The effect of gonadotropin-releasing hormone agonists on growth hormone secretion in adult premenopausal women*

Transcription

1 FERTILITY AND STERILITY Copyright 1990 The American Fertility Society Printed on acid-free paper in U.S.A. The effect of gonadotropin-releasing hormone agonists on growth hormone secretion in adult premenopausal women* R. Ann Word, M.D.t M. Janelle Odom, M.D. William Byrd, Ph.D. Bruce R. Carr, M.D. Division of Reproductive Endocrinology and The Cecil H. and Ida Green Center for Reproductive Biology Sciences, Department of Obstetrics and Gynecology, University of Texas Southwestern Medical Center at Dallas, Dallas, Texas Suppression of the pituitary-gonadal axis by the administration of gonadotropin-releasing hormone agonists (GnRH-a) has been used for a variety of endocrinological and gynecological disorders. The suppressive effect of GnRH-a on luteinizing hormone, follicle-stimulating hormone, and sex steroid production is well documented. However, little is known regarding the effect of GnRH -a on other aspects of pituitary function. The purpose of the present study was to determine the effect of GnRH -a treatment on growth hormone-releasing hormone (GH-RH)-stimulated GH release in premenopausal women. Eight control women and seven women, who were receiving a GnRH -a, were recruited. Before and after a bolus infusion of human GH-RH, blood samples were obtained over 3 hours and analyzed for GH by immunoassay. Basal GH and insulin-like growth factor levels were not statistically different between the two groups. However, basal levels of estradiol and the integrated GH response after GH-RH were significantly lower in the GnRH-a treated women. The reduction in GH-RH-stimulated GH release in GnRH-a treated women may be attributed to diminished endogenous estrogen secretion, or to direct pituitary suppression by GnRH-a, or both. Fertil Steril54:73, 1990 Suppression of the pituitary-ovarian axis after the administration of gonadotropin-releasing hormone agonists (GnRH-a) results in a reversible state of hypoestrogenism, which has been used as a treatment modality for a variety of gynecological conditions such as uterine leiomyoma and endometriosis. 1,2 The suppressive effect of GnRH-a on follicle-stimulating hormone, luteinizing hormone Received January 9, 1990; revised and accepted March 29, * Supported in part by grants' 5-T32-HD07190 and HD from the National Institutes of Health, Bethesda, Maryland. t Reprint requests: R. Ann Word, M.D., Department of Obstetrics and Gynecology, University of Texas Southwestern Medical Center at Dallas, 5323 Harry Hines Boulevard, Dallas, Texas (LH), and sex steroid production is well documented. Little is known, however, about the effect of GnRH-a on other aspects of pituitary function. Suppression of ovarian function in girls with central precocious puberty with a GnRH -a results in decreased height velocity and a decrease in nocturnal serum growth hormone (GH) and plasma insulin-like growth factor I (IGF-I) levels. 3 This suggests that GnRH-a may inhibit GH release in association with decreased endogenous estrogen secretion. In postmenopausal women, estrogen replacement is associated with an increase in basal and stimulated G H secretion. 4 Other investigators also suggest that estrogen may directly affect G H secretion. 5-8 The purpose of the present investigation was to evaluate the effect of GnRH-a on (1) basal estradiol (E 2 ) levels, (2) basal IGF-I levels, Word et al. Effect of GnRH-a on GH secretion 73

2 and (3) the release of GH after infusion of human growth hormone-releasing hormone (GH-RH) in adult premenopausal women. Subjects MATERIALS AND METHODS Eight ovulatory women experiencing regular cyclical menstruation and seven women receiving 1 mg/d of leuprolide acetate subcutaneously for at least 4 consecutive weeks were recruited for the present study. All women were between the ages of 21 and 40, body weight 49 to 77 kg, and had no significant past medical or surgical history. All subjects agreed to participate in this study and signed an informed consent approved by the Institutional Review Board of the University of Texas Southwestern Medical School at Dallas. Methods Human GH -RH 1-44 was obtained from Bachem Inc. (Torrance, CA). The peptide was dissolved in sterile water and sterilized by filtration (0.22 JIM; Millipore, Bedford, MA); aliquots were placed in sterile vials, lyophilized, and stored at -70 C until utilized. Samples were found to be pyrogen-free by Leberco Testing Inc. (Roselle Park, NJ). Immediately before injection of the peptides, each vial was reconstituted with pyrogen-free water plus 0.9% benzyl alcohol. Samples were obtained at 7:00 A.M. and 9:00 A.M. from women in the fasting state after 1 to 6 months of GnRH -a