Reduced Steroid Synthesis in the Follicular Fluid of MTHFR 677TT Mutation Carriers: Effects of Increased Folic Acid Administration.

Objective To compare steroid profiles in the follicular fluid (FF) from women homozygous for the methylenetetrahydrofolate reductase (MTHFR) 677C>T mutation and wildtype controls and to correlate it with the folic acid administration scheme applied at the time of oocyte retrieval. Design Retrospective single center study. Subjects and Methods Infertile patients treated by using assisted reproductive techniques were genotyped routinely for the MTHFR 677C>T mutation. In 2006 they had received folic acid supplementation doses of 400 µg daily per os. This group was designated Group-400 (n = 10). From 2008 onwards, all of our infertility patients received a daily dose of 800 µg folic acid per os. Women from this group were designated Group-800 (n = 28). FF were collected and a panel of steroid hormones (estradiol, estrone, estriol, cortisol, progesterone, 17-OH progesterone, testosterone, androstenedione, aldosterone, DHEA, and DHEA-S) was measured by isotope dilution liquid chromatography-tandem mass spectrometry employing atmospheric pressure photo ionization (APPI). Results In Group-400, the FF hormone profile confirmed a significant reduction of estradiol in homozygous 677TT carriers (0.52 ± 0.08-fold, exact p = 0.032) and for the first time also revealed significantly reduced estriol concentrations in these individuals (0.54 ± 0.05-fold, p = 0.016), as compared to wildtype controls. In Group-800, no significant differences were found for concentrations of any of the steroid hormones between homozygous 677TT carriers and wildtype controls. Conclusions The current findings support and extend previous reports on reduced concentrations of specific steroid hormones in follicular fluids of homozygous MTHFR 677C>T mutation carriers. The restoration of the FF hormone profile by elevated-dose folic acid supplementation might impact performing ART in infertile women with the MTHFR 677TT-genotype. Further adequately powered studies are necessary to verify our finding and to demonstrate the clinical effect of enhanced folic supplementation on ovarian function.

Subtle progesterone rise on the day of human chorionic gonadotropin administration is associated with lower live birth rates in women undergoing assisted reproductive technology: a retrospective study with 2,555 fresh embryo transfers.

OBJECTIVE: To evaluate the association between serum P levels on the day of hCG administration and pregnancy outcome in women undergoing controlled ovarian hyperstimulation, prevention of premature ovulation by GnRH analogues, and fresh ET after 5 days of embryo culture. DESIGN: Retrospective, observational, cohort study. SETTING: Private IVF center. PATIENT(S): A total of 2,555 women undergoing fresh ET on day 5 in 2,062 GnRH agonist and 493 GnRH antagonist cycles. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Live birth rate. RESULT(S): Live birth rate in cycles with GnRH agonists was significantly lower in women with P levels ≥ 2.0 ng/mL (17.4%) on the day of hCG administration as compared with women with P levels <1.5 ng/mL (24.6%) and 1.5-1.99 ng/mL (26.7%). No such significant differences in live birth rates in cycles with GnRH antagonist could be observed. CONCLUSION(S): A rise of serum P levels ≥ 2.0 ng/mL on the day of hCG administration is associated with impaired early embryo implantation and reduced live birth rate in cycles with GnRH agonists after day-5 fresh ET.

Divergent effects of the 677C>T mutation of the 5,10-methylenetetrahydrofolate reductase (MTHFR) gene on ovarian responsiveness and anti-Müllerian hormone concentrations.

OBJECTIVE: To investigate the influence of the 5,10-methylenetetrahydrofolate reductase (MTHFR) 677C>T mutation on serum anti-Müllerian hormone (AMH) concentrations and on the numbers of oocytes retrieved (NOR) following controlled ovarian hyperstimulation (COH). DESIGN: Prospective cohort study. SETTING: University-based infertility clinic. PATIENT(S): Two hundred and seventy women undergoing COH for IVF with or without intracytoplasmic sperm injection. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): AMH levels were determined from blood samples collected after 10 days of GnRH superagonist treatment and before COH. The MTHFR 677C>T genotype was characterized by a TaqMan 5' nuclease assay. RESULT(S): AMH serum concentrations correlated significantly with the NOR in all individuals studied. Average (±SD) AMH levels of TT carriers (2.85±2.23 ng/mL) were significantly higher than those of homozygous CC (1.91±1.59 ng/mL) or heterozygous CT individuals (2.23±1.74 ng/mL). When evaluated by multiple regression analysis, AMH had a significant positive effect on NOR, whereas age and MTHFR 677TT genotype had significant negative effects. CONCLUSION(S): The MTHFR 677TT genotype is associated with higher serum AMH concentrations and has a negative effect on NOR. This apparent paradox might be resolved in light of recent findings describing a negative feedback function of AMH in the coordination of follicle development.

Predictive value of early serum beta-hCG levels after single blastocyst transfer.

