Gene Regulation in The Human Immune System. Gene Expression Signatures of Th2 Cell Differentiation, Type 1 Diabetes, and Intrauterine Immune Adaptation

The human immune system is constantly interacting with the surrounding stimuli and microorganisms. However, when directed against self or harmless antigens, these vital defense mechanisms can cause great damage. In addition, the understanding the underlying mechanism of several human diseases caused by aberrant immune cell functions, for instance type 1 diabetes and allergies, remains far from being complete. In this Ph.D. study these questions were addressed using genome-wide transcriptomic analyses. Asthma and allergies are characterized by a hyperactive response of the T helper 2 (Th2) immune cells. In this study, the target genes of the STAT6 transcription factor in naïve human T cells were identified with RNAi for the first time. STAT6 was shown to act as a central activator of the genes expression upon IL-4 signaling, with both direct and indirect effects on Th2 cell transcriptome. The core transcription factor network induced by IL-4 was identified from a kinetic analysis of the transcriptome. Type 1 diabetes is an autoimmune disease influenced by both the genetic susceptibility of an individual and the disease-triggering environmental factors. To improve understanding of the autoimmune processes driving pathogenesis in the prediabetic phase in humans, a unique series of prospective whole-blood RNA samples collected from HLA-susceptible children in the Finnish Type 1 Diabetes Prediction and Prevention (DIPP) study was studied. Changes in different timewindows of the pathogenesis process were identified, and especially the type 1 interferon response was activated early and throughout the preclinical T1D. The hygiene hypothesis states that allergic diseases, and lately also autoimmune diseases, could be prevented by infections and other microbial contacts acquired in early childhood, or even prenatally. To study the effects of the standard of hygiene on the development of neonatal immune system, cord blood samples from children born in Finland (high standard of living), Estonia (rapid economic growth) and Russian Karelia (low standard of living) were compared. Children born in Russian Karelia deviated from Finnish and Estonian children in many aspects of the neonatal immune system, which was developmentally more mature in Karelia, resembling that of older infants. The results of this thesis offer significant new information on the regulatory networks associated with immune-mediated diseases in human. The results will facilitate understanding and further research on the role of the identified target genes and mechanisms driving the allergic inflammation and type 1 diabetes, hopefully leading to a new era of drug development.

Probiotic interventions in infancy: Benefit and Safety Assessment of Extended Applications

Immaturity of the gut barrier system in the newborn has been seen to underlie a number of chronic diseases originating in infancy and manifesting later in life. The gut microbiota and breast milk provide the most important maturing signals for the gut-related immune system and reinforcement of the gut mucosal barrier function. Recently, the composition of the gut microbiota has been proposed to be instrumental in control of host body weight and metabolism as well as the inflammatory state characterizing overweight and obesity. On this basis, inflammatory Western lifestyle diseases, including overweight development, may represent a potential target for probiotic interventions beyond the well documented clinical applications. The purpose of the present undertaking was to study the efficacy and safety of perinatal probiotic intervention. The material comprised two ongoing, prospective, double-blind NAMI (Nutrition, Allergy, Mucosal immunology and Intestinal microbiota) probiotic interventions. In the mother-infant nutrition and probiotic study altogether 256 women were randomized at their first trimester of pregnancy into a dietary intervention and a control group. The intervention group received intensive dietary counselling provided by a nutritionist, and were further randomized at baseline, double-blind, to receive probiotics (Lactobacillus rhamnosus GG and Bifidobacterium lactis) or placebo. The intervention period extended from the first trimester of pregnancy to the end of exclusive breastfeeding. In the allergy prevention study altogether 159 women were randomized, double-blind, to receive probiotics (Lactobacillus rhamnosus GG) or placebo 4 weeks before expected delivery, the intervention extending for 6 months postnatally. Additionally, patient data on all premature infants with very low birth weight (VLBW) treated in the Department of Paediatrics, Turku University Hospital, during the years 1997 - 2008 were utilized. The perinatal probiotic intervention reduced the risk of gestational diabetes mellitus (GDM) in the mothers and perinatal dietary counselling reduced that of fetal overgrowth in GDM-affected pregnancies. Early gut microbiota modulation with probiotics modified the growth pattern of the child by restraining excessive weight gain during the first years of life. The colostrum adiponectin concentration was demonstrated to be dependent on maternal diet and nutritional status during pregnancy. It was also higher in the colostrum received by normal-weight compared to overweight children at the age of 10 years. The early perinatal probiotic intervention and the postnatal probiotic intervention in VLBW infants were shown to be safe. To conclude, the findings in this study provided clinical evidence supporting the involvement of the initial microbial and nutritional environment in metabolic programming of the child. The manipulation of early gut microbial communities with probiotics might offer an applicable strategy to impact individual energy homeostasis and thus to prevent excessive body-weight gain. The results add weight to the hypothesis that interventions aiming to prevent obesity and its metabolic consequences later in life should be initiated as early as during the perinatal period.

