Abstract
Background:
Thyroglobulin, produced exclusively by the thyroid gland, has been proposed to be a more sensitive biomarker of iodine status than thyrotropin or the thyroid hormones triiodothyronine and thyroxine. However, evidence on the usefulness of thyroglobulin (Tg) to assess iodine status has not been extensively reviewed, particularly in pregnant women and adults.
Summary:
An electronic literature search was conducted using the Cochrane CENTRAL, Web of Science, PubMed, and Medline to locate relevant studies on Tg as a biomarker of iodine status. Since urinary iodine concentration (UIC) is the recommended method to assess iodine status in populations, only studies that clearly reported both Tg and UIC were included. For the purpose of this review, a median Tg <13 μg/L and a median UIC ≥100 μg/L (UIC ≥150 μg/L for pregnant women) were used to indicate adequate iodine status. We excluded studies conducted in subjects with either known thyroid disease or those with thyroglobulin antibodies. The search strategy and selection criteria yielded 34 articles of which nine were intervention studies. The majority of studies (six of eight) reported that iodine-deficient pregnant women had a median Tg ≥13 μg/L. However, large observational studies of pregnant women, including women with adequate and inadequate iodine status, as well as well-designed intervention trials that include both Tg and UIC, are needed. In adults, the results were equivocal because iodine-deficient adults were reported to have median Tg values of either <13 or ≥13 μg/L. Only studies in school-aged children showed that iodine-sufficient children typically had a median Tg <13 μg/L. Some of the inconsistent results may be partially explained by the use of different methodological assays and failure to assess assay accuracy using a certified reference material.
Conclusions:
These data suggest that Tg does hold promise as a biomarker of iodine deficiency. However, it is associated with limitations. A median Tg cutoff of 13 μg/L warrants further investigation, particularly in adults or pregnant women, as there is a lack of both observational and intervention studies in these groups.
Introduction
I
Thyroglobulin (Tg) plays an important role in the synthesis of thyroid hormones T3 and T4 (9). It is a glycoprotein comprising two 330 kDa protein chains synthesized in the thyrocyte (10). After synthesis, Tg is transported and stored in the follicular colloid of the thyrocyte (11). In the follicular lumen, the tyrosine residues of Tg undergo iodination to produce mono- (MIT) and di-iodotyrosines (DIT) catalyzed by thyroid peroxidase (12) and hydrogen peroxide (13). Subsequent coupling of these iodotyrosines produces T3 and T4 (14,15). Tg is pinocytosed into the thyroid cell (16) and undergoes proteolysis by lysosomes to release T3 and T4 (17), which are then secreted into the bloodstream (18).
When iodine intake is insufficient, low circulating levels of T4 stimulate the release of thyrotropin-releasing hormone from the pituitary gland, which subsequently increases the production of TSH. In addition to increasing the synthesis and proteolysis of Tg, TSH also stimulates the growth and division of the follicular cells, which causes the thyroid gland to enlarge (i.e., goiter) (19). In iodine deficiency, an increased amount of Tg is released into the blood (20), which is positively correlated with thyroid volume (21). For example, healthy adults have a mean Tg concentration ranging from 5 to 14 μg/L (22 –27). In contrast, adults with endemic goiter have a mean Tg ranging from 94 to 208 μg/L (28 –30). Recently, experts attending a National Institutes of Health workshop (31) recommended that Tg be used in the evaluation of iodine status.
The most common use of Tg is to monitor the treatment of patients with differentiated thyroid cancer (DTC) (32). Several review articles have focused on Tg monitoring in patients with DTC (11,33,34) or the performance of different assays used for monitoring DTC (35,36). The evidence on the usefulness of Tg in patients with DTC is well established. However, data on the effectiveness of Tg to assess iodine status in healthy populations is scarce. This review will report on: first, the analytical issues of Tg methods; second, observational studies measuring Tg to assess iodine status in healthy populations of pregnant women, newborns, children, and adults; and third, intervention studies investigating the effect of iodine supplementation on Tg in populations of pregnant women, newborns, children, and adults. This information will be used to determine if Tg can be used as a biomarker to assess iodine status.
