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.

Transition : en betydelsefull fas i den långtidssjuka ungas vård

The aim of this caring science thesis is to deepen the understanding for the long-term ill adolescents care. In particular it aims to reach a better understanding of transition in health care, with a focus on how young people perceive health care. The goal is to build a model, a consistent theoretical framework that makes it possible to listen to the adolescent´s own voice, providing a better understanding of what good care consists in from their point of view. This Master's thesis consists of a literature overview on the concept of transition and a themed interview with young patients regarding transition of health care. The overall issue is 1. What is good care during the transition according to the adolescents themselves? 2. How does transition affect the young people's health and lives? The theoretical perspective of this thesis lies in the caring science tradition that has been developed at the Åbo Akademi University department of caring science with the caritative nursing theory as a basis. The epistemological and ontological assumptions are based on the caritative nursing theory, which in itself is ethical. Understanding of the concept of transition is based on A. Meleis transitions theory. The methodological approach is hermeneutical. The theme interview has been analyzed by using S. Kvales method. The material consists of a thematic interview with 10 long-term ill adolescents, about their views of the care received. This thesis research even in how NOBABs standards appears in the litterature as well as how they are implemented from the point of view of young people. The findings of this thesis shows that good care for the adolescent is a true presence with the young and an understanding of the direction in which the young move toward during the transition of care. Good care for an adolescent consists in being recognized as a person who is seen, known, appreciated and understood. Good care for the young involves recognizing her strength and joy of life including the contrast she experiences between health and suffering. To recognize their personal strength and courage supports the adolescents during the process of transitions of care Good care for the young involves responsibility for the individual person and humility in approaching her, giving attention to her view of life and aspirations. Openness and humility in the approach lays the foundation for a trusting relation during the transition of care. A unified theoretical framework is needed to promote the long-term ill adolescents care during transition of care. When the young and her individual needs is the centerpiece of healthcare creates this the basis for empowering care that supports the young in her quest for health. This master thesis adds a given model to the long-term ill adolescents` healthcare.

Starting Insulin Treatment in Type 2 Diabetes

Type 2 diabetes is a disorder of glucose metabolism characterized by chronic hyperglycemia. Initially type 2 diabetes is characterized by insulin resistance and impaired function of beta cells, leading progressively to insulin deficiency. Type 2 diabetes is treated with diet and other lifestyle changes, and with medication modulating e.g. insulin resistance, liver glucose production and insulin secretion. Injectable insulin is added to the treatment when lifestyle changes and other medication are insufficient to maintain adequate control of hyperglycemia. The aim of the treatment is to remove the symptoms of diabetes and to prevent late complications of diabetes. Insulin was traditionally started at hospital wards, but from the early 1990’s also in outpatient care. The first substudy of this thesis examined retrospectively initiation practices and how successfully insulin treatment was introduced in 1990 – 1996 in Southwestern Finland. This study aimed also at identifying the best methods of controlling plasma glucose. It showed that in the 1990’s the incidence of insulin treatment increased and was initiated more often in outpatient care than previously. The use of combination treatment also increased, first with sulfonylureas and later with metformin as the oral drug. In combination therapy the insulin dose was smaller than with insulin monotherapy. HbA1c improved similarly in middle-aged and older age groups. Weight increase associated with insulin initiation was smaller when combined with oral agents. A prospective insulin initiation study (1994 – 1998) tested the hypothesis that hyperglycemia (fasting and postprandial hyperglycemia) may affect the outcome of insulin initiation. The type of hyperglycemia was determined by the relation of fasting plasma glucose to HbA1c. Treatment was initiated with insulin Lente or human NPH insulin. In patients treated with insulin monotherapy twice daily the decline in HbA1c was markedly greater for postprandial than fasting hyperglycemia patients suggesting that hyperglycemia type has significance in the selection of the insulin regimen. Another insulin initiation study showed that patients with fasting hyperglycemia starting on insulin (2004-2005) were significantly more prone to overweight than patients with postprandial hyperglycemia. Irrespective of the insulin preparation (insulin NPH or insulin glargine), patients with fasting hyperglycemia had a greater weight increase compared to patients with postprandial hyperglycemia. Special attention should be paid to prevention of weight increase in these patients.

Probing MST1 Kinase Sequence and Structure for Rational Drug Discovery (Targeting diabetes by blocking protein-protein interactions)

Apoptotic beta cell death is an underlying cause majorly for type I and to a lesser extent for type II diabetes. Recently, MST1 kinase was identified as a key apoptotic agent in diabetic condition. In this study, I have examined MST1 and closely related kinases namely, MST2, MST3 and MST4, aiming to tackle diabetes by exploring ways to selectively block MST1 kinase activity. The first investigation was directed towards evaluating possibilities of selectively blocking the ATP binding site of MST1 kinase that is essential for the activity of the enzymes. Structure and sequence analyses of this site however revealed a near absolute conservation between the MSTs and very few changes with other kinases. The observed residue variations also displayed similar physicochemical properties making it hard for selective inhibition of the enzyme. Second, possibilities for allosteric inhibition of the enzyme were evaluated. Analysis of the recognized allosteric site also posed the same problem as the MSTs shared almost all of the same residues. The third analysis was made on the SARAH domain, which is required for the dimerization and activation of MST1 and MST2 kinases. MST3 and MST4 lack this domain, hence selectivity against these two kinases can be achieved. Other proteins with SARAH domains such as the RASSF proteins were also examined. Their interaction with the MST1 SARAH domain were evaluated to mimic their binding pattern and design a peptide inhibitor that interferes with MST1 SARAH dimerization. In molecular simulations the RASSF5 SARAH domain was shown to strongly interact with the MST1 SARAH domain and possibly preventing MST1 SARAH dimerization. Based on this, the peptidic inhibitor was suggested to be based on the sequence of RASSF5 SARAH domain. Since the MST2 kinase also interacts with RASSF5 SARAH domain, absolute selectivity might not be achieved.