative (DN) thymocytes are the earliest T cell precursors in the T cell lineage. My results show that the Treg cell marker FOXP3 is up-regulated already in a subset of these DN thymocytes. FOXP3+ cells were also found among the more mature CD4+CD8+ double-positive (DP) cells and among the CD4+ and CD8+ single-positive (SP) thymocytes. The different developmental stages of the FOXP3+ thymocytes were isolated and their gene expression examined by quantitative PCR. T cell receptor (TCR) repertoire analysis was used to compare these different thymocyte populations. My data show that in humans commitment to the Treg cell lineage is an early event and suggest that the development of Treg cells follows a linear developmental pathway, FOXP3+ DN precursors evolving through the DP stage to become mature CD4+ Treg cells. Most T cells have only one kind of TCR on their cell surface, but a small fraction of cells expresses two different TCRs. My results show that the expression of two different TCRs is enriched among Treg cells. Furthermore, both receptors were capable of transmitting signals when bound by a ligand. By extrapolating flow cytometric data, it was estimated that the majority of peripheral blood Treg cells are indeed dual-specific. The high frequency of dual-specific cells among human Treg cells suggests that dual-specificity has a role in directing these cells to the Treg cell lineage. It is known that both genetic predisposition and environmental factors influence the development of autoimmunity. It is also known that the dysfunction or absence of Treg cells leads to the development of autoimmune manifestations. APECED (autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy) is a rare monogenic autoimmune disease, caused by mutations in the AIRE gene. In the absence of AIRE gene product, deletion of self-specific T cells is presumably disturbed and autoreactive T cells escape to the periphery. I examined whether Treg cells are also affected in APECED. I found that the frequency of FOXP3+ Treg cells and the level of FOXP3 expression were significantly lower in APECED patients than in controls. Additionally, when studied in cell cultures, the suppressive capacity of the patients' Treg cells was impaired. Additionally, repertoire analysis showed that the TCR repertoire of Treg cells was altered. These results suggest that AIRE contributes to the development of Treg cells in humans and the selection of Treg cells is impaired in APECED patients. In conclusion, my thesis elucidates the developmental pathway of Treg cells in humans. The differentiation of Tregs begins early during thymic development and both the cells dual-specificity and AIRE probably affect the final commitment of Treg cells.

