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Thyroxine derivatives in amniotic fluid
December, 1971 T h e Journal o/ P E D I A T R I C S
923
Thyroxine derivatives in amniotic fluid Fetal outcome in three patients with thyroid problems Amniotie fluid protein-bound iodine determinations have been measured in 20 euthyrold women in the sixteenth to fortieth week of gestation. The values ranged from 0.7 to 2.2 #g per I00 ml. with a mean of 1.3 t*g per 100 ml. T, by column values in 11 of the women ranged from 0.7 to 2.0 #g per 100 ml. with a mean of 1.2 #g per 100 ml. Two patients with abnormal thyroid function and one woman who had previously given birth to a child with cretinism were studied. The amniotic fluid protein-bound iodine level in each instance indicated the subsequent thyroid status of the infant. Amniocentesis is a potentially useful procedure in antenatal evaluation of thyroid function.
Dorothy R. Hollingsworth, M.D., and Elizabeth Austin, B.S., M.T. (ASCP) LEXINGTON~ KY.
T I-I E lu A T lg R I A L S in the amniotic fluid frequently give an indication of the state of the fetus? For example, a decrease in bilirubin and an increase in creatinine are indices of maturation, 2 and the measurement of bilirubin-heme chromagen in amniotic fluid aids in the management of pregnancy in Rh-sensitized women? "5 The antenatal detection of certain cytogenetic and biochemical abnormalities is also feasible by examination of cells from the amniotic fluid. 6-8 Rose and associates 9 first measured protein-bound iodine (PBI) in amniotic fluid obtained by direct needle aspiration through the partially dilated cervix. By a modification of the dry ash method they found only scant amounts (0.3 to 0.8 /~g per 100 ml.) of PBI in the amniotic fluid of 4 subjects; they concluded that amniotic fluid contains From the Department of Pediatrics, University of Kentucky Medical Center. Reprint address: Dorothy R. Holllngsworth, M.D., Department of Pediatrics, Universi.ty o] Kentucky Medical Center, Lexington, Ky. 40506.
no appreciable amount of PBI. Subsequently, Kaufman 1~ measured amniotic PBI from 18 Rh-negative multiparous women; his measurements, 1.6 to 3.8 /Lg per 100 ml., indicated that there was significant PBI in amniotic fluid. Recently, we evaluated two pregnant patients with hyperthyroidism and a third who had previously given birth to an infant with athyrotic cretinism. I n an attempt to ascertain prenatal information concerning the thyroid status of the fetuses, amniotic fluid was obtained and PBI measured. Twenty patients with normal thyroid function were studied and compared with the patients with thyroid problems. The results indicate that analysis of amniotic fluid might be useful in assessing abnormal thyroid function in the fetus.
METHODS Amniocentesis. This procedure was performed on outpatients by the obstetrics and Vol. 79, No. 6, pp. 923-929
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Hollingsworth and Austin
gynecology staff of the University of K e n tucky M e d i c a l Center. A f t e r voiding, the patient was p l a c e d in the supine position. T h e a b d o m e n was d r a p e d a n d p r e p a r e d with a noniodine-containing antiseptic solution. A local anesthetic was used a t the site of aspiration a n d a 20 gauge spinal needle inserted over the fetal small parts. A p p r o x i mately 10 to 20 ml. of fluid was w i t h d r a w n for chemical analysis. PBI values were d e t e r m i n e d by a modified a u t o m a t e d m e t h o d devised by Riley a n d G o c h m a n using the T e c h n i c o n AutoAnalyzer. 11 T h y r o x i n e (T4) by column was m e a sured by a modification of the Pileggi m e t h od. ~2 T h y r o i d - s t i m u l a t i n g h o r m o n e was det e r m i n e d by the Odell m e t h o d in the U n i versity of K e n t u c k y T h y r o i d Laboratory23 (Long-acting thyroid stimulator d e t e r m i n a tions were done by Dr. J. M. M c K e n z i e , M o n t r e a l , C a n a d a J 4) Control patients were euthyroid p r e g n a n t w o m e n in the obstetrics clinic who were having amniocentesis for clinical evaluation of R h sensitization of the fetus. CASE REPORTS
Case 1. At age I4 years, this patient (A. Mc.) was found to have exophthalmos and nervousness. A thyroidectomy was performed, and she remained asymptomatic for 20 years. In September, 1968, the patient, now a 34-year-old unmarried woman, was admitted to the University of Kentucky Medical Center with a short history of nervousness, heat intolerance, irritability, exophthalmos, and tachycardia. Laboratory studies revealed a PBI of 16.0 /~g per 100 ml. and an 1131 uptake of 61 per cent in 24 hours. Soon afterward, and with no knowledge of her early pregnancy, she received 10 mc. I T M and was started on propyhhiouracil, 300 mg. per day. Four months later she was found to be hypothyroid with a PBI of 2.6 /~g per 100 ml.; she was also found to be 7 months pregnant. The 1TM had been given in the ninth week of gestation. Amnlocentesls was performed and amniotic fluid PBI found to be 0.4/~g per 100 mh Propylthiouracil was discontinued and treatment with l-triiodothyronine (Cytomel), 100 #g per day, was started. Two months later a male infant weighing 2,825 Gin. was delivered. Cord blood studies revealed a PBI of 5.0 #g per 100 ml.,
The Journal o/ Pediatrics December 1971
butanol-extractable iodine of 4.5 /xg per 100 ml. and alkaline phosphatase of 11.0 BLB units. There was oniy faint ossification of the distal femoral epiphyses and no ossification of the proximal tibial epiphyses. Syndactyly of third and fourth fingers of the left hand and second and third toes of the left foot was present. There was no cretinoid facies, umbilical hernia, or neonatal jaundice. Desiccated thyroid was administered to the infant for i1 months. On re-evaluation two months after stopping medication, serum PBI (6.5 /~g per 100 ml.), T 4 by column (6.8 ,ag per 100 mI.), alkaline phosphatase (12 BLB), and 113z uptake (6 hours 13 per cent and 24 hours 25 per cent) were within normal limits. Bone age was within two standard deviations for age. The baby is euthyroid on no medication at the present time. T h e mother is also euthyroid and receiving no treatment. Case 2. Patient K. H., a 22-year-old unmarried Caucasian woman, who developed Graves disease one month after