treatment or from control women during days 1 to 10 of the menstrual cycle. The subjects were in the supine position and remained awake throughout the study. During the study, blood pressure and pulse were recorded. An 18-gauge intravenous (IV) catheter was placed in the forearm via a heparin lock to maintain patency. One-half hour after the catheter placement, blood samples were obtained at -15, 0, 15, 30, 45, 60, 90, 120, and 180 minutes relative to the bolus injection of GH RH. Each subject received one IV bolus injection containing 1 JIg/kg of human GH-RH. Samples were collected in chilled tubes (4 C) for serum or plasma. Samples were immediately centrifuged at 1,000 X g at 4 C for 10 minutes. The supernatant plasma or serum was aspirated, aliquoted, and stored at -70 C until assayed. Assays Quantitative human GH was determined by an immunoassay kit (Allegro HGH; Nichols Institute, San Juan Capistrano, CA). The intra-assay variation for GH was 4.3% and the interassay variation was 7.3%. Estradiol-17fJ was quantified using a solid phase, time-resolved fluoroimmunoassay (Delfia, Wallac Oy, Finald; Electro-Nucleonics, Columbia, MD). Unknown serum samples, internal standards, and standard curves were assayed on a LKB-Wallac Arcus spectrofluorometer (Electro-Nucleonics). The intra-assay variation was 7.6% and the interassay variation was 9.2%. Plasma IGF-I levels were determined by an immunoassay kit (Allegro Sm-C; Nichols Institute). The intra-assay variation for GH was 5.0% and the interassay variation was 9.0%. All tests were performed in duplicate, and all samples from each subject were measured in a single assay. Statistical Analysis Growth hormone secretion was evaluated by integrating the region circumscribed by plasma G H values during the test period of 180 minutes using a computerized Pharmacologic Calculation System (Pharm/PCS-Version 3.1). Because of heterogeneous variances of GH and E2 measurements (small variations in the GnRH-a treated women), differences in these values were analyzed for statistical significance using the nonparametric Mann Whitney U-test. Basal GH, E2, and IGF-I levels were compared using the two-tailed Mann-Whitney U-test. Probabilities of <0.05 were considered significant. RESULTS A summary of the clinical and laboratory findings of the seven women treated with GnRH -a is presented in Table 1. Two women were receiving treatment for uterine leiomyoma, and five subjects were receiving GnRH-a before ovarian stimulation for in vitro fertilization (lvf). All seven women experienced gonadal suppression, as determined by the development of amenorrhea, hot flushes, and suppressed E2 levels or suppression of ovarian follicles to <10 mm diameter (assessed by ultrasound in women who were to undergo IVF). The mean age and body weight of the GnRH -a treated subjects 74 Word et al. Effect of GnRH-a on GH secretion Fertility and Sterility

3 Table 1 Clinical and Laboratory Findings in Women Treated With GnRH-a Subject no. Age Weight Diagnosis PlasmaE 2 Integrated G H response Mean± SE 34.3 ± 1.5 b 8 Control women (n = 8) Mean± SE 30.0 ± 5.6 a IVF, in vitro fertilization. b P = y kg ± 3.3' 70.4 ± 5.0 nmil ng.ml-1.min-i Leiomyoma ,349 IVFa ,342 IVF IVF Leiomyoma ,498 IVF IVF 'P = d P< ± 0.012d ± ,100 ± d 2,915 ± 1,004.4 were not significantly different from control women (Table 1). The effect of GnRH -a treatment on G H -RHstimulated G H release is presented in Figures 1 and 2. The mean basal plasma GH level (-15 and 0 minutes) in control women was 7.95 ± 3.3 ng/ml (mean ± SE). The mean basal GH level in the Gn RH -a treated group was 6.0 ± 2.5 ng/ml, and these differences were not statistically significant. The mean peak GH response (15 minutes after the injection of GH-RH) was 24.9 ± 10.9 ng/ml in control women, whereas the mean peak GH response in the GnRH-a treated women was much lower (9.3 ± 2.1 ng/ml, P < 0.05). The eight control women experienced a variable increase in GH secretion after an IV bolus of G H -RH as shown in the shaded ~ 30 os '"., c: o E 20 ~ :I:.c: i o 10 i o t Time (min) Figure 1 Effect of GnRH-a on GH-RH-stimulated GH release. The shaded area represents mean ± SE for control women; the solid circles represent women receiving GnRH-a. The arrow indicates the time of injection of GH-RH. Growth hormone values are expressed as ng/ml, and each point represents mean ± SE. 180 area of Figure 1. In contrast to the control women, G H release over the 3 hours was suppressed and less variable in the GnRH -a treated women. As seen in Figure 2, the mean integrated G H secretion area was <50% ofthe control women (1,100 ± ng.ml-1.min-1 versus 2,915 ± 1,004 ng.ml- l. min-i, P < 0.05). All women who received GnRH-a had reduced basal serum E2 levels (0.031 ± nm), whereas control women had