The present study shows that GAD65A and IA-2A positivity at birth were associated with a 7.5-fold increased risk for developing type 1 diabetes in this cohort of Danish children. Every tenfold increase in GAD65A and IA-2A levels at birth increased the risk for development of type 1 diabetes by a factor 2.55. Thus, both presence and titers of GAD65A and IA-2A had a strong risk-increasing effect. Maternal type 1 diabetes was a confounder, but it was only responsible for approximately one-fifth of the risk increase associated with GAD65A and IA-2A presence. That the presence of islet autoantibodies at the time of birth may be predictive for later type 1 diabetes development was suggested by Lindberg et al
, who studied cord blood samples from 85 children who later developed type 1 diabetes and 321 control subjects. Our approach to test the validity of the previous observation (4)
was to utilize the unique repository of DBS samples collected for the Danish neonatal screening program since 1981 (17
). This repository, unique in that the DBSs have been stored at −25
°C, made it possible to study samples from 2023 type 1 diabetes cases and 4046 matched controls. The present study is to our knowledge the first to show in a large, validated, and population-based case–control investigation that islet autoantibodies at birth were associated with increased type 1 diabetes risk later in life.
Do the islet autoantibodies identified in the 5-day-old newborns have maternal origin or are they produced by the newborn themselves? It is well known that during second trimester of pregnancy, maternal immunoglobulin is actively transported across the placenta to the fetus (24)
. Antibody transport accelerates during the last trimester until birth, and IgG concentrations in the fetus exceed those of the mother (25)
. Neither maternal IgA nor IgM are transferred to the fetus. In the present study, detected GAD65A and IA-2A were mostly of the IgG subclass as Sepharose Protein A was used to separate free from antibody-bound labeled autoantigen. From vaccination studies (26)
, it is known that the ability of newborns to generate IgG responses increases over the first year, but adult responses to protein antigens are not achieved until 12–24 months of age. We assume that the GAD65A and IA-2A we measured were transferred from the mother to the child. It is therefore possible that subclinical islet autoimmunity in the pregnant mother may explain the increased risk for type 1 diabetes in her offspring.
Islet autoantibodies are strongly associated with HLA-conferred diabetes susceptibility both in families and in the general population (27)
. We asked whether autoantibody positivity is a surrogate marker of HLA-defined diabetes predisposition in the current study population. It is likely that the autoantibody-positive non-diabetic mothers carry HLA risk genotypes for type 1 diabetes at a higher frequency than the autoantibody-negative mothers. Accordingly, the autoantibody-positive mothers may also transfer such genotypes to their child resulting in an increased diabetes risk in the offspring. The type 1 diabetes risk ascribed to the presence of GAD65A and IA-2A was reduced but still remaining significant when HLA-DQB1
genotypes were included into the model. It cannot be excluded that other genetic factors, with lower risk, may contribute to the risk of type 1 diabetes in the offspring to islet autoantibody-positive mothers. We conclude that the presence of islet autoantibodies at the time of birth predicts type 1 diabetes risk regardless of HLA risk genotypes and maternal diabetes.
In the present study, only one-fifth of the increase in disease risk related to the presence of GAD65A and IA-2A was actually related to maternal diabetes. However, GAD65A- and IA-2A-positive cases without diabetic mothers may be the most intriguing observation. Follow-up studies of these mothers will be of interest as gestational islet autoantibodies may increase the risk for post partum
type 1 diabetes (28
). In these mothers, in utero
or other stressors (14)
could either have had direct impact on β cells or may have caused β-cell damage by initiating a strong proinflammatory immune response. Furthermore, about 50% of those children whose mothers have type 1 diabetes were still GAD65A and IA-2A negative at the time of birth. Mothers diagnosed with type 1 diabetes have variable duration of their diabetes, and may become negative for autoantibodies prior to or during their pregnancy (31)
A possible weakness to our study is that we may in particular detect cases that had an early age at diagnosis. However, the GAD65A- and IA-2A-related effect on type 1 diabetes risk was found to be similar in children below and above 10 years of age at type 1 diabetes onset. Proportions of GAD65A- and IA-2A-positive cases with young diagnosis out of all GAD65A- and IA-2A-positive cases, and of young GAD65A- and IA-2A-positive cases out of all cases, were similar. In the DIPP study (27)
, the majority of children who developed first multiple islet autoantibodies and then type 1 diabetes had an appearance of one or more islet autoantibody below the age of 5 years. In the DAISY study (11)
, no association could be shown between the presence of islet autoantibodies at birth and type 1 diabetes risk. In the BABY DIAB study (10)
, in contrast, the presence of maternal islet autoantibodies was suggested as protective against type 1 diabetes before the age of 8 years. Insulin autoantibodies (IAA) persisting after the age of 8 months of age (12)
, on the other hand, were associated with an increased type 1 diabetes risk. Owing to limited DBS material, we were also not able to analyze IAA in the present study. Much larger cohort studies will be needed to further analyze the risk of islet autoantibodies at the time when DBSs are obtained for perinatal screening of monogenic diseases. In such studies, it will also be important to obtain data on maternal islet autoantibodies as well as HLA
alleles and non-HLA genetic risk factors (2
). The individually matched case–control design implies that control subjects were selected as not having diabetes at the date of diagnosis of the corresponding case. None of the control subjects developed type 1 diabetes in the follow-up period; nevertheless, we cannot exclude some subjects of the control group, which may develop type 1 diabetes later in life. However, this is unlikely to affect the overall conclusion of the study.
In the present case cohort, specificity was very high, i.e. seven times as many future type 1 diabetes patients were GAD65A and/or IA-2A positive at birth compared to the control group. Because the number of cases and controls in our study population does not correspond to the real distribution in a background population (i.e. the ratio of cases to controls in the study is much higher), we are not able to draw conclusions for how specific and sensitive the presence of GAD65A and IA-2A at birth were for prediction of type 1 diabetes in the general population.
In conclusion, the present study represents a large population-based case–control study demonstrating that GAD65A and IA-2A positivity at birth suggest an increased risk of developing type 1 diabetes at any age before 23 years.