Prolactin modulates peripheral androgen metabolism*

Size: px

Start display at page:

Download "Prolactin modulates peripheral androgen metabolism*"

Isaac Ray
5 years ago
Views:

1 FERTILITY AND STERILITY Copyright 1986 The American Fertility Society Printed in USA, Prolactin modulates peripheral androgen metabolism* Paulo Serafini, M,D, t Rogerio A, Lobo, M,D,:j: Department of Obstetrics and Gynecology, University of Southern California (USC) School of Medicine and Women's Hospital, Los Angeles County/USC Medical Center, Los Angeles, California Although hyperprolactinemia may be associated with hyperandrogenism, if hirsutism develops, it is usually a mild form, This study was designed to investigate whether prolactin (PRL) modulates 5a-reductase activity (5a-RA), because 5a-RA is known to be a major factor influencing the manifestation of androgenicity, Compared with normal women, euprolactinemic hirsute and both hyperprolactinemic hirsute and nonhirsute women had elevated levels of unbound testosterone (ut), Serum 3a-androstanediol glucuronide (3a-diol-G) was elevated only in patients who were hirsute, and serum 3a-diol-GluT ratios were elevated in euprolactinemic hirsute patients and normal in hyperprolactinemic hirsute patients, Genital skin 5a-RA in vitro was elevated only in euprolactinemic hirsute women, The previously recognized positive correlation between 5a-RA and the severity of hirsutism was dissociated with hyperprolactinemia. Human PRL incubated in vitro with normal genital skin also inhibited 5a-RA. These data suggest that PRL modulates 5a-RA and peripheral androgen metabolism and that other factors may also be involved in the evolution of hirsutism in hyperprolactinemia. Fertil Steril45:41, 1986 Since the original observation by Forbes et al., 1 hyperprolactinemia has been linked with hyperandrogen ism and hirsutism in some women. Although multiple androgenic abnormalities have been associated with hyperprolactinemia,2-8 the mechanism for this relationship has not been determined. Recent studies 7, 8 have demonstrated the occurrence of elevated levels of free testosterone (T) or unbound T (ut) in hyperprolactinemic Received July 15, 1985; revised and accepted September 26, *Supported in part by grants HD A1 and RR-43 from the National Institutes of Health. tpresent address: Department of Obstetrics and Gynecology, University of California, Irvine, Orange, California. *Reprint requests: Rogerio A. Lobo, M.D., Division of Reproductive Endocrinology, Women's Hospital, Room 1M2, 124 North Mission Road, Los Angeles, California 933. women. Yet, despite this abnormality, not all hyperprolactinemic women develop hirsutism. Because it appears that tissue 5a-reductase activity (5a-RA) is extremely important in the manifestation of hirsutism in women,9-12 we postulated that hyperprolactinemia might modulate 5a-RA. Suggestive evidence for this association was our findings of decreased serum T/dihydrotestosterone (DHT) and ut/unbound DHT ratios in hyperprolactinemic women 7 and the similar effects of altered T/DHT formation in normal men given sulpiride 5 or bromocriptine. 6 In this study, we wished to extend these studies by investigating the influence of prolactin (PRL) on peripheral androgen metabolism and 5a-RA. We used measurements of serum 3a-androstane" diol glucuronide (3a-diol-G) as well as the ratio of 3a-diol-G/uT and studied in vitro skin 5a-RA in hyperprolactinemic patients. Finally, we assessed Serafini and Lobo Peripheral androgen metabolism 41