Genetic Basis of Pituitary Adenoma Predisposition

Much of the global cancer research is focused on the most prevalent tumors; yet, less common tumor types warrant investigation, since A rare disorder is not necessarily an unimportant one . The present work discusses a rare tumor type, the benign adenomas of the pituitary gland, and presents the advances which, during the course of this thesis work, contributed to the elucidation of a fraction of their genetic background. Pituitary adenomas are benign neoplasms of the anterior pituitary lobe, accounting for approximately 15% of all intracranial tumors. Pituitary adenoma cells hypersecrete the hormones normally produced by the anterior pituitary tissue, such as growth hormone (GH) and prolactin (PRL). Despite their non-metastasizing nature, these adenomas can cause significant morbidity and have to be adequately treated; otherwise, they can compromise the patient s quality of life, due to conditions provoked by hormonal hypersecretion, such as acromegaly in the case of GH-secreting adenomas, or due to compressive effects to surrounding tissues. The vast majority of pituitary adenomas arise sporadically, whereas a small subset occur as component of familial endocrine-related tumor syndromes, such as Multiple Endocrine Neoplasia type 1 (MEN1) and Carney complex (CNC). MEN1 is caused by germline mutations in the MEN1 tumor suppressor gene (11q13), whereas the majority of CNC cases carry germline mutations in the PRKAR1A gene (17q24). Pituitary adenomas are also encountered in familial settings outside the context of MEN1 and CNC, but unlike in the latter syndromes, their genetic background until recently remained elusive. Evidence in previous literature supported the notion that a tumor suppressor gene on 11q13, residing very close to but still distinct from MEN1, causes genetic susceptibility to pituitary tumors. The aim of the study was to identify the genetic cause of a low penetrance form of Pituitary Adenoma Predisposition (PAP) in families from Northern Finland. The present work describes the methodological approach that led to the identification of aryl hydrocarbon receptor interacting protein (AIP) as the gene causing PAP. Combining chip-based technologies (SNP and gene expression arrays) with traditional gene mapping methods and genealogy data, we showed that germline AIP mutations cause PAP in familial and sporadic settings. PAP patients were diagnosed with mostly adenomas of the GH/PRL-secreting cell lineage. In Finland, two AIP mutations accounted for 16% of all patients diagnosed with GH-secreting adenomas, and for 40% of patients being younger than 35 years of age at diagnosis. AIP is suggested to act as a tumor suppressor gene, a notion supported by the nature of the identified mutations (most are truncating) and the biallelic inactivation of AIP in the tumors studied. AIP has been best characterized as a cytoplasmic interaction partner of aryl hydrocarbon receptor (AHR), also known as dioxin receptor, but it has other partners as well. The mechanisms that underlie AIP-mediated pituitary tumorigenesis are to date largely unknown and warrant further investigation. Because AIP was identified in the genetically homogeneous Finnish population, it was relevant to examine its contribution to PAP in other, more heterogeneous, populations. Analysis of pituitary adenoma patient series of various ethnic origins and differing clinical settings revealed germline AIP mutations in all cohorts studied, albeit with low frequencies (range 0.8-7.4%). Overall, PAP patients were typically diagnosed at a young age (range 8-41 years), mainly with GH-secreting adenomas, without strong family history of endocrine disease. Because many PAP patients did not display family history of pituitary adenomas, detection of the condition appeared challenging. AIP immunohistochemistry was tested as a molecular pre-screening tool on mutation-positive versus mutation-negative tumors, and proved to be a potentially useful predictor of PAP. Mutation screening of a large cohort of colorectal, breast, and prostate tumors did not reveal somatic AIP mutations. These tumors, apart from being the most prevalent among men and women worldwide, have been associated with acromegaly, particularly colorectal neoplasia. In this material, AIP did not appear to contribute to the pathogenesis of these common tumor types and other genes seem likely to play a role in such tumorigenesis. Finally, the contribution of AIP in pediatric onset pituitary adenomas was examined in a unique population-based cohort of sporadic pituitary adenoma patients from Italy. Germline AIP mutations may account for a subset of pediatric onset GH-secreting adenomas (in this study one of seven GH-secreting adenoma cases or 14.3%), and appear to be enriched among young (≤25 years old) patients. In summary, this work reveals a novel tumor susceptibility gene, namely AIP, which causes genetic predisposition to pituitary adenomas, in particular GH-secreting adenomas. Moreover, it provides molecular tools for identification of individuals predisposed for PAP. Further elaborate studies addressing the functional role of AIP in normal and tumor cells will hopefully expand our knowledge on endocrine neoplasia and reveal novel cellular mechanisms of pituitary tumorigenesis, including potential drug targets.

Genetic studies on recurrent miscarriage

Recurrent miscarriage (RM) is defined as three consecutive pregnancy failures and is estimated to affect ~1% of couples trying to conceive. The cause of RM remains unknown in approximately 50% of cases. In this study, it was hypothesized that some of the underlying factors yet to be discovered are genetic. The aim was to search for mutations in genes AMN, EPCR, TM, and p53 known to cause miscarriage in mouse models and thereby find new genetic causes for unexplained miscarriages in humans. In addition, the mitochondrial genome was studied because mitochondria are involved in processes important in early development. Furthermore, sex chromosome characteristics suggested to underlie miscarriage were also studied. A total of 40 couples and 8 women with unexplained RM were collected for this study and screened for mutations in the candidate genes. Six interesting exonic or potential splice site disrupting variations were detected. However, their phenotypic effects cannot be determined without further investigations. Additionally, an association between the C11992A polymorphism of the p53 gene and RM was detected. The results indicate that women carrying the C/A or A/A genotype have a two-fold higher risk for RM than women with a C/C genotype. This strengthens the results of previous studies reporting that p53 sequence variations may cause miscarriage. The role of variation C11992A in embryonic development is, however, difficult to predict without further studies When screening the mitochondrial genome a heteroplasmic mtDNA variation was found in an unexpected high number of women, as heteroplasmic variations are reported to be rare. One novel variation and 18 previously reported polymorphisms were detected in the mitochondrial genome. Although the detected variations are likely to be neutral polymorphisms, a role in the aetiology of miscarriage cannot be excluded as some mtDNA variations may be pathogenic only when a threshold is reached. Recent publications have reported skewed X chromosome inactivation and Y chromosome microdeletions to be associated with RM. Therefore, these sex chromosome abnormalities in the context of RM were investigated. No associations between skewed X chromosome inactivation or Y chromosome microdeletions and RM in the Finnish patients were detected. Data on ancestral birthplaces of the patients were collected to study any possible geographic clustering, which would indicate a common predisposing factor. The results showed clustering of the birthplaces in eastern Finland in a subset of patients. This suggests a possibility of an enriched susceptibility gene which may contribute to RM.

