Article
Follicular-fluid anti-Mullerian hormone
concentrations are predictive of assisted
reproduction outcome in PCOS patients
Dr Recai Pabuccu
Dr Recai Pabuccu obtained his medical degree in 1975 from Istanbul University Faculty of Medicine, Turkey. After he completed his obstetrics and gynaecology residency, he worked as a clinical fellow in Reproductive Endocrinology and Infertility at the University of British Colombia, Canada. He has published more than forty articles in the field of assisted reproductive technologies and reproductive surgery. Professor Pabuccu is Director of the Department of Obstetrics and Gynecology, Ufuk University, Faculty of Medicine.
Recai Pabuccu1, Cemil Kaya1,3, Gamze Sinem Cag˘lar1, Efser Oztas1, Hakan Satiroglu2
1Department of Obstetrics and Gynecology, Faculty of Medicine, Ufuk University, Ankara, Turkey;2Department of Obstetrics and Gynecology, Faculty of Medicine, Ankara University, Ankara, Turkey
3
Correspondence: e-mail: kayacemil000@yahoo.com
Abstract
Serum anti-Mullerian hormone (AMH) concentrations constitute a sensitive marker for ovarian ageing. In addition, concentrations of AMH in the follicular fluid constitute a useful marker of embryo implantation in assisted repro-duction cycles. The present study measured serum and follicular-fluid AMH concentrations on the day of oocyte retrieval. These data showed that clinical pregnancy rates (25.0, 34.1 and 42.1%, respectively, P < 0.001), embryo implantation rates (24.3, 35.0 and 44.4%, respectively, P < 0.001) and fertilization rates (59.2, 70.9 and 79.5%, respectively, P < 0.001) were markedly different among the low, moderate and high follicular-fluid AMH groups but not among the different serum AMH concentration groups. Follicular-fluid AMH concentrations were nega-tively correlated with follicular-fluid oestradiol concentrations. The results of this study suggest that follicular-fluid AMH concentration on the day of oocyte retrieval would appear to better reflect the reproductive outcome in PCOS patients undergoing assisted reproduction.
Keywords: AMH, assisted reproduction cycles, follicular fluid, PCOS
Introduction
Anti-Mullerian hormone (AMH), a member of the trans-forming growth factor-b superfamily, is derived specifically from the granulosa cells of early developing pre-antral and antral follicles (Weenen et al., 2004). Polycystic ovary syn-drome (PCOS) is the main cause of anovulatory infertility (Franks, 1995). The pathogenesis of PCOS remains largely unknown although recent studies have suggested that AMH may have a role to play in the ovarian follicular sta-tus in PCOS (Pigny et al., 2003; Laven et al., 2004).
In anovulatory women with PCOS, granulosa cell function is abnormal (Franks et al., 2002). Therefore, the abnormal-ity of granulosa cells in PCOS may influence oocyte or embryo quality (Franks et al., 2002; Mulders, 2003) and
thus, high or low follicular-fluid AMH and serum AMH concentrations may affect reproductive outcome in PCOS patients undergoing assisted reproduction. PCOS exhibits the same number of primordial follicles, whereas the num-ber of 2–5 mm follicles is increased, compared with nor-moovulatory controls (Webber et al., 2003). This suggests differences in the process of follicles leaving the pool of non-growing follicles or a decline of the rate with which growing follicles go into atresia (van der Meer et al., 1998). This aberration of follicular maturation seems to indicate an abnormal endocrine environment, but whether there is an intrinsic abnormality of ovarian folliculogenesis remains unclear. Previously,Stubbs et al. (2005) demon-strated that AMH protein expression is reduced during the initial stages of follicle development in PCOS and this situation in PCOS may accelerate the process of primordial
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follicle recruitment and this may contribute to more small antral follicules in PCOS because AMH seems to inhibit the initiation of human primordial follicle growth (Stubbs et al., 2005). Therefore, AMH may be involved in the recruitment of FSH-sensitive follicles in the early antral stage (Durlinger et al., 2001). The early antral follicle count has been shown to reliably predict the fertility potential of women (Reuss et al., 1996) and their responsiveness to ovarian stimulation (Chang et al., 1998; Laszlo et al., 2002).
