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CASE REPORT
Potent thyrotrophin receptor-blocking antibodies: a cause of
transient congenital hypothyroidism and delayed thyroid
development
C Evans
1
, N J Jordan
1,2
, G Owens
3
, D Bradley
1
, M Ludgate
2
and R John
1
1
Department of Medical Biochemistry, University Hospital of Wales, Heath Park, Cardiff CF14 4XN, UK,
2
Department of Medicine,
University of Wales College of Medicine, Cardiff, UK and
3
Department of Paediatrics, Wrexham Maelor Hospital, Wrexham, UK
(Correspondence should be addressed to C Evans; Email: carol.evans@cardiffandvale.wales.nhs.uk)
C Evans and N J Jordan contributed equally to this work
Abstract
Objective: We describe an infant with surprisingly severe neonatal hypothyroidism due to transplacen-
tal passage of thyrotrophin receptor (TSH-R)-blocking antibodies (TBAb).
Design and methods: TBAb were detected using a cell line which stably expresses the human TSH-R and
a cAMP-responsive luciferase reporter by their ability to inhibit TSH-stimulated luciferase expression.
Potent TBAb were detected in maternal serum and initially in the infant’s serum but, in the latter,
TBAb decreased over time to within the reference range by 3– 4 months of age, illustrating the tran-
sient nature of this condition.
Results: The thyroid function of this child did not return to normal on withdrawal of thyroxine
therapy at 16 months of age when he developed transient compensated hypothyroidism.
Conclusions: We propose that the presence of potent TBAb in utero and in the first weeks of life may
have implications for the development of a normally sized thyroid gland. We have demonstrated the
presence of TBAb in the mother’s milk and, as far as we are aware, this is the first such report. How-
ever, the TBAb in the milk probably did not contribute significantly to hypothyroidism in the child,
given the reducing antibody titre in his circulation.
European Journal of Endocrinology 150 265–268
Introduction
Autoantibodies to the thyrotrophin receptor (TSH-R)
that block (TBAb) the effects of thyrotrophin (TSH)
(1, 2) have been described in the serum of some
patients with atrophic thyroiditis and Hashimoto’s thy-
roiditis. TBAb have a pathogenic role in the develop-
ment of hypothyroidism. During pregnancy, passage
of maternal TBAb across the placenta can cause tran-
sient hypothyroidism in the neonate (reviewed in 3).
Transient congenital hypothyroidism (CH) due to
TBAb generally occurs in infants of mothers with a
history of autoimmune hypothyroidism who are
taking thyroxine or have undiagnosed hypothyroidism,
and accounts for 1 –2% of cases of congenital
hypothyroidism (4, 5). TBAb are found in both
maternal and infant serum at birth but gradually
clear from the infant’s circulation after 3 –4 months.
This is usually accompanied by resumption of normal
thyroid function.
Materials and methods
Thyroid function tests
Serum concentrations of TSH, free thyroxine (T4) and
free tri-iodothyronine (T3) were measured on an
ADVIA Centaur automated immunoassay analyser
(Bayer plc, Newbury, Bucks, UK). Thyroid peroxidase
antibodies were measured by competitive immunoassay
on a Roche Diagnostics Elecsys 2010 automated immu-
noassay analyser (Roche Diagnostics GmbH, Mann-
heim, Germany). Thyroglobulin was measured using
the Pasteur thyroglobulin immunoradiometric assay
(Bio-Rad, Marnes La Coquette, France).
TBAb and TSH-R-stimulating antibodies
Serum concentrations of TBAb and TSH-R-stimulating
antibodies (TSAb) were measured using the previously
described lulu*cell line (Chinese hamster ovary cells
stably transfected with recombinant human TSH-R and
European Journal of Endocrinology (2004) 150 265–268 ISSN 0804-4643
q2004 Society of the European Journal of Endocrinology Online version via http://www.eje.org
a cAMP-dependent luciferase reporter) (6). Lulu*were
seeded at approximately 2 £10
4
cells per well in 96-
well plates in Ham’s F12 containing 10% charcoal
stripped calf serum. Thirty-six hours later cells were
switched to assay buffer (see below). To measure TBAb,
serum (10%) was incubated with lulu*in the presence
of bovine TSH (1mU/ml) in Ham’s F12 without 10%
fetal calf serum (FCS). To measure TSAb, serum (10%)
was incubated with lulu*in Hanks balanced salt solution
without sodium chloride (0.185 g/l CaCl
2
, 0.4 g/l KCl,
0.06 g/l KH
2
PO
4
, 0.1 g/l MgCl
2
, 0.1 g/l MgSO
2
,
0.35 g/l NaHCO
3
, 0.48 g/l Na
2
HPO
4
) containing
1.0 g/l D-glucose, 20 mM HEPES, 1.5% bovine serum
albumin, 280 mM sucrose and 5% polyethylene glycol).
All incubations were carried out for 5 h at 37 8Cin5%
CO
2
in air. Where dilutions of sera were assayed, the
total serum concentration was adjusted to a total of
10% by the addition of pooled euthyroid serum.
