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Hormonal Effects in Infants Conceived By Assisted Reproductive Technology

Posted on: Sunday, 10 July 2005, 03:00 CDT

ABSTRACT. Objective. The purpose of this report is to describe 7 infants conceived by assisted reproductive technology (ART) who presented with breast development and/or pubic hair. The clinical presentation in these infants raises awareness that an altered intrauterine hormonal milieu may impact the fetal and infant stages of children conceived by ART.

Methods. Between May 2001 and April 2004, 7 children between the ages of 5 and 21 months conceived by ART were referred by their pediatricians to the Division of Pediatric Endocrinology at the New York University School of Medicine for evaluation of possible precocious puberty. Patients were evaluated for the possibility of centrally mediated precocious puberty and pseudoprecocious puberty, with a possible ovarian or adrenal origin.

Results. Endocrine evaluation in all patients indicated sex- steroid and hormone levels in the prepubertal range; pelvic sonography confirmed prepubertal ovaries with unstimulated uteri. Clinical follow-up of our patients thus far has not revealed progression of breast development, pubarche, or elevation in sex steroids.

Conclusions. It is well established that the developing endocrine system in the fetus and maturation of endocrine-control systems are influenced by hormone concentrations in the fetus. Whether ART alters the intrauterine hormonal milieu for the growing fetus conceived by ART is as yet unknown and is an area of ongoing investigation. Patients conceived through ART, including our patients who presented with hormonal manifestations, will need to be monitored throughout childhood and into adolescence and adulthood to determine if any perturbation exists on the timing of puberty and later fertility. Pediatrics 2005;116:190-194; prenatal care, prenatal exposure, puberty, early-onset puberty, in vitro fertilization.

ABBREVIATIONS. ART, assisted reproductive technology; IVF, in vitro fertilization; FSH, follicle-stimulating hormone; LH, Iuteinizing hormone; DHEA, dehydroepiandrosterone; DHEA-S, dehydroepiandrosterone sulfate; r-FSH, recombinant follicle- stimulating hormone; r-hCG, recombinant human chorionic gonadotrophin; H-P-G, hypothalamic-pituitary-gonadal.

With the development of new techniques in assisted reproductive technology (ART), progress has been made in the ability of infertile couples to conceive a child.

Data from the American Society of Reproductive Medicine's national in vitro fertilization (IVF) registry indicate that in the year 1999, a total of 88 077 cycles of assisted reproductive treatment led to 21 904 deliveries, with the birth of 30 967 neonates.1 This represents a success rate (deliveries per transfer) of 30.5%, an increase from 17% in 1992. This increase is related to the higher rate of successful implantations as well as an increase in the number of multiple gestations. At present, ART is responsible for 1% to 2% of births in the United States.1

Numerous studies have explored the type and incidence of ART- related side effects in offspring. Included among the most common adverse effects are low birth weight (≤2500 g) among term (risk ratio: 2.6; 95% confidence interval: 2.4-2.8) and preterm (risk ratio: 1.3; 95% confidence interval: 1.2-1.4) singleton infants conceived by ART.2 This increased risk persists after adjustment for maternal age and parity, gestational age at delivery, multifetal reduction procedures, and cause of infertility. Although several studies do not report an increased risk of congenital malformations in children conceived through ART,3,4 others have shown that the prevalence of ≥1 major birth defects by the age of 1 year is twice as high in infants conceived by ART.5 In a Beckwith-Wiedemann registry of 65 children with this syndrome, 3 infants (5%) had been conceived by IVF, indicating a higher-than- expected rate (0.8%).6

Initial studies did not demonstrate an increase in cancer risk among children conceived by ART.7'8 More recently, several reports (including a study from the Netherlands9) conclude that the relative risk of retinoblastoma is significantly higher.

There are a range of possible factors associated with ART treatment that may contribute to potential adverse outcomes. These factors include the relatively advanced age of infertile couples seeking ART, the underlying causes of their infertility, the medications used to induce ovulation or to maintain the pregnancy in its early stages, and the ART procedures themselves.

The purpose of this report is to describe 7 infants born by ART who presented with breast development and/or pubic hair. The data available to the authors for this report were obtained through direct patient care and retrospective chart review.

