The process of β-cell destruction that leads to type 1 diabetes (T1D) is incompletely understood and it is believed to be a result of both genetic and environmental factors. Enterovirus (EV) infections of the β-cells have been proposed to be involved, however, the effects of EV infections on human β-cells have been little investigated. This thesis summarises studies of three different Coxsackie B4 virus strains that have previously been shown to infect human islets. The effects of infections with these EV were studied in vitro in human islets and in a rat insulin-producing cell line. In addition, a pilot study was performed on isolated human islets to investigate the ability to treat such infections with an antiviral compound. It was found that one of the virus strains replicated in human β-cells without affecting their main function for at least seven days, which in vivo may increase a virus’s ability to persist in islets. Nitric oxide was induced by synthetic dsRNA, poly(IC), but not by viral dsRNA in rat insulinoma cells in the presence of IFN-γ, suggesting that this mediator is not induced by EV infection in β-cells and that poly(IC) does not mimic an EV infection in this respect. All three virus strains were able to induce production of the T-cell chemoattractant interferon-γ-inducible protein 10 (IP-10) during infection of human islets, suggesting that an EV infection of the islets might trigger insulitis in vivo. Antiviral treatment was feasible in human islets, but one strain was resistant to the antiviral compound used in this study. To conclude, a potential mechanism is suggested for the involvement of EV infections in T1D. If EV infections induce IP-10 production in human islet cells in vivo, they might recruit immune cells to the islets. Together with viral persistence and/or virus-induced β-cell damage, this might trigger further immune-mediated β-cell destruction in vivo.

Type 1 diabetes (T1D) is a chronic disease caused by destruction of the insulin producing beta-cells in the pancreas. The incidence of T1D has increased rapidly, especially in the Western world and among young children. The pathogenesis of T1D is not fully understood, but the beta-cells are believed to be destroyed by an autoimmune process initiated years before the onset of T1D. During this pre-clinical period, autoantibodies to insulin (IAA), glutamic acid decarboxylase (GADA) and the tyrosine phosphatase-like protein IA-2 (IA-2A) can be detected and are used to identify individuals at risk of T1D. The major genetic determinant for T1D is the HLA class II genes, but also polymorphism in the insulin gene and CTLA-4 gene are associated with T1D. The risk genes cannot explain the rapid increase in incidence of T1D, therefore a role for different environmental factors has been suggested. The aim was to study the prevalence of beta-cell autoantibodies in children from the general population in relation to known genetic and environmental risk factors, and in young patients with T1D in high and low incidence areas. Short duration of breast-feeding was associated with an increased risk of developing beta-cell autoantibodies in children from the general population at 5-6 years of age. We found an association between positivity for GADA and/or IAA at the age of 5-6 years and a short duration of total breastfeeding, and also between positivity for GADA, IA-2A and/or IAA and a short duration of exclusive breast-feeding. Our findings suggest that breast-feeding has a long term protective effect on the risk of beta-cell autoimmunity in children from the general population. The T1D related risk genes were not associated with beta-cell autoantibodies other than GADA in children from the general population at 5-6 years of age. Children with the DR4-DQ8 haplotype were more often positive for GADA than children without this haplotype. We found no association of GADA with DR3-DQ2 haplotype or between these two haplotypes and any of the other autoantibodies. Our results suggest that beta-cell autoimmunity in children from the general population is not strongly associated with any risk genes of T1D other than DR4-DQ8. In the non-diabetic children with allergic heredity GADA was detectable in almost all children, IA-2A in about half and IAA in 10% of the children. The levels low of these autoantibodies fluctuated with age and different patterns of fluctuations were seen for GADA and IA-2A, which may reflect differences in the immune response to the autoantigens. In patients with newly diagnosed T1D, we found some differences between patients from a high incidence country (Sweden) and a country with a lower incidence (Lithuania). Among the Swedish patients, the prevalence of IAA and GADA or multiple autoantibodies was higher than in Lithuanian patients. The risk genes DR4-DQ8 and the heterozygous high risk combination DR4-DQ8/DR3-DQ2 was more common among the Swedish patients than Lithuanian patients. Patients with low levels of IAA had higher levels of HbA1c and ketones, indicating that patients without IAA or with low levels of IAA have a more severe onset of T1D. Our findings indicate that beta-cell autoimmunity is more pronounced in a high incidence area compared to an area with a lower incidence. In conclusion, short duration of breast-feeding is a risk factor for beta-cell autoantibodies in children from the general population, and the beta-cell autoantibodies in these children are not associated with specific risk genes. Children with newly diagnosed T1D in a high incidence area carry risk genes and have autoantibodies more often than newly diagnosed children from an area with a lower incidence, perhaps indicating different disease phenotypes.

