Hum. Reprod. [JOURNAL]
Sun
200 papers
Evidence for de novo synthesis of corticosteroids in the human ovary.
Johannsen ML, Styrishave B, Zheng M … +10 more , Mamsen LS, Poulsen LLC, Hay-Schmidt A, Anujan P, Franks S, Grøndahl ML, Kristensen SG, Macklon KT, Tanvig M, Yding Andersen C
Hum Reprod · 2026 Jul · PMID 42402373 · Publisher ↗
STUDY QUESTION: Does the human ovary possess the enzymatic machinery required for local corticosteroid synthesis during follicular development? SUMMARY ANSWER: The human ovary exhibits dynamic corticosteroid metabolism d...
STUDY QUESTION: Does the human ovary possess the enzymatic machinery required for local corticosteroid synthesis during follicular development? SUMMARY ANSWER: The human ovary exhibits dynamic corticosteroid metabolism during follicular development and expresses enzymes consistent with local corticosteroid synthesis. WHAT IS KNOWN ALREADY: Cortisol is a systemic glucocorticoid that binds to the glucocorticoid receptor (NR3C1), which is expressed in ovarian tissues. While excess systemic cortisol impairs follicular development, a surge in local cortisol activity around ovulation appears necessary for limiting the impact of the inflammatory process taking place. The preovulatory upregulation of 11β-hydroxysteroid dehydrogenase type 1 (HSD11B1) suggests local conversion of cortisone to active cortisol. However, whether the ovary can synthesize corticosteroids de novo remains uncertain. STUDY DESIGN, SIZE, DURATION: This observational study included samples from 34 women undergoing ovarian cryopreservation and samples from 50 women undergoing IVF/ICSI according to a standard antagonist protocol at a university hospital-affiliated fertility clinic in Denmark. PARTICIPANTS/MATERIALS, SETTING, METHODS: Women donated follicular fluid (FF), plasma, and granulosa cells. Steroid concentrations in FF and plasma were quantified using LC-MS/MS. Granulosa cell mRNA expression of steroidogenic enzymes (CYP11B1, CYP11B2, CYP21A2, HSD11B1, HSD11B2) was assessed by RT-qPCR. In addition, immunofluorescence (IF) was used to localize CYP21A2 in preantral and small antral follicles. FF samples were stratified according to follicle size (2-18 mm) and ovulatory status. Hormone dynamics and FF/plasma ratio were evaluated using linear mixed-effects models and non-parametric tests. MAIN RESULTS AND THE ROLE OF CHANCE: Intrafollicular cortisol levels increased with follicle size up to 8-10 mm, then declined until close to ovulation, where a sharp increase was observed (from <40 to ∼100 nM). In contrast, cortisone showed an inverse pattern, remaining elevated throughout the follicular phase but dropping sharply at ovulation. The cortisol/cortisone ratio shifted markedly at the time corresponding to ovulation. Cortisol precursors, particularly 11-deoxycortisol and 11-deoxycorticosterone, were substantially enriched in FF compared to plasma (FF/plasma ratios >10), suggesting local synthesis. This is supported by the expression of CYP21A2 and low-level CYP11B1 mRNA in GCs, and by IF localization of CYP21A2 in preantral and small antral follicles. In contrast, FF cortisol concentrations were consistently lower than in plasma (FF/plasma ratio <0.1), indicating active inactivation by HSD11B2, which was abundantly expressed. HSD11B1 expression was low but detectable in periovulatory follicles, consistent with local reactivation of cortisone to cortisol. Among mineralocorticoids, corticosterone was only detectable at ovulation, while aldosterone was undetectable. LIMITATIONS, REASONS FOR CAUTION: We were unable to localize CYP11B1 at the protein level due to limitations in available antibodies. Additionally, FF and plasma samples were obtained from different individuals, precluding direct paired comparisons. Some variability in plasma sample collection timing throughout the day also limits conclusions regarding diurnal fluctuations. WIDER IMPLICATIONS OF THE FINDINGS: These findings suggest that the human ovary possesses the capacity for partial corticosteroid synthesis, utilizing locally abundant precursors. Cortisol bioactivity appears to be temporally regulated through both inactivation and reactivation mechanisms, enabling a controlled, pro-resolving inflammatory response at ovulation. This mechanism may be essential for timely corpus luteum formation and maintenance of luteal phase integrity. Dysregulation of local cortisol dynamics may contribute to luteal phase insufficiency in a subset of women. The concept of a follicular "inflammatome" and its role in ovulatory success warrants further exploration. FUNDING: This research was supported by the University Hospital of Copenhagen (Rigshospitalet), The University of Copenhagen, and Novo Nordisk Foundation grant number NNF21OC00700556, as well as founded by the Interreg ÔKS V through ReproUnion (www.reprounion.eu) and a grant from Region Zealand Research Foundation. The funders had no role in study design, collection of data, analyses, writing of the manuscript, or the decision to submit it for publication. DISCLOSURES: During the preparation of this work, the author(s) used ChatGPT 4o for language assistance. After using this tool/service, the author(s) reviewed and edited the content as needed and take(s) full responsibility for the content of the published article. Furthermore, CYA has received consultant fees and honoraria for lectures from Ferring and IBSA. TRIAL REGISTRATION NUMBER: N/A.
Characterizing endometriosis and adenomyosis symptom clusters and their impact on quality of life in the All of Us Research Program.
Goroshchuk O, Pérez-Gómez N, Carmona F … +4 more , Taylor HS, Flores I, Saiz-Rodríguez M, Koller D
Hum Reprod · 2026 Jul · PMID 42381368 · Publisher ↗
STUDY QUESTION: Are there distinct symptom clusters in women with endometriosis and adenomyosis, and how do these clusters predict quality of life? SUMMARY ANSWER: Latent class analyses of 22 438 women with endometriosis...
STUDY QUESTION: Are there distinct symptom clusters in women with endometriosis and adenomyosis, and how do these clusters predict quality of life? SUMMARY ANSWER: Latent class analyses of 22 438 women with endometriosis identified four symptom phenotypes, including high pain-gastrointestinal and psychological-neurological clusters; adenomyosis cases were concentrated in high‑pain classes without a minimal‑symptom group, and high‑burden phenotypes had poorer quality of life. WHAT IS KNOWN ALREADY: Endometriosis and adenomyosis are heterogeneous conditions with diagnostic delays of 4-11 years. The classification based on surgical findings and imaging fails to capture the complex, systemic symptom combinations that affect patients' quality of life (QoL). STUDY DESIGN, SIZE, DURATION: Large-scale cross-sectional cohort study using data from the United States-based All of Us Research Program (Controlled Tier v8; May 2018-October 2023). PARTICIPANTS/MATERIALS, SETTING, METHODS: The dataset included 22 438 women with electronic health records (EHR) and/or self-reported endometriosis. Four Latent Class Analyses (LCA) were performed on different patient groups: (i) the full endometriosis cohort (n = 22 438); (ii) an age-corrected subset restricted to premenopausal women (aged 18-45; n = 5542) to minimize menopausal confounding; and two subgroups derived from the age-corrected cohort to test whether concomitant adenomyosis defines distinct symptomatic phenotypes: (iii) clinically confirmed endometriosis without adenomyosis (n = 1797) and (iv) adenomyosis with endometriosis (n = 643). LCA was performed using 19 self-reported and EHR-derived symptoms and comorbidities (e.g. chronic pelvic pain, migraine, depression, gastrointestinal symptoms) to identify patient subgroups. Multinomial logistic regression assessed sociodemographic (age, BMI, deprivation index) and clinical predictors (hormonal contraceptive use, surgical history) of class membership. The association between latent classes and QoL outcomes was evaluated using data from the 'Overall Health' and 'Health Care Access and Utilization' surveys. Data are available via the All of Us Researcher Workbench (https://workbench.allofus.org). MAIN RESULTS AND THE ROLE OF CHANCE: Latent class analysis of clinically confirmed endometriosis identified four distinct symptom phenotypes, most notably a severe 'High Pain & Gastrointestinal with Mood Symptoms' (High) cluster and a unique 'Predominantly Psychological/Neurological' cluster that challenges traditional gynecological-focused diagnostic frameworks. Patients with adenomyosis exhibited a distinct profile with two high-pain symptom classes and the absence of a fully asymptomatic group, indicating a higher overall disease burden. Membership in the High class was significantly associated with lower general health scores, reduced social satisfaction, and increased barriers to healthcare access. LIMITATIONS, REASONS FOR CAUTION: The All of Us is a United States-based research program, and the findings should be replicated in other independent cohorts to confirm generalizability across other geographical regions. The cross-sectional design limits causal inference regarding QoL outcomes. Also, the analyses rely on a mix of diagnostic approaches, including surgical and non-surgical clinical and self-reported diagnoses, which is both a strength and a limitation. The 'Minimal Symptom Burden' class may reflect under-documentation of symptoms in clinical records rather than a true lack of symptoms, and the High class may reflect to patients who have higher multimorbidity, hence have more hospital visits. WIDER IMPLICATIONS OF THE FINDINGS: These results support a shift toward personalized, interdisciplinary management of endometriosis that addresses mental health and gastrointestinal symptoms alongside pelvic pain. The identification of a 'Predominantly Psychological/Neurological' cluster suggests that young women presenting with non-classical symptoms (anxiety, migraine, depression) could be screened for endometriosis to reduce diagnostic delays and improve life-course potential. FUNDING: D.K. was supported by a 'Ramón y Cajal' fellowship from the Spanish Ministry of Science and Innovation (RYC2024-050099-I). O.G. was supported by a postdoctoral award from the Amy P Goldman Foundation. DISCLOSURES: I.F. is the co-founder and co-owner of Sur180 Therapeutics, which is developing a novel treatment for endometriosis, and the Chief Scientific Officer of Nura Health, which is developing a non-invasive diagnostic solution for the disease. I.F. has received ARPA-H grant no. ARPA-H-ICHUB-24-101-1035 as Principal Investigator, consulting fees from Nura Health and Sur180 Therapeutics, and stock and share options in both companies. I.F. is also the inventor on patents for 'Compositions and methods for treating endometriosis' (US10695341 and US10729693) and serves as an unpaid Board Member of the World Endometriosis Society and the Fundación Puertorriqueña de Pacientes con Endometriosis (ENDOPR). H.S.T. received speaker fees from Gedeon Richter, a grant from AbbVie through Yale University, consulting fees from Regeneron, and is a co-inventor on Yale's endometriosis biomarkers and treatments (including US11993816B2, US10982282B2, and US12286629B2). He also serves on the boards of the Environmental Health Trust and Environment and Human Health. TRIAL REGISTRATION NUMBER: N/A.