METHODS

Analyses

All patients underwent baseline hormone analysis. Patients who demonstrated both breast development and pubic hair were evaluated for the possibility of sexual precocity. In these patients, tests to determine the levels of follicle-stimulating hormone (FSH), luteinizing hormone (LH), estradiol, and dehydroepiandrosterone sulfate (DHEA-S) and ovarian sonography were performed (patients 1- 4). Patients presenting primarily with pubic hair (patients 5-7) were evaluated as well for causes of hyperandrogenism (eg, congenital adrenal hyperplasia). DHEA-S, serum testosterone, and 17- hydroxyprogesterone were performed in these patients (Table 1). In addition, a left-hand- and wrist-for-skeletal-age test was performed for each patient.

Laboratory analysis was performed at Quest Diagnostics (Teterboro, NJ) for patients 1, 3, and 5. Cortisol, estradiol, DHEA- S, and testosterone were measured by chemiluminescence; tests for 17- hydroxyprogesterone, A4-androstenedione, and estrone levels were performed by radioimmunoassay. LH and FSH measures were performed by microenzyme immunoassay. For patients 2, 4, 6, and 7, laboratory analysis of androgen levels was performed in-house. Radioimmunoassays for testosterone, 17-hydroxyprogesterone, and DHEA- S levels were performed in our laboratory using a well-described methodology.10 Radioimmunoassays for testosterone and 17- hydroxyprogesterone were performed after purification by Celite chromatography. The intraand interassay coefficients were 6% and 11%, respectively, for testosterone and 11% and 15%, respectively, for 17-hydroxyprogesterone. DHEA-S, however, was immunoassayed without purification.

Adrenocorticotrophic hormone-stimulation testing of patient 7 was performed by using cosyntropin 0.25 mg (Amphastar Pharmaceuticals, Inc, Rancho Cucamonga, CA) administered intravenously at O minutes. Serum sampling was performed at 60 minutes for serum testosterone and 17-hydroxyprogesterone.

Assessment of skeletal age was performed by well-established methods.11 Pelvic and adrenal sonography were performed at the Division of Pediatric Radiology at the New York University School of Medicine for all patients except patient 1, who underwent the procedure at Lenox Hill Hospital (New York, NY).

TABLE 1. Laboratory Data

With the exception of 2 patients (patients 2 and 4), who have been evaluated recently, all patients have been seen in follow-up consultation for a period of 3 months to as long as 20 months.

Patient Population

Between May 2001 and April 2004, 7 children between the ages of 5 and 21 months who were conceived by ART were referred by their pediatricians to the Division of Pediatric Endocrinology at the New York University School of Medicine for evaluation of breast development and/or pubic hair. Six patients were female, and 1 was male. The characteristics of the sample, including the clinical findings, are presented in Table 2.

All of the mothers had been treated with recombinant FSH (r-FSH) followed by recombinant human chorionic gonadotropin (r-hCG) and progesterone by established ART protocols.12^15WUh the exception of families 4 and 5, verbal reporting by the families indicated that maternal anovulation or an inability to conceive spontaneously prompted medical therapy with ART. The family of patient 5 underwent ART and treatment of paternal oligospermia. In addition to standard maternal hormonal therapy (for IVF), the father of patient 5 was treated with Teslac, clomiphene, and indomethacin. Patient 4 was conceived by IVF using a sperm donor. Family history for all patients was negative for congenital adrenal hyperplasia, polycystic ovary syndrome, or precocious puberty. All of the mothers denied using medication for systemic or endocrine disorders during the pregnancy.

Patients 1 and 2 were born at 7 months of gestation and had birth weights of 3.5 and 1.81 kg, respectively. Patients 3, 4, 6, and 7 were born at term and had birth weights that were appropriate for gestational age. The genetic backgrounds of the families were diverse, with no predominance within the patient population. Patient 5 was the 8-month product of a set of fraternal twins that was originally a quadruple pregnancy. According to the mother, the male twin was growing normally, with no evidence of pubarche.

On physical examination, all subjects were healthy appearing infants. Patients 1 through 4 presented with bilateral, well- developed breast tissue and pubic hair (Tanner stages II-III). Patients 5 and 6, both female, presented with Tanner III pubic hair and minimal breast tissue. There was noclitoral enlargement or posterior labial fusion in the female patients. Patient 7, a male infant, presented with terminal pubic hair along the scrotum but no gonadal or genital enlargement.