In each case, cAMP-dependent luciferase production
was determined by measuring light in the presence of
luciferin using a luciferase reporter assay (Promega
UK Ltd, Southampton, UK) on a Perkin Elmer Applied
Biosystems Tropix TR717 microplate luminometer
(Perkin Elmer, Norwalk, CT, USA). Experiments were
performed in duplicate or triplicate and the results
expressed as an average. TBAb activity was expressed
as an inhibition index (InI), calculated as follows: InI ¼
100 £½12lightðpatient sample þ
TSHÞ=lightðeuthyroidpool þTSHÞ:
InI .23 is considered positive (6). TSAb activity was
expressed as a stimulation index (SI): the ratio of light
output from the patient sample to light output from
the euthyroid pool. SI .1.5 is considered positive.
TBAb activity in milk was assessed by incubation of
varying amounts of patient or control unpasteurised
bovine milk with lulu*in the presence of bovine TSH
(1 mU/ml) and Ham’s F12 without 10% FCS. Luciferase
activity was measured as previously described.
All investigations were performed with the informed
consent of the mother.
Case report
A 4.57 kg male infant, the first child of a 31-year-old
mother, was born at 41 weeks of gestation after an
uneventful pregnancy. Neonatal screening revealed a
blood spot TSH of 330 mU/l (normal ,10) and serum
taken at 13 days of age confirmed primary hypothyroid-
ism with a low free T4 (2.7 pmol/l) and a grossly
increased TSH concentration (752 mU/l). He was jaun-
diced (bilirubin 381 mmol/l) and had coarse facial fea-
tures. X-ray of his knee showed neither the distal
femoral nor proximal tibial epiphyses present, consistent
with hypothyroidism in utero. A serum thyroglobulin
concentration of 215 mg/l confirmed the presence of
thyroid tissue. A normally placed, small thyroid gland
was found on ultrasound of the neck: left lobe length
0.7 cm, breadth 0.4 cm, right lobe length 0.7 cm,
breadth 0.6 cm (normative data for term infants (7):
lobe length (mean (S.D.) range) 1.94 (0.24) 0.9 –2.5 cm
and breadth 0.88 (0.16) 0.5 –1.4 cm). No uptake of tech-
netium-99 was observed. Thyroxine therapy was started
on day 13 of life at a dose of 50 mg/day.
The infant’s mother had a 14-year history of
atrophic hypothyroidism for which she was on thyrox-
ine replacement therapy (free T4 24.9 pmol/l, free T3
5.0 pmol/l and TSH 0.5 mU/l at the time of the infant’s
birth). Thyroid peroxidase antibodies were marginally
increased (38.9 kU/l (reference range ,32 kU/l)). She
was clinically well, with no goitre, no evidence of thyr-
oid eye disease or pretibial myxoedema. The maternal
history of atrophic hypothyroidism and the infant’s
negative radioisotope uptake scan raised the possibility
of transient neonatal hypothyroidism due to TBAb and
measurement of TBAb was therefore initiated.
Detection of TBAb in serum
TBAb activity (InI range 70 –80%) was detected in
maternal serum throughout the investigation (Fig. 1).
TBAb (InI 76%) were also detected in the infant’s
serum soon after birth, but decreased to within the
reference range by 4 months of age and were not
found in the infant’s serum thereafter (Fig. 1).
Since serum TBAb and TSAb may coexist (8) multiple
dilutions of maternal serum were assayed for both TBAb
and TSAb. Measurement of TSAb was performed in NaCl-
free buffer since detection of TSAb is improved in the
absence of salt. In contrast, TBAb activity was deter-
mined in culture medium containing physiological
NaCl concentrations in which TSH is more effective at sti-
mulating luciferase expression (6). Potent TBAb activity
was confirmed in the maternal serum (Fig. 2) and was
still detectable at a 100-fold dilution (0.1% maternal
serum in the bioassay equivalent to 15 mg/ml IgG).
TSAb activity was not detected at any dilution of the
maternal serum.
Figure 1 Measurement of TBAb in maternal and infant serum
after birth. TBAb activity was assessed by luminescent bioassay.
Serum (10%) was challenged against 1 mU/ml bovine TSH. InI^
S.E.M. was calculated for duplicate experiments (n¼3); maternal
serum (open bars) and infant serum (hatched bars). Euthyroid
patients (n¼31) had InI ,23% (shown by dotted line).
266 C Evans, N J Jordan and others EUROPEAN JOURNAL OF ENDOCRINOLOGY (2004) 150
www.eje.org
Detection of TBAb in milk
TSAb have been described in the milk of mothers with
Graves’ disease (9). We detected TBAb activity in milk
expressed by the mother (Fig. 3). A dose-dependent
inhibition of TSH stimulation was observed with
increasing amounts of breast milk assayed in contrast
to control milk, in which TSH stimulation was not sig-
nificantly different.
Withdrawal of thyroxine
Thyroxine therapy was required at birth, but the grow-
ing infant did not require an increase in dose to main-
tain TSH within the reference range and subsequent
reductions in the dose of thyroxine were well tolerated.
