Three-dimensional imaging and reconstruction of the whole ovary and testis: a new frontier for the reproductive scientist.

AbstractThe 3D functional reconstruction of a whole organ or organism down to the single cell level and to the subcellular components and molecules is a major future scientific challenge. The recent convergence of advanced imaging techniques with an impressively increased computing power allowed early attempts to translate and combine 2D images and functional data to obtain in-silico organ 3D models. This review first describes the experimental pipeline required for organ 3D reconstruction: from the collection of 2D serial images obtained with light, confocal, light-sheet microscopy or tomography, followed by their registration, segmentation and subsequent 3D rendering. Then, we summarise the results of investigations performed so far by applying these 3D image analyses to the study of the female and male mammalian gonads. These studies highlight the importance of working towards a 3D in-silico model of the ovary and testis as a tool to gain insights into their biology during the phases of differentiation or adulthood, in normal or pathological conditions. Furthermore, the use of 3D imaging approaches opens to key technical improvements, ranging from image acquisition to optimisation and development of new processing tools, and unfolds novel possibilities for multidisciplinary research

Editorial: 3D Modelling of Mammalian Embryos and Organs

The main scope of this Special issue was to gain understanding on how tissues and organs are arranged into integrative hierarchical levels of complexity, from the molecular to the morphological organization. To understand the underlying complexity of the relationships among these levels during morphogenesis or in the adult we must learn how to resolve single-cell spatial relationships in the three-dimensional (3D) organisation of tissues, organs and even of the whole organisms.Fil: Garagna, Silvia. Universita Degli Studi Di Pavia; ItaliaFil: Cebral, Elisa. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Biodiversidad y Biología Experimental y Aplicada. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Biodiversidad y Biología Experimental y Aplicada; ArgentinaFil: Aréchaga, Juan. Universidad del País Vasco; EspañaFil: Zuccotti, Maurizio. Universita Degli Studi Di Pavia; Itali

Gene Expression and Chromatin Organization during Mouse Oocyte Growth

AbstractMouse oocytes can be classified according to their chromatin organization and the presence [surrounded nucleolus (SN) oocytes] or absence [nonsurrounded nucleolus (NSN) oocytes] of a ring of Hoechst-positive chromatin around the nucleolus. Following fertilization only SN oocytes are able to develop beyond the two-cell stage. These studies indicate a correlation between SN and NSN chromatin organization and the developmental competence of the female gamete, which may depend on gene expression. In the present study, we have used the HSP70.1Luc transgene (murine HSP70.1 promoter + reporter gene firefly luciferase) to analyze gene expression in oocytes isolated from ovaries of 2-day- to 13-week-old females. Luciferase was assayed on oocytes after classification as SN or NSN type. Our data show that SN oocytes always exhibit a higher level of luciferase activity, demonstrating a higher gene expression in this category. Only after meiotic resumption, metaphase II oocytes derived from NSN or SN oocytes acquire the same level of transgene expression. We suggest that the limited availability of transcripts and corresponding proteins, excluded from the cytoplasm until GVBD in NSN oocytes, could explain why these oocytes have a lower ability to sustain embryonic development beyond the two-cell stage at which major zygotic transcription occurs. With this study we have furthered our knowledge of epigenetic regulation of gene expression in oogenesis

Spermatocyte apoptosis, which involves both intrinsic and extrinsic pathways, explains the sterility of Graomys griseoflavus Graomys centralis male hybrids

Spermatogenic impairment and the apoptotic pathways involved in establishing sterility of male hybrids obtained from crossing Graomys griseoflavus females with Graomys centralis males were studied. Testes from G. centralis, G. griseoflavus and hybrids were compared at different ages. Terminal transferase-mediated dUTP nick-end labelling assay (TUNEL), Fas, Bax and cytochrome c labelling were used for apoptosis evaluation, and calbindin D 28k staining as an anti-apoptotic molecule. In 1-month-old animals, spermatocytes were positive for all apoptotic markers, but moderate TUNEL (+) spermatocyte frequency was only found in G. centralis. At subsequent ages, the apoptotic markers were downregulated in testes from parental cytotypes, but not in hybrid testes. TUNEL (+) spermatocytes were present at 78% and 44% per tubule cross-section in 2- and 3-month-old hybrid animals, respectively. Pachytene spermatocyte death in adult hybrids occurs via apoptosis, as revealed by high caspase-3 expression. Calbindin was highly expressed in spermatocytes of adult hybrids, in which massive cell death occurs via apoptosis. Calbindin co-localisation with TUNEL or Fas, Bax and cytochrome c was very limited, suggesting an inverse regulation of calbindin and apoptotic markers. Hybrid sterility is due to breakdown of spermatogenesis at the pachytene spermatocyte stage. Both extrinsic and intrinsic pathways are involved in apoptosis of spermatocytes, which are the most sensitive cell type to apoptotic stimuli.Fil: Rodriguez, Valeria. Universidad Nacional de Córdoba. Facultad de Medicina. Cátedra de Bioquímica y Biología Molecular; ArgentinaFil: Díaz de Barboza, Gabriela Edith. Universidad Nacional de Córdoba. Facultad de Medicina. Cátedra de Bioquímica y Biología Molecular; ArgentinaFil: Ponce, Rubén Hugo. Universidad Nacional de Córdoba. Facultad de Odontología; ArgentinaFil: Merico, Valeria. Universita degli Studi di Pavia; ItaliaFil: Garagna, Silvia. Universita degli Studi di Pavia; ItaliaFil: Tolosa, Nori Graciela. Universidad Nacional de Córdoba. Facultad de Medicina. Cátedra de Bioquímica y Biología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones en Ciencias de la Salud. Universidad Nacional de Córdoba. Instituto de Investigaciones en Ciencias de la Salud; Argentin