TABLE 2. Clinical Data

Review of the linear growth trajectories indicated that all term infants were growing consistently along their centile channels. Patients 1, 2, and 5, who had been born prematurely, were demonstrating catch-up growth consistent with growth patterns of prematurity.

RESULTS

Results are presented in Table 1. Levels of LH, FSH, estradiol, DHEA-S, testosterone, A4-androstenedione, 17-hydroxyprogesterone, and estrone were in the prepubertal range in all patients. Pelvic sonography indicated the appearance of prepubertal ovaries (ovarian volumes measuring <1.2 cm^sup 3^) and a nonstimulated uterus in all females. The skeletal ages were within 2 SD for chronologic age in all patients (Table 1). During the follow-up visits, there was no evidence of pubertal progression or linear growth acceleration in any of the patients.

DISCUSSION

We present data on 7 patients conceived by ART, ages 5 to 21 months, referred for endocrine evaluation of possible precocious puberty and who presented with breast development and/or pubic hair. Patients who demonstrated both breast development and pubic hair were evaluated for the possibility of sexual precocity. Patients presenting primarily with pubic hair were evaluated as well for causes of hyperandrogenism (eg, congenital adrenal hyperplasia). Endocrine evaluation in all patients indicated hormonal levels in the prepubertal range; pelvic sonography confirmed prepubertal ovaries with unstimulated uteri. Clinical follow-up of our patients did not reveal progression of the breast development or an increase in pubarche. Linear growth velocity continued along prior centile channels.

Our patients were unusual in their clinical presentation. In those infants conceived by ART who were referred for evaluation of breast development, the size and maturation of breast glandular tissue exceeded that generally seen in our patient population with isolated thelarche, a benign but not well-explained condition seen in normal infants that may result from a delay in the transition from the active fetal hypothalamic-pituitary-gonadal (H-P-G) axis to the quiescent prepubertal H-P-G axis. The degree of breast tissue seen in our ART patients, either isolated or in association with pubic hair, raised the possibility of precocious puberty, either centrally mediated or of ovarian or adrenal origin.

By clinical history, it was apparent that the 1 common link among this group of patients was that they had all been conceived by ART. All mothers had received r-FSH, r-hCG, and progesterone as defined by standard ART protocols. 12~lft Review of the literature regarding the pharmacokinetics and pharmacodynamics of r-FSH and r-hCG suggests that theses compounds alone cannot be the cause of the clinical findings. Additionally, the timing of administration of both medications for ovulation induction renders it unlikely that there may be a direct effect of theses drugs on the fetal H-P-G axis.16

We reviewed the literature on the possible relation between ART procedures and the appearance of estrogen and/or androgen effect on offspring. Recent data indicate that maternal serum and amniotic fluid levels of βhCG are elevated in pregnancies conceived after ART.17-18 In addition, ART twin pregnancies have higher levels of βhCG than are seen in spontaneous twin pregnancies. The increase in βhCG is seen even in pregnancies after spontaneous frozenembryo transfer. Thus, the increase in βhCG seems not to be related to superovulation hormonal therapy.17,18 Although we cannot exclude other factors, we suspect that the elevation of maternal βhCG seen in ART may play an etiologic role. It is known that placental βhCG induces maternal and fetal adrenal steroidogenesis of DHEA.18-19 DHEA-S is then converted to Δ4- androstenedione in the placenta. DHEA-S and Δ4-androstenedione are the major precursors for placental estrogen production.18,19 Elevated levels of DHEA and Δ4-androstenedione are metabolized to estrogens and androgens that may impact the developing fetus.20- 22 We speculate that elevated levels of estrogen and androgens in utero may directly mediate the development of breast tissue and pubic hair and may further alter the maturation of the H-P-G axis in the developing fetus.