To date, the possible relationship between AMH produc-tion by an individual follicle and its funcproduc-tional quality remains to be demonstrated in women with PCOS undergo-ing IVF or intracytoplasmic sperm injection (ICSI) cycles. Therefore, this study investigated the relationship of serum and follicular-fluid AMH concentrations on the day of oocyte retrieval and reproductive outcome in PCOS patients undergoing assisted reproduction.
Materials and methods
Patients
A total of 80 infertile PCOS patients, 21–35 years of age, were prospectively recruited for this study between August 2007 and August 2008. All patients provided an informed consent and this investigation was approved by the institu-tional review board. The diagnosis of PCOS was made when two or three of the following criteria existed, as proposed at the Rotterdam Consensus Meeting: (i) oligomenorrhoea or amenorrhoea; (ii) clinical hyperandrogenism and/or hyper-androgenaemia; and (iii) polycystic ovaries (Rotterdam ESHRE/ASRM-Sponsored PCOS Consensus Workshop Group, 2004). The presence of polycystic ovarian appear-ance was determined ultrasonographically (Balen et al., 2003). Oligomenorrhoea (cycle intervals >35 days), amenor-rhoea (absence of menstruation for 3 consecutive months) and luteal-phase progesterone measurements less than 4 ng/ml in women with regular menstrual cycles were consid-ered indicative of oligoovulation. The inclusion criteria included: patients 35 years old or younger with patent Fallo-pian tubes (evaluated by hysterosalpingography), no previ-ous IVF attempts and partners with normal semen parameters. The semen quality was assessed using the World Health Organization guidelines (WHO) (World Health Organization, 1999; Guzick et al., 2001) (motility and con-centration) and Kruger’s (morphology) criteria (Kruger et al., 1988). All of the patients met the inclusion criteria. Diagnoses of congenital adrenal hyperplasia, Cushing’s syn-drome, androgen-producing tumours, hyperprolactinaemia and thyroid dysfunction were all excluded. In agreement with the inclusion criteria, no patients suffering any other aetiology of infertility were enrolled. A total 80 infertile PCOS patients underwent 80 ovarian stimulation-ICSI cycles.
Cycle monitoring and protocol for ovarian
stimulation
All patients received a standard gonadotrophin-releasing hormone (GnRH) agonist regimen starting on day 21 of a
spontaneous menstrual cycle. Leuprolide acetate (Lucrin daily injection; Abbott, Istanbul, Turkey) was administered for 10–14 days until complete pituitary desensitization was documented. Gonadotrophin stimulation (recombinant FSH, rFSH; 150 IU) was initiated on the third day of sub-sequent withdrawal bleeding and at that time the dose of leuprolide acetate was decreased to 0.5 mg/day. Further rFSH doses were determined according to the standard cri-terion of follicular maturation, assessed by ultrasound and serum oestradiol measurements. Human chorionic gonado-trophin (HCG; 10,000 IU; Pregnyl; Organon, Netherlands) was administered when at least three follicles had reached a diameter of >18 mm. Oocyte retrieval was performed 36 h later under transvaginal ultrasound guidance and intrave-nous sedation. All patients received luteal-phase support of 600 mg/daily of vaginally administered micronized pro-gesterone (Progestan, Kocßak, Istanbul, Turkey) daily start-ing from the day after oocyte retrieval. Oocytes were examined for pronuclei (PN) score 16–18 h after ICSI and only those in which two pronuclei were visualized were con-sidered to have fertilized normally (Tesarik et al., 2000). ICSI was performed to all cases. Embryos were transferred on day 3 after oocyte retrieval. Depending on the woman’s age and the embryo quality one to three embryos were transferred. All embryos were transferred using an Edwards Wallace catheter (Simcare Ltd., UK). Embryo transfers were performed under transabdominal sonographic guid-ance. Biochemical pregnancy was established when serum b-HCG concentration was found >20 IU/l on day 12 after embryo transfer and clinical pregnancy was defined as the presence of a gestational sac with fetal heart beat on ultra-sound performed at 6 weeks after embryo transfer. The implantation rate was calculated as the number of gesta-tional sacs/number of embryos transferred 100.