Animal model and molecular interactions of Cln5

Neuronal ceroid lipofuscinoses (NCLs) are a family of inherited pediatric neurodegenerative disorders, leading to retinal degeneration, death of selective neuronal populations and accumulation of autofluorscent ceroid-lipopigments. The clinical manifestations are generally similar in all forms. The Finnish variant late infantile neuronal ceroid lipofuscinosis (vLINCLFin) is a form of NCL, especially enriched in the Finnish population. The aim of this thesis was to analyse the brain pathology of vLINCLFin utilising the novel Cln5-/- mouse model. Gene expression profiling of the brains of already symptomatic Cln5-/- mice revealed that inflammation, neurodegeneration and defects in myelinization are the major characteristics of the later stages of the disease. Histological characterization of the brain pathology confirmed that the thalamocortical system is affected in Cln5-/- mice, similarly to the other NCL mouse models. However, whereas the brain pathology in all other analyzed NCL mice initiate in the thalamus and spread only months later to the cortex, we observed that the sequence of events is uniquely reversed in Cln5-/- mice; beginning in the cortex and spreading to the thalamus only months later. We could also show that even though neurodegeneration is inititated in the cortex, reactive gliosis and loss of myelin are evident in specific nuclei of the thalamus already in the 1 month old brain. To obtain a deeper insight into the disturbed metabolic pathways, we performed gene expression profiling of presymptomatic mouse brains. We validated these findings with immunohistological analyses, and could show that cytoskeleton and myelin were affected in Cln5-/- mice. Comparison of gene expression profiling results of Cln5-/- and Cln1-/- mice, further highlighted that these two NCL models share a common defective pathway, leading to disturbances in the neuronal growth cone and cytoskeleton. Encouraged by the evidence of this defected pathway, we analyzed the molecular interactions of NCL-proteins and observed that Cln5 and Cln1/Ppt1 proteins interact with each other. Furthermore, we demonstrated that Cln5 and Cln1/Ppt1 share an interaction partner, the F1-ATP synthase, potentially linking both vLINCLFIN and INCL diseases to disturbed lipid metabolism. In addition, Cln5 was shown to interact with other NCL proteins; Cln2, Cln3, Cln6 and Cln8, implicating a central role for Cln5 in the NCL pathophysiology. This study is the first to describe the brain pathology and gene expression changes in the Cln5-/- mouse. Together the findings presented in this thesis represent novel information of the disease processes and the molecular mechanisms behind vLINCLFin and have highlighted that vLINCLFin forms a very important model to analyze the pathophysiology of NCL diseases.

