Endocrine (2013) 44:750–755 DOI 10.1007/s12020-013-9925-y
ORIGINAL ARTICLE
Macroprolactinemia in hyperprolactinemic infertile women Krithika Thirunavakkarasu • Pinaki Dutta • Subbiah Sridhar • Lakhbir Dhaliwal G. R. V. Prashad • Shalini Gainder • Naresh Sachdeva • Anil Bhansali
•
Received: 2 December 2012 / Accepted: 12 March 2013 / Published online: 30 March 2013 Ó Springer Science+Business Media New York 2013
Abstract Hyperprolactinemia occurs in 15–20 % of women with menstrual disturbances and 30–40 % of infertile women and it can adversely affect the fertility. High molecular weight prolactin (macroprolactin) has long been known in hyperprolactinemic fertile women. However, the prevalence of macroprolactinemia in hyperprolactinemic infertile women is not known. This cross-sectional study was carried out during the period of June 2010 and June 2011 at a single tertiary care centre. All women who attended the infertility clinic during this period were screened for hyperprolactinemia and only women with hyperprolactinemia and infertility were further studied for the presence of macroprolactin by polyethylene glycol precipitation assay. We compared the clinical, hormonal profile and fertility outcome of infertile women with true hyperprolactinemia and macroprolacinemia using appropriate statistical tests. Of 1,163 infertile women, 183 (15.7 %) had hyperprolactinemia [134 (73 %) had primary infertility and 49 (27 %) had secondary infertility]. Out of these 183 women with hyperprolactinemia, one had microadenoma, 161 had true idiopathic hyperprolactinemia and 21 (11.5 %) women had macroprolactinemia. The prevalence of oligomenorrhea and galactorrhea were significantly higher in patients with true hyperprolactinemia than macroprolactinemia (46 vs. 14 %, p \ 0.008 and 30 vs. 5 %,
K. Thirunavakkarasu L. Dhaliwal (&) G. R. V. Prashad S. Gainder Department of Obstetrics and Gynecology, Post Graduate Institute of Medical Education and Research (PGIMER), Chandigarh, India e-mail:
[email protected] P. Dutta S. Sridhar N. Sachdeva A. Bhansali Department of Endocrinology, Post Graduate Institute of Medical Education and Research (PGIMER), Chandigarh, India e-mail:
[email protected]
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p = 0.01 respectively). Twenty-two patients (13.5 %) of true hyperprolactinemia and two (9 %) in macroprolactinemia became pregnant during the study period. Prolactin measurement should be a part of routine evaluation of couples referred to infertility clinics. Macroprolactin screening is mandatory when clinical features and serum PRL assay results are conflicting. Patients with macroprolactinemia should be investigated for causes of infertility other than hyperprolactinemia. Keywords Hyperprolactinemia Macroprolactin Infertility Galactorrhea
Introduction Hyperprolactinemia is associated with suppression of hypothalamic–pituitary–gonadal axis through the inhibition of the pulsatile secretion of gonadotropin releasing hormone (GnRH) [1]. It is one of the most common endocrine disorders causing female infertility [2] and occurs in 30–40 % of infertile women [3]. Human PRL (PRL) circulates in three different forms in the blood. Monomeric PRL (23.5 kDa—free form), which accounts for 85 % of the PRL, dimeric PRL (50 kDa) contributes 5–10 % and in the remaining higher molecular weight ([100 kDa) form [4, 5] contributes a small and variable amount. The high molecular weight form (big–big PRL) consists of an antigen antibody complex of monomeric PRL and immunoglobulin (IgG) [6]. When the predominant form of circulating PRL is [100 kDa, the condition is termed as macroprolactinemia [7]. The above three forms of PRL are indistinguishable by routine assays and the excessive requirement of PRL determination led to an important laboratory pitfall [8]. The
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gel-filtration chromatography is the gold standard method to confirm the presence of big–big PRL, but is a costly and time consuming process [9]. Polyethylene glycol (PEG) has been employed to precipitate macroPRL, is an excellent screening method, allowing its detection rapidly and in an inexpensive way [10, 11]. The predominant molecular form recovered (i.e., assayed after precipitation) is the highly biologically active monomeric PRL. Macroprolactinemia is present when the recovery of monomeric PRL following treatment with PEG is less than 40 % [12]. Several clinical studies demonstrated that macroprolactinemia occurring in 10–42 % of all cases of hyperprolactinemic sera [13–15]. However, the prevalence of macroprolactinemia in hyperprolactinemic infertile women is not known and hence the present study was planned.