A new regulatory framework: impact of the SoHO Regulation for medically assisted reproduction.
Bariani F, Alteri A, Santilli L … +4 more , Vicentini MT, Di Ciaccio P, Lombardini L, Feltrin G
Hum Reprod · 2026 Jun · PMID 42361301 · Publisher ↗
Regulation (EU) 2024/1938 on Substances of Human Origin (SoHO), adopted in June 2024 and entering into application from 7 August 2027, establishes a unified and legally binding framework for all human-derived materials u...
Regulation (EU) 2024/1938 on Substances of Human Origin (SoHO), adopted in June 2024 and entering into application from 7 August 2027, establishes a unified and legally binding framework for all human-derived materials used in clinical practice, explicitly encompassing gametes and embryos. By replacing the previous EU Tissues and Cells Directives, it introduces harmonized standards for authorization, vigilance, and traceability, while reinforcing donor, recipient, and offspring protection. This commentary examines the main regulatory innovations relevant to medically assisted reproduction (MAR) and critically assesses their expected impact through a SWOT (Strengths, Weaknesses, Opportunities, and Threats) analysis. The SoHO Regulation offers an unprecedented opportunity to strengthen safety, quality, and ethical governance in MAR through evidence-based oversight and harmonized technical standards. However, its implementation may also generate significant administrative burden, challenges in proportionality, and potential delays in innovation. Overall, the SoHO Regulation represents a milestone for reproductive medicine, offering the chance to elevate safety and equity across Europe, provided its implementation is proportionate and supported by adequate guidance and collaboration among competent authorities and MAR professionals.
Subfertility in relation to gestational weight gain and gestational diabetes: the Norwegian Mother, Father and Child Cohort Study.
Laursen ASD, Mikkelsen EM, Mitter VR … +1 more , Magnus MC
Hum Reprod · 2026 Jun · PMID 42334924 · Publisher ↗
STUDY QUESTION: Do gestational weight gain trajectories and risk of gestational diabetes mellitus (GDM) differ according to the underlying fecundability and use of ART? SUMMARY ANSWER: Women with ART pregnancies experien...
STUDY QUESTION: Do gestational weight gain trajectories and risk of gestational diabetes mellitus (GDM) differ according to the underlying fecundability and use of ART? SUMMARY ANSWER: Women with ART pregnancies experienced slightly different gestational weight gain trajectories than those who conceived spontaneously within 3 months, with a higher weight gain during the second trimester and lower weight gain during the third trimester. WHAT IS KNOWN ALREADY: Use of ART and prolonged time-to-pregnancy (TTP) have both been linked to adverse pregnancy outcomes, including GDM. However, little is known about how gestational weight gain trajectories differ by fecundability or ART conception. STUDY DESIGN, SIZE, DURATION: A cohort study including 69,491 singleton pregnancies contributed by 61,175 women participating in the Norwegian Mother, Father and Child Cohort Study conducted between 1999 and 2008. PARTICIPANTS/MATERIALS, SETTING, METHODS: Pregnancies were categorized by self-reported TTP (≤3 months, 4-6 months, 7-11 months, ≥12 months) and ART conception identified from the Medical Birth Registry of Norway. Body weight was self-reported at four points (pre-pregnancy, ∼16 weeks' gestation, ∼28 weeks, and at the end of pregnancy) and GDM identified in the Medical Birth Registry of Norway. Gestational weight gain trajectories according to TTP and ART were assessed using mixed-effects linear regression and the risk of GDM was estimated with log-binomial regression. MAIN RESULTS AND ROLE OF CHANCE: Across all pregnancies, the adjusted average weekly weight gain was -15 g during the first trimester, 620 g during the second trimester, and 491 g during the third trimester. ART pregnancies showed different weight gain trajectories than spontaneous conceptions, namely, stable weight during the first trimester, higher weight gain during the second trimester, and lower weight gain during the third trimester compared with pregnancies with TTP ≤3 months. A total of 583 pregnancies (0.8%) were complicated by GDM. Longer TTP and ART were associated with a higher risk of GDM. Compared with pregnancies with TTP ≤3 months, the adjusted risk ratio was 1.48 (95% CI: 1.15; 1.90) for TTP ≥12 and 2.39 (95% CI: 1.71; 3.35) for ART pregnancies. LIMITATIONS, REASONS FOR CAUTION: Weight and TTP were self-reported and subject to misclassification. Important metabolic and reproductive confounders, including PCOS, were unavailable. Unmeasured confounding may partly or fully explain the observed associations. WIDER IMPLICATIONS OF THE FINDINGS: Given the weak associations and unmeasured confounding, the results need confirmation before they have clinical implications. STUDY FUNDING/COMPETING INTEREST(S): The study was supported by funding from the European Research Council (ERC) under the European Union's Horizon 2020 Research and Innovation Programme (grant 947684). It was also partly supported by the Research Council of Norway (Women's fertility, grant 320656) and its Centres of Excellence Funding Scheme (grant 262700). The authors declare no conflict of interest. TRIAL REGISTRATION NUMBER: N/A.
Pathogenic variants in ACTRT3 lead to male infertility characterized by fertilization defects.
Zhu H, Shi R, Zou G … +13 more , Diao F, Yu R, Qu R, Liu R, Shi J, Zhang Z, Wu T, Wang W, Chen B, He L, Wang L, Mu J, Sang Q
Hum Reprod · 2026 Jun · PMID 42330090 · Publisher ↗
STUDY QUESTION: Are there new genetic factors responsible for male infertility characterized by fertilization defects despite normal sperm motility? SUMMARY ANSWER: We identified the novel pathogenetic gene actin-related...
STUDY QUESTION: Are there new genetic factors responsible for male infertility characterized by fertilization defects despite normal sperm motility? SUMMARY ANSWER: We identified the novel pathogenetic gene actin-related protein T3 (ACTRT3) as one of primary causes for male infertility characterized by fertilization defects. WHAT IS KNOWN ALREADY: To date, only a few genes have been reported to be responsible for fertilization defects in male infertility despite normal sperm motility. ACTRT3, as one of the actin-related proteins, is specifically expressed highly in murine testes and interacts with the protein Profilin3 to form a testis-specific complex during spermatogenesis. STUDY DESIGN, SIZE, DURATION: The ACTRT3 variants were identified by whole-exome sequencing in a cohort of 248 infertile couples characterized by fertilization defects (fertilization rate ≤ 30% in their IVF/ICSI attempts) despite normal sperm concentration (≥15 × 106/ml) and normal motility (progressive motility ≥32%; motility of PR (progressive)+NP (non-progressive) ≥40%). Thirty-six hours after cell transfection, immunoprecipitation was performed to detect the interaction between ACTRT3 and actin-like 7A (ACTL7A)/phospholipase C zeta 1 (PLCZ1) (n = 3 biological replicates). A knock-in (KI) mouse model was generated by CRISPR-Cas9. Sexually mature males (10 weeks old) were used for fertility testing (n = 3 mice per group, lasts 6-8 months), semen parameter analysis (n = 3 mice per group), western blot (n = 3 biological replicates), immunofluorescence (n = 3 biological replicates), IVF (n ≥ 3 biological replicates), ICSI (n = 3 biological replicates), scanning electron microscopy (SEM, n = 3 mice per group), transmission electron microscopy (TEM, n = 3 mice per group), Ca2+ oscillations (n ≥ 3 biological replicates), and other functional assays between 2022 and 2025. PARTICIPANTS/MATERIALS, SETTING, METHODS: The ACTRT3 variants were identified by whole-exome sequencing and confirmed by Sanger sequencing. ACTRT3 KI mouse was constructed to discover the pathogenic mechanism. We first checked ultrastructure of the sperm through periodic acid-schiff staining, SEM, and TEM. Western blotting, immunofluorescence, and immunoprecipitation were further performed to clarify the molecular mechanism of the ACTRT3 variants. MAIN RESULTS AND THE ROLE OF CHANCE: We identified compound-heterozygous (NM_032487.5: c. C41G (p. Ser14Trp) and c. C687delA (p. Lys229fs)) and homozygous variants (NM_032487.5: c.712C>T (p. Gln238Ter)) in ACTRT3 in two infertile males, and functional studies in transgenic KI mouse models confirmed the pathogenicity of the Actrt3 variants. Defects in ACTRT3 caused decreased protein levels of ACTL7A and disrupted localization of PLCZ1 in sperm, which resulted in acrosome detachment, further impacted the binding between sperm and the zona pellucida of oocytes and caused fertilization failure. In addition, sperm with defective ACTRT3 cannot initiate calcium oscillations, which leads to embryo arrest. LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: More cases are needed to confirm the genetic contribution of ACTRT3 variants to male infertility. In addition, it will be more persuasive and informative by adding the rescue experiments of artificial oocyte activation through SrCl2 which is used to active calcium ion oscillation. WIDER IMPLICATIONS OF THE FINDINGS: Our findings suggest the causal relationship between ACTRT3 variants and male infertility caused by fertilization defects, and partially revealing the pathological mechanisms of ACTRT3 variants. This provides new insights into the understanding of male infertility due to fertilization problems and provides references for future clinical treatment. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by the National Natural Science Foundation of China (82325021, 32130029, and 82201767) and the Fundamental and Interdisciplinary Disciplines Breakthrough Plan of the Ministry of Education of China (No. JYB2025XDXM508). TRIAL REGISTRATION NUMBER: N/A.
