Is paternal age associated with transfer day, developmental stage, morphology, and initial hCG-rise of the competent blastocyst leading to live birth? A multicenter cohort study.

In this study we investigated whether age of men undergoing assisted reproductive technology (ART) treatment was associated with day of transfer, stage, morphology, and initial hCG-rise of the competent blastocyst leading to a live birth? The design was a multicenter historical cohort study based on exposure (age) and outcome data (blastocyst stage and morphology and initial hCG-rise) from men whose partner underwent single blastocyst transfer resulting in singleton pregnancy/birth. The ART treatments were carried out at sixteen private and university-based public fertility clinics. We included 7246 men and women, who between 2014 and 2018 underwent controlled ovarian stimulation (COS) or Frozen-thawed Embryo Transfer (FET) with a single blastocyst transfer resulting in singleton pregnancy were identified. 4842 men with a partner giving birth were included, by linking data to the Danish Medical Birth Registry. We showed that the adjusted association between paternal age and transfer day in COS treatments was OR 1.06, 95% CI (1.00;1.13). Meaning that for every increase of one year, men had a 6% increased probability that the competent blastocyst was transferred on day 6 compared to day 5. Further we showed that the mean difference in hCG values when comparing paternal age group 30-34, 35-39 and 40-45 with the age group 25-29 in those receiving COS treatment, all showed significantly lower adjusted values for older men. In conclusion we hypothesize that the later transfer (day 6) in female partners of older men may be due to longer time spent by the oocyte to repair fragmented DNA of the sperm cells, which should be a focus of future research in men.

Association between women's age and stage, morphology, and implantation of the competent blastocyst: a multicenter cohort study.

OBJECTIVE: To study if the age of women undergoing assisted reproductive technology treatment associates with stage, morphology, and implantation of the competent blastocyst. DESIGN: Multicenter historical cohort study based on exposure (age) and outcome data (blastocyst stage and morphology and initial human chorionic gonadotrophin [hCG] rise) from women undergoing single blastocyst transfer resulting in singleton pregnancy/birth. SETTING: Sixteen private and university-based facilities. PATIENT(S): In this study, 7,246 women who, between 2014 and 2018, underwent controlled ovarian stimulation (COS) or frozen-thawed embryo transfer (FET) with a single blastocyst transfer resulting in singleton pregnancy were identified. Linking data to the Danish Medical Birth Registry resulted in a total of 4,842 women with a live birth being included. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): The competent blastocyst development stage (1-6), inner cell mass (A, B, C), trophectoderm (A, B, C), and initial serum hCG value. RESULT(S): Adjusted analysis of age and stage in COS treatments showed that for every 1-year increase in age there was a 5% reduced probability of the competent blastocyst assessed as being in a high stage at transfer. Comparison between hCG values in women 18-24 years and 25-29 years in both COS and FET showed significantly lower levels in the youngest women. CONCLUSION(S): The initial hCG rise was influenced by the age of the woman, with an identical pattern for hCG values in COS and FET treatments. In COS, the competent blastocyst had a reduced stage with increasing women's age.

Selective use of corifollitropin for controlled ovarian stimulation for IVF in patients with low anti-Müllerian hormone.

Corifollitropin, a long-acting follicle-stimulating hormone (FSH) analogue used for in vitro fertilization (IVF), does not allow individualization of dosage, and the ovarian response is similar to around 300 IU of daily recombinant FSH. This has raised concerns about the risk of ovarian hyperstimulation syndrome (OHSS) when used in standard patients. We administered corifollitropin selectively to patients with anticipated low to moderate ovarian response based on antimüllerian hormone levels in the lower quartile. The end points were oocyte distribution and occurrence of OHSS in women with AMH ≤15 pmol/L. The study included a cohort of 368 patients treated in 599 cycles. Post hoc the cohort was subdivided according to AMH. With increasing baseline AMH, the number of oocytes increased from a mean of 2.7 (range 0-8 with AMH <3 pmol/L) to 6.3 (range 0-15 with AMH 10-15 pmol/L) oocytes. Cancellations of retrievals and transfers decreased significantly with increasing AMH. Overall, the ongoing live pregnancy rate per started cycle was 15.2%. None developed OHSS. No cycles were cancelled or needed triggering of ovulation using a GnRH agonist due to risk of OHSS. Selective use of corifollitropin in patients with AMH in the lower quartile is a safe and appropriate way of optimising stimulation.