Research design and methods The study was conducted between June 2010 and June 2011 at the infertility clinic of the Department of Obstetrics and Gynaecology and Department of Endocrinology, Postgraduate Institute of Medical Education & Research, Chandigarh, India. The study was approved by Institute ethics committee. All patients provided written informed consent for participation in the trial. Infertility was defined as, failure to achieve a successful pregnancy after 12 months of regular unprotected intercourse in women \35 years and after 6 months in women[35 years [16]. All infertile women were worked up according to the WHO protocol (1992) which included screening for hyperprolactinemia at the initial visit. After excluding other causes of infertility like endometriosis, pelvic adhesions, and tubal abnormalities, only women with hyperprolactinemia and infertility were enrolled in the study. Eligibility criteria All newly referred infertile women in the age group of 21–40 years with serum PRL levels greater than 25 ng/ml (normal range 5–25 ng/ml) were included in the study. Women who were already under treatment for hyperprolactinemia, and/or hypothyroidism or had history of recent intake of metoclopramide, a-methyl dopa, ranitidine, chlorpromazine, risperidone, amitriptyline, fluoxetine, and haloperidol were excluded. The male partners of infertile women were investigated at initial visit for male infertility causes, and if found, those women were also excluded from the study. Study protocol All women in the study group underwent a comprehensive medical evaluation including detailed history (galactorrhea,
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menstrual irregularity, headache, visual disturbances, and drug intake) and physical examination. Only women with PRL levels more than 25 ng/ml on two occasions within 1 week interval were subjected to macroPRL screening. Women who had discrepant PRL values on two occasions (with increased and normal values) were subjected to reassessment of PRL and only if the two values were abnormal they were included. Magnetic resonance imaging (MRI) of the pituitary was done in all hyperprolactinemic patients with infertility. A baseline ultrasound was done to find out any associated gynecological abnormality. Transvaginal ultrasound was done between day 11 and 14 of the menstrual cycle to assess the size of the dominant follicle and endometrial thickness which was repeated after 2–3 days in the same cycle to look for follicular rupture. All hyperprolactinemic infertile women were treated with either cabergoline or bromocriptine if no spontaneous conception occurred after 3 months of follow-up. If the women remain anovulatory even after 3 months of treatment with cabergoline or bromocriptine, ovulation induction was done with clomiphene citrate. Hormonal profile Four millilitre of venous blood was collected in EDTA vacutainers twice at 20 min interval and pooled together between 0800 and 0900 h in the early follicular phase, either on day 2 or day 3 of patient’s menstrual cycle. Serum PRL (reference range (RR)—5.0 to 25 ng/ml; inter-assay CV, 3.1 to 5.0 %; intra-assay CV, 2.5 to 4.0 %), luteinizing hormone (LH), RR—1.7 to 8.6 mIU/ml; (inter-assay CV, 2.4 to 5.2 %; intra-assay CV, 0.3 to 1.8 %), follicle stimulating hormone (FSH), RR—1.5 to 12.4 mIU/ml; (interassay CV, 2.6 to 5.3 %; intra-assay CV, 0.4 to 2.0 %), estradiol, RR—7.63 to 42.6 pg/ml; (inter-assay CV, 3.5 to 6.2 %; intra-assay CV, 2.5 to 5.7 %), thyroid stimulating hormone (TSH), RR—0.27 to 4.2 ulU/ml (inter-assay CV, 2.5 to 3.2 %; intra-assay CV, 0.5 to 1.5 %) were determined at initial visit by electro-chemiluminiscence immunoassay (ELECSYS-2010, Roche-diagnostics, Mannheim, Germany). MacroPRL was estimated by PEG precipitation test. For macroPRL estimation, equal volumes (200 ll) of a 25 % aqueous PEG solution and patients’ sera were mixed in the ratio of 1:1 and centrifuged for 30 min. After centrifugation, the supernatant containing monomeric PRL was subjected to electro-chemiluminiscence immunoassay method. PRL levels were measured before and after PEG precipitation. The results were expressed as PRL recovery which is defined as immunoreactive PRL after PEG precipitation 9 100/immunoreactive PRL before PEG precipitation. If PRL recovery was less than 40 %, it suggested the presence of macroPRL [12].
