Human Reproduction, Vol. 17, No. 1, 88-91,
January 2002
© 2002 European Society of Human Reproduction and Embryology
The role of LH and FSH in ovarian androgen secretion and ovarian follicular development: Clinical studies in a patient with isolated FSH deficiency and multicystic ovaries: Case report
1 Departments of Obstetrics and Gynecology, University of Chicago, Chicago, IL 60637, 2 Emory University, Atlanta, GA 30322, 3 Department of Pediatrics and Medicine, University of Chicago, Chicago, IL 60637, 4 Department of Obstetrics and Gynecology, Medical College of Georgia, Augusta, GA 30912, USA
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Abstract |
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Inactivating mutations have proven to be instructive in elucidating the role of FSH in human ovarian function. We performed a detailed reproductive endocrine evaluation of a patient with inactivating mutations in the FSH ß-subunit gene who was hypo-estrogenic and had LH excess. The patient underwent a pelvic ultrasound and overnight frequent blood sampling followed by a human chorionic gonadotrophin (HCG) stimulation test. One month later she received human recombinant FSH, followed 24 h later by a second HCG stimulation test. Despite a mean LH serum concentration and LH pulse characteristics typical for polycystic ovaries (PCOS), baseline and dexamethasone-suppressed free testosterone were low–normal. The administration of HCG led to minimal stimulation of 17-hydroxyprogesterone and androgens. The patient had multicystic ovaries containing follicles 3–5 mm in diameter and responded to FSH with prompt increases in estradiol and inhibin B. There were no clinical or laboratory consequences of LH excess in this FSH-deficient woman. These findings support the hypothesis that excessive LH stimulation alone does not cause ovarian hyperandrogenism. We also found that follicular development was present in the absence of FSH. These antral follicles had apparently developed normally, since estradiol and inhibin B increased promptly after FSH administration.
Key words: androgens/FSH deficiency/LH excess/ovary
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Introduction |
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TopAbstractIntroductionCase reportResultsDiscussionAcknowledgementsReferences |
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Inactivating mutations have proven to be instructive in elucidating the role of FSH in human ovarian function (Layman and McDonough, 2000
; Themmen and Huhtaniemi, 2000). Our studies of a patient with compound heterozygous mutations of the FSH ß-subunit gene (Layman et al., 1997), undetectable serum FSH and elevated serum LH have elucidated two aspects of FSH action which challenge common wisdom. We report here evidence that FSH is not necessary for the development of small, healthy antral follicles readily responsive to FSH, which contrasts with studies in FSH-knockout mice (Kumar et al., 1997; Dierich et al., 1998). However, FSH is necessary for ovarian theca cell function, contrary to expectations from the 2-gonadotrophin, 2-cell model of ovarian steroidogenesis (Barnes et al., 2000). The latter finding has important implications for theories about the role of LH excess in the pathogenesis of polycystic ovary syndrome (PCOS). |
Case report |
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TopAbstractIntroductionCase reportResultsDiscussionAcknowledgementsReferences |
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Subject
The patient presented at 16 years of age with primary amenorrhoea and absent breast development. Her baseline and gonadotrophin-releasing hormone (GnRH)-stimulated FSH concentrations were undetectable by immunoassay, while LH concentrations at baseline (30 mIU/ml) and post-GnRH (150 mIU/ml) were elevated. The patient was subsequently treated with estrogen and underwent normal breast development. Analysis of the FSH ß-subunit gene demonstrated that she was a compound heterozygote for two different mutations, a two base-pair deletion in exon three at codon 61 (Val61X) inherited from her mother, and a missense mutation changing a cysteine to a glycine at codon 51 (Cys51Gly) inherited from her father. When these mutations were stably transfected into Chinese hamster ovary cells, they demonstrated no measurable FSH immuno- or bioactivity (Layman et al., 1997
).
Reproductive endocrine evaluation
The patient underwent a detailed reproductive endocrine evaluation at 21 years of age in the University of Chicago General Clinical Research Center (GCRC), after discontinuing the oral contraceptive pill for one month and giving informed consent. The study protocols were approved by the University of Chicago Institutional Review Board. Her height was 155 cm and her weight was 53.2 kg. There was no hirsutism (Ferriman and Gallway score of 4; normal <8), breasts and pubic hair were Tanner stage 5, and the pelvic examination was normal. An initial blood sample was drawn for immunoactive FSH, LH and free testosterone. The patient then received dexamethasone 0.5 mg four times daily for 4 days prior to and continuing throughout the endocrine evaluation to suppress adrenal function. Following admission to the GCRC, she underwent a transvaginal pelvic ultrasound followed by blood sampling every 10 min from 7.00 p.m. to 6.00 a.m. Serum immunoactive FSH and LH were determined for each sample; inhibin A and B, bioactive FSH and free 
-subunit were determined in a pooled sample. The following morning, a human chorionic gonadotrophin (HCG) test was performed. A blood sample was drawn for baseline testosterone, free testosterone and estradiol. The following steroid intermediates were also determined: 17-hydroxyprogesterone (17-Prog), androstenedione, 17-hydroxypregnenolone and dehydroepiandrosterone. HCG 5000 IU was then administered i.m. and blood was drawn 24 h later for repeat steroid measurements.