Diabetic long-term complications, despite intensive treatment, cause serious handicaps at relatively young age in diabetic patients. Diabetic nephropathy (DN) develops in up to 30% of patients with type 1 diabetes (T1D). Besides the eventual loss of kidney function, with need for dialysis treatment and transplantation, this complication also increases the risk of early death from cardiovascular disease. In addition to hyperglycaemia, the risk of developing DN is influenced by a number of life-style related factors, such as smoking and diet, but the mechanisms of action of these factors are largely unknown. The incidence of DN is not linearly related to diabetes duration. There is a peak incidence of DN at 15-20 years and this, together with results from family studies, shows that genetic factors are important contributors. Possible candidate genes are those involved in regulation of intraglomerular pressure and blood pressure, oxidative stress and inflammation. The main aims of this thesis were: ● To investigate the risk of DN associated with polymorphisms in; A) the endothelial NO-synthase gene (NOS3) and genes in the renin-angiotensin-system (RAAS) (all involved in the regulation of intraglomerular pressure). B) the manganese superoxide dismutase gene (SOD2) (involved in the regulation of oxidative stress). C) the ICAM1 gene (involved in activation and migration of lymphocytes) ● To investigate gene-smoking interactions ● To investigate the influence of normal diet on risk of microalbuminuria. The aims were addressed in different case-control settings, including 347 T1D patients from Sweden and 1163 patients from Finland, with or without DN, defined as; overt DN – having albumin excretion rate (AER) ≥200 μg/min, incipient DN – AER between 20 and 200 μg/min, non-DN controls – having AER <20 μg/min and at least 20 years of diabetes duration. In one study also non-diabetic healthy individuals were included to asses the risk of T1D associated with the ICAM1 gene. Results: The RAAS genes were investigated in the Swedish sample set and there was an association between a polymorphism in the angiotensin II type 1 receptor (AGTR1) gene and overt DN, when adjusting for age, duration of diabetes, HbA1c, sex and smoking (adjusted OR=3.04, 99% CI=1.02-9.06). Also a synergistic interaction with smoking was indicated. The ICAM1 gene was investigated in the Swedish sample set, but no association with DN was found. There were, however, associations between T1D and two polymorphisms in this gene, rs281432 (OR=1.64, 95% CI=1.14-2.38) and rs5498 (OR=2.46, 95% CI=1.59-3.80). In the combined Swedish/Finnish sample set, the Glu/Glu genotype of the Glu298Asp polymorphism in the NOS3 gene was associated with DN when age at diabetes onset, duration of diabetes, HbA1c, blood pressure, sex and smoking were taken into account (adjusted OR=1.46, 95% CI=1.12-1.91). There was also association between a polymorphism in the MnSOD gene and DN in this sample set. Homozygosity for the valine-allele of the Val16Ala polymorphism was associated with increased risk of DN in a model including age at diabetes onset, duration of diabetes, HbA1c, sex and smoking (adjusted OR=1.32, 95% CI=1.00-1.74). Smoking was associated with DN (OR=2.00, 95% CI=1.60-2.50) and in the Swedish sample set there were indications of interactions between smoking and the NOS3 and SOD2 genes, but these results could not be confirmed in the Finnish sample set. A high protein intake can enhance glomerular filtration rate and accelerate progression to DN, also other dietary components such as fat, fibres, vitamins and the ratio red/white meat have been discussed as important for DN development. In a nested case-control study including young T1D patients, the normal dietary intakes of protein and other nutrients were assessed using a semiquantitative questionnaire. The results showed that T1D patients consuming more than 6.5 g fish protein (>75th percentile) per day were at slightly lower risk to have microalbuminuria in both crude (OR=0.49, 95% CI=0.25-0.97) and adjusted analyses (OR=0.26, 95% CI=0.09-0.76, adjusted for age, duration of diabetes, sex, HbA1c, mean arterial pressure, BMI, region, smoking, energy intake and fish fat intake). Conclusions: The risk of having diabetic nephropathy is influenced by at least two genes controlling blood pressure and one gene protecting against oxidative stress. Smoking also increases the risk of DN and our findings indicate that smoking may accentuate the effect of the AGTR1, NOS3 and SOD2 genes. Normal dietary intake of protein was not associated with risk of having microalbuminuria in young T1D patients, on the other hand, an intake of fish protein above the 75th percentile decreased the risk of microalbuminuria.