Follicular fluid collection
To obtain the exact follicular-fluid AMH concentration within a single follicle and avoid contamination from blood or flush medium or mixed follicular fluid during oocyte retrieval, only the follicular fluid from the first retrieved fol-licle of bilateral ovaries was collected. Preference for each preovulatory follicle was given to those whose transvaginal accessibility was straightforward. This methodology was set to minimize the risk of blood contamination of follicular fluids. Rather than a pooling follicular study, this study was designed as an individual follicular study. In this study, the average AMH concentration of two samples as one sin-gle entry for that person were used from each patient. The two follicular-fluid samples (one from each ovary) from the same woman were aspirated and analysed. A total of 160 follicular-fluid samples were collected from 80 patients for analysis and the average value of the AMH concentration was used. The presence or absence of blood contamination was graded by visual inspection and samples that looked cloudy or blood stained were discarded. Meticulous care was taken to include only patients with uncontaminated samples. The follicular fluids contaminated with culture medium or blood was discarded. Follicular fluid was not collected from follicles <17 mm. The collected follicular flu-ids were processed by centrifugation at 3000 g for 15 min at 4°C to eliminate cellular elements and subsequently frozen
at 80°C for biochemical and hormonal analysis. All follic-ular-fluid assays for each patient were performed in dupli-cate. Time elapsed between follicular aspiration and follicular-fluid cryopreservation did not exceed 30 min.
Hormonal measurements in serum and
follicular fluid
Blood samples were obtained by venipuncture and pro-cessed within 1 h after withdrawal for AMH analyses in the morning just prior to oocyte retrieval. Serum was stored at 20°C and assayed for AMH. Serum and follicular-fluid AMH concentrations were determined using an ultrasensi-tive ELISA (AMH ELISA kit; Diagnostic System Labora-tories, Texas, USA). The coefficients of variability (CV) for AMH were functional sensitivity 0.2 ng/ml, intra-assay CV 4%, inter-assay CV 8%, and for FSH, functional sensitivity 0.1 mIU/ml, intra-assay CV 3% and inter-assay CV 5%. Results were expressed as ng/ml. Serum and follicular-fluid oestradiol concentrations were determined by an auto-mated chemiluminescence technique (Hitachi Elecsys 2010; Roche Diagnostics, Germany). The intraassay and inter-assay CV were 2.7% and 3%, respectively.
Statistical analysis
Data analysis was performed by using Statistical Package for Social Sciences (SPSS) for Windows, version 11.5 (SPSS Inc., USA). Continuous variables were assessed for nor-mally distribution using the Shapiro Wilk test. AMH con-centrations were divided into the following groups: 25th centile or below (low AMH, between the 25th and the 70th centiles (average AMH) or above the 70th centile (high AMH). Data were shown as median (25th–75th) centiles. The ovarian stimulation and clinical outcomes for the stud-ied population divided according to the <25th, 25–75th and >75th centile of serum and follicular-fluid AMH concentra-tions. Wilcoxon Sign Rank test was applied for intra-group comparisons. While mean differences among groups were compared by one-way analysis of variance (ANOVA), medians were tested by Kruskal–Wallis test. When the P value from the one-way ANOVA or Kruskal–Wallis test statistics were statistically significant, post-hoc Tukey or non-parametric multiple comparison tests, where appropri-ate, were used to establish which groups differed from which others. The degree of association between continuous variables was calculated by Spearman’s rho correlation coefficient. A P value less than 0.05 was considered statisti-cally significant.