conception, was referred to the University of Kentucky Medical Center. Her symptoms were characterized by heat intolerance, increased sweating, nervousness, and fatigue. She was found to have a rapid pulse (I40 per minute), systolic hypertension (blood pressure 160/70), exophthalmos, and a thyroid gland that was moderately enlarged. Amniotic fluid was aspirated and amniotic fluid PBt was found to be 3.8 p~g per 100 mI. A serum PBI obtained at the same time was 17.8 /~g per 100 mI. The patient was started on 600 mg. of propylthiouracil per day, but she was erratic in taking the medication. One week later she was admhted to the hospital because of uncontrolled hyperthyroidism. Propylthiouracil was increased te 900 rag. per day, and propranolol hydrochloride, 40 rag. per day, was also administered to control tachycardia. Two weeks later 100 /~g of l-triiodothyronine per day (Cytomel) was added and propylthiouracil decreased to 600 mg. per day. A second amnlocentesis revealed an amniotic fluid PBI of 5.7 k~g per 100 mt. but no measurable long-acting thyroid stimulator. A month later the patient's hyperthyroidism was stilI uncontrolled; she was severely toxic with exophthalmos, tremor, sweating, nervousness, and tachycardia (130 per minute). The fetal heart rate was 240 per minute. Amniotic fluid had been leaking per cervical os for 5 days. Lugol's solution was added to her treatment. Two days later, at 33 weeks' gestation, she delivered a boy weighing 2,310 Gm. The newborn
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Thyroxine derivatives in amniotic fluid
infant appeared malnourished and had a large goiter. Laboratory studies at birth included: cord blood T 4 (Murphy-Pattee) 5.6 /xg per 100 ml., T 4 by column 4.75 /zg per 100 ml., free T 4 1.6 /xg per 100 mI. These determinations were felt to reflect a summation of maternal medication with propylthiouracil, /-triiodothyronine, and Lugol's solution. The infant's serum at birth contained negligible thyroid-stimulatlng hormone
(3 m~). Long-acting thyroid stimulator assays by McKenzie's laboratory were as follows:
24 hr.% + S.D.
Date
5% Human serum albumin 10-23-69 Mother 10-23-69 Amniotic fluid 12-6-69 1-2-70
Cord blood Infant 4 wk.
Significance
72-+ 34 Control 282 + 52 Significant 77 + 6 Not significant 607 + 316 Significant 151 + 21 Significant
Initially the baby appeared euthyroid, but by t0 days he had weight loss, tachycardia (250 per minute), hyperactivity, and exophthalmos. A repeat Ta by column was > 20 /~g per 100 ml. Treatment was started with Lugol's solution and propylthiouracil (30 mg. per day). At 6 weeks of age these medications were tapered and stopped, but a relapse occurred and full treatment was resumed until the tenth week of life. He was discharged from the hospital at age 3 months receiving no medications and weighing 4,060 Gm. Three weeks later it was evident that his exophthalmos had increased and his weight had decreased (3,760 Gin.). T 4 by column was elevated (12.4 /zg per 100 ml.) and treatment was restarted with propylthiouracil 60 rag. per day. PBI remained ~levated (12.8 /~g per 100 ml.) and weight decreased from 4,840 Gm. at 7 months to 4,760 Gm. at 9 months. Medication was changed m methimazole (Tapazole) 5 mg. three times per day, following which he gained weight and became less hyperactive. At 11 months of age weight had increased to 6,280 Gm. and T , by column was 3.4 /xg per 100 ml. He continued to have a tachycardia of 180, prominent eyes, and a palpable thyroid measuring 4.0 cm. No tremor was noted and reflexes were 2+. At age one year bone age was normal and development appropriate for age. Both height and weight were below the tenth percentile (height 27 inches and weight 6,250 Gm.). Methimazote was stopped on his first birthday. Three months later he was
925
gaining poorly, had prominent eyes, a small goiter, and occasional diarrhea. PBI was normal (7.6 /xg per 100 ml.), and serum long-acting thyroid stimulator determination was now negative. Case 3. Patient B. J. was a 41-year-old married Caucasian woman who was seen at the University of Kentucky Medical Center in the sixth month of pregnancy. Six years earlier she had given birth to a 7 pound infant with athyrotic cretinism; she was concerned about the status of the present fetus. Physical examination was unremarkable aside from the 5 ~ month pregnancy. Serum PBI was 12.3 /~g per 100 ml. (normal for pregnancy 7 to 19 /xg per 100 ml.) and she appeared euthyroid. Amniotic fluid PBI was 1.6 /~g per 100 mt. Empirically she was started on /-triiodothyronine 75 /xg per day. She delivered a normal-appearing full-term male infant weighing 3,175 Gin. Cord blood PBI was 9.2 /xg per 100 ml. and the infant's femoral and tibial epiphyses appeared normal by roentgen examination. No medication was given to the infant. At age one year he had grown and developed normally and had normal thyroid function. RESULTS
Amniocentesis was p e r f o r m e d in 90 euthyroid p r e g n a n t women. A m n i o t i c fluid PBI d e t e r m i n a t i o n s r a n g e d from 0.7 to 2.2 /~g p e r 100 ml. with no t r e n d suggesting higher values with increasing gestational age (Fig. 1). In 11 of the patients a n amniotic fluid T4 by column was also done with values r a n g i n g f r o m 0.7 to 9.0 /~g p e r 100 ml. S e r u m PBI determinations in 12 patients d r a w n at the same time as the amniocentesis varied from 7.4 to 12.1 /zg p e r 100 ml. T a b l e I compares n o r m a l p r e g n a n c y values with those o b t a i n e d from p r e g n a n t women with hypo- or hyperthyroidism a n d from a n euthyroid w o m a n who h a d previously given birth to a child with cretinism. T h e first p a t i e n t was h y p o t h y r o i d after t r e a t m e n t of Graves' disease with I T M a n d propylthiouracil. S e r u m PBI a n d amniotic fluid PBI were very low. Because of the risk of fetal hypothyroidism, the m o t h e r was treated with l-triiodothyronine. T h e second patient, who h a d hyperthyroidism a n d a ser u m PBI of 17.8 /~g p e r 100 ml., h a d a
9 2 6 HolIingsworth and Austin
The ]ournal o[ Pediatrics December t971
T a b l e I. S e r u m PBI, amniotlc fluid PBI, a n d T4 b y column d u r i n g p r e g n a n c y
Sub}ects
Week o[ ~re~nancy
(PRI
Serum #,g/IO0
ml.)'
Amniot(c fluid T, by column PBI ~.~/I00 ml. #g/lO0 ml.