Type 1 diabetes is an autoimmune disorder determined both by genetic and environmental factors. The Non-obese diabetic (NOD) mouse is one of the best animal models of this disease. It spontaneously develops diabetes through a process resembling the human pathogenesis. The strong association of NOD Type 1 diabetes to the MHC region and the existence of other diabetes susceptibility loci are also in parallel with the human disease. The identity of the genetic factors and biological function mediated by these loci remain, however, largely unknown. Like in other autoimmune diseases, defects in tolerance mechanisms are thought to be at the origin of type 1 diabetes. Accordingly, defects in both central and peripheral tolerance mechanisms have been reported in the NOD mouse model. Using a subphenotype approach that aimed to dissect the disease into more simple phenotypes, we have addressed this issue. In paper I, we analyzed resistance to dexamethasone-induced apoptosis in NOD immature thymocytes previously mapped to the Idd6 locus. Using a set of congenic mice carrying B6-derived Idd6 regions on a NOD background and vice-versa we could restrict the Idd6 locus to an 8cM region on the telomeric end of chromosome 6 and the control of apoptosis resistance to a 3cM region within this area. In paper II, further analysis of diabetes incidence in these congenic mice separated the genes controlling these two traits, excluding the region controlling the resistance to apoptosis as directly mediating susceptibility to diabetes. These results also allowed us to further restrict the Idd6 locus to a 3Mb region. Expression analysis of genes in this chromosomal region highlighted the Lrmp/Jaw1 gene as a prime candidate for Idd6. Lrmp encodes an endoplasmatic reticulum resident protein. Papers III and IV relate to peripheral tolerance mechanisms. Several T cell populations with regulatory functions have been implicated in type 1 diabetes. In paper III, we analyzed NOD transgenic mice carrying a diverse CD1d-restricted TCR αVa3.2b9), named 24abNOD mice. The number of nonclassical NKT cells was found to be increased in these mice and almost complete protection from diabetes was observed. These results indicate a role for nonclassical NKT cells in the regulation of autoimmune diabetes. In paper IV, we studied the effects of introducing the diverse CD1d-restricted TCR (Va3.2b9) in immunodeficient NOD Rag-/- mice (24abNODRag-/- mice). This resulted in a surprising phenotype with inflammation of the ears and augmented presence of mast cells as well as spleenomegaly and hepatomegaly associated with extended fibrosis and increased numbers of mast cells and eosinophils in the tissues. These observations supported the notion that NKT cells constitute an “intermediary” cell type, not only able to elicit the innate immune system to mount an inflammatory response, but also able to interact with the adaptive immune system affecting the action of effector T cells in an autoimmune situation. In this context the 24abNODRag-/- mice provide an appropriate animal model for studying the interaction of NKT cells with both innate and adaptive components of the immune systemα.

This thesis deals with the design of a robust and safe control algorithm to aim at an artificial pancreas. More precisely we will be interested in controlling the stabilizing part of a classical cure. To meet this objective, the design of a robust nonlinear model predictive controller based on the solution of a saddle point optimization problem is considered. Also, to test the controller performances in a realistic case, numerical simulations on a FDA validated testing platform are envisaged.In a first part, we present an extension of the usual nonlinear model predictive controller designed to robustly control, in a sampled-data framework, systems described by nonlinear ordinary differential equations. This controller, which computes the best control input by considering the solution of a constrained saddle point optimization problem, is called saddle point model predictive controller (SPMPC). Using this controller, it is proved that the closed-loop is Ultimately Bounded and, with some assumptions on the problem structure, Input-to State practically Stable. Then, we are interested in numerically solving the corresponding control problem. To do so, we propose an algorithm inspired from the augmented Lagrangian technique and which makes use of adjoint model.In a second part, we consider the application of this controller to the problem of artificial blood glucose control. After a modeling phase, two models are retained. A simple one will be used to design the controller and a complex one will be used to simulate realistic virtual patients. This latter is needed to validate our control approach. In order to compute a good control input, the SPMPC controller needs the full state value. However, the sensors can only provide the value of blood glucose. That is why the design of an adequate observer is envisaged. Then, numerical simulations are performed. The results show the interest of the approach. For all virtual patients, no hypoglycemia event occurs and the time spent in hyperglycemia is too short to induce damageable consequences. Finally, the interest of extending the SPMPC approach to consider the control of time delay systems in a sampled-data framework is numerically explored.