Genetic Heterogeneity in Autism Spectrum Disorders in a Population Isolate

Positional cloning has enabled hypothesis-free, genome-wide scans for genetic factors contributing to disorders or traits. Traditionally linkage analysis has been used to identify regions of interest, followed by meticulous fine mapping and candidate gene screening using association methods and finally sequencing of regions of interest. More recently, genome-wide association analysis has enabled a more direct approach to identify specific genetic variants explaining a part of the variance of the phenotype of interest. Autism spectrum disorders (ASDs) are a group of childhood onset neuropsychiatric disorders with shared core symptoms but varying severity. Although a strong genetic component has been established in ASDs, genetic susceptibility factors have largely eluded characterization. Here, we have utilized modern molecular genetic methods combined with the advantages provided by the special population structure in Finland to identify genetic risk factors for ASDs. The results of this study show that numerous genetic risk factors exist for ASDs even within a population isolate. Stratification based on clinical phenotype resulted in encouraging results, as previously identified linkage to 3p14-p24 was replicated in an independent family set of families with Asperger syndrome, but no other ASDs. Fine-mapping of the previously identified linkage peak for ASDs at 3q25-q27 revealed association between autism and a subunit of the 5-hydroxytryptamine receptor 3C (HTR3C). We also used dense, genome-wide single nucleotide polymorphism (SNP) data to characterize the population structure of Finns. We observed significant population substructure which correlates with the known history of multiple consecutive bottle-necks experienced by the Finnish population. We used this information to ascertain a genetically homogenous subset of autism families to identify possible rare, enriched risk variants using genome-wide SNP data. No rare enriched genetic risk factors were identified in this dataset, although a subset of families could be genealogically linked to form two extended pedigrees. The lack of founder mutations in this isolated population suggests that the majority of genetic risk factors are rare, de novo mutations unique to individual nuclear families. The results of this study are consistent with others in the field. The underlying genetic architecture for this group of disorders appears highly heterogeneous, with common variants accounting for only a subset of genetic risk. The majority of identified risk factors have turned out to be exceedingly rare, and only explain a subset of the genetic risk in the general population in spite of their high penetrance within individual families. The results of this study, together with other results obtained in this field, indicate that family specific linkage, homozygosity mapping and resequencing efforts are needed to identify these rare genetic risk factors.

Molecular profiling of malignant pleural mesothelioma and pulmonary fibrosis

Malignant mesothelioma (MM) is a rare, usually incurable, disease mainly caused by former exposure to asbestos. Even though MM has a strong etiological link, genetic factors may play a role, since not all cases can be linked to former asbestos exposure. This thesis focuses on lung diseases, mainly malignant mesothelioma (MM), and idiopathic pulmonary fibrosis (IPF), which resembles asbestosis. The specific asbestos-related pathways associated with malignant as well as non-malignant lung diseases, still need to be clarified. Since most patients diagnosed with MM or asbestosis/fibrosis have a dismal prognosis and few therapeutic options are available, early diagnosis and better understanding of the disease pathogenesis are of the utmost importance. The first objective of this thesis was to identify asbestos specific differentially expressed genes. This was approached by using high-resolution gene expression arrays, and three different human lung cell lines, as well as with three different bioinformatics approaches. Since the first study aimed to elucidate potential early changes, the second study was used to screen DNA copy number changes in MM tumour samples. This was performed using genome wide microarrays for identification of DNA copy number changes characterstic for MM. Study III focused on the role of gremlin in the regulation of bone morphogenetic protein (BMPs) in IPF. Further studies were conducted in asbestos-exposed cell cultures as well as in an asbestos-induced mouse model. Furthermore, GATA-6 was studied in MM and metastatic pleural adenocarcinoma. The GATA transcription factors are important during embryonic development, but their role in cancer is still unclear. GATA-6 is a co-factor/target of thyroid transcription factor 1 (TTF-1), which is used in differential diagnostics of pleural MM and adenocarcinoma. Bioinformatics probed the genes and biological processes ordered in terms of significance, clusters, and highly enriched chromosomal regions. The study revealed several already identified targets, produced new ideas about genes which are central for asbestos exposure, as well as provided supplementary data for researchers to check their own novel findings or ideas. The analysis revealed DNA copy number changes characteristic for MM tumors. The most common regions of loss were detected in 1p, 3p, 6q, 9p, 13, 14, and 22, and gains at 17q. The histological features in asbestosis and IPF are very similar, wherefore IPF can be studied in asbestos models. The BMP antagonist gremlin was up-regulated by asbestos exposure in human epithelial cell lines, which was also observed in Study I. The transforming growth factor (TGF) -β and BMP expression and signaling activities were measured from murine and human fibrotic lungs. BMP-7 signaling was down-regulated in response to up-regulation of gremlin, and restoration of BMP-7 signaling prevented progression of fibrosis in mice. Therefore, the study suggests that the restoration of BMP-7 signaling in fibrotic lung could potentially aid in the treatment of IPF patients. Study IV revealed that GATA-6 was strongly expressed in the majority of the MM cases, and correlated statistically significant with longer survival in subgroups of MM.

