Ovarian control of pituitary sensitivity of luteinizing hormone secretion to gonadotropin-releasing hormone in women with the polycystic ovary syndrome This study investigated the ovarian control of LH responsiveness to GnRH in anovulatory women with the polycystic ovary syndrome (PCOS). It is suggested that the enhanced pituitary sensitivity of LH secretion to GnRH in anovulatory women with PCOS is not due to a reduced production but rather to a defect in the interaction of ovarian factors on the hypothalamic–pituitary system. (Fertil Steril 2009;92:1378–80. 2009 by American Society for Reproductive Medicine.)
Elevated plasma LH concentrations and hyper-responsiveness of the pituitary to GnRH have been found in the majority of women with the polycystic ovary syndrome (PCOS) (1–4). Whether this is related to a defect in the ovarian feedback mechanisms or to hypothalamic dysfunction has been a matter of discussion that remains unclarified. At present, the published studies have investigated the role of the ovaries under basal conditions and only after the exogenous administration of steroids. It is known that the ovaries, in addition to the steroids, secrete nonsteroidal substances, the activity of which becomes particularly evident under conditions of ovarian stimulation by exogenous FSH (5). Such a dynamic approach has been previously applied to normal women and has provided important information regarding the ovarian control of gonadotropin secretion (6, 7). The present study was undertaken to investigate the role of the ovaries in the regulation of the pituitary sensitivity to GnRH in terms of LH secretion in anovulatory women with PCOS to obtain further insight into the pathophysiology of the syndrome. Ten women with PCOS (aged 24.5 2.1 years) and eight women with normal menstrual cycles (aged 31.5 1.0 years) (controls) volunteered for the study and gave written informed consent. Institutional Review Board (IRB) approval of the study was obtained. Polycystic ovary syndrome was diagnosed according to the revised 2003 criteria (8). The women with PCOS had a body mass index (BMI) of 26 2.5 kg/m2 and the controls of 21 1.1 kg/m2. All women participated in the same experimental procedure (Exp) including the SC injection of a single dose 450 IU reReceived December 31, 2008; revised and accepted April 15, 2009; published online May 24, 2009. K.D. has nothing to disclose. C.V. has nothing to disclose. S.P. has nothing to disclose. A.K. has nothing to disclose. I.E.M. has nothing to disclose. Reprint requests: Ioannis E. Messinis, M.D., Ph.D., Department of Obstetrics and Gynecology, Medical School, University of Thessalia, 41110 Larissa, Greece (FAX: 30-2410-670096; E-mail: [email protected]
combinant FSH (Puregon 150 IU; Organon, Hellas, Greece) based on our previous studies (6, 7, 9). The day of the FSH injection was designated as day 1 (9 AM). The GnRH pulses, 10-mg each (Relefact LHRH, 0.1 mg/mL; Hoechst, Frankfurt, Germany) were injected before the injection of FSH, the next two every 12 hours and the remainder every 24 hours up to day 6. Blood samples in relation to each GnRH injection (time 0) were obtained at -15, 0, and 30 minutes. The 30-minute point shows a maximal response to pituitary sensitivity to GnRH (10). The Exp-control was performed during the early follicular phase of a spontaneous cycle in the eight normal women starting on cycle day 2. In the group of women with PCOS, Exp-1-PCOS and Exp-2-PCOS were performed, the first at least 15–20 days after a spontaneous menstrual period and the second at the end of a 20-day treatment with P (Utrogestan capsules, 100 mg of P; Faran, Athens, Greece) at the oral dose of 300 mg/day. Progesterone administration started the day after the end of Exp-1-PCOS. Follicle-stimulating hormone and LH were measured in all blood samples. Basal values of E2 and P were also measured at -15 and 0 minutes and on the 10th day of P administration. Hormonal measurements were performed using established commercial assays. Hormone values were normally distributed (one sample Kolmogorov-Smirnov test). Statistical analysis was performed by one-way analysis of variance (ANOVA) followed by Bonferroni post hoc testing, paired t-test, and repeated measures ANOVA with Geisser-Greenhouse adjustments followed by Bonferroni post hoc testing. All values are expressed as means standard error of the mean. An a-level of 0.05 was used to determine statistical significance. The statistical software package used was NCSS 2001 (Number Cruncher Statistical Systems, Kaysville, UT). The response of LH to GnRH was calculated as the net increase at 30 minutes (DLH) after their basal value (mean of the values at -15 and 0 minutes). The DLH values were significantly higher on day 1, before FSH injection, in Exp-1-PCOS than in Exp-2-PCOS and Exp-control (P<.05) (Fig. 1a). In the Exp-1-PCOS, DLH values
Fertility and Sterility Vol. 92, No. 4, October 2009 Copyright ª2009 American Society for Reproductive Medicine, Published by Elsevier Inc.
