The human being and other animals contain immensely complex biochemical processes that govern their function on a cellular level. It is estimated that several thousand small molecules (metabolites) are produced by various biochemical pathways in humans. Pathological processes can introduce perturbations in these biochemical pathways which can lead to changes in the amounts of some metabolites.Developments in analytical chemistry have made it possible measure a large number metabolites in a single blood sample, which gives a metabolic profile. In this thesis I have worked on establishing and understanding metabolic profiles from patients with rheumatoid arthritis (RA) and from animal models of the autoimmune diseases diabetes mellitus type 1 (T1D) and RA.Using multivariate statistical methods it is possible to identify differences between metabolic profiles of different groups. As an example we identified differences between patients with RA and healthy volunteers. This can be used to elucidate the biochemical processes that are active in a given pathological condition.Metabolite concentrations are affected by a many other things than the presence or absence of a disease. Both genomic and environmental factors are known to influence metabolic profiles. A main focus of my work has therefore been on finding strategies for ensuring that the results obtained when comparing metabolic profiles were valid and relevant. This strategy has included repetition of experiments and repeated measurement of individuals’ metabolic profiles in order to understand the sources of variation.Finding the most stable and reproducible metabolic effects has allowed us to better understand the biochemical processes seen in the metabolic profiles. This makes it possible to relate the metabolic profile differences to pathological processes and to genes and proteins involved in these.The hope is that metabolic profiling in the future can be an important tool for finding biomarkers useful for disease diagnosis, for identifying new targets for drug design and for mapping functional changes of genomic mutations. This has the potential to revolutionize our understanding of disease pathology and thus improving health care.

Enterovirus (EV) infections have been associated with the pathogenesis of Type 1 Diabetes (T1D). However, the pathway(s) by which EV may induce or accelerate diabetes is not well understood. The purpose of this thesis was to obtain new information on the mechanism by which EV infections, with different strains of EV, could cause damage to the insulin-producing β-cells in isolated human islets and in a rat insulin-producing cell line (RINm5F). Infection with EV strains isolated from T1D patients revealed replication/cell destruction in human islets and EV-like particles in the cytoplasm of the β-cell and infection with the isolates affected the release of insulin in response to glucose stimulation as early as three days post infection, before any decrease in cell viability was observed. A decrease in the induction/secretion of the chemokine RANTES in human islets during EV infection was also detected. When islets were cultured with nicotinamide (NA) the secretion of RANTES was increased irrespectively if the islets were infected or not. In addition, the degree of virus-induced cytolysis of human islets was reduced by NA, suggesting an antiviral effect of NA. Infection with EV strains revealed permissiveness to islet-derived cells. All EV strains used for infection were able to replicate in the RIN cell clusters (RCC) but not in the RIN cells that were cultured as a monolayer. This might be due to the differences in expression of the Coxsackie-adenovirus receptor (CAR), which only could be detected on the RCC. Infection of RCC with a CBV-4 strain did not affect cell viability and did not induce nitric oxide (NO) production alone or with the addition of IFN-γ. This was in contrast to the results obtained with synthetic dsRNA, poly(IC), which induced NO, suggesting that synthetic dsRNA does not mimic enteroviral intermediate dsRNA. During analyses performed with the samples from a family where the mother and one son where diagnosed with T1D on the same day, the results showed that the whole family had a proven EV infection at the time diagnosis. To conclude, the ability of EV strains to replicate in RIN cells is dependent on the growth pattern of the cells and this may be due to the upregulation and/or changed expression pattern of CAR in these cells. In the RIN cells, contrary to artificial dsRNA, viral dsRNA does not induce NO. The isolated EV virus strains used were able to infect and affect human pancreatic islets in vitro. The chemokine RANTES is reduced during an EV infection of human pancreatic islets and NA causes upregulation of RANTES in infected and uninfected islets.