significantly higher E2 levels (0.255 ± nm, P < 0.05) as expected for premenopausal women in the follicular phase of the menstrual cycle (Fig. 3). We observed a positive correlation between the mean integrated GH secretory area response to GH-RH and E 2levels (Spearman ranked r = 0.714, P < 0.05). Baseline IGF-I levels in control women (125.4 ± 55.4 mu/ml [mean ± SED were not signifi o L---L -'-_ Figure 2 Effect of GnRH-a on GH-RH-induced GH secretion: mean integrated area circumscribed by the curves. Data represent the mean area circumscribed by the' curve after bolus injection of GH-RH expressed in ng of GH ml-'.min-'. The closed bar represents GH secretion from women treated with GnRH-a and the open bar represents GH secretion in control women. * P < Word et al. Effect of GnRH-a on GH secretion 75

4 ~ i5 :t os 150! '3 100 c:! ]; w o Figure 3 (Left panel), The effect of GnRH-a on basal serum E2 levels. The open bar represents the mean ± SE baseline serum E2 in eight control women in the follicular phase of the menstrual cycle. The closed bar represents E2 levels in seven women receiving GnRH -a for a minimum of 4 weeks. * P < (Right panel), The effect of GnRH-a on basal IGF-I levels. The open bar represents mean ± SE basal serum IGF-I levels in milliunits per milliliters in control women. The closed bar represents IGF-I levels in women receiving GnRH-a. cantly different from baseline levels (176.8 ± 51.0 mu/ml) in the GnRH-a treated women (Fig. 3). There was no correlation between the G H -RH response and basal IGF-I levels, body weight, or age. DISCUSSION Suppression of the pituitary-gonadal axis by the administration of GnRH -a is now a widely accepted treatment modality for a variety of gynecological conditions (for review, see Santen and Bourguignon! and Forti2). The suppressive effects of GnRH -a on gonadotropins and gonadal function is well known. Recent evidence suggests that GnRH-a may act to regulate GH secretion in girls with precocious puberty.3 We examined the effects of a GnRH -a on G H secretion, serum E2 levels, and IGF-I levels in adult premenopausal women. We observed that GH-RH-stimulated GH release and E2 levels are significantly suppressed in women receiving GnRH -a. The effects of GnRH-a on GH secretion and IGF-I levels in young girls with central precocious puberty have been investigated as discussed previously.3 Nocturnal serum GH and plasma IGF-I levels were suppressed throughout the 2 years of gonadal suppression, and these values returned to pretreatment levels after discontinuation of therapy. Others observed that resting and exercise-induced GH secretion is lower in postmenopausal estrogen deficient women and increases after oral estrogen replacement therapy.4 In the present study, basal GH secretion is not significantly different be- tween control women and GnRH -a treated women. However, the GH response to a bolus infusion of GH-RH is significantly attenuated in women receiving a GnRH-a. The mean peak GH response of control ovulatory women (24.9 ng/ml) is similar to the mean peak G H response that is reported in young adult men after GH-RH infusion (20.9 ng/ml).9 The reported mean peak GH response to GH-RH is 15.3 ng/ml in postmenopausal women, which increases significantly to 23.0 ng/ml after estrogen replacement.4 The peak level of GH (9.3 ng/ml) in the GnRH-a treated group is less than that reported for estrogen deficient menopausal women and markedly reduced in comparison with the eight premenopausal control subjects of the present study.7 In the present study, an increase in the GH response to GH-RH is positively correlated with basal E2 levels. Therefore, the decreased release of GH in the GnRH-a treated women may be secondary to the hypoestrogenism induced by such treatment. There is in vitro and in vivo evidence to suggest that estrogen has a regulatory role in GH secretion. Estrogen increases GH content of rat pituitary cells in culture but has no effect on basal GH secretion by these cells.lo Sex steroid supplementation in men with hypogonadal hypogonadism results in increased G H levels. ll Our results are consistent with the view that estrogen acts to cause an increase in pituitary GH stores because treatment with a GnRH -a resulted in hypoestrogenism and in reduced GH release, despite normal baseline GH levels. Age and increased body fat also have been associated with decreased GH secretion in women.12,13 However, we observed no significant differences in age or body weight between the two study groups (Table 1). The role ofigf -I in the pathogenesis of a variety of gynecological conditions has been investigated.12,14,15 The reported levels of IGF-I after estrogen treatment differ. Insulin-like growth factor I levels were lower in postmenopausal women after estrogen therapy, despite the development of an increase in GH-RH responsiveness during estrogen therapy.4 Estradiol treatment in persons with acromegaly results in reduced immunoreactive IGF-I levels,16 and others have found a decrease in IGF-I activity in normal men after pharmacological estrogen therapy.5 Conversely, others have found that IGF-I levels increase with estrogen ther- 76 Word et al. Effect of GnRH-a on GH secretion Fertility and Sterility