Diet, microbes and gut immune responses in the pathogenesis of experimental type 1 diabetes

Type 1diabetes (T1D) is an autoimmune disease, which is influenced by a variety of environmental factors including diet and microbes. These factors affect the homeostasis and the immune system of the gut. This thesis explored the altered regulation of the immune system and the development of diabetes in non-obese diabetic (NOD) mice. Inflammation in the entire intestine of diabetes-prone NOD mice was studied using a novel ex-vivo imaging system of reactive oxygen and nitrogen species (RONS), in relation to two feeding regimens. In parallel, gut barrier integrity and intestinal T-cell activation were assessed. Extra-intestinal manifestations of inflammation and decreased barrier integrity were sought for by studying peritoneal leukocytes. In addition, the role of pectin and xylan as dietary factors involved in diabetes development in NOD mice was explored. NOD mice showed expression of RONS especially in the distal small intestine, which coincided with T-cell activation and increased permeability to macromolecules. The introduction of a casein hydrolysate (hydrolysed milk protein) diet reduced these phenomena, altered the gut microbiota and reduced the incidence of T1D. Extra-intestinally, macrophages appeared in large numbers in the peritoneum of NOD mice after weaning. Peritoneal macrophages (PM) expressed high levels of interleukin-1 receptor associated kinase M (IRAK-M), which was indicative of exposure to ligands of toll-like receptor 4 (TLR-4) such as bacterial lipopolysaccharide (LPS). Intraperitoneal LPS injections activated T cells in the pancreatic lymph nodes (PaLN) and thus, therefore potentially could activate islet-specific T cells. Addition of pectin and xylan to an otherwise diabetes-retarding semisynthetic diet affected microbial colonization of newly-weaned NOD mice, disturbed gut homeostasis and promoted diabetes development. These results help us to understand how diet and microbiota impact the regulation of the gut immune system in a way that might promote T1D in NOD mice.

Implementation of an Integrated Activity-Based Costing in a Holistic Database Management System and its Implications for Operations Management

Diplomityössä on tutkittu reaaliaikaisen toimintolaskennan toteuttamista suomalaisen lasersiruja valmistavan PK-yrityksen tietojärjestelmään. Lisäksi on tarkasteltu toimintolaskennan vaikutuksia operatiiviseen toimintaan sekä toimintojen johtamiseen. Työn kirjallisuusosassa on käsitelty kirjallisuuslähteiden perusteella toimintolaskennan teorioita, laskentamenetelmiä sekä teknisessä toteutuksessa käytettyjä teknologioita. Työn toteutusosassa suunniteltiin ja toteutettiin WWW-pohjainen toimintolaskentajärjestelmä case-yrityksen kustannuslaskennan sekä taloushallinnon avuksi. Työkalu integroitiin osaksi yrityksen toiminnanohjaus- sekä valmistuksenohjausjärjestelmää. Perinteisiin toimintolaskentamallien tiedonkeruujärjestelmiin verrattuna case-yrityksessä syötteet toimintolaskentajärjestelmälle tulevat reaaliaikaisesti osana suurempaa tietojärjestelmäintegraatiota.Diplomityö pyrkii luomaan suhteen toimintolaskennan vaatimusten ja tietokantajärjestelmien välille. Toimintolaskentajärjestelmää yritys voi hyödyntää esimerkiksi tuotteiden hinnoittelussa ja kustannuslaskennassa näkemällä tuotteisiin liittyviä kustannuksia eri näkökulmista. Päätelmiä voidaan tehdä tarkkaan kustannusinformaatioon perustuen sekä määrittää järjestelmän tuottaman datan perusteella, onko tietyn projektin, asiakkuuden tai tuotteen kehittäminen taloudellisesti kannattavaa.