Results
Table 1depicts the patient characteristics in the low, aver-age and high serum and follicular-fluid AMH groups. The concentrations of follicular-fluid AMH were higher than in serum (4.1 ± 1.2 versus 3.1 ± 0.9 ng/ml, P < 0.001). The mean ± SD value of follicular-fluid AMH was 4.1 ± 1.2 ng/ml and the range was 2.1–6.7 ng/ml with a median of 4.1 ng/ml. The mean ± SD value of serum AMH was 3.1 ± 0.9 ng/ml and the range was 1.5–4.9 ng/ ml with a median of 2.8 ng/ml. The women’s ages, body
mass indexes and menstrual cycle lengths (data not shown) were comparable in the two sets of AMH groups (serum and follicular fluid) and were not significantly correlated to either serum nor follicular-fluid AMH concentrations. The total dose of FSH used for maintaining dominant fol-licle growth was similar irrespective of AMH concentra-tions in the serum and follicular fluid on the day of oocyte retrieval. The ovarian stimulation and clinical out-comes for the studied population were divided according to the <25th, 50–75th and >75th centile of serum and follic-ular-fluid AMH. Cycle outcomes in the low, average and high serum and follicular-fluid AMH groups are presented in Table 2. As shown, the fertilization, implantation and clinical pregnancy rates per oocyte retrieval increased with an increase in follicular-fluid AMH concentration (P < 0.01, respectively) (Table 2). The day of oocyte retrieval serum AMH concentrations were positively and signifi-cantly correlated with follicular-fluid AMH concentrations. On the day of HCG administration, the mean size of the dominant follicle (data not shown) and serum oestradiol concentrations were comparable in the two sets of AMH groups and were not correlated with serum or follicular-fluid AMH concentrations.
On the day of oocyte retrieval, a negative correlation between follicular-fluid concentrations of AMH and oestra-diol was observed (r = 0.37; P < 0.0001). As expected, serum AMH concentrations (on the day of oocyte retrieval) and follicular-fluid AMH concentrations were correlated with early antral follicle counts on day 3 (r = 0.75, P < 0.0001; r = 0.71, P < 0.0001; and r = 0.29, P < 0.002, respectively). In addition, there were positive and signifi-cant correlations between follicular-fluid AMH concentra-tion and fertilizaconcentra-tion (r = 0.38; P < 0.001), implantaconcentra-tion (r = 0.59; P < 0.001) and clinical pregnancy rates (r = 0.34; P < 0.01) (Figure 1).
Discussion
In summary, the results of this study indicate that follicu-lar-fluid AMH concentration is a good predictor of fertil-ization, implantation and clinical pregnancy rates in PCOS patients during assisted reproduction. This is the first study concerning the role of serum and follicular-fluid AMH concentrations on the day of oocyte retrieval in reproductive outcome prediction in PCOS patients under-going assisted reproduction.
Previous studies showed that follicular fluid and serum of PCOS women contained increased AMH concentrations (Fallat et al., 1997; Cook et al., 2002; Pigny et al., 2003; La Marca et al., 2004; Laven et al., 2004; Eldar-Geva et al., 2005; Das et al., 2008). Initial studies suggested that the source of elevated serum AMH concentrations was due primarily to the increased number of small antral follicles, assuming that each follicle produces a normal amount of AMH (Cook et al., 2002; Pigny et al., 2003). In contrast, more recently,Das et al. (2008)have suggested an increased production of AMH by individual follicles and therefore the increase was not just due to the increased number of small antral follicles characteristically seen in PCOS. This suggests that actual AMH production by granulosa cells
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in anovulatory PCOS is significantly higher. The same authors suggested that the follicules in PCOS patients might have abnormal AMH production (Das et al., 2008). Therefore, PCOS granulosa cells may have an abnormal capacity to synthesize AMH in vivo. Oocytes and embryos are of poorer quality from patients with PCOS undergoing assisted reproduction (Pellicer et al., 1996; Cano et al., 1997; Ludwig et al., 1999; Franks et al., 2002; Plachot et al., 2003). Previously, a few studies reporting on IVF– ICSI cycles found a positive association between circulating
concentrations of AMH and fertilization rate, implantation rate and pregnancy outcome (Hazout et al., 2004). So far, the relationship between serum and follicular-fluid AMH concentrations on the day of oocyte retrieval and reproduc-tive outcome has not been specifically determined in PCOS patients undergoing IVF.
