The genotype frequencies were almost identical in all three Finnish populations of different ages, with no gender differences, and did not differ from corresponding figures for the Finnish blood donors. However, the PRNP codon 129 genotype distribution in Finland differed significantly from that of the British and the Irish blood donors and the previously published blood donor data on other Western Europeans and Americans.

SUMMARY Rotavirus, the most common cause of childhood gastroenteritis, has been implicated as one of the viral triggers of diabetes‐associated autoimmunity. To study the possible association between rotavirus infections and the development of diabetes‐associated autoantibodies, we measured the prevalence of rotavirus antibodies in serum samples collected at 3–6‐month intervals up to the age of 2 years from 177 children selected from consecutive newborns because they carried HLA‐DQB1 alleles associated with increased risk for type 1 diabetes. Twenty‐nine of the children developed at least two of four diabetes‐associated autoantibodies (ICA, IAA, GADA or IA‐2A) during the first 2 years of life (the cases), whereas 148 children remained autoantibody‐negative matched with the cases for date of birth, gender, living region and HLA‐DQB1 alleles. The temporal association between the development of the first‐appearing diabetes‐associated autoantibody and rotavirus infections was studied by analysing whether the cases had a diagnostic increase in rotavirus antibody titre more often during the 6‐month period that preceded seroconversion to autoantibody positivity than the controls. By the age of 12 months one of the 13 case children (7%), who had a serum sample drawn at that age and who had developed at least one type of diabetes‐associated autoantibodies, had experienced a rotavirus infection, while 12 of the 61 (20%) autoantibody‐negative control children had had a rotavirus infection. By 18 months, four of the 22 autoantibody‐positive cases (18%) and 18 of the 89 controls (20%) had rotavirus antibodies, and by the age of 24 months the respective numbers were five of the 27 cases (19%) and 32 of the 113 (28%) controls. A rotavirus infection occurred during the 6 months preceding the sample which was positive for an autoantibody in four of the 25 periods (16%) for which both necessary samples were available, while the controls had a rotavirus infection during 55 of the 370‐such periods (15%). Accordingly, our data suggest that rotavirus infections are unlikely triggers of beta‐cell autoimmunity in young children with genetic susceptibility to type 1 diabetes.

Timing of onset of autoimmunity is a prerequisite for unmasking triggers and pathogenesis of type 1 diabetes. We followed 4,590 consecutive newborns with 8 or 3% HLA-DQB1 conferred risk for type 1 diabetes at 3-, 6-, or 12-month intervals up to 5.5 years of age. Islet cell autoantibodies (ICAs) and, in the 137 children with ICAs, insulin autoantibodies (IAAs), GAD65 autoantibodies (GADAs), and IA-2 protein autoantibodies (IA2As) were measured. Children with high genetic risk developed ICAs more often than those with moderate risk (log-rank P ‫؍‬ 0.0015); 85 and 91% remained ICA negative by 5 years of age, respectively. The time of appearance of biochemical autoantibodies was then compared with the appearance of ICAs. IAAs and GADAs emerged usually before ICAs (means ؊1.8 and ؊1.5 months, respectively) and IA-2As after ICAs (mean 2.0 months). Ninety-five percent of all IAAs, GADAs, and IA-2As seroconversions occurred in a cluster (؊12 to 8 months) around the ICA seroconversion. We conclude that diabetes-associated autoantibodies emerged in children with predisposing HLA-DQB1 alleles after 3 months of age at a constant tempo, determined by the genetic risk level, usually in the order of IAA, GADA, ICA, and IA-2A. Seroconversion to multiple autoantibody positivity usually occurred tightly clustered in time. Diabetes 51:646 -651, 2002