1000 V distribution system and its potential in Russia

The application of the three voltage level 20/1/0.4 distribution system in Finland has proved to be an economic solution to enhance the reability of electricity distribution. By using 1 kV voltage level between medium and low voltage networks, the improvement in reability could be reached especially inaerial lines networks. Also considerable savings in investment and outage costscould be archieved compared to the traditional distribution system. This master's thesis is focused on the describing the situation in Russian distribution netwoks and consequent analyses the possibility of applying 1000V distribution system in Russia. The goal is to investigate on the basis of Finnish experience is any possible installation targets in Russia for the new system. Compatibility with Russian safety and quality standards are also studied in this thesis.

Deep-hole drilling process, monitoring system and sensors modifications

 In the drilling processes and especially deep-hole drilling process, the monitoring system and having control on mechanical parameters (e.g. Force, Torque,Vibration and Acoustic emission) are essential. The main focus of this thesis work is to study the characteristics of deep-hole drilling process, and optimize the monitoring system for controlling the process. The vibration is considered as a major defect area of the deep-hole drilling process which often leads to breakage of the drill, therefore by vibration analysis and optimizing the workpiecefixture, this area is studied by finite element method and the suggestions are explained. By study on a present monitoring system, and searching on the new sensor products, the modifications and recommendations are suggested for optimize the present monitoring system for excellent performance in deep-hole drilling process research and measurements.

Dimensioning DC-voltage capacitor for a DC-distribution system and dynamic analysis

Technical analysis of Low Voltage Direct Current (LVDC) distribution systems shows that in LVDC transmission the customer voltage quality is higher. One of the problems in LVDC distribution networks that converters both ends of the DC line are required. Because of the converters produce not pure DC voltage, but some fluctuations as well, the huge electrolytic capacitors are required to reduce voltage distortions in the DC-side. This thesis master’s thesis is focused on calculating required DC-link capacitance for LVDC transmission and estimation of the influence of different parameters on the voltage quality. The goal is to investigate the methods of the DC-link capacitance estimation and location in the transmission line.

The Influence of Diet and Microbes on Colonic Immune Regulation and their Implications on Type 1 Diabetes

Dietary and microbial factors are thought to contribute to the rapidly increasing prevalence of T1D in many countries worldwide. The impact of these factors on immune regulation and diabetes development in non-obese diabetic (NOD) mice are investigated in this thesis. Diabetes can be prevented in NOD mice through dietary manipulation. Diet affects the composition of intestinal microbiota, which may subsequently influence intestinal immune homeostasis. However, the specific effects of anti-diabetogenic diets on gut immunity and the explicit associations between intestinal immune disruption and type 1 diabetes onset remain unclear. The research presented herein demonstrates that newly weaned NOD mice suffer from a mild level of colitis, which shifts the colonic immune cell balance towards a proinflammatory status. Several aberrations can also be observed in the peritoneal B cells of NOD mice; an increase in activation marker expression, increased trafficking to the pancreatic lymph nodes and significantly higher antigen presenting cell (APC) efficiency towards insulin-specific T cells. A shift towards inflammation is likewise observed in the colon of germ-free NOD mice, but signs of peritoneal B cell activation are lacking in these mice. Remarkably, most of the abnormalities in the colon, peritoneal macrophages and the peritoneal B cell APC activity of NOD mice are abrogated when NOD mice are maintained on a diabetes-preventive, soy-based diet (ProSobee) from the time of weaning. Dietary and microbial factors hence have a significant impact on colonic immune regulation and peritoneal B cell activation and it is suggested that these factors influence diabetes development in NOD mice.

Researching Manufacturing Planning and Control system and Master Scheduling in a manufacturing firm.

