Risto Erkkola, M.D.t Kerttu Irjala, M.D. Kristiina Ruutiainen, M.D.tll

Transcription

1 FRTILITY AND STRILITY Copyright e 199 The American Fertility Society Vol. 58, No.4, October 199 Printed on acid-free paper in U.S.A. Serum androgen and gonadotropin levels decline after progestogeninduced withdrawal bleeding in oligomenorrheic women with or without polycystic ovaries Leena Anttila, M.D.t:j: Pertti Koskinen, M.D. Hanna-Leena Kaihola, Ph.D. Risto rkkola, M.D.t Kerttu Irjala, M.D. Kristiina Ruutiainen, M.D.tll University of Turku, Turku, Finland Objective: To examine the effect of short-term progestogen treatment on androgen, gonadotropin, and sex hormone-binding globulin (SHBG) levels in oligomenorrheic women. Design: Comparative study of changes in hormonal parameters in patients with or without ultrasonographically diagnosed polycystic ovarian disease (PCOD). Setting: Open patient clinic of reproductive endocrinology at University Central Hospital of Turku, Finland. Patients: Seventy-five oligomenorrheic women with (n = 51) or without (n = 4) PCOD. Main Outcome Measures: Serum concentrations of testosterone (T), androstenedione (A), dehydroepiandrosterone sulfate, luteinizing hormone (LH), follicle-stimulating hormone (FSH), andshbg. Results: The levels oft, A, LH, and the LH:FSH ratios decreased significantly after oral treatment with medroxyprogesterone acetate (1 mg/d for 1 days) in non-pcod women and in women with PCOD decreasing the frequencies of pathological laboratory findings, in particular elevated levels of LH:FSH ratio and A in PCOD women and of LH:FSH ratio in non-pcod women. The levels oft, A, and LH as well as the LH:FSH ratio were significantly higher in women with PCOD. Obesity was associated with high free androgen indices, low LH:FSH ratios, and low concentrations of LH, A,andSHBG. Conclusions: The serum samples for hormonal analyses used as an aid in diagnosing PCOD should be obtained without pretreatment with progestogen because it masks the biochemical findings of PCOD. Fertil Steril199;58:697-7 Key Words: Polycystic ovarian disease, oligomenorrhea, androgens, gonadotropins, medroxyprogesterone acetate Polycystic ovarian disease (PCOD) is a common cause of amenorrhea, oligomenorrhea, and infertility (1, ). The etiology and pathophysiology of this Received April, 199; revised and accepted June 4, 199. * Supported by grants from the Paulo Foundation, Helsinki, Finland, and the Foundation of the University of Turku, Turku, Finland. t Department of Obstetrics and Gynecology. :j: Reprint requests: Leena Anttila, M.D., Department of Obstetrics and Gynecology, Turku University Central Hospital, Kiinamyllynkatu 4-8, SF-5 Turku, Finland. Central Laboratory, Turku University Central Hospital. II Present address: Department of Obstetrics and Gynecology, The University of Maryland, Baltimore, Maryland. clinical entity are still incompletely understood. Likewise, several diagnostic criteria based on ovarian morphology (-4) or biochemical findings have been presented for PCOD (5-7). In most studies on androgen metabolism in eumenorrheic women, the sampling for hormonal analyses has been carried out in the early follicular phase. In amenorrheic and oligomenorrheic subjects, the results of the hormonal analyses are presumed to become more comparable with the normal early follicular phase values by taking the samples after a progestogen-induced withdrawal bleeding (8). However, it may be questioned whether such treatment would not change the androgen and gonado- Vol. 58, No.4, October 199 Anttila et al. MP A effect on hormones in oligomenorrhea 697