Aims/hypothesis. A reduced first-phase insulin response to intravenous glucose is perceived as a sign of far-advanced deterioration of beta-cell function during the development of Type I (insulin-dependent) diabetes mellitus, but data on insulin responses at the onset of diabetes-related autoimmunity are lacking. We studied the first-phase insulin responses of small children soon after observed seroconversion to autoantibody positivity. Methods. In the Type I Diabetes Prediction and Prevention Study newborn infants are screened for HLA-DQB1-associated genetic risk for Type I diabetes and those with increased risk are followed-up for the emergence of islet-cell antibodies. If antibodies are detected, autoantibodies to three other antigens (insulin, GAD65 and IA-2) are also measured. To measure first-phase insulin responses, intravenous glucose tolerance tests were carried out in 52 (1 to 5-year-old) children who had recently seroconverted to islet-cell antibody positivity.Results. The first-phase insulin response was subnormal (<38 mU/l, the 5 th percentile of insulin responses of 20 islet-cell antibody negative healthy children at this age) in 22 of the 52 children (42%). Stepwise multiregression analysis showed that islet-cell antibody greater than 20 JDFU (p=0.0005), insulin autoantibodies (p=0.0009) and an increasing number of positive autoantibodies (p=0.0011) were independent predictors of low first-phase insulin response. Conclusion/interpretation. A decreased first-phase insulin response could be an early phenomenon in the course of prediabetes in young children, implying a rapid autoimmune destruction or loss of function of beta cells as well as possible metabolic compensation mechanisms, since 11 out of the 22 high risk children remain nondiabetic for a considerable period of time despite low insulin responses. [Diabetologia (2002[Diabetologia ( ) 45:1639[Diabetologia ( -1648 Keywords Autoantibodies, HLA risk markers, insulin secretion, prediabetic state, Type I diabetes. Corresponding author: Dr. P. Keskinen, The JDRF Center for Prevention of Type I Diabetes in Finland. E-mail: paivi.keskinen@uta.fi Abbreviations: DIPP, Type I Diabetes Prediction and Prevention project; FPIR, first-phase insulin response; GAD65, 65 kilodalton isoform of glutamic acid decarboxylase; GADA, antibodies to GAD65; IAA, insulin autoantibodies; IA-2A, antibodies to the protein tyrosine phosphatase related IA-2

No abstract

The role of Fas ligand-induced lymphocyte apoptosis in the development of insulitis was studied in nonobese diabetic (NOD) mice. Fas ligand with a molecular weight of 45 kD appeared in the pancreas of NOD mice during the onset of insulitis as demonstrated by western blot analysis. In situ DNA end-labeling (ISEL) of the pancreases of NOD mice demonstrated positive cells in the islets of Langerhans, with an agedependent increase in the density and frequency of animals with islets containing ISEL-positive cells. In the pancreatic islets of BALB/c mice, no ISEL-positive cells were observed in any of the age groups studied. In ultrastructural analysis degenerating cells with condensed or fragmented nuclei and plasma membranes detached from neighboring cells were observed both in and around the islets. In some cases, these cells were being phagocytosed by the neighboring islet cells. Degenerating cells with characteristics of lymphocytes were seen in contact with healthy lymphocytes around the islets. Neutralizing Fas ligand antibodies affected 3H-thymidine incorporation of CD3+CD28+ pancreatic lymphocytes from 12-to 30-week-old NOD mice in one of three cultures. There was no difference in the effect of neutralizing Fas ligand antibodies between pancreatic and blood lymphocytes. The present results on an increase in density of apoptotic cells in pancreatic islets of NOD mice simultaneously with the onset of insulitis and appearance of Fas ligand suggest that the pancreas infiltrating lymphocytes may be destroyed by Fas ligand-induced apoptosis. Key Words: CD95 ligand-Fas ligand-ApoptosisAutoimmunity.Lymphocytes infiltrate the pancreas of nonobese diabetic (NOD) mice clearly earlier than the manifestations of diabetes occur (1). Thus, possibilities to suppress, delay, or prevent the immunological damage to the pancreatic islets exist. However, not much is known about the factors regulating lymphocytes locally in the pancreas.Recent evidence suggests that a novel member of the tumor necrosis factor polypeptide superfamily, Fas ligand, can suppress autoimmune processes in several organs, including the eye (2) and the testis (3). Fas ligand induces apoptosis in activated T cells, which express the Fas receptor (4). Data on the role of Fas ligand in the pancreatic immune regulation are limited.Fas ligand (CD95L) is a type I1 transmembrane protein with a molecular ratio of 36-43 kD (5). It has a soluble form 26 kD in size (6). CD95L belongs to the