Genomic and functional profiling of gastric cancer

Helicobacter pylori infection is a risk factor for gastric cancer, which is a major health issue worldwide. Gastric cancer has a poor prognosis due to the unnoticeable progression of the disease and surgery is the only available treatment in gastric cancer. Therefore, gastric cancer patients would greatly benefit from identifying biomarker genes that would improve diagnostic and prognostic prediction and provide targets for molecular therapies. DNA copy number amplifications are the hallmarks of cancers in various anatomical locations. Mechanisms of amplification predict that DNA double-strand breaks occur at the margins of the amplified region. The first objective of this thesis was to identify the genes that were differentially expressed in H. pylori infection as well as the transcription factors and signal transduction pathways that were associated with the gene expression changes. The second objective was to identify putative biomarker genes in gastric cancer with correlated expression and copy number, and the last objective was to characterize cancers based on DNA copy number amplifications. DNA microarrays, an in vitro model and real-time polymerase chain reaction were used to measure gene expression changes in H. pylori infected AGS cells. In order to identify the transcription factors and signal transduction pathways that were activated after H. pylori infection, gene expression profiling data from the H. pylori experiments and a bioinformatics approach accompanied by experimental validation were used. Genome-wide expression and copy number microarray analysis of clinical gastric cancer samples and immunohistochemistry on tissue microarray were used to identify putative gastric cancer genes. Data mining and machine learning techniques were applied to study amplifications in a cross-section of cancers. FOS and various stress response genes were regulated by H. pylori infection. H. pylori regulated genes were enriched in the chromosomal regions that are frequently changed in gastric cancer, suggesting that molecular pathways of gastric cancer and premalignant H. pylori infection that induces gastritis are interconnected. 16 transcription factors were identified as being associated with H. pylori infection induced changes in gene expression. NF-κB transcription factor and p50 and p65 subunits were verified using elecrophoretic mobility shift assays. ERBB2 and other genes located in 17q12- q21 were found to be up-regulated in association with copy number amplification in gastric cancer. Cancers with similar cell type and origin clustered together based on the genomic localization of the amplifications. Cancer genes and large genes were co-localized with amplified regions and fragile sites, telomeres, centromeres and light chromosome bands were enriched at the amplification boundaries. H. pylori activated transcription factors and signal transduction pathways function in cellular mechanisms that might be capable of promoting carcinogenesis of the stomach. Intestinal and diffuse type gastric cancers showed distinct molecular genetic profiles. Integration of gene expression and copy number microarray data allowed the identification of genes that might be involved in gastric carcinogenesis and have clinical relevance. Gene amplifications were demonstrated to be non-random genomic instabilities. Cell lineage, properties of precursor stem cells, tissue microenvironment and genomic map localization of specific oncogenes define the site specificity of DNA amplifications, whereas labile genomic features define the structures of amplicons. These conclusions suggest that the definition of genomic changes in cancer is based on the interplay between the cancer cell and the tumor microenvironment.