Education for teenagers with diabetes has had limited effects to date, especially regarding metabolic control. The development had been towards more patient-centred approaches, like empowerment, motivational interviews and family behavioural programmes. A patient-centred approach means that the patient is expected to take control of the management of the disease. The empowerment approach has been implemented in adults with diabetes, with some positive results. The aims of this thesis were to evaluate empowerment group education and education in a camp setting, and to validate the “Check your health” instrument which can be used to assess the effects of such education programmes on perceived health and the burden of diabetes. Thirty-two teenagers between 12 and 17 years of age were randomized to either an intervention group or a wait-listed control group. The intervention consisted of six group sessions with an empowerment approach as the theoretical frame. Thirty-one of the teenagers were interviewed two weeks after the empowerment education programme was completed. The programme did not have any positive effect on metabolic control between-groups, but within groups HbA1c and readiness to change increased. According to the definition of empowerment, which pinpoints decision-making, the interviews were analysed with that in focus. In the interviews the teenagers described five categories they perceived as important for decision-making competence: cognitive maturity, personal qualities, experience, social network and parent involvement. Parent involvement was described as both constructive and destructive. These categories were interpreted in the overall theme that “teenagers deserve respect and support for their shortcomings during the maturity process”. Ninety teenagers between 14 and 17 years of age attended education in a camp setting and were compared to a reference group, who had declined participation. The camp did not have any positive effect on metabolic control, but the frequency of insulin pump use after camp education increased. In a cross-sectional comparison, the campers had more positive attitudes towards diabetes and self-care than the non-campers did. The “Check your health” instrument was tested for reliability and validity in 199 teenagers between 12 and 17 years of age. The instrument was found to be reliable and valid for use on a group level in teenagers with diabetes. In conclusion, empowerment education programmes should be tailored to suit young people with diabetes, and should preferably be integrated into routine care. Teenagers who prefer individual education may be offered an individual education plan. Parents should be involved in all education of teenagers with diabetes, with the purpose of increasing teamwork and decreasing control and conflict. Continued assessment of teenagers perceived health and perceived burden of diabetes can be carried out using the “Check your health” instrument.

Syfte: Att undersöka hur kvinnor med diabetes typ-1 upplever informationen före och i samband med graviditet samt omvårdnaden i samband med graviditet. Metod: Semistrukturerade intervjuer med sju kvinnor genomfördes och innehållet analyserades med tematisk analysmetod för att få fram kategorier och subkategorier. Resultat: Majoriteten av kvinnorna fick information gällande graviditet i samband med behandlingen av sin diabetessjukdom av hälsovården innan de blev gravida. Många av kvinnorna upplevde att fokus under graviditeten låg på blodsockerkontrollerna och att barnmorskorna på specialistmödravården fokuserade på diabetessjukdomen och glömde mödravården. Informationen på specialistmödravården upplevdes som bristande eller saknades helt och kvinnorna valde att söka information själva. Omvårdnaden och mottagandet var bra på specialistmödravården fastän det framkom att väntetiderna ibland var långa och att personalen var ostrukturerad. Slutsats: De flesta kvinnorna upplever att informationen på specialistmödravården är bristande eller att den kommer för sent, medan de upplever att omvårdnaden i de flesta fall är bra. Vi anser att det finns utrymme för förbättringar genom att exempelvis utforma en broschyr med information gällande graviditet i samband med diabetes typ-1. Vi tycker att det är viktigt att vårdpersonalen förklarar för kvinnorna hur de kan minimera riskerna och poängtera att många kvinnor med diabetes typ-1 idag får friska barn. / Aim: To investigate how women with diabetes type-1 experienced information before and during pregnancy and nursing care during pregnancy. Method: Semi-structured interviews with seven women were carried out and the content was analyzed with a thematic analysis to generate categories and subcategories. Result: The majority of the women received information about pregnancy associated with diabetes type-1 in health care before they became pregnant. Many women felt that the focus during pregnancy was on blood sugar control and that the focused on diabetes and forgot about maternity care. The information received was seen as inadequate or missing entirely, and the women chose to seek information themselves. The care was good although it appeared that the waiting times were long and the staff was unstructured. Conclusion: Most women feel that the information is inadequate or that it comes too late, while they experience that the care in most cases is good. We believe that there is room for improvement, for example by designing a brochure with information regarding pregnancy associated with diabetes type-1. We think it is important that health professionals explain to women how they can minimize risks and point out that many women with diabetes type-1 have healthy children.

