RESUMEN
PURPOSE: Oocytes from women presenting primary ovarian insufficiency (POI) generate viable embryos at a lower rate than non-POI women, but the mechanisms responsible for the lower oocyte quality remain elusive. Due to the scarcity of human oocytes for research, animal models provide a promising way forward. We aimed at investigating the molecular events characterizing final maturation in POI oocytes in a well-defined POI-like bovine model. METHODS: Single-cell RNA-sequencing of bovine control and POI-like, GV, and MII oocytes (n = 5 per group) was performed. DEseq2 was used to identify differentially expressed genes. Further, a Gene set enrichment analysis and a transcriptomic meta-analysis between bovine and human oocytes were performed. RESULTS: In control cows, we found 2223 differentially expressed genes between the GV and MII stages. Specifically, the affected genes were related to RNA processing and transport, protein synthesis, organelle remodeling and reorganization, and metabolism. The meta-analysis with a set of young human oocytes at different maturation stages revealed 315 conserved genes through the GV-MII transition in cows and humans, mostly related to meiotic progression and cell cycle. Gene expression analysis between GV and MII of POI-like oocytes showed no differences in terms of differentially expressed genes, pointing towards a substantial failure to properly remodel the transcriptome in the POI model, and with the clustering analysis indicating that the cow's genetic background had a higher impact than the oocyte's maturation stage. CONCLUSION: Overall, we have identified and characterized a valuable animal model of POI, paving the way to identifying new molecular mechanisms involved in POI.
Asunto(s)
Meiosis , Oocitos , Insuficiencia Ovárica Primaria , Bovinos , Femenino , Insuficiencia Ovárica Primaria/genética , Insuficiencia Ovárica Primaria/patología , Animales , Oocitos/crecimiento & desarrollo , Oocitos/metabolismo , Oocitos/patología , Meiosis/genética , Humanos , Transcriptoma/genética , Modelos Animales de Enfermedad , Oogénesis/genéticaRESUMEN
In brief: Preantral follicles constitute the largest follicle reserve in the mammalian ovary. This study assesses a mechanical isolation method to maximize the number of follicles retrieved from a defined cortex volume. Abstract: Primordial, primary, and secondary follicles (collectively defined as preantral follicles) constitute the most abundant source of gametes inside the mammalian ovarian cortex. The massive isolation of preantral follicles and the refinement of stage-specific protocols for in vitro follicle growth would provide a powerful tool to boost the rescue and restoration of fertility in assisted reproduction interventions in human medicine, animal breeding, and vulnerable species preservation. Nevertheless, together with an efficient culture system, the most significant limitation to implementing in vitro follicle growth is the lack of an efficient method to isolate viable and homogeneous subpopulations of primordial, primary, and secondary follicles suitable for in vitro culture. Our study provides a strategy for high-yielding mechanical isolation of primordial, primary, and early secondary follicles from a limited portion of the ovarian cortex in the bovine animal model. In the first part of the study, we refined a mechanical isolation protocol of preantral follicles, adopting specific methodological strategies to separate viable and distinct subpopulations of primordial (oblate and prolate forms), primary, and early secondary follicles from 0.16 cm3 of the ovarian cortex. In the second part of the study, we tested the effectiveness of the isolation protocol, considering the individual's age as a critical factor, bearing in mind the progressive decrease in the ovarian reserve that naturally accompanies the reproductive life span. Our study provides a way for designing quantitative and conservative fertility preservation approaches to preserve organ function and minimize the invasiveness of the interventions, also considering age-related differences.
Asunto(s)
Folículo Ovárico , Animales , Femenino , Folículo Ovárico/citología , Folículo Ovárico/fisiología , Bovinos , Ovario/citología , Factores de Edad , Envejecimiento/fisiologíaRESUMEN
The mammalian ovary is a substantial source of oocytes arranged into follicles at various stages of folliculogenesis, from the primordial to the ovulatory ones. Primordial follicles constitute the most abundant source of gametes inside the mammalian ovary at any given time.The isolation of a high number of primordial follicles, together with the development of protocols for in vitro follicle growth, would provide a powerful tool to fully exploit the female reproductive potential and boost the rescue and restoration of fertility in assisted reproduction technologies in human medicine, animal breeding, and preservation of threatened species. However, the most significant limitation is the lack of efficient methods for isolating a healthy and homogeneous population of viable primordial follicles suitable for in vitro culture. Here, we provide a fast and high-yield strategy for the mechanical isolation of primordial follicles from limited portions of the ovarian cortex in the bovine animal model.
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Oocitos , Folículo Ovárico , Bovinos , Animales , Femenino , Humanos , Ovario , Mamíferos , Técnicas Reproductivas AsistidasRESUMEN
In the past four decades, the bovine model has been highly informative and inspiring to assisted reproductive technologies (ART) in other species. Most of the recent advances in ART have come from studies in cattle, particularly those unveiling the importance of several processes that must be recapitulated in vitro to ensure the proper development of the oocyte. The maintenance of structural and functional communications between the cumulus cells and the oocyte and a well-orchestrated chromatin remodeling with the gradual silencing of transcriptional activity represent essential processes for the progressive acquisition of oocyte developmental competence. These markers are now considered the milestones of physiological approaches to increase the efficiency of reproductive technologies. Different in vitro approaches have been proposed. In particular, the so-called "pre-IVM" or "prematuration" is a culture step performed before in vitro maturation (IVM) to support the completion of the oocyte differentiation process. Although these attempts only partially improved the embryo quality and yield, they currently represent a proof of principle that oocytes retrieved from an ovary or an ovarian batch shouldn't be treated as a whole and that tailored approaches can be developed for culturing competent oocytes in several species, including humans. An advancement in ART's efficiency would be desirable in carnivores, where the success is still limited. Since the progress in reproductive medicine has often come from comparative studies, this review highlights aspects that have been critical in other species and how they may be extended to carnivores.
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Técnicas Reproductivas Asistidas , Animales , Bovinos , HumanosRESUMEN
Programmed cell death (PCD) has been discussed and categorized endlessly over the years. Herein, we comment on the chaotic classification of PCD in the mammalian ovary and the compartmentalized ovarian follicle based on an unfiltered overview of the extent of publication about some of the different modes of PCD on PubMed.
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Apoptosis , Ovario/citología , Femenino , Humanos , Ovario/metabolismoRESUMEN
The genus Prometopidia Hampson, 1902 and its type-species P. conisaria Hampson, 1902 are redescribed and newly discovered morphological characters are explained. The female holotype of Prometopidia arenosa Wiltshire, 1961, was studied and the species redescribed, its correct position in Prometopidia is verified. The new species P. joshimathensis sp. nov. is described from Joshimath area in India, Uttarakhand province. Sympatric with P. conisaria at Joshimath, P. joshimathensis also occurs at Shimla, Punjab province, and in central and eastern Nepal. Morphological and genetic differences found in the specimens of Nepal are considered subspecific, justifying the new taxon P. joshimathensis yazakii ssp. nov. Types and specimens of Prometopidia across its whole range of distribution from Afghanistan to Nepal, habitats, genitalia, remarkable morphological characters and DNA barcoding-results are figured.