Patients 1. A, Mc. (Hypothyrold)
30
4.6
0.4,
2. K . H . (Hyperthyrold)
27 31
17.8
3.8 5.7
3. B. J. (Euthyrold mother of a child with cretinism)
26
12.3
1.6
1.22
Controls I. C . S . 2. E . W . 3. M . S . 4. C . S . 5. B . D . 6. E . B . 7. B . K . 8. S . H . 9. R . R . 10. J . B . 11. J . D . 12. A . H . 13. L . C . 14. S . S . 15. P . S . 16. E . R . 17. B . H . 18. B . A . 19. W . M . 20. H. LeC. Mean Range
16 19 20 20 24 27 27 28 30 32 32 32 33 34 35 36 36 38 40 40
greatly elevated amniotic fluid PBI on two occasions (3.8 a n d 5.7 /~g p e r 100 m l . ) . Alt h o u g h this infant a p p e a r e d euthyroid at birth, p r o b a b l y reflecting m a t e r n a l m e d i c a tions, he developed neonatal Graves' disease by 10 days of age. T h e third p a t i e n t h a d a n o r m a l serum P B I a n d a n o r m a l amniotic fluid P B I a n d T , by colunm. H e r infant was normal. DISCUSSION O u r findings in euthyroid patients supp o r t K a u f m a n ' s observation t h a t significant amounts of PBI are measurable in amniotic fluid. A l t h o u g h Fisher a n d associates 1~ have found low fetal serum T4 a n d free T4 between 11 a n d 18 weeks' gestation a n d a p r o gressive increase in values between 22 weeks
9.2 8.6 8.8 9.8 12.1 8.8 7.8 7.4 9.6 8.5 8.2 11.2 9.0 7.4 - 12.1
1.0 1.8 2.2 2.0 .7 1.7 1.6 1.2 1.0 1.2 1.2 1.5 1.3 1.4 1.8 1.0 1.4 1.8 1.6 1.3 1.33 0.7 - 2.2
2.0 1.4 1.2 1.7 1.2 1.8 1.8 1.1 0.7 1.0 1.1 0.7
1.26 0.7 - 2.0
a n d term, we have been unable to demonstrate increased amniotic fluid PBI with gestational age. Fisher a n d associates' study revealed low fetal serum thyrold-stimulating h o r m o n e values to 18 weeks a n d an a b r u p t increase of thyroid-stimulating h o r m o n e from 18 to 22 weeks. G r e e n b e r g a n d associates 1G have shown an increase in fetal free thyroxine from the tenth to twentieth week of gestation, with fetal levels reaching within one s t a n d a r d deviation of the term level by 20 weeks. All b u t two of our amniotic fluid levels were obtained after 20 weeks' gestation, w h e n fetal thyroid-stimulating h o r m o n e a n d feedback mechanisms were already functioning; this m a y explain the failure to note higher values with lengthening gestation. I n 3 patients serial determinations were done in the second a n d t h i r d
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Thyroxine derivatives in amniotic fluid
3.0 2.5 2.0
O
o_
05
9
1.5
1.0 .5
0
---~/
i
t
i
i
I
i
15
20
25
513
35
40
GESTATION (wks.)
Fig. 1. PBI (#g/100 ml. of amniotic fluid) plotted against gestational weeks for 29 samples in 20 normal patients.
trimesters and no rise was noted with fetal age. Previous studies have shown that by the third month of gestation the fetal thyroid is able to concentrate radioactive iodine17; thyroxine synthesis is known to occur by about 3 ~ months? 8 The studies of Fisher and associates 15 and Greenberg and associates 16 on abortion material have shown autonomous function of the fetal hypothalamic-pituitary control system to be evident by t2 weeks' gestation. French and Van Wyck,19 in discussing fetal hypothyroidism, have reviewed the pituitary regulation of fetal thyroid function. Maternal thyroidstimulating hormone does not cross the placenta, 2~ and in experimental animals goiters develop in the absence of a maternal pituitary, but not in the absence of a fetal pituitary. 22-23 Thus if the fetal pituitary-thyroid axis is normal, he can function adequately even though there is maternal hypothyroidism. Such cases have been reported, 2~-25 but prenatal data on infants at risk are lacking. Antenatal hypothyroidism causes irreversible impairment of brain development, and neonatal hyperthyroidism may be life threatening. It would be extremely helpful if some indication of fetal thyroid status might be obtained during pregnancy.
92 7
The first mother of our series was hypothyroid after having been given I T M at the ninth week of gestation, followed by propylthiouracil; we were concerned that possible fetal thyroid cellular damage might prevent a normal glandular response to fetal thyroidstimulating hormone. Because of the low amniotic fluid PBI the mother was given /-triiodothyronine for two months until delivery in the hope that enough placental transfer would occur to prevent intrauterine hypothyroidism. A full-term male infant was born without stigmata of cretinism, but with no ossification of the proximal tibial epiphyses and only faint ossification of the distal femoral epiphyses. Replacement therapy with desiccated thyroid was given for the first 9 months. The child was re-evaluated after having received no medication for 11 weeks and found to have normal thyroid function. The transient hypothyroidism may have been related to either propylthiouracil or I ~sl therapy. In the second case we were concerned about whether the baby was hyperthyroid in utero. A high amniotic fluid PBI was found which later turned out to reflect the baby's thyroid status. Long-acting thyroid stimulator was not demonstrable in the amniotic fluid, but was present in maternal serum, cord blood, and the baby's serum at birth. The child was born 6 weeks prematurely and had Graves' disease. Not all hyperthyroid mothers have hyperthyroid babies. Those who do often have demonstrable serum long-acting thyroid stimulator, a 7 S globulin, which may traverse the placenta and stimulate the fetus. Contrary to reports in the literature stating the half life of long-acting thyroid stimulator to be approximately 6 days, 26 excessive amounts were still measurable in our patient at one month of age and the baby has had prolonged rather than transient hyperthyroidism. He continues to have growth failure, prominent eyes, and a small goiter at 16 months, although he now has a normal PBI level and no demonstrable longacting thyroid stimulator. In the third case the patient was euthy-
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Hollingsworth and Austin
roid b u t h a d previously given b i r t h to a child with cretinism. I n this instance the amniotic fluid PBI was normal. This was reassuring b u t our experience with amniotic fluid PBI determinations was scant a n d the m o t h e r was given / - t r i i o d o t h y r o n i n e Y A n a p p a r e n t l y n o r m a l male infant was born at term. T h i s child remains n o r m a l at one y e a r of age. Decreased or absent fetal thyroid horm o n e formation m a y occur with iodine deprivation, with a n enzymatic defect in thyroxine synthesis, or with failure of m i g r a t i o n or absence of the thyroid gland. I n addition, i n a d v e r t e n t exposure to I ~sl at a critical time, excessive iodine or blocking agents m a y likewise depress fetal thyroid h o r m o n e synthesis. N o w t h a t new knowledge of pret e r m n o r m a l thyroid function has become available from abortion studies, an a t t e m p t should be m a d e to d o c u m e n t a b n o r m a l fetal thyroid function prenatally. Retrospectively, amnlotic fluid PBI determh~ations reflected the fetal thyroid status of each of our 3 babies at risk. T h e use of amniotic fluid PBI should be explored further a n d correlated with long-term observations on infants of mothers with thyroid disease a n d those who have previously given birth to children with cretinism. The authors wish to thank Dr. Philip K~ Bondy, Dr. C. Charlton Mabry, Dr. Jacquellne Noonan, and Dr. Warren Wheeler for their helpful comments and review of the manuscript. We are grateful to Dr. J. M. McKenzie of Montreal, Canada, for doing the long-acting thyroid stimulator assays.