This study, in contrast to serum AMH concentrations on the day of oocyte retrieval, found a positive and statistically significant correlation between follicular-fluid AMH Table 1. Patient characteristics in the low (<25th), average (25th–75th) and high (>75th)
anti-Mullerian hormone groups.
Variable dOPU serum AMH (ng/ml) dOPU follicular-fluid AMH (ng/ml)
<2.58 2.58–3.87 >3.87 <3.18 3.18–4.75 >4.74 Age (years) 29.7 ± 4.1 29.2 ± 4.6 29.3 ± 3.9 30.8 ± 3.7 28.9 ± 4.4 29.0 ± 4.5 BMI (kg/m2) 24.9 ± 4.0 25.0 ± 3.5 25.7 ± 4.6 23.6 ± 3.4 24.4 ± 4.1 24.1 ± 4.2 FSH (IU/l) 5.3 ± 1.5 5.7 ± 1.4 5.6 ± 1.7 5.7 ± 1.7 5.4 ± 1.4 5.8 ± 1.6 LH (IU/l) 8.2 ± 1.8 8.7 ± 2.3 8.4 ± 2.9 8.2 ± 2.1 8.3 ± 2.4 9.0 ± 2.5 >14 mm follicle (n) 11.0 ± 1.8 11.1 ± 2.5 11.1 ± 1.7 10.8 ± 1.8 11.0 ± 2.2 11.4 ± 2.3 Total oocytes (n) 19.6 ± 5.1 20.2 ± 5.7 18.2 ± 5.0 19.7 ± 5.2 20.3 ± 4.7 19.3 ± 5.3 MII oocytes (n) 13.2 ± 2.7 17.3 ± 2.8 13.1 ± 3.1 13.2 ± 3.0 17.7 ± 4.1 13.3 ± 3.4 Total FSH (IU) 1989 ± 397 2086 ± 254 2088 ± 299 1991 ± 371 2014 ± 252 2121 ± 285 rFSH (IU)/day 11.1 ± 1.1 10.7 ± 1.3 10.4 ± 1.2 10.9 ± 1.1 10.9 ± 1.3 10.2 ± 1.3 >17 mm follicle (n) 7.7 ± 3.2 7.5 ± 2.1 8.0 ± 2.9 7.8 ± 2.9 7.8 ± 2.5 7.4 ± 2.5 Oocytes retrieved (n) 18.6 ± 5.3 17.9 ± 4.9 18.9 ± 5.0 17.2 ± 3.9 19.2 ± 5.8 17.4 ± 4.1 Oestradiol (pg/ml) on HCG day 2764 ± 478 3078 ± 521 3119 ± 626 2715 ± 404 3070 ± 518 3146 ± 638 Follicular-fluid oestradiol (pg/ml) on dOPU 2318 ± 790 1767 ± 733 1275 ± 489 2102 ± 840 1804 ± 677 1425 ± 806 Serum oestradiol (pg/ml) on dOPU 2524 ± 822 2319 ± 675 2400 ± 520 2330 ± 872 2309 ± 631 2439 ± 846 Serum progesterone pg/ml 13.7 ± 5.7 10.8 ± 3.3 7.6 ± 2.4 14.1 ± 5.5 10.7 ± 3.4 7.5 ± 2.0
Values are mean ± SD. dOPU = day of oocyte pick-up; AMH = anti-Mullerian hormone; BMI = body mass index; HCG = human chorionic gonadotrophin; MII = metaphase II; rFSH = recombinant FSH. Differences among groups are not statistically significant.
Table 2. Outcomes of assisted reproduction cycles in low (<25th), average (25th–75th) and high (>75th) anti-Mullerian hormone concentration groups.