A large retrospective study on 1PN embryo transfer supports the need for updated harmonized best practice guidelines.
McPherson NO, Ussher C, Holden S … +10 more , Hesketh N, Murray A, Lee A, Bui D, Traversa MV, Grkovic S, Dickson R, Bowman M, Marren A, Morbeck DE
Hum Reprod · 2026 Jun · PMID 42330074 · Publisher ↗
STUDY QUESTION: Do the genetic and clinical outcomes of monopronuclear blastocysts (MPBs) differ between standard insemination and intracytoplasmic sperm injection cycles, and what do these differences imply for risk str...
STUDY QUESTION: Do the genetic and clinical outcomes of monopronuclear blastocysts (MPBs) differ between standard insemination and intracytoplasmic sperm injection cycles, and what do these differences imply for risk stratification and individualized clinical decision-making? SUMMARY ANSWER: IVF-derived MPBs demonstrate significantly lower rates of uniparental inheritance than ICSI-derived MPBs (96.9% vs 65.9%) biparental inheritance (P < 0.001), with comparable clinical and neonatal outcomes to 2PN blastocysts following transfer of euploid biparental embryos. WHAT IS KNOWN ALREADY: A proportion of monopronuclear (1PN) zygotes can develop into euploid blastocysts and, following transfer, result in healthy live births, yet these embryos are widely discarded following fertilization check due to atypical pronucleation. The mechanisms underlying 1PN formation are varied and include asynchronous pronuclear formation, early pronuclear fusion, and premature pronuclear breakdown, meaning a subset may represent normally fertilized diploid zygotes missed at static assessment. STUDY DESIGN, SIZE, DURATION: Retrospective cohort study of 1PN embryos (N = 13 203) derived from IVF (n = 5266) or ICSI (n = 5464) inseminations across 10 730 cycles performed at multiple Australian clinics between January 2010 and December 2023. PARTICIPANTS/MATERIALS, SETTING, METHODS: Embryos were defined as 1PN by the appearance of a single pronucleus at fertilization check 16-18 h post-insemination. Time-lapse footage was reviewed on Day 3 to identify late appearing 1PNs and exclude late second pronucleus appearance. Suitable blastocysts underwent trophectoderm biopsy for pre-implantation genetic testing for aneuploidy (PGT-A) and short tandem repeat (STR)-based biparental inheritance testing; only euploid embryos with confirmed biparental inheritance were available for frozen embryo transfer. Outcomes assessed included ploidy, biparental inheritance, blastocyst development, utilization, morphokinetics, pregnancy, live birth, and maternal and neonatal outcomes. MAIN RESULTS AND THE ROLE OF CHANCE: IVF-derived MPBs had similar aneuploidy rates to two pronuclei (2PN) embryos (37.3% vs 33.9%) and 440/454 (96.9%) demonstrated biparental inheritance. ICSI-derived MPBs had higher aneuploidy rates (45.5% vs 31.5%, P < 0.05) and only 108/164 (65.9%) had biparental inheritance. Uniparental inheritance was predominantly maternal (IVF 92.8%; ICSI 94.6%). Both IVF and ICSI MPBs were less likely to reach blastocyst stage by Day 5 than 2PN embryos (IVF 19.3% vs 63.3%; ICSI 9.7% vs 60.6%, P < 0.05), and biparental IVF-1PN zygotes were more likely to have more nucleoli compared with uniparental IVF-1PN zygotes (P = 0.008). For embryos with confirmed biparental inheritance, there was no significant difference in clinical pregnancy, ongoing pregnancy, live birth rates, or neonatal outcomes compared with 2PN blastocysts. In approximately one in six cycles containing a 1PN embryo, no utilizable 2PN embryo were available (IVF 15.6%; ICSI 16.7%), with the 1PN embryo representing the sole option for embryo utilization. LIMITATIONS, REASONS FOR CAUTION: Retrospective single-entity design introduces potential selection bias and limits generalizability. Uniform protocols across sites preclude the level of evidence required for formal guideline revision. Differential use of time-lapse imaging for ICSI versus static assessment for IVF embryos may contribute to differences in 1PN identification rates between fertilization methods. The STR-based biparental classification platform has not been validated against an orthogonal technology for parental origin calling in 1PN embryos, and the possibility of triploid misclassification or absorption into unreported inconclusive outcomes cannot be excluded. WIDER IMPLICATIONS OF THE FINDINGS: These findings support a risk-stratified approach to MPB management based on fertilization method. IVF-derived MPBs meeting specific morphological and developmental criteria demonstrate a low-risk profile that warrants reconsideration of genetic testing requirements and may inform individualized consent discussions, particularly where 2PN embryos are unavailable. ICSI-derived MPBs carry a substantially higher risk of uniparental inheritance and comprehensive genetic testing remains indicated. STUDY FUNDING/COMPETING INTEREST(S): No funding was attached to this study. The authors declare no conflict of interest. TRIAL REGISTRATION NUMBER: N/A.
Explainability of artificial intelligence (AI) tools in assisted reproductive technologies (ARTs): promise, clinical reality, and conceptual limits.
Freour T, Anichini G, Gaillard M … +1 more , Mouchere H
Hum Reprod · 2026 Jun · PMID 42323849 · Publisher ↗
Artificial intelligence (AI) is increasingly integrated into clinical medicine, offering new opportunities for decision support, prediction, and personalization of care. However, many high-performing AI systems, particul...
Artificial intelligence (AI) is increasingly integrated into clinical medicine, offering new opportunities for decision support, prediction, and personalization of care. However, many high-performing AI systems, particularly those based on deep learning, operate as 'black boxes', raising concerns about transparency, trust, and professional accountability. Explainable artificial intelligence (XAI) has been proposed as a solution to these challenges, yet its clinical value remains contested. In this Perspective, we critically examine the role of explainability in medical AI using assisted reproductive technologies (ARTs) as a paradigmatic clinical context. ART is characterized by biological complexity, probabilistic decision-making, and growing reliance on AI tools for embryo assessment, gamete selection, and outcome prediction. Drawing on interdisciplinary expertise in medicine, AI, philosophy, and social sciences, we briefly review current AI applications in ART, clarify key conceptual distinctions between interpretability, explainability, and transparency, and analyze the potential benefits and limitations of XAI. We argue that explainability should not be regarded as an intrinsic requirement for all clinical AI systems. Instead, its value is context-dependent and must be weighed against other critical factors such as transparent reporting, rigorous validation, and demonstrated clinical impact. Moving beyond a binary opposition between black-box and explainable models, we advocate for a pragmatic, evidence-based approach to the integration of AI into digital medicine.
Ultra-rapid versus conventional oocyte vitrification: a comparative study of clinical and molecular outcomes with single-cell transcriptomic insights.
Aydin B, Baltaci V, Aktuna S … +10 more , Ozer L, Dorofeyeva U, Kubašková TM, Babaríková V, Lazar I, Kmet'ová T, Evin E, Baysan M, Vives Perelló A, Unsal E
Hum Reprod · 2026 Jun · PMID 42323848 · Publisher ↗
STUDY QUESTION: By reducing the duration of the vitrification-warming procedure to 1 min does ultra-rapid vitrification-warming (URV/W) effectively maintain oocyte viability and improve clinical outcomes compared to conv...