However, complete withdrawal of thyroxine at 16
months resulted in a transient compensated hypothyr-
oidism (Table 1). At this time, he had a normal
response to a perchlorate discharge test and remained
clinically well. TBAb block the action of TSH and the
uptake of radioactive isotope by the thyroid (references
in 3) and account for lack of technetium-99 uptake in
the earlier scan. By 2 years of age, his thyroid function
had fully recovered and TSH levels returned to within
the reference range.
Discussion
We have described an infant with profound hypothyr-
oidism at birth and in utero due to transplacental pas-
sage of TBAb. Potent TBAb were detected in maternal
serum and in the infant’s serum at birth using a lucifer-
ase-based bioassay. TBAb activity in the infant’s serum
decreased to within the reference range by 3 – 4
months of age, illustrating the transient nature of CH
due to maternal TBAb. The thyroid function of this
child did not return to normal on withdrawal of thyrox-
ine therapy when he developed a transient compensated
hypothyroidism. We speculate that this slow recovery of
normal thyroid function was caused by the impairment
of the normal growth and development of the thyroid
gland by the TBAb present before and after birth.
Most cases of hypothyroidism due to TBAb are transient,
but there has been one previous report of an infant with
permanent thyroid damage due to TBAb (10). Hence it
is important to monitor the response to thyroxine with-
drawal in these infants carefully.
Transplacental passage of TBAb resulted in delayed
development of this infant’s thyroid gland, which was
found to be small by ultrasound scan shortly after
Figure 3 TBAb activity in milk. The graph shows the inhibition of
stimulation by 1 mU/ml bovine TSH, due to varying concentrations
of breast milk (shaded bars) and non-pasteurised bovine milk
(open bars). Mean relative light units (RLU) for a duplicate exper-
iment are shown.
Figure 2 TBAb activity determined by luminescent bioassay for
various dilutions of maternal serum. Serum (made up to a total of
10% with euthyroid serum) was challenged against 1mU/ml
bovine TSH. Inl calculated for a duplicate experiment are shown.
Euthyroid patients had InI ,23% (shown by dotted line).
Table 1 Biochemical investigations in the neonate.
Age of
infant
TSH
(mU/l)
Free T4
(pmol/l)
Free T3
(pmol/l) Comments
13 days 752 2.7 Thyroxine commenced
(50 mg)
26 days 1.1 35.3 9.0
1 month 0.59 9.1 Thyroxine dose reduced
to 40 mg
2 months 0.35 24.8 6.3
3 months 0.25 6.1
4 months 0.2 16.8 6.6
6 months 1.5 17.4 6.1
9 months 1.2 15.5 5.9
11 months 2.0 14.5 5.8
15 months Thyroxine dose reduced
to 20 mg
16 months 3.0 15.9 5.0 Thyroxine stopped
17 months 10.8 13.9 7.0
18 months 8.6 12.2 8.6
20 months 7.3 16.5 5.7
21 months 7.3 16.1 7.1
24 months 5.1 11.6 5.6
26 months 3.5 14.4 5.6
Thyrotrophin receptor-blocking antibodies in transient congenital hypothyroidism 267EUROPEAN JOURNAL OF ENDOCRINOLOGY (2004) 150
www.eje.org
birth. Hypoplastic thyroid glands have also been
described in some patients with TSH-R loss-of-function
mutations (references in 11). Therefore it seems that
TSH/TSH-R signalling is of importance for the develop-
ment of a normally sized thyroid gland in humans in
utero, although it has recently been demonstrated
that TSH or a functional TSH-R is not required for
the development of a normal sized thyroid gland in
mice in utero (11).
Although functional TSH/TSH-R signalling seems to
be required for production of a normal sized gland, the
gland of the infant described here and those of pre-
viously described patients with inactivating mutations
of the TSH receptor are normally sited and serum
thyroglobulin is increased. These observations support
the hypothesis that migration and development of the
thyroid gland is TSH independent, although functional
TSH-R signalling may be required to achieve a fully dif-
ferentiated thyroid phenotype (12).
We have presented evidence for TBAb activity in
breast milk. Others (9) have previously observed the
presence of TSAb in breast milk but the clinical signifi-
cance is not clear. We speculate that the TBAb in the
milk did not contribute significantly to hypothyroidism
in the child, since clearing of the antibodies from his
circulation was evident over time.
This case has demonstrated that transplacental pas-
sage of TBAb can result in profound neonatal hypothyr-
oidism at birth. It is essential to start thyroxine
replacement without delay. The transient nature of
the hypothyroidism can then be determined by
measurement ideally of TBAb or indirectly through
measurement of TSH-binding inhibitory immunoglobu-
lins in maternal serum. Affected infants require thyrox-
ine therapy until TBAb clears from their circulation, so
allowing normal thyroid function to resume. However,
potent TBAb in the infant’s circulation may damage
or delay development of the thyroid gland, therefore
thyroid function should be carefully monitored when
thyroxine is withdrawn.
Acknowledgements
We thank Dr C Williams (Wrexham Maelor Hospital)
and Dr M Lewis (UWCM) for their assistance and the
Welsh Office of Research and Development for Health
and Social Care for a studentship for N J J.
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Received 7 August 2003
Accepted 21 November 2003
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www.eje.org