Search Strategy
We conducted an electronic literature search using the Cochrane CENTRAL, Web of Science, PubMed, and Medline (OvidSP) to locate relevant studies published in English between January 1960 and October 2013 using Tg as a biomarker of iodine status. We used the following combined keywords: serum thyroglobulin, thyroglobulin, blood, children, infants, adults, pregnant women, pregnancy, maternal iodine status, iodine status, iodine deficiency, iodine insufficiency, iodine sufficiency, and iodine supplementation. We also located additional studies from references in the retrieved articles. Since UIC is the recommended biomarker of iodine status in populations (5), only studies that clearly report both Tg and UIC were included. We excluded studies conducted in subjects with either known thyroid disease or those with thyroglobulin antibodies (TgAb) because such subjects can have falsely low or high Tg that are not caused by insufficient iodine intake. The search resulted in 34 articles (i.e., 38 studies) being selected (Table 1). Of these, nine were randomized controlled trials, two were nonrandomized controlled trials, three were cohort observational studies, 23 were cross-sectional studies (10 multicenter), and one was a monitoring report of iodization programs that included a measurement before the introduction of iodized salt to a measurement after the introduction of iodized salt. In order to investigate the consistency of the relationship between iodine status as determined by UIC and Tg more clearly, for those studies that reported this information for more than one group, we considered each of these groups separately (i.e., one study of pregnant women and their newborns (37); one study of pregnant women and adults (38); one study of children and adults (30); three studies of children living in different regions (39) or countries (40,41); and seven studies of adults living in different regions (21,25,42–44) or countries (45,46)).
Included pregnant women and adults.
Included pregnant women and newborns.
Counted as one article.
Included children and adults.
AA, agglutination assay; ECLIA, electrochemiluminescence immunoassay; ELISA, enzyme-linked immunosorbent assay; FIA, fluoroimmunoassay; HA, hemagglutination assay; ICMA, immunochemiluminescence assay; ILMA, immunoluminometric assay; IRMA, immunoradiometric assay; NA, not assessed; NR, not reported; RIA, radioimmunoassay; RIPA, radioimmunoprecipitation assay; Tg, thyroglobulin; TgAb, thyroglobulin antibodies; UIC, urinary iodine concentration.
Discussion
Methods to measure Tg concentration
Tg can be measured using either immunometric assay (IMA) or radioimmunoassay (RIA) (35). Of the 34 articles measuring Tg (Table 1), the predominant Tg assay used was RIA (27%), followed by various IMAs, including immunoluminometric assay (22%), immunochemiluminescence assay (21%), immunoradiometric assay (12%), fluoroimmunoassay (10%), enzyme-linked immunosorbent assay (3%), electrochemiluminescence immunoassay (3%), and not reported (3%). Only one article (21) measured Tg using two different types of assays. A dried blood spot method using fluoroimmunoassay (FIA) (40) has been developed by Zimmermann et al. to assess Tg in children (5). Though Tg obtained from a dried blood spot was well correlated with serum samples (r=0.98, p<0.0001) in healthy children (n=29) (47), this relationship has yet to be validated in populations of adults including pregnant women. Furthermore, the dried blood spot method has not been reproduced in other laboratories.
Studies of Tg using RIA were first published in the 1960s. Several of these early studies (48 –50) reported that Tg was undetectable in some healthy participants. For example, a small study conducted by Hjort et al. (48) used a RIA with a limit of detection (LoD) of 50 μg/L and found that Tg was undetected in all 12 healthy subjects, indicating that these subjects would likely have had Tg concentrations ≤50 μg/L. In contrast, Torrigiani et al. (49) detected Tg in 60–70% of healthy subjects (n=111) when they used a RIA with a LoD of 10 μg/L; van Harle et al. (50) detected Tg in 74% of healthy subjects (n=95) using a RIA with a LoD of 1.6 μg/L. Therefore, early RIAs had a relatively poorer functional sensitivity compared with first-generation Tg assays (0.5–1.0 μg/L) developed in the 1980s (51,52) and second-generation Tg assays (≤0.1 μg/L) in use since the early 2000s (53,54); studies using first-generation Tg assays (21,55) did not report undetectable Tg in any healthy subjects.