2 the effect of PRL on 5u-RA directly by adding PRL to incubations of skin in vitro. SUBJECTS MATERIALS AND METHODS Twenty-three nonhirsute euprolactinemic women (NW), 28 to 45 years of age with a Ferriman-Gallwey (FG) score13 of 5.2 ±.1 standard error (SE), were selected as controls for the evaluation of 5u-RA,11 as were 2 hirsute euprolactinemic women (HW), 2 to 42 years of age with FG scores averaging 2.3 ±.8. The HW had mildly elevated androgen levels and no thyroid or adrenal disorders and were not virilized. Although these patients did not all meet strict criteria for the diagnosis of polycystic ovary syndrome, their clinical presentation, which included oligomenorrhea, was most compatible with this diagnosis. Five hyperprolactinemic hirsute women (HP-HW) 24 to 32 years of age with FG scores averaging 14.2 ±.3 and eight hyperprolactinemic nonhirsute women (HP-NW) with FG scores averaging 4.7 ±.4 participated in this study after giving informed consent. Hyperprolactinemia was defined as two or more measurements of serum PRL > 2 ng/ml. The mean serum PRL in the HP-NW was 19 ± 2 ng/ml and in the HP-HW was 48 ± 6 ng/ml. Among the eight HP-NW patients, three had radiographic findings consistent with the diagnosis of microadenoma; in the HP-HW group, two patients were found to have a microadenoma. The HP-HW group were also oligomenorrheic. They were similar in age and weight to the other HW but were less severely hirsute (1.2 ±.3 versus 2.3 ±.8, P <.5). In all patients, genital skin was obtained from the inferior margin of the labia majora. In NW, skin was obtained during the course of vaginal surgery for,benign disease ll and in HW and HP HW by outpatient biopsy procedure under local anesthesia. Obtaining biopsy specimens from HP NW could not be justified and was not carried out for ethical considerations. METHODS Blood was obtained from all subjects between 8: A.M. and 11: A.M. Serum was stored at - 2 C until it was analyzed for T, ut, 3u-diol-G, dehydroepiandrosterone sulfate (DHEA-S), and PRL by radioimmunoassay.3, Serafini and Lobo Peripheral androgen metabolism Genital skin biopsies were assayed for 5u-RA with minces incubated in RPMI-164 media (GIBCO Laboratories, Santa Clara, CA) with 14C_ T in the presence of 95% O 2, 5% CO 2 for 2 hours as previously describedy In brief, the skin specimens were placed in a chilled container with RPMI-164 medium, dissected free of subcutaneous fat and hair follicles, and minced at 4 C. Four nanomoles of 14C_ T were added to fresh media, and the minces were incubated in a Dubbnoff incubator (Precision Scientific Company, Chicago, IL) at 37 C. After the incubation, steroids were extracted with ether and separated by chromatography. Radioactivity in specific eluates (DHT, androstenedione [~4A], and 3u-diol) were determined. The conversion ratios of 14C_T to DHT, ~4A, and 3u-diol obtained in 2-hour incubations, based on 2 mg of skin, were used as indices of skin 5u-RA (T/DHT) and of 5u-keto and 3u-keto reductase activities (T/3u-diol) and 17-keto pathway (T/~4A). EFFECT OF PROLACTIN IN VITRO The effect ofprl on in vitro 5u-RA was evaluated in normal human genital skin obtained from nine additional NW. Skin minces from each subject were divided into 2- to 7-mg portions for separate incubations with 14C_T without and with increasing concentrations of PRL (1 to 14 ng/2 ml of media). Highly purified human PRL was generously supplied by Dr. Salvatore Raiti (National Institute of Arthritis, Diabetes, and Digestive and Kidney Diseases; PRL AFP 2284 C-2). All data were analyzed by Student's t-test, paired t-test, and Mann-Whitney U-test. Linear regression analysis was calculated by the method of least squares. RESULTS Mean serum T levels were elevated only for the hirsute patients. They were significantly greater in HW and HP-HW (53.9 ± 4 and 65.6 ± 3 ng/dl), compared with NW (26.1 ± 3.6 ng/dl; P <.1). Serum DHEA-S concentrations were also significantly greater in HW (2.7 ±.4 j.lg/ml) and HP HW (2.6 ±.2 j.lg/ml), compared with NW (1.24 ±.27 j.lg/ml; P <.1). Serum T and DHEA-S levels were comparable in the two hirsute groups. Both hirsute groups (HW and HP-HW), as well as the HP-NW who had normal total T levels, had Fertility and Sterility