The patient remained off any sex steroids for another month, then returned to the GCRC for a second HCG study which was like the first except she received 300 IU of recombinant human FSH s.c. in the morning, 24 h prior to the HCG test. A blood sample was drawn 24 h later for steroids and inhibin A and B, and HCG 5000 IU was administered. She then returned 24 h after HCG (48 h after FSH) for a final blood sample for steroid measurements (Figure 1).
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All samples for steroids were frozen at –20°C and measured in the same assay using previously published methods (Barnes et al., 1989
; Rosenfield et al., 1994). Serum immunoactive LH and FSH were measured using a ß-subunit-specific assay (DelfiaTM, Wallach, Finland; lower limit of sensitivity 0.15 mIU/ml for LH and 0.2 mIU/ml for FSH). Bioactive FSH, inhibin A, inhibin B and free -subunit were measured using previously published methods (Christin-Maitre et al., 1996; Lavoie et al., 1998; Welt et al., 1999). LH pulse analysis was performed using the ULTRA program (Van Cauter and Copinschi, 1981). |
Results |
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The ovarian ultrasound prior to FSH treatment is shown in Figure 2
. The ovaries were multicystic containing follicles 3–5 mm in diameter, but unlike typical polycystic ovaries (Adams et al., 1986), there was no dense stroma and the follicles were scattered throughout the ovary rather than being predominately subcapsular.
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Immunoactive LH was 46 mIU/ml and FSH was undetectable (<0.2 mIU/ml) in the single sample drawn before dexamethasone. There was no immunoactive FSH detected in the samples collected overnight (Table I
), and no bioactive FSH was detected in the pooled sample. The mean immunoactive LH in the overnight samples was 46 mIU/ml; mean pulse amplitude was 15.8 mIU/ml and nine pulses were detected in 11 h (Figure 3). This LH profile is similar to that reported for PCOS (Waldstreicher et al., 1988). Free -subunit measured in the pooled 11 h sample was in the postmenopausal range (350–1740 pg/ml; Table I).
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Although LH concentrations were elevated, the patient had no laboratory evidence of ovarian hyperandrogenism. The patient's baseline free testosterone was 6 pg/ml (normal range 3–10 pg/ml; ovarian hyperandrogenism >10 pg/ml) (Rosenfield et al., 1994
). After 4 days of dexamethasone suppression the free testosterone fell to 2 pg/ml (normal range <8 pg/ml; ovarian hyperandrogenism 
8 pg/ml; Rosenfield et al., 1994). The 17-Prog concentration 24 h after HCG was 40 ng/dl which was 2.7 standard deviations (SD) below the mean in women with ovarian hyperandrogenism (288 ng/dl) (Levrant et al., 1997) and 1.5 SD below the mean of normal follicular phase women (139 ng/dl: Levrant et al., 1997).
One month later, during the second GCRC study, estradiol increased substantially 24 h after FSH administration (28–80 pg/mL). Inhibin B was near the lower limit of the normal range at baseline and rose to above the range for menstruating women 24 h after FSH injection (Table I
). In contrast, inhibin A was present at low concentrations.
The response of testosterone and the steroid intermediates to the two HCG tests have been previously reported (Barnes et al., 2000). In brief, all steroid concentrations 24 h after the second HCG injection (48 h after FSH) were greater than at 24 h after the first HCG injection (HCG alone). The difference between the two HCG tests was most dramatic for testosterone, which was unaffected by the first HCG test (12 ng/dl before and after HCG), but increased from 16 to 41 ng/dl after the second, FSH-primed HCG test.
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Discussion |
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TopAbstractIntroductionCase reportResultsDiscussionAcknowledgementsReferences |
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This patient, with well characterized inactivating mutations in the FSH ß-subunit gene, was hypo-estrogenic with secondary LH excess. Two features were noteworthy: antral follicular development was present in spite of the lack of FSH, and there were no clinical or laboratory consequences of the LH excess.