Healthy term pregnancy requires precisely timed coordination of multiple systems, including reproductive, neuroendocrine, immune and cardiovascular. Dynamic maternal alterations occur systemically as well as locally within the reproductive tract. Systemic cardiovascular changes during gestation are relatively conserved in mammals, permitting comparison. These physiological changes are relatively acute and reversible, in contrast to the pathological changes seen during cardiovascular disease development. Gestational hypertensive disorders, such as preeclampsia, are the leading causes of maternal and fetal morbidity and mortality. The pathogenesis of preeclampsia is not fully elucidated, but perturbation of the immune system is a fundamental component. The angiogenic and vascular properties of uterine NK lymphocytes have been well studied in mice and women, but their relationships to gestational blood pressure regulation and cardiovascular adaptations have not been addressed. In non-pregnant women and mice, T cells, but not B cells, have been found to alter cardiovascular functioning. NK cells in humans also possess these capabilities, but no functional studies have been completed. The aim of this thesis was to define the role of NK and T lymphocytes in cardiovascular adaptations during mouse gestation. Using chronic radiotelemetry, histology, post-mortem and other techniques, female inbred mice of differing genotypes that lack specific lymphocyte subsets were compared before and across gestation. In normal, immune competent mice, a five-phase gestational blood pressure profile was found. This dynamic profile corresponded to stages of placental development. In mice with a compound deficit in arterial modification and lymphocytes, no gestational hypertension was observed. To elevate the maternal challenge of pregnancy, studies of pregnant, autoimmune Type 1 Diabetic mice were conducted. Impaired spiral artery remodeling, dysfunctional lymphocytes and growth-restricted fetuses were identified. From mid-gestation, diabetic pregnant mice were hypotensive and bradycardic and showed signs of pre-renal failure (proteinuria and electrolyte imbalances). In pregnant mice lacking T cells, tachycardia was observed despite otherwise normal gestational outcomes. In pregnant mice lacking T cells with impaired NK cells, blood pressure was blunted and tachycardia was observed. These findings support the conclusion that impaired spiral artery remodeling is insufficient to cause gestational hypertension in mice. The data further identify a role for T and NK cells in cardiac function during gestation. / Thesis (Ph.D, Anatomy & Cell Biology) -- Queen's University, 2010-09-01 20:56:15.648

Abstract   Aims and objectives. To examine whether an intervention with carbohydrate- counting at onset of type 1 diabetes in children, has had any effect on metabolic indicators such as HbA1c , BMI-sds and total daily insulin / kg, 2 months and 1 year after onset by age and gender .Background. The pediatric department at the university hospital of Norrland in Umeå (NUS), has since several years a higher average HbA1c among children 0-17 years, compare with pediatric departments in the rest of Sweden.  Therefore, in autumn 2011, an intervention was made to teach children with diabetes and their parents to count carbohydrates from the onset of diabetes. Using this method they can regulate their insulin doses depending on the amount of carbohydrates they are eating. Design. This is an empirical registry study done on the basis of a quantitative method. It is retrospective controlled non- randomized with comparison group .Methods. The study included 46 children aged 0-17 years diagnosed with type 1 diabetes. Those who fell ill in 2009 (n=22) did not learn to use carbohydrate counting and served as a control group. The intervention group were diagnosed 2011-2012, (n=24), began using carbohydrate counting at the onset of diabetes.Results. The study shows with significant security that carbohydrate counting lowers HbA1c two months and one year after onset of type 1 diabetes. No significant difference was found in BMI-sds. The study shows no significant difference in HbA1c between the sexes.Conclusion.  Carbohydrate-counting from onset of diabetes gives a significant reduction of HbA1c without affecting BMI-sds significantly.Relevance to clinical practice. It is important that health professionals continue to teach and support families in carbohydrate counting from the onset of diabetes.Keywords. children, HbA1c, BMI- sds , type 1 diabetes , carbohydrate counting

Summary: Background: Type 1 diabetes is a chronic disease and Sweden tops the statistics for the worst affected countries in the world. The main part of treatment takes place alone in the home when the sufferer administers insulin to the body via an injection or pump. The sufferer must adapt their lifestyle to supplying