2 tropin status of a patient with menstrual irregularities, thereby masking diagnostic hormonal findings. Cycle-regulating progestagens (e.g., medroxyprogesterone acetate [MPA)) are known to suppress testosterone (T) (9), luteinizing hormone (LH) (9-11), and sex hormone-binding globulin (SHBG) levels (1). The aim of the present study was to investigate the effect of progestogen treatment on the levels of serum androgens, gonadotropins, and SHBG in oligomenorrheic women with or without morphologically diagnosed PCOD. Subjects MATRIALS AND MTHODS The study group consisted of 75 oligomenorrheic women. Oligomenorrhea was defined as menstrual cycles of >6 weeks during the previous 6 to 1 months. Patients with hyperprolactinemia or thyroid dysfunction were excluded. All subjects attending the study bled in response to a progestogen challenge indicating an estrogen ()-primed endometrium. The diagnosis of PCOD was based on ovarian morphology assessed by the same investigator (L.A.) using a 7.5-MHz vaginal ultrasonographic vaginal probe (Combison 31; Kretztechnik, Zipf, Austria) and diagnostic criteria by Adams (3). The body mass index (BMI, weight [kg]/height [m]) was measured. Hair growth was assessed using a modification of the criteria of Ferriman and Gallwey (13). In hirsute and hyperandrogenic women, a dexamethasone suppression test was performed to exclude Cushing's syndrome. Adrenocorticotropin stimulation test was performed when needed to exclude late onset adrenal hyperplasia. Polycystic ovaries fulfilling diagnostic criteria (enlarged ovaries with multiple small subcortical follicles and increased stroma) were found in 51 women. Their age ranged from 14 to 4 years (7.9 ± 5.5 years, mean ± SD) and the BMI from 17 to 56 kg/m (8.8 ± 7.7 kg/m). The remaining 4 women had normal (n = 18) or multicystic (n = 6) ovaries on ultrasound. The age of the non-pcod women ranged from 19 to 4 years ( years) and the BMI from 18 to 48 kg/m (8.4 ± 9. kg/m). Thirty-one women with and 1 without PCOD were obese (BMI > 5 kg/m). Hirsutism (total score> 9) were found in 11 women with PCOD. None of the women without PCOD were hirsute. Hormone Sampling The blood samples were collected after an overnight fast between 7 and 9 A.M. The first samples were taken during oligomenorrhea. The sampling was repeated after an MPA-induced (MPA 1 mg/d for 1 days) withdrawal bleeding (days 3 to 7 after the onset of induced menstrual flow). Hormone and SHBG Assays Serum T and androstenedione (A) were measured with in-house radioimmunoassays (RIAs) as described previously (14). Dehydroepiandrosterone sulfate (DHAS) was measured with an RIA (151_ DHAS RIA kit; Baxter Dade AG, Dudingen, Germany). Luteinizing hormone and follicle-stimulating hormone (FSH) were measured with time-resolved immunofluorometric assays (Delfia; Wallac Oy, Turku, Finland). Sex hormone-binding globulin was determined with a ligand-binding assay (14). The free androgen index was calculated as the quotient T (nmoljl)/shbg (nmoljl) X 1,. The early follicular phase reference intervals for all the analytes were determined in 4 healthy female volunteers (14). Statistical Methods Paired differences were evaluated with the paired t-test. The effects of PCOD and obesity in the entire study group and hirsutism in the subgroup with PCOD on the hormone and SHBG levels were assessed with ANCOV A. The changes in the distribution were analyzed with McNemar's Xtest. RSULTS The serum concentrations of T, A, and LH as well as the LH:FSH ratios decreased significantly after treatment with MPA in women with PCOD as well as in non-pcod women (Table 1, Fig. 1). The levels ofdhas and the free androgen indices decreased significantly only in PCOD women (Table 1, Fig. 1). The levels of T, A, and LH as well as the LH: FSH ratios and the free androgen indices were higher in women with PCOD than in non-pcod women both in the oligomenorrheic state and after MPA-induced bleeding (Table 1). The PCOD and non-pcod women did not differ with respect to serum DHAS concentrations (Table 1). Figure shows the distributions of the women according to normal (outside the appropriate circle) or elevated (inside the circle) LH:FSH ratios, free androgen indices, and A concentrations in oligomenorrhea and after MPA-induced bleeding. The overlaping areas present those with more than 698 Anttila et at. MPA effect on hormones in oligomenorrhea Fertility and Sterility