OBJECTIVE: Analysis of serum beta-human chorionic gonadotropin (beta-hCG) levels on standardized days after oocyte retrieval (OR) following single blastocyst transfer to predict pregnancy outcome after assisted reproductive technology. DESIGN: Retrospective study. SETTING: Private fertility center. POPULATION: A total of 230 women with an intact intrauterine pregnancy 21 days after OR and a vital term delivery (n = 191) or a miscarriage (< 10+0 gestational weeks) (n = 39) in the course of the pregnancy following single blastocyst transfer between 1999 and 2008. METHODS: Blood was sampled 11, 14 and partly 21 days after OR. The serum concentration of total beta-hCG was measured by an automated quantitative immunoassay. Receiver operating characteristic curves were generated to calculate sensitivity and specificity. MAIN OUTCOME MEASURES: A total of 509 beta-hCG measurements. RESULTS: The mean beta-hCG concentrations on days 11, 14 and 21 after OR were significantly lower in the group of patients with a miscarriage compared to the group with a term delivery (p = 0.02, < 0.001 and < 0.001 respectively). With the cut-off values of 15 mIU/mL, 80 mIU/mL and 1500 mIU/mL on days 11, 14 and 21, the positive predictive value was 89%, 93% and 92%. The negative predictive value was 28%, 40% and 54%. CONCLUSIONS: The predictive value of a single beta-hCG measurement for pregnancy outcome after single blastocyst transfer increases between days 11 and 21 after OR. A single beta-hCG measurement on day 14 may balance the accuracy of prediction and the necessity of an early reassuring test.

Common 677C-->T mutation of the 5,10-methylenetetrahydrofolate reductase gene affects follicular estradiol synthesis.

OBJECTIVE: To investigate the influence of the 5,10-methylenetetrahydrofolate reductase 677C-->T mutation on the E(2) synthesis in human granulosa cells (GCs). DESIGN: In vitro cell culture study. SETTING: Research laboratory of a university hospital. PATIENT(S): Follicular fluids (n = 139) and GCs (n = 66) were obtained from patients undergoing controlled ovarian hyperstimulation for IVF with or without ICSI. INTERVENTION(S): Granulosa cells were cultured for a total of 5 days. On day 3, the cells either were stimulated with recombinant (r-) FSH or r-LH (80 IU/L for 48 h) or were sham stimulated. MAIN OUTCOME MEASURE(S): Estradiol and protein content were measured in the pooled follicular fluids of each individual. At the end of each GC-culturing period, the concentrations of E(2) were measured in the supernatants of triplicate cultures by immunoassays. The 5,10-methylenetetrahydrofolate reductase 677C-->T genotype was determined by RFLP analysis. RESULT(S): The E(2)-protein ratio of homozygous T/T carriers was significantly lower compared with that of homozygous C/C individuals. Furthermore, basal and r-FSH- as well as r-LH-stimulated E(2) synthesis of GC obtained from homozygous T/T patients was significantly reduced, compared with GC from heterozygous C/T and homozygous C/C subjects. CONCLUSION(S): Decreased E(2) in follicular fluid and decreased E(2) synthesis of GC from homozygous T/T individuals suggest that reduced follicular E(2) is a result of impaired E(2) production of human GC.

Quantitative evaluation of spermatozoa ultrastructure after acupuncture treatment for idiopathic male infertility.

OBJECTIVE: To evaluate the ultramorphologic sperm features of idiopathic infertile men after acupuncture therapy. DESIGN: Prospective controlled study. SETTING: Christian-Lauritzen-Institut, Ulm, IVF center Munich, Germany, and Department of General Biology, University of Siena, Siena, Italy. PATIENT(S): Forty men with idiopathic oligospermia, asthenospermia, or teratozoospermia. INTERVENTION(S): Twenty eight of the patients received acupuncture twice a week over a period of 5 weeks. The samples from the treatment group were randomized with semen samples from the 12 men in the untreated control group. MAIN OUTCOME MEASURE(S): Quantitative analysis by transmission electron microscopy (TEM) was used to evaluate the samples, using the mathematical formula based on submicroscopic characteristics. RESULT(S): Statistical evaluation of the TEM data showed a statistically significant increase after acupuncture in the percentage and number of sperm without ultrastructural defects in the total ejaculates. A statistically significant improvement was detected in acrosome position and shape, nuclear shape, axonemal pattern and shape, and accessory fibers of sperm organelles. However, specific sperm pathologies in the form of apoptosis, immaturity, and necrosis showed no statistically significant changes between the control and treatment groups before and after treatment. CONCLUSION(S): The treatment of idiopathic male infertility could benefit from employing acupuncture. A general improvement of sperm quality, specifically in the ultrastructural integrity of spermatozoa, was seen after acupuncture, although we did not identify specific sperm pathologies that could be particularly sensitive to this therapy.