Maternal Oct-4 is a potential key regulator of the developmental competence of mouse oocytes

Background The maternal contribution of transcripts and proteins supplied to the zygote is crucial for the progression from a gametic to an embryonic control of preimplantation development. Here we compared the transcriptional profiles of two types of mouse MII oocytes, one which is developmentally competent (MIISN oocyte), the other that ceases development at the 2-cell stage (MIINSN oocyte), with the aim of identifying genes and gene expression networks whose misregulated expression would contribute to a reduced developmental competence. Results We report that: 1) the transcription factor Oct-4 is absent in MIINSN oocytes, accounting for 2) the down-regulation of Stella, a maternal-effect factor required for the oocyte-to-embryo transition and of which Oct-4 is a positive regulator; 3) eighteen Oct-4-regulated genes are up-regulated in MIINSN oocytes and are part of gene expression networks implicated in the activation of adverse biochemical pathways such as oxidative phosphorylation, mitochondrial dysfunction and apoptosis. Conclusion The down-regulation of Oct-4 plays a crucial function in a sequence of molecular processes that leads to the developmental arrest of MIINSN oocytes. The use of a model study in which the MII oocyte ceases development consistently at the 2-cell stage has allowed to attribute a role to the maternal Oct-4 that has never been described before. Oct-4 emerges as a key regulator of the molecular events that govern the establishment of the developmental competence of mouse oocytes

OCT4 and the acquisition of oocyte developmental competence during folliculogenesis

The role that the transcription factor OCT4 plays during oocyte growth is yet unknown. In this review, we summarise the data on its potential role in the acquisition of oocyte developmental competence in the mouse. These studies describe the presence in MII oocytes and 2-cell embryos of an OCT4 transcriptional network that might be part of the molecular signature of maternal origin on which the inner cell mass and the embryonic stem cell-associated pluripotency is assembled and shaped. The Oct4-gene regulatory network thus provides a connection between eggs, early preimplantation embryos and embryonic stem cells

A half-century of studies on a chromosomal hybrid zone of the house mouse

The first natural chromosomal variation in the house mouse was described nearly 50 years ago in Val Poschiavo on the Swiss side of the Swiss–Italian border in the Central Eastern Alps. Studies have extended into neighboring Valtellina, and the house mice of the Poschiavo-Valtellina area have been subject to detailed analysis, reviewed here. The maximum extent of this area is 70 km, yet it has 4 metacentric races and the standard 40-chromosome telocentric race distributed in a patchwork fashion. The metacentric races are characterized by highly reduced diploid numbers (2n = 22–26) resulting from Robertsonian fusions, perhaps modified by whole-arm reciprocal translocations. The races hybridize and the whole Poschiavo-Valtellina area can be considered a “hybrid zone.” The studies of this area have provided insights into origin of races within hybrid zones, gene flow within hybrid zones and the possibility of speciation in hybrid zones. This provides a case study of how chromosomal rearrangements may impact the genetic structure of populations and their diversification.Fil: Giménez, Mabel Dionisia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Nordeste. Instituto de Biología Subtropical. Instituto de Biología Subtropical - Nodo Posadas | Universidad Nacional de Misiones. Instituto de Biología Subtropical. Instituto de Biología Subtropical - Nodo Posadas; Argentina. University of York; Reino UnidoFil: Förster, Daniel W.. Leibniz-institute For Zoo And Wildlife Research; Alemania. University of York; Reino UnidoFil: Jones, Eleanor P.. University of York; Reino Unido. Fera Science; Reino UnidoFil: Jóhannesdóttir, Fríđa. University of York; Reino Unido. Cornell University; Estados UnidosFil: Gabriel, Sofia I.. Universidade de Lisboa; Portugal. University of York; Reino UnidoFil: Panithanarak, Thadsin. Burapha University; Tailandia. University of York; Reino UnidoFil: Scascitelli, Moira. University of York; Reino UnidoFil: Merico, Valeria. Universita Di Pavia; ItaliaFil: Garagna, Silvia. Universita Di Pavia; ItaliaFil: Searle, Jeremy B.. University of York; Reino Unido. Cornell University; Estados UnidosFil: Hauffe, Heidi C.. Instituto Agrario San Michele All'adige Fondazione Edmund Mach. Centro Ricerca E Innovazione; Italia. University of York; Reino Unid