INKT cells are non-conventional T lymphocytes that are restricted to glycolipid presenting CD1d molecule. iNKT cells express an invariant TCR a chain (Va14-Ja18 in mice and Va28-Ja18 in humans). Their particularity is to rapidly produce copious amounts of cytokines (IFN-? and IL-4) after activation and to activate other cells of the immune system such as dendritic cells, NK cells and T lymphocytes. iNKT cells, therefore, form a bridge between innate and adaptive immune responses. Type 1 diabetes is an autoimmune disease characterized by the destruction of pancreatic ß cells whose role is to produce insulin. While diabetes development can clearly be associated with genetic polymorphisms, environmental factors were also implicated in the etiology of the disease. Numerous studies suggest that viral infections, particularly infections with Coxsackievirus B4 (CVB4), could be implicated in the development of type 1 diabetes. Our study was performed with NOD mice that develop type 1 diabetes around 15 weeks of age and with proinsulin 2 knockout NOD mice (Pro-ins2-/-) which become diabetic around 8 weeks of age. Our results show that CVB4 infection induces accelerated diabetes in around half of NOD and Pro-ins2-/- mice compared to uninfected mice. However, the activation of iNKT cells with their agonist, aGalactosylceramide (aGalCer), at the time of infection greatly decreases diabetes incidence. CVB4 infection induces a strong recruitment of macrophages into the pancreas. Interestingly, iNKT cell activation modifies the function of these macrophages. Indeed, pancreatic macrophages of CVB4 infected mice strongly express IL-1, IL-6 and TNF-a, indicating their pro-inflammatory character. On the contrary, macrophages of mice infected with CVB4 and treated with aGalCer express low levels of these cytokines, but strong levels of suppressive enzymes iNOS (inducible NO synthase), IDO (Indoleamine 2,3-dioxygenase) and arginase I. The use of inhibitors of these enzymes showed that diabetes prevention is induced by IDO. We have also observed that autoreactive T cells strongly infiltrate the pancreatic islets after CVB4 infection. It is interesting to note that the high diabetes incidence of CVB4 infected mice is associated with an increased frequency of IFN-? producing autoreactive T cells in pancreatic islets. On the contrary, the frequency of these cells is very low in infected mice treated with aGalCer. The inhibition of IFN-? production is dependent on IDO enzyme, since the use of its inhibitor strongly increases IFN-? production by anti-islet T cells and diabetes incidence. To summarize, our results show that iNKT cell activation during the infection with CVB4 induces immunosuppressive macrophages in the pancreas. These cells inhibit the function of autoreactive T cells and prevent diabetes development.

Diabetes mellitus typ 1 är en kronisk sjukdom och en av de vanligaste barnsjukdomarna som påverkar det endokrina- och metaboliska systemet. I dagsläget beräknas 497 100 barn i världen ha sjukdomen. När ett barn diagnostiseras får föräldrarna ett stort ansvar då sjukdomen kan vara livshotande om inte behandlingen sköts på rätt sätt. Syftet med denna litteraturstudie var att beskriva föräldrars upplevelser när deras barn diagnostiseras med diabetes mellitus typ 1. En transitionsteori användes för att strukturera föräldrars upplevelser i tre faser. Resultatet visade att föräldrarna upplevde det påfrestande att barnen fick sjukdomen. Det visade sig att det inte bara var de behandlingsrelaterade faktorerna som påverkade deras vardag. Det handlade även om förändring av relationer, ökad oro och svårigheter att lita på att andra tog ansvar. I slutet av föräldrarnas transitionsprocess var barnen gamla nog att ta över ansvaret själva. Föräldrarna behövde släppa kontrollen och låta barnen vara självständiga vilket var svårt för dem. Det är viktigt att forskning i ämnet kontinuerligt förnyas då föräldrarna behöver mycket stöd i denna svåra situation. Sjuksköterskan bör vara väl införstådd i upplevelser som kan förekomma hos föräldrar för att på så sätt kunna utveckla sin förmåga i att stötta dem i deras dagliga diabetesvård. / Type 1 diabetes mellitus is a chronic disease and one of the most common childhood illnesses that affect the endocrine- and metabolic system. In the current situation estimated 497 100 children worldwide have the disease. When a child is diagnosed, parents get a big responsibility since the disease can be fatal if treatment is not done properly. The purpose of this study was to describe parents’ experiences when their children get diagnosed with type 1 diabetes mellitus. A transition theory was used to structure the parents’ experiences in three phases. The results showed that parents experienced pain when their children got the disease. It turned out that it was not only the treatment-related factors that affected their daily lives. It was also about changes in relationships, increased anxiety and difficulty in trusting others to take responsibility. At the end of the parental transition process, the children were old enough to take over the responsibility themselves. Parents needed to let go of control and allow the children to be independent, which was hard for them. It is important that the research on the topic is continually renewed since parents need much support in this difficult situation. The nurse should have a good understanding of the experiences that may affect parents in order to be able to develop capacity to support the parents in their daily diabetes care.