REFERENCES I. Fuchs, F.: Amniotie fluid, Clin. Obstet, Gynecol. 9: 425, 1966. 2. Mandelbaum, B., and Evans, T. N." Life in the amniotie fluid, Am. J. Obstet. Gynecol, 104: 365, 1969. 3. Freda, V. J.: Rh problem in obstetrics and a new concept of its management using amniocentesis and spectrophotometric scanning of amniotic fluid, Am. J. Obstet. Gynecol. 92; 341, 1965. 4. Queenan, J. T., and Adams, D. W.: Amniocentesis for prenatal diagnosis of erythroblastosis fetalis, Obstet. Gynecol. 25: 302, 1965.
The Journal o/ Pediatrics December 1971
5. Queenan, J. T., and Wyatt, R. H.: Intrauterine transfusion of fetus for severe erythroblastosis fetalis, Am. J. Obstet. Gynecol. 92: 796, 1967. 6. Jacobson, C. B., and Barter, R. H.: Intrauterine diagnosis and management of genetic defects, Am. J. Obstet. Gynecol. 99: 796, 1967. 7. Jeffcoate, T. N. A., Fliegner, J. R. H., Russell, S. H., Davis, J. C., and Wade, A. P.: Diagnosis of adrenogenital syndrome before birth, Lancet 2: 553, 1965. 8. Frantantoni, J. C., Neufeld, E. F., Uhlendorf, W., and ffacobson, C. B.: Intrauterine diagnosis of Hurler and Hunter syndromes, N. Engl. J. Med. 280: 686, 1969. 9. tl.ose, H., Russell, K. P., and Starr, P.: Serum protein-bound iodine of mothers and infants at delivery in premature and term pregnancies, Am. J. Obstet. Gynecol. 86: 767, 1963. 10. Kaufman, S.: Protein-bound iodine (PBI) in human amniotic fluid, J. PEDIATR. 68: 990, 1966. 11. Austin, E , and Koepke, J. A.: An automated procedure for total organic iodine, Am. J. Clin~ Pathol. 45: 344, 1966. 12. Kessler, G., and Pileggi, V. J.: A Semiautomated nonincineration technique for determining serum thyroxine, Clin. Chem. 16: 382, 1970. 13. Odell, W. D., Wilbur, J. F., and Paul, W. E.: Radioimmunoassay of thyrotrophin in human serum, J. Clin. Endocrinol Metab. 25: 1179, 1965. 14. McKenzie, J. M., and Williamson, A.: Experience with the blo-assay of the long acting thyroid stimulator, J. Clin. Endocrinol. Metab. 26" 518, 1966. 15. Fisher D. A., Hobel, C. J., Garza, R., and Pierce, C. A.: Thyroid function in the preterm fetus, Pediatrics 46: 208, 1970. 16. Greenberg, A. H., Czernichow, P., Reba, R. C., Tyson, J., and Blizzard, R. M.: Observations on the maturation of thyroid function in early fetal life, J. Clin. Invest. 49: 1790, 1970. 17. Hodges, R. 2., Evans, T. C., Bradbury, J. T., and Keettel, W. C.: The accumulation of radioactive iodine by human fetal thyroids, J. Clin. Endocrinol. Metab. 15: 661, 1955. 18. Shepherd, T. H.: Onset of function in the human fetal thyroid: Biochemical and radioautographic studies from organ culture, J. CIin. Endocrinol. Metab. 27: 954, 1967. 19. French, F. S., and Van Wyk, J. J.: Fetal hypothyroidism, J. P~DIATR. 64: 589, 1964. 20. Postel, S.: Placental transfer of perchIorate and triiodothyronine in the guinea pig, Endocrinology 60: 50, 1957. 21. Peterson, R. R., and Young, W. G.: The problem of placental permeability for thyrotropin, propylthiouracil and thyroxine in the guinea pig, Endocrinology 50: 218, 1952. 22. Nikitovitch, M., and Knobil, E.: Placental transfer of TSH in the rat, J. Clin. Endocrinol. Metab. 15: 837, 1955.
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23. Sobel, E. H., Hamburg, M., and Koblln, R.: Development of the fetal thyroid in rats; Evidence for a placental transfer of thyroxine, Am. J. Dis. Child. 100: 709, 1960. 24. Lister, L. M., and Ashe, J. R., Jr.: Pregnancy and myxedema: Report of a case, Obstet. Gynecol. 6: 436, 1955. 25. Parkin, G., and Green, J. A.: Pregnancy occurring in cretinism and in juvenile and adult myxedema, J. Clin. Endoerinol. Metab. 3: 466, 1943.
Thyroxine derivatives in amniotic fluid
9 29
26. Sunshine, P., Kusumoto, H., and Krlss, J. P.: Survival time of LATS in neonatal thyrotoxicosis: Implications for diagnosis and therapy of the disorder, Pediatrics 38: 869, 1965. 27. Dussault, J., Row, V. V., Lickrish, G., and Vocpe, R.: Studies of serum triiodothyronine concentration in maternal and cord blood: Transfer of trliodothyronine across the human placenta, J. Clin. Endoerinol. Metab. 29: 595, 1969.