5 apy/7-19 which is consistent with the results of Mansfield et al.3 who demonstrated decreased levels of IGF-I during gonadal suppression with GnRH-a in children with precocious puberty. Basal IGF-I levels were much higher in the children with precocious puberty (2,000 to 7,500 mu/ ml) compared with the IGF-I levels of adult women in the present study (177 mu/ml). In the present study, basal IGF-I levels were slightly higher in the GnRH -a treated group than in the control group; however, this difference was not statistically significant. In addition to the effects of GnRH-a on estrogen and IGF-I levels, there is evidence that GnRH-a may have direct effects on pituitary cells other than gonadotropes. Gonadotropin-releasing hormone stimulates thyroid-stimulating hormone (TSH) secretion from pituitary thyrotrope tumors in vitro, and the thyrotrope tumor secretes G H and ex-subunit in addition to TSH.20 Moreover, others have demonstrated at least three peptides are coreleased with LH when pituitary cell aggregates from the rat are stimulated with GnRH.21 Intraperitoneal injection of GnRH acted to cause an elevated circu- 1ating G H level in female goldfish, and GnRH stimulated dose-dependent, reversible GH secretion from goldfish pituitary fragments in vitro.22 These results suggest that GH secretion may be directly regulated by GnRH. Gonadotropin-releasing hormone infusion in normal men and women acts to cause an increase in serum GH and TSH,23 and GnRH stimulates G H secretion in subjects with active acromegaly.24 Taken together, these studies suggest direct regulation of pituitary GH secretion by GnRH. The possibility then exists that GnRH-a may have direct inhibitory effects on G H secretion from the anterior pituitary, in addition to the indirect effects induced by hypoestrogenism. In summary, GH-RH-stimulated GH release is decreased in hypoestrogenic women receiving Gn RH-a. Decreased GH secretion may be a direct effect ofthe agonists on the pituitary, or an indirect effect secondary to hypoestrogenism, or both. The effect of GnRH -a on GH secretion or on other pituitary hormones and growth factors may contribute to their success in the treatment of various gynecological conditions. Acknowledgments. We thank Ms. Sandy Nance for her expert editorial assistance. We also thank Micki Roark, R.N., and Ms. Nora Cline for assisting in the blood sampling and hormone assays, respectively. REFERENCES 1. Santen RJ, Bourguignon JP: Gonadotropin-releasing hormone: physiological and therapeutic aspects, agonists and antagonists. Horm Res 28:88, Forti G: Clinical applications of GnRH analogs. 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6 romegaly: reduction of immunoreactive somatomedin-c and improvement in metabolic status. Am J Med 69:571, Meyer W, Furlanetto RW, Walker PA: The effect of sex steroids on radioimmunoassayable plasma somatomedin-c concentrations. J Clin Endocrinol Metab 55:1184, Ross JL, Cassoria FG, Skuda MC, Valk 1M, Loriairz KL, Cutler GB: A preliminary study of the effect of estrogen dose on growth in Turner's syndrome. N Engl J Med 309: 1104, Copeland KC, Johnson DM, Kuehl T J, Castracane VD: Estrogen stimulates growth hormone and somatomedin-c in castrate and intact female baboons. J Clin Endocrinol Metab 58:698, Simard M, Mirell CJ, Pekary AE, Drexler J, Kovacs K, Hershman JM: Hormonal control of thyrotropin and growth hormone secretion in a human thyrotrope pituitary adenoma studied in vitro. Acta Endocrinol (Copenh) 119: 283, Sion B, Chanat E, Duval J, Thieulant ML: Peptides coreleased with luteinizing hormone by perifused pituitary cell aggregates. Mol Cell Endocrinol60:151, Marchant TA, Chang JP, Nahorniak CS, Peter RE: Evidence that gonadotropin-releasing hormone also functions as a growth hormone-releasing factor in the goldfish. Endocrinology 124:2509, Amsterdam JD, Winokur A, Lucki I, Snyder P, Harris RI, Caroff S, Rickels K: Growth hormone prolactin and thyrotropin responses to gonadotropin-releasing hormone in depressed patients and healthy volunteers. Psychoneuroendocrinology 7:177, Rubin AL, Levin SR, Bernstein RI, Tyrrel TB, Noacco C, Forsham PH: Stimulation of growth hormone by luteinizing hormone-releasing hormone in active acromegaly. J Clin Endocrinol Metab 37:160, Word et al. Effect of GnRH-a on GH secretion Fertility and Sterility