Studiens syfte var att beskriva upplevelse av hälsa och välbefinnande hos vuxna personer med Typ 1 Diabetes Mellitus (T1DM). Jämförelser gjordes även mellan kön och åldrar. Urvalet gjordes från medlemmar i Gävleborgs Diabetesförbund och bestod utav 56 personer (kvinnor n 30, män n 26) med ett åldersspann mellan 19-76. Använd datainsamlingsmetod var en förkortad och översatt version av mätinstrumentet Ryff’s Psychological Well being Scale. Enkäten bestod utav 18 frågor inom de 6 dimensioner som instrumentet består utav: Självacceptans, positiva relationer med andra, autonomi, levnadskontroll, meningen med livet samt personlig utveckling. Resultatet visade att gruppen T1DM skattade sig generellt högt inom hälsa och välbefinnande men lägst inom dimensionen autonomi och högst inom positiva relationer med andra. Statistisk signifikans framkom vid jämförelse av åldersgrupperna 19-39 och 40-60 inom dimensionerna meningen med livet samt personlig utveckling där gruppen 19-39 skattade sig högst inom båda dimensionerna. Statistisk signifikans kunde även påvisas vid jämförelse av samtliga 3 åldersgrupper inom dimensionen meningen med livet där gruppen 19-39 skattade sig högst och 61+ lägst. Slutligen kunde statistisk signifikans påvisas mellan könen inom dimensionen positiva relationer med andra där kvinnorna skattade sig högre än männen. Vidare forskning behövs inom området som rör personer med T1DM för att förbättra allmän hälsa och välbefinnande. / The aim of this study was to describe the experience health and well-being among adults with type 1 diabetes mellitus (T1DM). Comparison was made regarding sex and age. The sample was taken from Gävleborgs Diabetesförbund and consisted of 56 persons (women n 30, men n 26) aged between 19 and 76. Health and well-being was measured with a shortened translated 18 item version of the instrument Ryff’s Psychological Well being Scale. The 18 items describe the 6 dimensions of the instrument which it is built on: self-acceptance, positive relationships with others, autonomy, environmental mastery, purpose in life and personal growth. The result shows that the group T1DM estimated themselves generally high in health and well-being but lowest in the dimension autonomy and highest in positive relations with others. Statistical significance was shown in comparison between age groups 19-39 and 40-60 in the dimensions purpose in life and personal growth, where the group 19-39 estimated themselves highest in both dimensions. Statistical significance was also demonstrated in comparison between all age groups in the dimension purpose in life, where the group 19-39 estimated themselves highest and 61+ lowest. Finally, statistical significance was shown between the sexes in the dimension positive relations with others, where the women estimated themselves higher than the men. Further research is needed in the area that involves persons with T1DM to improve their general health and well-being.

In this thesis interactions in complex fluids, such as serum and meat juice, were analysed with optical biosensor techniques. Panels of lectins immobilised on gold surfaces were used for investigation of differences in protein glycosylation pattern in sera and meat juices between various species. The present panel was also used for investigation of global glycosylation changes of serum proteins in type 1 diabetes patients. Biorecognition was evaluated with null ellipsometry and scanning ellipsometry combined with multivariate data analysis techniques (MVDA). Principal component analysis (PCA) showed that the lectin panel enabled discrimination between sera from the different species as well as for the different meat juices. The results also indicate that there is a measurable global alteration in glycosylation pattern of serum proteins in type 1 diabetic patients compared to healthy subjects. Using an artificial neuronal net (ANN), it was also possible to correctly categorise unknown serum samples into their respective class or group. The analytical potential of combining information from lectin panels with multivariate data analysis was thereby demonstrated. Also, a sensitive and specific method based on surface plasmon resonance (SPR) for detection of insulin autoantibodies (IAA) in serum samples from individuals at high risk of developing type 1 diabetes (T1D) has been developed. When measuring trace molecules, such as autoantibodies, in undiluted sera with label-free techniques like SPR, non-specific adsorption of matrix proteins to the sensor surface is often a problem, since it causes a signal that masks the analyte response. The developed method is an indirect competitive immunoassay designed to overcome these problems. Today, IAA is mainly measured in radio immunoassays (RIAs), which are time consuming and require radioactively labelled antigen. With our SPR-based immunoassay the overall assay time is reduced by a factor of >100 (from 4 days to 50 min), while sensitivity is maintained at a level comparable to that offered by RIA. Finally, the assay was used in a screening study of newly diagnosed type 1 diabetes patients and non-diabetic subjects.