The objective of this thesis is to research Manufacturing Planning and Control (MPC) system and Master Scheduling (MS) in a manufacturing firm. The study is conducted at Ensto Finland Corporation, which operates on a field of electrical systems and supplies. The paper consists of theoretical and empirical parts. The empirical part is based on weekly operating at Ensto and includes inter-firm material analysis, learning and meetings. Master Scheduling is an important module of an MPC system, since it is beneficial on transforming strategic production plans based on demand forecasting into operational schedules. Furthermore, capacity planning tools can remarkably contribute to production planning: by Rough-Cut Capacity Planning (RCCP) tool, a MS plan can be critically analyzed in terms of available key resources in real manufacturing environment. Currently, there are remarkable inefficiencies when it comes to Ensto’s practices: the system is not able to take into consideration seasonal demand and react on market changes on time; This can cause significant lost sales. However, these inefficiencies could be eliminated through the appropriate utilization of MS and RCCP tools. To utilize MS and RCCP tools in Ensto’s production environment, further testing in real production environment is required. Moreover, data accuracy, appropriate commitment to adapting and learning the new tools, and continuous developing of functions closely related to MS, such as sales forecasting, need to be ensured.

Generation and Characterization of Knockout Mice and Analysis of Patient Material Demonstrates a Role for Tissue Inhibitor of Metalloproteinases 4 (Timp4) in the Cardiovascular System and Tumor Metastasis

This thesis focuses on tissue inhibitor of metalloproteinases 4 (TIMP4) which is the newest member of a small gene and protein family of four closely related endogenous inhibitors of extracellular matrix (ECM) degrading enzymes. Existing data on TIMP4 suggested that it exhibits a more restricted expression pattern than the other TIMPs with high expression levels in heart, brain, ovary and skeletal muscle. These observations and the fact that the ECM is of special importance to provide the cardiovascular system with structural strength combined with elasticity and distensibility, prompted the present molecular biologic investigation on TIMP4. In the first part of the study the murine Timp4 gene was cloned and characterized in detail. The structure of murine Timp4 genomic locus resembles that in other species and of the other Timps. The highest Timp4 expression was detected in heart, ovary and brain. As the expression pattern of Timp4 gives only limited information about its role in physiology and pathology, Timp4 knockout mice were generated next. The analysis of Timp4 knockout mice revealed that Timp4 deficiency has no obvious effect on the development, growth or fertility of mice. Therefore, Timp4 deficient mice were challenged using available cardiovascular models, i.e. experimental cardiac pressure overload and myocardial infarction. In the former model, Timp4 deficiency was found to be compensated by Timp2 overexpression, whereas in the myocardial infarct model, Timp4 deficiency resulted in increased mortality due to increased susceptibility for cardiac rupture. In the wound healing model, Timp4 deficiency was shown to result in transient retardation of re-epithelialization of cutaneous wounds. Melanoma tumor growth was similar in Timp4 deficient and control mice. Despite of this, lung metastasis of melanoma cells was significantly increased in Timp4 null mice. In an attempt to translate the current findings to patient material, TIMP4 expression was studied in human specimens representing different inflammatory cardiovascular pathologies, i.e. giant cell arteritis, atherosclerotic coronary arteries and heart allografts exhibiting signs of chronic rejection. The results showed that cardiovascular expression of TIMP4 is elevated particularly in areas exhibiting inflammation. The results of the present studies suggest that TIMP4 has a special role in the regulation of tissue repair processes in the heart, and also in healing wounds and metastases. Furthermore, evidence is provided suggesting the usefulness of TIMP4 as a novel systemic marker for vascular inflammation.

The Roots of Neurofibromas in Neurofibromatosis 1 — A Question of Multipotency, Differentiation and Hiding

Neurofibromatosis type 1 (NF1) is an autosomal dominant cancer predisposition syndrome that affects about 1 in 3500 individuals worldwide. NF1 is caused by mutations in the NF1 gene that encodes the tumor suppressor protein neurofibromin, an inactivator of the Ras oncogene. The hallmarks of NF1 include pigmentary lesions of the skin, Lisch nodules of the iris and cutaneous neurofibromas. Cutaneous neurofibromas are benign tumors composed of all the cell types of normal peripheral nerve. The traditional view of neurofibroma development has been that cutaneous neurofibromas arise from the disruption of the small nerve tributaries of the skin and subsequent proliferation of the resident cells. The second hit mutation in the NF1 gene has been considered as a prerequisite for neurofibroma development. The second hit is detectable in a subpopulation of primary Schwann cells cultured from neurofibromas. This thesis challenges the traditional concept of neurofibroma development. The results show that cutaneous neurofibromas are intimately associated with hair follicular structures and contain multipotent precursor cells (NFPs), suggesting that neurofibromas may arise from the multipotent cells which reside in hair follicles. Furthermore, this study presents that neurofibroma-derived Schwann cells that harbor bi-allelic inactivation in the NF1 gene express HLA class II genes and may act as nonprofessional antigen presenting cells. The CD4- and FoxP3-positive cells detected in cutaneous neurofibromas suggest that these cells may represent regulatory T cells (Tregs) which interact with HLA II –positive cells and aid the tumor cells in hiding from the immune system and are thus mediators of immune tolerance. This thesis also investigated neurofibroma development in the oral cavity and the use of different biomarkers to characterize cellular differentiation in neurofibromas. The results revealed that oral neurofibromas are not rare, but they usually appear as solitary lesions contrary to multiple cutaneous neurofibromas and present high heterogeneity within and between tumors. The use of class III beta-tubulin as a marker for neuronal differentiation led to an unexpected finding showing that multiple cell types express class III beta-tubulin during mitosis. The increased understanding of the multipotency of tumor cells, cellular differentiation and ability to hide from immune system will aid in the development of future treatments. Specifically, targeting Tregs in NF1 patients could provide a novel therapeutic approach to interfere with the development of neurofibromas.