Although the etiology of the elevation in βhCG in ART pregnancies is uncertain, it is known that maternal βhCG levels are higher in twin pregnancies than in singleton pregnancies. Therefore, it is conceivable that, in our patient population, multiple embryo implantations early in gestation could have been a factor in increasing maternal βhCG levels and altering the maternal-fetal hormonal milieu. We should note, however, that 1 study did not find any elevation consistent with the number of implantation and posterior multifetal reduction.23 Additionally, the impact of artificial fertilization or the embryo culture and the effect of progesterone on the fetus are also unknown.

The impact on the developing fetus of an altered hormonal milieu is unclear. It is known that fetal life is an early and important stage in the development of the H-P-G axis that culminates in adult life with the achievement of full sexual maturation and fertility. In both humans and primates, the fetal hypothalamic gonadotrophin- releasing hormone pulse generator is oprant in the fetus by the end of the first trimester. Studies suggest that hormonal imprinting or programming occurs in fetal life during a critical period of maturation24 and may modulate gene expression and nuclear and/or plasma membrane receptors.25 Evidence exists to indicate that prenatal androgens program the timing of neuroendocrine puberty in sheep; the higher the dose of prenatal testosterone, the earlier the initiation of the pubertal LH rise.26 It has been shown that estrogen administration to pregnant rats during the last third of gestation produces cryptorchid male offspring and may suppress spermatogenesis permanently in adult males. Additionally, perinatal estrogen administration to the developing female rodent produced long-term effects including persistent vaginal cornification, hyperplastic vaginal lesions, and cervicovaginal cancer; synthetic nonsteroidal estrogens (diethylstilbestrol) had similar effects.25,27

Whatever the mechanisms, the developing endocrine system in the fetus and maturation of endocrine-control systems are influenced by the hormone concentrations in the fetus. Patients conceived through ART, including our patients who presented with hormonal manifestations, will need to be monitored throughout childhood and into adolescence and adulthood to determine what impact, if any, exists on the timing of puberty and, later, fertility.

We recognize the limitations of our study. Our data represent a compilation of our experience in ART-conceived infants referred for evaluation of clinical signs of precocious puberty. Although we postulate that elevation in maternal hCG may play a critical role in the development of a hormonal effect in these infants, serial maternal hCG levels are not available for our study. Additionally, the number of ART-conceived infants presenting with clinical signs of puberty is extremely small when compared to the hundreds of infants conceived by ART within our referral area. Well-controlled, prospective studies to include fetal ovarian sonography, hormonal levels in amniotic fluid (when available), and careful ongoing evaluation of the ART-conceived infant will provide additional insight into the mechanism of this entity in infants conceived by ART.

ACKNOWLEDGMENT

We acknowledge the contributions of Dr V. K. Prasad, associate director of our pediatric endocrine laboratory, for the in-house steroid-hormone assays.

REFERENCES

1. American Society for Reproductive Medicine; Society for Assisted Reproductive Technology Registry. Assisted reproductive technology in the United States: 1999 results generated from the American Society for Reproductive Medicine/Society for Assisted Reproductive Technology Registry. Fertil Steril. 2002;78:918-931

2. Schieve LA, Meikle SF, Ferre C, Peterson HB, Jeng G, Wilcox LS. Low and very low birth weight in infants conceived with the use of assisted reproductive technology. Nf Engl } Mai. 2000;346:731- 737

3. Pregnancies and births resulting from in vitro fertilization: French national registry, analysis of data 1986 to 1990. FIVNAT (French In Vitro National). Fertil Steril. 1995;64:746-756

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7. Klip H, Burger CW, de Kraker J, van Leeuwen FE; OMEGA-project group. Risk of cancer in the offspring of women who underwent ovarian stimulation for IVF. Hum Reprod. 2001;16:2451-2458

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9. Moll AC, Imhof SM, Cruysberg JR, Schouten-van Meeteren AY, Boers M, van Leeuwen FE. Incidence of retinoblastoma in children born after in-vitro fertilisation, lancet. 2003;361:309-310

10. Manlimos F, Abraham GE. Chromatographie purification of tritiated steroids prior to use in radioimmunoassay. Anal Lett. 1975;8:403-41\1

11. Greulich WW, Pyle SI. Radiographie Atlas of Skeletal Development of the Hand and Wrist. Stanford, CA: Stanford University Press; 1959