Tg has been reported to be method dependent (56 –58), and the interassay variation can vary between 43% and 65% in healthy subjects (35,57,59). To overcome interassay variation and allow for comparisons between studies, a certified Tg reference material (i.e., CRM-457) has been produced as a quality-control material for assay standardisation (60). Some but not all types of Tg assays have been standardized against CRM-457 in-house by the manufacturers (61). However, Tg CRM-457 only reduces interassay variation by 14–27% (59). It is suggested that this is because current Tg assays are unable to identify the heterogeneity of Tg epitopes (52,62). Of 34 articles measuring Tg (Table 1), only four (40,41,63,64) used Tg CRM-457 as an external quality control.
Another issue with regard to the measurement of Tg is the presence of TgAb. When a RIA is used, a subject positive for TgAb will most likely have a higher Tg value, while IMA tends to lower Tg in TgAb-positive subjects (33,65). Thus, subjects who have a positive test for TgAb should be excluded from the results if Tg is used as a biomarker of iodine status in a population. In adults, studies (66 –69) have found that 3–13% of adults have TgAb. However, in children, the prevalence of TgAb is lower (70,71), and Zimmermann et al. (47) suggest that screening for TgAb in this age group is not necessary. Twenty-two of 34 studies measured TgAb prior to Tg measurement (Table 1). Of these, the predominant TgAb assay used was a RIA (58%), followed by the radioimmunoprecipitation assay (14%), immunochemiluminescence assay (12%), Tg recovery (5%), hemagglutination assay (5%), agglutination assay (5%), and FIA (2%). Only two articles (21,72) measured TgAb using two different types of TgAb assays.
In a large multicenter study of healthy children aged 5–14 years, Zimmermann et al. reported a reference range for Tg of 4–40 μg/L as determined by FIA (40). This is similar to reference ranges reported for adults of 3–40 μg/L using both RIA and IMA methods (65,68). We did not identify any consistent effects of age or sex on Tg. Only one study (73) reported that Tg decreased with advancing age. In 1994, the WHO/ICCIDD/UNICEF suggested that a median Tg concentration <10 μg/L indicates adequate iodine status in populations of school-age children. However, in 2007, the WHO/ICCIDD/UNICEF, although acknowledging that Tg could be used an indicator of iodine status, did not provide a cutoff for Tg. More recently, Zimmermann et al. (41) conducted a large multicenter study of children (n=2512) from 12 countries with varying iodine status, and suggest that a median Tg concentration <13 μg/L and/or <3% of Tg values >40 μg/L be used as a biomarker of adequate iodine status in children and, with caution, in adults. To date, the cutoff of 13 μg/L and/or <3% of Tg values >40 μg/L has not been examined in younger children or pregnant women. Because no studies have reported the percentage of Tg values >40 μg/L in populations, for the purpose of this review, a median Tg <13 μg/L and a median UIC ≥100 μg/L (UIC ≥150 μg/L for pregnant women) were used to indicate adequate iodine status.
Pregnant women
Eight observational studies measuring Tg in iodine-deficient pregnant women aged between 15 and 46 years were identified (Table 2). Six of eight studies (37,38,74 –77) reported that iodine-deficient pregnant women (either first, second, or third trimester, or at delivery) had a median Tg ≥13 μg/L (range 16–67 μg/L). Two of the eight studies (78,79) assessed Tg concentration in iodine-deficient women throughout their pregnancy (i.e., in each trimester); in one study (79), a median Tg <13 μg/L was observed in all three trimesters, and in one study (78), a Tg ≥13 μg/L was reported in the first and third trimesters, but it was <13 μg/L in the second trimester. Although six of eight studies (37,38,74,75,77,79) collected information on the use of iodine supplements in pregnancy, of these, only one study (37) reported that the Tg concentration of women taking iodine supplements was significantly lower compared with women who did not take supplements (i.e., difference of ∼15 μg/L). We are unaware of any published studies of pregnant women with adequate iodine status that include measures of both UIC and Tg.
Range used unless mean reported.
Only subjects with no known thyroid disease or negative for TgAb were included.
Median used unless mean or geometric mean reported.
UIC reported as μg/g creatinine.
Tg was reported in Costeira et al. (79); UIC and the data on supplement use were reported in Costeira et al. (93). These two studies were counted as one study (79).
Geometric mean.