Implementing an open data system and showing its benefits

Open data refers to publishing data on the web in machine-readable formats for public access. Using open data, innovative applications can be developed to facilitate people‟s lives. In this thesis, based on the open data cases (discussed in the literature review), Open Data Lappeenranta is suggested, which publishes open data related to opening hours of shops and stores in Lappeenranta City. To prove the possibility of creating Open Data Lappeenranta, the implementation of an open data system is presented in this thesis, which publishes specific data related to shops and stores (including their opening hours) on the web in standard format (JSON). The published open data is used to develop web and mobile applications to demonstrate the benefits of open data in practice. Also, the open data system provides manual and automatic interfaces which make it possible for shops and stores to maintain their own data in the system. Finally in this thesis, the completed version of Open Data Lappeenranta is proposed, which publishes open data related to other fields and businesses in Lappeenranta beyond only stores‟ data.

Pertussis: infection, vaccination and gene polymorphisms

Pertussis or whooping cough is a human respiratory tract infection and a vaccine-preventable disease that is caused by Bordetella pertussis bacteria. Pertussis vaccination has been part of the Finnish national vaccine program since 1952. Despite extensive vaccinations, the incidence of pertussis has increased in many countries during the last decades. Large epidemics have been observed also in countries with high vaccine coverage. Inter-individual variation in immune responses is always encountered after vaccination. Low vaccine responses may cause vulnerability to pertussis even straight after vaccination. Reasons for low responses are not fully understood. The innate immune system is responsible for the initial recognition of pathogens and vaccine antigens. The role of innate immunity on pertussis immunity has not been thoroughly investigated. Mannose-binding lectin (MBL) and toll-like receptor 4 (TLR4) are important molecules of the innate immune system and in the recognition of pathogens. Cytokines form a signaling network that have a notable role in immune responses after infections as well as after vaccinations. Single nucleotide polymorphism (SNP) is common in genes encoding these molecules and the polymorphisms have been reported to affect vaccine response after viral and bacterial vaccines. This study investigated the gene polymorphisms of MBL2, TLR4 and interleukin (IL)-10 promoter and their association with vaccine responses after acellular pertussis (aP) vaccination in Finnish adolescents and infants. Cell-mediated immune responses were investigated ten years after the previous pertussis vaccinations in young adults. In addition, the role of MBL deficiency in pertussis infection susceptibility was evaluated. The results of this study show that subjects with TLR4 polymorphism had lower antibody production and persistence after aP vaccination compared with normal allele. A specific SNP in the TLR4 gene was associated with decreased antibody responses and persistence in adolescents after aP booster vaccination. Cell-mediated immune responses were partly detected ten years after the previous vaccination; booster vaccine clearly enhanced the responses. In addition, subjects with IL-10 polymorphism had altered cell-mediated immune responses. MBL deficiency was found to be more frequent in pertussis patients than healthy controls but the polymorphism of MBL2 was not associated with antibody responses after acellular pertussis vaccination. The novel finding of this study was that genetic variation in the innate immune system seems to play a role in altered pertussis vaccine responses as well as in pertussis infection. These new findings enlighten the mechanisms behind the low responses after pertussis vaccination and help to predict risk factors related to this phenomenon.