| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
Laboratoire de Biochimie et Hormonologie (P.P.), Parc Eurasanté; Service de Gynécologie Endocrinienne et de Médecine de la Reproduction (S.J., D.D.), Hôpital Jeanne de Flandre; and Service de Radiologie, Hôpital Jeanne de Flandre (Y.R.), Centre Hospitalier Régional Universitaire de Lille, F-59037 Lille, France
Address all correspondence and requests for reprints to: P. Pigny, Laboratoire de Biochimie et Hormonologie, Parc Eurasanté, Centre Hospitalier Régional Universitaire, 59037 Lille cedex, France. E-mail: p-pigny{at}chru-lille.fr
 |
Abstract |
|---|
 Top Abstract IntroductionPatients and MethodsResultsDiscussionReferences |
|---|
Objective: The aim of our study was to check whether anti-Müllerian hormone (AMH) measurement in the serum could be a surrogate for antral follicle count in the diagnostic criteria of PCOS.
Design, Setting, and Patients: Serum AMH was measured with a second-generation immunoassay in a cohort of 73 PCOS patients and 96 controls, and its diagnostic power was evaluated by receiver operating characteristic curves. PCOS was diagnosed according to the Rotterdam definition.
Results: Serum AMH levels were 3-fold higher in PCOS patients than in controls (81.6 vs. 33.5 pmol/liter; P < 0.001) and were significantly related to the follicle number in the two groups. The area under the receiver operating characteristic curve for the AMH assay was 0.851, indicating a good diagnostic potency. Setting the threshold at 60 pmol/liter offered the best compromise between specificity (92%) and sensitivity (67%).
Conclusions: The serum AMH level is an accurate marker of the ovarian early antral follicle number and offers a good diagnostic potency. In situations where accurate ultrasonographic data are not available, AMH could thus be used instead of the follicle count as a diagnostic criterion and incorporated as such in the Rotterdam definition of PCOS.
|
Introduction |
|---|
|
TopAbstractIntroductionPatients and MethodsResultsDiscussionReferences |
|---|
For many years, different combinations of clinical (irregular menstrual cycles, hirsutism, and acne), biological (elevated serum testosterone or androstenedione levels or increased LH/FSH ratio), and ultrasound (U/S) criteria have been proposed, with very little international consensus. Indeed, the conservative definition for PCOS that was issued from a conference held in the National Institutes of Health in 1990 (1) did not satisfy many authors because it omitted the U/S criteria. More recently, during another consensus conference held in Rotterdam in 2003 (2) it has been proposed to include in the definition of PCOS the U/S criteria that are considered at the present time as the most specific, namely an increased ovarian volume (>10 ml) and/or the presence of 12 or more follicles in each ovary measuring 2–9 mm (3). Indeed, using a threshold of 12 for the follicle number per ovary (FNPO), we showed that 75% of PCOS patients were diagnosed whereas 99% of the normal women were under this cut off value (4).
Many participants at the Rotterdam consensus conference felt that the introduction of those U/S criteria would help standardizing the diagnosis of PCOS worldwide. However, others are still reluctant, mainly because the counting of the FNPO may not be easy to obtain with sufficient reliability by every group, and also because it is still debated whether a FNPO greater than 12 is specific of polycystic ovaries (PCO) (5). For these reasons, we investigated in this study whether the assay of the anti-Müllerian hormone (AMH) could be a surrogate for the FNPO in the Rotterdam definition of PCOS. Indeed, increased AMH serum levels were found in PCOS patients, in close relationship with the excess of the 2- to 9-mm FNPO (6, 7). This was expected because this member of the TGFß family is produced by the granulosa cells of early developing preantral and small antral follicles (8), whose number is 2- to 3-fold in PCO compared with normal ovaries (4).
Therefore, providing it offers similar specificity and sensitivity, AMH would present itself as a biological marker for PCO that would be easier to obtain than FNPO and would avoid using U/S in some patients. We took the opportunity of having access to a new second-generation AMH ultrasensitive assay to check this hypothesis.
|
Patients and Methods |
|---|
|
TopAbstractIntroductionPatients and MethodsResultsDiscussionReferences |
|---|
Controls.