3 Table 1 Gonadotropins, Androgens, and SHBG in the Study Groups* Cycle phase PCODwomen Probabilityt Non-PCOD LH (lull) Oligomenorrhea 1.8 ± 5.5 (1. to 9):1: < ± 5.5 (1 to 18) After MPA 7.4 ± 4.5 (1.4 to 8):1: < ±.8 (1.4 to 1) LH:FSH ratio Oligomenorrhea.4 ± 1.1 (.9 to 5.6):1: < ± 1.1 (.3 to 3.8) After MPA 1.7 ±.8 (.4 to 4.):1: < ±.6 (.3 to.1) T (nmol/l) Oligomenorrhea 3.1 ±.9 (1.9 to 5.9):1: <.1. ±.4 (1.5 to 3.3) After MPA.5 ±.7 (1.3 to 4.7):1: < ±.3 (1. to.4) SHBG (nmol/l) Oligomenorrhea 35. ±.5 (7 to 91) II NS1f 4. ±.6 (18 to 94) II After MPA 34.8 ±.8 (6 to 11) II NS 38.4 ± 19. (19 to 88) II Free androgen Oligomenorrhea 1Z ±83 (35 to 475):1: <.1 56 ± 4 ( to 111) II index After MPA 98 ± 64 (6 to 3):1: <.1 55 ± 4 (19 to 1) II A (nmol/l) Oligomenorrhea 13.3 ± 3.9 (7. to.6):1: < ±.5 (3.8 to 15.9) After MPA 1.3 ± 3.4 (5.1 to.1):1: < ± 1.4 (4.9 to 1.8) DHAS Oligomenorrhea 7. ± 3.4 (. to 16.3) NS 5.8 ±.9 (1.7 to 1.5) II (ltmol/l) After MPA 7.6 ± 4.1 (. to 16.6) NS 6. ±.3 (1.6 to 9.7) II * Values are means ± SD with ranges in parentheses. t The difference between PCOD and non-pcod women. :I: The difference between the results obtained during oligomenorrhea and after MPA-induced bleeding, P <.1. The difference between the results obtained during oligomenorrhea and after MPA-induced bleeding, P <.1. II The difference between the results obtained during oligomenorrhea and after MPA-induced bleeding is not significant. 1f NS, not significant. one abnormal finding. Treatment with MP A decreased the frequency of pathological laboratory findings, in particular elevated levels of LH:FSH ratio (from 66% to 38%, P <.1) and A (from 66% to 34%, P <.1) in PCOD women and elevated levels of LH:FSH ratio (from 57% to 13%, P <.5) in non-pcod women. Seventy-seven percent of PCOD women had a combination of more than one pathological finding. The frequency decreased to 51 % after MP A in the PCOD group (P <.1). Among the non-pcod group, elevated LH:FSH ratio together with elevated free androgen index was found in 4 % of subjects in the non PCOD group during oligomenorrhea. Only one of the non-pcod women had high A during amenorrhea (Fig. 1). Free androgen index was elevated in 83% of PCOD patients and 59% of non-pcod patients, in whom mainly obese subjects had elevated free androgen indicies (Fig. ). No significant difference was found in BMI between PCOD and non-pcod women. Obesity was associated with high levels of free androgen index (P <.1, Fig. 3) and low levels of LH (P <.1), LH:FSH ratio (P <.1, Fig. 3), A (P <.5, Fig. 3), and SHBG (P <.1) in oligomenorrheic state. The levels of SHBG were significantly lower (9.7 ± 18.3 nmoljl versus 4. ± 1.1 nmoljl, P <.5) and the levels of free androgen index significantly higher (13.8 ± 73.9 versus 89.7 ± 45., P <.5) in hirsute than in nonhirsute PCOD women. No significant difference was found in BMI between the hirsute and nonhirsute PCOD groups. DISCUSSION We have previously shown that the timing of the hormone sampling is important when exploring the androgen and particularly the gonadotropin status in regularly menstruating women (14). During a normal menstrual cycle, T and SHBG values are greater in the luteal phase, but despite the cyclic variation, serum androgen concentrations and especially free androgen index are confined within narrow limits (14). However, the levels of the LH: FSH ratio as measured by novel fluoroimmunoassays vary more in the luteal phase (range.3 to 3.5; coefficient variation [CV] 6%) than in the early follicular phase (range. to 1.7; CV 3%) (14). Identifying excessive androgen production resulting in hyperandrogenism and altered gonadotropin secretion resulting in a high LH:FSH ratio has widely been used in the biochemical diagnosis of PCOD (5-7). To get adequate information of gonadotropin and androgen status is problematic