SUMMARYEnterovirus infections are a potential environmental trigger of the autoimmune process leading to clinical type 1 diabetes. It has been suggested that the risk of virus-induced beta-cell damage might be connected with a defect in humoral immune responsiveness to enteroviruses. In the present study we assessed whether such a defect in IgG responsiveness to coxsackievirus B4 antigen existed in young children who developed diabetes-associated autoantibodies during prospective observation from birth until the age of 18 months. IgG levels and maturation of antibody avidity were analysed in 21 children with autoantibodies and 41 control children who had experienced an equal number of enterovirus infections and were additionally matched for age, sex and HLA-DQB1 risk alleles for type 1 diabetes but had not produced diabetes-associated autoantibodies. IgG levels to coxsackievirus B4 were high in cord serum reflecting the presence of maternal antibodies. Mean IgG levels gradually decreased but began to increase after the age of 6 months, showing no significant difference between autoantibody positive and control children. The avidity of antibodies was strong in cord serum and decreased gradually during the first year of life when maternal antibodies disappeared. The avidity indices, which varied considerably from child to child, did not differ between the autoantibody-positive and -negative subjects. In conclusion, our data suggest that children affected by a beta-cell damaging autoimmune process show normal responses to coxsackievirus B4 antigens.

Aims/hypothesis. We evaluated the role of enterovirus infections in the pathogenesis of Type I (insulindependent) diabetes mellitus by monitoring enterovirus antibody levels in prediabetic children who turned positive for diabetes-associated autoantibodies in a prospective birth cohort study. Methods. Serial serum samples taken during prospective observation starting at birth were analysed for IgG and IgA class antibodies against enterovirus antigens including purified coxsackievirus B4, echovirus 11, poliovirus 1 and a synthetic enterovirus peptide antigen using enzyme immunoassay. Maternal samples taken at the end of the third month of pregnancy were also studied. Analyses were done from 21 childen who developed autoantibodies and from 104 autoantibody-negative control children who were matched for the time of birth, gender and HLA susceptibility alleles. For comparison, adenovirus antibodies were also analysed from all samples collected. Results. IgG class enterovirus antibody levels were high in maternal samples and in cord blood in both case and control children. After birth the IgG levels decreased reaching a nadir at the age of 6 months. No IgA class antibodies were detected at birth but started to emerge postnatally. Antibody levels did not differ between the autoantibody positive and the control children during the first 6 months of life. From 6 months to 24 months of age, the autoantibody positive children had higher IgG and IgA levels against coxsackievirus B4, echovirus 11 and the synthetic enterovirus peptide antigens than control children but poliovirus 1 and adenovirus antibodies were closely similar in the two groups. The difference between children with autoantibodies and control children was predominantly seen among boys and among those with the HLA-DQB1*0302/x genotype. Conclusions/interpretation. Our data show that children who seroconverted for diabetes-associated autoantibodies develop stronger humoral immune responses to coxsackievirus B4, echovirus 11 and a synthetic enterovirus peptide antigen than children who remained negative for autoantibodies. Poliovirus antibodies induced by uniform vaccinations did not differ between the prediabetic and control children suggesting that the regulation of antibody responses to enteroviruses is not disturbed. Accordingly, the results imply a stronger enterovirus exposure in prediabetic children supporting the role of enteroviruses in the pathogenesis of Type I diabetes. [Diabetologia (2001) 44: 818±823]