The fatty acid composition in follicles is related to the developmental potential of oocytes up to the blastocyst stage: a single-centre cohort study.

BACKGROUND: Advanced maternal age and obesity are associated with impaired female fertility. Moreover, fatty acids (FA) in follicular fluid (FF) play important roles in oocyte maturation and embryo development. However, the effects of body mass index (BMI), age, and FF FA composition on embryo development between days 3 and 5 and blastocyst stage on day 5 are still unclear. METHODS: This study included 138 patients undergoing assisted reproductive technology (ART), which were divided into three BMI groups (18.5-24.9 kg/m2 vs. 25.0-29.9 kg/m2 vs. ≥ 30.0 kg/m2) and three age-related groups (20-30 years vs. 31-34 years vs. ≥ 35 years) which were compared for ART outcomes. Further, observations were divided into quartiles based on either of three parameters related to embryo outcome, i.e. (i) embryos developing between days 3 and 5 (ED3-5) and (ii) expanded blastocysts on day 5 (EB5), both expressed proportionally to the number of oocytes with two pronuclei (2PN), as well as (iii) the embryo utilization rate (EUR). Proportions of FF FA were then compared between Q1 and Q4, representing the quartile with the worst vs. the best embryo outcome, respectively. Finally, regression models were created to assess the relationships between BMI, age, FF total FA (TFA) concentration, relative proportions of specific FA and embryo outcome. RESULTS: Patients of Q1 had higher proportions of FF C20:5n-3, C22:6n-3 and total n-3 PUFA than Q4 patients. Furthermore, Q4 patients tended to be younger than Q1 patients. Within the whole cohort, the proportion of C20:5n-3 negatively correlated with ED3-5/2PN and EUR, while EB5/2PN tended to be negatively correlated with age. Regression models within the overweight and obese group confirmed the negative relation between C20:5n-3 and ED3-5/2PN, but also indicated additional associations: C18:1n-9 and C20:4n-6 were positively associated with ED3-5/2PN and EUR, respectively while the proportion of C18:0 was negatively associated with EUR. CONCLUSION: The proportions of n-3 PUFA, particularly C20:5n-3 and C22:6n-3 were reduced in the patients' quartile with the best embryo outcome. This group of patients was also younger. However, the embryo quality parameters of overweight/obese patients were not associated with age but were positively associated with FF C18:1n-9 and negatively with the proportions of C18:0 or C20:5n-3. TRIAL REGISTRATION: This study' registration number was B670201627735.

Activin A-derived human embryonic stem cells show increased competence to differentiate into primordial germ cell-like cells.

Protocols for specifying human primordial germ cell-like cells (hPGCLCs) from human embryonic stem cells (hESCs) remain hindered by differences between hESC lines, their derivation methods, and maintenance culture conditions. This poses significant challenges for establishing reproducible in vitro models of human gametogenesis. Here, we investigated the influence of activin A (ActA) during derivation and maintenance on the propensity of hESCs to differentiate into PGCLCs. We show that continuous ActA supplementation during hESC derivation (from blastocyst until the formation of the post-inner cell mass intermediate [PICMI]) and supplementation (from the first passage of the PICMI onwards) is beneficial to differentiate hESCs to PGCLCs subsequently. Moreover, comparing isogenic primed and naïve states prior to differentiation, we showed that conversion of hESCs to the 4i-state improves differentiation to (TNAP [tissue nonspecific alkaline phosphatase]+/PDPN [podoplanin]+) PGCLCs. Those PGCLCs expressed several germ cell markers, including TFAP2C (transcription factor AP-2 gamma), SOX17 (SRY-box transcription factor 17), and NANOS3 (nanos C2HC-type zinc finger 3), and markers associated with germ cell migration, CXCR4 (C-X-C motif chemokine receptor 4), LAMA4 (laminin subunit alpha 4), ITGA6 (integrin subunit alpha 6), and CDH4 (cadherin 4), suggesting that the large numbers of PGCLCs obtained may be suitable to differentiate further into more mature germ cells. Finally, hESCs derived in the presence of ActA showed higher competence to differentiate to hPGCLC, in particular if transiently converted to the 4i-state. Our work provides insights into the differences in differentiation propensity of hESCs and delivers an optimized protocol to support efficient human germ cell derivation.

DSD: A Discussion at the Crossroads of Medicine, Human Rights, and Politics.