in patients with occasional ovulations and/or periods, as often found in PCOD patients. It has been recommended that the hormone samples should be taken in amenorrheic and oligomenorrheic patients after a progestogen-induced bleeding (8). This view relies on the assumption that progestogen administration would mimic the luteal phase, rendering the results more comparable with the early follicular phase values. In this study, we found highly significant declines in the levels of LH and androgens after a short-term Vol. 58, No.4, October 199 Anttila et al. MPA effect on hormones in oligomenorrhea 699

4 ... % II) u. ::: %...J '" ::: ". 15 " e 1 «" 5. II) 5, ::: 4 ". Cl 3 :I: '" II) ::: ". M M M M M M ob le.n ob.,. I n obi I n hinute "onhinut. nonhirlute pedd non-pcoo M M M M M M ob... I.,n ob... I.. n ob.,. le.n hinut. nonhinut. "orhinut. pedd non-pcod Ssil l; '" M M M M M M obese I n obese lean obese I n hirsute nonhinut. "onhinut. peed non-pedd Figure 1 Serum LH:FSH ratios, free androgen indices, and A concentrations in women with PCOD and non-pcod women before () and after (M) MPA-induced withdrawal bleeding. Rectangles indicate the reference intervals. ment suppresses T (9), LH (9), and SHBG (1) levels. Homburg and his co-workers (1) found an intramuscular (1M) short-term MPA (5 mgjd for 5 days) to markedly influence the gonadotropin release. Furthermore, 7 of 1 clomiphene citrate (CC) resistant PCOD women responded to CC with ovulation after such pretreatment (1). Petsos et al. (11) found MPA treatment (5 mg two times daily for 14 days) to suppress LH and FSH levels. In this work, even a short-term MPA treatment masked the biochemical findings of PCOD in several patients (Fig. ). The rather long half-life ofmpa (4 to 5 hours) may explain the changes in hormonal levels even though the samples for endocrinological evaluation were collected approximately 6 to 1 days after the last MP A dose. Despite the mean levels of LH, the LH:FSH ratios' and all other androgen levels except DHAS were significantly higher in the PCOD group, we found considerable heterogeneity of biochemical findings within PCOD patients (Fig. ). This finding is in accordance with the current concept of the heterogeneous etiology of PCOD. Fox et al. () have recently suggested that a positive progestogen challenge test (5 mg oral MPA for 5 days) alone can predict PCOD in oligoamenorrheic subjects. Moreover, they found that the combination of LH and free androgen index was the best biochemical test to predict PCOD in oligoamenorrheic subjects. In this study, all subjects had a positive progestogen challenge test, indicating an -primed endometrium. Within the non-pcod group, 57% of LH/FSH In oligomenorrhea lhffsh Androstenedione LH/FSH In oligomenorrhea lh/fsh Androstenedione Androstenedione oral treatment with MPA (Table 1, Fig. 1). The abnormal gonadotropin secretion in the PCOD patients is presumed to be reversible because of chronic deprivation of progesterone (P) (15). Progesterone acts at hypothalamic levels, decreasing LH pulse frequency (16). Similarly, vaginal P supplementation decreases LH pulse frequency in anovulatory PCOD women (17). Furthermore, in PCOD women ovulation is followed by a temporary normalization of the LH:FSH ratio (18, 19). Long-term MPA-treat- Afler MPA-induced Withdrawal bleeding PCOD After MPA-Induced Withdrawal bleeding non-pcod Figure The distribution (%) ofpcod and non-pcod women according to elevated (inside the appropriate circle) or normal (outside the circle) levels of serum A (upper limit of the reference interval 11 nmol/l), free androgen index (T/SHBG X 1,, upper limit ofthe reference interval 55) and LH:FSH ratio (upper limit of the reference intervall. 7). Percentages of subjects are shown in the states of oligomenorrhea and after MPA-induced withdrawal bleeding. The overlaping areas present those women with more than one pathological finding. 7 Anttila et a1. MPA effect on hormones in oligomenorrhea Fertility and Sterility