The control population consisted of 96 women who were referred to our department for in vitro fertilization because of tubal and/or male infertility. Exclusion criteria were a history of menstrual disturbances (i.e. cycle length either <25 d or >35 d), hirsutism, abnormal serum level of prolactin or androgens (i.e. serum testosterone above 0.7 ng/ml and/or 
4-androstenedione above 2.2 ng/ml), ovarian volume greater than 10 ml and FNPO greater than 12 at U/S, and hormonal treatment during the 3 months before the study.
Patients with PCOS.
Seventy-three women were enrolled in this study. According to the Rotterdam criteria (2), the diagnosis of PCOS was based on the association of at least two of the three following criteria: 1) ovulatory disturbance, mainly oligomenorrhea or amenorrhea, 2) hyperandrogenism (HA) as defined either clinically by hirsutism (modified Ferriman and Gallwey score > 6), or severe acne/seborrhea, and/or biologically by a testosterone serum level greater than 0.7 ng/ml and/or 4-androstenedione greater than 2.2 ng/ml, and 3) more than 12 follicles in the 2- to 9-mm range in each ovary at U/S and/or an ovarian volume higher than 10 ml. U/S examination was performed with a 7-MHz transvaginal transducer (Logic 400; General Electric, Milwaukee, WI). U/S measurements were taken in real time, according to a standardized protocol, as previously described (9). This study was approved by the Institutional Review Board of the University Hospital of Lille. All patients and controls gave an informed consent before their inclusion in this study.
Hormonal immunoassays
Blood sampling was performed in the early follicular phase, between d 2 and 7 after the last menstrual period both in PCOS patients and in controls. In PCOS patients, the last menstrual period was either spontaneous or induced by the administration of didrogesterone (10 mg/d for 7 d). Any patient with at least one follicle with a diameter greater than 9 mm or a serum estradiol level above 80 pg/ml was excluded from the study. Serum AMH levels were assessed using the second-generation enzyme immunoassay AMH-EIA (reference A16507) provided by Immunotech (a Beckman Coulter company from Marseille, France) according to the supplier’s instructions. A recombinant human AMH was used as a calibration standard to build a standard curve ranging from 0–150 pmol/liter. Intra- and interassay coefficients of variation were less than 12.3 and 14.2%, respectively. The functional sensitivity (i.e. the lowest concentration of AMH that could be measured with an interassay coefficient of variation less than 20%) is 2.50 pmol/liter (Taieb, J., and P. Pigny, unpublished data). LH and FSH were measured using chemiluminescent two-site immunoassays on a multiparameter system (Axsym; Abbott Laboratories, Rungis, France) whereas testosterone was measured in duplicate using a RIA (Coat-a-Count, total testosterone kit) provided by Diagnostic Products Corp. (La Garenne Colombes, France). Androstenedione was also measured by RIA using the active androstenedione kit provided by Diagnostic Systems Laboratories (Cergy-Pontoise, France).
Statistical methods
Receiver operating characteristic (ROC) curves were constructed to examine the diagnostic test performance, i.e. its capacity to discriminate between controls and patients with PCOS. Sensitivity (y-axis) against [1-specificity (x-axis)] was plotted at each threshold level, and the area under the curve (AUC) was computed by the nonparametric Wilcoxon test. AUC represents the probability of correctly identifying controls and patients with PCOS. A value of 0.5 means that the test result is no better than chance. Comparisons of two independent groups were performed using the Student t test for normally distributed variables and by Mann-Whitney test for variables not normally distributed. The Kruskal-Wallis nonparametric ANOVA was used for comparisons of AMH levels between subgroups of patients (n > 2). Significant relationships between AMH and the various parameters were evaluated by the nonparametric Spearman correlation coefficient. All statistical analyses were performed using Statview 4.5 (Abacus Concepts Inc., Berkeley, CA) and SPSS 11.5 (SPSS Inc., Chicago, IL).
|
Results |
|---|
|
TopAbstractIntroductionPatients and MethodsResultsDiscussionReferences |
|---|
. As expected, patients with PCOS had higher body mass index values and higher serum levels of LH, testosterone, and androstenedione than controls. There was a trend toward lower FSH values in PCOS patients than in controls. The mean serum AMH level was 3-fold higher in PCOS patients than in controls (P < 0.001) as already reported by ourselves with the previous first-generation assay (6). Likewise, the mean 2- to 9-mm FNPO at U/S was 3-fold higher in PCOS patients than in controls (P < 0.001). It was significantly related to the serum AMH levels, both in controls (r = 0.537; P < 0.01) and in patients (r = 0.274; P < 0.05). AMH was also positively related to the serum testosterone level in both groups (r = 0.225 and P < 0.03 in controls; r = 0.360 and P < 0.003 in patients; see Fig. 1). No significant correlation was observed between FSH and AMH levels in PCOS patients.