Type 1 Diabetes (T1D) is an autoimmune disorder caused by both genetic and environmental factors. The non-obese diabetic (NOD) mouse is one of the best and most commonly studied animal models for T1D. This mouse strain spontaneously develops diabetes through a process that closely resembles the human pathogenesis. More than 20 insulin dependent susceptibility (Idd) loci have been identified in the NOD mouse, contributing to disease susceptibility; however, the contribution of each of the various factors to disease pathogenesis is largely unknown. The aim of this thesis was to identify and functionally characterize candidate genes mediating susceptibility to murine T1D. Cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) is a negative regulator of T-cell activation and has been shown to be associated with autoimmune diseases. Genetic analyses of the NOD mouse have identified the Ctla-4 gene as a major candidate for the Idd5.1 diabetes susceptibility locus and NOD mice have been found to display an impaired expression of CTLA-4 upon anti-CD3 stimulation in vitro. In Paper I, we showed that a novel locus (Ctex) in the distal part of the chromosome 1 together with the Idd3 (Il-2) locus on chromosome 3, constitute the major factors conferring the observed difference in CTLA-4 expression levels. Moreover, we also demonstrated that the defective expression of CTLA-4 in NOD T-cells can in part be overcome by the addition of exogenous interleukin-2 (IL-2). In Paper II, using congenic mice, we confirmed that the Ctex locus contributes to decreased expression of CTLA-4 observed in NOD mice and restricted the region of interest to a 28.8 Mb region containing the Cd3ζ gene. We also demonstrated a phenotypic correlation between strains carrying the NOD versus C57BL/6 alleles of Cd3ζ, respectively and showed that expression of CD3ζ is impaired in activated NOD CD4+ T cells. The NOD allele of the Cd3ζ region was found to confer impaired T cell activation and the defective CD3 signalling could be surpassed by PMA plus ionomycin stimulation supporting the notion of CD3ζ as a prime candidate gene for Ctex. NOD lymphocytes display relative resistance to various apoptosis-inducing signals, which have been proposed to contribute to the pathogenesis of diabetes. Resistance to dexamethasone-induced apoptosis in NOD immature thymocytes has been mapped to the Idd6 locus. In Paper III we restricted the Idd6 locus to an 8 cM region on the telomeric end of chromosome 6 using a set of congenic mice. In addition, we could confirm that the Idd6 region controls apoptosis resistance in immature thymocytes and restricted the control of apoptosis resistance to a 3 cM region within the Idd6 locus. In Paper IV, we further restricted the Idd6 locus to a 3 Mb region and excluded the region controlling the resistance to apoptosis as directly mediating susceptibility to diabetes. We also showed that defective expression of the Lrmp/Jaw1 gene, encoding an endoplasmatic reticulum resident protein, is controlled by the Idd6 locus making it the prime candidate for Idd6.  Together, these results contribute to the identification and functional characterization of candidate genes that may confer susceptibility to T1D in the NOD mouse. These results offer important insights into the pathophysiological processes underlying this disease.