Variable dOPU serum AMH (ng/ml)
a
dOPU follicular-fluid AMH (ng/ml)a 2.58 2.58–3.87 >3.87 <3.18 3.18–4.75 >4.74 Fertilization rate (%) 149/224 (66.5) 493/690 (71.4) 231/324 (71.3) 142/240 (59.2) 436/615 (70.9)c 272/342 (79.5)b,d Implantation rate (%) 12/35 (34.3) 16/45 (35.6) 17/46 (37.0) 9/37 (24.3) 21/60 (35.0)c 16/36 (44.4)b,d Clinical pregnancy rate (%) 6/19 (31.6) 12/37 (32.4) 9/24 (37.5) 5/20 (25.0) 14/41 (34.1)c 8/19 (42.1)b,d
AMH = anti-Mullerian hormone; dOPU = day of oocyte pick-up. Statistical significance was defined as P < 0.05.
a
Data are shown as low (<25th), median (25th–75th) and high (>75th) percentiles.
b
Overall P values determined by Kruskal–Wallis test.
c
Kruskal–Wallis multiple comparison test was used to know which group differ from which others: P < 0.001, low vs average.
d
Kruskal–Wallis multiple comparison test was used to know which group differ from which others: P 0.002 average vs high.
concentrations after FSH stimulation and fertilization rate. Fertilization rate was significantly higher in the highest fol-licular-fluid AMH concentrations than in the group with lowest concentrations. These results suggest that follicu-lar-fluid AMH concentrations on the day of oocyte retrie-val play a role in the maturation and development of oocytes in women with PCOS. AMH concentrations in fol-licular fluid might be indicative of oocyte quality. More recently, it has been demonstrated that the higher follicu-lar-fluid AMH concentrations on the day of oocyte retrieval positively correlated with successful oocyte fertil-ization and constitute a useful follicular marker of embryo implantation (Takahashi et al., 2008). In addition, clinical pregnancy rates and embryo implantation rates were signif-icantly higher in patients with follicles containing high AMH concentrations (Fanchin et al., 2007). Ebner et al. (2006)demonstrated an association between AMH concen-tration and the quality of the oocytes. In that study, the dark central granulation of the cytoplasm was the most dominant anomaly in the lowest AMH group. Oocyte mat-uration consists of two separate processes, nuclear and cytoplasmic maturation. Patients who received embryos derived from granular oocytes, following embryonic trans-fer, exhibited low pregnancy outcome. Therefore, AMH may affect cytoplasmic maturation of the oocyte.Lekamge (2007)also showed that low AMH may be associated with poor oocyte quality as supported by diminished fertilization rates.
Serum AMH on the day of oocyte retrieval seems to result from the follicular pool and its production is independent of the gonadotrophin-dependent indicators of ovarian reserve. The differential predictability of follicular-fluid ver-sus serum AMH concentrations on the day of oocyte retrieval may be explained by the fact that circulating AMH concen-trations reflect the growing follicular pool on day of oocyte retrieval and are less effective to discriminate per-follicle AMH production. Therefore, high follicular-fluid AMH concentrations positively correlated with successful oocyte fertilization, which indicates that follicular-fluid AMH may play a more important role in the development of oocytes and fertilization in PCOS patients undergoing assisted repro-duction. Follicular-fluid AMH concentrations were higher than serum concentrations which may indicate that AMH has a paracrine effect on the process of oocyte development in women with PCOS undergoing IVF or ICSI. Based on these data, the follicular-fluid AMH concentrations after stimulation by exogenous FSH did reflect fertilization that resulted in high-quality embryos in PCOS patients undergo-ing assisted reproduction.