|
|
, the area under the ROC curve for AMH reached a value of 0.851 (0.796–0.905, 95% confidence interval). Several threshold values of serum AMH level were analyzed in terms of specificity and sensitivity from the ROC curve data. Table 2 showed that the best compromise between specificity (92%) and sensitivity (67%) was obtained with a cutoff value of 60 pmol/liter. This value is slightly higher than the 90th percentile of controls.
|
|
). As expected, the 2- to 9-mm FNPO followed the same trend in these three subgroups (ANOVA, P < 0.05), whereas age did not exert a statistically significant effect (Table 3).
|
). On the contrary, the 2- to 9-mm FNPO was not significantly different between HA-positive and HA-negative PCOS patients (Fig. 3). Therefore, these data suggest that HA is associated with higher serum AMH levels in PCOS patients with irregular cycles.
|
|
Discussion |
|---|
|
TopAbstractIntroductionPatients and MethodsResultsDiscussionReferences |
|---|
In this study, the diagnostic performance of AMH was assessed using the ROC curves. The AUC of serum AMH assay yielded a satisfying value of 0.851 (0.796–0.907), lower however than the one obtained with the 2- to 9-mm follicle count (0.937) in our previous study (4). With a cutoff value of 60 pmol/liter, the serum AMH level had a good specificity of 92% but a relatively poor sensitivity of 67%, meaning that a third of PCOS patients, as defined by the Rotterdam criteria, would be misdiagnosed if one used exclusively the measurement of serum AMH. With a sensitivity of 75% and a specificity of 100% for a cutoff value at 12 (4), the follicle count appears as a better diagnostic tool than AMH measurement, at least in our hands. However, it must be stressed that comparing both studies is somewhat artificial because the latter data were obtained in a different series of patients, using different inclusion criteria. Indeed, the comparison of the diagnostic performance between AMH and the 2- to 9-mm follicle count could not be done in the present study because the latter was used as inclusion and exclusion criteria (see Patients and Methods). Whatsoever, it can be proposed from the present data to perform an AMH assay in medical centers where the U/S count of small antral follicles is not available. Indeed the single measurement of AMH in the early follicular phase appeared to be a valuable surrogate for the FNPO and it could therefore be used in place of the U/S data in the Rotterdam definition.
However, it must be kept in mind that neither a high value of AMH nor a high FNPO is per se sufficient to ascertain PCOS. Both criteria need to be incorporated into the Rotterdam definition, and it indicates PCOS only if associated with HA and/or oligo-anovulation. In addition, as recommended by the Rotterdam consensus conference (2), it must be remembered that this classification can be used only after the step of exclusion of other etiologies. This is particularly important to allow distinguishing PCOS from other diseases that are accompanied by multifollicular ovaries at U/S (12), such as recovering anorexia nervosa or functional hypothalamic anovulation. The AMH assay is not helpful for solving this issue because we previously found increased serum AMH levels in functional hypothalamic anovulation patients (13). This emphasizes the need for a careful exclusion diagnosis lying mainly on clinical history, before using the Rotterdam definition including either the FNPO or the serum AMH level.
Lastly, we showed in this study that the serum AMH level was also strongly associated with the main phenotypic features of PCOS, namely ovulation disorder and HA. This is line with Laven et al. (7), who also reported a positive correlation between the cycle duration, the serum androgens levels, and AMH in PCOS patients. Our results also agree with those of La Marca et al. (14) showing that amenorrheic women with PCOS had a higher mean serum AMH level than their oligomenorrheic counterparts. Altogether, these studies suggest that higher AMH levels are observed in PCOS patients in whom the process of terminal follicle maturation is the most impaired. Interestingly, we observed the same trend with the FNPO, in agreement with data obtained by others on ovarian cortical biopsies (15). Moreover, we also showed that in patients with irregular cycles, the presence of HA was associated with a further increase of the AMH value, in agreement with the previous report of Eldar-Geva et al. (16). This lends support to our hypothesis that AMH may be involved in the ovulatory disorder of PCOS patients (17) and may have effects distinct from HA, putatively through inhibition of the FSH-dependant cyclic recruitment (8) and/or aromatase activity (18).