Culture of human oocytes with granulocyte-macrophage colony-stimulating factor has no effect on embryonic chromosomal constitution.

The effect on ploidy rate in donated human oocytes after in-vitro culture with recombinant human granulocyte-macrophage colony-stimulating factor (GM-CSF; 2 ng/ml) from fertilization until day 3 was examined in a multicentre, prospective placebo-controlled and double-blinded study including 73 women donating 86 oocytes. The primary endpoint was to investigate the chromosomal constitution of human embryos (fluorescence in-situ hybridization analysis for chromosomes 13, 16, 18, 21, 22, X and Y) cultured with or without GM-CSF. The secondary endpoints were number of top-quality embryos (TQE) and number of normally developed embryos evaluated morphologically on day 3. The cytogenetic analyses demonstrated non-inferiority and therefore the chromosomal constitution of human embryos cultured in vitro in the presence of 2 ng/ml GM-CSF was no worse than the control group cultured without GM-CSF. In-vitro culture of human embryos in the presence of 2 ng/ml GM-CSF resulted in 34.8% (8/23) uniformly normal embryos. Culture without 2 ng/ml GM-CSF resulted in 33.3% (9/27) uniformly normal embryos. A trend towards a higher number of TQE in the test group was observed; however, due to lack of TQE in the control group, this was considered a random finding.

Diet-derived polyphenol metabolite enterolactone is a tissue-specific estrogen receptor activator.

Numerous dietary compounds can modify gene expression by binding to the members of the nuclear receptor superfamily of transcription factors. For example, dietary polyphenols, such as soy isoflavones genistein and daidzein, modulate the activity of the estrogen receptors (ERs)-alpha and ERbeta. An additional class of dietary polyphenols that modulate cellular signaling pathways are lignans, compounds that are common constituents of Western diets. In this study, we show that a metabolite of dietary lignans, enterolactone, at physiological concentrations, activates ER-mediated transcription in vitro with preference for ERalpha. The effects of enterolactone are mediated by the ER ligand binding domain and are susceptible to antiestrogen treatment. Furthermore, the affinity of enterolactone toward ERalpha, measured by a novel ligand binding assay, is augmented in cell culture conditions. Moreover, our results demonstrate for the first time that enterolactone has estrogenic activity in vivo. In transgenic estrogen-sensitive reporter mice, enterolactone induces tissue-specific estrogen-responsive reporter gene expression as well as promotes uterine stromal edema and expression of estrogen-responsive endogenous genes (CyclinD1 and Ki67). Taken together, our data show that enterolactone is a selective ER agonist inducing ER-mediated transcription both in vitro in different cell lines and in vivo in the mouse uterus.

Eggs forever?

A group of scientists from Harvard Medical School (Johnson et al., 2004) claims to have "established the existence of proliferative germ cells that sustain oocyte and follicle production in the postnatal mammalian ovary," expressing no doubts about their methods, results and conclusion. Johnson et al. based their conclusions of oocyte and follicular renewal from existing germline stem cells (GSC) in the postnatal mouse ovary on three types of observations: (1) A claimed discordance in follicle loss versus follicle atresia in the neonatal period and in the following pubertal and adult period; (2) immunohistochemical detection of proliferating GSC with meiotic capacity using combined markers for meiosis, germline, and mitosis; and (3) neo-folliculogenesis in ovarian chimeric grafting experiments with adult mice. Oogenesis is the process that transforms the proliferative oogonium into an oocyte through meiosis, followed by folliculogenesis and follicular and oocyte maturation. The most crucial part in producing a functional oocyte is firstly, initiation and completion of the first meiotic prophase, and secondly, enclosure of the resulting diplotene oocyte in a follicle. Neither of these two events has been shown to take place in Johnson et al.'s study of the postnatal mouse ovary. We hereby address the observations underpinning their hypothesis and conclude that it is premature to replace the paradigm that adult mammalian neo-oogenesis/folliculogenesis does not take place.

