Measurement of serum anti-Müllerian hormone by revised Gen II or automated assay: Reproducibility under various blood/serum storage conditions.

OBJECTIVE: We investigated the agreement between anti-Müllerian hormone (AMH) levels measured with revised Gen II (rev-Gen II) and automated AMH (Access) assays and evaluated the reproducibility of each method under various blood/serum storage conditions. METHODS: AMH levels in blood samples from 74 volunteers were measured by rev-Gen II and Access assays under various conditions: immediate serum separation and AMH measurement (fresh control); serum stored at -20 °C and AMH measured after 48 hours, 1 week, and 2 years; serum stored at 0 to 4 °C and AMH measured after 48 hours and 1 week; and blood kept at room temperature and delayed serum separation after 48 hours and 1 week, with immediate AMH measurement. RESULTS: In fresh controls, all rev-Gen II-AMH values were higher than comparable Access-AMH values (difference, 8.3% to 19.7%). AMH levels measured with the two methods were strongly correlated for all sample conditions (r=0.977 to 0.995, all p<0.001). For sera stored at -20 °C or 0 to 4 °C for 48 hours, Access-AMH values were comparable to control measurements, but rev-Gen II-AMH values were significantly lower. AMH levels in sera stored at -20 °C or 0 to 4 °C for 1 week were significantly lower than in fresh controls, irrespective of method. Across methods, long-term storage at -20 °C for 2 years yielded AMH measurements significantly higher than control values. When serum separation was delayed, rev-Gen II-AMH values were significantly lower than control measurements, but Access-AMH values varied. CONCLUSION: The rev-Gen II and Access-AMH assays showed varying reproducibility across blood/serum storage conditions, but automated Access yielded superior stability to rev-Gen II.

Expression of the genes for peroxisome proliferator-activated receptor-γ, cyclooxygenase-2, and proinflammatory cytokines in granulosa cells from women with polycystic ovary syndrome.

OBJECTIVE: To identify differences in the expression of the genes for peroxisome proliferator-activated receptor (PPAR)-γ, cyclooxygenase (COX)-2, and the proinflammatory cytokines interleukin (IL)-6 and tumor necrosis factor (TNF)-α in granulosa cells (GCs) from polycystic ovary syndrome (PCOS) patients and controls undergoing controlled ovarian stimulation. METHODS: Nine patients with PCOS and six controls were enrolled in this study. On the day of oocyte retrieval, GCs were collected from pooled follicular fluid. Total mRNA was extracted from GCs. Reverse transcription was performed and gene expression levels were quantified by realtime quantitative polymerase chain reaction. RESULTS: There were no significant differences in age, body mass index, and total gonadotropin dose, except for the ratio of luteinizing hormone to follicle-stimulating hormone between the PCOS and control groups. PPAR-γ and COX-2 mRNA was significantly downregulated in the GCs of PCOS women compared with controls (p=0.034 and p=0.018, respectively), but the expression of IL-6 and TNF-α mRNA did not show significant differences. No significant correlation was detected between the expression of these mRNA sequences and clinical characteristics, including the number of retrieved oocytes, oocyte maturity, cleavage, or the good embryo rate. Positive correlations were found among the PPAR-γ, COX-2, IL-6, and TNF-α mRNA levels. CONCLUSION: Our data may provide novel clues regarding ovarian GC dysfunction in PCOS, and indirectly provide evidence that the effect of PPAR-γ agonists in PCOS might result from alterations in the ovarian follicular environment. Further studies with a larger sample size are required to confirm these proposals.

Reference Values for the Revised Anti-Müllerian Hormone Generation II Assay: Infertile Population-based Study.

Anti-Müllerian hormone (AMH) is now accepted as an important clinical marker of ovarian reserve and is increasingly measured as an initial evaluation at infertility clinics. The aim of this study was to establish reference values for the revised second generation (Gen II) assay using population-based data. In this population-based cohort study, AMH data from unselected infertile women aged 25-45 years from June 2013 to June 2014 (n = 15,801) were collected. The AMH values were measured using the revised Gen II assay. We established and validated 5 AMH-age regression models. Based on the optimal AMH-age model, reference values and centile charts were obtained. The quadratic model (log AMH = 0.410 × age -0.008 × age² -3.791) was the most appropriate for describing the age-dependent decrease in AMH measured using the revised Gen II assay. This is the largest population-based study to establish age-specific reference values of AMH using the revised Gen II assay. These reference values may provide more specific information regarding the ovarian reserve estimation of infertile women.

The relationship between the number of CGG repeats and serum level of anti-Müllerian hormone in women without FMR1 premutation.