3 ns/dl 14 UNBOUND T * pco.os between the FG score, serum androgen levels, and skin enzymatic activities. The effect of increasing concentrations of PRL on decreasing the normal CR of T to DHT from normal skin is depicted in Figure 6. When minces of genital skin were incubated repeatedly with media alone, the coefficient of variation for the CRs of T to DHT was found to be 9%.11 The percentage decrement in the CR of T to DHT after the addition of PRL, compared with two coefficients of variation of this CR, is depicted on Figure 6. A significant decrement in the CR occurred at PRL concentrations of 1 f-lg (P <.1), and a 5% decrease in the normal ratio occurred with 1 f-lg/incubation (P <.1). NW HW HP-NW HP-HW Figure 1 Mean ± SE serum ut levels in normal women (NW), hirsute women (HW), hyperprolactinemic normal (HP-NW) and hirsute women (HP-HW). Asterisks indicate the difference from NW. significantly higher ut levels, compared with NW (P <.5) (Fig. 1). The hyperprolactinemic groups had a two- to fivefold increase in ut, and these two levels were not statistically different. Serum 3a-diol-G was significantly elevated in the hirsute patients (HW and HP-HW, P <.5) and was normal in HP-NW (Fig. 2). We used the ratio of 3a-diol-G/uT indirectly to assess peripheral androgen activity and 5a-RA. 1O This ratio only was elevated in HW (58.1 ± 11.9, P <.1) (Fig. 3). Serum 3a-diol-G/uT ratios in NW, HP-NW, and HP-HW were similar (12 ± 2.7,19.7 ± 3.5, and 17.1 ± 2.7, respectively). HW had significantly higher conversion ratios (CRs) of T to DHT (P <.1), T to 3a-diol, and T to a 4 A (P <.5), compared with both NW and HP-HW (Fig. 4). The CRs of T to DHT were similar in HP-HW and NW, as were CRs of T to 3a-diol and T to a 4 A. These ratios were elevated in HW (Fig. 4). That a positive correlation exists between 5a-RA and the FG score in NW and HW has been documented. 12 However, the values for the CR of T to DHT and the FG score in HP-HW did not correlate and are dissociated from this previously calculated relationship12 (Fig. 5). In this group (HP-HW), no correlations could be demonstrated DISCUSSION The observation of an elevated serum ut in hyperprolactinemic patients confirms previous reports from our laboratory 7 and others. 8 The levels of ut, which reflect the biologically active moiety of T, were equally high in HP-NW and HW. These data suggest that for the manifestation of hirsutism, other factors are important, such as the metabolism of T in peripheral tissues. Further, our data support the notion that PRL may also play a modulatory role in this process. SERUM 3a DIOL-G NW HW HP-NW HP-HW * * pc.5 Figure 2 Mean ± SE serum 3a-diol-G levels in normal women (NW), hirsute women (HW), hyperprolactinemic normal (HP-NW) and hirsute women (HP-HW). Asterisks indicate the difference from NW. Serafini and Lobo Peripheral androgen metabolism 43