FIGURE 1 The DLH values (a) and serum FSH (b), LH (c), E2 (d), and P (e) concentrations (mean SE) during the experimental period in women with PCOS (n ¼ 10) (green line) before (Exp-1-PCOS) and (red line) after treatment with exogenous P (Exp-2-PCOS) and (blue line) in normal women (n ¼ 8) (Exp-control). (a) *P< .05, higher in Exp1-PCOS than in Exp-2-PCOS and Exp-control. þ P< .05, higher in Exp-1-PCOS and in Exp-control than in Exp2-PCOS. (c) * P< .05, higher in Exp-1-PCOS than in Exp-2-PCOS and Exp-control. The LH levels in Exp-2-PCOS and Exp-control decreased significantly up to day 3 and increased significantly thereafter (P< .05). (d) * P< .001, higher in Exp-1-PCOS and in Exp-2-PCOS than in Exp-control. þ P< .05, higher in Exp-1-PCOS than in Exp-2PCOS and Exp-control. See text for abbreviations.
Dafopoulos. Correspondence. Fertil Steril 2009.
showed a trend for a decrease until day 2, increasing significantly thereafter (P<.05) (Fig. 1a). In Exp-2-PCOS and Exp-control, DLH values decreased significantly until days 2 and 3 (P<.05), increasing significantly thereafter (P<.05) (Fig. 1a). The DLH values remained higher during Exp-1-PCOS compared with Exp-control (P<.05) and Exp-2-PCOS (P<.05) (Fig. 1a). The DLH values were almost identical in Exp-2-PCOS and Exp-control (Fig. 1a). The FSH values increased significantly in the first 24 hours after the FSH injection (P<.05) in all experiments (Fig. 1b). Basal LH values before the onset of the experiments in the PCOS group were significantly higher than Fertility and Sterility
in the control group (P<.05), but after the treatment with P they decreased, becoming similar to those in the controls (Fig. 1c). The LH values in all experiments decreased gradually after the injection of FSH until day 3 (P<.05) (Fig. 1c), but in Exp-2-PCOS and Exp-control, they increased significantly thereafter (P<.05) (Fig. 1c). Serum E2 values increased significantly after the FSH injection in all three experiments (Fig. 1d) until day 3 or 4 (P<.05). Serum P levels increased significantly in the women with PCOS (6.4 1.5 ng/mL on the 10th day of its administration), but were stable throughout the three experimental periods, with no significant differences between them (Fig. 1e).
In the present study, we used a P regimen to normalize the LH response to GnRH (11, 12) and a dynamic approach with the exogenous administration of a single high dose FSH to increase the activity of steroidal and nonsteroidal ovarian substances and assess the behavior of the pituitary to such changes (6, 7). Under these conditions, the response of LH to GnRH during a simulated follicular phase after the injection of FSH in the women with PCOS was almost identical with that in the early follicular phase of normal women. Although the normalization of LH secretion in response to GnRH on day 1 of the simulated follicular phase (Fig. 1) was related to the action of P, the further significant reduction in the LH response during the next 2 days was due to the FSH-mediated increased activity of ovarian substances in a way similar to that in the normal early follicular phase (6, 7). Estradiol is the main ovarian factor that sensitizes the pituitary gonadotrophs to GnRH (5). Accumulating evidence has indicated that a nonsteroidal ovarian factor, named gonadotropin surge attenuating factor, is possibly involved by antagonizing the sensitizing effect of E2 (5). Recently, we provided interesting information regarding the characterization of this factor (13). It is tempting, therefore, to speculate that under the present experimental conditions, the amount of gonadotropin surge attenuating factor produced by the polycystic ovaries (PCO) was similar to that produced by the normal ovaries. The present results are consistent with a differential action of E2 on the pituitary during the two experimental periods in the patients with PCOS. The levels of this steroid were significantly higher in women with PCOS before the administration of P, in agreement with previous data (14). Although a similar E2 response to FSH was seen during the two periods, the long-term exposure of the pituitary to the unopposed action of this steroid could overcome the antagonizing effect of ovarian substances on the pituitary sensitivity before but not after treatment with P. The mechanism of this difference is not clear, although the administered P might have modulated the action of E2 centrally by decreasing the number of E2 receptors (15, 16). Other substances, such as inhibin and leptin, are less likely to have affected this interaction (17, 18). In conclusion, the present results suggest for the first time that the enhanced pituitary sensitivity to GnRH in terms of LH secretion during the periods of anovulation in women with PCOS is not due to a reduced production but to a defect in the interaction of different ovarian factors on the hypothalamic–pituitary system. Konstantinos Dafopoulos, M.D.a Christos Venetis, M.D.a Spyros Pournaras, M.D.b Athanasios Kallitsaris, M.D.a Ioannis E. Messinis, M.D., Ph.D.a 1380
Dafopoulos et al.