STUDY QUESTION: By reducing the duration of the vitrification-warming procedure to 1 min does ultra-rapid vitrification-warming (URV/W) effectively maintain oocyte viability and improve clinical outcomes compared to conventional vitrification-warming (CV/W) timing (14 min) and what is the effect of the two procedures on cellular stress? SUMMARY ANSWER: URV/W shortens procedure time, reduces cryoprotectant exposure, improves oocyte survival, and is associated with fewer transcriptomic alterations in oocytes than CV/W. WHAT IS KNOWN ALREADY: CV/W techniques require extended cryoprotectant exposure and micromanipulation, which elevate cellular stress and the risk of cryoinjury. URV/W procedures address these challenges by reducing exposure and manipulation, though comparative data on clinical outcomes and transcriptomic signatures in human oocytes remain limited. STUDY DESIGN, SIZE, DURATION: The study was performed between August 2024 and December 2024 and included a clinical cohort and a transcriptomic cohort to provide a thorough assessment of the effects of cryopreservation methodology on oocyte and embryo viability, development, and cellular stress. The clinical cohort comprised 1077 oocytes used to evaluate clinical outcomes following CV/W and URV/W. The study also included a prospective analysis involving 68 oocytes from 4 donors in the transcriptomic cohort. Oocytes and trophectoderm biopsy samples from blastocyst-stage embryos were assigned to three groups: fresh, CV/W, and URV/W. PARTICIPANTS/MATERIALS, SETTING, METHODS: The clinical study was performed using a total of 1077 oocytes from 46 donors, which were matched and allocated for the comparison of clinical outcomes between CV/W (n = 519) and URV/W (n = 558). Following vitrification-warming, the oocytes were fertilized via ICSI for assessment of embryo development and clinical outcomes. In the transcriptomic cohort of the study, transcriptomic testing and analysis were performed as part of the prospective analysis to compare CV/W and URV/W in terms of their effectiveness in oocyte freezing and warming, their impact on embryo development, and their effects at the molecular level. A total of 68 samples were obtained from 4 donors, including eight oocytes and eight trophectoderm biopsy samples from each of the three groups. These samples were analyzed to investigate the cellular effects of cryopreservation-induced stress through transcriptomic profiling. The single-cell transcriptomic study included preparation of cDNA libraries followed by next-generation sequencing to investigate differential gene expression across oocytes and embryonic trophectoderm cells in the three groups. This method allowed for the comprehensive monitoring of transcriptional activity, enabling the detection and quantification of mRNA molecules to evaluate the transcriptomic signatures of the study groups. In addition, functional enrichment analyses of up- and downregulated genes were conducted and classified based on gene ontology to identify potential pathways associated with embryo quality and cellular stress. MAIN RESULTS AND THE ROLE OF CHANCE: Transcriptomic analysis indicated that gene expression patterns following URV/W were more similar to fresh oocytes than those undergoing CV/W. Differences in gene expression patterns among blastocysts from the three groups were minimal, as the total number of differentially expressed genes was limited. Oocytes subjected to URV/W demonstrated a significantly higher survival rate, lower post-ICSI degeneration, and produced more high-quality Day-5 blastocysts compared to those vitrified using CV/W methods (P < 0.001 for all comparisons). These findings are unlikely to be solely attributable to differences in procedural timing, as fertilization, cleavage, euploidy, clinical pregnancy, and live birth rates were comparable between groups (P > 0.05 for all comparisons). LIMITATIONS, REASONS FOR CAUTION: All oocyte samples donated (1145) for research purposes were included in the study without quality-based selection. Only embryos reaching the blastocyst stage on Days 5 or 6 using the URV/W method were included in the transcriptomic analysis. Embryos not reaching these stages were excluded. Consequently, only data from embryos with optimal development and good quality were analyzed, and compared among the three groups. Obtaining cDNA in single-cell studies is a novel and challenging process, particularly due to the limited availability of RNA. While advanced clinical evaluations can still be conducted on biopsied oocytes and embryos, the failure to obtain cDNA from the same samples may result in incomplete data, which can limit the overall scope and outcomes of the study. WIDER IMPLICATIONS OF THE FINDINGS: The comprehensive single-cell transcriptomic data obtained from this expanded dataset will help delineate pathways altered by different vitrification methods, providing critical insights into their efficacy and potential risks in clinical practice. Additionally, the findings will illuminate key genes and pathways involved in embryonic stress, enabling the development of more cost-effective, targeted gene panels for evaluating these factors. STUDY FUNDING/COMPETING INTEREST(S): This study was funded by the Sanatórium pre liečbu neplodnosti SPLN's, Ovogene and Mikrogen Genetic Diagnosis Laboratory's own resources. A.V.P. is a Medical Science Liaison for Kitazato Corporation-Dibimed. His role in the study was strictly scientific and unrelated to any commercial interest. All the other co-authors declare no conflicts of interest. TRIAL REGISTRATION NUMBER: n/a.
Natural versus artificial cycle for endometrial preparation in ovulatory women undergoing frozen-thawed embryo transfer: a randomised controlled trial.
Geysenbergh B, Debrock S, Lie Fong S … +11 more , Vanhie A, Tomassetti C, Bafort C, Meeuwis L, Neyens S, Pelckmans S, Laenen A, Vriens J, De Loecker P, Dancet E, Peeraer K
Hum Reprod · 2026 Jun · PMID 42315758 · Publisher ↗
STUDY QUESTION: Does a natural cycle result in higher clinical pregnancy rates (CPR) with foetal heartbeat compared to an artificial cycle for frozen-thawed embryo transfer (FET) preparation in ovulatory women? SUMMARY A...
STUDY QUESTION: Does a natural cycle result in higher clinical pregnancy rates (CPR) with foetal heartbeat compared to an artificial cycle for frozen-thawed embryo transfer (FET) preparation in ovulatory women? SUMMARY ANSWER: The CPR with foetal heartbeat did not differ between natural and artificial cycles in ovulatory women undergoing FET. WHAT IS KNOWN ALREADY: Several protocols for endometrial preparation have been developed for the increasing number of FET cycles in reproductive medicine. In natural cycle FET (NC-FET), spontaneous ovulation is used to time the embryo thawing and transfer, while in artificial cycle FET (AC-FET), endometrial preparation involves sequential oestrogen and progesterone administration. Current evidence on pregnancy rates does not favour one regimen over the other in women with regular ovulatory cycles. STUDY DESIGN, SIZE, DURATION: We conducted a multicentre, open-label, randomised trial comparing NC-FET with AC-FET across five Belgian fertility centres. Between October 2018 and October 2024, 561 women were randomised (1:1) using a computer-generated allocation after written informed consent. The primary outcome was CPR with foetal heartbeat per cycle analysed on an intention-to-treat basis. Secondary outcomes included endometrial thickness, number of clinic visits for FET cycle monitoring and the rates of (biochemical) pregnancy, ongoing pregnancy, live birth, miscarriage, ectopic pregnancy, multiple pregnancy and cycle cancellation. Obstetric outcomes were recorded for all ongoing pregnancies. PARTICIPANTS/MATERIALS, SETTING, METHODS: Women aged 18-45 with regular ovulatory cycles and normal uterine cavities undergoing FET after vitrification and warming of one or two day-3 embryos or blastocysts were included. In NC-FET, spontaneous ovulation was detected by serial ultrasounds and serum LH and oestradiol measurements. In AC-FET, oestradiol valerate (6 mg/day) was administered from cycle day 2 (increased to 8 mg/day after 1 week if endometrial thickness was <7 mm) and micronized progesterone (600 mg/day) was initiated once endometrial thickness reached ≥7 mm. MAIN RESULTS AND THE ROLE OF CHANCE: Clinical pregnancy rates with foetal heartbeat were similar between NC-FET and AC-FET in both intention-to-treat (33.1% [94/284] vs 28.9% [80/277]; RR 1.15, 95% CI 0.89, 1.47) and as-treated analyses (34.3% [97/283] vs 31.8% [77/242]; RR 1.08, 95% CI 0.85, 1.38). Cycle cancellations were less frequent in NC-FET (3.9% vs 9.4%; RR 0.41, 95% CI 0.21, 0.82), but NC-FET required more monitoring visits (mean 3.0 vs 2.4; P = 0.0002). Caesarean section was more common after AC-FET (42.7% vs 20.2%; P = 0.003). Other reproductive, obstetric, and perinatal outcomes did not differ significantly between groups. LIMITATIONS, REASONS FOR CAUTION: The sample size was powered to detect a difference in CPR with foetal heartbeat, not for comparing live birth rates or obstetric outcomes. The study was performed in ovulatory women undergoing FET, and therefore, results may not be applicable to other populations. WIDER IMPLICATIONS OF THE FINDINGS: This RCT demonstrates no significant difference in CPR with foetal heartbeat in NC-FET and AC-FET in ovulatory women. Given recent data suggesting higher obstetric risks with AC-FET, NC-FET may be preferred in women with normal ovulatory cycles. STUDY FUNDING/COMPETING INTEREST(S): The study was an investigator-initiated study supported by internal KU Leuven funding (Project number C14/18/106 and C14/24/152) and funding from the Research Foundation Flanders (G.084515N and G.0B1819N to J.V.). K.P. has received grants from Ferring. C.B. has received speaker fees from Gedeon Richter, paid to her institution; travel support from Gedeon Richter, Ferring Pharmaceuticals, and Intuitive; and holds a leadership role as Senior Deputy of the ESHRE SIG Endometriosis and Endometrial Disorders. C.T. has received grants from Merck SA, paid to her institution; consulting fees and speaker fees from Gedeon Richter; travel support from Ferring and Gedeon Richter; and holds leadership roles as Deputy Editor of JNIG and as a board member. A.V. has received speaker fees from Gedeon Richter, paid to his institution, and holds a leadership role as Senior Deputy of the ESHRE SIG Endometriosis and Endometrial Disorders. The other authors declare that there is no conflict of interest to disclose with respect to the content of this article. TRIAL REGISTRATION NUMBER: This trial has been registered at ClinicalTrials.gov (NCT03642665). TRIAL REGISTRATION DATE: 17 July 2018. DATE OF FIRST PATIENT’S ENROLMENT: 27 October 2018.
A population-based study of the co-occurrence of hypospadias and congenital heart defects among boys conceived with in vitro fertilization†.
Richard MA, Luke B, Jaime E … +11 more , Fisher SC, Betancourt D, Yazdy MM, Stone SL, Forestieri NE, Wantman E, Baker VL, Eisenberg ML, Williams C, Sutcliffe AG, Lupo PJ
Hum Reprod · 2026 Jun · PMID 42314752 · Publisher ↗
STUDY QUESTION: Is there an association of co-occurring hypospadias and congenital heart defects (CHD) with method of conception? SUMMARY ANSWER: There is a small excess risk for co-occurring hypospadias and CHD among bo...
