Objective
The goal of this work is to expand the usefulness of antimüllerian hormone (AMH) in predicting in vitro fertilization cycle outcome by demonstrating that AMH concentration obtained in an ongoing treatment cycle predicts both oocyte number and pregnancy.
Study Design
Serum samples were obtained from 190 in vitro fertilization patients at onset of follicle-stimulating hormone stimulation. These were analyzed retrospectively during a single cycle in which clinicians were blinded to the results. Our major outcome measures were the number of oocytes obtained and ongoing pregnancy.
Results
Patients with an initial AMH concentration of >3 ng/mL were found to produce a mean of 19.8 oocytes and had an ongoing pregnancy rate of 60.3%. In contrast, those with AMH values of ≤1 ng/mL yielded a mean of 6.2 oocytes and had an ongoing pregnancy rate of 23.4% ( P < .0001 for both).
Conclusion
Greater AMH serum concentration strongly predicts an increased number of oocytes and ongoing pregnancy ( P ≤ .0001).
Many clinicians use endocrine and/or ultrasound parameters to predict patient response to–and possibly to alter–stimulation during in vitro fertilization (IVF) treatment. Pretreatment baseline endocrine studies such as follicle-stimulating hormone (FSH), estradiol, inhibin B, antimüllerian hormone (AMH), and ultrasound measurements of antral follicle count, ovarian volume, and parameters of ovarian blood flow have been recommended as useful markers of potential cycle performance, as are functional endocrine assays such as the clomiphene challenge test. AMH is unique in this group of indicators in that its level is largely uninfluenced by menstrual cycle day and in that it is arguably the most powerful tool available for such use. All of these studies, however, have been used prior to the actual IVF cycle.
We have attempted to expand the usefulness of AMH for predicting cycle outcome by measuring the AMH concentration found in IVF patients during the actual cycle of stimulation after ovarian suppression with either a gonadotropin-releasing hormone (GNRH) agonist or combined estrogen/progestin tablets in the treatment cycle. We hypothesized that AMH measured during the cycle would predict both the number of oocytes retrieved and the probability of ongoing pregnancy. Accordingly, we have measured AMH levels initially and at the conclusion of FSH stimulation, ie, at the time of human chorionic gonadotropin (HCG) administration, 36 hours prior to oocyte retrieval and correlated these levels with cycle outcome.
Materials and Methods
We recruited 190 patients receiving FSH stimulation during a single IVF treatment cycle at the Women and Infants Hospital Center for Reproduction and Infertility (from October 2007 through February 2009) who consented to assessment of serum concentration of AMH in an institutional review board–approved protocol. No patients initiating IVF treatment were excluded. In all, 161 of the 190 patients had undergone prior down-regulation with luteal leuprolide acetate. A smaller group (n = 27) had been treated with oral contraceptive tablets for 15-20 days before starting stimulation with FSH; these patients received GNRH antagonist therapy when the lead follicle diameter averaged 13 mm and/or serum estradiol reached 250 pg/mL. Two patients were stimulated with leuprolide acetate and FSH in the so-called microdose flare protocol.
The usual dose of FSH used was 225 IU administered daily but adjustments to dosage were made based on history of response when known and on in-cycle observations when thought appropriate by the managing physician. HCG was administered also at the discretion of the physician, but usually when at least 2 follicles with mean diameter of 18 mm were observed. Embryo transfers were done at both cleavage and blastocyst stage, depending on the number and morphologic quality of embryos observed. Transfer number varied with age and followed guidelines recommended by the American Society for Reproductive Medicine. Physicians were unaware of the AMH values during treatment and therefore these values had no influence on outcome in the present study.
Patients were monitored by serial ultrasounds and serum estradiol measurements using the Immulite method (Diagnostic Products Corp, Los Angeles, CA). FSH was measured prior to cycle initiation by a chemiluminescent assay also obtained from Diagnostic Products Corp. Oocytes were aspirated by ultrasound guided transvaginal retrieval 36 hours after HCG administration and were then inseminated by standard IVF or by intracytoplasmic sperm injection. Ongoing pregnancy was documented by ultrasound done 5-6 weeks after embryo transfer demonstrating a fetus with a heartbeat. AMH concentration was measured on day 1 of FSH stimulation, utilizing an assay obtained from Diagnostic Systems Laboratories (DSL-14400; Webster, TX). A second AMH determination was obtained on the final day of FSH administration (the day of HCG injection), 36 hours before anticipated oocyte retrieval utilizing the same assay. The sensitivity of the assay is 0.1 ng/mL. At a level of 3 ng/mL, the interassay coefficient of variation was 5.2% and the intraassay coefficient of variation was 7.1%.
Primary outcome measures included the number of oocytes obtained at retrieval and achievement of an ongoing pregnancy (ultrasound demonstration of at least 1 normal fetal heart at 5-6 weeks after transfer). Demographic data on each patient including age, body mass index (BMI), and infertility diagnosis were recorded. Additional variables observed were peak levels of serum estradiol, prior basal FSH values, total quantity of FSH used for stimulation and details of stimulation protocol and dosage. All patients were studied in a single cycle only.
Statistical power calculations were based on the detection of modest effect sizes at a 2-sided 5% significance level. In all, 190 patients provided >90% power to detect a correlation of 0.25 between baseline AMH and oocyte count and a 0.25 SD difference in AMH by pregnancy status at the end of the cycle. The data were analyzed by χ 2 tests, correlation coefficients, and analysis of variance with Tukey correction for multiple comparisons, as appropriate. AMH values below the limit of sensitivity were analyzed as 0.1 ng/mL (n = 1 at onset and n = 3 at completion of ovarian stimulation). Continuous variables were examined graphically for deviations from normality. The number of oocytes retrieved appeared normally distributed, whereas both AMH measurements were skewed, including 1 undiagnosed patient with very high values at both time points. Therefore, AMH measurements were compared by the appropriate nonparametric statistics. Lowess curves (locally weighted smoothed scatterplots) showed a nonlinear association between baseline AMH and both outcomes that tapered off >3 ng/mL. For this reason, AMH was categorized as <1, 1-3, and >3 ng/mL–approximately the tertiles of the distribution–for the primary analyses. Adjusted estimates for AMH groups were calculated by analysis of covariance for oocyte count and multiple logistic regression for achievement of pregnancy.
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