Background/aim. The incidence of microvascular complications has been reported to be unchanged the last decades. However, in randomized clinical trials it has been shown that improved metabolic control can reduce the development of long term complications. It has been debated whether it is possible to achieve the same results in an unselected population. In a previous study we found a decreased incidence of overt nephropathy, but unchanged incidence of severe laser treated retinopathy in a population of patients with type 1 diabetes diagnosed in childhood. The aim of the present study was to investigate the incidence 10 years later in the same population and to analyse the importance of possible risk factors. In another previous study we found a high prevalence of subclinical neuropathy among young diabetic patients despite intensive insulin therapy since diagnosis. The aim of the present study was to examine if intensive treatment is more effective in preventing early diabetic complications other than neuropathy. The incidence of type 1 diabetes has doubled in Sweden the last decades. The reason must be environmental factors. These, as well as more intensive insulin regimens from onset of diabetes, might also lead to different disease process. We wanted to analyse if clinical characteristics at onset had changed the last 25 years and if there was any secular trend of C-peptide secretion. We also intended to investigate if longer persistence of C-peptide secretion could be of importance for prevention of long term complications. Methods. The whole study population consisted of all 478 patients with type 1 diabetes diagnosed before the age of 15 during the years 1961 - 2000, living in the catchment area of the Paediatric Clinic, University Hospital, Linköping, Sweden. For the statistical analysis the population was divided into five–year cohorts according to time of onset of diabetes. The cumulative proportion of severe retinopathy and overt nephropathy in 269 patients with onset of diabetes 1961 - 1985 was computed with survival analysis. Multivariable regression models were used to analyse the importance of metabolic control, diabetes duration, blood pressure, smoking, BMI, lipids and persisting C-peptide secretion. The prevalence of all grades of retinal changes, nephropathy and neuropathy, defined as abnormal nerve conduction, was estimated in the late 1990s in a subgroup of 80 children and adolescents with mean 13 years of diabetes duration. Clinical characteristics at onset, duration of partial remission and regularly measurements of fasting and stimulated C-peptide secretion the first five years after onset were analysed in 316 patients with onset of diabetes 1976 - 2000. Results. The cumulative proportion of severe laser treated retinopathy showed a significant declining trend the last decades. The decrease was significant between the oldest cohort with diabetes onset 1961 - 1965 and the cohorts with diabetes onset 1971 - 1975 and 1976 - 1980. The cumulative proportion of overt nephropathy also declined with a significant decrease between the oldest cohorts and all the following cohorts. After 25 years of diabetes duration it was 30% and 8% in the two oldest cohorts respectively and remained largely unchanged after 30 years. Diabetes duration and long term HbA1c were the only significant independent risk factors for both retinopathy and nephropathy. The risk of overt nephropathy increased substantially when HbA1c was above 8.5%, while the risk of severe retinopathy increased already when HbA1c exceeded 7.5%. The prevalence of neuropathy was 59%, of retinopathy 27% and of nephropathy 5% in the population of young patients after mean 13 years of diabetes duration. During the last 25 years the clinical characteristics at onset were unchanged as well as duration of partial remission and magnitude and persistence of C-peptide secretion. Conclusions. In this unselected population the cumulative proportion of severe retinopathy and overt nephropathy decreased over the last decades. Diabetic nephropathy has probably been prevented and not just postponed. Good glycaemic control was the most important factor to avoid complications, with the necessity of a lower level of HbA1c to escape retinopathy than nephropathy. Intensive insulin regimens from diabetes onset was not sufficient to entirely escape early diabetic complications after mean 13 years of diabetes duration, even if the prevalence of retinopathy and especially nephropathy was lower than usually reported. The clinical picture at onset of diabetes was unchanged the last 25 years. There was no secular trend of partial diabetes remission or C-peptide secretion during the first years after diagnosis.

Background: Type 1 diabetes is a chronic disease for which there is currently no cure, and high quality care is essential if acute and long-term complications are to be avoided. Many children and adolescents have inadequate metabolic control with increased risk for complications later in life, and adolescent girls have reported low quality of life. Differences in metabolic control between treatment centres have been found but the reasons for this are unclear. Diabetes is a largely self-managed disease. Patient education is central to successful self-management but little is known about how to make best use of diabetes communities on the Internet and integrate them into a practitioner-driven service. Aim: The main objective of this thesis was to gain better understanding of how to improve the quality of diabetes care for children and adolescents, aiming to have near-normal blood glucose, to prevent both acute and late complications and to have good quality of life. Methods: The geographic populations of two paediatric centres (n=400) received validated questionnaires on perceived quality of care and Health-Related Quality of Life (HRQOL). An intervention with a web portal containing diabetes-related information and social networking functions was carried out within the same population. Clinical variables from 18 651 outpatient visits registered in the Swedish paediatric diabetes quality registry, SWEDIABKIDS were analysed. Using data from SWEDIABKIDS, five centres with the lowest mean HbA1c, five with the highest, and five with the largest decrease in centre mean HbA1c between 2003 and 2007 were identified. Team members (n=128) were asked about structure, process, policy, and the messages given to patients about important diabetes issues. Results: Specific areas that were identified as needing improvement included information about self-care, waiting time at outpatient clinics and for treatment, and access to care. Diabetes seemed to reduce HRQOL. Subjects with better metabolic control and with higher frequency of injections reported slightly higher HRQOL, as did those living with both parents compared to those with separated parents. Only 35% of children and adolescents with diabetes in Sweden had an HbA1c level below the treatment target value. Mean HbA1c showed a correlation with mean insulin dose, diabetes duration, and age. A difference between centres was found, but this could not be explained by differences in insulin dose, diabetes duration, or age. Adolescent girls reported lower HRQOL, as did parents of girls aged < 8 years. Girls also had poorer metabolic control, especially during adolescence. In teams with the lowest and the most decreased mean HbA1c, members gave a clear message to patients and parents and had a lower HbA1c target value. Members of these teams appeared more engaged, with a more positive attitude and a greater sense of working as a team. Members of teams with the highest mean HbA1c gave a vaguer message, felt they needed clearer guidelines, and had a perception of poor collaboration within the team. High insulin dose, large centre population, and larger teams also seemed to characterize diabetes centres with low mean HbA1c. The most frequently visited pages on the web portal were the social networking pages, such as blogs, stories and discussions, followed by the diabetes team pages. Those who used the portal most actively were younger, had shorter diabetes duration, and lower HbA1c, and were more often girls. The web portal was not found to have any significant beneficial or adverse effects on HRQOL, empowerment or metabolic control. Conclusions: The quality of diabetes care for children and adolescents in Sweden is not sufficiently good and needs to improve further if complications in later life are to be avoided. Psychosocial support for children and adolescents with diabetes should be appropriate for age and gender. The attitudes of the members in the diabetes care team and the message they give to patients and their parents seem to influence metabolic control in children and adolescents. A clear and consistent message from a unified team appears to have beneficial effects on metabolic control. A web portal that includes comprehensive information about diabetes, and the opportunity to communicate with other people with diabetes and with healthcare professionals may be a useful complement to traditional patient education tools. Members of the diabetes team should encourage its use.

Syftet med detta arbete är att undersöka om det föreligger någon skillnad mellan hur personer med diabetes typ 1 med MDI- respektive IP-behandling skattar sin hälsorelaterade livskvalitet, behandlingstillfredsställelse och uppfattning av behandlingsformens påverkan på livskvaliteten. Vidare är syftet att undersöka om det föreligger någon skillnad mellan män och kvinnor, samt mellan kvinnor respektive män i MDI- respektive IP-gruppen, avseende hälsorelaterade livskvalitet, behandlingstillfredsställelse och uppfattning av behandlingsformens påverkan på livskvaliteten. Som instrument användes enkäten SF-36 samt en egenkonstruerad enkät med bakgrundsfrågor rörande skattning av behandlingstillfredsställelse och uppfattning av behandlingsformens påverkan på livskvaliteten. I MDI-gruppen deltog 40 deltagare och i IP-gruppen 37 deltagare. Studien har en deskriptiv jämförande design. Resultatet visade att det inte finns någon signifikant skillnad mellan MDI- och IP-gruppens skattning av hälsorelaterad livskvalitet. Skillnader kunde påvisas mellan behandlingsformerna gällande behandlingstillfredsställelse och uppfattningen av behandlingsformens påverkan på livskvalitet. IP-gruppen har en signifikant högre behandlingstillfredsställelse samt anser att behandlingsformen påverkar deras livskvalitet mer än MDI-gruppen. Kvinnor i MDI-gruppen skattade sin hälsorelaterade livskvalitet signifikant lägre än män i MDI-gruppen, ingen skillnad kunde dock påvisas mellan männen och kvinnorna i IP-gruppen. Männen i MDI-gruppen skattar sin hälsorelaterade livskvalitet signifikant högre än männen i IP-gruppen. Ingen skillnad kunde påvisas mellan männen i de båda grupperna gällande behandlingsstillfredsställelse eller uppfattningen av behandlingsformens påverkan på livskvaliteten. Kvinnorna i IP-gruppen skattade signifikant högre gällande behandlingsstillfredsställelse och uppfattningen av behandlingsformens påverkan på livskvaliteten än kvinnorna i MDI-gruppen. Kvinnorna i IP-gruppen har likartad hälsorelaterad livskvalitetsskattning som männen i samma grupp och har även en högre behandlingstillfredsställelse än kvinnor i MDI-gruppen. Slutsatsen kan vara att IP är en lämplig behandling för kvinnor