923
Thyroxine derivatives in amniotic fluid Fetal outcome in three patients with thyroid problems Amniotie fluid protein-bound iodine determinations have been measured in 20 euthyrold women in the sixteenth to fortieth week of gestation. The values ranged from 0.7 to 2.2 #g per I00 ml. with a mean of 1.3 t*g per 100 ml. T, by column values in 11 of the women ranged from 0.7 to 2.0 #g per 100 ml. with a mean of 1.2 #g per 100 ml. Two patients with abnormal thyroid function and one woman who had previously given birth to a child with cretinism were studied. The amniotic fluid protein-bound iodine level in each instance indicated the subsequent thyroid status of the infant. Amniocentesis is a potentially useful procedure in antenatal evaluation of thyroid function.
Dorothy R. Hollingsworth, M.D., and Elizabeth Austin, B.S., M.T. (ASCP) LEXINGTON~ KY.
T I-I E lu A T lg R I A L S in the amniotic fluid frequently give an indication of the state of the fetus? For example, a decrease in bilirubin and an increase in creatinine are indices of maturation, 2 and the measurement of bilirubin-heme chromagen in amniotic fluid aids in the management of pregnancy in Rh-sensitized women? "5 The antenatal detection of certain cytogenetic and biochemical abnormalities is also feasible by examination of cells from the amniotic fluid. 6-8 Rose and associates 9 first measured protein-bound iodine (PBI) in amniotic fluid obtained by direct needle aspiration through the partially dilated cervix. By a modification of the dry ash method they found only scant amounts (0.3 to 0.8 /~g per 100 ml.) of PBI in the amniotic fluid of 4 subjects; they concluded that amniotic fluid contains From the Department of Pediatrics, University of Kentucky Medical Center. Reprint address: Dorothy R. Holllngsworth, M.D., Department of Pediatrics, Universi.ty o] Kentucky Medical Center, Lexington, Ky. 40506.
no appreciable amount of PBI. Subsequently, Kaufman 1~ measured amniotic PBI from 18 Rh-negative multiparous women; his measurements, 1.6 to 3.8 /Lg per 100 ml., indicated that there was significant PBI in amniotic fluid. Recently, we evaluated two pregnant patients with hyperthyroidism and a third who had previously given birth to an infant with athyrotic cretinism. I n an attempt to ascertain prenatal information concerning the thyroid status of the fetuses, amniotic fluid was obtained and PBI measured. Twenty patients with normal thyroid function were studied and compared with the patients with thyroid problems. The results indicate that analysis of amniotic fluid might be useful in assessing abnormal thyroid function in the fetus.
METHODS Amniocentesis. This procedure was performed on outpatients by the obstetrics and Vol. 79, No. 6, pp. 923-929
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Hollingsworth and Austin
gynecology staff of the University of K e n tucky M e d i c a l Center. A f t e r voiding, the patient was p l a c e d in the supine position. T h e a b d o m e n was d r a p e d a n d p r e p a r e d with a noniodine-containing antiseptic solution. A local anesthetic was used a t the site of aspiration a n d a 20 gauge spinal needle inserted over the fetal small parts. A p p r o x i mately 10 to 20 ml. of fluid was w i t h d r a w n for chemical analysis. PBI values were d e t e r m i n e d by a modified a u t o m a t e d m e t h o d devised by Riley a n d G o c h m a n using the T e c h n i c o n AutoAnalyzer. 11 T h y r o x i n e (T4) by column was m e a sured by a modification of the Pileggi m e t h od. ~2 T h y r o i d - s t i m u l a t i n g h o r m o n e was det e r m i n e d by the Odell m e t h o d in the U n i versity of K e n t u c k y T h y r o i d Laboratory23 (Long-acting thyroid stimulator d e t e r m i n a tions were done by Dr. J. M. M c K e n z i e , M o n t r e a l , C a n a d a J 4) Control patients were euthyroid p r e g n a n t w o m e n in the obstetrics clinic who were having amniocentesis for clinical evaluation of R h sensitization of the fetus. CASE REPORTS
Case 1. At age I4 years, this patient (A. Mc.) was found to have exophthalmos and nervousness. A thyroidectomy was performed, and she remained asymptomatic for 20 years. In September, 1968, the patient, now a 34-year-old unmarried woman, was admitted to the University of Kentucky Medical Center with a short history of nervousness, heat intolerance, irritability, exophthalmos, and tachycardia. Laboratory studies revealed a PBI of 16.0 /~g per 100 ml. and an 1131 uptake of 61 per cent in 24 hours. Soon afterward, and with no knowledge of her early pregnancy, she received 10 mc. I T M and was started on propyhhiouracil, 300 mg. per day. Four months later she was found to be hypothyroid with a PBI of 2.6 /~g per 100 ml.; she was also found to be 7 months pregnant. The 1TM had been given in the ninth week of gestation. Amnlocentesls was performed and amniotic fluid PBI found to be 0.4/~g per 100 mh Propylthiouracil was discontinued and treatment with l-triiodothyronine (Cytomel), 100 #g per day, was started. Two months later a male infant weighing 2,825 Gin. was delivered. Cord blood studies revealed a PBI of 5.0 #g per 100 ml.,
The Journal o/ Pediatrics December 1971
butanol-extractable iodine of 4.5 /xg per 100 ml. and alkaline phosphatase of 11.0 BLB units. There was oniy faint ossification of the distal femoral epiphyses and no ossification of the proximal tibial epiphyses. Syndactyly of third and fourth fingers of the left hand and second and third toes of the left foot was present. There was no cretinoid facies, umbilical hernia, or neonatal jaundice. Desiccated thyroid was administered to the infant for i1 months. On re-evaluation two months after stopping medication, serum PBI (6.5 /~g per 100 ml.), T 4 by column (6.8 ,ag per 100 mI.), alkaline phosphatase (12 BLB), and 113z uptake (6 hours 13 per cent and 24 hours 25 per cent) were within normal limits. Bone age was within two standard deviations for age. The baby is euthyroid on no medication at the present time. T h e mother is also euthyroid and receiving no treatment. Case 2. Patient K. H., a 22-year-old unmarried Caucasian woman, who developed Graves disease one month after conception, was referred to the University of Kentucky Medical Center. Her symptoms were characterized by heat intolerance, increased sweating, nervousness, and fatigue. She was found to have a rapid pulse (I40 per minute), systolic hypertension (blood pressure 160/70), exophthalmos, and a thyroid gland that was moderately enlarged. Amniotic fluid was aspirated and amniotic fluid PBt was found to be 3.8 p~g per 100 mI. A serum PBI obtained at the same time was 17.8 /~g per 100 mI. The patient was started on 600 mg. of propylthiouracil per day, but she was erratic in taking the medication. One week later she was admhted to the hospital because of uncontrolled hyperthyroidism. Propylthiouracil was increased te 900 rag. per day, and propranolol hydrochloride, 40 rag. per day, was also administered to control tachycardia. Two weeks later 100 /~g of l-triiodothyronine per day (Cytomel) was added and propylthiouracil decreased to 600 mg. per day. A second amnlocentesis revealed an amniotic fluid PBI of 5.7 k~g per 100 mt. but no measurable long-acting thyroid stimulator. A month later the patient's hyperthyroidism was stilI uncontrolled; she was severely toxic with exophthalmos, tremor, sweating, nervousness, and tachycardia (130 per minute). The fetal heart rate was 240 per minute. Amniotic fluid had been leaking per cervical os for 5 days. Lugol's solution was added to her treatment. Two days later, at 33 weeks' gestation, she delivered a boy weighing 2,310 Gm. The newborn
Volume 79 Number 6
Thyroxine derivatives in amniotic fluid
infant appeared malnourished and had a large goiter. Laboratory studies at birth included: cord blood T 4 (Murphy-Pattee) 5.6 /xg per 100 ml., T 4 by column 4.75 /zg per 100 ml., free T 4 1.6 /xg per 100 mI. These determinations were felt to reflect a summation of maternal medication with propylthiouracil, /-triiodothyronine, and Lugol's solution. The infant's serum at birth contained negligible thyroid-stimulatlng hormone
(3 m~). Long-acting thyroid stimulator assays by McKenzie's laboratory were as follows:
24 hr.% + S.D.