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Statistical analysis The statistical analysis was carried out using SPSS 15. All quantitative variables were expressed as mean ± SD unless specified. Normality of data was checked by Kolmogorov–Smirnnov test. Categorical data were compared using v2 test and continuous normally distributed data using independent t test and skewed data using Mann– Whitney U test. A p value of \ 0.05 was considered statistically significant.
Results Prevalence of macroprolactinemia A total of 1,163 women attended the infertility clinic during the study period of 1 year; of these 183 had hyperprolactinemia (15.7 %). The mean age of women with hyperprolactinemia was 28.7 ± 4.6 years (range; 20–40 years). One hundred thirty-four women (73 %) had primary infertility while 49 (27 %) had secondary infertility. The duration of infertility ranged from 12 to 240 months, with mean duration of infertility being 58.7 ± 43.7 months. One hundred and three (56 %) patients with hyperprolactinemia were infertile for less than 4 years, 61 patients (33 %) were infertile for 4–8 years and for the remaining 19 patients (11 %) the duration of infertility was[8 years. In those with secondary infertility the mean duration was 51.4 months (range; 24–98 months, median 72). Of 183 women with hyperprolactinemia, macroprolactinemia was detected in 21 (11.5 %). Overall prevalence of macroprolactinemia in infertile women was 1.8 %, but it was higher among those who had hyperprolactinemia (11.5 %, p = 0.001). Only one of the total 183 hyperPRLemic women had evidence of micro adenoma in MRI and the rest were labeled as idiopathic hyperprolactinemia.
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(p = 0.01, Fig. 2). Most of the patients with macroprolactinemia had regular menstrual cycles, whereas regular cycles were present only in 50 % of patients with true hyperprolactinemia (86 vs. 50 %, p = 0.002, Table 1; Fig. 1). No significant differences in the hormonal levels (TSH, LH, and FSH) were found (Table 2). The prevalence of polycystic ovaries was comparable in both of the groups [(24 (15 %) vs. 3 (14 %), p = 1]. More than half of the patients (53 %) with true hyperprolactinemia did not show any evidence of ovulation on ultrasound, whereas in the macroprolactinemia group only four patients (19 %) had anovulation (p = 0.004). Two patients with true hyperprolactinemia had PRL level [100 ng/ml and one of them had a microadenoma on MRI. The remaining 182 patients were labeled as idiopathic hyperprolactinemia. In the two patients with PRL level
Fig. 1 Presenting complaints and associated conditions in patients with true hyperprolactinemia
Clinical characteristics of women with true hyperprolactinemia and macroprolactinemia The median serum PRL level in true hyperprolactinemia was 31.1 ng/ml (range; 25.1–475), whereas in macroprolactinemia group it was 46.3 ng/ml (range; 25.6–98; p = 0.7). The majority of women had PRL level between 25 and 50 ng/ml (88 % of true hyperprolactinemia and 70 % of macroprolactinemia). Serum PRL level [50 ng/ml was found in onethird (30 %) of patients with macroprolactinemia while only 19 (12 %) patients with true hyperprolactinemia had serum PRL levels [50 ng/ml, the difference between the PRL levels was significant (p = 0.001). Thirty percent of hyperprolactinemic women had galactorrhea (Fig 1), whereas only 5 % patients with macroprolactinemia had galactorrhea
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Fig. 2 Presenting complaints and associated conditions in patients with macroprolactinemia
Endocrine (2013) 44:750–755 Table 1 Showing comparison of symptoms and associated conditions in women with true hyperprolactinemia and macroprolactinemia
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Parameters
True hyperprolactinemia (n = 162)
Macroprolactinemia (n = 21)
p value
1.