5 s : I 4 en..s l<i C) 3 m I 3 o o II!::J en I... -' " (; ODD l1li.,.s en I- 1 c9 c7l BMI (kg/m) c BMI (kg/) o 6 5.s., c. "., c!! " C «CD en o 1 o o cg :. db --.. '""- c9c J ClIO o o cji CO 6 Figure 3 Relationship between BMI and LH:FSH ratios, free androgen index (T/SHBG X 1,), and A in hirsute (_) and nonhirsute women (D) with PCOD and non-pcod oligomenorrheic women (). the women had elevated LH:FSH ratio during oligomenorrhea; thus the elevation of LH is not a pathognomonic feature of PCOD. Furthermore, free androgen index was elevated in 59% of non-pcod patients and, in accordance with earlier reports (1, ), elevated free androgen index was associated with obesity within non-pcod patients (Figs. 1 and 3). The combination of elevated free androgen index together with high LH:FSH ratio was found in 4% of non-pcod patients. Within PCOD women, elevated levels of free androgen index were found in both obese and lean subjects (together 83%). Highest free androgen indexes and lowest SHGB values were found in hirsute PCOD women. Among the PCOD women, 77% had a combination of more than one pathological finding. Interestingly, elevated A concentration, presumably reflecting the ovarian overproduction, was found only in the PCOD group (66%). We recently have demonstrated that obesity is related to low/normal bioactive LH concentrations within PCOD women (3). In this study, we measured the gonadotropins with highly sensitive fluoroimmunoassays, which have a good correlation with the bioassay (4). Our results concerning the association of obesity to lower LH concentrations within PCOD women is in agreement with that previously stated by Laatikainen and his co-workers (5), who found normal LH levels in massively obese PCOD patients. Furthermore, obesity has significant effect on free androgen indexes, LH:FSH ratios, and A concentrations. Thus, the common practice to compare the hormonal findings simply between lean (BMI < 5) and obese (BMI > 5) subjects may obscure the effects of varying degrees of obesity. The ultrasonographically diagnosed PCOD group showed considerable heterogeneity as concerning the biochemical findings. Which of the biochemical changes is the most efficient diagnostic analyte for the disease obviously depends on the pathogenetic mechanism that primarily prevails behind each individual case of PCOD. In conclusion, the pretreatment with MP A lowered significantly androgen and LH levels both in PCOD and non-pcod patients. Thus the samples for hormonal analyses used as an aid in diagnosing PCOD should be obtained in native state because progestagen treatment masks biochemical findings of PCOD. Acknowledgments. We are indebted to Katariina Korkeila, M.D., and Jaana Lepisto, M.D., for their help in collecting the data. RFRNCS 1. Hull MGR. pidemiology of infertility and polycystic ovarian disease: endocrinological and demographic studies. Gynecol ndocrinol 1987;1: Adams J, Polson DW, Franks S. Prevalence of polycystic ovaries in women with anovulation and hirsutism. Br Med J 1986;93: Adams J, Polson DW, Abdul Wahid N, Morris DV, Franks S, Mason HD, et al. Multifollicular ovaries: clinical and endocrine features and responses to pulsatile gonadotropin releasing hormone. Lancet 1985;1: Ardaens Y, Robert Y, Lemaitre L, Fossati P, Dewailly D. Polycystic ovarian disease: contributions of vaginal endosonography and reassessment of ultrasonic diagnosis. Fertil SteriI1991;55: Yen SSC. The polycystic ovary syndrome. Clin ndocrinol (Oxf) 198;1: Rebar R, Judd HL, Yen SSC, Rakoff J, Vandenberg G, Naftolin F. Characterization of the inappropriate gonadotropin secretion in polycystic ovary syndrome. J Clin Invest 1979;57: Lobo RA, Grager L, Gobelsmann U, Mishell DR Jr. levations of unbound serum estradiol as a possible mechanism Vol. 58, No.4, October 199 Anttila et al. MPA effect on hormones in oligomenorrhea 71

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