In vivo imaging of activated estrogen receptors in utero by estrogens and bisphenol A.

Environmental estrogens are of particular concern when exposure occurs during embryonic development. Although there are good models to study estrogenic activity of chemicals in adult animals, developmental exposure is much more difficult to test. The weak estrogenic activity of the environmental estrogen bisphenol A (BPA) in embryos is controversial. We have recently generated transgenic mice that carry a reporter construct with estrogen-responsive elements coupled to luciferase. We show that, using this in vivo model in combination with the IVIS imaging system, activation of estrogen receptors (ERs) by maternally applied BPA and other estrogens can be detected in living embryos in utero. Eight hours after exposure to 1 mg/kg BPA, ER transactivation could be significantly induced in the embryos. This was more potent than would be estimated from in vitro assays, although its intrinsic activity is still lower than that of diethylstilbestrol and 17beta-estradiol dipropionate. On the basis of these results, we conclude that the estrogenic potency of BPA estimated using in vitro assays might underestimate its estrogenic potential in embryos.

In vitro antiestrogenic effects of aryl methyl sulfone metabolites of polychlorinated biphenyls and 2,2-bis(4-chlorophenyl)-1,1-dichloroethene on 17beta-estradiol-induced gene expression in several bioassay systems.

Methylsulfonyl (MeSO(2)) metabolites of polychlorinated biphenyls (PCBs) and 2,2-bis(4-chlorophenyl)-1,1-dichloroethene (4,4'-DDE), itself a metabolite of the insecticide 4,4'-DDT, are emerging as a major class of contaminants in the tissues of wildlife and humans. We investigated the antiestrogenic capacity and potencies of 3'- and 4'-MeSO(2)-2,2',4,5,5'-pentachlorobiphenyl (CB101) and -2,2',4,5'-tetrachlorobiphenyl (CB49), which are among the most environmentally persistent MeSO(2)-PCBs, and 3-MeSO(2)-4,4'-DDE on estrogen receptor (ER)-dependent gene expression in four cell-based bioassay systems. Congener- and concentration-dependent antagonism of 17beta-estradiol (E2)-induced gene expression, rather than induction of ER-dependent gene expression, was observed for the MeSO(2)-PCBs on lucifierase activity in stably transfected human breast adenocarcinoma T47D cells (ER-CALUX) and vitellogenin (vtg) production in primary hepatocytes from male carp fish (Cyprinus carpio) (CARP-HEP/vtg). 4'-MeSO(2)-CB101 and -CB49 had the highest antagonistic potency (i.e., maximum inhibition of about 70%, LOECs of 1.0 microM and 2.5 microM), whereas 3'-MeSO(2)-CB101 and -CB49 were less antagonistic; the precursor CB101 and MeSO(2)-PCB analog MeSO(2)-2,5-dichlorobenzene had no effect. Relative to the 4-MeSO(2)-PCBs, tamoxifen (IC(50), 0.06 microM and 0.7 microM) was about 40 and 7 times more potent in the ER-CALUX and CARP-HEP/vtg assays, respectively. Congener- and concentration-dependent effects on aryl hydrocarbon receptor-mediated induction of EROD activity (carp hepatocytes), luciferase expression (H4IIE rat hepatoma [H4IIE.luc] cell line), or cell viability were not observed. 3-MeSO(2)-4,4'-DDE was neither estrogenic nor antiestrogenic in either of the bioassays. Inhibitory trends for the MeSO(2)-PCBs in a bioassay based on stably transfected human embryonic kidney cell (HEK293-ERalpha-ERE) were similar to the ER-CALUX and CARP-HEP/vtg bioassays, whereas the antagonism was weaker in a related HEK293-ERbeta-ERE bioassay. Our findings suggest that the 4'-MeSO(2)-PCBs are antiestrogenic in vitro via a reversible or surmountable interaction with fish or human ER, and that the interaction with human ERalpha is apparently favored over ERbeta. MeSO(2)-PCB metabolites are persistent and bioaccumulative contaminants, and therefore, could be potentially active as environmental antiestrogens in wildlife and humans.