Selenium and its effects on growth, yield and tuber quality in potato

Selenium (Se) has been demonstrated to be an essential trace element for maintenance of animal and human health. Although it has not been confirmed to be an essential micronutrient in higher plants, there is increasing evidence that Se functions as an antioxidant in plants. Selenium has been shown to exert a beneficial effect on crop growth and promotes stress tolerance at low concentrations. However, the specific physiological mechanisms that underlie the positive effects of Se in plants have not been clearly elucidated. The aims of this study were to determine the Se concentration in potato (Solanum tuberosum L.) and the effects of Se on the accumulation of carbohydrates, growth and yield in potato plants. An additional aim was to study the impact of Se on the total glycoalkaloid concentration in immature potato tubers. The distribution of Se in different biochemical Se fractions and the effect of storage on the Se concentration were studied in Se-enriched tubers. Furthermore, the effect of Se on raw darkening and translocation of Se from seed tubers to the next tuber generation was investigated. Due to the established anti-ageing properties of Se, it was of interest to study if Se affects physiological age and growth vigour of seed tubers. The Se concentrations in the upper leaves, roots, stolons and tubers of potato increased with increasing Se supplementation. The highest Se concentration was reached in young upper leaves, roots and stolons, indicating that added selenate was efficiently utilized and taken up at an early stage. During the growing period the Se concentration declined in the aerial parts, roots and stolons of potato plants whereas an intensive accumulation took place in immature and mature tubers. Selenium increased carbohydrate accumulation in the young upper leaves and in stolons, roots and tubers at maturity. This could not be explained by increased production of photoassimilates as net photosynthesis did not differ among Se treatments. The Se treated plants produced higher tuber yields than control plants, and at the highest Se concentration (0.3 mg kg-1) lower numbers of larger tubers were harvested. Increased yield of Se treated plants suggested that Se may enhance the allocation of photoassimilates for tuber growth, acting as a strong sink for both Se and for carbohydrates. Similarly as for other plant species, the positive impact of Se on the yield of potato plants could be related to its antioxidative effect in delaying senescence. The highest Se supplementation (0.9 mg kg-1) slightly decreased the glycoalkaloid concentration of immature tubers. However, at this level the Se concentration in tubers was about 20 µg g-1 DW. A 100 g consumption of potato would provide about 500 mg of Se, which exceeds the upper safe intake level of 400 µg per day for human dietary. The low Se applications (0.0035 and 0.1 mg kg-1) diminished and retarded the degree of raw darkening in tubers stored for one and eight months, which can be attributed to the antioxidative properties of Se. The storage for 1 to 12 months did not affect the Se concentrations of tubers. In the Se enriched tubers Se was allocated to the organic Se fraction, indicating that it was incorporated into organic compounds in tubers. Elevated Se concentration in the next-generation tubers produced by the Se enriched seed tubers indicated that Se could be translocated from the seed tubers to the progeny. In the seed tubers stored for 8 months, at high levels, Se had some positive effects on the growth vigour of sprouts, but Se had no consistent effect on the growth vigour of seed tubers of optimal physiological age. These results indicate that Se is a beneficial trace element in potato plants that exerts a positive effect on yield formation and improves the processing and storage quality of table potato tubers. These positive effects of Se are, however, dependent on the Se concentration and the age of the potato plant and tuber.

Egg production in furnished cages

In the European Union, conventional cages for laying hens will be faded out at the beginning of 2012. The rationale behind this is a public concern over animal welfare in egg production. As alternatives to conventional cages, the European Union Council Directive 1999/74/EC allows non-cage systems and enriched (furnished) cages. Layer performance, behavior, and welfare in differently sized furnished cages have been investigated quite widely during recent decades, but nutrition of hens in this production system has received less attention. This thesis aims to compare production and feed intake of laying hens in furnished and conventional cages and to study the effects of different dietary treatments in these production systems, thus contributing to the general knowledge of furnished cages as an egg production system. A furnished cage model for 8 hens was compared with a 3-hen conventional cage. Three consecutive experiments each studied one aspect of layer diet: The first experiment investigated the effects of dietary protein/energy ratio, the second dietary energy levels, and the third the effects of extra limestone supplementation. In addition, a fourth experiment evaluated the effects of perches on feed consumption and behavior of hens in furnished cages. The dietary treatments in experiments 1 3 generally had similar effects in the two cage types. Thus, there was no evidence supporting a change in nutrient requirements for laying hens when conventional cages are replaced with small-group furnished cages. Moreover, the results from nutritional experiments conducted in conventional cages can be applied to small-group furnished cage systems. These results support the view that production performance comparable with conventional cages can be achieved in furnished cages. All of the advantages of cages for bird welfare are sustained in the small-group furnished cages used here. In addition, frequent use of perches and nests implies a wider behavioral repertoire in furnished cages than in conventional cages. The increase observed in bone ash content may improve bird welfare in furnished cages. The presence of perches diminished feed consumption during the prelaying period and enhanced the feed conversion ratio during the early laying period in furnished cages. However, as the presence or absence of perches in furnished cages had no significant effect on feed consumption after the prelaying period, the lower feed consumption observed in furnished cages than in conventional cages could be attributed to other factors, such as the presence of wood shavings or a nest box. The wider feed trough space per hen in conventional than in furnished cages may partly explain the higher feed consumption observed in conventional cages.