Date
5% Human serum albumin 10-23-69 Mother 10-23-69 Amniotic fluid 12-6-69 1-2-70
Cord blood Infant 4 wk.
Significance
72-+ 34 Control 282 + 52 Significant 77 + 6 Not significant 607 + 316 Significant 151 + 21 Significant
Initially the baby appeared euthyroid, but by t0 days he had weight loss, tachycardia (250 per minute), hyperactivity, and exophthalmos. A repeat Ta by column was > 20 /~g per 100 ml. Treatment was started with Lugol's solution and propylthiouracil (30 mg. per day). At 6 weeks of age these medications were tapered and stopped, but a relapse occurred and full treatment was resumed until the tenth week of life. He was discharged from the hospital at age 3 months receiving no medications and weighing 4,060 Gm. Three weeks later it was evident that his exophthalmos had increased and his weight had decreased (3,760 Gin.). T 4 by column was elevated (12.4 /zg per 100 ml.) and treatment was restarted with propylthiouracil 60 rag. per day. PBI remained ~levated (12.8 /~g per 100 ml.) and weight decreased from 4,840 Gm. at 7 months to 4,760 Gm. at 9 months. Medication was changed m methimazole (Tapazole) 5 mg. three times per day, following which he gained weight and became less hyperactive. At 11 months of age weight had increased to 6,280 Gm. and T , by column was 3.4 /xg per 100 ml. He continued to have a tachycardia of 180, prominent eyes, and a palpable thyroid measuring 4.0 cm. No tremor was noted and reflexes were 2+. At age one year bone age was normal and development appropriate for age. Both height and weight were below the tenth percentile (height 27 inches and weight 6,250 Gm.). Methimazote was stopped on his first birthday. Three months later he was
925
gaining poorly, had prominent eyes, a small goiter, and occasional diarrhea. PBI was normal (7.6 /xg per 100 ml.), and serum long-acting thyroid stimulator determination was now negative. Case 3. Patient B. J. was a 41-year-old married Caucasian woman who was seen at the University of Kentucky Medical Center in the sixth month of pregnancy. Six years earlier she had given birth to a 7 pound infant with athyrotic cretinism; she was concerned about the status of the present fetus. Physical examination was unremarkable aside from the 5 ~ month pregnancy. Serum PBI was 12.3 /~g per 100 ml. (normal for pregnancy 7 to 19 /xg per 100 ml.) and she appeared euthyroid. Amniotic fluid PBI was 1.6 /~g per 100 mt. Empirically she was started on /-triiodothyronine 75 /xg per day. She delivered a normal-appearing full-term male infant weighing 3,175 Gin. Cord blood PBI was 9.2 /xg per 100 ml. and the infant's femoral and tibial epiphyses appeared normal by roentgen examination. No medication was given to the infant. At age one year he had grown and developed normally and had normal thyroid function. RESULTS
Amniocentesis was p e r f o r m e d in 90 euthyroid p r e g n a n t women. A m n i o t i c fluid PBI d e t e r m i n a t i o n s r a n g e d from 0.7 to 2.2 /~g p e r 100 ml. with no t r e n d suggesting higher values with increasing gestational age (Fig. 1). In 11 of the patients a n amniotic fluid T4 by column was also done with values r a n g i n g f r o m 0.7 to 9.0 /~g p e r 100 ml. S e r u m PBI determinations in 12 patients d r a w n at the same time as the amniocentesis varied from 7.4 to 12.1 /zg p e r 100 ml. T a b l e I compares n o r m a l p r e g n a n c y values with those o b t a i n e d from p r e g n a n t women with hypo- or hyperthyroidism a n d from a n euthyroid w o m a n who h a d previously given birth to a child with cretinism. T h e first p a t i e n t was h y p o t h y r o i d after t r e a t m e n t of Graves' disease with I T M a n d propylthiouracil. S e r u m PBI a n d amniotic fluid PBI were very low. Because of the risk of fetal hypothyroidism, the m o t h e r was treated with l-triiodothyronine. T h e second patient, who h a d hyperthyroidism a n d a ser u m PBI of 17.8 /~g p e r 100 ml., h a d a
9 2 6 HolIingsworth and Austin
The ]ournal o[ Pediatrics December t971
T a b l e I. S e r u m PBI, amniotlc fluid PBI, a n d T4 b y column d u r i n g p r e g n a n c y
Sub}ects
Week o[ ~re~nancy
(PRI
Serum #,g/IO0
ml.)'
Amniot(c fluid T, by column PBI ~.~/I00 ml. #g/lO0 ml.