4 6 SERUM 3a DIOL-G/UNBOUND T * * pc.1 NW HW HP-NW HP-HW Figure 3 Mean ± SE serum 3a-diol-G/uT ratios in normal women (NW), hirsute women (HW), hyperprolactinemic normal (HP NW) and hirsute women (HP-HW). Asterisks indicate the difference from NW. Serum 3a-diol-G is a marker of peripheral androgen metabolism and of 5a-RA. Because serum 3a-diol-G was elevated only in the hirsute groups (HW and HP-HW), peripheral androgen metabolism of secreted androgens appears to be an important factor for the occurrence of hirsutism. The degree of hirsutism by the FG score was less in HP-HW than in HW with normal PRL levels. In using the ratio of serum 3a-diol-G/uT to assess tissue 5a-RA indirectly, we found that this ratio was normal in HP-NW. Direct assessment of genital skin 5a-RA suggested that the CR oft to DHT and oft to 3a-diol was abnormally elevated only in the euprolactinemic HW and was normal in HP-HW. The elevated conversion of T to!::j.4a in HW also confirmed our previous observation ll that hirsutism is associated with accelerated androgen metabolism in general. However, because this 17-keto pathway was normal in HP-HW, we suggest that hyperprolactinemia is not associated with altered androgen metabolism. Although direct assessment of genital skin 5a-RA from HP-NW would be extremely interesting, we could not justify obtaining biopsy specimens from these patients. We speculate that 5a-RA would be extremely low in this group. We previously demonstrated a close relationship between hirsutism (FG score) and 5a-RA. 12 Because HP-HW had less severe hirsutism than the euprolactinemic hirsute group, we plotted their FG scores and 5a-RA along with those we reported previously (Fig. 6). We observed that the relationship between the FG score and 5a-RA was somewhat dissociated in the HP-HW group. HP-HW had normal 5a-RA as well as serum 3adiol-G/uT ratios, despite elevated ut levels. Thus these findings suggest that with hyperprolactinemia, a more attenuated form of hirsutism may result and that 5a-RA and peripheral androgen metabolism appear to be normal. Because genital skin 5a-RA and serum 3a-diol-G/uT ratios appear to be normal despite the presence of hirsutism, albeit in a milder form, the development of hirsutism in hyperprolactinemic patients may involve other mechanisms as well. Nevertheless, taken together, these findings suggest that PRL modulates 5a-RA. In evaluating the effect ofprl on normal genital skin, we found a significant decrease in 5a-RA with PRL concentrations of 1 /-Lg/incubation. This response was noted to occur in vitro, but the mechanisms remain illusive. It is unclear whether this is mediated through a receptor or by other indirect means. The latter possibility appears to be more plausible. These findings, how N.. 1!IIi C.. > c u 5 ** * pc.5 ** pc.1 Figure 4 Mean ± SE ers oft to DHT, 3a-diol, and 114A in NW, HW, and HP-HW. Asterisks indicate significant changes, compared with NW. -= NW HWHP-HW NW HWHP-HW NW HWHP-HW T- DHT T- 3a diol T-.6.4 A 44 Serafini and Lobo Peripheral androgen metabolism Fertility and Sterility