Microfiltration in cheese and whey processing

Milk microfiltration (0.05-0.2 um) is a membrane separation technique which divides milk components into casein-enriched and native whey fractions. Hitherto the effect of intensive microfiltration including a diafiltration step for both cheese and whey processing has not been studied. The microfiltration performance of skimmed milk was studied with polymeric and ceramic MF membranes. The changes caused by decreased concentration of milk lactose, whey protein and ash content for cheese milk quality and ripening were studied. The effects of cheese milk modification on the milk coagulation properties, cheese recovery yield, cheese composition, ripening and sensory quality as well as on the whey recovery yield and composition by microfiltration were studied. The functional properties of whey protein concentrate from native whey were studied and the detailed composition of whey protein concentrate powders made from cheese wheys after cheese milk pretreatments such as high temperature heat treatment (HH), microfiltration (MF) and ultrafiltration (UF) were compared. The studied polymeric spiral wound microfiltration membranes had 38.5% lower energy consumption, 30.1% higher retention of whey proteins to milk retentate and 81.9% lower permeate flux values compared to ceramic membranes. All studied microfiltration membranes were able to separate main whey proteins from skimmed milk. The optimal lactose content of Emmental cheese milk exceeded 3.2% and reduction of whey proteins and ash content of cheese milk with high concentration factor (CF) values increased the rate of cheese ripening. Reduction of whey protein content in cheese milk increased the concentration of caseinomacropeptide (CMP) of total proteins in cheese whey. Reduction of milk whey protein, lactose and ash content reduces milk rennet clotting time and increased the firmness of the coagulum. Cheese yield calculated from raw milk to cheese was lower with microfiltrated milks due to native whey production. Amounts of a-lactalbumin (a-LA) and b-lactoglobulin (b-LG) were significantly higher in the reference whey, indicating that HH, MF and UF milk pretreatments decrease the amounts of these valuable whey proteins in whey. Even low CF values in milk microfiltration (CF 1.4) reduced nutritional value of cheese whey. From the point of view of utilization of milk components it would be beneficial if the amount of native whey and the CMP content of cheese whey could be maximized. Whey protein concentrate powders made of native whey had excellent functional properties and their detailed amino acid composition differed from those of cheese whey protein concentrate powders.

Phosphorus in agricultural soils of Finland - characterization of reserves and retention in mineral soil profiles