12. Rowell P, Braude P. Assisted conception. I-General principles. BMJ. 2003;327:799-801

13. Rowell P, Brande P. Assisted conception. II-in vitro fertilisation and intracytoplasmic sperm injection. BM]. 2003;327:852-855

14. Daya S, Gunby J, Hughes EG, Collins JA, Sagle MA. Follicle- stimulating hormone versus human menopausal gonadotropin for in vitro fertilization cycles: a meta-analysis. Fertil Steril. 1995;64:347-354

15. Filicori M, Cognigni GE, Pocognoli P, Ciampaglia W. Choice of ovarian stimulation regimens in assisted reproduction: finding the thread in the gonadotropin maze. Fertil Steril. 2003;80:1114-1116

16. Ie Cotonnec JY, Loumaye E, Porchet HC, Beltrami V, Munafo A. Pharmacokinetic and pharmacodynamic interactions between recombinant human luteinizing hormone and recombinant human folliclestimulating hormone. Fertil Steril. 1998;69:201-209

17. Perheentupa A, Ruokonen A, Tuomivaara L, Ryynancn M, Martikainen H. Maternal serum beta-HCG and alpha-fetoprotein concentrations in singleton pregnancies following assisted reproduction. Hum Reprod. 2002;17:794-797

18. Hui PW, Lam YH, Tang MH, Ng EH, Yeung WS, Ho PC. Amniotic fluid human chorionic gonadotrophin and alpha-fetoprotein levels in pregnancies conceived after assisted reproduction. Prencit Diagn. 2003;23: 484-487

19. Braunstein G. Endocrine changes in pregnancy. In: Larsen PR, Kronenberg HM, Melmed S, Polonsky K, eds. Wilinnis Textbook of Endocrinology. 10th ed. Philadelphia, PA: WB Saunders; 2003:800-803

20. Miller WL. Steroid hormone biosynthesis and actions in the maternofeto-placental unit. Clin Perinatal. 1998;25:799-817

21. Vaskivuo TE, Aittomki K, Anttonen M, et al. Effects of folliclestimulating hormone (FSH) and human chorionic gonadotropin in individuals with an inactivating mutation of the FSH receptor. Fcrtil Steril. 2002;78:108-113

22. Barnes RB, Rosenfield RL, Namnoum A, Layman LC. Effect of folliclestimulating hormone on ovarian androgen production in a woman with isolated follicle-stimulating hormone deficiency. N Engl ] Med. 2000;343: 1197-1198

23. Rotmensch S, Celentano C, Shalev J, et al. Midtrimester maternal serum screening after multifetal pregnancy reduction in pregnancies conceived by in vitro fertilization. J Assist Reprod Genet. 1999;16:8-12

24. Grumbach MM. The neuroendocrinology of human puberty revisited. Harm Res. 2002;57:2-T4

25. Csaba G. Receptor ontogeny and hormonal imprinting. Experientia. 1986;42:750-759

26. Kosut SS, Wood RI, Herbosa-Encarnacion C, Foster DL. Prenatal androgens time neuroendocrine puberty in the sheep: effect of testosterone dose. Endocrinology. 1997;138:1072-1077

27. Bern HA, Talamentes FJ Jr. Neonatal mouse models and their relation to disease in the human female. In: Herbst AL, Bern HA, eds. Developmental Effects of Dietliylstilbestrol (DES) in Pregnancy. New York, NY: Thieme-Stratton; 1981:129-147

Patricia Martin Rojas-Marcos, MD*; Raphael David, MD[double dagger]; and Brenda Kohn, MD[double dagger]

From the *Department of Endocrinology and Nutrition, Hospital Clinico San Carlos, C/Martin Lagos s/n Madrid, Spain; and [double dagger]Department of Pediatrics, Division of Pediatric Endocrinology, New York University School of Medicine, New York, New York.

Accepted for publication Feb 28, 2005.

doi:10.1542/peds.2004-2553

No conflict of interest declared.

Address correspondence to Brenda Kohn, MD, Department of Pediatrics, New York University School of Medicine, 550 First Ave, New York, NY 10016. E-mail: brenda.kohn@med.nyu.edu

PEDIATRICS (ISSN 0031 4005). Copyright 2005 by the American Academy of Pediatrics.

Copyright American Academy of Pediatrics Jul 2005

Source: Pediatrics

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