Patients 1. A, Mc. (Hypothyrold)
30
4.6
0.4,
2. K . H . (Hyperthyrold)
27 31
17.8
3.8 5.7
3. B. J. (Euthyrold mother of a child with cretinism)
26
12.3
1.6
1.22
Controls I. C . S . 2. E . W . 3. M . S . 4. C . S . 5. B . D . 6. E . B . 7. B . K . 8. S . H . 9. R . R . 10. J . B . 11. J . D . 12. A . H . 13. L . C . 14. S . S . 15. P . S . 16. E . R . 17. B . H . 18. B . A . 19. W . M . 20. H. LeC. Mean Range
16 19 20 20 24 27 27 28 30 32 32 32 33 34 35 36 36 38 40 40
greatly elevated amniotic fluid PBI on two occasions (3.8 a n d 5.7 /~g p e r 100 m l . ) . Alt h o u g h this infant a p p e a r e d euthyroid at birth, p r o b a b l y reflecting m a t e r n a l m e d i c a tions, he developed neonatal Graves' disease by 10 days of age. T h e third p a t i e n t h a d a n o r m a l serum P B I a n d a n o r m a l amniotic fluid P B I a n d T , by colunm. H e r infant was normal. DISCUSSION O u r findings in euthyroid patients supp o r t K a u f m a n ' s observation t h a t significant amounts of PBI are measurable in amniotic fluid. A l t h o u g h Fisher a n d associates 1~ have found low fetal serum T4 a n d free T4 between 11 a n d 18 weeks' gestation a n d a p r o gressive increase in values between 22 weeks
9.2 8.6 8.8 9.8 12.1 8.8 7.8 7.4 9.6 8.5 8.2 11.2 9.0 7.4 - 12.1
1.0 1.8 2.2 2.0 .7 1.7 1.6 1.2 1.0 1.2 1.2 1.5 1.3 1.4 1.8 1.0 1.4 1.8 1.6 1.3 1.33 0.7 - 2.2
2.0 1.4 1.2 1.7 1.2 1.8 1.8 1.1 0.7 1.0 1.1 0.7
1.26 0.7 - 2.0
a n d term, we have been unable to demonstrate increased amniotic fluid PBI with gestational age. Fisher a n d associates' study revealed low fetal serum thyrold-stimulating h o r m o n e values to 18 weeks a n d an a b r u p t increase of thyroid-stimulating h o r m o n e from 18 to 22 weeks. G r e e n b e r g a n d associates 1G have shown an increase in fetal free thyroxine from the tenth to twentieth week of gestation, with fetal levels reaching within one s t a n d a r d deviation of the term level by 20 weeks. All b u t two of our amniotic fluid levels were obtained after 20 weeks' gestation, w h e n fetal thyroid-stimulating h o r m o n e a n d feedback mechanisms were already functioning; this m a y explain the failure to note higher values with lengthening gestation. I n 3 patients serial determinations were done in the second a n d t h i r d
Volume 79 Number 6
Thyroxine derivatives in amniotic fluid
3.0 2.5 2.0
O
o_
05
9
1.5
1.0 .5
0
---~/
i
t
i
i
I
i
15
20
25
513
35
40
GESTATION (wks.)
Fig. 1. PBI (#g/100 ml. of amniotic fluid) plotted against gestational weeks for 29 samples in 20 normal patients.
trimesters and no rise was noted with fetal age. Previous studies have shown that by the third month of gestation the fetal thyroid is able to concentrate radioactive iodine17; thyroxine synthesis is known to occur by about 3 ~ months? 8 The studies of Fisher and associates 15 and Greenberg and associates 16 on abortion material have shown autonomous function of the fetal hypothalamic-pituitary control system to be evident by t2 weeks' gestation. French and Van Wyck,19 in discussing fetal hypothyroidism, have reviewed the pituitary regulation of fetal thyroid function. Maternal thyroidstimulating hormone does not cross the placenta, 2~ and in experimental animals goiters develop in the absence of a maternal pituitary, but not in the absence of a fetal pituitary. 22-23 Thus if the fetal pituitary-thyroid axis is normal, he can function adequately even though there is maternal hypothyroidism. Such cases have been reported, 2~-25 but prenatal data on infants at risk are lacking. Antenatal hypothyroidism causes irreversible impairment of brain development, and neonatal hyperthyroidism may be life threatening. It would be extremely helpful if some indication of fetal thyroid status might be obtained during pregnancy.
92 7
The first mother of our series was hypothyroid after having been given I T M at the ninth week of gestation, followed by propylthiouracil; we were concerned that possible fetal thyroid cellular damage might prevent a normal glandular response to fetal thyroidstimulating hormone. Because of the low amniotic fluid PBI the mother was given /-triiodothyronine for two months until delivery in the hope that enough placental transfer would occur to prevent intrauterine hypothyroidism. A full-term male infant was born without stigmata of cretinism, but with no ossification of the proximal tibial epiphyses and only faint ossification of the distal femoral epiphyses. Replacement therapy with desiccated thyroid was given for the first 9 months. The child was re-evaluated after having received no medication for 11 weeks and found to have normal thyroid function. The transient hypothyroidism may have been related to either propylthiouracil or I ~sl therapy. In the second case we were concerned about whether the baby was hyperthyroid in utero. A high amniotic fluid PBI was found which later turned out to reflect the baby's thyroid status. Long-acting thyroid stimulator was not demonstrable in the amniotic fluid, but was present in maternal serum, cord blood, and the baby's serum at birth. The child was born 6 weeks prematurely and had Graves' disease. Not all hyperthyroid mothers have hyperthyroid babies. Those who do often have demonstrable serum long-acting thyroid stimulator, a 7 S globulin, which may traverse the placenta and stimulate the fetus. Contrary to reports in the literature stating the half life of long-acting thyroid stimulator to be approximately 6 days, 26 excessive amounts were still measurable in our patient at one month of age and the baby has had prolonged rather than transient hyperthyroidism. He continues to have growth failure, prominent eyes, and a small goiter at 16 months, although he now has a normal PBI level and no demonstrable longacting thyroid stimulator. In the third case the patient was euthy-
928
Hollingsworth and Austin
roid b u t h a d previously given b i r t h to a child with cretinism. I n this instance the amniotic fluid PBI was normal. This was reassuring b u t our experience with amniotic fluid PBI determinations was scant a n d the m o t h e r was given / - t r i i o d o t h y r o n i n e Y A n a p p a r e n t l y n o r m a l male infant was born at term. T h i s child remains n o r m a l at one y e a r of age. Decreased or absent fetal thyroid horm o n e formation m a y occur with iodine deprivation, with a n enzymatic defect in thyroxine synthesis, or with failure of m i g r a t i o n or absence of the thyroid gland. I n addition, i n a d v e r t e n t exposure to I ~sl at a critical time, excessive iodine or blocking agents m a y likewise depress fetal thyroid h o r m o n e synthesis. N o w t h a t new knowledge of pret e r m n o r m a l thyroid function has become available from abortion studies, an a t t e m p t should be m a d e to d o c u m e n t a b n o r m a l fetal thyroid function prenatally. Retrospectively, amnlotic fluid PBI determh~ations reflected the fetal thyroid status of each of our 3 babies at risk. T h e use of amniotic fluid PBI should be explored further a n d correlated with long-term observations on infants of mothers with thyroid disease a n d those who have previously given birth to children with cretinism. The authors wish to thank Dr. Philip K~ Bondy, Dr. C. Charlton Mabry, Dr. Jacquellne Noonan, and Dr. Warren Wheeler for their helpful comments and review of the manuscript. We are grateful to Dr. J. M. McKenzie of Montreal, Canada, for doing the long-acting thyroid stimulator assays.
