Study Design

TEDDY is a prospective cohort study involving six clinical research centers — three in the United States (Colorado, Georgia, and Washington) and three in Europe (Finland, Germany, and Sweden). The primary objective of TEDDY is to identify genetic, gestational, and environmental risk factors for islet autoantibodies, type 1 diabetes, or both in children at increased risk for type 1 diabetes on the basis of their two HLA haplotypes (HLA genotype).3 Because the major HLA genotypes that confer a risk of type 1 diabetes also confer a risk of celiac disease, we explored the genetic and environmental contributions to the development of celiac disease autoimmunity and celiac disease in this cohort.

In TEDDY, all newborns underwent HLA genotyping, and those who were found to carry high-risk genotypes for type 1 diabetes were enrolled before 4.5 months of age, with plans for follow-up until the age of 15 years. Serum samples were obtained from all children every 3 months until the age of 48 months and every 6 months thereafter. The HLA genotypes of interest2 are shown in Table S1 in the Supplementary Appendix, available with the full text of this article at NEJM.org. The presence of the DR4–DQ8/DR8–DQ4 genotype allowed the assessment of the effect of one copy of DR4–DQ8 in the absence of DR3–DQ2.

Study Participants

From September 2004 through February 2010, we screened 424,788 newborns. Among these infants, 21,589 had one of nine HLA genotypes of interest to TEDDY investigators. Of the infants carrying a targeted HLA genotype, 8677 were enrolled for prospective study; of these infants, 6403 carried one of the four HLA genotypes reported in this study and were tested for serum tissue transglutaminase (tTG) antibodies, a marker for celiac disease. The characteristics of the eligible children who were enrolled and those who were not enrolled in the TEDDY cohort have been described previously.4 (Data regarding screening and eligibility according to clinical center are provided in Table S2 in the Supplementary Appendix.)

Testing for celiac disease autoimmunity started at the 24-month visit and continued yearly. If a child tested positive at 24 months, earlier blood samples that had been collected from birth onward were also analyzed to determine the age at which tTG antibodies first became detectable. The persistence of tTG antibodies was confirmed by testing of the next available sample from each study participant identified as having a positive test at any time point.

Measurement of tTG Antibodies

We used radioligand binding assays to measure tTG antibody levels in two laboratories.5,6 All serum samples in the United States were screened for tTG IgA antibodies at the Barbara Davis Center for Childhood Diabetes at the University of Colorado in Denver (normal index, <0.05 units).

In European centers, all serum samples were tested at the University of Bristol in the United Kingdom with the use of an assay that detected both IgA and IgG antibodies against tTG (normal index, <1.3 units). Results from these two laboratories had previously shown high levels of sensitivity and specificity for celiac disease and concordance in an international tTG autoantibody workshop that included direct comparison with commercial enzyme-linked immunosorbent assay kits for the detection of IgA antibodies against tTG.7 To harmonize the protocol, and on the basis of quality-control data,8 we sent all samples with tTG antibody levels above 0.01 as assessed in the Denver laboratory for quantification of tTG antibodies in the Bristol laboratory, the reference laboratory for the study. Results were expressed in arbitrary units derived from a standard curve consisting of dilutions of serum taken from a patient with celiac disease. Samples were considered to be positive for tTG antibodies if the value was 1.3 or more units. The interassay coefficient of variation was 22% at both 6 units and 20 units.

Celiac Disease Autoimmunity and Celiac Disease

Celiac disease autoimmunity was defined as the presence of tTG antibodies on two consecutive tests at least 3 months apart, as measured by the Bristol laboratory. Children meeting this primary outcome in the TEDDY cohort were referred to a gastroenterologist at the clinical discretion of their usual physician. The decision about whether to perform a biopsy and, if so, when, was outside the purview of the study protocol.

Celiac disease, the secondary outcome of the study, was defined as an intestinal biopsy sample showing a Marsh score of 2 or higher (on a scale of 0 to 3, with 0 indicating normal intestinal mucosa and 3 indicating atrophied villi and elongated crypts).9 In addition, children who did not undergo biopsy but who had a mean tTG antibody level of 100 units or higher on two consecutive tests at least 3 months apart were also considered to have celiac disease for the purposes of the study. This threshold was selected on the basis of an internal review of all children who underwent biopsy, in order to achieve a disease specificity of 95% or higher, so that only children who did not undergo biopsy but who were very likely to have celiac disease were included.

Statistical Analysis

We analyzed two outcomes: the age at which celiac disease autoimmunity developed and the age at which celiac disease developed. The primary outcome was defined as the first positive result on testing for tTG antibodies, and the right-censored time (i.e., censoring when the event had not yet occurred at the time of measurement) was the age at which the last blood sample was collected for testing of tTG antibodies. The secondary outcome was defined as a positive result on biopsy or the first high-level result on tTG antibody testing (for participants who did not undergo biopsy), and the right-censored time was the age of the child at the last clinic visit at which celiac disease had not been diagnosed. A Cox proportional-hazards model was used for the analysis, with adjustment for country, sex, and presence or absence of a family history of celiac disease (i.e., in a first-degree relative). We used Fisher's exact test to compare proportions and the log-rank test to compare Kaplan–Meier estimates. A P value of less than 0.05 was considered to indicate statistical significance, without adjustment for multiple testing. All analyses were performed with the use of SAS software, version 9.2 (SAS Institute).