An overwhelming majority of all the research on soil phosphorus (P) has been carried out with soil samples taken from the surface soils only, and our understanding of the forms and the reactions of P at a soil profile scale is based on few observations. In Finland, the interest in studying the P in complete soil profiles has been particularly small because of the lack of tradition in studying soil genesis, morphology, or classification. In this thesis, the P reserves and the retention of orthophosphate phosphorus (PO4-P) were examined in four cultivated mineral soil profiles in Finland (three Inceptisols and one Spodosol). The soils were classified according to the U.S. Soil Taxonomy and soil samples were taken from the genetic horizons in the profiles. The samples were analyzed for total P concentration, Chang and Jackson P fractions, P sorption properties, concentrations of water-extractable P, and for concentrations of oxalate-extractable Al and Fe. Theoretical P sorption capacities and degrees of P saturation were calculated with the data from the oxalate-extractions and the P fractionations. The studied profiles can be divided into sections with clearly differing P characteristics by their master horizons Ap, B and C. The C (or transitional BC) horizons below an approximate depth of 70 cm were dominated by, assumingly apatitic, H2SO4-soluble P. The concentration of total P in the C horizons ranged from 729 to 810 mg kg-1. In the B horizons between the depths of 30 and 70 cm, a significant part of the primary acid-soluble P has been weathered and transformed to secondary P forms. A mean weathering rate of the primary P in the soils was estimated to vary between 230 and 290 g ha-1 year-1. The degrees of P saturation in the B and C horizons were smaller than 7%, and the solubility of PO4-P was negligible. The P conditions in the Ap horizons differed drastically from those in the subsurface horizons. The high concentrations of total P (689-1870 mg kg-1) in the Ap horizons are most likely attributable to long-term cultivation with positive P balances. A significant proportion of the P in the Ap horizons occurred in the NH4F- and NaOH-extractable forms and as organic P. These three P pools, together with the concentrations of oxalate-extractable Al and Fe, seem to control the dynamics of PO4-P in the soils. The degrees of P saturation in the Ap horizons were greater (8-36%) than in the subsurface horizons. This was also reflected in the sorption experiments: Only the Ap horizons were able to maintain elevated PO4-P concentrations in the solution phase − all the subsoil horizons acted as sinks for PO4-P. Most of the available sorption capacity in the soils is located in the B horizons. The results suggest that this capacity could be utilized in reducing the losses of soluble P from excessively fertilized soils by mixing highly sorptive material from the B horizons with the P-enriched surface soil. The drastic differences in the P characteristics observed between adjoining horizons have to be taken into consideration when conducting soil sampling. Sampling of subsoils has to be made according to the genetic horizons or at small depth increments. Otherwise, contrasting materials are likely to be mixed in the same sample; and the results of such samples are not representative of any material present in the studied profile. Air-drying of soil samples was found to alter the results of the sorption experiments and the water extractions. This indicates that the studies on the most labile P forms in soil should be carried out with moist samples.

HPLC analysis of plant sterol oxidation products

Increased interest in the cholesterol-lowering effect of plant sterols has led to development of plant sterol-enriched foods. When products are enriched, the safety of the added components must be evaluated. In the case of plant sterols, oxidation is the reaction of main concern. In vitro studies have indicated that cholesterol oxides may have harmful effects. Due their structural similarity, plant sterol oxidation products may have similar health implications. This study concentrated on developing high-performance liquid chromatography (HPLC) methods that enable the investigation of formation of both primary and secondary oxidation products and thus can be used for oxidation mechanism studies of plant sterols. The applicability of the methods for following the oxidation reactions of plant sterols was evaluated by using oxidized stigmasterol and sterol mixture as model samples. An HPLC method with ultraviolet and fluorescence detection (HPLC-UV-FL) was developed. It allowed the specific detection of hydroperoxides with FL detection after post-column reagent addition. The formation of primary and secondary oxidation products and amount of unoxidized sterol could be followed by using UV detection. With the HPLC-UV-FL method, separation between oxides was essential and oxides of only one plant sterol could be quantified in one run. Quantification with UV can lead to inaccuracy of the results since the number of double bonds had effect on the UV absorbance. In the case of liquid chromatography-mass spectrometry (LC-MS), separation of oxides with different functionalities was important because some oxides of the same sterol have similar molecular weight and moreover epimers have similar fragmentation behaviour. On the other hand, coelution of different plant sterol oxides with the same functional group was acceptable since they differ in molecular weights. Results revealed that all studied plant sterols and cholesterol seem to have similar fragmentation behaviour, with only relative ion abundances being slightly different. The major advantage of MS detection coupled with LC separation is the capability to analyse totally or partly coeluting analytes if these have different molecular weights. The HPLC-UV-FL and LC-MS methods were demonstrated to be suitable for studying the photo-oxidation and thermo-oxidation reactions of plant sterols. The HPLC-UV-FL method was able to show different formation rates of hydroperoxides during photo-oxidation. The method also confirmed that plant sterols have similar photo-oxidation behaviour to cholesterol. When thermo-oxidation of plant sterols was investigated by HPLC-UV-FL and LC-MS, the results revealed that the formation and decomposition of individual hydroperoxides and secondary oxidation products could be studied. The methods used revealed that all of the plant sterols had similar thermo-oxidation behaviour when compared with each other, and the predominant reactions and oxidation rates were temperature dependent. Overall, these findings showed that with these LC methods the oxidation mechanisms of plant sterols can be examined in detail, including the formation and degradation of individual hydroperoxides and secondary oxidation products, with less sample pretreatment and without derivatization.