STUDY QUESTION: Is there an association of co-occurring hypospadias and congenital heart defects (CHD) with method of conception? SUMMARY ANSWER: There is a small excess risk for co-occurring hypospadias and CHD among boys conceived through in vitro fertilization (IVF) not fully explained by other risk factors. WHAT IS KNOWN ALREADY: We previously reported a higher prevalence of major CHDs among boys born with hypospadias. Our previous analyses leveraged 3.7 million pregnancies recorded through birth defect registries in 11 US states but could not evaluate the role of IVF in the risks for these pregnancy outcomes. Simultaneously, in registry data linked to fertility parameters, we observed excess risk for hypospadias and CHD with conception by IVF. The rarity of co-occurring birth defects requires large sample sizes with key information on both IVF treatment parameters and maternal characteristics to differentiate their roles in birth defects. STUDY DESIGN, SIZE, DURATION: We conducted a population-based study of live births in four US states (Massachusetts, New York, North Carolina, and Texas) from 2004 to 2018 and linked to IVF cycles reported to the Society for Assisted Reproductive Technology Clinic Outcome Reporting System (SART CORS). Information on the treatment cycle included the use of autologous or donor oocytes, fresh or frozen embryos, male infertility diagnosis, and the use of ICSI. Non-IVF births were sampled 10:1 at the time of IVF births. For the purposes of evaluating isolated and co-occurring hypospadias and CHD, our study included 731 838 boys born over the study period. PARTICIPANTS/MATERIALS, SETTING, METHODS: State birth defect registries reporting British Pediatric Association codes were used to identify major birth defects. We formed four outcome groups: (i) boys with CHD alone, (ii) boys with hypospadias alone, (iii) boys with both (hypospadias-CHD), and (iv) boys without birth defects (comparison group). We used unadjusted and adjusted logistic regression to associate IVF conception with each outcome group relative to the common comparison group. Adjustments in each outcome model included backward elimination for birth year, plurality, maternal race/ethnicity, age, education, diabetes, hypertension, and parity. We additionally calculated the prevalence of CHD and hypospadias by IVF treatment parameters. MAIN RESULTS AND THE ROLE OF CHANCE: Among 731 838 boys, we identified 267 who had both hypospadias and a major CHD, 65 conceived through IVF and 202 controls. We observed strong unadjusted associations of IVF with hypospadias (OR 1.49, 95% CI 1.37-1.62), CHD (OR 1.64, 95% CI 1.49-1.81), and hypospadias-CHD (OR 2.63, 95% CI 1.39-4.69). The associations of IVF with isolated hypospadias and isolated CHD were attenuated, but not fully explained, after adjustment for key covariates (hypospadias adjusted OR 1.15, 95% CI 1.03-1.28; CHD adjusted OR 1.14, 95% CI 1.003-1.29). The adjusted association of IVF with hypospadias-CHD suggests a stronger association than for either defect alone (OR 2.16, 95% CI 1.17-3.99), and calculating the observed-expected multiplicative effect supports an excess risk for hypospadias-CHD co-occurrence among boys conceived through IVF (observed ratio 1.65 > expected ratio 1). Among boys with hypospadias, 4.4% (95% CI 4.0-4.8) of boys conceived without IVF also had a CHD, whereas 6.9% (95% CI 5.8-8.0) of boys conceived using IVF also had a CHD. We found that prevalence of CHD was highest among boys with the most severe form of hypospadias among those conceived with IVF (14.1% with CHD, 95% CI 5.5-22.6) or naturally conceived (10.8% with CHD, 95% CI 6.6-15.0). Calculating number needed to screen (NNS) suggests that incorporating information on hypospadias and method of conception could allow for targeted CHD screening opportunities (NNS range: 8-99). Description of fertility treatment parameters suggests the prevalence of hypospadias or CHD among boys conceived using frozen embryos (autologous or donor oocytes) is 2.8%, and is 2.7% among boys whose fathers were diagnosed with male infertility. For co-occurring hypospadias-CHD, the prevalence was highest among boys conceived using donor oocytes (both fresh or frozen embryos) and among boys whose fathers were diagnosed with male infertility, though the absolute risk remained low (0.09-0.12%). When further evaluating male infertility by the use of ICSI, we found that ICSI use increased the risk for isolated or co-occurring defects regardless of an underlying diagnosis of male infertility. LIMITATIONS, REASONS FOR CAUTION: Based on the rarity of co-occurring birth defects (0.08% prevalence of hypospadias-CHD among boys conceived using IVF), we were not able to fully evaluate IVF treatment parameters or subfertility for their role in the co-occurrence of hypospadias and CHD. However, we adjusted for maternal characteristics related to both birth defect risks and fertility. We further sought to characterize treatment parameters that warrant follow-up in future studies of health outcomes among children conceived using IVF. WIDER IMPLICATIONS OF THE FINDINGS: The use of IVF is associated with increased risk for co-occurrence of two of the most common birth defects in boys, hypospadias and CHD, but the absolute risk remains low. Additional investigations of combinations of birth defects, other childhood outcomes in relationship to method of conception, and mechanisms to explain these associations are warranted. STUDY FUNDING/COMPETING INTEREST(S): This project was supported by grant R01 HD112081 from the National Institute of Child Health and Human Development, USA (Barbara Luke and Philip Lupo, Multiple Principal Investigators). The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institute of Child Health and Human Development, or the National Institutes of Health, nor any State Departments of Health which contributed data. M.L.E. declares stock/shares from their role as advisor to Swim Club, VSeat, Doveras, Legacy, Illumicell, Hannah, and HisTurn. E.W. is employed by Redshift Technologies, Inc., which is the data vendor for SART, and has received consulting fees from SART. The other authors declare no competing interests. TRIAL REGISTRATION NUMBER: N/A.
Endometriosis and the mental health burden: a registry-based study of redeemed prescriptions and psychiatric hospital contacts before and after diagnosis of endometriosis.
Josiasen M, Melgaard A, Bech BH … +2 more , Hansen KE, Rytter D
Hum Reprod · 2026 Jun · PMID 42295201 · Publisher ↗
STUDY QUESTION: Do women with endometriosis have a higher use of antidepressants and anxiolytics, and more frequent psychiatric hospital contacts due to depression or anxiety, before and after receiving their endometrios...
STUDY QUESTION: Do women with endometriosis have a higher use of antidepressants and anxiolytics, and more frequent psychiatric hospital contacts due to depression or anxiety, before and after receiving their endometriosis diagnosis, compared with women without endometriosis? SUMMARY ANSWER: Women with an endometriosis diagnosis redeemed more prescriptions for antidepressants and anxiolytics and had more psychiatric hospital contacts due to depression or anxiety both before and after endometriosis diagnosis compared with women without endometriosis. WHAT IS KNOWN ALREADY: Endometriosis is associated with an increased risk of anxiety and depression, but evidence on patterns before and after diagnosis is limited. STUDY DESIGN, SIZE, DURATION: This national, retrospective, registry-based study included 136 842 Danish women aged 15-55 years between 1 January 2000 and 31 December 2019. Each woman was followed for up to 10 years before and 10 years after the index date. PARTICIPANTS/MATERIALS, SETTING, METHODS: We identified 22 807 women with a first-time hospital-based diagnosis of endometriosis using density sampling. Each woman with endometriosis was matched on age at the date of diagnosis (index date) to five women without diagnosed endometriosis (n = 114 035). Information on redeemed prescriptions and psychiatric hospital contacts was obtained from national registries and compared using negative binomial regression. MAIN RESULTS AND THE ROLE OF CHANCE: Women with an endometriosis diagnosis redeemed more prescriptions for antidepressants and anxiolytics and had more psychiatric hospital contacts due to depression or anxiety in the 20-year study window. Adjusted incidence rate ratios (IRRs) for antidepressants were 1.29 (95% CI 1.23-1.36) before and 1.40 (95% CI 1.34-1.47) after the index date, when adjusting for age, region of residence, highest educational level, household type, labor market affiliation, origin, and parity. For anxiolytics, IRRs were 1.16 (95% CI 1.05-1.28) before and 1.46 (95% CI 1.29-1.64) after the index date. For psychiatric hospital contacts, IRRs were 1.28 (95% CI 1.09-1.49) before and 1.41 (95% CI 1.22-1.63) after the index date. The association for antidepressants was quite stable over the study period, while the association for anxiolytics showed a significant increase right before the index date, and the associations for psychiatric hospital contacts increased around 5 years before the index date. LIMITATIONS, REASONS FOR CAUTION: It was not possible to include women who were treated for suspected endometriosis in general practice or women diagnosed with endometriosis solely by private gynecologists. Therefore, the results are only applicable to hospital-based diagnoses of endometriosis. Not all women with depression or anxiety receive pharmacological treatment or hospital-based care. Hence, we might not have identified all women with depression and anxiety. However, we expect this under-ascertainment of depression and anxiety to be similar among women with and without endometriosis and thereby independent of endometriosis diagnosis. WIDER IMPLICATIONS OF THE FINDINGS: These findings align with existing literature but provide new insights into the mental health burden in the pre-diagnostic period. Future studies should investigate the causal pathways and potential mediating factors of the association. A better understanding of the link between endometriosis, depression, and anxiety and improved awareness can help healthcare professionals identify affected women more easily and improve treatment strategies. STUDY FUNDING/COMPETING INTEREST(S): This article is supported by grants from the project 'Finding Endometriosis using Machine Learning' (FEMaLe/101017562) funded by the European Union's Horizon 2020 research and innovation program, and from Helsefonden (21-B-0141). The authors have no conflicts of interest to declare. TRIAL REGISTRATION NUMBER: N/A.
Oocyte aging, the mitochondrial downward spiral, and the path to responsible innovation.
Mastenbroek S, Hamer G
Hum Reprod · 2026 Jun · PMID 42275567 · Publisher ↗
Abstract loading — click title to view on PubMed.
ESHRE PGT Consortium data collection XXII-XXIV: PGT analyses from 2019 to 2021†.
Spinella F, Christopikou D, De Rycke M … +7 more , Florensa M, Heijligers M, Toft CLF, Constantinou E, Viville S, Goossens V, Dimitriadou E
Hum Reprod · 2026 Jun · PMID 42263100 · Publisher ↗
STUDY QUESTION: What are the trends and developments in preimplantation genetic testing (PGT) in 2019-2021 as compared to previous years? SUMMARY ANSWER: The main trend observed in the 22nd to 24th datasets on PGT is tha...