The present study found that high follicular-fluid AMH concentrations correlated with high implantation and clini-cal pregnancy rates. Implantation and cliniclini-cal pregnancy rates were significantly higher in the highest follicular-fluid AMH concentration group compared with the group with the lowest concentrations. High follicular-fluid AMH con-centrations constitute a useful marker of embryo implanta-tion. That follicular-fluid AMH concentration on the day of oocyte retrieval may be related to embryo quality could contribute to an explanation for the high correlation with Figure 1. Correlation between follicular-fluid
anti-Mulle-rian hormone concentration and (a) fertilization, (b) implantation and (c) clinical pregnancy rates. CPR = clin-ical pregnancy rate; dOPU = day of oocyte pick-up;
pregnancy probability, because embryo quality is crucial for clinical success. These results confirm those found by
Silberstein et al. (2006). The same authors reported a signif-icant correlation between serum AMH concentrations on the day of oocyte retrieval and embryo morphology score. They concluded that AMH >2.7 ng/ml indicated good oocyte quality as reflected by a higher implantation rate and a trend towards a better clinical pregnancy rate. More recently,Fanchin et al. (2007)demonstrated that follicular-fluid AMH concentrations were positively correlated to embryo implantation and clinical pregnancy rates after IVF–embryo transfer. This study’s results also demon-strated a strong association between follicular-fluid AMH concentrations on the day of oocyte retrieval and implanta-tion and clinical pregnancy rates. Follicular fluids may play an important role in the endocrine balance of PCOS by its effect on the relation between theca and granulosa cells. An abnormal interaction between PCOS granulosa cells and their own follicular fluids may be implicated in the altered steroidal response of the granulosa (Teissier et al., 2000). Successful implantation depends on the synchronized devel-opment of both embryos and endometrium (Paulson et al., 1990). In this study, follicular AMH concentrations were negatively and significantly correlated with follicular oest-radiol concentrations. As concentrations of follicular AMH increased, linear decreases in follicular oestradiol concentrations were observed. In PCOS patients, aroma-tase activity may be decreased because follicles from PCOS do not produce large amounts of oestradiol. Exogenous AMH did inhibit the biosynthesis of aromatase in cultured rat granulosa cells (di Clemente et al., 1994). Therefore, it can be speculated AMH might constitute this follicular-fluid-derived inhibitory factor of oestradiol. This data con-firms that follicular AMH is influential on follicular oestra-diol in PCOS patients. These findings reveal that there is a direct relationship between follicular AMH concentrations and follicular oestradiol in patients with PCOS. Oocyte and embryo quality may also be influenced by serum oest-radiol secretion (Dor et al., 1992). There are inconsistent results related to the detrimental and beneficial effects of oestradiol in assisted reproduction cycles. Some investiga-tors have noted no adverse effects, while others demon-strated evidence of significant decreases in fertilization, implantation and pregnancy rates. Nevertheless, the rela-tionship between supraphysiological oestradiol concentra-tions and reproductive outcome has not yet been extensively investigated. Adverse effects of supraphysiolog-ical oestradiol concentrations may include alterations in both endometrial receptivity and oocyte/embryo quality. However, Valbuena et al. (2001)showed that high oestra-diol concentrations are deleterious to embryo adhesion in vitro, mainly because they have a direct toxic effect on the embryo that may occur at the cleavage stage. The effect of AMH on the oocyte and embryo may be mediated through oestradiol. For future studies, it would be interest-ing to explore the impact of follicular AMH concentration embryo implantation markers in assisted reproduction cycles. Another potential mechanism could be a direct AMH action on the developing embryo. Future studies focusing on the association between follicular-fluid or serum AMH and embryo implantation may help to clarify this matter further. Therefore, it can be speculated that
higher AMH concentrations lead to decreases in follicular oestradiol which can improve fertilization and implantation rates in PCOS assisted reproduction cycles.
In conclusion, the present study found that fertilization, implantation and clinical pregnancy rates were significantly higher in the highest follicular-fluid AMH concentration group than in the group with the lowest concentrations in PCOS patients undergoing assisted reproduction. There-fore, the follicular-fluid AMH concentration on the day of oocyte retrieval would appear to better reflect the repro-ductive outcome in PCOS patients undergoing assisted reproduction.
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Received 20 November 2008; refereed 28 April 2009; accepted 3 June 2009.
