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.

The effect of human embryonic stem cells (hESCs) long-term normoxic and hypoxic cultures on the maintenance of pluripotency.

The maintenance of pluripotency of human embryonic stem cells (hESCs) requires a high efficiency of self-renewal. During in vitro propagation, however, spontaneous differentiation occurs frequently, and there is also a risk of chromosomal changes. In this study, we assessed the properties of hESCs after long-term culture at ambient air and 5% oxygen growth conditions. The hESC lines were grown for up to 42 and 18 mo in normoxic and hypoxic conditions, respectively, and their proliferation; expression of Oct4, SSEA1, Nanog, and Notch1; karyotype; telomerase activity; and differentiation potential in vitro were evaluated. In contrast to cultures at 20% oxygen, where the central zones of the colonies underwent spontaneous differentiation, during exposure to 5% oxygen, the hESC colonies maintained a homogenous and flat morphology that was consistent with the presence of Oct4-positive undifferentiated phenotype. Irrespective of oxygen concentration, the undifferentiated cells expressed high levels of Nanog and Oct4 transcripts, normal karyotype, and high telomerase activity. When assayed for differentiation potential, they yielded derivatives of all three embryonic germ layers. Our data thus indicate that hypoxic exposure has the capacity to sustain enhanced long-term self-renewal of hESCs. The hESC lines described in the current paper can be obtained for research purposes from the Laboratory for Stem Cell Research, Aalborg University.

[Egg-donation in Denmark]. / Aegdonation i Danmark.

Egg-donation is an efficient treatment of female infertility caused by ovarian failure. Although the use of volunteer donors has been allowed in Denmark since 1 January 2007, there is a shortage of oocyte donors, which is in contrast to the increasing demand from couples in need of donor eggs. In this paper we discuss possibilities to enhance the recruitment of volunteer donors. In order to increase the number of egg-donations, the Danish legislation should be changed to allow the use of crossed anonymous donation.

Derivation and characterization of four new human embryonic stem cell lines: the Danish experience.

In September 2003, legislation approved in Denmark legalized work on surplus human embryos from IVF for clinical purposes to establish human embryonic stem (ES) cell cultures. The aim of this study was to establish such stem cell lines. Fresh surplus embryos were donated after informed consent from the donors. Embryos were cultured into blastocysts and using the immunosurgery procedure, inner cell masses were isolated and cultured on irradiated human foreskin fibroblasts in KnockOut D-MEM supplemented with KnockOut Serum Replacement, bFGF, and LIF. Within a period of 12 months, 198 embryos were donated. Four isolated inner cell masses developed into putative ES cell lines, CLS1, CLS2, CLS3, CLS4, which have now been continuously cultured for eight months, corresponding to 30 passages. These cells expressed markers for undifferentiated human ES cells: stage-specific embryonic antigen-4, tumour-related antigen (TRA)-1-60, TRA-1-81, OCT4, NANOG, SOX2, and FGF4. The cells expressed high levels of telomerase activity, had a normal karyotype, and have been successfully cryopreserved and thawed. Finally, the cells displayed the potential to differentiate in vitro into cell types originating from all three germ layers. It is thought that the cell lines described in this study are the first human ES cells established in Denmark.

Mutational analysis of the human FATE gene in 144 infertile men.

The FATE gene maps to Xq28 where one case of a translocation breakpoint has been found in an infertile man. Moreover, the FATE promoter contains a putative SF-1-binding site, and FATE has been proposed as representing a target gene of SF-1 in testicular development or germ cell differentiation. This study presents a complete mutational screening of the FATE gene in a random group of 144 infertile males. Four polymorphisms and two mutations were found. Three of the polymorphisms, viz., 741C-->T, 905A-->C, and 3985C-->T, occurred in exon 5 and intron 2 and did not alter the deduced polypeptide. One polymorphism resulted in the conservative amino acid exchange, A10 V, in 16.0% of the patients. This substitution occurred with similar frequencies in the control groups, indicating that the mutation does not affect fertility in men or women. The two mutations caused the non-conservative amino acid substitutions S125R (patient 1) and I34T (patient 2). A family study (patient 1) revealed, however, that S125R was inherited and that a fertile male family member carried the mutation. Patient 2 did not have relevant family members who could be examined. Thus, this study has shown that only 1.4% of infertile men have mutations in the FATE gene, and that some of these mutations do not singly cause infertility. Hence, FATE may not play an important role in the disease-state of infertile men attending fertility clinics. However, FATE mutations cannot be excluded as being a contributing factor in some cases of male infertility.