STUDY QUESTION: What are the trends and developments in preimplantation genetic testing (PGT) in 2019-2021 as compared to previous years? SUMMARY ANSWER: The main trend observed in the 22nd to 24th datasets on PGT is that the implementation of trophectoderm biopsy with comprehensive whole-genome testing is most often applied for PGT for aneuploidies (PGT-A) and concurrent PGT-M/SR/A, while for PGT for monogenic disorders (PGT-M) and PGT for chromosomal structural rearrangements (PGT-SR), single-cell targeted testing with PCR and FISH still prevails. WHAT IS KNOWN ALREADY: Since it was established in 1997, the PGT Consortium within the European Society for Human Reproduction (ESHRE) has been collecting and analysing data from mainly European PGT centres. To date, 21 datasets and an overview of the first 10 years of data collections have been published. STUDY DESIGN, SIZE, DURATION: The data for PGT analyses performed between 1 January 2019 and 31 December 2021 with a 2-year follow-up, after analysis were provided by participating centres on a voluntary basis. Data were collected using an online platform; the data reported on a per genetic analysis basis as opposed to the former cycle-based database. PARTICIPANTS/MATERIALS, SETTING, METHODS: Data on biopsy methods, diagnostic technologies, genetic diseases, and clinical outcomes were submitted by 45 centres. Records with multiple PGT-M and/or PGT-SR analyses or inconsistent PGT data were excluded. Embryo transfers performed within 2 years of analysis were included to calculate clinical outcomes, including stratification by biopsy day and cumulative pregnancy rates. Data analysis, calculations, and preparation of figures and tables were carried out by expert co-authors. MAIN RESULTS AND THE ROLE OF CHANCE: A total of 14 731 PGT analyses were reported: 3782 for PGT-M, 874 for PGT-SR, 8492 for PGT-A, and 1583 for combined PGT-M/SR with PGT-A, henceforth referred to as combined testing. The use of trophectoderm biopsy increased for PGT-M (33% in 2018 to 49% in 2019-2021), remained stable for PGT-SR (33-36%), and was consistently high for PGT-A (98%) and combined cases (96%). The adoption of genome-wide or targeted technologies following whole-genome amplification (WGA) remained relatively stable for PGT-M (12-14%) and PGT-SR (44-45%), while genome-wide analysis performed on whole-genome-amplified DNA became the standard for PGT-A (99%). The combination of PGT-M/SR with PGT-A was performed either by applying a technology following WGA (74%) or by targeted methods (PCR or FISH, 24%). Diagnostic efficiency was 87% for PGT-M, 92% for PGT-SR, 98% for PGT-A, and 90% for combined testing. Pregnancy rates per embryo transfer were 26% for PGT-M, 26% for PGT-SR, 35% for PGT-A, and 36% for combined testing. Across all groups, Day 5 biopsies yielded better outcomes than Day 3 or Day 6 biopsies. LIMITATIONS, REASONS FOR CAUTION: The findings refer to data submitted by 45 participating centres and do not reflect global trends in PGT. Information on the health of babies born following PGT was not included in this manuscript. It should also be noted that final outcomes may be influenced by the number and type of centres submitting data in different years. WIDER IMPLICATIONS OF THE FINDINGS: The Consortium datasets provide a valuable resource for monitoring trends in PGT practice, both from a technological perspective and in terms of clinical outcomes. STUDY FUNDING/COMPETING INTEREST(S): The study has no external funding, and all costs are covered by ESHRE. There are no competing interests declared. TRIAL REGISTRATION NUMBER: NA.
Novel variants in YTHDC2 cause non-obstructive azoospermia by disrupting the mitotic-to-meiotic transition in humans and mice.
Zhi A, Li M, Zubair M … +13 more , Abbas M, Shah W, Mansoor A, Rahim F, Ali I, Raza Y, Murtaza G, Ahmad N, Abideen Z, Jiang H, Shi B, Zhang H, Shi Q
Hum Reprod · 2026 Jun · PMID 42249589 · Publisher ↗
STUDY QUESTION: Do variants in YTH N6-methyladenosine RNA binding protein C2 (YTHDC2) cause male infertility in humans, and what is the underlying pathogenic mechanism? SUMMARY ANSWER: Biallelic pathogenic missense varia...
STUDY QUESTION: Do variants in YTH N6-methyladenosine RNA binding protein C2 (YTHDC2) cause male infertility in humans, and what is the underlying pathogenic mechanism? SUMMARY ANSWER: Biallelic pathogenic missense variants in YTHDC2 disrupt the mitotic-to-meiotic transition, causing meiotic arrest and non-obstructive azoospermia (NOA) or severe oligozoospermia in humans. WHAT IS KNOWN ALREADY: YTHDC2 is a male germ cell-specifically expressed RNA helicase essential for meiotic progression. In mice, loss of Ythdc2 leads to meiotic arrest at the early prophase. However, clinical evidence linking YTHDC2 variants to human male infertility and the underlying mechanisms involved remains to be established. STUDY DESIGN, SIZE, DURATION: This study utilized a large cohort comprising 56 consanguineous families and 89 sporadic infertile men diagnosed with NOA or severe oligozoospermia. Through extensive genetic screening, we specifically identified five infertile men from three unrelated families who harbored candidate pathogenic variants in the YTHDC2 gene. The overall study design encompassed genetic screening followed by in vivo functional validation using a knock-in mouse model. PARTICIPANTS/MATERIALS, SETTING, METHODS: Whole-exome sequencing (WES) and bioinformatic analyses were performed on the patient cohort to screen for candidate pathogenic variants. Human meiotic defects were characterized via histological analyses and immunofluorescence staining of testicular sections. To validate the pathogenicity of the identified variant, a knock-in mouse model harboring the equivalent variant found in patients was generated by CRISPR/Cas9 technology, and analyzed for spermatogenesis and meiosis using spermatocyte spreading and immunofluorescence staining, quantitative real-time PCR, and western blotting. MAIN RESULTS AND THE ROLE OF CHANCE: Two homozygous missense variants in YTHDC2 were identified in four NOA patients from two unrelated consanguineous families (MT1: c.3491A>T, p. E1164V; MT2: c.2639G>A, p. R880H), and compound-heterozygous missense variants were identified in a sporadic patient with severe oligozoospermia (MT3: c.1145A>G, p. D382G; MT4: c.292A>G, p. R98G). The MT1 variant (p.E1164V) is not located in any annotated domains, the other three variants reside within known functional domains of YTHDC2 protein. The knock-in mouse model carrying the MT1 variant recapitulated the patient's phenotype, with both exhibiting meiotic prophase arrest during spermatogenesis. Mechanistically, significantly decreased levels of MEIOC and RBM46, two YTHDC2-interacting proteins required for meiotic transcriptome reprogramming, were observed in the patient's testes. Concurrently, mitotic cell cycle regulators such as CCNA2, CCND1, and WEE1 were aberrantly upregulated in the patient's testicular cells expressing meiosis markers, indicating a failure to silence the mitotic program upon meiotic entry. LIMITATIONS, REASONS FOR CAUTION: This study is limited by the small sample size of patients with pathogenic YTHDC2 variants. While the MT1 variant was functionally validated in vivo, the specific pathogenic mechanisms of the other variants identified require further investigation. WIDER IMPLICATIONS OF THE FINDINGS: Our findings provide direct clinical evidence establishing the pathogenicity of YTHDC2 variants in human NOA. The study reveals a conserved role for YTHDC2 in safeguarding the mitotic-to-meiotic transition by suppressing mitotic gene expression while maintaining the meiotic program. These findings expand the genetic spectrum of male infertility and suggest YTHDC2 screening as a promising approach for the genetic diagnosis of male infertility. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by the National Key Research and Developmental Program of China (2022YFA0806303 to H.Z., and 2024YFC2706801 to H.J.); the National Natural Science Foundation of China (32470898 to H.Z., W2412028 and 32330032 to Q.S., 32470915 to B.S.); the State Key Laboratory of Reproductive Medicine and Offspring Health, Nanjing Medical University (SKLRM-K202405); and the Open Research Project of Fuyang Normal University (FYKFKT24023). The authors declare no competing interests. TRIAL REGISTRATION NUMBER: N/A.
Remodeling lesions locate at sites of strong extravillous trophoblast invasion and are associated with neutrophil presence in the human first-trimester decidua.
Moser G, Kummer D, Gruber M … +13 more , Hartmann T, Forstner D, Guettler J, Siwetz M, Sundl M, Daxboeck C, Kienesberger S, Kargl J, Deutsch A, Fessler J, Huppertz B, Feichtinger J, Gauster M
Hum Reprod · 2026 Jun · PMID 42248577 · Publisher ↗
STUDY QUESTION: Does the degree of extravillous trophoblast (EVT) invasion influence decidual tissue architecture and immune cell distribution in first-trimester decidua? SUMMARY ANSWER: Areas of pronounced morphological...