In conclusion, the assay of serum AMH in women with HA and/or oligo-anovulation could indicate to the clinicians the presence of PCO when reliable U/S is not available. Additional studies are warranted to confirm that AMH could also be a prognostic marker of the extent of the ovarian dysfunction in PCOS patients and, in particular, whether it can predict response to ovulation induction.
|
Acknowledgments |
|---|
|
Footnotes |
|---|
Abbreviations: AMH, Anti-Müllerian hormone; AUC, area under the curve; FNPO, follicle number per ovary; HA, hyperandrogenism; PCOS, polycystic ovary syndrome; ROC, receiver operating characteristic; U/S, ultrasound.
Received September 19, 2005.
Accepted December 12, 2005.
|
References |
|---|
|
TopAbstractIntroductionPatients and MethodsResultsDiscussionReferences |
|---|
This article has been cited by other articles:
 |
|
 |
|
  R. Homburg Androgen circle of polycystic ovary syndrome Hum. Reprod., July 1, 2009; 24(7): 1548 - 1555. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S.M. Carlsen, E. Vanky, and R. Fleming Anti-Mullerian hormone concentrations in androgen-suppressed women with polycystic ovary syndrome Hum. Reprod., July 1, 2009; 24(7): 1732 - 1738. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. La Marca, F.J. Broekmans, A. Volpe, B.C. Fauser, N.S. Macklon, and on behalf of the ESHRE Special Interest Group for Anti-Mullerian hormone (AMH): what do we still need to know? Hum. Reprod., June 11, 2009; (2009) dep210v1. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 E. Diamanti-Kandarakis, A. Piouka, S. Livadas, C. Piperi, I. Katsikis, A. G Papavassiliou, and D. Panidis Anti-mullerian hormone is associated with advanced glycosylated end products in lean women with polycystic ovary syndrome Eur. J. Endocrinol., May 1, 2009; 160(5): 847 - 853. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 P A Johnson, T R Kent, M E Urick, L S Trevino, and J R Giles Expression of anti-Mullerian hormone in hens selected for different ovulation rates Reproduction, May 1, 2009; 137(5): 857 - 863. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R.L. Thomson, J.D. Buckley, L.J. Moran, M. Noakes, P.M. Clifton, R.J. Norman, and G.D. Brinkworth The effect of weight loss on anti-Mullerian hormone levels in overweight and obese women with polycystic ovary syndrome and reproductive impairment Hum. Reprod., April 20, 2009; (2009) dep101v1. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. Piouka, D. Farmakiotis, I. Katsikis, D. Macut, S. Gerou, and D. Panidis Anti-Mullerian hormone levels reflect severity of PCOS but are negatively influenced by obesity: relationship with increased luteinizing hormone levels Am J Physiol Endocrinol Metab, February 1, 2009; 296(2): E238 - E243. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 C. Rico, S. Fabre, C. Medigue, N. d. Clemente, F. Clement, M. Bontoux, J.-L. Touze, M. Dupont, E. Briant, B. Remy, et al. Anti-Mullerian Hormone Is an Endocrine Marker of Ovarian Gonadotropin-Responsive Follicles and Can Help to Predict Superovulatory Responses in the Cow Biol Reprod, January 1, 2009; 80(1): 50 - 59. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. Catteau-Jonard, S. P. Jamin, A. Leclerc, J. Gonzales, D. Dewailly, and N. di Clemente Anti-Mullerian Hormone, Its Receptor, FSH Receptor, and Androgen Receptor Genes Are Overexpressed by Granulosa Cells from Stimulated Follicles in Women with Polycystic Ovary Syndrome J. Clin. Endocrinol. Metab., November 1, 2008; 93(11): 4456 - 4461. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Das, D.J. Gillott, E. Saridogan, and O. Djahanbakhch Anti-Mullerian hormone is increased in follicular fluid from unstimulated ovaries in women with polycystic ovary syndrome Hum. Reprod., September 1, 2008; 23(9): 2122 - 2126. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. Yding Andersen, M. Rosendahl, and A. G. Byskov Concentration of Anti-Mullerian Hormone and Inhibin-B in Relation to Steroids and Age in Follicular Fluid from Small Antral Human Follicles J. Clin. Endocrinol. Metab., June 1, 2008; 93(6): 2344 - 2349. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. Gnoth, A.N. Schuring, K. Friol, J. Tigges, P. Mallmann, and E. Godehardt Relevance of anti-Mullerian hormone measurement in a routine IVF program Hum. Reprod., June 1, 2008; 23(6): 1359 - 1365. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M.-J. Chen, W.-S. Yang, C.-L. Chen, M.-Y. Wu, Y.-S. Yang, and H.-N. Ho The relationship between anti-Mullerian hormone, androgen and insulin resistance on the number of antral follicles in women with polycystic ovary syndrome Hum. Reprod., April 1, 2008; 23(4): 952 - 957. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. Codner, G. Iniguez, C. Villarroel, P. Lopez, N. Soto, T. Sir-Petermann, F. Cassorla, and R. A. Rey Hormonal Profile in Women with Polycystic Ovarian Syndrome with or without Type 1 Diabetes Mellitus J. Clin. Endocrinol. Metab., December 1, 2007; 92(12): 4742 - 4746. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Catteau-Jonard, P. Pigny, A.-C. Reyss, C. Decanter, E. Poncelet, and D. Dewailly Changes in Serum Anti-Mullerian Hormone Level during Low-Dose Recombinant Follicular-Stimulating Hormone Therapy for Anovulation in Polycystic Ovary Syndrome J. Clin. Endocrinol. Metab., November 1, 2007; 92(11): 4138 - 4143. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. J. Moran, M. Noakes, P. M. Clifton, and R. J. Norman The Use of Anti-Mullerian Hormone in Predicting Menstrual Response after Weight Loss in Overweight Women with Polycystic Ovary Syndrome J. Clin. Endocrinol. Metab., October 1, 2007; 92(10): 3796 - 3802. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. Dewailly, S. Catteau-Jonard, A.-C. Reyss, C. Maunoury-Lefebvre, E. Poncelet, and P. Pigny The excess in 2-5 mm follicles seen at ovarian ultrasonography is tightly associated to the follicular arrest of the polycystic ovary syndrome Hum. Reprod., June 1, 2007; 22(6): 1562 - 1566. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. S. Wachs, M. S. Coffler, P. J. Malcom, and R. J. Chang Serum Anti-Mullerian Hormone Concentrations Are Not Altered by Acute Administration of Follicle Stimulating Hormone in Polycystic Ovary Syndrome and Normal Women J. Clin. Endocrinol. Metab., May 1, 2007; 92(5): 1871 - 1874. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M.L. Hendriks, J.C.F. Ket, P.G.A. Hompes, R. Homburg, and C.B. Lambalk Why does ovarian surgery in PCOS help? Insight into the endocrine implications of ovarian surgery for ovulation induction in polycystic ovary syndrome Hum. Reprod. Update, May 1, 2007; 13(3): 249 - 264. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 F. H. Thomas, E. E. Telfer, and H. M. Fraser Expression of Anti-Mullerian Hormone Protein during Early Follicular Development in the Primate Ovary in Vivo Is Influenced by Suppression of Gonadotropin Secretion and Inhibition of Vascular Endothelial Growth Factor Endocrinology, May 1, 2007; 148(5): 2273 - 2281. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. Pellatt, L. Hanna, M. Brincat, R. Galea, H. Brain, S. Whitehead, and H. Mason Granulosa Cell Production of Anti-Mullerian Hormone Is Increased in Polycystic Ovaries J. Clin. Endocrinol. Metab., January 1, 2007; 92(1): 240 - 245. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. Y. Andersen and A. G. Byskov Estradiol and Regulation of Anti-Mullerian Hormone, Inhibin-A, and Inhibin-B Secretion: Analysis of Small Antral and Preovulatory Human Follicles' Fluid J. Clin. Endocrinol. Metab., October 1, 2006; 91(10): 4064 - 4069. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. Sir-Petermann, E. Codner, M. Maliqueo, B. Echiburu, C. Hitschfeld, N. Crisosto, F. Perez-Bravo, S. E. Recabarren, and F. Cassorla Increased Anti-Mullerian Hormone Serum Concentrations in Prepubertal Daughters of Women with Polycystic Ovary Syndrome J. Clin. Endocrinol. Metab., August 1, 2006; 91(8): 3105 - 3109. [Abstract] [Full Text] [PDF]
|
|
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
| Endocrinology | Endocrine Reviews | J. Clin. End. & Metab. |
| Molecular Endocrinology | Recent Prog. Horm. Res. | All Endocrine Journals |