Department of Obstetrics and Gynecology and b Department of Microbiology, Medical School, University of Thessalia, Larissa, Greece
REFERENCES 1. Balen AH, Conway GS, Kaltsas G, Techatrasak K, Manning PJ, West C, et al. Polycystic ovary syndrome: the spectrum of the disorder in 1741 patients. Hum Reprod 1995;10:2107–11. 2. Hendriks ML, Brouwer J, Hompes PG, Homburg R, Lambalk CB. LH as a diagnostic criterion for polycystic ovary syndrome in patients with WHO II oligo/amenorrhoea. Reprod Biomed Online 2008;16:765–71. 3. Mortimer RH, Lev-Gur M, Freeman R, Fleischer N. Pituitary response to bolus and continuous intravenous infusion of luteinizing hormonereleasing factor in normal women and women with polycystic ovarian syndrome. Am J Obstet Gynecol 1978;130:630–4. 4. Patel K, Coffler MS, Dahan MH, Malcom PJ, Deutsch R, Chang RJ. Relationship of GnRH-stimulated LH release to episodic LH secretion and baseline endocrine-metabolic measures in women with polycystic ovary syndrome. Clin Endocrinol 2004;60:67–74. 5. Messinis IE. Ovarian feedback, mechanism of action and possible clinical implications. Hum Reprod Update 2006;12:557–71. 6. Messinis IE, Lolis D, Papadopoulos L, Tsahalina T, Papanikolaou N, Seferiadis K, et al. Effect of varying concentrations of follicle stimulating hormone on the production of gonadotrophin surge attenuating factor (GnSAF) in women. Clin Endocrinol 1993;39:45–50. 7. Messinis IE, Lolis D, Zikopoulos K, Tsahalina E, Seferiadis K, Templeton AA. Effect of an increase in FSH on the production of gonadotrophin-surge-attenuating factor in women. J Reprod Fertil 1994;101:689–95. 8. The Rotterdam ESHRE/ASRM-sponsored PCOS consensus workshop group 2004. Revised (2003) consensus on diagnostic criteria and longterm health risks related to polycystic ovary syndrome. Hum Reprod 2003;19:41–7. 9. Lolis DE, Tsolas O, Messinis IE. The follicle-stimulating hormone threshold level for follicle maturation in superovulated cycles. Fertil Steril 1995;63:1272–7. 10. Wang CF, Lasley BL, Lein A, Yen SC. The functional changes of the pituitary gonadotrophs during the menstrual cycle. J Clin Endocrinol Metab 1976;42:718–28. 11. Fiad TM, Cunningham SK, McKenna TJ. Role of progesterone deficiency in the development of luteinizing hormone and androgen abnormalities in polycystic ovary syndrome. Eur J Endocrinol 1996;135:335–9. 12. Dafopoulos K, Kotsovassilis CG, Milingos S, Kallitsaris A, Galazios G, Zintzaras E, et al. Changes in pituitary sensitivity to GnRH in estrogentreated post-menopausal women: evidence that gonadotrophin surge attenuating factor plays a physiological role. Hum Reprod 2004;19:1985–92. 13. Karligiotou E, Kollia P, Kallitsaris A, Messinis IE. Expression of human serum albumin (HSA) mRNA in human granulosa cells: potential correlation of the 95 amino acid long carboxyl terminal of HSA to gonadotrophin surge-attenuating factor. Hum Reprod 2006;21:644–50. 14. Anderson RA, Groome NP, Baird DT. Inhibin A and inhibin B in women with polycystic ovarian syndrome during treatment with FSH to induce mono-ovulation. Clin Endocrinol 1998;48:577–84. 15. Smanik EJ, Young HK, Muldoon TG, Mahesh VB. Analysis of the effect of progesterone in vivo on estrogen receptor distribution in the rat anterior pituitary and hypothalamus. Endocrinology 1983;113:15–22. 16. Blaustein JD, Brown TJ. Progesterone decreases the concentration of hypothalamic and anterior pituitary estrogen receptors in ovariectomized rats. Brain Res 1984;304:225–36. 17. Wachs DS, Coffler MS, Malcom PJ, Chang RJ. Comparison of folliclestimulating-hormone-stimulated dimeric inhibin and estradiol responses as indicators of granulosa cell function in polycystic ovary syndrome and normal women. J Clin Endocrinol Metab 2006;91:2920–5. 18. Messinis IE, Milingos S, Zikopoulos K, Kollios G, Seferiadis K, Lolis D. Leptin concentrations in the follicular phase of spontaneous cycles and cycles superovulated with follicle stimulating hormone. Hum Reprod 1998;13:1152–6.
Vol. 92, No. 4, October 2009