Over the past years, the topic of "disorder/differences in sex development (DSD)" or "intersex" people has become subject of the international political agenda. In 2017, a resolution of the Parliamentary Assembly of the Council of Europe argued that the practice of surgically modifying intersex children's genitals without medical necessity and without consent of the person concerned is a human rights violation. This resolution and related statements might impact heavily on pediatric urologists and their practice. While this resolution concerns a form of soft law and is not directly enforceable in member states, it might impact the national debates concerning legislation and medical guidelines on DSD. Consequently, this article reflects on this discussion by elaborating on the importance of human rights in our evolving understanding and legislation on DSD and other gender and sexuality issues in general. It constitutes a plea for a dialogue between medical professionals, lawmakers and human rights scholars which would lead to legislation and medical guidelines that take a holistic and rights-based approach.

Constructing and enacting kinship in sister-to-sister egg donation families: a multi-family member interview study.

Although intra-familial egg donation has been practiced for more than 15 years in several countries, little is known about family relationships in this family type. Framed within the new kinship studies, this article focuses on the experiential dimension of kinship in sister-to-sister egg donation families: how is kinship 'unpacked' and 'reconstructed' in this specific family constellation? Qualitative data analysis of interviews with receiving parents, their donating sisters and the donor children revealed six themes: (1) being connected as an extended family; (2) disambiguating motherhood; (3) giving and receiving as structuring processes; (4) acknowledging and managing the 'special' link between donor and child; (5) making sense of the union between father and donor; and (6) kinship constructions being challenged. This study showed the complex and continuous balancing of meanings related to the mother-child dyad, the donor-child dyad and the donor-father dyad. What stood out was the complexity of, on the one hand cherishing the genetic link with the child allowed by the sisters' egg donation, while, on the other, managing the meanings related to this link, by, for instance, acknowledging, downsizing, symbolising, and differentiating it from the mother-child bond. (A Virtual Abstract of this paper can be accessed at: https://www.youtube.com/channel/UC_979cmCmR9rLrKuD7z0ycA).

Detailed method description for noninvasive monitoring of differentiation status of human embryonic stem cells.

The (non)differentiation status of human embryonic stem cells (hESCs) is usually analyzed by determination of key pluripotency defining markers (e.g., OCT4, Nanog, SOX2) by means of reverse transcription quantitative polymerase chain reaction (RT-qPCR), flow cytometry (FC), and immunostaining. Despite proven usefulness of these techniques, their destructive nature makes it impossible to follow up on the same hESC colonies for several days, leading to a loss of information. In 2003, an OCT4-eGFP knock-in hESC line to monitor OCT4 expression was developed and commercialized. However, to the best of our knowledge, the use of fluorescence microscopy (FM) for monitoring the OCT4-eGFP expression of these cells without sacrificing them has not been described to date. Here, we describe such a method in detail, emphasizing both its resolving power and its complementary nature to FC as well as the potential pitfalls in standardizing the output of the FM measurements. The potential of the method is demonstrated by comparison of hESCs cultured in several conditions, both feeder free (vitronectin, VN) and grown on feeder cells (mouse embryonic fibroblasts, MEFs).

A maternally inherited autosomal point mutation in human phospholipase C zeta (PLCζ) leads to male infertility.

BACKGROUND: Male factor and idiopathic infertility contribute significantly to global infertility, with abnormal testicular gene expression considered to be a major cause. Certain types of male infertility are caused by failure of the sperm to activate the oocyte, a process normally regulated by calcium oscillations, thought to be induced by a sperm-specific phospholipase C, PLCzeta (PLCζ). Previously, we identified a point mutation in an infertile male resulting in the substitution of histidine for proline at position 398 of the protein sequence (PLCζ(H398P)), leading to abnormal PLCζ function and infertility. METHODS AND RESULTS: Here, using a combination of direct-sequencing and mini-sequencing of the PLCζ gene from the patient and his family, we report the identification of a second PLCζ mutation in the same patient resulting in a histidine to leucine substitution at position 233 (PLCζ(H233L)), which is predicted to disrupt local protein interactions in a manner similar to PLCζ(H398P) and was shown to exhibit abnormal calcium oscillatory ability following predictive 3D modelling and cRNA injection in mouse oocytes respectively. We show that PLCζ(H233L) and PLCζ(H398P) exist on distinct parental chromosomes, the former inherited from the patient's mother and the latter from his father. Neither mutation was detected utilizing custom-made single-nucleotide polymorphism assays in 100 fertile males and females, or 8 infertile males with characterized oocyte activation deficiency. CONCLUSIONS: Collectively, our findings provide further evidence regarding the importance of PLCζ at oocyte activation and forms of male infertility where this is deficient. Additionally, we show that the inheritance patterns underlying male infertility are more complex than previously thought and may involve maternal mechanisms.