STUDY QUESTION: Does the degree of extravillous trophoblast (EVT) invasion influence decidual tissue architecture and immune cell distribution in first-trimester decidua? SUMMARY ANSWER: Areas of pronounced morphological changes have been identified at sites of strong EVT invasion in the decidua basalis-defined as 'remodeling lesions'-and are associated with a substantially reshaped immune cell landscape. WHAT IS KNOWN ALREADY: During early human placental development, EVTs invade the decidua to facilitate placental attachment and nutrient supply to the fetus. EVT-driven decidual tissue restructuring and vascular adaptation are essential for establishing a functional fetal-maternal interface, to which the decidual microenvironment is thought to contribute substantially. However, the precise impact of the degree of EVT invasion on decidual architecture and immune cell distribution remains poorly understood, underscoring the need to elucidate how varying EVT abundances shape the morphological and cellular landscape of the decidual microenvironment. STUDY DESIGN, SIZE, DURATION: First-trimester decidual tissue (n = 23, gestational age Weeks 7-9) was analyzed from women undergoing elective terminations of pregnancy between 2011 and 2023. Additionally, hematoxylin and eosin-stained sections from archival specimens (n = 11), obtained from three different sources, were included in the study. PARTICIPANTS/MATERIALS, SETTING, METHODS: Matched first-trimester decidua basalis and decidua parietalis samples from the same donors were analyzed through a comprehensive approach combining spatial transcriptomics, histomorphological characterization, and quantitative image analysis. Decidua sections were (i) categorized according to the degree of invasion, (ii) subjected to spatial transcriptomics, including integration with a previously published single-cell RNA-seq dataset, and (iii) quantitatively assessed on the protein level for selected immune cell populations with immunostaining and semi-automated image analysis. The study was complemented by (iv) an observer-based histological evaluation and (v) comprehensive staining series of consecutive decidua sections. MAIN RESULTS AND THE ROLE OF CHANCE: Analyses revealed a characteristic tissue restructuring of the decidua and distinct spatial patterns of immune cell abundance in relation to the degree of EVT invasion. In strongly invaded decidual areas, we identified regions with pronounced morphological changes-defined as 'remodeling lesions'. These remodeling lesions typically displayed compromised tissue integrity, eroded blood vessels, extravasal erythrocytes, fibrin deposits, and a distinct gene expression profile, reflecting coagulation, fibrinolysis, and tissue restructuring. While we observed a decline in local immune cell populations-specifically T cells, macrophages, and decidual natural killer cells-with increasing EVT density, neutrophils were almost exclusively located within or in close proximity to remodeling lesions, indicating a substantially reshaped immune landscape. LARGE SCALE DATA: Spatial transcriptomics data are available in the Gene Expression Omnibus repository under accession number GSE301306. LIMITATIONS, REASONS FOR CAUTION: Studies using first-trimester placental tissues from elective terminations are inherently limited by surgical disruption of the intact (in toto) anatomical architecture of the tissue and the unknown pregnancy outcome. Spatial transcriptomics was performed on a limited number of tissue sections, and histological tissue sections represent just a snapshot, highlighting the limitations of such tissue-based analyses. WIDER IMPLICATIONS OF THE FINDINGS: While blood leakage into the stromal tissue compartment has typically been documented for pathological conditions-such as large atherosclerotic plaques and tumors-we report such a scenario under physiological conditions for the early invaded decidua. We propose that strong EVT invasion induces remodeling lesions in the decidua basalis and also shapes the surrounding immune cell landscape. We further suggest that the occurrence of these remodeling lesions contributes to the establishment of a stable yet flexible basal plate and is thus necessary for a reliable connection between mother and placenta/fetus. It can be speculated that inadequate decidual tissue restructuring and vascular adaptation lead to pregnancy pathologies and complications such as placental abruption. STUDY FUNDING/COMPETING INTEREST(S): G.M. was supported by the Austrian Science Fund (FWF): PAT9611123. M.G. was supported by the Austrian Science Fund (FWF): 10.55776/P35118 and 10.55776/I6907. This project has received funding from the European Union's Horizon Europe research and innovation programme under the Marie Skłodowska-Curie grant agreement No 101169308 (funding supported M.G.). M.G., G.M., and J.F. were supported by the Medical University of Graz through the PhD program MolMed. J.F. and G.M. were supported by the COMET center acib: Next Generation Bioproduction (Project #98.311 and #95.802) is funded by BMIMI, BMWET, SFG, Standortagentur Tirol, Government of Lower Austria and Vienna Business Agency in the framework of COMET-Competence Centers for Excellent Technologies. The COMET-Funding Program is managed by the Austrian Research Promotion Agency FFG. The authors declare that they have no conflicts of interest related to this work.
Correction to: Revisiting the promise and pitfalls of mitochondrial replacement therapies.
Hum Reprod · 2026 Jun · PMID 42243637 · Publisher ↗
Abstract loading — click title to view on PubMed.
Deep immune profiling of endometrial and peripheral blood cells in endometriosis.
Kisovar A, Buttenschoen M, Obrigewitch Q … +6 more , Powell K, Klenerman P, Zondervan K, Becker CM, Granne IE, Southcombe JH
Hum Reprod · 2026 Jun · PMID 42243626 · Publisher ↗
STUDY QUESTION: How is endometrial and systemic immunity modulated throughout the menstrual cycle and are there changes in women with endometriosis? SUMMARY ANSWER: Endometriosis is associated with reduced endometrial ea...
STUDY QUESTION: How is endometrial and systemic immunity modulated throughout the menstrual cycle and are there changes in women with endometriosis? SUMMARY ANSWER: Endometriosis is associated with reduced endometrial early natural killer (NK) cells and increased mucosal-associated invariant T (MAIT)-like CD8+ T cells, with cyclical variation. WHAT IS KNOWN ALREADY: The endometrial mucosa contains innate and adaptive immune cells that fluctuate across the menstrual cycle. Immune dysregulation is found in endometriosis, however few studies have broadly assessed endometrial immune single-cell proteome phenotypes. STUDY DESIGN, SIZE, DURATION: This observational cross-sectional immune phenotyping study included 40 participants (28 with surgically confirmed endometriosis and 12 controls). PARTICIPANTS/MATERIALS, SETTING, METHODS: Endometrial and peripheral blood samples were analysed by spectral flow cytometry using a 36-parameter immune phenotyping panel and a 13-parameter MAIT-specific panel. Totals of 1 950 292 circulating and 1 023 215 endometrial immune cells were profiled. Full-thickness uterine biopsies (n = 3) underwent multiplex immunohistochemical imaging to assess spatial organization across the menstrual cycle. MAIN RESULTS AND THE ROLE OF CHANCE: Compared with controls, patients with endometriosis exhibited decreased endometrial early NK cells (P.adj = 0.006, log2FC = -1.369) and increased MAIT-like CD8+ T cells (CD161+CD8+CD3+) (P.adj = 0.033, log2FC = 1.415). The MAIT cells (CD161+Va7.2+CD3+) peaked during ovulation and the implantation window (P.adj < 0.05). Peripheral immunity also showed cyclical variation with increased early NK cells (P.adj = 0.001, log2FC = 1.052) and decreased effector CD4 T (P.adj = 0.002/log2FC = -2.010) and effector CD8 T cells (P.adj = 0.002, log2FC = -1.180) in the endometriosis group. LIMITATIONS, REASONS FOR CAUTION: The cytometric panel design was biased towards acquired immunity, and the endometriosis patient sample size prevented subtype analysis. WIDER IMPLICATIONS OF THE FINDINGS: MAIT cell dysregulation represents a novel feature of endometriosis, potentially contributing to subfertility and providing new avenues for therapeutic development. STUDY FUNDING/COMPETING INTEREST(S): This study was funded by the Nuffield Department of Women's and Reproductive Health, University of Oxford, and also supported from the University of Oxford Medical Sciences HIDI Internal Fund Award (0010398), Academy of Medical Science Award (SBF007\100078) and, British Society for Immunology Career Enhancing Grant. C.M.B. has a consultancy role with ObsEva, Theramex, Roche Diagnostics, Sumitovant, Gedeon Richter, Gesyntha, and they have Research Grants from Bayer, Gesyntha, and Serac Life Services. K.Z. is a Board member (non-remunerated) of the World Endometriosis Research Foundation. She has consultancy status with Roche Diagnostics and Gedeon Richter. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. TRIAL REGISTRATION NUMBER: N/A.
A comprehensive review for defining cut-off values of oocyte, zygote, and embryo maturation arrest (OZEMA) due to maternal-effect genes: towards the establishment of clinical criteria.
Van Der Kelen A, Okutman Ö, Serdarogullari M … +10 more , Dequeker BJH, Javey E, Capalbo A, Lang C, Kakourou G, Konstantinou E, Mulas F, Sermon K, Verpoest W, Viville S
Hum Reprod · 2026 Jun · PMID 42222905 · Publisher ↗
STUDY QUESTION: Which clinical definition of oocyte, zygote, and embryo maturation arrest (OZEMA) merits genetic investigation? SUMMARY ANSWER: The diagnosis OZEMA is considered when: only two mature oocytes are obtained...
STUDY QUESTION: Which clinical definition of oocyte, zygote, and embryo maturation arrest (OZEMA) merits genetic investigation? SUMMARY ANSWER: The diagnosis OZEMA is considered when: only two mature oocytes are obtained from six cumulus-oocyte complexes (COCs), only one zygote is obtained from six metaphase II oocytes (MIIs), or no blastocysts are obtained from six two-pronuclei zygotes (2PNs). WHAT IS KNOWN ALREADY: OZEMA is a cover term for various defects, including oocyte abnormalities, fertilization failure, cleavage arrest, and abnormal embryo development, often associated with specific genetic variants. While some variants in specific genes are associated with distinct phenotypes, others commonly display variable expressivity across cycles and patients. Currently, there are no standardized cut-off values defining the OZEMA disorder. STUDY DESIGN, SIZE, DURATION: This is a pooled secondary analysis of aggregated data extracted from published studies. A PubMed search was conducted to identify peer-reviewed original studies in the human reporting genetically confirmed cases of OZEMA. Studies were included if patients had displayed OZEMA in at least two medically assisted reproduction (MAR) cycles. In total, clinical and genetic data were extracted from 132 publications, including 521 patients and 39 genes. PARTICIPANTS/MATERIALS, SETTING, METHODS: Clinical phenotypes from 132 publications were classified into oocyte maturation arrest (OMA), zona pellucida abnormality (ZPA), fertilization failure (FF), zygote arrest (ZA), early embryo arrest (EEA), mixed phenotypes, and other abnormalities. Data at both cycle and patient levels were evaluated using ratios corresponding to key developmental milestones (ratio of MII/COC, 2PN/MII, cleaved embryos/2PN, blastocysts/2PN). These ratios were compared with the expected values Vienna Consensus binomial modeling to assess whether the observed proportions were significantly lower than the established competency thresholds (P < 0.05). MAIN RESULTS AND THE ROLE OF CHANCE: This review identified 39 OZEMA-associated genes. Analysis of 668 cycles from 253 patients carrying variants in 35 genes showed abnormal developmental outcomes in 93%, based on binomial modeling. Reliable diagnosis of OZEMA requires a sufficient number of COCs, MIIs, or 2PNs to distinguish true developmental impairment from normal biological variation. When these minimal thresholds are not reached, statistical power is insufficient to reliably confirm developmental arrest. Based on our findings, OZEMA is diagnosed when only two or fewer mature oocytes form from six COCs, one or no zygote forms from six MIIs, or no blastocysts form from six 2PNs. LIMITATIONS, REASONS FOR CAUTION: This study is limited by its retrospective design and reliance exclusively on published data from women selected for poor IVF outcomes and the presence of variants in candidate OZEMA genes, which may introduce publication bias and reporting heterogeneity. Therefore, this study does not report on genetic yield from women with normal IVF outcomes. Variability in clinical protocols and genetic testing methods, limited ancestral diversity, and missing data across studies could contribute to confounding factors. WIDER IMPLICATIONS OF THE FINDINGS: This study is an important step toward understanding the clinical and genetic spectrum of OZEMA, highlighting the variability of and overlap between affected subphenotypes. Establishing cut-off values against consensus oocyte and embryo developmental competency levels provides a benchmark for more accurate classification. Importantly, this represents the most comprehensive analysis of published OZEMA cases with genetic variants to date, strengthening the reliability of the proposed clinical thresholds. These findings may guide personalized treatment strategies and genetic counseling, ultimately improving outcomes in MAR. STUDY FUNDING/COMPETING INTEREST(S): No funding was received for this study. The authors declare no competing interests. REGISTRATION NUMBER: Not applicable.