Galactorrhea
49 (30 %)
1 (5 %)
0.01
2.
Menstrual pattern 81 (50 %)
18 (86 %)
0.002
Regular cycles
3.
Oligomenorrhea
74 (46 %)
3 (14 %)
0.008
Amenorrhea
7 (4 %)
–
0.605
86 (53 %)
4 (19 %)
0.004
76 (47 %)
17 (81 %)
0.004
Ovulation pattern Anovulation by ultrasound Ovulatory cycle
4.
5.
Table 2 Showing comparison of hormonal levels between true hyperprolactinemia and macroprolactinemia
Associated conditions Polycystic ovaries
24 (15 %)
3 (14 %)
1
Fibroid
8 (5 %)
1 (4.7 %)
1
Endometriosis
12 (7.4 %)
0
0.36
No pathology
118 (72.6 %)
17 (81.3)
0.5
Fertility outcome
22 (13.5 %)
2 (9 %)
0.9
Parameters
True hyperprolactinemia (n = 162)
Macroprolactinemia (n = 21)
p value
PRL levelsa
31.1 ng/ml
46.2 ng/ml
0.7
(Range)
(25.1–475)
(25.6–98)
Other hormonal profileb
a b
Median value Mean value
TSH (lIU/ml)
3.5 ± 0.7
2.9 ± 0.4
1
FSH (mIU/ml)
7.3 ± 1.4
6.1 ± 1.8
0.3
LH (mIU/ml)
8.9 ± 1.3
7.9 ± 1.2
0.8
Estradiol (pg/ml)
82.1 ± 12.1
58.7 ± 9.2
0.09
of [100 ng/ml, there was no evidence of overlap of simultaneous hyperprolactinemia and macroprolactinemia. None of the macroprolactinemic patients had neither PRL level more than 100 ng/ml nor MRI abnormalities. In total 24 patients became pregnant during the study period, 22 (13.5 %) with true hyperprolactinemia and two (9 %) in macroprolactinemia group. Eleven patients (6.8 %) with true hyperprolactinemia and one (4 %) of macroprolactinemia conceived without any treatment. Among the 12 patients who conceived spontaneously, 7 patients (58 %) had regular cycles and evidence of ovulation was observed in ultrasound between day 12 and day 14 of menstrual cycle. One of the macroprolactinemic patients and the 11 hyperprolactinemic patients conceived following cabergoline/bromocriptine and/or clomiphene citrate treatment. Eight (5 %) patients in true hyperprolactinemia group and one in macroPRL group conceived following clomiphene citrate therapy. The mean PRL level in hyperprolactinemic patients who conceived spontaneously was 40 ng/ml (range 26–52).