REFERENCES I. Fuchs, F.: Amniotie fluid, Clin. Obstet, Gynecol. 9: 425, 1966. 2. Mandelbaum, B., and Evans, T. N." Life in the amniotie fluid, Am. J. Obstet. Gynecol, 104: 365, 1969. 3. Freda, V. J.: Rh problem in obstetrics and a new concept of its management using amniocentesis and spectrophotometric scanning of amniotic fluid, Am. J. Obstet. Gynecol. 92; 341, 1965. 4. Queenan, J. T., and Adams, D. W.: Amniocentesis for prenatal diagnosis of erythroblastosis fetalis, Obstet. Gynecol. 25: 302, 1965.
The Journal o/ Pediatrics December 1971
5. Queenan, J. T., and Wyatt, R. H.: Intrauterine transfusion of fetus for severe erythroblastosis fetalis, Am. J. Obstet. Gynecol. 92: 796, 1967. 6. Jacobson, C. B., and Barter, R. H.: Intrauterine diagnosis and management of genetic defects, Am. J. Obstet. Gynecol. 99: 796, 1967. 7. Jeffcoate, T. N. A., Fliegner, J. R. H., Russell, S. H., Davis, J. C., and Wade, A. P.: Diagnosis of adrenogenital syndrome before birth, Lancet 2: 553, 1965. 8. Frantantoni, J. C., Neufeld, E. F., Uhlendorf, W., and ffacobson, C. B.: Intrauterine diagnosis of Hurler and Hunter syndromes, N. Engl. J. Med. 280: 686, 1969. 9. tl.ose, H., Russell, K. P., and Starr, P.: Serum protein-bound iodine of mothers and infants at delivery in premature and term pregnancies, Am. J. Obstet. Gynecol. 86: 767, 1963. 10. Kaufman, S.: Protein-bound iodine (PBI) in human amniotic fluid, J. PEDIATR. 68: 990, 1966. 11. Austin, E , and Koepke, J. A.: An automated procedure for total organic iodine, Am. J. Clin~ Pathol. 45: 344, 1966. 12. Kessler, G., and Pileggi, V. J.: A Semiautomated nonincineration technique for determining serum thyroxine, Clin. Chem. 16: 382, 1970. 13. Odell, W. D., Wilbur, J. F., and Paul, W. E.: Radioimmunoassay of thyrotrophin in human serum, J. Clin. Endocrinol Metab. 25: 1179, 1965. 14. McKenzie, J. M., and Williamson, A.: Experience with the blo-assay of the long acting thyroid stimulator, J. Clin. Endocrinol. Metab. 26" 518, 1966. 15. Fisher D. A., Hobel, C. J., Garza, R., and Pierce, C. A.: Thyroid function in the preterm fetus, Pediatrics 46: 208, 1970. 16. Greenberg, A. H., Czernichow, P., Reba, R. C., Tyson, J., and Blizzard, R. M.: Observations on the maturation of thyroid function in early fetal life, J. Clin. Invest. 49: 1790, 1970. 17. Hodges, R. 2., Evans, T. C., Bradbury, J. T., and Keettel, W. C.: The accumulation of radioactive iodine by human fetal thyroids, J. Clin. Endocrinol. Metab. 15: 661, 1955. 18. Shepherd, T. H.: Onset of function in the human fetal thyroid: Biochemical and radioautographic studies from organ culture, J. CIin. Endocrinol. Metab. 27: 954, 1967. 19. French, F. S., and Van Wyk, J. J.: Fetal hypothyroidism, J. P~DIATR. 64: 589, 1964. 20. Postel, S.: Placental transfer of perchIorate and triiodothyronine in the guinea pig, Endocrinology 60: 50, 1957. 21. Peterson, R. R., and Young, W. G.: The problem of placental permeability for thyrotropin, propylthiouracil and thyroxine in the guinea pig, Endocrinology 50: 218, 1952. 22. Nikitovitch, M., and Knobil, E.: Placental transfer of TSH in the rat, J. Clin. Endocrinol. Metab. 15: 837, 1955.
Volume 79 Number 6
23. Sobel, E. H., Hamburg, M., and Koblln, R.: Development of the fetal thyroid in rats; Evidence for a placental transfer of thyroxine, Am. J. Dis. Child. 100: 709, 1960. 24. Lister, L. M., and Ashe, J. R., Jr.: Pregnancy and myxedema: Report of a case, Obstet. Gynecol. 6: 436, 1955. 25. Parkin, G., and Green, J. A.: Pregnancy occurring in cretinism and in juvenile and adult myxedema, J. Clin. Endoerinol. Metab. 3: 466, 1943.
Thyroxine derivatives in amniotic fluid
9 29
26. Sunshine, P., Kusumoto, H., and Krlss, J. P.: Survival time of LATS in neonatal thyrotoxicosis: Implications for diagnosis and therapy of the disorder, Pediatrics 38: 869, 1965. 27. Dussault, J., Row, V. V., Lickrish, G., and Vocpe, R.: Studies of serum triiodothyronine concentration in maternal and cord blood: Transfer of trliodothyronine across the human placenta, J. Clin. Endoerinol. Metab. 29: 595, 1969.