OBJECTIVE: To investigate the relationship between the number of CGG repeats in the fragile X mental retardation 1 (FMR1) gene and serum levels of anti-Müllerian hormone (AMH) in Korean infertility patients without premutation. STUDY DESIGN: A retrospective study of 228 infertile women who received fertility treatment in a single private in vitro fertilization (IVF) clinic from May 2010 to August 2012 was performed. Serum FSH and AMH were measured on menstrual day 3 and the number of CGG repeats was evaluated. RESULTS: The mean age of the study population was 33.3±3.8 years. No significant correlation was observed between CGG repeat count in both alleles and the serum FSH, AMH or multiples of median (MoM) of AMH in whole study subjects. In women with age ≥35 years, however, there was an increasing tendency in the MoM of AMH with increasing number of CGG repeats in allele 2 (R(2)=0.075, p=0.008). This correlation was not observed in patients aged less than 35 years. CONCLUSION: We observed a positive correlation between MoM of AMH and number of CGG repeats in allele 2 in women aged over 35 years. Our findings are discordant with other reports, and therefore further studies are needed to determine whether this discrepancy is due to ethnic differences.

Application of sperm selection using hyaluronic acid binding in intracytoplasmic sperm injection cycles: a sibling oocyte study.

The purpose of this study was to investigate whether sperm selection by hyaluronic acid (HA) binding could improve fertilization rate and embryo quality in intracytoplasmic sperm injection (ICSI) cycles. Two hundred nineteen oocytes obtained from eighteen women were injected with either HA-bound (n = 107) or conventionally selected spermatozoa (n = 112) in a randomized way. All of the participants were infertile couples who had normal sperm parameters but low fertilization rate in previous in vitro fertilization (IVF) cycle (n = 5) or experienced multiple IVF failures (n = 13). Lower fertilization (75.7% vs 83.0%) and cleavage rate on day 2 (72.9% vs 83.0%) was observed in oocytes injected with HA-bound spermatozoa than the conventional group, but the difference was not significant. Significantly lower cleavage rate was observed on day 3 in HA group (56.0% vs 69.6%, P = 0.038). Blastocyst formation rate and the number of transferred embryos were similar in both groups. In multiple IVF failure patients, significantly reduced fertilization rate (71.8% vs 85.3%, P = 0.046) and cleavage rate on day 2 (70.4% vs 85.3%, P = 0.029) and day 3 (53.5% vs 77.3%, P = 0.002) were noticed in HA group. Five women achieved pregnancy continuing more than 12 weeks after transfer (27.8%). Success of ICSI was not related with the number of embryos fertilized by HA-bound spermatozoa. Application of ICSI by sperm selection using HA binding is not helpful in couples with repeated poor fertilization or implantation despite normal sperm parameters.

Outcomes of preimplantation genetic diagnosis using either zona drilling with acidified Tyrode's solution or partial zona dissection.

OBJECTIVE: To review the outcomes of preimplantation genetic diagnosis (PGD) using zona drilling with acid Tyrode's solution (chemical zona pellucida drilling, chemical ZD) and those of partial zona dissection (PZD). METHODS: Clinical outcomes of seventy-one couples undergoing 85 PGD cycles from January 2005 to December 2010 were included. Blastocyst formation and the hatching rate, clinical pregnancy rate, ongoing pregnancy rate, implantation rate, and fetal gender ratio of the PZD and chemical ZD groups were compared. RESULTS: Application of PZD resulted in a significantly higher rate of clinical pregnancy (40.7% vs. 15.4%, p=0.022), ongoing pregnancy (35.6% vs. 11.5%, p=0.023), and implantation (18.1% vs. 5.7%, p=0.007) compared with chemical ZD. Among non-transferred embryos, the rate of blastocyst formation on day 5 (49.1% vs. 39.5%, p=0.016) and hatching on day 6 (47.2% vs. 26.5%, p<0.001) were also significantly higher in the PZD group. CONCLUSION: The mechanical zona dissection method showed better outcomes than chemical ZD in terms of the blastocyst development and pregnancy rate. In this study, the fact that chemical ZD was conducted in different period from mechanical method should be considered in interpreting the result.

Age-related distributions of anti-Müllerian hormone level and anti-Müllerian hormone models.

OBJECTIVE: To determine age-specific reference values for anti-Müllerian hormone (AMH) and to set up an optimal model for AMH changes by age for infertility investigations. DESIGN: Prospective study. SETTING: Several infertility clinics and two university hospitals. SAMPLE: A total of 21 226 AMH samples were obtained. METHODS: Data on patients' age, race/ethnicity, and AMH levels were available from the laboratory center data registry between November 2008 and January 2011. MAIN OUTCOME MEASURES: The distribution of AMH levels by age. From 16 972 women aged between 25 and 45 years, we established and validated five AMH-age regression models. RESULTS: The overall mean AMH level was 4.09 ± 3.71 ng/mL (median: 3.13 ng/mL). There was an inverse relation between AMH level and age. Among multiple regression models, the quadratic model was most appropriate to describe AMH-age relation (log AMH = 0.205 × age - 0.005 × age(2) - 0.047). CONCLUSIONS: AMH levels show a progressive decline with increasing age. Age-specific AMH values may provide more specific information useful for patients and clinicians. AMH-age models could play a role as a basic step to approach more accurate ovarian reserve estimation.