I, iodine; PP, postpartum.
Three intervention studies investigating the effect of iodine supplementation on Tg in iodine-deficient pregnant women were identified (Table 3). One of the studies (80) assessed Tg concentration in the first trimester before supplementation and then again at two weeks postpartum; one study (81) assessed Tg in the first and third trimesters; and one study (82) assessed Tg in all three trimesters and again 12–24 weeks postpartum. Tg concentrations in women in the first trimester (i.e., at baseline before supplementation) ranged from 13 to 25 μg/L, and postpartum, in women that had received any type of additional iodine (i.e., supplements or iodized salt), Tg ranged from 8 to 18 μg/L. Of the two studies with postpartum data (80,82), only one study (80) reported that women taking iodine supplementation in pregnancy had a postpartum median Tg <13 μg/L. However, the interpretation of these findings is confounded by differences in study designs, including a lack of a placebo group, relatively small sample sizes (n=66–131), varying levels and types of supplemental iodine (iodized salt or supplements containing 150–300 μg iodine per day), duration of follow-up (2–24 weeks postpartum), and use of different Tg assays.
Range used unless mean reported.
Only subjects with no known thyroid disease or negative for thyroglobulin antibody were included.
Actual quantity of iodine from supplement; Liesenkötter et al. (80) and Santiago et al. (82) used iodine supplements in the form of KI.
Median used unless mean reported.
UIC reported as μg/g creatinine.
Estimated value from a figure.
Mean.
Estimated value from a table.
IS, iodized salt; KI, potassium iodide.
In summary, it appears that the majority studies typically report that iodine-deficient pregnant women have a median Tg ≥13 μg/L. Furthermore, iodine supplementation does not consistently decrease Tg below this cutoff either during pregnancy or postpartum, although this may reflect inadequate supplementation, as UIC did not reach recommended cutoffs. More large observational studies of pregnant women, including women with adequate and inadequate iodine status, as well as good intervention trials that include both Tg and UIC, are required before conclusions can be drawn about the usefulness of Tg as a biomarker of iodine status in pregnancy. Another consideration is whether Tg in pregnancy needs to be trimester specific, as is suggested for thyroid hormones such as TSH (83) and T4 (84).
Newborns
Three studies that measured Tg in cord blood from newborns were identified (Tables 4 and 5). Two of the three studies (85,86) were supplementation trials of mothers during pregnancy. The Tg concentration of newborns born to mothers receiving a placebo or who did not take supplements in pregnancy ranged from 62 to 113 μg/L, while in the newborns of mothers who took iodine supplements, Tg ranged from 31–65 μg/L. The usefulness of measuring Tg in newborn cord blood is questionable. A more commonly used and relatively accessible biomarker to assess iodine status in newborns is neonatal TSH collected by heel prick two to three days after birth (5).
Range used unless mean reported.
Only subjects with no known thyroid disease or negative for TgAb.
Median used unless mean or geometric mean reported.
Geometric mean.
Mean.
UIC reported as urinary iodine excretion (μg/day).
UIC reported as μg/g creatinine.
Range used unless mean reported.
Only subjects with no known thyroid disease or negative for TgAb.
Median used unless mean or geometric mean reported.
Mean.
Geometric mean.
UIC reported as urinary iodine excretion (μg/day).
Se, selenium.
Children
Six observational studies measuring Tg in children aged between 5 and 14 years were identified (Table 4). Four studies (30,39,41,73) found that iodine-deficient children had a median Tg ≥13 μg/L (range 13–59 μg/L), while two studies (40,41) reported that iodine-sufficient children also had a median Tg ≥13 μg/L (range 13–19 μg/L). Four of six studies (39 –41,55) reported that children with adequate iodine status had a median Tg <13 μg/L. However, one study (39) reported that iodine-deficient children had a median Tg <13 μg/L. The study by Zimmermann et al. (41) included 2512 children from 12 countries with severe iodine deficiency (i.e., median UIC <20 μg/L), mild iodine deficiency (i.e., median UIC 50–99 μg/L), adequate iodine status (i.e., median UIC 100–299 μg/L), and iodine excess (i.e., median UIC ≥300 μg/L). It showed that median Tg appeared to follow a U-shaped curve with the nadir at an UIC of 100–300 μg/L. When iodine intake is very high, excess iodide transiently inhibits the activity of thyroid peroxidase and proteolysis of Tg, which subsequently reduces the synthesis and secretion of thyroid hormones (i.e., the Wolff–Chaikoff effect) (87). However, when prolonged excess iodine intake occurs, Tg could increase because the thyroid gland fails to escape from the Wolff–Chaikoff effect (88). Nonetheless, close examination of data reported by Zimmermann et al. (41) suggests that the relationship between UIC and Tg is highly variable. It is not known, however, how much of this variability is associated with UIC and/or Tg because a single UIC can be confounded by the hydration status, dietary intake, and diurnal variation (4).