5 It 32 % 24 t ~ iii III: > Z 8 u x x 8 Normal women Xx H irsut. women X X f PRL Hirsute I I I I I FERRIMAN GALLWEY SCORE Figure 5 Correlation of the conversion oft to DHT (5a-RA) with the FG score in NW and HW, as previously described (r =.66, P <.1). Added to this correlation figure and not included in the calculations for the regression line are the dissociated values of hirsute patients with elevated ( t ) PRL levels, denoted by crosses. ever, complement our clinical data in skin and serum, which suggests the modulation between PRL and 5a-RA. Although the level of PRL showing inhibition by direct incubation experiments is higher than in PRL concentrations in the patients studied, these differences may be accounted for by the fact J: o t c o.. ~ II > C o ~ -4 1 c g -8 ~ u "" 1 **p<o.ol Concentration of PRL in n91 incubation Figure 6 Percentage change in the normal CR of T to DHT (5a-RA) with added concentrations of human PRL at concentrations ranging from 1 ng to 1... g/incubation. The shaded area represents two coefficients of variation for the in vitro assay. Asterisks indicate significant reduction in 5a-RA. that patients are exposed on a daily basis to PRL, whereas the in vitro experiment was carried out only over 2 hours. Thus, a constant lower concentration of PRL may be all that is necessary to modulate 5a-RA. The level of PRL was lower in the HP-HW, compared with the HP-NW, but all patients had documented hyperprolactinemia on repeated evaluation, and, although 5a-RA was normal in these groups, this activity was not different. Further, no statistical correlation could be found between the serum PRL level and skin 5a RA. These findings suggest further that although PRL appears to modulate 5a-RA, its effect appears to be indirect. This relationship may, in turn, be influenced by other factors. REFERENCES 1. Forbes A, Henremou PA, Griswold GC, Albright F: Syndrome characterized by galactorrhea, amenorrhea, and low urinary FSH: comparison with acromegaly and normal lactation. J Clin Endocrinol Metab 14:265, Vermeulen A, Suy E, Rubens R: Effect of prolactin on plasma DHEA(S) levels. J Clin Endocrinol Metab 44:1222, Lobo R, Kletzky, Kaptein E, Goebelsmann U: Prolactin modulation of dehydroepiandrosterone sulfate secretion. Am J Obstet Gynecol 138:632, Higuchi K, Nawata H, Maki T, Higashizima M, Kato K, Ibayashi H: Prolactin has a direct effect on adrenal androgen secretion. J Clin Endocrinol Metab 59:714, Magrini G, Ebiner J, Burckhardt P, Felber J: Study on the relationship between plasma prolactin levels and androgen metabolism in man. J Clin Endocrinol Metab 43:944, Lackritz RM, Bartke A: The effect of prolactin on androgen response to human chorionic gonadotropin in normal men. Fertil Steril 34:14, Lobo R, Kletzky : Normalization of androgen and sexhormone-binding globulin levels after treatment of hyperprolactinemia. J Clin Endocrinol Metab 56:562, Glickman SP, Rosenfield RL, Bergenstal RM, Helke J: Multiple androgenic abnormalities, including elevated free testosterone, in hyperprolactinemic women. J Clin Endocrinol Metab 55:251, Kuttenn F, Mowsowicz I, Schaison G, Mauvais-Jarvis P: Androgen production and skin metabolism in hirsutism. J Endocrinol 75:83, Lobo RA, Goebelsmann U, Horton R: Evidence for the importance of peripheral tissue events in the development of hirsutism in polycystic ovary syndrome. J Clin Endocrinol Metab 57:393, Serafini P, Ablan F, Lobo RA: 5a-reductase activity in the genital skin of hirsute women. J Clin Endocrinol Metab 6:349, Serafini P, Lobo RA: Increased 5a-reductase activity in idiopathic hirsutism. Fertil Steril 43:74, Ferriman D, Gallwey JD: Clinical assessment of body hair growth in women. J Clin Endocrinol Metab 21:144, 1961 Serafini and Lobo Peripheral androgen metabolism 45

6 14. Goebelsmann U, Arce JJ, Thorneycroft IH, Mishell DR Jr: Serum testosterone concentrations in women throughout the menstrual cycle and following hcg administration. Am J Obstet Gynecol119:445, Stumpf P, Nakamura R, Mishell DR Jr: Changes in the physiologically free pools of estradiol and testosterone during exposure to levonorgestrel. J Clin Endocrinol Metab 52:138, Morimoto I, Edmiston A, Hawks D, Horton R: Studies on the origin of androstanediol and androstenediol glucuronide in young and elderly men. J Clin Endocrinol Metab 52:772, Horton R, Endres D, Galmarini M: Ideal conditions for hydrolysis of androstane 3n 17[3 diol glucuronide in plasma. J Clin Endocrinol Metab 59:127, Serafini and Lobo Peripheral androgen metabolism Fertility and Sterility

Gynecology-endocrinology

Gynecology-endocrinology FERTILITY AND STERILITY Copyright (fj 1991 The American Fertility Society Vol. 55, No, 5, May 1991 Printed on acid-free paper in U.S.A. Serum androsterone conjugates differentiate