The human zona pellucida (ZP) is a highly organized glycoprotein matrix that encircles oocytes and plays an essential role in successful reproduction. Previous studies have reported that mutations in human ZP1, ZP2 and ZP3 influence their functions and result in a lack of ZP or in an abnormal oocytes and empty follicle syndrome, which leads to female infertility. Here, we performed whole‐exome sequencing in two probands with primary infertility whose oocytes lacked a ZP, and we identified a heterozygous mutation in ZP1 (NM_207341:c.326G>A p.Arg109His), which is situated in the N‐terminus, and a heterozygous mutation in ZP3 (NM_001110354:c.400G>A p.Ala134Thr), which is situated in the ZP domain. The effects of the mutations were investigated through structure prediction and in vitro studies in HeLa cells. The results, which were in line with the phenotype, suggested that these mutations might impede the function of cross‐linking and secretion of ZP proteins. Our study showed that the two mutations in ZP1 and ZP3 influenced the formation of the ZP, causing female infertility. Meanwhile, these data highlight the importance of the ZP1 N‐terminus in addition to the conserved domains for ZP1 function and ZP formation. Additionally, the patient with the ZP1 mutation delivered a baby following intracytoplasmic sperm injection (ICSI); thus, we suggest the targeted genetic diagnosis of ZP genes to choose appropriate fertilization methods and improve the success rate of assisted reproductive technology (ART) treatments.

Purpose: To elucidate the characterization of extracellular vesicles (EVs) in the follicular fluid-derived extracellular vesicles (FF-EVs) and discover critical molecules and signaling pathways associating with the etiology and pathobiology of PCOS, the differentially expressed miRNAs (DEmiRNAs) and differentially expressed proteins profiles (DEPs) were initially explored and combinedly analyzed. Methods: First, the miRNA and protein expression profiles of FF-EVs in PCOS patients and control patients were compared by RNA-sequencing and tandem mass tagging (TMT) proteomic methods. Subsequently, Gene Ontology and the Kyoto Encyclopedia of Genes and Genomes were used to analyze the biological function of target genes of DEmiRNAs and DEPs. Finally, to discover the functional miRNA-target gene-protein interaction pairs involved in PCOS, DEmiRs target gene datasets and DEPs datasets were used integratedly. Results: A total of 6 DEmiRNAs and 32 DEPs were identified in FF-EVs in patients with PCOS. Bioinformatics analysis revealed that DEmiRNAs target genes are mainly involved in thiamine metabolism, insulin secretion, GnRH, and Apelin signaling pathway, which are closely related to the occurrence of PCOS. DEPs also closely related to hormone metabolism processes such as steroid hormone biosynthesis. In the analysis integrating DEmiRNAs target genes and DEPs, two molecules, GRAMD1B and STPLC2, attracted our attention that are closely associated with cholesterol transport and ceramide biosynthesis, respectively. Conclusion: Dysregulated miRNAs and proteins in FF-EVs, mainly involving in hormone metabolism, insulin secretion, neurotransmitters regulation, adipokine expression, and secretion, may be closely related to PCOS. The effects of GRAMD1B and STPLC2 on PCOS deserve further study.

Comparative transcriptome analysis reveals a regulatory network of microRNA-29b during mouse early embryonic development

This study was to evaluate the relationship between morphology or developmental rate of blastocysts and ploidy status in preimplantation genetic testing for aneuploidy (PGT-A) cycles, or live birth rates in single euploid frozen-thawed embryo transfer (FET) cycles. We focused on infertile patients who underwent PGT-A procedures, following with single euploid FET cycles in the assisted reproduction center from January 2016 to December 2022. Blastocysts were categorized for biopsy, and euploid embryos would be selected for transfer. Multivariate logistic regression was used to assess the effects of blastocoel expansion degree, inner cell mass (ICM) and trophectoderm (TE) grades and developmental stage (Day 5, 6 and 7) on euploidy and live birth rates. A total of 4172 blastocysts from 941 PGT-A cycles were included. Expansion 4 were associated with lower euploidy rate than expansion 5 (P = 0.011) and 6 (P = 0.001). Better ICM (P < 0.05 for A compared to B grade) increased blastocyst euploidy. Euploidy rate was significantly associated with A grade TE (P < 0.001). However, no relationship existed between blastocyst euploidy and developmental rate. Furthermore, live birth rates had no significant effect on most of morphological parameters in euploidy blastocyst FET cycles, only A grade TE was shown higher live birth rate than C grade (P = 0.024). The rate of euploidy in morphologically poor blastocysts is low in the cohort, but the developmental rate does not associate with euploidy. Moreover, only TE grades take association with live birth rates, when the single euploidy blastocyst was transplanted.

Background: Oocyte maturation arrest results in female infertility and the genetic etiology of this phenotype remains largely unknown. Previous studies have proven that cyclins play a significant role in the cell cycle both in meiosis and mitosis. Cyclin B3 (CCNB3) is one of the members of the cyclin family and its function in human oocyte maturation is poorly understood. Methods: 118 infertile patients were recruited and WES was performed for 68 independent females that experienced oocyte maturation arrest. Four mutations in CCNB3 were found and effects of these mutations were validated by Sanger sequencing and in vitro functional analyses. Results: We found these mutations altered the location of cyclin B3 which affected the function of cyclin dependent kinase 1 (CDK1) and led to mouse oocyte arrested at germinal vesicle (GV) stage. And then, low CDK1 activity influenced the degradation of cadherin 1 (CDH1) and the accumulation of cell division cycle 20 (CDC20) which are two types of anaphase-promoting complex/cyclosome (APC/C) activators and act in different stages of the cell cycle. Finally, APC/C activity was downregulated due to insufficient CDC20 level and resulted in oocyte metaphase I (MI) arrest. Moreover, we also found that the addition of PP1 inhibitor Okadic acid and CDK1 inhibitor Roscovitine at corresponding stages during oocyte in vitro maturation (IVM) significantly improved the maturation rates in CCNB3 mutant cRNAs injected oocytes. The above experiments were performed in mouse oocytes. Conclusion: Here, we report five independent patients in which mutations in CCNB3 may be the cause of oocyte maturation arrest. Our findings shed lights on the critical role of CCNB3 in human oocyte maturation.

Polycystic ovary syndrome (PCOS) adversely impacts oocyte quality, underscoring effective interventions as a central focus of research. Our study demonstrates that nicotinamide mononucleotide (NMN) not only restores NAD ⁺ homeostasis in oocytes from PCOS mice but also enhances the developmental rate of PCOS oocytes during in vitro maturation (IVM). Transcriptomic analysis and experimental validation suggest that NMN helps reverse the decline in PCOS oocyte quality by mitigating oxidative stress, improving mitochondrial function, and preserving spindle morphology. Mechanistically, we found that NMN supplementation upregulates SIRT1 expression, which responds to fluctuations in NAD ⁺ levels through the NAD ⁺ salvage pathway regulated by nicotinamide mononucleotide adenylyltransferase (NMNAT). In conclusion, our findings highlight the potential of NMN in improving PCOS oocyte quality, offering valuable insights for clinical PCOS treatment and the advancement of assisted reproductive technology.