Discussion In the present study, we have examined the prevalence of macroprolactinemia in infertile women. It showed that 16 % of infertile women are hyperprolactinemic and 12 % of these women had macroprolactinemia. The prevalence of oligomenorrhea and galactorrhea was significantly higher in patients with true hyperprolactinemia than in patients with macroprolactinemia. Macroprolactin is a complex of PRL and IgG that may account for a significant proportion of idiopathic hyperprolactinemia because of the slow clearance rate rather than increased production [17]. From various studies, the estimated prevalence of macroprolactinemia in unselected women of reproductive age group is 0.3–3.68 % [18, 19] and in the selected sub group of prolactinoma patients, the prevalence is 10–42 % [15, 20, 21]. In the current study in infertile women with hyperprolactinemia the prevalence of macroprolactinemia was 16 %. Hyperprolactinemia is present in 15–20 % of women with infertility [1], and our
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study confer this observation. These data suggest that this condition should be considered as an important cause of infertility in women of reproductive age group. The clinical significance of macroprolactinemia is controversial. While there are some reports of macroprolactinemia have an association with galactorrhea, menstrual irregularities, and anovulation [7, 14, 15], few other studies have suggested macroprolactinemia to be asymptomatic [22, 23]. Lower biologic activity of macroprolactinoma and its bigger size (150 kDa), restrict it crossing from vascular endothelium, has been suggested as the reason for the lack of symptoms typical of hyperprolactinemia [14]. This has been supported by in vitro studies comparing the bioactivity of monomeric PRL with macroPRL in the NB2 lymphoma cell bioassay [14]. The incidence of galactorrhea in macroprolactinemic infertile women was 5 % in the present study compared to 39 % in a study by Isik et al. [14], 0.5 % by Leslie et al. [15], and 20 % in Donadio et al. [7]. The prevalence of oligomenorrhea/amenorrhea was only 14 % in patients with macroprolactinemia while compared to 50 % of women with true hyperprolactinemia, which is in consonance with other studies [15]. The possible mechanism of menstrual irregularities in patients with macroprolactinemia is intermittent dissociation of monomeric PRL from the low affinity, high capacity IgG antibody, may be a contributory factor to the development of symptoms of PRL excess [19]. On the other hand this could be due to associated polycystic ovary syndrome (PCOS) or merely coincidental findings that were not attributable to macroprolactinemia. In the previous studies, symptoms related to PRL excess were found in about 40–50 % of individual from the macroprolactinemia group and in 80–90 % of patients with monomeric hperprolactinemia [7, 20]. Circumstances leading to the diagnosis of macroprolactinemia included menstrual disorders (39 %), infertility (29 %), and galactorrhea (46 %) [7]. Although oligomenorrhea/amenorrhea and galactorrhea were more common in patients with true hyperprolactinemia, macroprolactinemic patients could not be differentiated from true hyperprolactinemic patients on the basis of clinical features alone[14]. The frequency of pituitary microadenoma in two large unselected series of patient comprising macroprolactinemia is 2–26 % [15, 21] although none of them were proven by histology as prolactinomas. In the present study only one out of 183 patients had pituitary microadenoma and rest all were having idiopathic hyperprolactinemia, which is too low and could be due to selection bias. The overall fertility rate in our study is 13.1 % and in macroprolactinemic patients it is 9 %. In a previous study by Vallette et al. [20] 68 % of the patients had preserved fertility in macroprolactinemic patients with uneventful pregnancies. In the present study selected sub group of patients with infertility were taken
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while in the above study unselected women with macroprolactinemia were taken. The duration of follow up also was short (1 year) to assess the fertility prospects. .Macroprolactinemia may be associated with other causes of hyperprolactinemia like hypothyroidism and drugs (metoclopramide, a-methyl dopa, ranitidine, chlorpromazine, risperidone, amitriptyline, fluoxetine, and haloperidol). However, we excluded the possible secondary causes in our study. The present study shows the usefulness of macroprolactin screening in infertile women with hyperprolactinemia, as also recommended by Pituitary Society guidelines [24]. The strengths of the present study include first study of macroprolactinemia in the context of infertility, involving large cohort of homogenous study population and optimal assessment of other causes of infertility (including MRI) have been evaluated as they also may be contributory for macroprolactinemia, while the limitations were many patients could not be followed for their pregnancy outcome since the study duration was short and macroprolactin level was estimated by PEG (Roche assay) instead of gel-filtration chromatography. The limitation of the previously published series was small number of subjects in addition to different study population including patients of prolactinoma. In conclusion, PRL measurement should be a part of routine evaluation of couples referred to infertility clinics. Macroprolactin screening is mandatory when clinical features and serum prolactin assay results are conflicting. Patients with macroprolactinemia should be investigated for causes of infertility other than hyperprolactinemia. Acknowledgments We are thankful to Miss Pooja Kundal for manuscript typing and edition. Conflict of interest
No conflict of interest to disclose.
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