Psychological and physical effects of short-term discontinuation of feminizing gender-affirming hormone therapy among older transgender women: a within-subject clinical trial.
van Heesewijk JO, Geels RES, den Heijer M … +2 more , Dreijerink KMA, Kreukels BPC
Hum Reprod · 2026 May · PMID 42217199 · Publisher ↗
STUDY QUESTION: What are the psychological and physical effects of short-term discontinuation and subsequent reinitiation of feminizing gender-affirming hormone therapy (fGAHT) in older transgender women receiving long-t...
STUDY QUESTION: What are the psychological and physical effects of short-term discontinuation and subsequent reinitiation of feminizing gender-affirming hormone therapy (fGAHT) in older transgender women receiving long-term therapy? SUMMARY ANSWER: Short-term fGAHT discontinuation increased estradiol deprivation-related symptoms while reinitiation showed opposite effects, and psychological well-being was not affected. WHAT IS KNOWN ALREADY: With growing numbers of older transgender individuals, questions arise regarding the necessity of long-term continuation of fGAHT, particularly since cisgender women have low estradiol concentrations after menopause. However, data on the effects of fGAHT discontinuation in older transgender women are lacking. STUDY DESIGN, SIZE, DURATION: This within-subject clinical trial included 19 participants and was conducted in 2024 and 2025. PARTICIPANTS/MATERIALS, SETTING, METHODS: Transgender women aged ≥55 years (median age 66 years), on fGAHT for ≥5 years (median 23 years), were recruited from the Center of Expertise on Gender Dysphoria, Amsterdam, the Netherlands. Estradiol deprivation-related symptoms, sleep quality, quality of life, mental health, and body image were assessed via questionnaires at baseline, 12 weeks after fGAHT discontinuation, and 12 weeks after reinitiation. Outcomes were analyzed using multilevel models. Differences were considered substantial based on 95% CIs and ≥10% change between measurements. Perceived psychological and physical changes and willingness to permanently discontinue fGAHT were also assessed via open-ended questions. MAIN RESULTS AND THE ROLE OF CHANCE: Experiences varied, but after discontinuation, unfavorable mean changes were found for somato-vegetative symptoms (β 0.4, 95% CI -0.7 to 1.5, %Δ 11.5), sleep disturbance (OR 1.6, 95% CI 0.2 to 10.3, %Δ 10.7), and happiness to a lesser extent (β -0.5, 95% CI -0.9 to -0.1, %Δ -6.4). Reinitiation showed favorable changes for somato-vegetative and total estradiol deprivation-related symptoms (β -0.8, 95% CI -1.9 to 0.3, %Δ -22.1; β -3.0, 95% CI -4.8 to -1.2, %Δ -36.3), total sleep quality (β -0.7, 95% CI -1.5 to 0.2, %Δ -10.3), sleep onset latency (β -2.8, 95% CI -8.1 to 2.5, %Δ -14.9), and sleep disturbance (OR 0.1, 95% CI 0.0 to 1.0, %Δ -46.6). No mean differences were found for depressive symptoms, anxiety, quality of life, or body satisfaction. Perceived psychological and physical changes were diverse and generally aligned with known menopause-related changes. One participant wished to permanently stop fGAHT. LIMITATIONS, REASONS FOR CAUTION: The study was not blinded or placebo-controlled, which may have introduced expectancy bias. The small clinical sample may limit generalizability to the broader population of transgender women. WIDER IMPLICATIONS OF THE FINDINGS: Transgender women, like cisgender women, experience symptoms in response to estradiol deprivation, which may support the continuation of fGAHT in older age. This study did not identify psychological or body image barriers to discontinuation for those who wish to stop. STUDY FUNDING/COMPETING INTEREST(S): Funded by an Amsterdam UMC PhD scholarship to J.O.v.H. and by departmental funds from the Department of Endocrinology and Metabolism, Amsterdam UMC, location VUmc. K.M.A.D. declares receipt of consulting fees from Recordati, paid to their institution. The other authors declare no competing interests. TRIAL REGISTRATION NUMBER: This study was registered in the EU Clinical Trials Register (EUCTR2023-505143-39-00).
Third-generation sequencing enables accurate preimplantation genetic testing of monogenic defects for de novo variants and incomplete pedigrees.
Fang Y, Wang Y, Zhan Y … +14 more , Zhu X, Kuo Y, Wang N, Guan S, Song S, Ma M, Lian Y, Huang J, Zhi X, Liu P, Li R, Yan Z, Qiao J, Yan L
Hum Reprod · 2026 May · PMID 42217180 · Publisher ↗
STUDY QUESTION: Can preimplantation genetic testing for monogenic defects (PGT-M) be achieved by performing third-generation sequencing (TGS) only on the proband for families with de novo variants or incomplete pedigrees...
STUDY QUESTION: Can preimplantation genetic testing for monogenic defects (PGT-M) be achieved by performing third-generation sequencing (TGS) only on the proband for families with de novo variants or incomplete pedigrees? SUMMARY ANSWER: Whole-genome TGS facilitates a simplified PGT-M workflow by establishing reliable haplotypes solely from proband sequencing involving de novo variants or incomplete pedigrees. WHAT IS KNOWN ALREADY: PGT-M enables the accurate exclusion of embryos carrying pathogenic variants. However, its application to de novo variants or incomplete pedigrees is hindered by haplotype phasing. Moreover, direct variant detection suffers from detection failure and erroneous genotyping due to uneven coverage and allele dropout caused by whole-genome amplification. Current solutions, such as gamete or embryo analysis and targeted TGS, remain constrained by procedural complexity and lack of universality across different genes and mutation types. STUDY DESIGN, SIZE, DURATION: This prospective study enrolled 16 families requiring PGT-M with de novo variants or incomplete pedigrees at the Reproductive Medicine Center of Peking University Third Hospital from July 2023 to August 2025. PARTICIPANTS/MATERIALS, SETTING, METHODS: This study included 9 families with incomplete pedigrees and 7 families with de novo variants, covering 10 distinct disease-causing genes or regions. To assess the capability of TGS for haplotype phasing, we evaluated its performance regarding genomic coverage and the retrieval of informative single-nucleotide polymorphisms (SNPs). Haplotypes were constructed using proband TGS data, and linkage analysis was performed by integrating linked heterozygous SNPs with next-generation sequencing data from the couple and embryos to determine pathogenic status. Subsequently, we developed a simplified strategy that inferred inheritance by comparing heterozygous SNPs from the proband's haplotype directly against corresponding homozygous sites in the embryos. The diagnostic outcomes of this simplified workflow were statistically evaluated and compared with those of the standard TGS strategy to assess concordance. MAIN RESULTS AND THE ROLE OF CHANCE: Phase blocks generated by TGS achieved >75% coverage for the vast majority of OMIM genes, most of which contained more than 100 heterozygous informative SNPs located in the gene body and their 1 Mb flanking regions, indicating a wide range of applicability in a variety of gene variants. Haplotypes were successfully constructed for all 16 enrolled families using TGS data, with 14 families having completed embryo testing, while the 2 families withdrew due to personal reasons. To date, prenatal diagnosis via amniocentesis in three families has confirmed the fetuses to be free of pathogenic variants. A simplified strategy was further applied to 14 families that completed the embryo testing process. This approach achieved applicability rates of 91.9% and 80.0% in embryos from non-D4Z4 and D4Z4 families, respectively. While diagnosis was precluded in a subset of embryos due to aneuploidy or insufficient SNP retrieval, the diagnostic outcomes for all remaining embryos were fully concordant with those of the standard TGS strategy. LIMITATIONS, REASONS FOR CAUTION: The applicability of this approach is primarily contingent upon embryo chromosomal euploidy and sufficient retrieval of informative SNPs. Additionally, the relatively high cost of whole-genome TGS remains a barrier to widespread adoption. Given the limited cohort size (n = 16) of this study, the applicability of this method necessitates further validation in larger clinical populations. WIDER IMPLICATIONS OF THE FINDINGS: Direct haplotype construction via proband whole-genome TGS provides an effective clinical strategy to expand the applicability of PGT-M, particularly for families with de novo variants or incomplete pedigrees. Furthermore, the simplified TGS workflow demonstrates the potential to improve clinical efficiency and reduce costs relative to the standard TGS protocol within its applicable scope. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by the National Natural Science Foundation of China (82125013, 82288102, 825B2046). The authors declare no competing interests. TRIAL REGISTRATION NUMBER: N/A.
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