The point at which FSH is necessary for follicular development in the human ovary is debated (Gougeon, 1996; McGee and Hsueh, 2000). Despite having no detectable FSH, our patient had multicystic ovaries with follicles up to 5 mm in diameter. Similar sized follicles have been reported in some women with FSH receptor mutations (Aittomaki et al., 1996; Beau et al., 1998); however, those FSH receptor mutations may not be completely inactivating. These findings are in contrast to FSH ß- and FSH receptor-knockout mice, in which antral follicles are not maintained (Kumar et al., 1997; Dierich et al., 1998). In studies of human ovarian xenografts transplanted into the kidney capsule of immunodeficient and hypogonadotrophic mice, FSH was required for the growth of follicles beyond the two-layer granulosa cell stage (Oktay et al., 1998), which is about the point at which the FSH receptor gene is first expressed in human follicles (Oktay et al., 1997). Although the xenograft data are in contrast to our findings, it is likely that the endocrine and growth factor milieu of the in-situ human ovary is very different than that in the immunodeficient, hypogonadotrophic mouse.
The prompt estradiol and inhibin B responses 24 h after exogenous FSH suggest that our patient's antral follicles contained granulosa cells that had developed normally in the absence of FSH. Our findings are similar to those in two other patients with isolated FSH deficiency who ovulated and had a successful pregnancy after ~14 days of menotrophin therapy (Rabinowitz et al., 1979; Matthews et al., 1993). Ovulation after a short exposure to menotrophins implies that some healthy follicles had reached the point of recruitability without FSH exposure, since ~14 days are required for an ovulatory follicle to develop from follicles a few millimeters in diameter, in contrast to the 3 months required to develop from the pre-antral stage (Gougeon, 1996). These earlier reports and our findings suggest that in the complete absence of FSH stimulation, human ovarian antral follicles can develop up to 5 mm in diameter.
Our patient had no clinical or laboratory evidence of ovarian hyperandrogenism despite a mean LH concentration, LH pulse characteristics and ovarian follicular sizes typical for PCOS. On ultrasound her multicystic ovaries lacked the excess stroma of classic polycystic ovaries. Indeed, her ovaries produced little, if any, androgen. Her baseline and dexamethasone-suppressed free testosterone were low–normal. The administration of HCG led to minimal stimulation of 17-Prog or other thecal cell steroids. However, we have previously shown that exogenously administered FSH augmented her LH- and HCG-stimulated production of testosterone and all steroids of thecal cell origin (Barnes et al., 2000). This suggests that FSH action, probably via granulosa cell-produced paracrine intermediates, is necessary for thecal cells to respond to LH. One of many such paracrine factors is inhibin B, which increased markedly with FSH administration in this patient (Barnes 1998).
These findings are relevant to the role of LH excess in the pathogenesis of the ovarian hyperandrogenism of PCOS. It has been reported that 75% of women with clinical evidence of PCOS have an elevated LH concentration and 94% have an increased LH/FSH ratio (Taylor et al., 1997). These gonadotrophin secretory abnormalities have been thought to play an important role in the development of the ovarian hyperandrogenism characteristic of PCOS (Hall, 1993). In a related study, we found that a woman with a constitutively activating mutation of the LH receptor, identified because she was the mother of two sons with gonadotrophin-independent precocious puberty, had no clinical or laboratory evidence of ovarian hyperandrogenism (Rosenthal et al., 1996). Thus, increased LH stimulation, even in the presence of FSH, appears to be insufficient to induce the hyperandrogenism and stromal hyperplasia of PCOS. The findings in the previous, as well as the current, case report support the hypothesis that thecal cell androgen secretion in response to excessive LH stimulation is strictly limited by intra-ovarian factors. Ovarian hyperandrogenism is more likely a result of escape from down-regulation by these intra-ovarian factors than a result of elevated LH concentrations (Ehrmann et al., 1995). Taken together, our studies support the hypotheses that normal ovarian androgen production depends on both FSH and LH and that excessive ovarian androgen production is a result of abnormal intra-ovarian regulation, and not of excessive LH stimulation.
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Acknowledgements |
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We are greatly indebted to Patrick Sluss, Reproductive Endocrine Unit, The Massachusetts General Hospital, Boston, MA, who performed the FSH bioassay, FAS, inhibin A, and inhibin B assays; to Zubie Sheikh, Department of Obstetrics and Gynecology, University of Chicago, for performing the transvaginal ultrasound; and to Jennifer M.Cunningham, Department of Medicine, University of Chicago, for performing the LH pulse analysis. We also appreciate the helpful discussions of J.Larry Jameson from the Center for Endocrinology, Metabolism, and Molecular Medicine, Northwestern University, Chicago, IL and William F.Crowly Jr from the Reproductive Endocrine Unit and the National Center for Infertility Research, The Massachusetts General Hospital, Boston, MA.
L.C.L. was supported by NIH grant support PHS NICHD HD33004; R.L.R. was supported by NIH grant support RR-00055 (CRC).
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Notes |
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5 To whom correspondence should be addressed at: Dept of Obstetrics and Gynecology, University of Chicago, 5841 Maryland Avenue, Chicago, IL 60637, USA. E-mail: rbarnes{at}babies.bsd.uchicago.edu
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References |
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Submitted on February 28, 2001; accepted on August 21, 2001.
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