Four intervention studies investigating the effect of iodine supplementation on Tg in iodine-deficient children aged 5–14 years for a duration of 5–12 months were identified (Table 5). Three of the four studies (40,47,64) were more than six months long, and reported that median Tg decreased significantly and fell below 13 μg/L when UIC increased from <100 to ≥100 μg/L. The remaining study (89) included five treatment groups but not a control group, was only five months long, and had fewer children in each group. In the three groups where the children became iodine sufficient, Tg was <13 μg/L in only one group.
The majority of observational and intervention studies in school children appear to support the 13 μg/L cutoff proposed by Zimmermann et al. (41) to assess iodine status in this age group. However, the relationship between UIC and Tg is not always consistent, suggesting that Tg alone should not be used to assess iodine status in this group.
Adults
Twelve observational studies measuring Tg in adults aged between 18 and 97 years were identified (Table 4). Seven studies (21,25,30,38,43,45,46) showed that iodine deficient adults had a median Tg ≥13 μg/L (range 16–69 μg/L), while only one study (45) reported that iodine-sufficient adults had a median Tg ≥13 μg/L. However, 8 of 12 studies (21,25,42 –44,72,90,91) reported that adults who were categorized as iodine deficient had a median Tg <13 μg/L. One of these studies (42) included adults with iodine excess (i.e., median UIC ≥300 μg/L) who, in contrast to the findings of Zimmermann et al. (41) in schoolchildren, had a median Tg <13 μg/L.
Two intervention studies investigating the effect of iodine supplementation on Tg in adults were identified (Table 5). One study included iodine-sufficient middle-aged adults supplemented with additional iodine for 8–12 weeks (92); at baseline, the median Tg was <13 μg/L, which decreased, but not significantly, after supplementation. Another study of older adults (60–80 years) (63) who were moderately iodine deficient reported a median Tg ≥13 μg/L at baseline. Although iodine status improved, the subjects remained mildly iodine deficient, which likely explains that, although Tg concentration significantly decreased after supplementation, it remained ≥13 μg/L.
Based on these observational studies, it is difficult to conclude that the Tg cutoff of 13 μg/L suggested by Zimmermann et al. (41) for children can be used as a biomarker of iodine status in adults. Furthermore, there are no randomized placebo-controlled trials in adults that have shown an improvement in iodine status (indicated by an increase in baseline UIC from <100 to ≥100 μg/L) results in a concomitant fall in Tg concentration from ≥13 to <13 μg/L.
Summary and Conclusions
Tg does hold promise as a biomarker of iodine deficiency. However, it is also associated with limitations. The variety of methods used to analyze Tg makes it difficult to compare studies. It would be helpful if studies that measured Tg standardized their assays with CRM-457. Furthermore, particularly in adult populations, subjects should be screened for TgAb. Despite these problems, the studies included in this review support the use of Tg as a biomarker of iodine status in school children using the <13 μg/L cutoff as suggested by Zimmermann et al. (41). However, it is not possible to draw conclusions regarding the efficacy of Tg in adults because the data are equivocal, while there are no studies of pregnant women with adequate iodine status that also include data on Tg concentration. In particular, few intervention studies have investigated the diagnostic performance of Tg assays and its clinical relevance in assessing iodine status in healthy populations. Well-designed randomized placebo-controlled trials are required to investigate further the effect of iodine supplementation on Tg in mild to moderately iodine-deficient populations, particularly in adults and pregnant women.
Footnotes
Author Disclosure Statement
The authors declare that they have no conflict of interest.