The Foundation for Blood Research administers a statewide, second-trimester prenatal serum screening program for open neural-tube defects and Down's syndrome in Maine.12,13 Aliquots of serum that remain after screening are routinely coded and stored at –20°C. Outcome information is available through a contract with the state's Bureau of Vital Records. The current study cohort was limited to women with viable singleton pregnancies, who were screened between January 1987 and March 1990, and their infants whose birth weight was at least 1500 g. The serum from the mothers was shipped to the New England Newborn Screening Program in Boston, where serum thyrotropin was measured. Samples from 25,216 women were analyzed in five batches over a two-year period.

Selection of Women with Hypothyroidism and Control Subjects

We recruited women with hypothyroidism during pregnancy, as determined by a high serum thyrotropin concentration, without regard to treatment status, and we tested their children between March 1996 and December 1997. We contacted 55 of the 75 pregnant women with serum thyrotropin concentrations at or above the 99.7th percentile of the values for all the pregnant women; 47 (85 percent) agreed to participate. In 2 of the 75 pregnancies, the women were enrolled through a previous pregnancy. Of the 18 women not contacted, 3 had moved to another state, 1 had died, and for 1, the child was a ward of the state. The remaining 13 were not contacted because of limited funds. At the urging of a grant review panel, we recruited 18 more women to represent a range of milder cases, defined by a serum thyrotropin concentration between the 98th and 99.6th percentiles, inclusive, and a serum thyroxine concentration below 7.75 μg per deciliter (99.7 nmol per liter). To identify this second subgroup, we measured serum thyroxine concentrations in 247 pregnant women with serum thyrotropin concentrations between the 98th and 99.7th percentiles. Fifteen of the 18 women identified (83 percent) agreed to participate. After recruitment, we measured thyroxine, free thyroxine, and antithyroid peroxidase antibodies in the original serum samples from all women in the study.

For each woman with hypothyroidism, we identified potential control subjects who had serum thyrotropin concentrations below the 98th percentile and who were matched according to the following criteria: mother's age at delivery (within one year), number of years of education of the mother (within one year), gestational age at the time of sampling (same completed week), duration of storage of serum sample (within one month), and sex of the child. From this group, two women were randomly selected and recruited for each woman with hypothyroidism. Additional matched control women were available from the same list, if one initially declined participation.

The protocols for the additional assays and the follow-up study were approved by the institutional review board at the Foundation for Blood Research. Enrollment began with a telephone call to the woman and a letter describing the study. Then an appointment was arranged, at which informed consent was requested and, if consent was granted, testing was performed on the child. The neuropsychological test results were provided to the family within one month after the child's testing was completed.

At the end of the study, we contacted the women with hypothyroidism and the matched control women again to determine whether hypothyroidism had been clinically diagnosed since the pregnancy in those who had not received a diagnosis of hypothyroidism at the time of pregnancy. The contact was initially by a letter, which also included information about the thyrotropin concentrations in the stored serum samples. The letter was followed by a telephone call, during which a questionnaire was administered and a blood test for measuring thyrotropin was offered. For those who agreed to be tested, blood spots were collected on filter paper by a finger prick.

Study Procedures

We collected standardized information about socioeconomic status and medical history from all women enrolled in the study, using the Four Factor Index of Social Status (the Hollingshead score).14 Each woman and her husband or partner were assigned an education code ranging from 1 (corresponding to less than seven years of schooling) to 7 (corresponding to graduate or professional training) and an occupation code ranging from 1 (e.g., “farm worker”) to 9 (e.g., “higher executive”). Each couple's individual education scores were multiplied by 3, the occupation scores were multiplied by 5, and the two values were then added together. The final score was the average of the scores of the woman and her partner. When one partner was not employed, the final score was taken to be the score of the employed partner. The woman was also asked whether her child had repeated a grade and about her child's school performance, including whether the child had had learning problems or other difficulties in school.

Neuropsychological testing of the women's children included assessment of intelligence, attention, language, reading ability, school performance, and visual–motor performance. One of two certified psychologists performed the testing, and the project's consulting psychologist supervised the testing and rescored the tests. The staff involved in the neuropsychological testing did not know whether the children's mothers were women with hypothyroidism or control women. Intelligence was measured with use of the Wechsler Intelligence Scale for Children, third edition,15 the most widely used intelligence test. This test provides a full-scale IQ score and subscale scores (range, 40 to 160) for verbal skills, performance, and freedom from distractibility. To measure hearing deficits in the children, the staff administered subtests on word discrimination and word articulation from the Test of Language Development, second edition16 (range of scores, 1 to 20). We used the norms for children 8 years 11 months of age, because the version for older children did not have scales for word discrimination or articulation. The Peabody Individual Achievement Test, revised (PIAT-R),17 was used to measure reading recognition and reading comprehension (range of scores, 55 to 145).

The staff administered the Conners' Continuous Performance Test to measure sustained vigilance and attention,18 using a computer program that employs a go–no go paradigm (range of overall index score, 1 to 30). The Developmental Test of Visual-Motor Integration19 was administered to provide a standard measure of visual perception and fine motor skills (range, 55 to 145). The grooved pegboard test20 was administered to assess visual–motor coordination and dexterity by measuring the time required to insert pegs with both the preferred and nonpreferred hand (for this test, it is recommended that normative data be derived from control children in the study).

Assay Methods

We measured serum thyrotropin using a coated-tube radioimmunoassay (Diagnostic Products, Los Angeles). Thyrotropin was measured on dried blood spots with a time-resolved immunofluorometric assay (Wallac Oy, Turku, Finland). Serum thyroxine was measured with a solid-phase radioimmunoassay21 or a time-resolved immunofluorometric assay (Wallac Oy); serum free thyroxine was measured with a time-resolved immunofluorometric assay. We measured serum antithyroid peroxidase antibodies using the Kalibre enzyme-linked immunosorbent assay (Kronus, San Clemente, Calif.) (normal concentration, ≤2 U per milliliter).

Statistical Analysis

The serum thyrotropin, thyroxine, and free thyroxine concentrations were logarithmically transformed before analysis. We used geometric means and logarithmic standard deviations to summarize the results (after censoring seven measurements that were more than 3 SD above or below the group mean). We compared categorical variables using an exact test of significance or odds ratios, and we compared continuous variables using the Student's t-test. When necessary, the t-test was modified to allow for unequal variances. The primary analysis was of all 62 women with hypothyroidism and all 124 control women; we preserved matching by comparing the result from the child of a woman with hypothyroidism with the average result from the two matched control children. No adjustment was made for multiple comparisons. All statistical tests were two-sided.